Available via license: CC BY 4.0
Content may be subject to copyright.
1
1Effect of heavy metals on gut barrier integrity and gut microbiota
2
3Sweta Ghosh1, Syam P. Nukavarpu2 and Venkatakrishna Rao Jala1*
4
51Department of Microbiology and Immunology, Brown Cancer Center, Center for Microbiomics,
6Inflammation and Pathogenicity, University of Louisville, Louisville, Kentucky, United States of
7America.
82Department of Biomedical Engineering, and Department of Materials Science & Engineering,
9University of Connecticut, Storrs, Connecticut, United States of America
10
11
12 *Address for correspondence
13 Venkatakrishna Rao JALA, Ph. D
14 Department of Microbiology and Immunology,
15 UofL-Brown Cancer Center,
16 505 South Hancock Street # 323
17 Louisville, Kentucky – 40202, USA
18 Tel: 1-502 852-5523
19 Fax: 1-502 852-2123
20 Email: jvrao001@louisville.edu
21
22 ORCID: 0000-0002-4206-7305.
23
24
25
26
27
28 Abstract
Page 1 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
2
29 Exposure to environmental pollutants such as heavy metals lead to significant damage in intestinal
30 epithelial barrier, loss of microbial and immune homeostasis. The intestinal epithelial barrier
31 protects and regulates the responses against several endogenous and exogenous factors including
32 inflammatory cytokines, pathogens, toxins, and pollutants. Intestinal epithelial barrier dysfunction,
33 immune dysregulation and microbial dysbiosis are associated with several gastro-intestinal (GI)-
34 related disorders including inflammatory bowel disease (IBD). The mechanisms and consequences
35 of exposure of environmental toxins on gut barrier function and mucosal immune system are not
36 fully understood. This review explores some of the recent findings of heavy metals and their effect
37 on intestinal barrier function, microbiota, and their contributions to human health and pathogenesis
38 of GI-related disorders such as IBD.
39
40
41
42 Keywords
43 Heavy metal toxicity, pollution, intestinal barrier dysfunction, tight junctional proteins, gut
44 microbiota, microbial metabolites.
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59 Introduction
Page 2 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
3
60 The industrial revolution caused a significant increase in the release of several toxic
61 chemicals into environment that became part of our food cycle. Epidemiological studies suggested
62 that an association between metal pollution and worst health outcomes, and increased risk for
63 numerous disorders (Tchounwou et al., 2012, Chowdhury et al., 2018). Exposure of heavy metals
64 through water, air and foods affects different organs, and causes nervous system disorders, skin
65 lesions, vascular damage, immune system dysfunction, birth defects, cancer and gastro-intestinal
66 (GI) and kidney dysfunction. Additionally, exposure of multiple metals may exert cumulative
67 adverse effects on overall health.
68 The effects of metals on target organs vary from each other based on their route of
69 exposure, amounts of metals (low vs high), duration of exposure as well as species. For instance,
70 mice develop intestinal adenomas and carcinomas upon chronic exposure of high concentration of
71 hexavalent chromium in drinking water, but not in rats (Thompson et al., 2013). Similarly, acute
72 exposure of high-dose mercury and lead may induce kidney failure, abdominal colic pain and
73 bloody diarrhea. In contrast, chronic exposure at low doses regularly can cause complications such
74 as neuropsychiatric disorders including fatigue, anxiety, and detrimental impacts on intelligence
75 quotient (IQ) and intellectual function in children (Balali-Mood et al., 2021). Mechanistically, high
76 dose exposure of heavy metals leads to DNA damage, disrupts proteins and mechanisms involved
77 in DNA synthesis and repair. Commonly, heavy metal-induced reactive oxygen species,
78 suppression of oxidative stress and inactivation of critical metabolic enzymes are responsible for
79 heavy metal mediated-adverse effects. However, the mechanism of actions of each metal may vary
80 based on organ, tissue, and the cell types involved. Current review focuses on how heavy metal(s)
81 exposure show impact on gut microbiota and gut barrier functions leading to GI-related disorders.
82 Recent studies highlighted that the exposure of environmental pollutants led to significant
83 changes in the composition of gut microbiota (Shao and Zhu, 2020, Breton et al., 2013b), epithelial
84 barrier function, and increased intestinal inflammation (Celebi Sozener et al., 2020, Mitamura et
85 al., 2021, Lindell et al., 2022). Microbial dysbiosis, gut barrier dysfunction and immune
86 dysregulation are associated with gastro-intestinal (GI)-related disorders including inflammatory
87 bowel disease (IBD) (Martini et al., 2017, Guan, 2019), which comprises of ulcerative colitis (UC)
88 and Crohn’s disease (CD). Gut barrier consists of single layer of intestinal epithelial cells that
89 selectively allows trans-migration of nutrients and protects from external challenges and
90 pathogenic bacteria (Ghosh et al., 2021). Additionally, gut barrier mediates the cross talk between
Page 3 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
4
91 commensal microbes and immune system, and provides the first line of defense against pathogens,
92 toxins, and environmental pollutants. Gut barrier dysfunction results in leakiness of bacteria and
93 toxins leading to immune imbalance at gut mucosal sites that potentially promotes pathogenesis
94 of GI-related disorders including IBD. The mechanisms and long-term impact of environmental
95 toxins on gut barrier functions and related disorders are not fully understood. Gut microbiota and
96 their metabolites can play an important role in regulation of the environmental toxins-mediated
97 etiology and pathogenesis of gastrointestinal disorders (Claus et al., 2016, Bist and Choudhary,
98 2022). The use of probiotics has become as an attractive strategy to reduce the adverse effects of
99 toxic metals (Duan et al., 2020). In this manuscript, we reviewed the structure of intestinal barrier
100 and its impact upon exposure to heavy metals. The review also explored some of the possible
101 mechanisms through which heavy metals interact and affect gut barrier function, microbiome and
102 microbial metabolites.
Page 4 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
5
103 1. Gut barrier dysfunction and systemic responses
104
105 1.1. Intestinal epithelial cells
106 The gastrointestinal (GI) barrier consists of three key components: the mucus layer, the
107 intestinal epithelial cell layer, and the immunological barrier. Goblet cells (GC) are responsible
108 for secretion of mucins, which create protective mucus layer (Johansson et al., 2013, Kim and Ho,
109 2010). This mucus layer, composed of an outer and an inner layer, serves as the primary barrier
110 against luminal microorganisms and foreign antigens (Bansil and Turner, 2006, Thornton and
111 Sheehan, 2004, Leal et al., 2017). The mucus layer consists of glycoprotein sheets, featuring a
112 densely packed inner layer and a less dense outer layer that serve as a niche for various intestinal
113 bacteria, acting as a carbon source for microbial metabolism (Pelaseyed et al., 2014, Johansson et
114 al., 2013). The mucus functions as a barrier for hydrophilic solutes, which can only be transported
115 via specific transporters (Turner, 2009, Li et al., 2020). Mucins and their glycosylation status play
116 a critical role in regulating gut barrier functions. Deficiencies in mucin production lead to defective
117 gut barrier activities and promote GI-related disorders, including inflammatory bowel disease
118 (IBD), irritable bowel syndrome (IBS), and cancer.
119 The intestinal epithelium is composed of various cellular subtypes, such as enterocytes,
120 Paneth cells, M cells, endocrine cells, and tuft cells, which collectively play essential roles in
121 digestion, nutrient absorption, and protection against pathogens, among other functions (reviewed
122 elsewhere (Peterson and Artis, 2014, Clevers, 2013, Okumura and Takeda, 2017)). Intestinal
123 epithelial cell (IEC) junctions inclduing tight junctions (TJ), adherens junction (AJ), desmosomes
124 and gap Junctions (GJ) are specialized structures present in the epithelial cell membranes. They
125 establish contacts between intestinal epithelial cells and regulate the transport of molecules based
126 on their size and charge through the paracellular space (Zihni et al., 2016, González-Mariscal et
127 al., 2003, Van Itallie and Anderson, 2014). Defects in paracellular permeability are associated with
128 several GI-related disorders (Vermette et al., 2018, Odenwald and Turner, 2013, Vanuytsel et al.,
129 2021, Bischoff et al., 2014).
130 Enterocytes, comprising over 80% of intestinal epithelial cells, are polarized cells with
131 microvilli expanding the absorptive surface. These cells are interconnected via several proteins
132 that enable cell to cell adherence through formation of junctions. Enterocytes undergo apoptosis,
133 and are replaced by crypt-derived stem cells. It is known that the enterocytes regulate water and
Page 5 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
6
134 nutrient absorption, contribute to intestinal layer formation, macromolecular transportation and
135 digestion(Snoeck et al., 2005). They are involved in macromolecular transport via receptor-
136 mediated endocytosis(Stern and Walker, 1984, Snoeck et al., 2005). Increased inflammation, such
137 as elevated TNF-α, accelerates enterocyte turnover and proliferation, leading to heightened
138 shedding and apoptosis, potentially compromising the intestinal barrier and promoting bacterial
139 translocation, particularly in conditions like IBD.
140 Paneth cells are another subset of differentiated secretory cells located at the base o f the
141 crypts of Lieberkuhn. Paneth cells are essential for enteric immune homeostasis and actively
142 secrete antimicrobial peptides such as alpha defensins, lysozyme, and phospholipases A2 that limit
143 bacterial numbers. Infants with necrotizing enterocolitis (NEC) have significantly decreased levels
144 of Paneth cells compared to age-matched controls(McElroy et al., 2013, Underwood, 2012). It was
145 shown that depletion or dysfunction of Paneth cells in mouse models results in a NEC-like
146 phenotype, indicating the importance of Paneth cell function in immature intestine(Lueschow et
147 al., 2018, Lueschow and McElroy, 2020, Sampath et al., 2017). Paneth cells can directly sense gut
148 commensals and control intestinal barrier penetration in a myeloid differentiation marker 88
149 (MyD88)-dependent manner at the intestinal host-microbial interface(Vaishnava et al., 2008).
150 Paneth cells depend on autophagy to control their secretion capacity of anti-microbial peptides.
151 Mutations in genes like Atg16L1 disrupt this process, leading to reduced Paneth cell function,
152 imbalanced gut microbiota, compromised barrier integrity, and an elevated risk of diseases such
153 as Crohn's disease in humans (Cray et al., 2021).
154 M cells are specialized epithelial cells found in the gut-associated lymphoid tissue (GALT)
155 of Peyer’s patches of the small intestine, isolated lymphoid follicles, colonic patches and
156 nasopharyngeal associated lymphoid tissues (NALT) (Dillon and Lo, 2019). These cells are
157 involved in immune responses, especially antigen sampling, uptake of microorganisms and interact
158 with dendritic cells or lymphocytes to initiate adaptive immunity. M cells deliver samples of
159 foreign material from the lumen to organized mucosal lymphoid tissues. They interact closely with
160 immune cells of Peyer’s patches and play an important role in the initiation of immunological
161 response and tolerance. During chronic inflammation, increased levels of M cells were observed
162 along with selective apoptosis of M cells, leading to elevated uptake of microorganisms and
163 inflammation (Kucharzik et al., 2000b). Increased apoptosis of M cells during ileitis conditions
Page 6 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
7
164 lead to breakdown of intestinal barrier resulting translocation bacteria and enhanced inflammation
165 (Kucharzik et al., 2000a) .
166 Enteroendocrine cells representing 1% of the intestinal epithelium are also a type of
167 intestinal secretary cells that mediate hormone release and are critical for digestion.
168 Enteroendocrine cells are a major component of a specialized chemosensory system that can sense
169 the intestinal microbiota and their metabolites. These cells secrete peptide hormones and classical
170 cytokines to the surrounding immune cells and modulate both innate and adaptive immune
171 systems. Enteroendocrine cells possess cytoplasmic processes in close proximity to the enteric
172 nerve terminals and mediate several physiological functions including visceral hyperalgesia,
173 intestinal motility, and synaptic transmission. Importantly, enteroendocrine hormones can
174 modulate the intestinal epithelial barrier function through both transcellular and paracellular
175 pathways (Yu et al., 2019).
176 Tuft cells are chemosensory sentinel cells that sense signals from the local milieu and
177 communicate to immune cells within the intestine. These cells have been recently described to be
178 critical in exerting an immune response against helminths (Allaire et al., 2018). The underlying
179 mechanisms of these sensory signals are yet to be established. It is known that Tuft cells secrete
180 IL-25 continuously to maintain type 2 innate lymphoid cells (ILC2) homeostasis. Helminths
181 infection leads to increased levels of IL-25 production by Tuft cells, which directly acts on ILC2
182 to release IL-13, and in turn IL-13 acts on tuft cells and goblet cells to promote hyperplasia in an
183 IL-13Rα1/IL-4αR-dependent manner. These series of activities lead to an increase in mucin levels
184 to expel the parasite from the GI tract and protect the gut barrier (von Moltke et al., 2016, Ting
185 and von Moltke, 2019).
186
187
188 1.2. Tight junction (TJ) proteins in gut epithelium.
189
190 Claudins are 20–27 kDa tetraspanmembrane proteins contain four hydrophobic
191 transmembrane domains with two extracellular loops and N- and C-terminal cytoplasmic domains
192 (Van Itallie and Anderson, 2006). The extracellular loops are responsible for homophilic and/or
193 heterophilic TJ protein-protein interactions and the formation of ion-selective channels. The
194 intracellular C-terminal domain anchors claudin to the cytoskeleton through interactions with
195 PDZ-binding domain proteins including ZO-1, -2 and -3 (Morita et al., 1999). Thus far, at least 24
Page 7 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
8
196 members of the claudin family have been reported. Claudins play an important role in ‘pore
197 formation’ and ‘sealing (barrier forming)’ of the epithelium in the GI tract and classified as ‘pore’
198 versus ‘barrier forming’ TJ proteins (Günzel and Yu, 2013). The ‘pore’ forming claudins
199 specifically increase paracellular permeability either for molecules of a certain size or of a certain
200 charge (or both), and decrease transepithelial resistance but leave the epithelial barrier function
201 against macromolecules intact (Günzel and Yu, 2013). Similarly, ‘barrier’ forming claudins
202 decrease paracellular permeability and increase transepithelial resistance. It was also demonstrated
203 that treatment with IL-17A protected against TNF-α-mediated barrier disruption in Caco-2 cells
204 through regulating occludin, suggesting a direct role for IL-17 and IL-17R signaling on epithelial
205 cells (Lee et al., 2015). In an independent study, Dr. Rao’s group reported that the deficiency of
206 occludin increased the susceptibility to ethanol-induced colonic mucosal barrier permeability and
207 liver damage in mice (Mir et al., 2016). Moreover, inflammatory cytokines like TNF-α and IFN-γ
208 are reported to downregulate intestinal epithelial barrier function by reorganizing several TJ
209 proteins such as zonula occludens-1 (ZO-1), claudin-1, claudin-4, occludin and JAM-A
210 (Zolotarevsky et al., 2002). Th2 cytokines like IL-4 and IL-13 also cause an increase in intestinal
211 permeability through induction of epithelial apoptosis and expression of the pore-forming tight
212 junction protein claudin-2 (Madden et al., 2002, Ceponis et al., 2000, Berin et al., 1999).
213
214 1.3. Adherens Junction (AJ) proteins in gut epithelium:
215 Adherens junctions, also known as zonula adherens, are located on the lateral membrane of
216 epithelial cells and maintain cell-to-cell contact, cell polarity, motility, and proliferation (Hartsock
217 and Nelson, 2008, Perez-Moreno et al., 2003). Interconnections between transmembrane proteins,
218 intracellular adaptor proteins, and the cytoskeleton form AJ protein complexes. AJs are reported
219 to present beneath TJs and are required for the assembly of TJs. TJs along with AJs are linked to
220 the peri-junctional ring of cellular actin and myosin leading to junction regulation via the
221 cytoskeleton. AJs primarily consist of cadherin and catenin proteins that are linked to several
222 intracellular cytoskeletal protein domains (Perez-Moreno et al., 2003). One of the most
223 characterized AJ protein, E-cadherin binds to repeat regions of -catenin, -catenin, or -catenin
224 (plakoglobin) at varying affinities based on phosphorylation states (Perez-Moreno and Fuchs,
225 2006, Gumbiner, 1996, Halbleib and Nelson, 2006). Moreover, the functions of AJ and TJ proteins
226 are regulated by myosin and actin proteins that form a dense ring, which encircles the cell.
Page 8 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
9
227 Activation of actomyosin contraction is regulated by phosphorylation of myosin II regulatory light
228 chain (MLC) by MLC kinase (MLCK) and is implicated in AJ and TJ protein regulation. It has
229 been demonstrated that MLCK-driven MLC phosphorylation is involved in signal transduction
230 pathways that influence gut barrier function in response to diverse stimuli (Turner, 2000,
231 Cunningham and Turner, 2012, Shen et al., 2006). In the case of IBD, decreased E-cadherin–
232 catenin complexes lead to a loss of cell-to-cell adhesions along with impairment of the integrity of
233 the mucosal barrier. Exposure of luminal substances to the mucosal immune system leads to
234 inflammation and further increased gut barrier permeability (Mehta et al., 2015). Furthermore,
235 modulation of intestinal permeability by oxidative stress leads to redistribution of E-cadherin and
236 β-catenin (Rao, 2008, Rao et al., 2002).
237
238 1.4. Desmosomes in gut epithelium:
239 Desmosomes are located on the basolateral membrane and are known to provide mechanical
240 support due to connections to the intracellular cytoskeletal framework (Gumbiner, 1996, Green
241 and Simpson, 2007, Nekrasova and Green, 2013). Desmosomes form links between adjacent cells
242 and provide connection between intermediate filaments of the cell cytoskeletons. Desmosomes
243 consist of the cadherin family of proteins, desmoglein and desmocollins (Gumbiner, 1996,
244 Nekrasova and Green, 2013). Together with AJs, desmosomes maintain the integrity of the
245 epithelium by imparting strong adhesive bonds. Studies revealed that Desmoglein 2 (Dsg2), which
246 is expressed in enterocytes, potentially contributes to the pathogenesis of CD (Spindler et al.,
247 2015). It was demonstrated that Dsg2 regulates intestinal epithelial barrier function in enterocytes
248 by modulating the p38MAPK signaling cascade (Ungewiss et al., 2017).
249
250 1.5. Gap Junctions (GJ) in gut epithelium:
251 Gap junctions form tiny connections between adjacent cells allowing passage of small
252 molecules, ions and electrical signals (Goodenough and Paul, 2009). Small molecules and ions
253 diffuse through GJ channels that span the bilayers of both cells and the extracellular space that
254 separate them. GJ are formed by head-to-head docking of proteins called connexins. Connexins
255 are hexameric protein transmembrane complex that spans between membranes of adjacent cells
256 (Goodenough et al., 1996). The gap inside the hexameric GJ protein passes the respective signal
257 or molecule to the next cells. Ey et al. showed that during IEC injury, activated Toll like receptor-2
Page 9 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
10
258 (TLR2) drives Connexin-43 (Cx43) protein synthesis and subsequently increases gap junctional
259 intercellular communication leading to increased epithelial permeability and related diseases (Ey
260 et al., 2009).
261 In general, the dysfunction of gut epithelial cells can have far-reaching consequences,
262 primarily stemming from disruptions in the mucus layer and junctional protein complexes within
263 the gut epithelium. This dysfunction ultimately leads to an increase in intestinal permeability,
264 allowing bacteria and bacterial endotoxins to leak through. This, in turn, triggers an elevation in
265 the levels of inflammatory cytokines and recruitment of inflammatory cells. These interconnected
266 events culminate in the development of microbial dysbiosis, a substantial rise in mucosal and
267 systemic inflammation. These conditions, in turn, play a pivotal role in the onset and progression
268 of various disorders, such as IBD, diabetes, neurological disorders, liver diseases, and various
269 types of cancers, among others.
270
271 2. Microbiota is essential for gut barrier function
272 The gut microbiota is often referred as an essential ‘organ’ due to the vast density and richness
273 of microbial life that exist in the gut lumen (Eckburg et al., 2005). The composition of microbiota
274 is influenced by several factors including but not limited to host genetics, age, dietary habits, drugs,
275 environmental and lifestyles. Microbiota can be also inheritable from mother to child with slight
276 differences between delivery and postpartum birth (Dominguez-Bello et al., 2010, Palmer et al.,
277 2007, Ursell et al., 2012). Germ free (GF) mice exhibit significant deficiency in gut barrier function
278 suggesting critical role of microbiota in development of gut barrier components (Wang et al., 2021,
279 Parker et al., 2018). The inner mucus layer of GF mice is more penetrable to bacteria-sized beads
280 compared to conventionally raised mice suggesting defects in mucin layer (Johansson et al., 2015).
281 Introducing fecal commensal bacteria collected from conventionally raised mice into GF mice
282 induces a normal colonic barrier structure by increasing the thickness of the colonic mucus layer
283 and mucin glycosylation (Hayes et al., 2018). These studies demonstrated the importance of gut
284 microbiota in developing gut mucus barrier and maintenance of homeostasis. It was shown that
285 GF mice are more susceptible to epithelial injury as a result of an impaired intestinal barrier via
286 downregulation of claudin 4, occludin, TFF3, and MUC3 protein expression and IL-22 secretion
287 (Hernández-Chirlaque et al., 2016). It was shown that the mice treated with broad-spectrum
288 antibiotics, such as ampicillin, streptomycin, or clindamycin for 14 days led to reduced bacterial
Page 10 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
11
289 diversity and richness. These mice displayed an increase in gut permeability and decreased
290 expression of intestinal tight junction proteins such as ZO-1, occludin and claudin-1 (Feng et al.,
291 2019), re-enforcing the importance of microbiota in gut barrier function. Now there is an
292 unequivocal evidence for the critical role of microbiota in the development and maintenance of
293 gut barrier function. Hence, rebuilding a healthy gut microbiota, while maintaining the known
294 resident commensals, is critical (Schmidt et al., 2018). Although the determination of ‘beneficial’
295 microbes seems to be a challenge, pro-, pre- and syn-biotics may provide opportunities to restore
296 intestinal homeostasis and enhance gut barrier function in addition to blocking of unwarranted
297 inflammation and dysregulation (Gibson and Roberfroid, 1995, Schrezenmeir and de Vrese, 2001,
298 Ghosh et al., 2022d, Ghosh et al., 2022c). Impact of microbial metabolites on gut barrier was
299 reviewed elsewhere (Ghosh et al., 2021). Members of the gut microbiota influence the host
300 metabolic and immune status by modulating nutrient metabolism, xenobiotic and drug metabolism,
301 and production of antimicrobial metabolites that limit numbers of competing microbes for the same
302 niche. Gut microbes and metabolites influence the structure of the gastrointestinal tract, integrity
303 of the gut barrier and differentiation of various immune cell subsets (Jandhyala et al., 2015,
304 Thursby and Juge, 2017). However, most of the functions of individual species and metabolites
305 remain unclear.
306 The mucosal immune system profoundly impacts gut barrier functions both in health and
307 disease conditions (reviewed in (Arpaia et al., 2013, Konieczna et al., 2013, Mazmanian et al.,
308 2005, Kayama and Takeda, 2020, Zuo et al., 2020, Allaire et al., 2018)). Microbial metabolites
309 regulate host immunity by exploiting metabolite-specific immune cell receptors such as aryl
310 hydrocarbon receptor (AhR), Farnesoid X receptor (FXR), pregnane X receptor (PXR), membrane
311 bile acid receptor (M-BAR/TGR5), purinergic receptor (P2X7), and G-protein coupled receptors
312 (GPR 41, GPR43, GPR109A) (Liu et al., 2022c, Liu et al., 2022a). These receptors play crucial
313 roles in host-microbiota interactions, are expressed at various levels in different cell types such as
314 intestinal epithelial cells, innate lymphoid cells, macrophages, T cells and dendritic cells
315 (Furusawa et al., 2013). Functional immune responses often modulate gut barrier integrity; hence,
316 the fine tuning of gut microbiota to evoke necessary immune responses to alter intestinal barrier
317 function is critical. Major confounding factors that disturb the composition of gut microbiota and
318 gut barrier functions include exposure of heavy metals and associated disorders. The following
Page 11 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
12
319 sections review the current understanding of metal exposure and their impact on gut barrier
320 functions.
321
322 2. The role of heavy metals in regulation of gut microbiota and gut barrier function.
323 Humans are exposed daily to numerous environmental chemicals, heavy metals, pesticides,
324 organic pollutants, mycotoxins, food additives and other contaminants (Celebi Sozener et al.,
325 2020). Exposure of the heavy metals such as arsenic (As), lead (Pb), mercury (Hg), cadmium (Cd)
326 and chromium (Cr) results in adverse health effects in humans and animals (Balali-Mood et al.,
327 2021). Around 60% of ingested metals are absorbed by intestine resulting in severe oxidative
328 stress, gut barrier damage leading to increased intestinal inflammation (Shao and Zhu, 2020,
329 Assefa and Kohler, 2020, Feng et al., 2018) (Figure 1). The functional consequences of exposure
330 to these environmental contaminants on the human gut microbiota are yet to be investigated. Table
331 1 summarizes the known effects of heavy metals on host health and gut microbiota. Importance of
332 gut microbiota in metabolism of metals was demonstrated using GF mice. It was shown that heavy
333 metals significantly accumulated in blood and target organs of GF mice compared to normal mice
334 following exposure (Breton et al., 2013a).
335
336 2.1. Arsenic (As)
337 About 225 million people in over 70 countries in the world are chronically exposed to heavy metal
338 arsenic (Naujokas et al., 2013, Podgorski and Berg, 2020), making it as an environmental health
339 crisis with no known treatment. In USA, about 3 million individuals are exposed to arsenic mostly
340 through unregulated domestic well water used for drinking purposes (Ayotte et al., 2017). Arsenic
341 is a Group I human carcinogen and chronic exposure leads to toxic effects in multiple organs like
342 liver, kidney, bladder, skin, and central nervous system (Banerjee, 2011, IARC., 2012, Hong et
343 al., 2014, Garza-Lombo et al., 2019, Hunt et al., 2014, Coryell et al., 2019). While skin has been
344 reported as the major target organ for arsenic with precancerous and cancerous outcomes (Hunt et
345 al., 2014), recent data is shedding light on its adverse on the lifestyle disorders including diabetes,
346 cardiovascular diseases and obesity (Moon et al., 2018, Moon et al., 2012, Young et al., 2018,
347 Farkhondeh et al., 2019, Grau-Perez et al., 2017, Navas-Acien et al., 2008, Chen and Karagas,
348 2013, Bulka et al., 2017). Recent studies highlighted that chronic exposure of arsenic has
349 significant impact on gastrointestinal system leading to irritation, nausea, pain and vomiting
Page 12 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
13
350 (Jomova et al., 2011) as well as perturbation of gut microbiome (Choiniere and Wang, 2016b).
351 Previous reports suggested that the chronic arsenic exposure abrogates gut barrier function
352 exhibiting symptoms like dyspepsia, gastroenteritis and chronic diarrhea (Choiniere and Wang,
353 2016a, Fernández Fernández et al., 2019, Chiocchetti et al., 2019b, Chiocchetti et al., 2019a, Guha
354 Mazumder and Dasgupta, 2011). Rodents exposed to arsenic metabolite (monomethylarsonic acid)
355 target GI tract leading to enlargements of the intestinal wall, edemas, hemorrhages, necrosis and
356 ulcerations as well as increase in the incidence of squamous metaplasia of absorptive epithelial
357 cells of the colon and rectum (Arnold et al., 2003). Sub-chronic exposure of arsenic led to
358 disruption of colonic epithelial structure and barrier function (Chiocchetti et al., 2018). The
359 transmission electron microscope (TEM) image of Caco-2 cells after arsenic exposure (As
360 (III) ≥ 0.075 mg/L; As (V) ≥ 0.75 mg/L) for 14 days showed the disruption in microvilli structures
361 resulting increased paracellular transport and barrier dysfunction (Chiocchetti et al., 2018).
362 Trivalent forms of arsenic significantly induce inflammatory cytokines such as IL-6, IL-8 and
363 TNF-α as well as oxidative stress in colon epithelial cells (Calatayud et al., 2013, Calatayud et al.,
364 2014). Recently, it was shown that gut microbiota is required for full protection against acute
365 arsenic toxicity using mouse models (Coryell et al., 2018). In this study, the authors showed that
366 when mice were given the antibiotic cefoperazone (Cef) before being exposed to inorganic sodium
367 arsenate (iAsV), the iAs accumulated significantly in their organs. These mice also exhibited lower
368 levels of iAs in their stools compared to the mice that didn't receive antibiotics. Furthermore, in
369 line with these findings, mice without gut microbiota (GF mice) exposed to iAs also showed higher
370 levels of iAs in their organs compared to mice with normal microbiota. In essence, these studies
371 suggest that gut microbiota is necessary for effectively processing/metabolizing iAs for excretion
372 from the host's body. This study also demonstrated that transplantation of human microbiota, into
373 GF mice protected from arsenic-induced adverse effects. It was shown that functional arsenic
374 detoxification enzyme (As3mt) and specific bacterium called ‘Faecalibacterium’ are essential for
375 protection against acute arsenic toxicity in mouse models (Coryell et al., 2018). iAs speciation is
376 significantly impacted by microbial-dependent iAs metabolism (reviewed in (Tsai et al.,
377 2009),(Kruger et al., 2013)). The iAs is transported through glycerol or phosphate transporters in
378 bacteria. Arsenate can be reduced arsenite which may be expelled by ArsAB. Alternatively,
379 arsenite can be methylated by ArsM to MMAs (III), DMAs (III) and TMAs. Briefly,
380 biotransformation of iAs by microbes can be divided into four functional groups: (i) As(III)
Page 13 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
14
381 oxidation (ii) iAs(V) reduction (iii) iAs methylation and demethylation, and (iv) iAs
382 transport(Zhao et al., 2019). The toxic forms of As(III)/MMAs(III) can be actively expelled from
383 bacterial cells using arsenic transporters encoded by arsB, arsP, or acr3 (associated with
384 resistance). Arsenic methylation, governed by arsM (related to resistance), results in the release
385 of arsenic through volatilization or the formation of products like mono-, di-, or trimethylarsines.
386 It is possible that lack of microbes in the host (either in anti-biotic treated or germ-free mice) failed
387 to metabolize iAs and expel from host body leading to the accumulation of iAs in organs, and this
388 accumulation may have resulted in target organs including GI-tract.
389 Gut microbiome analysis of individuals exposed to arsenic revealed that abundance of
390 pathogenic bacteria is positively, and commensal gut bacteria is negatively correlated with
391 increased arsenic concentration (Brabec et al., 2020, Chen et al., 2021). It was shown by Chi et al
392 that exposure of environmental relevant levels of As (100 ppb) to the mice led to significant
393 changes in the functional metagenome. These include increase in the genes involved in energy
394 metabolism, LPS synthesis, oxidative stress responses, and DNA repair (Chi et al., 2017).
395 Additionally, arsenic exposure also enriched genes that encode conjugative transposon proteins,
396 components of the multidrug efflux system, and the synthesis of multiple vitamins (Chi et al.,
397 2017). Human gut microbiota can biochemically transform arsenic-containing compounds
398 (arsenicals) leading to arsenic speciation and bioavailability (Lu et al., 2014, Lu et al., 2013, Yin
399 et al., 2022). It is possible that either microbiota metabolizes arsenic to produce less-toxic
400 arsenicals or microbial metabolites can counteract against arsenic-induced toxicity or organ
401 damage. Our group showed that gut microbial metabolite, urolithin A (UroA) protected colon
402 epithelial cells against inorganic arsenic exposure by reducing inorganic trivalent arsenic (iAs3+)-
403 induced oxidative stress and enhancing tight junctional proteins (Ghosh et al., 2022a, Ghosh et al.,
404 2022b). We have demonstrated that UroA protected from arsenic-induced cell death in colon
405 epithelial cells and alleviated iAs3+-induced barrier dysfunction in both colon epithelial cell
406 monolayers and a human 3D small intestinal tissue model. Importantly, UroA treatment
407 significantly protected from arsenic-induced gut barrier permeability by enhancing tight junction
408 proteins such as zonula occludens-1, occludin, and claudin-4. Mechanistically, we showed that
409 UroA treatment reduced iAs3+-induced reactive oxygen species (ROS) through regulating genes
410 involved oxidative stress pathways. Further in vivo studies are required to determine the impact of
411 metabolites in ameliorating arsenic-induced toxicities. Moreover, use of probiotics like
Page 14 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
15
412 Bifidobacterium and Lactobacillus modulated arsenic transformation abilities by enhancing
413 excretion and detoxification (Liu et al., 2022b). It is pertinent to recall the epidemiological
414 observations in people from Northport, WA who suffered from chronic exposure of iAs have
415 reported 15 times more IBD cases than national average (Pynn). Therefore, targeting gut barrier
416 dysfunction and inflammation simultaneously by beneficial microbiota and microbial metabolites
417 would offer better protection against arsenic-induced adverse events.
418
419 2.2. Lead (Pb)
420 A non-essential heavy metal, Lead (Pb) exerts its toxic effects on several organs including liver,
421 kidney, nervous system, cardiovascular, gastrointestinal, and reproductive systems (Flora et al.,
422 2012, Liu et al., 2021, Yu et al., 2021b). Soil contamination with gasoline, paint exposes humans
423 to Pb related toxification. An epidemiological study by Lanphear et al determined the
424 concentrations of Pb in blood of adults aged 20 years or older and confirmed that low-level
425 exposure of Pb was associated with risk factors for cardiovascular disease mortality in USA
426 (Lanphear et al., 2018). Pb-induced oxidative stress by ROS is one of the causes for Pb-poisoning
427 and related adverse health effects. Hence, the modulation of cellular thiols by antioxidants against
428 ROS production has emerged as a key therapeutic approach (Flora et al., 2012). Several studies
429 elaborated the adverse effects of Pb on gastrointestinal tract and gut microbiota in several species.
430 Mice exposed to low-dose (chronic model) or high-dose (acute model) Pb exhibited reduced
431 colonic MUC2 levels, tight junctional proteins such as ZO-1, Claudin-1 and Occludin and
432 increased intestinal permeability suggesting its direct impact on GI functions (Zhai et al., 2019b).
433 Intestinal epithelial cell damage was observed in honeybees when they were exposed to lead oxide
434 (PbO) and/or cadmium oxide (CdO) nanoparticles for 9 days (Dabour et al., 2019). In these studies,
435 they also showed that CdO and PbO induced the cytological alterations in intestinal epithelial cells
436 using TEM methodologies (Dabour et al., 2019). Xia et al showed that chronic exposure of Pb (15
437 weeks) induced gut microbial dysbiosis and metabolic disorder in mice (Xia et al., 2018).
438 Especially, Pb exposure caused significant increase in the levels of hepatic triglycerides, total
439 cholesterol as well as genes involved in lipid metabolism. Further, Pb exposure led to change in
440 the structure and richness of the gut microbiota, especially relative abundance of Firmicutes and
441 Bacteroidetes was altered compared to normal control mice (Xia et al., 2018). Even early exposure
442 to Pb during developmental period enhanced the risk for obesity in adulthood by altering the gut
Page 15 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
16
443 microbiota independent of gender (Wu et al., 2016). Gao et al. demonstrated by using multi-omics
444 approaches that Pb exposure affected gut microbiome trajectories including metabolic pathways
445 in C57BL/6 mice (Gao et al., 2017). Fecal microbiota transplantation (FMT) from donors
446 supplemented with galactooligosaccharide (GOS) altered the gut microbiota composition and
447 improved the recovery of the gut barrier function in mice after Pb exposure. Zhai et al showed that
448 the mice treated with antibiotics exhibited accumulation of higher levels of Pb in their blood and
449 primary organs, along with reduced Pb levels in their feces (Zhai et al., 2019a) indicating
450 importance of microbiota in clearance of Pb from host. Typically, certain gut microorganisms can
451 boost the healing of intestinal mucosal injuries, support the balance of gut immunity, and
452 contribute to the reduction of gut inflammation. It was shown that the Pb exposure leads to gut
453 dysbiosis resulting in decrease in beneficial bacteria such as Akkermansia muciniphila,
454 Faecalibacterium prausnitzii, and Oscillibacter ruminantium (Zhai et al., 2017). Interestingly, oral
455 administration of these Pb-intolerant gut microbes, A. muciniphila, F. prausnitzii, and O.
456 ruminantium reversed the Pb-induced toxicity in mouse models(Zhai et al., 2019b, Zhai et al.,
457 2019a). Oral administration of these bacteria led to reduction in Pb burden in target organs and
458 blood stream, and increased fecal Pb excretion in chronically Pb exposed mice (Zhai et al., 2017).
459 Moreover, oral supplementation of these Pb-intolerant gut microbes (especially F. prausnitzii, and
460 O.ruminantium) enhanced gut barrier function by upregulating TJPs such as ZO-1, occludin and
461 claudin-1 proteins in the colon and small intestine. Further, these treatments increased the levels
462 of short chain fatty acids (SCFAs) that have been shown to have significant impact on gut barrier
463 function. Mechanistically, treatment with these microbes significantly reduced Pb-induced hepatic
464 and renal oxidative stress in the mice. These studies further showed that the three Pb-intolerant gut
465 microbe strains showed a high adsorption capacity for Pb2+ compared with E. coli K12(Zhai et al.,
466 2017). In summary, these findings indicate that gut microbiota can limit the absorption and
467 accumulation of Pb in host tissues. When Pb-intolerant intestinal microbes like F. prausnitzii and
468 O. ruminantium were orally administered to mice, there was a significant decrease in Pb
469 accumulation and alleviate Pb toxicity. This underscores the significance of adjusting gut
470 microbiota as a potential strategy for minimizing the Pb toxicity in the host. Pb exposure to GF
471 mice resulted in increased accumulation of Pb in blood and organs compared to conventional SPF
472 mice suggesting gut microbiota play a major role in excretion of heavy toxic metals (Breton et al.,
473 2013a). Recently, a human cohort study revealed that increased urinary Pb is associated with the
Page 16 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
17
474 presence of Proteobacteria suggesting environmental toxins potentially induce microbial
475 dysbiosis leading to adverse health in humans (Eggers et al., 2019). Further in-depth studies are
476 required to define cause or consequence and mechanisms of actions of Pb in various disease
477 conditions.
478
479 2.3. Mercury (Hg)
480 Mercury (Hg) is a well-known biohazard that exists in the environment in its elemental form, as
481 inorganic mercury, or as organic mercury (Rice et al., 2014). The most common and major source
482 of Hg in the ecosystems is Methylmercury (MeHg) i.e., Hg in its organic methylated form. MeHg
483 is highly toxic as it covalently binds to the glutathione and cystine residues of any protein in the
484 host system and causes alterations in the protein structure and function (Pinto et al., 2020). Mercury
485 exposure causes DNA damage, induction of oxidative stress, mitochondrial dysfunction, increased
486 lipid peroxidation, altered calcium homeostasis at cellular level. Mercury poisoning affects all the
487 organs as it can readily pass into the circulation system, ultimately leading to death. Likewise, gut
488 barrier permeability and microbial dysbiosis has been closely associated to exposure to mercury.
489 Vázquez et al. showed that inorganic divalent mercury, Hg (II) and MeHg can exert to toxicity to
490 intestinal epithelial cells and mucosa by generating redox imbalance (Vazquez et al., 2014).
491 Generation of ROS or reactive nitrogen species (RNS) and decrease in glutathione by mercury
492 redistributed the F-actin and ZO-1 protein in the intestinal cells resulting in elevated barrier
493 permeability (Vazquez et al., 2014). It is shown that Hg exposure significantly down regulated
494 expression of intercellular junction proteins like claudin 1, occludin, ZO-1 and junctional adhesion
495 molecule 1 (JAM1) in colon epithelial cells. Hg also increased the intestinal cell volume and
496 membrane permeability without any loss in cell viability, thus it promotes the uptake of other toxic
497 metals in same conditions (Aduayom et al., 2005). Bolan et al. reported that gut microbes like
498 Escherichia coli and Lactobacillus acidophilus, or chelating agents (EDTA and DMPS)
499 significantly reduced the Hg-induced permeability in in vitro gastrointestinal/Caco-2 cell intestinal
500 epithelium model (Bolan et al., 2021). It was suggested that microbiota-dependent protection
501 against heavy metal-induced intestinal permeability may be associated with indirect intestinal
502 sequestration of metal by the gut bacteria through adsorption on bacterial surface (Bolan et al.,
503 2021). Moreover, probiotics strain Lactobacillus brevis 23017 exhibits strong mercury binding
504 capacity, which may potentially reduce the Hg exposure to the host cells and protects the gut
Page 17 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
18
505 barrier integrity against Hg-induced toxicity (Jiang et al., 2018). It was shown that the L. brevis
506 23017 reduced the Hg-induced inflammation and oxidative stress through regulating MAPK and
507 NF-κB pathways (Jiang et al., 2018). Additionally, L. brevis 23017 supplementation maintained a
508 normal mucosal barrier via modulation of tight junction proteins. Depletion of gut microbiota by
509 antibiotics significantly increased the levels of Hg accumulation in several organs like cerebellum,
510 liver, and lungs in mice that are exposed to Hg in comparison with the mice that are harboring
511 normal microbiota (Seki et al., 2021). It was shown that MeHg is captured and inactivated by the
512 hydrogen sulfide and hydrogen persulfide (by forming sulfur adducts to excrete out) produced by
513 microbes which resulted in reduced MeHg toxicity (Seki et al., 2021). Furthermore, gut microbes
514 also regulate Hg biotransformation and bioaccumulation in freshwater fish (Micropterus
515 salmoides), marine fish (Acanthopagrus latus) and polar bear (Ursus maritimus) (Tan et al., 2022,
516 Watson et al., 2021, Yang et al., 2021b). Zhao et al showed that subchronic oral mercury caused
517 loss in body weight, intestinal injury and led to changes in gut microbiota, aggravated apoptosis in
518 mice (Zhao et al., 2020). In case reports, it was shown that occupation exposure of mercury vapors
519 led to the increased episodes of disease reactivation in patient with chronic ulcerative colitis
520 (Cummings and Rosenman, 2006). It is interesting recall that mercury released from dental ‘silver’
521 fillings provoked an increase in mercury and anti-biotic resistant bacteria in oral and intestines
522 (Summers et al., 1993). These mercury- and antibiotic-resistant isolates include the Staphylococci,
523 the Enterococci, and the members of the family Enterobacteriaceae. Mercury exposure led to the
524 generation of mercury- and antibiotic resistance plasmids in the normal microbiota of primates
525 (Summers et al., 1993). Further studies are required to define effects of mercury on human GI
526 system and its role in promoting GI-related disorders.
527
528 2.4. Cadmium (Cd)
529 Cadmium (Cd) is a highly toxic environmental pollutant which majorly gets accumulated from
530 cigarette smoking and diet. Cd exposure is related to several diseases like diabetes, chronic kidney
531 disease, osteoporosis, obesity, liver disease, cardiovascular diseases and cancer (Tinkov et al.,
532 2018, Tellez-Plaza et al., 2012, Tinkov et al., 2017). Cd induces cytotoxicity by enhancing
533 inflammation, gut barrier dysfunction, oxidative stress, endoplasmic reticulum stress, endocrine
534 disruption and genomic instability (Rafati Rahimzadeh et al., 2017). Several studies reported that
535 exposure to Cd-induced leaky gut with an irregular distribution and reduced expression of tight
Page 18 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
19
536 junctional proteins like in ZO-1, ZO-2, JAM-A, occludin, and claudin-1 in intestinal epithelial
537 cells and mice (Rusanov et al., 2015, Zhai et al., 2016, Duizer et al., 1999, Liu et al., 2020a).
538 Relative changes in transepithelial/transendothelial electrical resistance (TEER) after high dose
539 (100 and 300 μM) of Cd exposure concentration were result of damaged tight intercellular
540 junctions and loss of the monolayer integrity (Rusanov et al., 2015). Even low dose exposure of
541 Cd significantly caused the gut microbial dysbiosis in mice and aggravated the liver injury by
542 increased intestinal permeability (Liu et al., 2020a). In this study, they showed that Cd exposure
543 significantly down regulated the expression of tight junction proteins such as ZO-1, JAM-A and
544 Occludin led to the increased intestinal permeability and liver injury. Importantly, it was shown
545 that low Cd exposure led to decrease in the Akkermansia muciniphila, which known commensal
546 to protect the gut barrier integrity suggesting adverse impacts of Cd exposure in regulating gut
547 microbiota composition (Liu et al., 2020a). Cd interfered with gut mucosa and goblet cells in dose
548 and site -dependent manner in zebrafish (Danio rerio) (Motta et al., 2022). Cd altered the mucosal
549 efficiency by changing localization and distribution of glycan residues, metallothionein expression
550 in intestinal cells (Motta et al., 2022). Moreover, oral intake of Cd changed the adaptive immune
551 activities in mice that was associated with increased gut permeability, intestinal tissue damage and
552 inflammation. Importantly, Cd exposure reduced the beneficial commensal Lactobacillus strain
553 (Ninkov et al., 2015). It was shown that Cd exposure modified the gut-liver axis along with gut
554 dysbiosis in ApoE4-KI males, which are the most susceptible mice strain to neurological damages
555 (Zhang et al., 2021). Exposure of Cd in mice also modulated the levels of gut microbiota like
556 Eisenbergiella, Blautia, Clostridium_XlVa, Lactobacillus, Bifidobacterium and decreased levels
557 of microbial metabolites such as short chain fatty acids (e.g., butyrate) (Li et al., 2019, Liu et al.,
558 2014). It was that Cd exposure to mice led to significant alteration in the gut microbiota population
559 including several butyrate-producers along with changes in the bile acid fraction of the gut
560 metabolome (Li et al., 2019). Moreover, Cd exposure caused decrease in the thickness of inner
561 mucus layer and reduced the growth of Bacteroidetes, Lactobacillus and Bifidobacterium followed
562 by altered SCFAs metabolism (Liu et al., 2014). Yang et al. showed that Cd can show impact on
563 liver, kidney and ovary functions in adolescent rats along with downregulation in Prevotella and
564 Lachnoclostridium microbiota and upregulation in Escherichia coli and Shigella (Yang et al.,
565 2021a).Whereas use of gut microbes like Escherichia coli and Lactobacillus acidophilus reduced
566 the Cd-mediated intestinal epithelial cell permeability (Bolan et al., 2021). Probiotic Lactobacillus
Page 19 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
20
567 plantarum CCFM8610 also reduced that Cd-induced toxicity and intestinal motility dysfunction
568 in mice with decreased Cd content in the tissues and blood of animals along with enhanced fecal
569 cadmium excretion (Liu et al., 2020b). Moreover, Lactobacillus plantarum CCFM8610 exhibited
570 excellent Cd binding and antioxidative capacity, which helped in protection of the Cd mediated
571 disruption of tight junctions in the intestinal epithelial cells. Overall, Cd exposure leads to gut
572 microbial dysbiosis and exerts its imp act on gut barrier integrity leading to increased gut
573 permeability and promoting GI-related disorders and supplementation of beneficial certain
574 probiotics could protect from Cd-induced adverse effects.
575
576 2.5. Chromium (Cr)
577 Chromium, a known mutagenic, metallic contaminant gets bioaccumulated in animals from solid
578 waste disposal, pesticides, fertilizers, mining activities and residues from industrial productions.
579 Cr is available in multiple oxidation state, but trivalent and hexavalent forms are most stable
580 (Balali-Mood et al., 2021). Cr in trivalent state is required for lipid metabolism, protein metabolism
581 and as a cofactor for insulin action, whereas hexavalent Cr is responsible for disease pathogenesis
582 like organ failure, rhinitis, asthma, gut barrier dysfunction and cancer. A recent meta-analysis from
583 human study showed that Cr exposure increased mortality and occurrence of larynx, lung, thyroid,
584 bone, kidney, bladder, and testicular cancer (Deng et al., 2019). Another human study reported
585 that groundwater contamination of Cr caused severe GI distress and digestive problems in both
586 male and females along with changes in hematological parameters like RBC and platelet counts
587 (Sharma et al., 2012). Studies reveled that long-term exposure to Cr exaggerated gastrointestinal
588 symptoms and body weight loss in colorectal cancer model with change in microbial abundance
589 (Zhang et al., 2020). Moreover, Cr affected the gastrointestinal tract of earthworm (Eisenia fetida)
590 via nuclear damage in gut epithelia with hemorrhage and ulceration (Tang et al., 2019). The TEM
591 images reveled that the Cr exposed earthworm contained subcellular injury with short, messy and
592 rough gut villus and damaged organelles. A recent study showed that exposure of hexavalent
593 chromium in ducks led to gut barrier damage through downregulation of ZO-1, occludin, claudin-
594 1, and MUC2 expression (Xing et al., 2022). Activation of NLRP3 inflammasome and generation
595 of oxidative stress contributed to the shortening of the intestinal villi and gut barrier dysfunction.
596 To alleviate the Cr induced damage, use of Lactobacillus plantarum TW1-1 reduced Cr
597 accumulation and restored gut bacterial homeostasis in mice (Wu et al., 2017). Cr exposure
Page 20 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
21
598 effectively dysregulated that Bacteroidetes and Firmicutes homeostasis and increased abundance
599 of S24-7, Prevotella, and Clostridiales, but lowered Lachnospiraceae. The lower butyrate
600 producer Lachnospiraceae hampered the development of intestinal epithelial cells leading to
601 inflammation and oxidative stress after Cr exposure in mice (Wu et al., 2017). Long term Cr
602 exposure also increased the presence of Firmicutes and Actinobacteria and removed the presence
603 of Deferribacteres, Intestinimonas, Butyricimonas, Butyricicoccus,
604 Lachnospiraceae_FCS020_group and Ruminococcaceae_V9D2013_group in chickens (Li et al.,
605 2021). Several studies in Wister rats and chickens proved that gut microbiota is essential for first
606 line of defense against Cr toxicity and microbes may act as a prebiotic or probiotic to attenuate Cr
607 toxicity (Shrivastava et al., 2005, Wang et al., 2022b, Li et al., 2022). Upreti et al developed
608 resistance to Cr up to 64 ppm in probiotic Lactobacilli stain with chronological chronic exposures.
609 These probiotic strains showed no antibiotic resistance but ameliorated Cr induced GI-toxicity
610 (Upreti et al., 2011).Thus, this study showed that alteration of microbial composition can be
611 therapeutic for Cr induced GI damage and toxicity.
612 Moreover, some metals like Aluminum (Al) despite not having high density like ‘heavy
613 metals’ considered toxic due to the level of toxicity. Being most abundant metal element in the
614 Earth's crust, continuous exposure of Al leads to bioaccumulation and severe toxicity in several
615 tissues including gut (Vignal et al., 2016). Studies have reported that excessive Al induced the
616 apoptosis of intestinal epithelial cells, disrupted the TJ proteins leading to increased intestinal
617 permeability and gut barrier dysfunction (Vignal et al., 2016, Hao et al., 2022). Damage of the gut
618 barrier due to Al exposure leads to impaired immune system in affected individuals. Yu et al.
619 reported that dietary Al exposure altered the human gut microbiota composition and
620 supplementation with probiotics like L. plantarum CCFM639 abrogated the toxicity of Al(Yu et
621 al., 2021a). In Wistar rats dietary intake of Al diminished the natural gut microbiota diversity and
622 affected the physiological homeostasis (Wang et al., 2022a).
623
624 Outstanding questions and Major challenges
625 What are the molecular and cellular mechanisms of heavy metals-mediated gut microbial
626 dysbiosis and gut barrier dysfunction? Especially, how do heavy metals disrupt epithelial
627 junctional proteins? Is the metal-induced oxidative stress solely responsible for long-term
628 adverse effects that were observed in the GI tract?
Page 21 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
22
629 How does microbiota regulate metal toxicity and is there any specific bacterium or bacterial
630 consortium responsible for detoxification of specific metal toxicity? For example, is there
631 any bacterial specificity for heavy metals that either increase or decrease toxicity? If so,
632 what are the mechanisms for metal-specificity?
633 What are interactions of microbial metabolites with heavy metals and their implications on
634 gut barrier functions in humans? Especially, it will be interesting to determine whether
635 direct supplementation of microbial metabolites render the benefits against heavy metal
636 toxicity and overcome the microbial dysbiosis?
637 Effects of heavy metal exposure on skin and lung microbiome is less explored. It is
638 important to evaluate as metals directly influence the pathogenesis of skin and respiratory
639 disorders.
640 Developing appropriate models to investigate metal toxicity is a major challenge in this
641 area of research. For instance, recapitulation of human exposure (low vs high) of heavy
642 metals in animal models is a major constraint due to limitations in duration of exposure
643 and controlling route of administration may not provide complete array of effects observed
644 in humans. Secondly, the metabolism rates of metals are significantly different between
645 humans and animal models (mice or rats), adds another layer of complexity. Thirdly, the
646 effects of heavy metals on gut environment may not be detectable in early stages, but metal-
647 induced gut barrier dysfunction may promote and cause other organ damage due to leakage
648 of constant bacterial endotoxins and inflammatory cytokines. Therefore, developing
649 methods and biomarkers to detect gut barrier function significantly aid in the design and
650 development of therapeutics.
651
652 Summary
653 The gut epithelial barrier is the most important system for nutrition absorption and required for
654 protection from exogenous pathobionts and environmental pollutants (Ghosh et al., 2021, Gillois
655 et al., 2018). In homeostatic conditions, the gut microbiota and gut barrier play a protective role
656 against toxic effects of environmental pollutants and reduce the risk factors for GI-related
657 disorders. Nevertheless, several exogenous and endogenous factors such as diet, xenobiotics,
658 heavy metals and microbial dysbiosis as well as increased inflammation contribute to the enhanced
Page 22 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
23
659 barrier dysfunction and systemic toxicity. The extent and duration of exposure are crucial factors
660 in determining the hazards that may jeopardize the host's overall health. Failure to promptly
661 address these issues could result in the continuous deterioration of epithelial barriers, posing an
662 increased risk for a range of gastrointestinal and non-gastrointestinal disorders. Alterations in gut
663 physiology, encompassing but not limited to the degradation of the mucus layer, disruption of
664 junctional proteins, heightened intestinal permeability, inflammation, and microbial imbalances,
665 are the underlying causes of gut barrier dysfunction and immune irregularities. Heavy metal
666 exposure impairs the metabolic activity of the gut microbiome, giving rise to inflammatory
667 responses and cellular damage. In contrast, the restoration of microbial equilibrium and the
668 sequestration of heavy metals by specific microbiota hold potential benefits for the host. The
669 concept of "Intestinal Bioremediation" is emerging as a valuable tool for immobilizing toxic metals
670 by selectively employing relevant microbes to shield against their harmful effects (George et al.,
671 2021), although the precise mechanisms underpinning this microbiota-mediated defense against
672 environmental pollutants continue to be under investigation. Recent research has unveiled that gut
673 bacteria communicate with the host signaling system through metabolites, which in turn regulate
674 intestinal immunity and barrier defense (Figure 2). The vital role played by the gut microbiota in
675 maintaining gut homeostasis can thwart the non-essential toxicity of heavy metals. Therefore,
676 leveraging beneficial microbiota and their metabolites as a therapeutic approach holds great
677 promise in treating pollutant-induced gut barrier dysfunction.
Page 23 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
24
678 Figure legends
679
680 Figure 1: Effect of heavy metals on gut. Heavy metals such as Arsenic (As), Lead (Pb), Mercury
681 (Hg), Cadmium (Cd) and Chromium (Cr) leads to increased oxidative stress, altered gut microbial
682 composition and inflammation potentially leading to gut leakiness and gut barrier dysfunction.
683
684 Figure 2: Effect of heavy metals on human health. Environmental pollutants such as Arsenic
685 (As), Lead (Pb), Mercury (Hg), Cadmium (Cd) and Chromium (Cr) lead to several adverse health
686 effects. Heavy metals-induced loss of gut barrier integrity and microbial balance initiate
687 inflammation and toxicity in host tissue and organs. Consumption of healthy diets, treatment with
688 beneficial gut microbiota or microbial metabolites potentially mitigate the adverse effects of heavy
689 metal toxicity and restores gut homeostasis.
Page 24 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
25
691 Abbreviations
692 AhR: aryl hydrocarbon receptor
693 AJ: Adherens junction
694 DMPS: Dimercaptopropanesulfonic acid
695 EDTA: Ethylenediaminetetraacetic acid
696 FMT: Fecal microbiota transplantation
697 FXR: Farnesoid X receptor
698 GC: Goblet cell
699 GF: Germ free
700 GJ: Gap junctions
701 GI: Gastrointestinal tract
702 IBD: Inflammatory bowel disease
703 IBS: Irritable bowel syndrome
704 IEC: Intestinal epithelial cells
705 IL: Interleukin
706 JAM: Junctional adhesion molecule
707 M Cell: Microfold cells
708 MUC: Mucin
709 NLRP: Nod-like receptor protein
710 PXR: Pregnane X receptor
711 ROS: Reactive oxygen species
712 TEM: Transmission electron microscopy
713 TGR5: G-protein-coupled bile acid receptor
714 TNF-α: Tumor necrosis factor alpha
715 TJ: Tight junction
716 UroA: Urolithin A
717 ZO: Zonula occludens
718
719
Page 25 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
26
720 Acknowledgements: VRJ is supported by NIH/NCI (CA191683), NIH/NIGMS CoBRE grant
721 (P20GM125504-01), NIH/NIEHS, (P30ES030283), The Jewish Heritage Fund for Excellence
722 Research Enhancement Grant and UofL-Health Brown Cancer Center. The authors thank Dr.
723 Bodduluri Haribabu for poof reading the manuscript and insightful discussions. Images were
724 prepared using Biorender.com
725
726
727 Declarations
728 Funding: VRJ is supported by NIH/NCI (CA191683), NIH/NIGMS CoBRE grant
729 (P20GM125504-01), NIH/NIEHS (P30ES030283), The Jewish Heritage Fund for Excellence
730 Research Enhancement Grant and UofL Health-BCC.
731
732 Author contribution statement:
733 SG and VRJ collected the literature, conceptualized and wrote the review article. SP contributed
734 by conceptualizing, proof reading and editing the article.
735
736 Conflict of Interest: VRJ is one of the scientific co-founders of Artus Therapeutics. SG and SPN
737 have no conflicts of interest to declare.
738
Page 26 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
27
739 References:
740
741 ADUAYOM, I., DENIZEAU, F. & JUMARIE, C. 2005. Multiple effects of mercury on cell
742 volume regulation, plasma membrane permeability, and thiol content in the human
743 intestinal cell line Caco-2. Cell Biol Toxicol, 21, 163-79.
744 ALLAIRE, J. M., CROWLEY, S. M., LAW, H. T., CHANG, S. Y., KO, H. J. & VALLANCE, B.
745 A. 2018. The Intestinal Epithelium: Central Coordinator of Mucosal Immunity. Trends
746 Immunol, 39, 677-696.
747 ARNOLD, L. L., ELDAN, M., VAN GEMERT, M., CAPEN, C. C. & COHEN, S. M. 2003.
748 Chronic studies evaluating the carcinogenicity of monomethylarsonic acid in rats and mice.
749 Toxicology, 190, 197-219.
750 ARPAIA, N., CAMPBELL, C., FAN, X., DIKIY, S., VAN DER VEEKEN, J., DEROOS, P., LIU,
751 H., CROSS, J. R., PFEFFER, K., COFFER, P. J. & RUDENSKY, A. Y. 2013. Metabolites
752 produced by commensal bacteria promote peripheral regulatory T-cell generation. Nature,
753 504, 451-5.
754 ASSEFA, S. & KOHLER, G. 2020. Intestinal Microbiome and Metal Toxicity. Curr Opin Toxicol,
755 19, 21-27.
756 AYOTTE, J. D., MEDALIE, L., QI, S. L., BACKER, L. C. & NOLAN, B. T. 2017. Estimating
757 the High-Arsenic Domestic-Well Population in the Conterminous United States. Environ
758 Sci Technol, 51, 12443-12454.
759 BALALI-MOOD, M., NASERI, K., TAHERGORABI, Z., KHAZDAIR, M. R. & SADEGHI, M.
760 2021. Toxic Mechanisms of Five Heavy Metals: Mercury, Lead, Chromium, Cadmium,
761 and Arsenic. Front Pharmacol, 12, 643972.
762 BANERJEE, M., BHATTACHARJEE, P., GIRI, A.K. 2011. Arsenic-induced Cancers: A Review
763 with Special Reference to Gene, Environment and Their Interaction. Genes and
764 Environment, 33, 128-140.
765 BANSIL, R. & TURNER, B. S. 2006. Mucin structure, aggregation, physiological functions and
766 biomedical applications. Current Opinion in Colloid & Interface Science, 11, 164-170.
767 BERIN, M. C., YANG, P. C., CIOK, L., WASERMAN, S. & PERDUE, M. H. 1999. Role for
768 IL-4 in macromolecular transport across human intestinal epithelium. Am J Physiol, 276,
769 C1046-52.
770 BISCHOFF, S. C., BARBARA, G., BUURMAN, W., OCKHUIZEN, T., SCHULZKE, J.-D.,
771 SERINO, M., TILG, H., WATSON, A. & WELLS, J. M. 2014. Intestinal permeability – a
772 new target for disease prevention and therapy. BMC Gastroenterology, 14, 189.
773 BIST, P. & CHOUDHARY, S. 2022. Impact of Heavy Metal Toxicity on the Gut Microbiota and
774 Its Relationship with Metabolites and Future Probiotics Strategy: a Review. Biol Trace
775 Elem Res.
776 BOLAN, S., SESHADRI, B., KEELY, S., KUNHIKRISHNAN, A., BRUCE, J., GRAINGE, I.,
777 TALLEY, N. J. & NAIDU, R. 2021. Bioavailability of arsenic, cadmium, lead and mercury
778 as measured by intestinal permeability. Sci Rep, 11, 14675.
779 BRABEC, J. L., WRIGHT, J., LY, T., WONG, H. T., MCCLIMANS, C. J., TOKAREV, V.,
780 LAMENDELLA, R., SHERCHAND, S., SHRESTHA, D., UPRETY, S., DANGOL, B.,
781 TANDUKAR, S., SHERCHAND, J. B. & SHERCHAN, S. P. 2020. Arsenic disturbs the
782 gut microbiome of individuals in a disadvantaged community in Nepal. Heliyon, 6, e03313.
Page 27 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
28
783 BRETON, J., DANIEL, C., DEWULF, J., POTHION, S., FROUX, N., SAUTY, M., THOMAS,
784 P., POT, B. & FOLIGNE, B. 2013a. Gut microbiota limits heavy metals burden caused by
785 chronic oral exposure. Toxicol Lett, 222, 132-8.
786 BRETON, J., MASSART, S., VANDAMME, P., DE BRANDT, E., POT, B. & FOLIGNE, B.
787 2013b. Ecotoxicology inside the gut: impact of heavy metals on the mouse microbiome.
788 BMC Pharmacol Toxicol, 14, 62.
789 BULKA, C. M., MABILA, S. L., LASH, J. P., TURYK, M. E. & ARGOS, M. 2017. Arsenic and
790 Obesity: A Comparison of Urine Dilution Adjustment Methods. Environ Health Perspect,
791 125, 087020.
792 CALATAYUD, M., DEVESA, V. & VÉLEZ, D. 2013. Differential toxicity and gene expression
793 in Caco-2 cells exposed to arsenic species. Toxicol Lett, 218, 70-80.
794 CALATAYUD, M., GIMENO-ALCAÑIZ, J. V., VÉLEZ, D. & DEVESA, V. 2014. Trivalent
795 arsenic species induce changes in expression and levels of proinflammatory cytokines in
796 intestinal epithelial cells. Toxicol Lett, 224, 40-6.
797 CELEBI SOZENER, Z., CEVHERTAS, L., NADEAU, K., AKDIS, M. & AKDIS, C. A. 2020.
798 Environmental factors in epithelial barrier dysfunction. J Allergy Clin Immunol, 145, 1517-
799 1528.
800 CEPONIS, P. J., BOTELHO, F., RICHARDS, C. D. & MCKAY, D. M. 2000. Interleukins 4 and
801 13 increase intestinal epithelial permeability by a phosphatidylinositol 3-kinase pathway.
802 Lack of evidence for STAT 6 involvement. J Biol Chem, 275, 29132-7.
803 CHEN, F., LUO, Y., LI, C., WANG, J., CHEN, L., ZHONG, X., ZHANG, B., ZHU, Q., ZOU, R.,
804 GUO, X., ZHOU, Y. & GUO, L. 2021. Sub-chronic low-dose arsenic in rice exposure
805 induces gut microbiome perturbations in mice. Ecotoxicol Environ Saf, 227, 112934.
806 CHEN, Y. & KARAGAS, M. R. 2013. Arsenic and cardiovascular disease: new evidence from
807 the United States. Ann Intern Med, 159, 713-4.
808 CHI, L., BIAN, X., GAO, B., TU, P., RU, H. & LU, K. 2017. The Effects of an Environmentally
809 Relevant Level of Arsenic on the Gut Microbiome and Its Functional Metagenome.
810 Toxicological Sciences, 160, 193–204.
811 CHIOCCHETTI, G. M., DOMENE, A., KÜHL, A. A., ZÚÑIGA, M., VÉLEZ, D., DEVESA, V.
812 & MONEDERO, V. 2019a. In vivo evaluation of the effect of arsenite on the intestinal
813 epithelium and associated microbiota in mice. Arch Toxicol, 93, 2127-2139.
814 CHIOCCHETTI, G. M., VELEZ, D. & DEVESA, V. 2019b. Inorganic arsenic causes intestinal
815 barrier disruption. Metallomics, 11, 1411-1418.
816 CHIOCCHETTI, G. M., VÉLEZ, D. & DEVESA, V. 2018. Effect of subchronic exposure to
817 inorganic arsenic on the structure and function of the intestinal epithelium. Toxicol Lett,
818 286, 80-88.
819 CHOINIERE, J. & WANG, L. 2016a. Exposure to inorganic arsenic can lead to gut microbe
820 perturbations and hepatocellular carcinoma. Acta pharmaceutica Sinica. B, 6, 426-429.
821 CHOINIERE, J. & WANG, L. 2016b. Exposure to inorganic arsenic can lead to gut microbe
822 perturbations and hepatocellular carcinoma. Acta Pharm Sin B, 6, 426-429.
823 CHOWDHURY, R., RAMOND, A., O'KEEFFE, L. M., SHAHZAD, S., KUNUTSOR, S. K.,
824 MUKA, T., GREGSON, J., WILLEIT, P., WARNAKULA, S., KHAN, H.,
825 CHOWDHURY, S., GOBIN, R., FRANCO, O. H. & DI ANGELANTONIO, E. 2018.
826 Environmental toxic metal contaminants and risk of cardiovascular disease: systematic
827 review and meta-analysis. BMJ, 362, k3310.
Page 28 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
29
828 CLAUS, S. P., GUILLOU, H. & ELLERO-SIMATOS, S. 2016. The gut microbiota: a major
829 player in the toxicity of environmental pollutants? NPJ Biofilms Microbiomes, 2, 16003.
830 CLEVERS, H. 2013. The intestinal crypt, a prototype stem cell compartment. Cell, 154, 274-84.
831 CORYELL, M., MCALPINE, M., PINKHAM, N. V., MCDERMOTT, T. R. & WALK, S. T.
832 2018. The gut microbiome is required for full protection against acute arsenic toxicity in
833 mouse models. Nat Commun, 9, 5424.
834 CORYELL, M., ROGGENBECK, B. A. & WALK, S. T. 2019. The Human Gut Microbiome's
835 Influence on Arsenic Toxicity. Curr Pharmacol Rep, 5, 491-504.
836 CRAY, P., SHEAHAN, B. J. & DEKANEY, C. M. 2021. Secretory Sorcery: Paneth Cell Control
837 of Intestinal Repair and Homeostasis. Cellular and Molecular Gastroenterology and
838 Hepatology, 12, 1239-1250.
839 CUMMINGS, C. E. & ROSENMAN, K. D. 2006. Ulcerative colitis reactivation after mercury
840 vapor inhalation. Am J Ind Med, 49, 499-502.
841 CUNNINGHAM, K. E. & TURNER, J. R. 2012. Myosin light chain kinase: pulling the strings of
842 epithelial tight junction function. Annals of the New York Academy of Sciences, 1258, 34-
843 42.
844 DABOUR, K., AL NAGGAR, Y., MASRY, S., NAIEM, E. & GIESY, J. P. 2019. Cellular
845 alterations in midgut cells of honey bee workers (Apis millefera L.) exposed to sublethal
846 concentrations of CdO or PbO nanoparticles or their binary mixture. Sci Total Environ,
847 651, 1356-1367.
848 DENG, Y., WANG, M., TIAN, T., LIN, S., XU, P., ZHOU, L., DAI, C., HAO, Q., WU, Y., ZHAI,
849 Z., ZHU, Y., ZHUANG, G. & DAI, Z. 2019. The Effect of Hexavalent Chromium on the
850 Incidence and Mortality of Human Cancers: A Meta-Analysis Based on Published
851 Epidemiological Cohort Studies. Front Oncol, 9, 24.
852 DILLON, A. & LO, D. D. 2019. M Cells: Intelligent Engineering of Mucosal Immune
853 Surveillance. Frontiers in Immunology, 10.
854 DOMINGUEZ-BELLO, M. G., COSTELLO, E. K., CONTRERAS, M., MAGRIS, M.,
855 HIDALGO, G., FIERER, N. & KNIGHT, R. 2010. Delivery mode shapes the acquisition
856 and structure of the initial microbiota across multiple body habitats in newborns. Proc Natl
857 Acad Sci U S A, 107, 11971-5.
858 DUAN, H., YU, L., TIAN, F., ZHAI, Q., FAN, L. & CHEN, W. 2020. Gut microbiota: A target
859 for heavy metal toxicity and a probiotic protective strategy. Sci Total Environ, 742, 140429.
860 DUIZER, E., GILDE, A. J., VERSANTVOORT, C. H. & GROTEN, J. P. 1999. Effects of
861 cadmium chloride on the paracellular barrier function of intestinal epithelial cell lines.
862 Toxicol Appl Pharmacol, 155, 117-26.
863 ECKBURG, P. B., BIK, E. M., BERNSTEIN, C. N., PURDOM, E., DETHLEFSEN, L.,
864 SARGENT, M., GILL, S. R., NELSON, K. E. & RELMAN, D. A. 2005. Diversity of the
865 human intestinal microbial flora. Science, 308, 1635-8.
866 EGGERS, S., SAFDAR, N., SETHI, A. K., SUEN, G., PEPPARD, P. E., KATES, A. E.,
867 SKARLUPKA, J. H., KANAREK, M. & MALECKI, K. M. C. 2019. Urinary lead
868 concentration and composition of the adult gut microbiota in a cross-sectional population-
869 based sample. Environ Int, 133, 105122.
870 EY, B., EYKING, A., GERKEN, G., PODOLSKY, D. K. & CARIO, E. 2009. TLR2 mediates gap
871 junctional intercellular communication through connexin-43 in intestinal epithelial barrier
872 injury. J Biol Chem, 284, 22332-43.
Page 29 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
30
873 FARKHONDEH, T., SAMARGHANDIAN, S. & AZIMI-NEZHAD, M. 2019. The role of arsenic
874 in obesity and diabetes. J Cell Physiol, 234, 12516-12529.
875 FENG, P., YE, Z., KAKADE, A., VIRK, A. K., LI, X. & LIU, P. 2018. A Review on Gut
876 Remediation of Selected Environmental Contaminants: Possible Roles of Probiotics and
877 Gut Microbiota. Nutrients, 11.
878 FENG, Y., HUANG, Y., WANG, Y., WANG, P., SONG, H. & WANG, F. 2019. Antibiotics
879 induced intestinal tight junction barrier dysfunction is associated with microbiota
880 dysbiosis, activated NLRP3 inflammasome and autophagy. PLoS One, 14, e0218384.
881 FERNÁNDEZ FERNÁNDEZ, N., ESTEVEZ BOULLOSA, P., GÓMEZ RODRÍGUEZ, A. &
882 RODRÍGUEZ PRADA, J. I. 2019. A Rare Cause of Gastric Injury: Arsenic Intake. Am J
883 Gastroenterol, 114, 1193.
884 FLORA, G., GUPTA, D. & TIWARI, A. 2012. Toxicity of lead: A review with recent updates.
885 Interdiscip Toxicol, 5, 47-58.
886 FURUSAWA, Y., OBATA, Y., FUKUDA, S., ENDO, T. A., NAKATO, G., TAKAHASHI, D.,
887 NAKANISHI, Y., UETAKE, C., KATO, K., KATO, T., TAKAHASHI, M., FUKUDA,
888 N. N., MURAKAMI, S., MIYAUCHI, E., HINO, S., ATARASHI, K., ONAWA, S.,
889 FUJIMURA, Y., LOCKETT, T., CLARKE, J. M., TOPPING, D. L., TOMITA, M., HORI,
890 S., OHARA, O., MORITA, T., KOSEKI, H., KIKUCHI, J., HONDA, K., HASE, K. &
891 OHNO, H. 2013. Commensal microbe-derived butyrate induces the differentiation of
892 colonic regulatory T cells. Nature, 504, 446-50.
893 GAO, B., CHI, L., MAHBUB, R., BIAN, X., TU, P., RU, H. & LU, K. 2017. Multi-Omics Reveals
894 that Lead Exposure Disturbs Gut Microbiome Development, Key Metabolites, and
895 Metabolic Pathways. Chem Res Toxicol, 30, 996-1005.
896 GARZA-LOMBO, C., PAPPA, A., PANAYIOTIDIS, M. I., GONSEBATT, M. E. & FRANCO,
897 R. 2019. Arsenic-induced neurotoxicity: a mechanistic appraisal. J Biol Inorg Chem, 24,
898 1305-1316.
899 GEORGE, F., MAHIEUX, S., DANIEL, C., TITECAT, M., BEAUVAL, N., HOUCKE, I., NEUT,
900 C., ALLORGE, D., BORGES, F., JAN, G., FOLIGNE, B. & GARAT, A. 2021.
901 Assessment of Pb(II), Cd(II), and Al(III) Removal Capacity of Bacteria from Food and Gut
902 Ecological Niches: Insights into Biodiversity to Limit Intestinal Biodisponibility of Toxic
903 Metals. Microorganisms, 9.
904 GHOSH, S., BANERJEE, M., BODDULURI, H. & JALA, V. R. 2022a. Microbial metabolite
905 mitigates arsenic induced oxidative stress, inflammation, and barrier dysfunction in gut
906 epithelia. The FASEB Journal, 36.
907 GHOSH, S., BANERJEE, M., HARIBABU, B. & JALA, V. R. 2022b. Urolithin A attenuates
908 arsenic-induced gut barrier dysfunction. Arch Toxicol.
909 GHOSH, S., MOORTHY, B., HARIBABU, B. & JALA, V. R. 2022c. Cytochrome P450 1A1 is
910 essential for the microbial metabolite, Urolithin A-mediated protection against colitis.
911 Frontiers in Immunology, 13.
912 GHOSH, S., SINGH, R., VANWINKLE, Z. M., GUO, H., VEMULA, P. K., GOEL, A.,
913 HARIBABU, B. & JALA, V. R. 2022d. Microbial metabolite restricts 5-fluorouracil-
914 resistant colonic tumor progression by sensitizing drug transporters via regulation of
915 FOXO3-FOXM1 axis. Theranostics, 12, 5574-5595.
916 GHOSH, S., WHITLEY, C. S., HARIBABU, B. & JALA, V. R. 2021. Regulation of Intestinal
917 Barrier Function by Microbial Metabolites. Cell Mol Gastroenterol Hepatol, 11, 1463-
918 1482.
Page 30 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
31
919 GIBSON, G. R. & ROBERFROID, M. B. 1995. Dietary modulation of the human colonic
920 microbiota: introducing the concept of prebiotics. J Nutr, 125, 1401-12.
921 GILLOIS, K., LEVEQUE, M., THEODOROU, V., ROBERT, H. & MERCIER-BONIN, M. 2018.
922 Mucus: An Underestimated Gut Target for Environmental Pollutants and Food Additives.
923 Microorganisms, 6.
924 GONZÁLEZ-MARISCAL, L., BETANZOS, A., NAVA, P. & JARAMILLO, B. E. 2003. Tight
925 junction proteins. Prog Biophys Mol Biol, 81, 1-44.
926 GOODENOUGH, D. A., GOLIGER, J. A. & PAUL, D. L. 1996. Connexins, connexons, and
927 intercellular communication. Annu Rev Biochem, 65, 475-502.
928 GOODENOUGH, D. A. & PAUL, D. L. 2009. Gap junctions. Cold Spring Harb Perspect Biol, 1,
929 a002576.
930 GRAU-PEREZ, M., KUO, C. C., GRIBBLE, M. O., BALAKRISHNAN, P., JONES SPRATLEN,
931 M., VAIDYA, D., FRANCESCONI, K. A., GOESSLER, W., GUALLAR, E.,
932 SILBERGELD, E. K., UMANS, J. G., BEST, L. G., LEE, E. T., HOWARD, B. V., COLE,
933 S. A. & NAVAS-ACIEN, A. 2017. Association of Low-Moderate Arsenic Exposure and
934 Arsenic Metabolism with Incident Diabetes and Insulin Resistance in the Strong Heart
935 Family Study. Environ Health Perspect, 125, 127004.
936 GREEN, K. J. & SIMPSON, C. L. 2007. Desmosomes: new perspectives on a classic. J Invest
937 Dermatol, 127, 2499-515.
938 GUAN, Q. 2019. A Comprehensive Review and Update on the Pathogenesis of Inflammatory
939 Bowel Disease. J Immunol Res, 2019, 7247238.
940 GUHA MAZUMDER, D. & DASGUPTA, U. B. 2011. Chronic arsenic toxicity: studies in West
941 Bengal, India. Kaohsiung J Med Sci, 27, 360-70.
942 GUMBINER, B. M. 1996. Cell adhesion: the molecular basis of tissue architecture and
943 morphogenesis. Cell, 84, 345-57.
944 GÜNZEL, D. & YU, A. S. 2013. Claudins and the modulation of tight junction permeability.
945 Physiol Rev, 93, 525-69.
946 HALBLEIB, J. M. & NELSON, W. J. 2006. Cadherins in development: cell adhesion, sorting, and
947 tissue morphogenesis. Genes Dev, 20, 3199-214.
948 HAO, W., HAO, C., WU, C., XU, Y. & JIN, C. 2022. Aluminum induced intestinal dysfunction
949 via mechanical, immune, chemical and biological barriers. Chemosphere, 288, 132556.
950 HARTSOCK, A. & NELSON, W. J. 2008. Adherens and tight junctions: structure, function and
951 connections to the actin cytoskeleton. Biochim Biophys Acta, 1778, 660-9.
952 HAYES, C. L., DONG, J., GALIPEAU, H. J., JURY, J., MCCARVILLE, J., HUANG, X.,
953 WANG, X. Y., NAIDOO, A., ANBAZHAGAN, A. N., LIBERTUCCI, J., SHERIDAN,
954 C., DUDEJA, P. K., BOWDISH, D. M. E., SURETTE, M. G. & VERDU, E. F. 2018.
955 Commensal microbiota induces colonic barrier structure and functions that contribute to
956 homeostasis. Sci Rep, 8, 14184.
957 HERNÁNDEZ-CHIRLAQUE, C., ARANDA, C. J., OCÓN, B., CAPITÁN-CAÑADAS, F.,
958 ORTEGA-GONZÁLEZ, M., CARRERO, J. J., SUÁREZ, M. D., ZARZUELO, A.,
959 SÁNCHEZ DE MEDINA, F. & MARTÍNEZ-AUGUSTIN, O. 2016. Germ-free and
960 Antibiotic-treated Mice are Highly Susceptible to Epithelial Injury in DSS Colitis. J
961 Crohns Colitis, 10, 1324-1335.
962 HONG, Y. S., SONG, K. H. & CHUNG, J. Y. 2014. Health effects of chronic arsenic exposure. J
963 Prev Med Public Health, 47, 245-52.
Page 31 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
32
964 HUNT, K. M., SRIVASTAVA, R. K., ELMETS, C. A. & ATHAR, M. 2014. The mechanistic
965 basis of arsenicosis: pathogenesis of skin cancer. Cancer Lett, 354, 211-9.
966 IARC. 2012. Special Report: Policy, A review of Human Carcinogens—Part C: Metals, Arsenic,
967 Dusts, and Fibres. IARC Monogr Eval Carcinog Risks Hum.
968 JANDHYALA, S. M., TALUKDAR, R., SUBRAMANYAM, C., VUYYURU, H., SASIKALA,
969 M. & NAGESHWAR REDDY, D. 2015. Role of the normal gut microbiota. World J
970 Gastroenterol, 21, 8787-803.
971 JIANG, X., GU, S., LIU, D., ZHAO, L., XIA, S., HE, X., CHEN, H. & GE, J. 2018. Lactobacillus
972 brevis 23017 Relieves Mercury Toxicity in the Colon by Modulation of Oxidative Stress
973 and Inflammation Through the Interplay of MAPK and NF-kappaB Signaling Cascades.
974 Front Microbiol, 9, 2425.
975 JOHANSSON, M. E., JAKOBSSON, H. E., HOLMEN-LARSSON, J., SCHUTTE, A.,
976 ERMUND, A., RODRIGUEZ-PINEIRO, A. M., ARIKE, L., WISING, C., SVENSSON,
977 F., BACKHED, F. & HANSSON, G. C. 2015. Normalization of Host Intestinal Mucus
978 Layers Requires Long-Term Microbial Colonization. Cell Host Microbe, 18, 582-92.
979 JOHANSSON, M. E., SJÖVALL, H. & HANSSON, G. C. 2013. The gastrointestinal mucus
980 system in health and disease. Nat Rev Gastroenterol Hepatol, 10, 352-61.
981 JOMOVA, K., JENISOVA, Z., FESZTEROVA, M., BAROS, S., LISKA, J., HUDECOVA, D.,
982 RHODES, C. J. & VALKO, M. 2011. Arsenic: toxicity, oxidative stress and human
983 disease. J Appl Toxicol, 31, 95-107.
984 KAYAMA, H. & TAKEDA, K. 2020. Manipulation of epithelial integrity and mucosal immunity
985 by host and microbiota-derived metabolites. Eur J Immunol, 50, 921-931.
986 KIM, Y. S. & HO, S. B. 2010. Intestinal goblet cells and mucins in health and disease: recent
987 insights and progress. Curr Gastroenterol Rep, 12, 319-30.
988 KONIECZNA, P., FERSTL, R., ZIEGLER, M., FREI, R., NEHRBASS, D., LAUENER, R. P.,
989 AKDIS, C. A. & O'MAHONY, L. 2013. Immunomodulation by Bifidobacterium infantis
990 35624 in the murine lamina propria requires retinoic acid-dependent and independent
991 mechanisms. PLoS One, 8, e62617.
992 KRUGER, M. C., BERTIN, P. N., HEIPIEPER, H. J. & ARSÈNE-PLOETZE, F. 2013. Bacterial
993 metabolism of environmental arsenic--mechanisms and biotechnological applications.
994 Appl Microbiol Biotechnol, 97, 3827-41.
995 KUCHARZIK, T., LÜGERING, A., LÜGERING, N., RAUTENBERG, K., LINNEPE, M.,
996 CICHON, C., REICHELT, R., STOLL, R., SCHMIDT, M. A. & DOMSCHKE, W. 2000a.
997 Characterization of M cell development during indomethacin-induced ileitis in rats.
998 Aliment Pharmacol Ther, 14, 247-56.
999 KUCHARZIK, T., LÜGERING, N., RAUTENBERG, K., LÜGERING, A., SCHMIDT, M. A.,
1000 STOLL, R. & DOMSCHKE, W. 2000b. Role of M cells in intestinal barrier function. Ann
1001 N Y Acad Sci, 915, 171-83.
1002 LANPHEAR, B. P., RAUCH, S., AUINGER, P., ALLEN, R. W. & HORNUNG, R. W. 2018.
1003 Low-level lead exposure and mortality in US adults: a population-based cohort study.
1004 Lancet Public Health, 3, e177-e184.
1005 LEAL, J., SMYTH, H. D. C. & GHOSH, D. 2017. Physicochemical properties of mucus and their
1006 impact on transmucosal drug delivery. Int J Pharm, 532, 555-572.
1007 LEE, J. S., TATO, C. M., JOYCE-SHAIKH, B., GULEN, M. F., CAYATTE, C., CHEN, Y.,
1008 BLUMENSCHEIN, W. M., JUDO, M., AYANOGLU, G., MCCLANAHAN, T. K., LI, X.
Page 32 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
33
1009 & CUA, D. J. 2015. Interleukin-23-Independent IL-17 Production Regulates Intestinal
1010 Epithelial Permeability. Immunity, 43, 727-38.
1011 LI, A., DING, J., SHEN, T., HAN, Z., ZHANG, J., ABADEEN, Z. U., KULYAR, M. F., WANG,
1012 X. & LI, K. 2021. Environmental hexavalent chromium exposure induces gut microbial
1013 dysbiosis in chickens. Ecotoxicol Environ Saf, 227, 112871.
1014 LI, A., WANG, Y., HAO, J., WANG, L., QUAN, L., DUAN, K., FAKHAR, E. A. K. M., ULLAH,
1015 K., ZHANG, J., WU, Y. & LI, K. 2022. Long-term hexavalent chromium exposure disturbs
1016 the gut microbial homeostasis of chickens. Ecotoxicol Environ Saf, 237, 113532.
1017 LI, X., BREJNROD, A. D., ERNST, M., RYKAER, M., HERSCHEND, J., OLSEN, N. M. C.,
1018 DORRESTEIN, P. C., RENSING, C. & SORENSEN, S. J. 2019. Heavy metal exposure
1019 causes changes in the metabolic health-associated gut microbiome and metabolites.
1020 Environ Int, 126, 454-467.
1021 LI, X., LIU, L., CAO, Z., LI, W., LI, H., LU, C., YANG, X. & LIU, Y. 2020. Gut microbiota as
1022 an "invisible organ" that modulates the function of drugs. Biomed Pharmacother, 121,
1023 109653.
1024 LINDELL, A. E., ZIMMERMANN-KOGADEEVA, M. & PATIL, K. R. 2022. Multimodal
1025 interactions of drugs, natural compounds and pollutants with the gut microbiota. Nat Rev
1026 Microbiol.
1027 LIU, S., KANG, W., MAO, X., GE, L., DU, H., LI, J., HOU, L., LIU, D., YIN, Y., LIU, Y. &
1028 HUANG, K. 2022a. Melatonin mitigates aflatoxin B1-induced liver injury via modulation
1029 of gut microbiota/intestinal FXR/liver TLR4 signaling axis in mice. J Pineal Res, 73,
1030 e12812.
1031 LIU, W., FENG, H., ZHENG, S., XU, S., MASSEY, I. Y., ZHANG, C., WANG, X. & YANG, F.
1032 2021. Pb Toxicity on Gut Physiology and Microbiota. Front Physiol, 12, 574913.
1033 LIU, X., WANG, J., DENG, H., ZHONG, X., LI, C., LUO, Y., CHEN, L., ZHANG, B., WANG,
1034 D., HUANG, Y., ZHANG, J. & GUO, L. 2022b. In situ analysis of variations of arsenicals,
1035 microbiome and transcriptome profiles along murine intestinal tract. J Hazard Mater, 427,
1036 127899.
1037 LIU, Y., KANG, W., LIU, S., LI, J., LIU, J., CHEN, X., GAN, F. & HUANG, K. 2022c. Gut
1038 microbiota-bile acid-intestinal Farnesoid X receptor signaling axis orchestrates cadmium-
1039 induced liver injury. Sci Total Environ, 849, 157861.
1040 LIU, Y., LI, Y., LIU, K. & SHEN, J. 2014. Exposing to cadmium stress cause profound toxic
1041 effect on microbiota of the mice intestinal tract. PLoS One, 9, e85323.
1042 LIU, Y., LI, Y., XIA, Y., LIU, K., REN, L. & JI, Y. 2020a. The Dysbiosis of Gut Microbiota
1043 Caused by Low-Dose Cadmium Aggravate the Injury of Mice Liver through Increasing
1044 Intestinal Permeability. Microorganisms, 8.
1045 LIU, Y., WU, J., XIAO, Y., LIU, Q., YU, L., TIAN, F., ZHAO, J., ZHANG, H., CHEN, W. &
1046 ZHAI, Q. 2020b. Relief of Cadmium-Induced Intestinal Motility Disorder in Mice by
1047 Lactobacillus plantarum CCFM8610. Front Immunol, 11, 619574.
1048 LU, K., CABLE, P. H., ABO, R. P., RU, H., GRAFFAM, M. E., SCHLIEPER, K. A., PARRY,
1049 N. M., LEVINE, S., BODNAR, W. M., WISHNOK, J. S., STYBLO, M., SWENBERG, J.
1050 A., FOX, J. G. & TANNENBAUM, S. R. 2013. Gut microbiome perturbations induced by
1051 bacterial infection affect arsenic biotransformation. Chem Res Toxicol, 26, 1893-903.
1052 LU, K., MAHBUB, R., CABLE, P. H., RU, H., PARRY, N. M., BODNAR, W. M., WISHNOK,
1053 J. S., STYBLO, M., SWENBERG, J. A., FOX, J. G. & TANNENBAUM, S. R. 2014. Gut
Page 33 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
34
1054 microbiome phenotypes driven by host genetics affect arsenic metabolism. Chem Res
1055 Toxicol, 27, 172-4.
1056 LUESCHOW, S. R. & MCELROY, S. J. 2020. The Paneth Cell: The Curator and Defender of the
1057 Immature Small Intestine. Front Immunol, 11, 587.
1058 LUESCHOW, S. R., STUMPHY, J., GONG, H., KERN, S. L., ELGIN, T. G., UNDERWOOD,
1059 M. A., KALANETRA, K. M., MILLS, D. A., WONG, M. H., MEYERHOLZ, D. K.,
1060 GOOD, M. & MCELROY, S. J. 2018. Loss of murine Paneth cell function alters the
1061 immature intestinal microbiome and mimics changes seen in neonatal necrotizing
1062 enterocolitis. PLoS One, 13, e0204967.
1063 MADDEN, K. B., WHITMAN, L., SULLIVAN, C., GAUSE, W. C., URBAN, J. F., JR.,
1064 KATONA, I. M., FINKELMAN, F. D. & SHEA-DONOHUE, T. 2002. Role of STAT6
1065 and mast cells in IL-4- and IL-13-induced alterations in murine intestinal epithelial cell
1066 function. J Immunol, 169, 4417-22.
1067 MARTINI, E., KRUG, S. M., SIEGMUND, B., NEURATH, M. F. & BECKER, C. 2017. Mend
1068 Your Fences: The Epithelial Barrier and its Relationship With Mucosal Immunity in
1069 Inflammatory Bowel Disease. Cell Mol Gastroenterol Hepatol, 4, 33-46.
1070 MAZMANIAN, S. K., LIU, C. H., TZIANABOS, A. O. & KASPER, D. L. 2005. An
1071 immunomodulatory molecule of symbiotic bacteria directs maturation of the host immune
1072 system. Cell, 122, 107-18.
1073 MCELROY, S. J., UNDERWOOD, M. A. & SHERMAN, M. P. 2013. Paneth cells and necrotizing
1074 enterocolitis: a novel hypothesis for disease pathogenesis. Neonatology, 103, 10-20.
1075 MEHTA, S., NIJHUIS, A., KUMAGAI, T., LINDSAY, J. & SILVER, A. 2015. Defects in the
1076 adherens junction complex (E-cadherin/ beta-catenin) in inflammatory bowel disease. Cell
1077 Tissue Res, 360, 749-60.
1078 MIR, H., MEENA, A. S., CHAUDHRY, K. K., SHUKLA, P. K., GANGWAR, R., MANDA, B.,
1079 PADALA, M. K., SHEN, L., TURNER, J. R., DIETRICH, P., DRAGATSIS, I. & RAO,
1080 R. 2016. Occludin deficiency promotes ethanol-induced disruption of colonic epithelial
1081 junctions, gut barrier dysfunction and liver damage in mice. Biochim Biophys Acta, 1860,
1082 765-74.
1083 MITAMURA, Y., OGULUR, I., PAT, Y., RINALDI, A. O., ARDICLI, O., CEVHERTAS, L.,
1084 BRUGGEN, M. C., TRAIDL-HOFFMANN, C., AKDIS, M. & AKDIS, C. A. 2021.
1085 Dysregulation of the epithelial barrier by environmental and other exogenous factors.
1086 Contact Dermatitis, 85, 615-626.
1087 MOON, K., GUALLAR, E. & NAVAS-ACIEN, A. 2012. Arsenic exposure and cardiovascular
1088 disease: an updated systematic review. Curr Atheroscler Rep, 14, 542-55.
1089 MOON, K. A., OBEROI, S., BARCHOWSKY, A., CHEN, Y., GUALLAR, E., NACHMAN, K.
1090 E., RAHMAN, M., SOHEL, N., D'IPPOLITI, D., WADE, T. J., JAMES, K. A., FARZAN,
1091 S. F., KARAGAS, M. R., AHSAN, H. & NAVAS-ACIEN, A. 2018. A dose-response
1092 meta-analysis of chronic arsenic exposure and incident cardiovascular disease. Int J
1093 Epidemiol, 47, 1013.
1094 MORITA, K., FURUSE, M., FUJIMOTO, K. & TSUKITA, S. 1999. Claudin multigene family
1095 encoding four-transmembrane domain protein components of tight junction strands. Proc
1096 Natl Acad Sci U S A, 96, 511-6.
1097 MOTTA, C. M., CALIFANO, E., SCUDIERO, R., AVALLONE, B., FOGLIANO, C., DE
1098 BONIS, S., RAGGIO, A. & SIMONIELLO, P. 2022. Effects of Cadmium Exposure on
1099 Gut Villi in Danio rerio. Int J Mol Sci, 23.
Page 34 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
35
1100 NAUJOKAS, M. F., ANDERSON, B., AHSAN, H., APOSHIAN, H. V., GRAZIANO, J. H.,
1101 THOMPSON, C. & SUK, W. A. 2013. The broad scope of health effects from chronic
1102 arsenic exposure: update on a worldwide public health problem. Environ Health Perspect,
1103 121, 295-302.
1104 NAVAS-ACIEN, A., SILBERGELD, E. K., PASTOR-BARRIUSO, R. & GUALLAR, E. 2008.
1105 Arsenic exposure and prevalence of type 2 diabetes in US adults. JAMA, 300, 814-22.
1106 NEKRASOVA, O. & GREEN, K. J. 2013. Desmosome assembly and dynamics. Trends Cell Biol,
1107 23, 537-46.
1108 NINKOV, M., POPOV ALEKSANDROV, A., DEMENESKU, J., MIRKOV, I., MILEUSNIC,
1109 D., PETROVIC, A., GRIGOROV, I., ZOLOTAREVSKI, L., TOLINACKI, M.,
1110 KATARANOVSKI, D., BRCESKI, I. & KATARANOVSKI, M. 2015. Toxicity of oral
1111 cadmium intake: Impact on gut immunity. Toxicol Lett, 237, 89-99.
1112 ODENWALD, M. A. & TURNER, J. R. 2013. Intestinal permeability defects: is it time to treat?
1113 Clin Gastroenterol Hepatol, 11, 1075-83.
1114 OKUMURA, R. & TAKEDA, K. 2017. Roles of intestinal epithelial cells in the maintenance of
1115 gut homeostasis. Experimental & Molecular Medicine, 49, e338-e338.
1116 PALMER, C., BIK, E. M., DIGIULIO, D. B., RELMAN, D. A. & BROWN, P. O. 2007.
1117 Development of the human infant intestinal microbiota. PLoS Biol, 5, e177.
1118 PARKER, A., LAWSON, M. A. E., VAUX, L. & PIN, C. 2018. Host-microbe interaction in the
1119 gastrointestinal tract. Environ Microbiol, 20, 2337-2353.
1120 PELASEYED, T., BERGSTRÖM, J. H., GUSTAFSSON, J. K., ERMUND, A.,
1121 BIRCHENOUGH, G. M., SCHÜTTE, A., VAN DER POST, S., SVENSSON, F.,
1122 RODRÍGUEZ-PIÑEIRO, A. M., NYSTRÖM, E. E., WISING, C., JOHANSSON, M. E.
1123 & HANSSON, G. C. 2014. The mucus and mucins of the goblet cells and enterocytes
1124 provide the first defense line of the gastrointestinal tract and interact with the immune
1125 system. Immunol Rev, 260, 8-20.
1126 PEREZ-MORENO, M. & FUCHS, E. 2006. Catenins: keeping cells from getting their signals
1127 crossed. Dev Cell, 11, 601-12.
1128 PEREZ-MORENO, M., JAMORA, C. & FUCHS, E. 2003. Sticky business: orchestrating cellular
1129 signals at adherens junctions. Cell, 112, 535-48.
1130 PETERSON, L. W. & ARTIS, D. 2014. Intestinal epithelial cells: regulators of barrier function
1131 and immune homeostasis. Nature Reviews Immunology, 14, 141-153.
1132 PINTO, D. V., RAPOSO, R. S., MATOS, G. A., ALVAREZ-LEITE, J. I., MALVA, J. O. &
1133 ORIA, R. B. 2020. Methylmercury Interactions With Gut Microbiota and Potential
1134 Modulation of Neurogenic Niches in the Brain. Front Neurosci, 14, 576543.
1135 PODGORSKI, J. & BERG, M. 2020. Global threat of arsenic in groundwater. Science, 368, 845-
1136 850.
1137 PYNN, L. Harvard study concludes elevated rates of Colitis/Crohn’s in Northport [Online].
1138 Available: https://northportproject.com/2013/11/29/northport-the-town-that-could-help-
1139 cure-ibd-2/ [Accessed].
1140 RAFATI RAHIMZADEH, M., RAFATI RAHIMZADEH, M., KAZEMI, S. &
1141 MOGHADAMNIA, A. A. 2017. Cadmium toxicity and treatment: An update. Caspian J
1142 Intern Med, 8, 135-145.
1143 RAO, R. 2008. Oxidative stress-induced disruption of epithelial and endothelial tight junctions.
1144 Front Biosci, 13, 7210-26.
Page 35 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
36
1145 RAO, R. K., BASUROY, S., RAO, V. U., KARNAKY, K. J., JR. & GUPTA, A. 2002. Tyrosine
1146 phosphorylation and dissociation of occludin-ZO-1 and E-cadherin-beta-catenin
1147 complexes from the cytoskeleton by oxidative stress. Biochem J, 368, 471-81.
1148 RICE, K. M., WALKER, E. M., JR., WU, M., GILLETTE, C. & BLOUGH, E. R. 2014.
1149 Environmental mercury and its toxic effects. J Prev Med Public Health, 47, 74-83.
1150 RUSANOV, A. L., SMIRNOVA, A. V., POROMOV, A. A., FOMICHEVA, K. A., LUZGINA,
1151 N. G. & MAJOUGA, A. G. 2015. Effects of cadmium chloride on the functional state of
1152 human intestinal cells. Toxicol In Vitro, 29, 1006-11.
1153 SAMPATH, V., BHANDARI, V., BERGER, J., MERCHANT, D., ZHANG, L., LADD, M.,
1154 MENDEN, H., GARLAND, J., AMBALAVANAN, N., MULROONEY, N., QUASNEY,
1155 M., DAGLE, J., LAVOIE, P. M., SIMPSON, P. & DAHMER, M. 2017. A functional
1156 ATG16L1 (T300A) variant is associated with necrotizing enterocolitis in premature
1157 infants. Pediatr Res, 81, 582-588.
1158 SCHMIDT, T. S. B., RAES, J. & BORK, P. 2018. The Human Gut Microbiome: From Association
1159 to Modulation. Cell, 172, 1198-1215.
1160 SCHREZENMEIR, J. & DE VRESE, M. 2001. Probiotics, prebiotics, and synbiotics--approaching
1161 a definition. Am J Clin Nutr, 73, 361s-364s.
1162 SEKI, N., AKIYAMA, M., YAMAKAWA, H., HASE, K., KUMAGAI, Y. & KIM, Y. G. 2021.
1163 Adverse effects of methylmercury on gut bacteria and accelerated accumulation of mercury
1164 in organs due to disruption of gut microbiota. J Toxicol Sci, 46, 91-97.
1165 SHAO, M. & ZHU, Y. 2020. Long-term metal exposure changes gut microbiota of residents
1166 surrounding a mining and smelting area. Sci Rep, 10, 4453.
1167 SHARMA, P., BIHARI, V., AGARWAL, S. K., VERMA, V., KESAVACHANDRAN, C. N.,
1168 PANGTEY, B. S., MATHUR, N., SINGH, K. P., SRIVASTAVA, M. & GOEL, S. K.
1169 2012. Groundwater contaminated with hexavalent chromium [Cr (VI)]: a health survey and
1170 clinical examination of community inhabitants (Kanpur, India). PLoS One, 7, e47877.
1171 SHEN, L., BLACK, E. D., WITKOWSKI, E. D., LENCER, W. I., GUERRIERO, V.,
1172 SCHNEEBERGER, E. E. & TURNER, J. R. 2006. Myosin light chain phosphorylation
1173 regulates barrier function by remodeling tight junction structure. J Cell Sci, 119, 2095-106.
1174 SHRIVASTAVA, R., KANNAN, A., UPRETI, R. K. & CHATURVEDI, U. C. 2005. Effects of
1175 chromium on the resident gut bacteria of rat. Toxicol Mech Methods, 15, 211-8.
1176 SNOECK, V., GODDEERIS, B. & COX, E. 2005. The role of enterocytes in the intestinal barrier
1177 function and antigen uptake. Microbes Infect, 7, 997-1004.
1178 SPINDLER, V., MEIR, M., VIGH, B., FLEMMING, S., HUTZ, K., GERMER, C. T.,
1179 WASCHKE, J. & SCHLEGEL, N. 2015. Loss of Desmoglein 2 Contributes to the
1180 Pathogenesis of Crohn's Disease. Inflamm Bowel Dis, 21, 2349-59.
1181 STERN, M. & WALKER, W. A. 1984. Food proteins and gut mucosal barrier I. Binding and
1182 uptake of cow's milk proteins by adult rat jejunum in vitro. Am J Physiol, 246, G556-62.
1183 SUMMERS, A. O., WIREMAN, J., VIMY, M. J., LORSCHEIDER, F. L., MARSHALL, B.,
1184 LEVY, S. B., BENNETT, S. & BILLARD, L. 1993. Mercury released from dental "silver"
1185 fillings provokes an increase in mercury- and antibiotic-resistant bacteria in oral and
1186 intestinal floras of primates. Antimicrob Agents Chemother, 37, 825-34.
1187 TAN, S., XU, X., CHENG, H., WANG, J. & WANG, X. 2022. The alteration of gut microbiome
1188 community play an important role in mercury biotransformation in largemouth bass.
1189 Environ Res, 204, 112026.
Page 36 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
37
1190 TANG, R., LI, X., MO, Y., MA, Y., DING, C., WANG, J., ZHANG, T. & WANG, X. 2019. Toxic
1191 responses of metabolites, organelles and gut microorganisms of Eisenia fetida in a soil with
1192 chromium contamination. Environ Pollut, 251, 910-920.
1193 TCHOUNWOU, P. B., YEDJOU, C. G., PATLOLLA, A. K. & SUTTON, D. J. 2012. Heavy
1194 Metal Toxicity and the Environment. In: LUCH, A. (ed.) Molecular, Clinical and
1195 Environmental Toxicology: Volume 3: Environmental Toxicology. Basel: Springer Basel.
1196 TELLEZ-PLAZA, M., NAVAS-ACIEN, A., MENKE, A., CRAINICEANU, C. M., PASTOR-
1197 BARRIUSO, R. & GUALLAR, E. 2012. Cadmium exposure and all-cause and
1198 cardiovascular mortality in the U.S. general population. Environ Health Perspect, 120,
1199 1017-22.
1200 THOMPSON, C. M., PROCTOR, D. M., SUH, M., HAWS, L. C., KIRMAN, C. R. & HARRIS,
1201 M. A. 2013. Assessment of the mode of action underlying development of rodent small
1202 intestinal tumors following oral exposure to hexavalent chromium and relevance to
1203 humans. Crit Rev Toxicol, 43, 244-74.
1204 THORNTON, D. J. & SHEEHAN, J. K. 2004. From mucins to mucus: toward a more coherent
1205 understanding of this essential barrier. Proc Am Thorac Soc, 1, 54-61.
1206 THURSBY, E. & JUGE, N. 2017. Introduction to the human gut microbiota. Biochem J, 474,
1207 1823-1836.
1208 TING, H. A. & VON MOLTKE, J. 2019. The Immune Function of Tuft Cells at Gut Mucosal
1209 Surfaces and Beyond. Journal of Immunology, 202, 1321-1329.
1210 TINKOV, A. A., FILIPPINI, T., AJSUVAKOVA, O. P., AASETH, J., GLUHCHEVA, Y. G.,
1211 IVANOVA, J. M., BJORKLUND, G., SKALNAYA, M. G., GATIATULINA, E. R.,
1212 POPOVA, E. V., NEMERESHINA, O. N., VINCETI, M. & SKALNY, A. V. 2017. The
1213 role of cadmium in obesity and diabetes. Sci Total Environ, 601-602, 741-755.
1214 TINKOV, A. A., GRITSENKO, V. A., SKALNAYA, M. G., CHERKASOV, S. V., AASETH, J.
1215 & SKALNY, A. V. 2018. Gut as a target for cadmium toxicity. Environ Pollut, 235, 429-
1216 434.
1217 TSAI, S. L., SINGH, S. & CHEN, W. 2009. Arsenic metabolism by microbes in nature and the
1218 impact on arsenic remediation. Curr Opin Biotechnol, 20, 659-67.
1219 TURNER, J. R. 2000. Show me the pathway! Regulation of paracellular permeability by Na(+)-
1220 glucose cotransport. Adv Drug Deliv Rev, 41, 265-81.
1221 TURNER, J. R. 2009. Intestinal mucosal barrier function in health and disease. Nat Rev Immunol,
1222 9, 799-809.
1223 UNDERWOOD, M. A. 2012. Paneth cells and necrotizing enterocolitis. Gut Microbes, 3, 562-5.
1224 UNGEWISS, H., VIELMUTH, F., SUZUKI, S. T., MAISER, A., HARZ, H., LEONHARDT, H.,
1225 KUGELMANN, D., SCHLEGEL, N. & WASCHKE, J. 2017. Desmoglein 2 regulates the
1226 intestinal epithelial barrier via p38 mitogen-activated protein kinase. Sci Rep, 7, 6329.
1227 UPRETI, R. K., SINHA, V., MISHRA, R. & KANNAN, A. 2011. In vitro development of
1228 resistance to arsenite and chromium-VI in Lactobacilli strains as perspective attenuation of
1229 gastrointestinal disorder. J Environ Biol, 32, 325-32.
1230 URSELL, L. K., CLEMENTE, J. C., RIDEOUT, J. R., GEVERS, D., CAPORASO, J. G. &
1231 KNIGHT, R. 2012. The interpersonal and intrapersonal diversity of human-associated
1232 microbiota in key body sites. J Allergy Clin Immunol, 129, 1204-8.
1233 VAISHNAVA, S., BEHRENDT, C. L., ISMAIL, A. S., ECKMANN, L. & HOOPER, L. V. 2008.
1234 Paneth cells directly sense gut commensals and maintain homeostasis at the intestinal host-
1235 microbial interface. Proc Natl Acad Sci U S A, 105, 20858-63.
Page 37 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
38
1236 VAN ITALLIE, C. M. & ANDERSON, J. M. 2006. Claudins and epithelial paracellular transport.
1237 Annu Rev Physiol, 68, 403-29.
1238 VAN ITALLIE, C. M. & ANDERSON, J. M. 2014. Architecture of tight junctions and principles
1239 of molecular composition. Semin Cell Dev Biol, 36, 157-65.
1240 VANUYTSEL, T., TACK, J. & FARRE, R. 2021. The Role of Intestinal Permeability in
1241 Gastrointestinal Disorders and Current Methods of Evaluation. Front Nutr, 8, 717925.
1242 VAZQUEZ, M., VELEZ, D. & DEVESA, V. 2014. In vitro evaluation of inorganic mercury and
1243 methylmercury effects on the intestinal epithelium permeability. Food Chem Toxicol, 74,
1244 349-59.
1245 VERMETTE, D., HU, P., CANARIE, M. F., FUNARO, M., GLOVER, J. & PIERCE, R. W. 2018.
1246 Tight junction structure, function, and assessment in the critically ill: a systematic review.
1247 Intensive Care Medicine Experimental, 6, 37.
1248 VIGNAL, C., DESREUMAUX, P. & BODY-MALAPEL, M. 2016. Gut: An underestimated
1249 target organ for Aluminum. Morphologie, 100, 75-84.
1250 VON MOLTKE, J., JI, M., LIANG, H.-E. & LOCKSLEY, R. M. 2016. Tuft-cell-derived IL-25
1251 regulates an intestinal ILC2–epithelial response circuit. Nature, 529, 221-225.
1252 WANG, B., WU, C., CUI, L., WANG, H., LIU, Y. & CUI, W. 2022a. Dietary aluminium intake
1253 disrupts the overall structure of gut microbiota in Wistar rats. Food Sci Nutr, 10, 3574-
1254 3584.
1255 WANG, G., LI, X., ZHOU, Y., FENG, J. & ZHANG, M. 2022b. Effects of Dietary Chromium
1256 Picolinate on Gut Microbiota, Gastrointestinal Peptides, Glucose Homeostasis, and
1257 Performance of Heat-Stressed Broilers. Animals (Basel), 12.
1258 WANG, R., MONIRUZZAMAN, M., WONG, K. Y., WIID, P., HARDING, A., GIRI, R., TONG,
1259 W., CREAGH, J., BEGUN, J., MCGUCKIN, M. A. & HASNAIN, S. Z. 2021. Gut
1260 microbiota shape the inflammatory response in mice with an epithelial defect. Gut
1261 Microbes, 13, 1887720.
1262 WATSON, S. E., MCKINNEY, M. A., PINDO, M., BULL, M. J., ATWOOD, T. C., HAUFFE,
1263 H. C. & PERKINS, S. E. 2021. Diet-driven mercury contamination is associated with polar
1264 bear gut microbiota. Sci Rep, 11, 23372.
1265 WU, G., XIAO, X., FENG, P., XIE, F., YU, Z., YUAN, W., LIU, P. & LI, X. 2017. Gut
1266 remediation: a potential approach to reducing chromium accumulation using Lactobacillus
1267 plantarum TW1-1. Sci Rep, 7, 15000.
1268 WU, J., WEN, X. W., FAULK, C., BOEHNKE, K., ZHANG, H., DOLINOY, D. C. & XI, C.
1269 2016. Perinatal Lead Exposure Alters Gut Microbiota Composition and Results in Sex-
1270 specific Bodyweight Increases in Adult Mice. Toxicol Sci, 151, 324-33.
1271 XIA, J., JIN, C., PAN, Z., SUN, L., FU, Z. & JIN, Y. 2018. Chronic exposure to low concentrations
1272 of lead induces metabolic disorder and dysbiosis of the gut microbiota in mice. Sci Total
1273 Environ, 631-632, 439-448.
1274 XING, C., YANG, F., LIN, Y., SHAN, J., YI, X., ALI, F., ZHU, Y., WANG, C., ZHANG, C.,
1275 ZHUANG, Y., CAO, H. & HU, G. 2022. Hexavalent Chromium Exposure Induces
1276 Intestinal Barrier Damage via Activation of the NF-kappaB Signaling Pathway and NLRP3
1277 Inflammasome in Ducks. Front Immunol, 13, 952639.
1278 YANG, J., CHEN, W., SUN, Y., LIU, J. & ZHANG, W. 2021a. Effects of cadmium on organ
1279 function, gut microbiota and its metabolomics profile in adolescent rats. Ecotoxicol
1280 Environ Saf, 222, 112501.
Page 38 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
39
1281 YANG, T. T., LIU, Y., TAN, S., WANG, W. X. & WANG, X. 2021b. The role of intestinal
1282 microbiota of the marine fish (Acanthopagrus latus) in mercury biotransformation. Environ
1283 Pollut, 277, 116768.
1284 YIN, N., CAI, X., WANG, P., FENG, R., DU, H., FU, Y., SUN, G. & CUI, Y. 2022. Predictive
1285 capabilities of in vitro colon bioaccessibility for estimating in vivo relative bioavailability
1286 of arsenic from contaminated soils: Arsenic speciation and gut microbiota considerations.
1287 Sci Total Environ, 818, 151804.
1288 YOUNG, J. L., CAI, L. & STATES, J. C. 2018. Impact of prenatal arsenic exposure on chronic
1289 adult diseases. Syst Biol Reprod Med, 64, 469-483.
1290 YU, L., DUAN, H., KELLINGRAY, L., CEN, S., TIAN, F., ZHAO, J., ZHANG, H., GALL, G.
1291 L., MAYER, M. J., ZHAI, Q., CHEN, W. & NARBAD, A. 2021a. Lactobacillus
1292 plantarum-Mediated Regulation of Dietary Aluminum Induces Changes in the Human Gut
1293 Microbiota: an In Vitro Colonic Fermentation Study. Probiotics Antimicrob Proteins, 13,
1294 398-412.
1295 YU, L., YU, Y., XIAO, Y., TIAN, F., NARBAD, A., ZHAI, Q. & CHEN, W. 2021b. Lead-induced
1296 gut injuries and the dietary protective strategies: A review. Journal of Functional Foods,
1297 83, 104528.
1298 YU, Y., YANG, W., LI, Y. & CONG, Y. 2019. Enteroendocrine Cells: Sensing Gut Microbiota
1299 and Regulating Inflammatory Bowel Diseases. Inflammatory Bowel Diseases, 26, 11-20.
1300 ZHAI, Q., LI, T., YU, L., XIAO, Y., FENG, S., WU, J., ZHAO, J., ZHANG, H. & CHEN, W.
1301 2017. Effects of subchronic oral toxic metal exposure on the intestinal microbiota of mice.
1302 Sci Bull (Beijing), 62, 831-840.
1303 ZHAI, Q., QU, D., FENG, S., YU, Y., YU, L., TIAN, F., ZHAO, J., ZHANG, H. & CHEN, W.
1304 2019a. Oral Supplementation of Lead-Intolerant Intestinal Microbes Protects Against Lead
1305 (Pb) Toxicity in Mice. Front Microbiol, 10, 3161.
1306 ZHAI, Q., TIAN, F., ZHAO, J., ZHANG, H., NARBAD, A. & CHEN, W. 2016. Oral
1307 Administration of Probiotics Inhibits Absorption of the Heavy Metal Cadmium by
1308 Protecting the Intestinal Barrier. Appl Environ Microbiol, 82, 4429-40.
1309 ZHAI, Q., WANG, J., CEN, S., ZHAO, J., ZHANG, H., TIAN, F. & CHEN, W. 2019b.
1310 Modulation of the gut microbiota by a galactooligosaccharide protects against heavy metal
1311 lead accumulation in mice. Food Funct, 10, 3768-3781.
1312 ZHANG, A., MATSUSHITA, M., ZHANG, L., WANG, H., SHI, X., GU, H., XIA, Z. & CUI, J.
1313 Y. 2021. Cadmium exposure modulates the gut-liver axis in an Alzheimer's disease mouse
1314 model. Commun Biol, 4, 1398.
1315 ZHANG, Z., CAO, H., SONG, N., ZHANG, L., CAO, Y. & TAI, J. 2020. Long-term hexavalent
1316 chromium exposure facilitates colorectal cancer in mice associated with changes in gut
1317 microbiota composition. Food Chem Toxicol, 138, 111237.
1318 ZHAO, Y., SU, J. Q., YE, J., RENSING, C., TARDIF, S., ZHU, Y. G. & BRANDT, K. K. 2019.
1319 AsChip: A High-Throughput qPCR Chip for Comprehensive Profiling of Genes Linked to
1320 Microbial Cycling of Arsenic. Environ Sci Technol, 53, 798-807.
1321 ZHAO, Y., ZHOU, C., WU, C., GUO, X., HU, G., WU, Q., XU, Z., LI, G., CAO, H., LI, L.,
1322 LATIGO, V., LIU, P., CHENG, S. & LIU, P. 2020. Subchronic oral mercury caused
1323 intestinal injury and changed gut microbiota in mice. Sci Total Environ, 721, 137639.
1324 ZIHNI, C., MILLS, C., MATTER, K. & BALDA, M. S. 2016. Tight junctions: from simple
1325 barriers to multifunctional molecular gates. Nature Reviews Molecular Cell Biology, 17,
1326 564-580.
Page 39 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
40
1327 ZOLOTAREVSKY, Y., HECHT, G., KOUTSOURIS, A., GONZALEZ, D. E., QUAN, C., TOM,
1328 J., MRSNY, R. J. & TURNER, J. R. 2002. A membrane-permeant peptide that inhibits
1329 MLC kinase restores barrier function in in vitro models of intestinal disease.
1330 Gastroenterology, 123, 163-72.
1331 ZUO, L., KUO, W. T. & TURNER, J. R. 2020. Tight Junctions as Targets and Effectors of
1332 Mucosal Immune Homeostasis. Cell Mol Gastroenterol Hepatol, 10, 327-340.
1333
Page 40 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
Table 1 – Effect of heavy metals in host health and gut microbiota:
Heavy metal
Effect on health
Changes in Gut microbiota
Increase
Decrease
Arsenic
Pathological changes in skin; gut barrier dysfunction;
inflammation; carcinogenesis; microbial dysbiosis;
cardiovascular diseases
Bacteroidetes; Bifidobacterium;
Faecalibaculum; Enterobacteriaceae
Firmicutes; Gammaproteobacteria;
Enterobacteriaceae
Lead
Increased bodyweight; reduced MUC2, ZO-1,
Occludin, and Claudin-1; increased gut permeability;
elevated oxidative stress and inflammation;
dysregulated hepatic metabolism; microbial dysbiosis;
hepatotoxicity; nephrotoxicity
Bacteroidetes; Clostridiaceae;
Ruminococcus; Oscillibacter;
Parabacteroides; Desulfovibrionaceae;
ClostridiumXIVb; Barnesiella
Firmicutes; Proteobacteria; Turicibacter;
Akkermansia; Dehalobacterium;
Lactococcus; Enterorhabdus;
Caulobacterales
Mercury
Neurotoxicity; oxidative stress and inflammation;
leaky gut; mitochondrial dysfunction; increased lipid
peroxidation; altered calcium homeostasis
Firmicutes/Bacteroidetes ratio;
Akkermansia
Lactobacillus; Protobacteria;
Cadmium
Inflammation; microbial dysbiosis; intestinal damage;
junctional protein, mucus, and glycan distribution
alteration; hepatotoxicity; energy metabolism
dysregulation; endocrine disruption; genome
instability
Bacteroidetes;
Akkermansia muciniphila; Prevotella
spp; Escherichia coli_Shigella
Firmicutes; γ- Proteobacteria; A.
Mucinipbila; Clostridium cocleatum;
Lachnoclostridium
Chromium
Oxidative stress; cancer; GI distress;
microbial dysbiosis; DNA damage; lipid peroxidation;
liver toxicity
Bacteroidetes; Tenericutes; Prevotella;
Clostridiales; S24-7Actinobacteria
Firmicutes; Lachnospiraceae
Page 41 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
Figure 1: Effect of heavy metals on gut. Heavy metals such as Arsenic (As), Lead (Pb), Mercury (Hg),
Cadmium (Cd) and Chromium (Cr) leads to increased oxidative stress, altered gut microbial composition and
inflammation potentially leading to gut leakiness and gut barrier dysfunction.
645x645mm (236 x 236 DPI)
Page 42 of 43Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
Figure 2: Effect of heavy metals on human health. Environmental pollutants such as Arsenic (As), Lead (Pb),
Mercury (Hg), Cadmium (Cd) and Chromium (Cr) lead to several adverse health effects. Heavy metals-
induced loss of gut barrier integrity and microbial balance initiate inflammation and toxicity in host tissue
and organs. Consumption of healthy diets, treatment with beneficial gut microbiota or microbial metabolites
potentially mitigate the adverse effects of heavy metal toxicity and restores gut homeostasis.
645x534mm (236 x 236 DPI)
Page 43 of 43 Accepted Manuscript published as MAH-23-0015.R1. Accepted for publication: 12-Dec-2023
Copyright © 2023 the authors
Downloaded from Bioscientifica.com at 12/13/2023 12:52:18PM
via Open Access. This work is licensed under a Creative Commons Attribution
4.0 International License
http://creativecommons.org/licenses/by/4.0/deed.en_GB
