ArticlePDF Available

The neonicotinoid insecticide Clothianidin adversely affects immune signaling in a human cell line

Springer Nature
Scientific Reports
Authors:
  • IPSP-CNR Institute for Sustainable Plant Protection, National Research Council - Piazzale Enrico Fermi 1, 80055 Portici, Naples, Italy

Abstract and Figures

Clothianidin is a widely used neonicotinoid insecticide, which is a potent agonist of the nicotinic acetylcholine receptor in insects. This neurotoxic compound has a negative impact on insect immunity, as it down-regulates the activation of the transcription factor NF-κB. Given the evolutionary conserved role of NF-κB in the modulation of the immune response in the animal kingdom, here we want to assess any effect of Clothianidin on vertebrate defense barriers. In presence of this neonicotinoid insecticide, a pro-inflammatory challenge with LPS on the human monocytic cell line THP-1 results both in a reduced production of the cytokine TNF-α and in a down-regulation of a reporter gene under control of NF-κB promoter. This finding is corroborated by a significant impact of Clothianidin on the transcription levels of different immune genes, characterized by a core disruption of TRAF4 and TRAF6 that negatively influences NF-κB signaling. Moreover, exposure to Clothianidin concurrently induces a remarkable up-regulation of NGFR, which supports the occurrence of functional ties between the immune and nervous systems. These results suggest a potential risk of immunotoxicity that neonicotinoids may have on vertebrates, which needs to be carefully assessed at the organism level.
This content is subject to copyright. Terms and conditions apply.
1
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
www.nature.com/scientificreports
The neonicotinoid insecticide
Clothianidin adversely aects
immune signaling in a human cell
line
Gennaro Di Prisco, Marco Iannaccone, Flora Ianniello, Rosalba Ferrara, Emilio Caprio ,
Francesco Pennacchio & Rosanna Capparelli
Clothianidin is a widely used neonicotinoid insecticide, which is a potent agonist of the nicotinic
acetylcholine receptor in insects. This neurotoxic compound has a negative impact on insect immunity,
as it down-regulates the activation of the transcription factor NF-κB. Given the evolutionary conserved
role of NF-κB in the modulation of the immune response in the animal kingdom, here we want to assess
any eect of Clothianidin on vertebrate defense barriers. In presence of this neonicotinoid insecticide, a
pro-inammatory challenge with LPS on the human monocytic cell line THP-1 results both in a reduced
production of the cytokine TNF-α and in a down-regulation of a reporter gene under control of NF-κB
promoter. This nding is corroborated by a signicant impact of Clothianidin on the transcription levels
of dierent immune genes, characterized by a core disruption of TRAF4 and TRAF6 that negatively
inuences NF-κB signaling. Moreover, exposure to Clothianidin concurrently induces a remarkable up-
regulation of NGFR, which supports the occurrence of functional ties between the immune and nervous
systems. These results suggest a potential risk of immunotoxicity that neonicotinoids may have on
vertebrates, which needs to be carefully assessed at the organism level.
Neonicotinoids are among the most widely used insecticides in agriculture, which are eective at low dosage
and show poor anity for the nicotinic acetylcholine receptor of mammalian species1,2. e limited impact on
non-target higher animals is, however, challenged by a growing number of studies, which support a negative
eect of these systemic and persistent insecticides on several non-target organisms and ecosystem services3,4. In
particular, pollinators seem to be particularly aected. Indeed, in spite of the fact that acute lethal eects are rarely
observed5, there are a number of reports on sub-lethal eects, such as impaired honeybee learning or homing
behavior68, and a stronger impact on pollinators of various pathogens913. is latter eect is in part due to the
immunosuppressive action exerted by neonicotinoids14,15 which further exacerbates the negative impact that viral
pathogens and Varroa destructor have on honeybee defense barriers1619.
e molecular mechanism underlying the negative eect of the neonicotinoid Clothianidin on insect immune
response has been recently reported14. Basically, this insecticide is able to exert a negative eect on the activation
of the nuclear factor-κB (NF-κB) and of the downstream immune barriers, which promotes uncontrolled viral
replication in honeybees bearing covert infections14. Moreover, other immune responses controlled by this tran-
scription factor, both cellular and humoral, are down-regulated by neonicotinoids15, suggesting the occurrence of
a wider impact of these insecticides on immunity.
NF-κB has a central role in the immune response by animals20, and, therefore, any defense pathway, conserved
across distant evolutionary lineages, under control of this transcription factor could be inuenced by a shared
negative regulation of its activation. is could account for the proposed link between the use of neonicotinoids
and the increasing incidence of pathologies in dierent animal groups4,19. It does not require a leap of imagination
to speculate that neonicotinoids may have possible negative eects on human health, by similarly interfering with
the regulation of the immune system. is is a hypothesis that certainly merits to be investigated, as part of a more
Department of Agricultural Sciences, University of Napoli “Federico II” – Via Università 100, 80055 Portici, Napoli,
Italy. Gennaro Di Prisco and Marco Iannaccone contributed equally to this work. Correspondence and requests for
materials should be addressed to F.P. (email: f.pennacchio@unina.it) or R.C. (email: capparel@unina.it)
Received: 6 July 2017
Accepted: 18 September 2017
Published: xx xx xxxx
OPEN
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
2
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
comprehensive eort towards a thorough characterization of neonicotinoid impact on human health, which,
surprisingly, is still in its infancy21,22.
Here we contribute to ll this gap of knowledge, by focusing our attention on the impact of the neonicotinoid
Clothianidin on the human immune response, using an in vitro model system to characterize the eects that this
molecule has on gene expression prole upon immune challenge. is has been done by RNA sequencing in
the monocytic human cell line THP-1, as aected by Clothianidin exposure, and by studying how this latter can
inuence pro-inammatory cytokine release upon immune challenge.
Results
Clothianidin disrupts NF-κB signaling. Insect immune response is negatively modulated by
Clothianidin, which disrupts NF-κB signaling by up-regulating a negative modulator of this transcription factor14.
Because NF-κB signaling underpins the modulation of several immune reactions in animals, we wanted to assess
if this alteration induced by Clothianidin occurred also in humans. To test this hypothesis, we focused our atten-
tion on the eect of Clothianidin on the expression prole of the Tumor Necrosis Factor Alpha (TNF-α), a
pro-inammatory cytokine regulated by NF-κB via TLR-423, both at transcriptional and translational level, using
an immune cell line (THP-1), which expresses the nicotinic acetylcholine receptor24. Our data clearly indicate that
exposure to Clothianidin disrupts the LPS-mediated induction of TNF-α expression, both in terms of transcript
level (Fig.1a) (One-Way ANOVA: F = 148.09; df = 3; p < 0.001) and protein production (Fig.1b) (One-Way
ANOVA: F = 183.61; df = 3; p < 0.001). e experimental concentration of Clothianidin used (100 ng/ml) did not
have any cytotoxic eect on THP1 cells, as demonstrated by lactate dehydrogenase (LDH) release across a range
of dierent doses of this insecticide (Fig.S1). Collectively, these results demonstrate that Clothianidin inhibits
TNF-α expression, which is under NF-κB control.
To unequivocally demonstrate that Clothianidin exposure interferes with NF-κB activation, we stably trans-
fected the THP-1 cell line with lentiviral particles carrying a NF-κB-responsive luciferase-expressing reporter
gene (CignalLentiReporters, SABiosciences). e cells were incubated overnight, in presence or absence of
Clothianidin, at the same concentration indicated above, and then treated with LPS or le unchallenged. When
LPS challenge was performed in presence of Clothianidin, a signicant (One-Way ANOVA: F = 137.09; df = 3;
p < 0.001) inhibition of LPS-induced enhancement of the reporter gene expression was observed, indicating the
occurrence of a negative eect of this neonicotinoid insecticide on NF-κB signaling (Fig.2).
Clothianidin alters the transcriptome of an immune cell line. In order to identify the molecular
networks underlying the inhibition of NF-κB activation induced by Clothianidin, we performed a transcriptomic
analysis by RNA-Seq of the human THP-1 cell line exposed overnight to this neonicotinoid, at a concentration
of 100 ng/ml, the same used in the experiments described above. Aer trimming and quality control of the
obtained sequences, about 97% resulted as high quality reads (Supplementary TableS1), and were mapped against
Homo sapiens reference genome (Ensembl, GRCh38). Principal component analysis (PCA) was applied to the
dataset showing two distinct clusters, conrming replicate uniformity (Supplementary FigureS2). Dierential
expression analysis by false discovery rate (FDR) (P < 0.05) showed that 2,833 and 2,678 genes were signi-
cantly up- and down-regulated, respectively, in Clothianidin treated cells (Supplementary FigureS3a,b). To
select the most dierentially expressed genes, we applied a more stringent lter for Log2 fold change of >+1
or <−1, which allowed the identication of 36 genes up-regulated and 54 down-regulated; both categories
included immune genes under NF-κB transcriptional control, such as TNF receptor-associated factor 4 (TRAF4),
TNF receptor-associated factor 6 (TRAF6), Fork head box protein O4 (FOXO4), Interleukin-18-binding pro-
tein(IL18BP) and Interleukin-17 receptor (IL17R) (Fig.3). The concurrent up-regulation of the negative
Figure 1. Clothianidin inhibits TNF-α expression induced by LPS treatment. In human THP-1 cells TNF-α
transcription rate was measured by qRT-PCR (a) and TNF-α secreted protein in cell free supernatant by ELISA
(b), aer overnight incubation with Clothianidin (100 ng/ml), followed by LPS stimulation for 1 h (1 µg/ml), and
compared with values obtained in untreated cells or exposed to Clothianindin but le unchallenged. Data are
reported as mean ± SEM and are representative of 3 independent experiments, with 3 replicates each (One-Way
ANOVA, all p < 0.05).
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
3
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
modulator of NF-κB activation TRAF4 and the down-regulation ofTRAF6, exerting an opposite activity, well
account for the reduced expression of genes controlled by this transcription factor. e entire collection of raw
data is available on public database with BioProject Number PRJNA392257 (National Center of Biotechnology
Information, U.S. National Library).
To analyze the putative interactions among proteins encoded by genes found to be dierentially expressed and,
then, of particular functional importance to face the stress induced by Clothianidin exposure, we used the String
soware25. is approach allowed the identication of a network of interactions among nerve growth factor
receptor (NGFR), TRAF4 and TRAF6, which indicate how the core alteration of NF-κB signaling in the immune
cell line THP-1 induced by Clothianidin is associated with a transcriptional change related to neural functions.
To validate the RNA-Seq analysis, the expression of the immune-related genes, found to be markedly
modulated, was further assessed in an independent qRT-PCR experiment, where THP-1 cells were treated
with Clothianidin overnight, at the same concentration used for RNA-Seq analysis. TRAF4 and NGFR were
signicantly up-regulated (Student’s t test: TRAF4, t = 17.064, df = 4, P < 0.001; NGFR, t = 12.275, df = 2,
P = 0.007), while TRAF6, FOXO4, IL18BP and IL17R were down-regulated, as expected on the basis of RNA-Seq
analysis (Students t test: TRAF6, t = 5.438, df = 4, P = 0.006; FOXO4,t = 5.444, df = 4, P = 0.006; IL18BPt = 7.995,
df = 4, P = 0.001; IL17R,t = 8.976, df = 4, P = 0.001) (Fig.4).
Collectively, these results allow to conclude that exposure to Clothianidin of the human cell line THP-1
determines a negative modulation of NF-κB signaling, associated with an up-regulation of TRAF4 and a
down-regulation of TRAF6, a negative and a positive modulator, respectively, of NF-κB activation, which partly
account for the observed immunosuppressive eects.
Discussion
In this study we demonstrate that human THP-1 cells treated with the neonicotinoid insecticide Clothianidin
react to an inammatory stimulus by showing a lower expression of the cytokine TNF-α, due to a reduced activa-
tion of NF-κB, which controls its transcription. e negative impact of this neonicotinoid on NF-κB signaling has
been recently reported in insects, and thought to be one of the stress elements that can contribute to the reduced
ecacy of antiviral immune barriers controlling DWV replication in honeybees bearing covert infections of this
viral pathogen14. e observed similar eects of Clothianidin on immune responses by cells of organisms in phy-
logenetically distant lineages indicate the occurrence of conserved mechanisms of cross-modulation between the
nervous and immune system2628. e nervous and the immune systems are traditionally thought to be separate
functional entities and, as such, are separately studied. However, it is increasingly evident that this is not the case
and their intimate interaction is a fascinating research area that continuously generates novel information on the
subtle mechanisms involved28 and on their wide occurrence in the animal kingdom27. is conceptual framework
nicely accounts for the observed immunomodulation by the acetylcholine agonist Clothianidin, even though the
underlying molecular network that modulates this response remain still largely unexplored14. ese conserved
pathways of neuroimmune regulation and the fact that Clothianidin binds, even though with much lower anity
than in insects, to the human α4β2nicotinic acetylcholine receptor (α4β2AchR)29 were the major elements stim-
ulating the present study, aiming to discover any immunotoxic eect that neonicotinoids may have on vertebrates.
Using STRING software, we highlighted a tight interaction between TRAF4, TRAF6 and the NGFR
(Supplementary FigureS4). A previous study has reported that co-expression of NGFR with TRAF6 enhances
expression of NF-κB, while TRAF4 negatively interferes with this process30. is further reinforces the tight
relation between the nervous and immune systems. Indeed, the NGF in vertebrates can act as a homologue of the
y Toll ligand Spaetzle in eliciting immune reactions31, and its perception by THP-1 cells seems to be inuenced
Figure 2. Clothianidin inhibits the expression of a NF-κB responsive reporter gene. NF-κB induction as
aected by insecticide exposure was measured in human THP-1 cells, using a NF-κB luciferase reporter
aer incubation with Clothianidin overnight (100 ng/ml), followed by LPS stimulation for 1 h (1 µg/ml), and
compared with values obtained in untreated cells or exposed to Clothianidin but le unchallenged. Data are
reported as mean ± SEM and are representative of 3 independent experiments, with 5 replicates each (One-Way
ANOVA, all p < 0.05).
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
4
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
by the acetylcholine agonist Clothianidin, which enhances the transcription of NGFR. en, the observed tran-
scriptional regulation may potentially inuence cross-communication between the nervous and the immune
systems28.
Our experimental data on THP-1 cells clearly indicate that exposure to Clothianidin is detrimental for the
expression of genes under NF-κB control, as similarly observed in insects14. Indeed, both the RNA-Seq and
qRT-PCR data revealed marked eects of Clothianidin exposure on the expression of genes linked to immune
response. In particular, TRAF4 and TRAF6, which are members of the TRAF protein family, largely associated
with the immune response32, resulted up-regulated and down-regulated, respectively. ese proteins are involved
Figure 3. List of signicantly (FDR <0.05) up- and down-regulated genes with Log2 fold change higher than 1
and lower than 1, respectively.
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
5
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
in a number of transduction pathways of immune signaling molecules, with TRAF6 promoting NF-κB activation,
while TRAF4 is a negative modulator of this transcription factor, as it competes for its binding sites on signal
transduction proteins recruiting TRAF633. e concurrent transcriptional down-regulation of IL17R induced by
Clothianidin, associated with the TRAF6 and TRAF4 changes mentioned above, suggest that IL17 immune sign-
aling is negatively inuenced by neonicotinoids in THP-1 cells. Moreover, we can assume the occurrence of sim-
ilar immune disruption pathways, largely driven by the same mechanism, that can be further aggravated by the
down-regulation of proteins involved in NF-κB activation, such as IL18BP34. In contrast, the down-regulation of
FOXO4 is not easy to interpret. is transcription factor is a member of the FOXO protein family, which is central
in the integration of growth factor signaling, oxidative stress and inammation35. Recent work has demonstrated
that knockdown of FOXO4 does not aect NF-κB activation, suggesting that FOXO4 acts downstream in the
signaling pathway36. en, its down-regulation by Clothianidin treatment could further reinforce the inhibition
of NF-κB activation. is is a likely hypothesis that merits to be investigated. Future studies will have to address
these intriguing neuroimmune pathways, which are poorly known, in particular in non-vertebrate model sys-
tems, in order to fully appreciate the whole impact of neurotoxic molecules on the immune system. is is an
important research area that is currently not adequately considered in toxicological studies.
In conclusion, our data show that the insecticide Clothianidin negatively inuences in a human cell line the
expression of immune related genes, under control of the transcription factorNF-κB, as similarly observed in
Figure 4. Relative gene expression in THP-1 cell treated with Clothianidin: NGFR (a); TRAF4 (b); TRAF6 (c);
FOXO4 (d); IL18BP (e); IL17R (f). Data are reported as a mean ± SEM of 3 independent experiments, with 5
replicates each. (Student’s t test, all p < 0.05).
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
6
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
insects. ese ndings are interesting, but their relevance will have to be assessed at the organism level to see if
they may represent a signicant threat for human health.
Methods
Cell culture. e human monocytic cell line THP-1 (ATCC number: TIB-202; LGC Standards GmbH) was
cultured in RPMI 1640 (Life technologies) medium supplemented with 10% FCS (Life technologies) and 1 mM of
L-glutamine (Life technologies), under 5% CO2 at 37 °C.
Total RNA extraction and cDNA Synthesis. Total RNA extraction from THP- 1 cells was carried out
by using TRIzol (ermoScientic), according to the manufacturer’s instructions. e RNA yield and A260/280
ratio were monitored with a NanoDrop ND 100 spectrometer (NanoDrop Technologies), and RNA integrity was
veried using the 2100 Bioanalyzer (Agilent Technologies). cDNA synthesis was carried out starting from 1 μg
of total RNA and using the High Capacity cDNA Reverse Transcription Kit(ermoScientic), according to the
manufacturer’s protocol.
RNA-Seq. An amount of 106/well cells (control and Clothianidin treated, 3 biological replicates each) were
processed for RNA-Seq analysis. Indexed libraries were prepared using 1 µg of each RNA puried with TruSeq
Stranded mRNA Sample Prep Kit (Illumina), according to the manufacturer’s instructions. Libraries were quanti-
ed using the Agilent 2100 Bioanalyzer (Agilent Technologies) and pooled, so that each index-tagged sample was
present in equimolar amounts, with a nal concentration of the pooled samples of 2 nM. e pooled samples were
subjected to cluster generation and sequencing, using an Illumina HiSeq. 2500 System (Illumina) in a 2 × 100
paired-end format, at a nal concentration of 8 pmol. e raw sequence les generated (.fastq les) underwent
quality control analysis, using FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Raw reads were checked for quality with FastQC v0.11.3 and then trimming and removal of adapters were
performed with Trimmomatic v0.33 (minimum quality score 25, minimum length 35). e obtained reads were
then mapped against Homo sapiens reference genome (Ensembl, GRCh38) with STAR (v2.5.0b), providing the
reference gene annotation le with known transcripts. FeatureCounts (v1.4.5-p1) was used to perform read sum-
marization at gene level, with the strand-specic option “reversely stranded”. Statistical analysis of the read counts
was performed with R, using the HTSFilter package, to remove low expressed genes, and the NOIseq package, to
perform dierential expression analysis. Gene Ontology enrichment analysis of the dierentially expressed genes
was performed with the GOStat package.
qRT-PCR. e expression prole of the immune genes that showed the most pronounced transcriptional
regulation was also analyzed by TaqManqRT-PCR, using specific primers and probes:TNF-α (AssayID:
Hs00174128_m1), FOXO4 (Assay ID: Hs00172973_m1), NGFR (Assay ID: Hs00609976_m1), IL18BP (Assay ID:
Hs00271720_m1), TRAF4 (Assay ID: Hs01030628_g1), TRAF6 (Assay ID: Hs00939742_g1) and IL17R (Assay
ID: Hs01056316_m1). All probes were normalized to Gapdh (Assay ID: Hs02786624_g1) as internal control
(Applied Biosystems). All fold changes were calculated using the ΔΔCt method (Livack et al., 2001) and com-
pared with untreated cell. Amplications were performed with ABI PRISM 7900HT (Applied Biosystems).
Clothianidin eect on TNF-α expression. Clothianidin was obtained from Sigma (Cat No: 33589) and
used as follows: 106/well cells were seeded in a 24 well plate and were pre-treated with Clothianidin (100 ng/
ml) overnight, then, stimulated for 1 hour with LPS 1 μg/ml (Sigma), and compared with unchallenged cells and
untreated basal controls. Experimental cells were washed in PBS before RNA extraction. A qRT-PCR analysis was
performed to measure the transcription rate of TNF-α gene in THP-1 cell treated as described above. TNF-α pro-
tein secretion was measured in cell free supernatant using TNF-α DuoSet ELISA development kit (R&D system),
following manufacturers procedure.
NF-κB reporter gene assay. THP-1 cells were infected with 10 µL lentiviral particles carrying a NF-κB
responsive luciferase-expressing reporter gene (CignalLenti Reporters, SABiosciences), according to the protocol
provided by the manufacturer, followed by selection with puromycin. Once the cell line was established, THP-1
were incubated with Clothianidin (100 ng/ml) overnight and then treated with LPS (1 μg/ml) for 1 hour. NF-κB
activity was measured using Dual Glo Luciferase assay (Promega), according to the manufacturer’s procedure.
Luciferase activity was normalized for all samples with total amount of proteins.
Cytotoxicity assay. THP-1 cell were treated overnight with dierent dose of Clothianidin and cytotoxic-
ity was evaluated by measuring lactate dehydrogenase (LDH) release in the supernatant, using a CytoTox 96®
Non-Radio cytotoxicity assay kit (Promega, Madison, WI, USA), according to the manufacturer’s instructions.
Statistical analysis. Normality of data was checked with Shapiro-Wilk test, while homoscedasticity was
tested with Levene’s procedure. Dierences in the relative expression of TNF-α, secreted TNF-α protein and
Luciferase activity of the NF-κB responsive reporter gene were analyzed with One-Way ANOVA followed by
Games-Howell post-hoc test (parametric and non-homoscedastic procedure).
Two-tailed parametric non-homoscedastict-test was used to analyze dierences in relative expression of
NGFR, while for TRAF4, TRAF6, FOXO4, IL18BP and IL17R gene expression was analyzed with the two-tailed
parametric homoscedastic t-test. Dierences in the LDH amounts released by cells exposed to Clothianidin were
analysed by One-Way ANOVA, followed by LSD post-hoc test (parametric and homoscedastic procedure). ese
analyses were performed by using Prism v.5 for Mac OSX (GraphPad soware, San Diego, CA, USA). All statisti-
cal data are available in the Supplementary Table2.
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
7
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
References
1. Simon-Delso, N. et al. Systemic insecticides (neonicotinoids and pronil): trends, uses, mode of action and metabolites. Environ Sci
Pollut es Int 22, 5–34, https://doi.org/10.1007/s11356-014-3470-y (2015).
2. Tomizawa, M. & Casida, J. E. Neonicotinoid insecticide toxicology: mechanisms of selective action. Annu ev Pharmacol Toxicol 45,
247–268, https://doi.org/10.1146/annurev.pharmtox.45.120403.095930 (2005).
3. Elb ert, A., Haas, M., Springer, B., ielert, W. & Nauen, . Applied aspects of neonicotinoid uses in crop protection. Pest Manag Sci
64, 1099–1105, https://doi.org/10.1002/ps.1616 (2008).
4. EASAC. Ecosystem services, agriculture and neonicotinoids. EASAC policy report 26 (2015).
5. Pistorius, J., Bischo, G., Heimbach, U. & Stähler, M. Bee poisoning incidents in Germany in spring 2008 caused by abrasion of
active substance from treated seeds during sowing of maize. ulius-ühn-Archiv 423 (2009).
6. Yang, E. C., Chuang, Y. C., Chen, Y. L. & Chang, L. H. Abnormal foraging behavior induced by sublethal dosage of imidacloprid in
the honey bee (Hymenoptera: Apidae). J Econ Entomol 101, 1743–1748 (2008).
7. Han, P., Niu, C. Y., Lei, C. L., Cui, J. J. & Desneux, N. Use of an innovative T-tube maze assay and the proboscis extension response
assay to assess sublethal eects of GM products and pesticides on learning capacity of the honey bee Apis mellifera L. Ecotoxicology
19, 1612–1619, https://doi.org/10.1007/s10646-010-0546-4 (2010).
8. Henry, M. et al. A common pesticide decreases foraging success and survival in honey bees. Science 336, 348–350, https://doi.
org/10.1126/science.1215039 (2012). science.1215039 [pii].
9. Alaux, C. et al. Interactions between Nosema microspores and a neonicotinoid weaen honeybees (Apis mellifera). Environmental
Microbiology 12, 774–782, https://doi.org/10.1111/j.1462-2920.2009.02123.x (2010).
10. Aufauvre, J. et al. Parasite-insecticide interactions: a case study of Nosema ceranae and pronil synergy on honeybee. Scientic
eports 2, 326, https://doi.org/10.1038/srep00326 (2012).
11. Pettis, J. S., vanEngelsdorp, D., Johnson, J. & Dively, G. Pesticide exposure in honey bees results in increased levels of the gut
pathogen Nosema. Naturwissenschaen 99, 153–158, https://doi.org/10.1007/s00114-011-0881-1 (2012).
12. Fauser-Misslin, A., Sadd, B. M., Neumann, P. & Sandroc, C. Inuence of combined pesticide and parasite exposure on bumblebee
colony traits in the laboratory. Journal of Applied Ecology 51, 450–459, https://doi.org/10.1111/1365-2664.12188 (2014).
13. Doublet, V., Labarussias, M., de Miranda, J. ., Moritz, . F. & Paxton, . J. Bees under stress: sublethal doses of a neonicotinoid
pesticide and pathogens interact to elevate honey bee mortality across the life cycle. Environ Microbiol 17, 969–983, https://doi.
org/10.1111/1462-2920.12426 (2015).
14. Di Prisco, G. et al. Neonicotinoid clothianidin adversely aects insect immunity and promotes replication of a viral pathogen in
honey bees. Proc Natl Acad Sci USA 110, 18466–18471, https://doi.org/10.1073/pnas.1314923110 (2013).
15. Brandt, A. et al. e neonicotinoids thiacloprid, imidacloprid, and clothianidin aect the immunocompetence of honey bees (Apis
mellifera L.). J Insect Physiol 86, 40–47, https://doi.org/10.1016/j.jinsphys.201j6.01.001 (2016).
16. Nazzi, F. et al. Synergistic parasite-pathogen interactions mediated by host immunity can drive the collapse of honeybee colonies.
PLoS Pathog 8, e1002735, https://doi.org/10.1371/journal.ppat.1002735 (2012).
17. Nazzi, F. & Pennacchio, F. Disentangling multiple interactions in the hive ecosystem. Trends Parasitol 30, 556–561, https://doi.
org/10.1016/j.pt.2014.09.006 (2014).
18. Di Prisco, G. et al. A mutualistic symbiosis between a parasitic mite and a pathogenic virus undermines honey bee immunity and
health. Proc Natl Acad Sci USA 113, 3203–3208, https://doi.org/10.1073/pnas.1523515113 (2016).
19. Sánchez-Bayo, F. et al. Are bee diseases lined to pesticides? — A brief review. Environment International 89, 7–11, https://doi.
org/10.1016/j.envint.2016.01.009 (2016).
20. Spatuzza, C. et al. Physical and functional characterization of the genetic locus of IBt, an inhibitor of Bruton’s tyrosine inase:
evidence for three protein isoforms of IBt. Nucleic Acids es 36, 4402–4416, https://doi.org/10.1093/nar/gn413 (2008).
21. Cimino, A. M., Boyles, A. L., Thayer, . A. & Perry, M. J. Effects of Neonicotinoid Pesticide Exposure on Human Health: A
Systematic eview. Environmental Health Perspectives 125, 155–162, https://doi.org/10.1289/ehp515 (2017).
22. Seltenrich, N. Catching Up with Popular Pesticides: More Human Health Studies Are Needed on Neonicotinoids. Environ Health
Perspect 125, A41–A42, https://doi.org/10.1289/ehp.125-A41 (2017).
23. Aira, S. & Taeda, . Toll-lie receptor signalling. Nat ev Immunol 4, 499–511, https://doi.org/10.1038/nri1391 (2004).
24. Costantini, T. W. et al. A Human-Specic alpha7-Nicotinic Acetylcholine eceptor Gene in Human Leuocytes: Identication,
egulation and the Consequences of CHFAM7A Expression. Mol Med 21, 323–336, https://doi.org/10.2119/molmed.2015.00018
(2015).
25. Szlarczy, D. et al. STINGv10: protein-protein interaction networs, integrated over the tree of life. Nucleic Acids es 43,
D447–452, https://doi.org/10.1093/nar/gu1003 (2015).
26. Tracey, . J. eex control of immunity. Nat ev Immunol 9, 418–428, https://doi.org/10.1038/nri2566 (2009).
27. Tracey, . J. Ancient Neurons egulate Immunity. Science (New Yor, N.Y.) 332, 673–674, https://doi.org/10.1126/science.1206353
(2011).
28. Talbot, S., Foster, S. L. & Woolf, C. J. Neuroimmunity: Physiology and Pathology. Annu ev Immunol 34, 421–447, https://doi.
org/10.1146/annurev-immunol-041015-055340 (2016).
29. Li, P., Ann, J. & A, G. Activation and modulation of human alpha4beta2 nicotinic acetylcholine receptors by the neonicotinoids
clothianidin and imidacloprid. J Neurosci es 89, 1295–1301, https://doi.org/10.1002/jnr.22644 (2011).
30. Ye, X. et al. TAF family proteins interact with the common neurotrophin receptor and modulate apoptosis induction. J Biol Chem
274, 30202–30208 (1999).
31. Hepburn, L. et al. A Spaetzle-lie role for Nerve Growth Factor β in vertebrate immunity to Staphylococcus aureus. Science (New
Yor, N.Y.) 346, 641–646, https://doi.org/10.1126/science.1258705 (2014).
32. Aggarwal, . & Silverman, N. Positive and negative regulation of the Drosophila immune response. BMB ep 41, 267–277 (2008).
33. Xie, P. TAF molecules in cell signaling and in human diseases. J Mol Signal 8, 7, https://doi.org/10.1186/1750-2187-8-7 (2013).
34. Dinarello, C. A. & Fantuzzi, G. Interleuin-18 and host defense against infection. J Infect Dis 187(Suppl 2), S370–384, https://doi.
org/10.1086/374751 (2003).
35. Hedric, S. M., Michelini, . H., Doedens, A. L., Goldrath, A. W. & Stone, E. L. FOXO transcription factors throughout T cell
biology. Nature reviews. Immunology 12, https://doi.org/10.1038/nri3278 (2012).
36. Oteiza, A. & Mechti, N. Control of FoxO4 Activity and Cell Survival by TIM22 Directs TL3-Stimulated Cells Toward IFN Type I
Gene Induction or Apoptosis. J Interferon Cytoine es 35, 859–874, https://doi.org/10.1089/jir.2015.0020 (2015).
Acknowledgements
e research work reported in this paper was supported by POR Campania FESR 2007–2013, Bio Industrial
Processes-BIP (to FP and RC) and by EU Seventh Framework Program (FP7/2007–2013), under Grant 613960
(SMARTBEES) (to FP). Dr. Marco Iannaccone was supported by research funding from Fondazione con il Sud
(Project no. 2011-PDR-18, ‘Biosensori piezoelettrici a risposta in tempo reale per applicazioni ambientali e agro-
alimentari’).
Content courtesy of Springer Nature, terms of use apply. Rights reserved
www.nature.com/scientificreports/
8
Scientific REPORTs | 7: 13446 | DOI:10.1038/s41598-017-13171-z
Author Contributions
R.C. and F.P. conceived the study and designed the research plan; G.D.P., M.I., F.I., R.F. and E.C. performed the
experiments; G.D.P. and M.I. analyzed data; F.P. and R.C. wrote the paper, which was revised and approved by all
co-authors.
Additional Information
Supplementary information accompanies this paper at https://doi.org/10.1038/s41598-017-13171-z.
Competing Interests: e authors declare that they have no competing interests.
Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and
institutional aliations.
Open Access This article is licensed under a Creative Commons Attribution 4.0 International
License, which permits use, sharing, adaptation, distribution and reproduction in any medium or
format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Cre-
ative Commons license, and indicate if changes were made. e images or other third party material in this
article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the
material. If material is not included in the article’s Creative Commons license and your intended use is not per-
mitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the
copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
© e Author(s) 2017
Content courtesy of Springer Nature, terms of use apply. Rights reserved
1.
2.
3.
4.
5.
6.
Terms and Conditions
Springer Nature journal content, brought to you courtesy of Springer Nature Customer Service Center GmbH (“Springer Nature”).
Springer Nature supports a reasonable amount of sharing of research papers by authors, subscribers and authorised users (“Users”), for small-
scale personal, non-commercial use provided that all copyright, trade and service marks and other proprietary notices are maintained. By
accessing, sharing, receiving or otherwise using the Springer Nature journal content you agree to these terms of use (“Terms”). For these
purposes, Springer Nature considers academic use (by researchers and students) to be non-commercial.
These Terms are supplementary and will apply in addition to any applicable website terms and conditions, a relevant site licence or a personal
subscription. These Terms will prevail over any conflict or ambiguity with regards to the relevant terms, a site licence or a personal subscription
(to the extent of the conflict or ambiguity only). For Creative Commons-licensed articles, the terms of the Creative Commons license used will
apply.
We collect and use personal data to provide access to the Springer Nature journal content. We may also use these personal data internally within
ResearchGate and Springer Nature and as agreed share it, in an anonymised way, for purposes of tracking, analysis and reporting. We will not
otherwise disclose your personal data outside the ResearchGate or the Springer Nature group of companies unless we have your permission as
detailed in the Privacy Policy.
While Users may use the Springer Nature journal content for small scale, personal non-commercial use, it is important to note that Users may
not:
use such content for the purpose of providing other users with access on a regular or large scale basis or as a means to circumvent access
control;
use such content where to do so would be considered a criminal or statutory offence in any jurisdiction, or gives rise to civil liability, or is
otherwise unlawful;
falsely or misleadingly imply or suggest endorsement, approval , sponsorship, or association unless explicitly agreed to by Springer Nature in
writing;
use bots or other automated methods to access the content or redirect messages
override any security feature or exclusionary protocol; or
share the content in order to create substitute for Springer Nature products or services or a systematic database of Springer Nature journal
content.
In line with the restriction against commercial use, Springer Nature does not permit the creation of a product or service that creates revenue,
royalties, rent or income from our content or its inclusion as part of a paid for service or for other commercial gain. Springer Nature journal
content cannot be used for inter-library loans and librarians may not upload Springer Nature journal content on a large scale into their, or any
other, institutional repository.
These terms of use are reviewed regularly and may be amended at any time. Springer Nature is not obligated to publish any information or
content on this website and may remove it or features or functionality at our sole discretion, at any time with or without notice. Springer Nature
may revoke this licence to you at any time and remove access to any copies of the Springer Nature journal content which have been saved.
To the fullest extent permitted by law, Springer Nature makes no warranties, representations or guarantees to Users, either express or implied
with respect to the Springer nature journal content and all parties disclaim and waive any implied warranties or warranties imposed by law,
including merchantability or fitness for any particular purpose.
Please note that these rights do not automatically extend to content, data or other material published by Springer Nature that may be licensed
from third parties.
If you would like to use or distribute our Springer Nature journal content to a wider audience or on a regular basis or in any other manner not
expressly permitted by these Terms, please contact Springer Nature at
onlineservice@springernature.com
... Activation of NF-kB leads to the production of inflammatory cytokines, which are part of the bodyʹs response to harmful stimuli. Inflammation, closely linked to the immune system, is a pathological response to such stimuli [23]. Additionally, the transcription factor nuclear factor, erythroid 2-like-2 (NFE2L2/NRF2) plays a significant role in cytoprotection by stimulating the expression of AOX and detoxifying enzymes, including NAD(P)H:quinone oxidoreductase-1 (NQO-1), glutathione S-transferase (GST), and hemeoxygenase-1 (HO-1) [24]. ...
Preprint
Full-text available
The thymus, a central lymphoid organ in animals serves as the site for T cell development, differentiation and maturation, vital to adaptive immunity. Thymus is critical for maintaining tissue homeostasis to protect against tumors and tissue damage. Overactive or prolonged immune response can lead to oxidative stress from increased production of reactive oxygen species. Heat stress induces oxidative stress and overwhelms the natural antioxidant defense mechanisms. The study objectives were to investigate the protective properties of astaxanthin against heat-induced oxidative stress and apoptosis in the chicken thymus, by comparing the growth performance and gene signaling pathways among three groups- thermal neutral, heat stress and heat stress with astaxanthin. The thermal neutral temperature conditions were 21-22°C and heat stress temperature was 32-35°C. Both heat stress groups experienced reduced growth performance, while the astaxanthin treated group showed a slightly lesser decline. The inflammatory response and antioxidant defense system were activated by the upregulation of the NF-kB, NFE2L2, PPARα, cytoprotective capacity and apoptotic gene pathways in the heat stress compared to the thermal neutral group. Conversely, expression levels were without significance between the thermal neutral and heat stress with antioxidant groups, suggesting astaxanthin antioxidant effectiveness to mitigate inflammation and oxidative stress damage.
... Activation of NF-kB leads to the production of inflammatory cytokines, which are part of the body's response to harmful stimuli. Inflammation, closely linked to the immune system, is a pathological response to such stimuli [23]. Additionally, the transcription factor nuclear factor, erythroid 2-like-2 (NFE2L2/NRF2) plays a significant role in cytoprotection by stimulating the expression of AOX and detoxifying enzymes, including NAD(P)H:quinone oxidoreductase-1 (NQO-1), glutathione S-transferase (GST), and hemeoxygenase-1 (HO-1) [24]. ...
Preprint
Full-text available
The thymus, a central lymphoid organ in animals serves as the site for T cell development, differentiation and maturation, vital to adaptive immunity. Thymus is critical for maintaining tissue homeostasis providing protection against tumors and tissue damage. Overactive or prolonged immune response can lead to oxidative stress due to increased production of reactive oxygen species. Heat stress induces oxidative stress and overwhelms the natural antioxidant defense mechanisms. The objectives of the study were to investigate the protective properties of astaxanthin against heat-induced oxidative stress and apoptosis in the chicken thymus, by comparing the growth performance and gene signaling pathways among three groups- thermal neutral, heat stress and heat stress with astaxanthin, under two temperature conditions of 21-22°C and 32-35°C. Both treatments under heat stress experienced reduced growth performance, while the group treated with astaxanthin showed a slightly lesser decline. The inflammatory response and antioxidant defense system were activated by the upregulation of the NF-kB, NFE2L2, PPARα, cytoprotective capacity and apoptotic gene pathways in the heat stress compared to the thermal neutral group. Conversely, expression levels were without significance between the thermal neutral and heat stress with antioxidant groups, suggesting astaxanthin antioxidant effectiveness to mitigate inflammation and oxidative stress damage.
... Neonicotinoids have low affinity for vertebrate nicotinic receptors as compared to that of the insects. Hence, they generally show low acute toxicity to mammals, birds, and fish as compared to traditional insecticides 8 , but some recent studies also display that neonicotinoid pesticides indeed cause some toxicity in amphibians, fish and mammals including humans [9][10][11][12][13][14][15][16] . In recent years, neonicotinoids and their metabolites have been detected in various environmental and biological samples 17 . ...
Article
Full-text available
The ubiquitous use of insecticides leads to detrimental effects on non-target organisms due to accidental exposure. Neonicotinoid insecticides are popularly used worldwide for their high affinity for arthropod nicotinic receptors which effectively kill insect pests. Low affinity towards vertebrate nicotinic receptors, make them safer as compared to traditional insecticides. Recent studies demonstrated that neonicotinoid exposure can cause some toxicity in vertebrates including humans. Zebrafish is one of the popular model organisms to study ecotoxicity. This is the first study on adult zebrafish to report the effect of novel neonicotinoid, Clothianidin on mortality, liver antioxidant stress profile, liver function profile and brain acetylcholinesterase (AChE). Observations were made over two treatment periods, 96 hours, and 21 days, in five groups exposed to varying concentrations of Clothianidin viz. 30mg/L, 50mg/L, 70mg/L, 90mg/L, 110mg/L and a control. Although no mortality was observed, Clothianidin exposure led to increased activity of superoxide dismutase (SOD) enzyme, lipid peroxidation and decreased catalase (CAT), glutathione-S-transferase (GST). Treated groups also showed increased concentrations of liver enzyme alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) and acid phosphatase (ACP). These results indicate that Clothianidin causes disturbances in the hepatic function. A prominent decreasing trend observed in brain AChE shows that Clothianidin inhibits AChE in adult zebrafish. Further investigations on DNA damage and gene expression studies could be conducted to understand the exact mechanism of action of Clothianidin
Chapter
Humans are continuously exposed to insecticides through various routes, including dietary and drinking water intake or inhalation of pollen and dust. Chronic exposure to insecticides such as neonicotinoids has raised concerns about their potential toxic effects on non-target organisms. After ingestion, much of the neonicotinoids is excreted in the urine, but a portion of the parent compounds and their metabolites are distributed throughout the body where they may have potential adverse effects. Although neonicotinoids have a low affinity for human nicotinic acetylcholine receptors (nAChRs), compared to insect receptors, the high density and ubiquitous distribution of these receptors in the human nervous system makes them a potential target for toxic effects of neonicotinoids. Several in vivo and in vitro studies, especially with animals, have linked exposure to neonicotinoids with the development of toxic effects in mammals, including neurological symptoms, liver and kidney abnormalities, or metabolic changes, among others. This chapter will review human exposure to neonicotinoids and summarize the existing evidence on their potential impact on different systems of the human body. Due to the scarcity of studies on the toxic effects of neonicotinoids in humans, the information will be complemented by the available evidence on their effects in rodents.
Article
Full-text available
The thymus, a central lymphoid organ in animals, serves as the site for T cell development, differentiation and maturation, vital to adaptive immunity. The thymus is critical for maintaining tissue homeostasis to protect against tumors and tissue damage. An overactive or prolonged immune response can lead to oxidative stress from increased production of reactive oxygen species. Heat stress induces oxidative stress and overwhelms the natural antioxidant defense mechanisms. This study’s objectives were to investigate the protective properties of astaxanthin against heat-induced oxidative stress and apoptosis in the chicken thymus, by comparing the growth performance and gene signaling pathways among three groups: thermal neutral, heat stress, and heat stress with astaxanthin. The thermal neutral temperature was 21–22 °C, and the heat stress temperature was 32–35 °C. Both heat stress groups experienced reduced growth performance, while the astaxanthin-treated group showed a slightly lesser decline. The inflammatory response and antioxidant defense system were activated by the upregulation of the NF-kB, NFE2L2, PPARα, cytoprotective capacity, and apoptotic gene pathways during heat stress compared to the thermal neutral group. However, expression levels showed no significant differences between the thermal neutral and heat stress with antioxidant groups, suggesting that astaxanthin may mitigate inflammation and oxidative stress damage.
Chapter
The increase in the world population is projected to reach 9.3 billion by the year 2050 and will no doubt require a significant and continued increase in food production to meet the food needs. The adoption of intensive agriculture began in the second half of the twentieth century and is dependent on diverse agrochemicals. Agricultural intensification practices involve the enlargement of small farms into large ventures, the concentration on the culture of single species of exotic cash crops, and the use of pesticides and fertilizers. The results from these practices have been very encouraging in terms of the amount of food produces but not without a price on biodiversity and environmental integrity. For instance, it has constituted a source of threat to wildlife habitats, niche functionality, and ecosystem processes and services all over the world with the outcome depending on the type and amount of agrochemical commonly used in the locale. This chapter attempts to collate evidence from previous studies on the extent of information on the detrimental effects of agricultural intensification through agrochemical use on the various environments and flora and fauna diversity around the world. The harmful effects of these modern agricultural practices are taking a negative toll on diverse important aspects of biodiversity and indirectly affecting human sustenance on Earth. To address this problematic trend, policies such as the adoption of sustainable agricultural practices are crucial. An example of such is organic agriculture which has less hazardous effects on biodiversity. These practices must be put in place by authorities and stakeholders in the agriculture industry so that food can be secured and conservation of biodiversity will be of major interest.KeywordsAgricultural intensificationAgrochemicalsBiodiversityEcosystemWildlife habitatsHabitat loss
Article
Full-text available
Background: Numerous studies have identified detectable levels of neonicotinoids (neonics) in the environment, adverse effects of neonics in many species including mammals, and pathways through which human exposure to neonics could occur, yet little is known about the human health effects of neonic exposure. Objective: This systematic review sought to identify human population studies on the health effects of neonics. Methods: Studies published in English between 2005 and 2015 were searched using PubMed, Scopus, and Web of Science databases. No restrictions were placed on the type of health outcome assessed. Risk of bias was assessed using guidance developed by the National Toxicology Program's Office of Health Assessment and Translation. Results: Eight studies investigating the human health effects of exposure to neonics were identified. Four examined acute exposure: three neonic poisoning studies reported two fatalities (n=1280 cases) and an occupational exposure study of 19 forestry workers reported no adverse effects. Four general population studies reported associations between chronic neonic exposure and adverse developmental or neurological outcomes, including tetralogy of Fallot (AOR 2.4, 95% CI: 1.1-5.4), anencephaly (AOR 2.9, 95% CI: 1.0-8.2), autism spectrum disorder (AOR 1.3, 95% CrI: 0.78-2.2), and a symptom cluster including memory loss and finger tremor (OR 14, 95% CI: 3.5-57). Reported odds ratios were based on exposed compared to unexposed groups. Conclusions: The studies conducted to date were limited in number with suggestive but methodologically weak findings related to chronic exposure. Given the wide-scale use of neonics, more needs to be known about their human health effects.
Article
Full-text available
Significance The parasitic mite Varroa destructor and the deformed wing virus (DWV) are linked in a mutualistic symbiosis. The mite acts as vector of the viral pathogen, whereas the DWV-induced immunosuppression in honey bees favors mite feeding and reproduction. This functional interaction underpins a rapidly escalating immunosuppression, which can be primed and/or aggravated by a wealth of stress factors that co-trigger colony losses. Our experimental results explain the pivotal role proposed for the Varroa –DWV association in the induction of honey bee colony losses. Here we provide a functional framework for studying the dynamics of this multifactorial syndrome and defining effective strategies to reduce its negative impact on the beekeeping industry.
Article
Full-text available
Evolution has yielded multiple complex and complementary mechanisms to detect environmental danger and protect tissues from damage. The nervous system rapidly processes information and coordinates complex defense behaviors, and the immune system eliminates diverse threats by virtue of mobile, specialized cell populations. The two systems are tightly integrated, cooperating in local and systemic reflexes that restore homeostasis in response to tissue injury and infection. They further share a broad common language of cytokines, growth factors, and neuropeptides that enables bidirectional communication. However, this reciprocal cross talk permits amplification of maladaptive feedforward inflammatory loops that contribute to the development of allergy, autoimmunity, itch, and pain. Appreciating the immune and nervous systems as a holistic, coordinated defense system provides both new insights into inflammation and exciting opportunities for managing acute and chronic inflammatory diseases. Expected final online publication date for the Annual Review of Immunology Volume 34 is May 20, 2016. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.
Article
Full-text available
The human genome contains a variant form of the α7-nicotinic acetylcholine receptor (α7nAChR) gene that is uniquely human. This CHRFAM7A gene arose during human speciation and recent data suggests that its expression alters ligand tropism of the normally homopentameric human α7-AChR ligand-gated cell surface ion channel that is found on the surface of many different cell types. To understand its possible significance in regulating inflammation in humans, we investigated its expression in normal human leukocytes and leukocyte cell lines, compared CHRFAM7A expression to that of the CHRNA7 gene, mapped its promoter and characterized the effects of stable CHRFAM7A over-expression. We report here that CHRFAM7A is highly expressed in human leukocytes but that the levels of both CHRFAM7A and CHRNA7 mRNAs were independent and varied widely. To this end, mapping of the CHRFAM7A promoter in its 5'-untranslated region (UTR) identified a unique 1kb sequence that independently regulates CHRFAM7A gene expression. Because over-expression of CHRFAM7A in THP1 cells altered the cell phenotype and modified the expression of genes associated with focal adhesion (e.g. FAK, P13K, Akt, rho, GEF, Elk1, CycD), leukocyte trans-epithelial migration (Nox, ITG, MMPs, PKC) and cancer (kit, kitL, ras, cFos cyclinD1, Frizzled and GPCR), we conclude that CHRFAM7A is biologically active. Most surprisingly however, stable CHRFAM7A overexpression in THP1 cells up-regulated CHRNA7, which in turn, led to increased binding of the specific α7nAChR ligand, bungarotoxin on the THP1 cell surface. Taken together, these data confirm the close association between CHRFAM7A and CHRNA7 expression, establish a biological consequence to CHRFAM7A expression in human leukocytes and support the possibility that this human-specific gene might contribute to, and/or gauge, a human-specific response to inflammation.
Article
Full-text available
Many key components of innate immunity to infection are shared between Drosophila and humans. However, the fly Toll ligand Spaetzle is not thought to have a vertebrate equivalent. We have found that the structurally related cystine-knot protein, nerve growth factor β (NGFβ), plays an unexpected Spaetzle-like role in immunity to Staphylococcus aureus infection in chordates. Deleterious mutations of either human NGFβ or its high-affinity receptor tropomyosin-related kinase receptor A (TRKA) were associated with severe S. aureus infections. NGFβ was released by macrophages in response to S. aureus exoproteins through activation of the NOD-like receptors NLRP3 and NLRC4 and enhanced phagocytosis and superoxide-dependent killing, stimulated proinflammatory cytokine production, and promoted calcium-dependent neutrophil recruitment. TrkA knockdown in zebrafish increased susceptibility to S. aureus infection, confirming an evolutionarily conserved role for NGFβ-TRKA signaling in pathogen-specific host immunity.
Article
A strong immune defense is vital for honey bee health and colony survival. This defense can be weakened by environmental factors that may render honey bees more vulnerable to parasites and pathogens. Honey bees are frequently exposed to neonicotinoid pesticides, which are being discussed as one of the stress factors that may lead to colony failure. We investigated the sublethal effects of the neonicotinoids thiacloprid, imidacloprid, and clothianidin on individual immunity, by studying three major aspects of immunocompetence in worker bees: total hemocyte number, encapsulation response, and antimicrobial activity of the hemolymph. In laboratory experiments, we found a strong impact of all three neonicotinoids. Thiacloprid (24 h oral exposure, 200 lg/l or 2000 lg/l) and imidacloprid (1 lg/l or 10 lg/l) reduced hemocyte density, encapsulation response, and antimicrobial activity even at field realistic concentrations. Clothianidin had an effect on these immune parameters only at higher than field realistic concentrations (50–200 lg/l). These results suggest that neonicotinoids affect the individual immunocompetence of honey bees, possibly leading to an impaired disease resistance capacity. � 2016 Elsevier Ltd. All rights reserved.
Article
Activation of innate immune response, induced after the recognition of double-stranded RNA (dsRNA), formed during replication of most viruses, results in intracellular signaling cascades ultimately culminating in the expression of type I interferon (IFN). In this study, we provide the first evidence that FoxO4 triggers the activation of the innate immune signaling pathway in coupling stimulation of TLR3 and RIG-like receptors by the synthetic dsRNA analog, poly(I:C), to IFN-β and IFN-induced gene induction, whereas knockdown of FoxO4 had opposite effects. Similar effects of FoxO4 were observed during paramyxovirus-mediated IFN-β transcriptional induction. We further found that knockdown of FoxO4 did not affect IRF3 and NF-κB activation by poly(I:C), suggesting that FoxO4 would act downstream in the signaling pathway. In addition, we show that the IFN-induced TRIM22 ubiquitin ligase targets FoxO4 and antagonizes its activity through an unrelated ubiquitin/autophagosomic-lysosomal pathway. Unexpectedly, TRIM22 knockdown strongly sensitizes cells to dsRNA-induced caspase-dependent apoptosis, as early as 2 h after poly(I:C) stimulation, concomitantly to the inhibition of the expression of the antiapoptotic protein, Bcl-2, indicating that TRIM22 might be a key factor for controlling the cell survival after TLR3 stimulation. Taken together, our data demonstrate that the regulation of FoxO4 protein expression and cell survival by TRIM22 controls TLR3-mediated IFN type I gene induction, preventing excessive antiviral response through dsRNA-induced apoptosis.
Article
The widespread losses of honeybee colonies recorded over the past number of years in the northern hemisphere represent a major concern for the beekeeping industry and, more importantly, may have a severe impact on ecological services and biodiversity. There is now a general consensus about the multifactorial origin of colony losses, but the mechanistic basis of this complex phenomenon still remains largely elusive. In this review, we propose a functional framework for interpreting how different stress agents can interact to adversely affect bee immunity and health. This provides a new background rationale in which to develop an integrated approach to bee protection, as part of a more comprehensive strategy for the conservation of insect pollinators. Copyright © 2014 Elsevier Ltd. All rights reserved.