Article

Ellagitannins from Pomegranate Ameliorates 5-Fluorouracil-Induced Intestinal Mucositis in Rats while Enhancing Its Chemotoxicity against HT-29 Colorectal Cancer Cells through Intrinsic Apoptosis Induction

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Abstract

Worldwide, colorectal cancer (CRC) is a deleterious disease causing millions of death annually. 5-Fluorouracil (5-FU) is a first-line chemotherapy for CRC, but chemoresistance and gastrointestinal mucositis limit its efficacy. Polyphenol-rich foods are increasingly popular due to their potential beneficial role in cancer. Ellagitannins is a group of phenolic compounds commonly found in pomegranate, strawberries, raspberries, etc. The objective of this study was to explore whether ellagitannins from pomegranate (PETs) could ameliorate 5-FU-induced intestinal mucositis and enhance its efficacy against CRC. The results showed that PETs (100 mg/kg) counteracted 5-FU-induced intestinal mucositis in rats. The number of apoptotic cells per crypt was reduced from 1.50±0.21 to 0.85±0.18 (P<0.05). Moreover, PETs induced HT-29 CRC cell death through intrinsic apoptosis as demonstrated by dissipation of mitochondrial membrane potential, increased Bax to Bcl-2 ratio, and cleavage of caspase 9 and caspase 3. PETs and 5-FU combination treatments exhibited synergistic cytotoxicity against HT-29 cells with a weighted combination index of 0.3494. PETs (80 µg/mL) and 5-FU (40 µg/mL) treatments for 48 h induced 14.03±0.76% and 16.42±1.15% of HT-29 cells to undergo apoptosis while the combination treatment further increased apoptosis cells to 34.00±1.54% (P<0.05). Combination treatment of the cells also enhanced S phase cell cycle arrest as compared with PETs or 5-FU monotherapy (P<0.05). These results suggest that dietary ellagitannins from pomegranate could alleviate intestinal mucositis in rats induced by 5-FU while enhancing its toxicity against HT-29 cells through potentiation of apoptosis and cell cycle arrest.

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... Polyphenols can become pro-oxidants to generate this additional oxidative stress in cancer cells, primarily through the generation of reactive oxygen species (ROS) (Khan et al., 2014). We have previously reported that polyphenols from Chinese herbal medicine pomegranate peels induce apoptosis in colorectal cancer cells (Chen et al., 2018). ...
... The extracts were combined and diluted to 10 ml with water for further analysis. Preparation procedures for the punicalagin fraction as well as punicalagin and granatin B fraction were previously described (Chen et al., 2018). From 20 g of pomegranate peel powder, 1.32 ± 0.14 g of punicalagin and 0.82 ± 0.25 g of the granatin B and punicalagin rich fraction were obtained, respectively. ...
... LC-MS/MS analysis was performed by an Agilent 1290 infinity LC system with API 3200 Q Trap and controlled by Analyst software (Chen et al., 2018). Mobile phase A was 0.2% acetic acid aqueous solution and B was ACN. ...
Article
Background : Colorectal cancer ranks among the most common cancers. 5-Fluorouracil (5-FU) based first-line chemotherapy for colorectal cancer treatment often leads to chemoresistance and gastrointestinal mucositis. Purpose : This study aimed to find potential therapeutic agents from herbal medicine with anti-colorectal cancer and anti-mucositis activities. Methods : Chinese medicine theory, network pharmacology analyses, and antioxidant activity coupled with liquid chromatography tandem mass spectrometry analyses were used to identify potential bioactive compounds. HT-29 human colorectal cancer cell culture and xenograft tumor models were employed to study anti-colorectal cancer efficacy. Lipopolysaccharide-induced RAW 264.7 and 5-FU treated Dark Agouti rats were used to evaluate anti-inflammatory and anti-mucositis activities. Histological staining, immunofluorescence imaging, western blots, and flow cytometric analyses were employed to explore the underlying mechanisms. Results : Both Chinese medicine theory and network pharmacology analyses indicated pomegranate peels as a potential anti-colorectal cancer and anti-mucositis agent. Antioxidant activity coupled with liquid chromatography tandem mass spectrometry analyses revealed granatin B and punicalagin as the most potent antioxidant compounds in pomegranate peels. Granatin B and punicalagin demonstrated superior anti-colorectal cancer activities in both cell culture and xenograft tumor models. Granatin B and punicalagin also exhibited strong anti-inflammatory activities in lipopolysaccharide-induced RAW264.7 cells and anti-mucositis activities in 5-FU-treated rats. Mechanistic studies revealed that granatin B and punicalagin induced reactive oxygen species-mediated S-phase cell cycle arrest and apoptosis in HT-29 cells. Moreover, these compounds sensitized HT-29 cells to 5-FU-induced cell death and S-phase cell cycle arrest. Conclusion : We report that granatin B and punicalagin exhibit superior anti-colorectal cancer and anti-mucositis activities. To the best of our knowledge, these results are novel and suggest that utilizing phenols from herbal medicine, such as granatin B and punicalagin, to target reactive oxygen species may be an innovative therapy to treat colorectal cancer and intestinal mucositis.
... Thus, considering the possible role of the mediators evaluated here also in intestinal mucositis associated with chemotherapy, the effect of PPJE was also studied in IECs treated with 5-FU. Previous studies already suggested a protective effect of pomegranate juice in intestinal mucositis, at the level of intestinal epithelial cells [58], but the novelty of this study rests, in particular, on the use of a nontumorigenic cell line and in the evaluation of parameters potentially involved in the five-phase model of mucositis development, proposed by Sonis [8]. In brief, the stages of this model include the phases of (1) initiation, (2) upregulation, (3) signal amplification, (4) ulceration and (5) healing. ...
... The reduction of apoptotic rate was also associated with a significant reduction in caspase-3 expression. These results agree with previous data reporting the ability of ellagitannins from pomegranate to ameliorate intestinal mucositis and reduce apoptosis of intestinal cells induced by 5-FU in rats [58]. In most cases, mucositis is an acute phenomenon that is self-resolving once cancer therapy ends. ...
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Intestinal epithelial cells (IECs) play a pivotal role in maintaining intestinal homeostasis. Different noxious agents, among them also anticancer therapies, can impair intestinal epithelial integrity triggering inflammation and oxidative stress. A frequent complication of chemotherapy is gastrointestinal mucositis, strongly influencing the effectiveness of therapy, increasing healthcare costs, and impairing patients’ quality of life. Different strategies are used to treat gastrointestinal mucositis, including products from natural sources. Our study focused on the effect of pomegranate (Punica granatum L.) juice extract on IEC-6 cells, both during inflammatory conditions and following treatment with 5-fluorouracil (5-FU). The polyphenolic profile of pomegranate juice was characterized in detail by Online Comprehensive two dimensional Liquid Chromatography-Mass Spectrometry. The evaluation of pomegranate juice extract in IEC-6 indicates a significant inhibition in proinflammatory factors, such as cytokines release, cyclooxygenase-2 and inducible nitric oxide synthase expression, and nitrotyrosine formation. Pomegranate also inhibited oxidative stress and adhesion protein expression. In 5-FU-treated IEC-6, pomegranate also inhibited both inflammatory and oxidative stress parameters and apoptosis. It promoted wound repair and tight junction expression. These results suggest a potential use of pomegranate as an adjuvant in the treatment of intestinal inflammatory and oxidative stress states, which also occur during chemotherapy-induced mucositis.
... However, treatment with 5-FU not only destroys neoplastic cells, it also affects cell populations of healthy tissue throughout the body (Duncan and Grant, 2003;Chen et al., 2018). More than 50% of patients treated with 5-FU suffer from oral and intestinal mucositis, which are accompanied by pain, bacteremia, and malnutrition (Decker-Baumann et al., 1999;Liu et al., 2012;Tang et al., 2017). ...
... After cooling to room temperature, the supernatant was used. Western blotting and data analysis were performed according to Chen et al. (2018). Antibodies used (all from Solarbio) were anti-NLRP3 polyclonal antibody (rabbit, 1:1,000), anti-Bcl2 polyclonal antibody (rabbit, 1:1,000) and anti-β-actin polyclonal antibody (rabbit, 1:1,000) and goat anti-rabbit IgG/horseradish peroxidase antibody (rabbit, 1:5,000). ...
Article
5-Fluorouracil (5-FU) is widely used as a chemotherapeutic drug for the treatment of cancer but it has toxic side effects. It can induce severe intestinal damage and even lead to death. The purpose of this study was to investigate whether milk fermented with Lactobacillus rhamnosus FLRH93 could alleviate intestinal damage induced by 5-FU. The results of injury intervention in a mouse model showed that milk fermented with Lb. rhamnosus FLRH93 significantly ameliorated intestinal injury caused by 5-FU. The results of hematoxylin and eosin staining showed that mice fed Lb. rhamnosus FLRH93 preserved the villus/crypt ratio and reduced the loss of goblet cells in ileum sections of 5-FU-treated animal. Further, administration of fermented milk upregulated expression of Bcl-2 in the intestinal tract and downregulated the expression of NLRP3, thus reducing the production of inflammatory factors interleukin 1-β and tumor necrosis factor-α. The survival rate of mice treated with fermented milk was twice that of mice not fed fermented milk after continuous oral administration of 5-FU. In conclusion, Lb. rhamnosus FLRH93 has positive effects on body injury and could be used to prevent intestinal damage caused by cancer chemotherapy.
... Several studies pointed to the potential neuroprotective effect of many constituents present in PP, that is, ellagitannins (Tejada et al., 2017) ellagic acid to prevent against myelin-associated sphingolipid loss in EAE, and further its metabolite urolithin, which stimulated ceramide biosynthesis in glioma (Busto et al., 2018), and to suggest that more studies should be conducted on PP as a neuropro- thyroid Liu et al., 2017), uterine, breast (Dikmen et al., 2011;Fazio et al., 2018), bladder (Chang et al., 2018), prostate (Deng et al., 2017), leukemia (Panth et al., 2017), and osteosarcoma (Li et al., 2014). Moreover, its ellagitannins enhanced the chemotoxicity of 5-fluorouracil in colorectal cancer cell lines through apoptosis potentiation and to lessen its side effects in rats (Chen, Lam, et al., 2018). The synergism between natural extracts and chemotherapeutic agents has been extensively reported in the literature (Atteya et al., 2017) and extend herein to include pomegranate peel. ...
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Pomegranate is an ancient shrub, globally distributed nowadays. It has been used in the middle east as a medicinal food and traditional medicine for thousands of years. Pomegranate peel (PP) constitutes about 50% of the total fruit, however, it has been previously regarded as a waste. Recent research points to PP as a rich source of phenolics (e.g., ellagitannins, flavonoids, and anthocyanins), polysaccharides, in addition to its biotransformed metabolites viz. urolithins making it a valuable waste with promising pharmacological actions. Compared to the pulp and the juice, PP exhibited stronger antioxidant and antimicrobial activities. Besides, it inhibited inflammation in several conditions, including colitis, arthritis, hepatitis, contact dermatitis, and lung inflammation. Moreover, it displayed anti-osteoporosis, anti-hyperglycemic, antidiabetic, antihypertensive, vasculoprotective, hepatoprotective, neuroprotective, and immunomodulatory effects. Additionally, it was effective as a prebiotic and in obesity control, besides it promoted wound healing. Furthermore, PP demonstrated anticancer effects against different cancer types, for example, colon, liver, thyroid, uterine, breast, bladder, prostate, leukemia, and osteosarcoma. Despite PP safety, it may interfere with the metabolism of other drugs because it inhibits cytochromes (CYP) changing their bioavailability, effectiveness, and toxicity. PP biowaste valorization not only avoids against its environmental and economic burden but can also provide a promising platform to produce novel or improved nutraceuticals. This study provides a comprehensive overview of PP biological activities with the reported action mechanisms related to its phytochemicals and further biotransformed metabolites inside the body. Future research prospects to unravel the merits of such waste and optimize its use are discussed. Practical application Pomegranate is widely distributed throughout the world. Although its peel was previously considered a waste, recent research regards it as a rich source of bioactive compounds with promising biological activities. Its recycling not only overcomes the bio-waste problems, but also provides a source of valuable compounds with several health benefits. In recent years, PP has been demonstrated to exhibit excellent pharmacological bioactivities, for example, antioxidant, anti-inflammatory, antimicrobial, antiosteoporosis, antihyperlipidemic, and anticancer activities. Its health-promoting power is mostly attributed to the phenolic and polysaccharide content, in addition to its amazing biotransformed metabolites. The underlying action mechanisms of such pharmacological activities are discussed and related to its chemical content. This review presents the latest research progress on the role of PP in the prevention and treatment of various chronic diseases, and its protective health effects for future research to be used in nutraceuticals.
... Banerjee et al. [105] demonstrated that polyphenolic extract from pomegranate juice exerted cytotoxic, anti-inflammatory and antiangiogenesis effects on HT-29 cells via upregulation of caspase-3, PARP cleavage, downregulation of COX-2, vascular cell adhesion molecule 1 (VCAM-1), VEGF, and NF-B p65, reduction in phosphorylation of phosphatidylinositol 3kinase/protein kinase B (PI3K/Akt) and increase in the expression of miRNA-126. Recently, Chen et al. [106] showed that both pomegranate peel-derived ellagitannins alone and ellagitannins in combination with 5-fluorouracil (5-FU) significantly inhibited HT-29 cell proliferation and induced S-phase cell cycle arrest. Concomitant mechanistic studies showed induction of intrinsic apoptosis via dissipation of mitochondrial membrane potential, increased Bax to Bcl-2 ratio, and cleavage of caspase-3 and caspase-9. ...
Article
Cancer remains to be the second highest cause of mortality in our society, falling just short of heart disease. Despite major advancement in cancer therapy over the past decade, momentum has been gaining for an alternative approach of using naturally-occurring and dietary agents for cancer prevention and management. Research on pomegranate (Punica granatum L.), a fruit of the Punicaceae family, has shown enormous potential for cancer prevention and intervention. In addition to a rich source of polyphenols, including flavonoids and ellagitannins, in its juice, pomegranate also houses hundreds of other phytochemicals in its pericarp, seed, flower, bark, flowers and leaves, These phytochemicals provide powerful antiproliferative, anti-inflammatory, antioxidant, anti-invasive, antimigratory, anti-angiogenic and anti-metastatic effects without significant toxicity. This makes the use of its various extracts a very attractive strategy to our current battle against cancer. This review article presents a systematic, comprehensive and critical review of research on pomegranate-derived products in both cancer prevention and intervention. It discusses the chemical constituents of pomegranate, the results of both preclinical (in vitro, ex vivo and in vivo) and clinical studies on the anticancer effect of pomegranate phytochemicals and molecular targets in numerous types of cancers, such as breast, gastrointestinal tract (oral, colon, liver and pancreas), gynecological (uterine and ovarian), hematological (lymphoma, leukemia and myeloma), lung, neurological (glioma), urogenital (bladder and prostate), bioavailability, pharmacokinetics and safety of pomegranate constituents. In order to guide the direction of future research, we have also included current limitations and challenges in the field and our post analysis recommendation.
... They used lactate dehydrogenase, 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium bioassays, acridine orange-ethidium bromide, and terminal deoxyribonucleotidyl transferasemediated dUTP nick-end labeling to study the cytotoxic and growth inhibition effects of pomegranate extracts and genistein on MCF-7 cancer cells. Similar studies were also carried out by Dikmen et al. (2011) to demonstrate that the methanolic extract of pomegranate peel at varied concentrations (25,50,100,200, and 300 μg/ml) Ameliorates 5-FU-induced intestinal mucositis Chen et al. (2018) HT-29 CRC cell line Induces intrinsic apoptosis with a decrease in mitochondrial potential, increases bcl-2-like protein 4 (BAX) to Bcl-2 ratio, and cleaves caspase-9 and caspase-3 Apc-mutated Pirc rats Stimulates type I procollagen synthesis and inhibits matrix metalloproteinase-1 (MMP-1; interstitial collagenase) production by dermal fibroblasts that promote the regeneration of dermis (Aslam et al. 2006) (continued) decreased cell proliferation and stimulated apoptosis in MCF-7 cancer cells. This was evident from the enhanced expression of pre-apoptotic gene Bax and decrease in the expression of anti-apoptotic gene Bcl-2. ...
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Cancer is one of the prominent death causing diseases around the globe. About 1 in 6 deaths is due to cancer and its related diseases. Cancer mortality can be reduced by early diagnosis and screening, implementing effective treatments. A precise cancer identification is vital for effective treatment, because each cancer type requires a definite treatment procedure, such as radiotherapy, surgery, and chemotherapy. The brisk expansion of herbal therapy and escalating ongoing clinical studies are becoming trendy and useful in the drug development against cancer. Pomegranate (Punica granatum) is a prehistoric fruit with illustrious dietary and remedial properties in alternative traditional systems of medicine. The current chapter is aiming to understand various model systems (in silico, in vitro, and in vivo), employed for studying its anti-cancerous properties and diverse molecular effects exhibited by the pomegranate peel and its phytoconstituents. It also highlights the importance of secondary metabolites of P. granatum, especially ellagitannins and their anticancer properties. Although there are enormous in vitro and preclinical data, human clinical trials are sorely lacking. The major focus is on up-to-date investigations into the outcomes of previously reported pomegranate peel components against a diverse type of cancers.
... Therefore, anticancer drugs may cause some side effects on immune system. 5-Fluorouracil (5-FU) is an effective antitumor drug and it is still in first-line chemotherapy to treat various kinds of cancers, including lung , breast (Boncel et al., 2016), liver (Qi et al., 2016), colorectal (Chen et al., 2018), and pancreatic cancers (Wang et al., 2014). Moreover, 5-FU has important side effects such as myelosuppression, cardiotoxicity and gastrointestinal toxicity (Potikanond et al., 2015;Takimoto et al., 1996). ...
Article
Due to the toxicity and resistance to treatment with anticancer drugs, various methods are used to improve their efficacy in cancer treatment. In this present study, in order to overcome the limitation of 5-Fluorouracil (5-FU), prodrug strategy has been pursued with using density functional theory (DFT) and molecular dynamics simulation (MDs). The main objective of this study is to examine the mechanisms of drug release from its prodrug form by using the intrinsic reaction coordinate (IRC) calculations. The reaction mechanisms of 5-FU prodrug (EMC-5-FU) in the presence of lactic acid (LA) and water molecule were theoretically studied. The IRC calculations were carried out at the M06-2X/6-311G** level in the aqueous phase through the mechanism of ester hydrolysis to obtain energies, the geometry optimization of all stationary points along the potential energy surfaces (PES), and also to determine the harmonic vibrational frequencies. The results herein presented suggest that three reaction pathways and transition states TS1 to TS2 are involved along the calculated potential energy surface. We found that the drug molecule is released in the third step and this occurs by separation CH2O group in the presence of water molecule with the highest energy barrier about 25.9 kcal/mol. Since the carbon nanotubes (CNTs) can act as drug delivery vehicles and deliver anticancer drugs directly to the target cells. Therefore in DFT section, the interaction mechanism of CNTs with 5-FU prodrug is studied by means of DFT method. The atoms in molecules (AIM) and the non-covalent interactions (NCI) between the CNTs and prodrug are used in order to examine the strength and type of interaction between them. The result of negative binding energy values of CNT-prodrug interaction show the stability of these complexes. Our theoretical results show that the more favorable interaction occurs when the prodrug is located inside the carbon nanotube. Furthermore, for design and development of intracellular drug delivery systems, steered molecular dynamics (SMD) simulations was used to investigate the possibility of encapsulated prodrug-CNT penetration through a (1-palmitoyl-2-oleoyl phosphatidylcholine) POPC lipid bilayer. For this purpose, the forces of penetration and the free energies of rupture of POPC bilayer with a Prodrug-CNT were studied. Our simulation results show that encapsulated prodrug-carbon nanotube does not permanently destroy the POPC membrane structure.
... Aynı zamanda yapılan son çalışmalar incelendiğinde nar kabuğunun potansiyel bir anti-karsinojen olabileceğini görülmektedir 15 . Kemoterapötikler ile nar kabuğu ekstresinin kombine tedavisinin tedaviyi desteklediği ve daha güçlü anti kanser etki oluştuğu da önceki çalışmalarda gösterilmiştir 16 . Ancak nar kabuğunun kanser kemoterapisine bağlı gelişebilecek nörotoksisiteye karşı nasıl bir etki göstereceği konusunda bir bilgi bulunmamaktadır. ...
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Introduction: This study pointed to validate the phytochemistry, antioxidant and antimutagenic activities of two common plant peels; Punica granatum L. and Opuntia ficus-indica L. Material and methods: HPLC analysis was performed for identification of phenolics and flavonoids, beside to isolation of some triterpenes and phenolics from both plant peels. Results: P. granatum peels composed of 16 flavonoids and 18 phenolics, while O. ficus-indica comprised of 18 flavonoids and 10 phenolics. α-Amyrin acetate (1), friedelin (2), lup-20(29)-en-3β-ol (3), quercetin-3,4'-dimethyl ether-7-O-α-Larabinofuranosyl β-D-glucopyranoside (4), punicaflavanol (5), and two hydrolyzabl tannins (6&7) were isolated from P. granatum peels, while friedelin (8), 24-Methylene-ergosta-5-en-3β-ol (9), apigenin-7-O-glucoside (10), isorhamnetin 3-O-β-D-glucopyranoside (11), and betanin pigment (12) were isolated from O. ficus-indica peels. Compounds (1-5 and 8, 10, 11, 12) were isolated for the first time from both plant peels. P. granatum and O. ficus-indica peel extracts have relatively significant antioxidant, cytotoxic and antimutagenic effects. Discussion and conclusion: The tested plant peel extracts could be a reliable source as natural antioxidant, antimutagenic and cytotoxic agents with a high level of safety. The novelty of this study is the comparison of such activities of the peels under study.
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Chemotherapeutic medications, including 5 – fluorouracil (5FU), are the same old technique to most cancers and are associated with numerous peripheral toxicities. We investigated exopolysaccharide (EPSST) produced from the isolated streptomycete of the Mediterranean Sea for the capability to lower the severity of mucositis in vivo. The streptomycete was isolated from Mediterranean Sea sediment from the beaches of Port Said Governorates, Egypt and identified morphologically, physiologically, and biochemically and confirmed by molecularly 16S rDNA analysis. The EPSST was extracted from the supernatant of streptomycete by using 4 volumes chilled ethanol and then the functional groups, MW, and chemical evaluation have been detected via Fourier-transform infrared (FTIR), and high-performance liquid chromatography (HPLC). In addition, antioxidant activity was measured through the usage of 2, 2-diphenyl-1-picrylhydrazyl (DPPH). Thirty-two male rats (180–200 g) were randomly divided into a control group (normal saline), intraperitoneal injection of 5-fluorouracil (5-FU, 150 mg/kg), normal rats were treated with EPSST and 5-FU + EPSST group. These groups were continued up to the day of sacrifice (28 days post treatments). The isolated strain became recognized based totally on 16S rDNA sequence as Streptomyce sp. with accession number SAMN08349905. The chemical evaluations of EPSST were galacturonic, glucose, galactose, mannose, and arabinose with a relative ratio of 2.1: 1: 5.37: 1.62: 1.29 individually, with an average molecular weight (Mw) 9.687 × 10³ g/mol. Also, the EPSST contained uronic acid (16%) and sulfate (12.149%) and no protein was detected. EPSST inhibited the DPPH radical activity. The findings of this study propose that EPSST inhibits 5-FU-induced mucositis through adjustment of oxidative stress, apoptosis, inflammatory factors, activation of antioxidant enzymes. The clinical administration of EPSST may recover the chemotherapy-induced intestinal dysfunction, consequently increasing the clinical efficiency of chemotherapy. In addition, the administration of EPSST reduced 5-FU-induced histopathological incongruities such as neutrophil infiltration, loss of cellular integrity, and villus and crypt distortion. The clinical administration of EPSST may recover the chemotherapy-induced intestinal dysfunction, consequently increasing the clinical efficiency of chemotherapy.
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Colorectal cancer is a leading cause of cancer-related mortality worldwide, with Fluorouracil (5-FU)-based chemotherapy as the major treatment for advanced disease. Many patients with advanced colorectal cancer eventually succumb to the disease despite some patients responded initially to chemotherapy. Thus, identifying molecular mechanisms responsible for chemotherapy resistance will help design novel strategies to treat colorectal cancer. In this study, we established an acquired 5-FU resistant cell line, LoVo-R, from LoVo cells. Through exome sequencing, we discovered that elevated GLI1 signaling axis is a major genetic alteration in the 5-FU resistant cells. Hh signaling, a pathway essential for embryonic development, is an important regulator for residual cancer cells. We demonstrated that knockdown of GLI1 or GLI2 sensitized LoVo-R cells to 5-FU treatment, reduced cell invasiveness. The relevance of our studies to colorectal cancer patients is reflected by our discovery that high expression of GLI1 signaling molecules was associated with a high incidence of cancer relapse and a shorter survival in a larger cohort of colorectal cancer patients who underwent chemotherapy (containing 5-FU). Taken together, our data demonstrate the critical role of the GLI1 signaling axis for 5-FU resistance in colorectal cancer. Electronic supplementary material The online version of this article (doi:10.1186/s13578-017-0145-7) contains supplementary material, which is available to authorized users.
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Proanthocyanidins (PAs) have been shown to have potential health benefits. However, no data exist concerning their dietary intake. Therefore, PAs in common and infant foods from the U.S. were analyzed. On the bases of our data and those from the USDA's Continuing Survey of Food Intakes by Individuals (CSFII) of 1994-1996, the mean daily intake of PAs in the U.S. population (>2 y old) was estimated to be 57.7 mg/person. Monomers, dimers, trimers, and those above trimers contribute 7.1, 11.2, 7.8, and 73.9% of total PAs, respectively. The major sources of PAs in the American diet are apples (32.0%), followed by chocolate (17.9%) and grapes (17.8%). The 2- to 5-y-old age group (68.2 mg/person) and men >60 y old (70.8 mg/person) consume more PAs daily than other groups because they consume more fruit. The daily intake of PAs for 4- to 6-mo-old and 6- to 10-mo-old infants was estimated to be 1.3 mg and 26.9 mg, respectively, based on the recommendations of the American Academy of Pediatrics. This study supports the concept that PAs account for a major fraction of the total flavonoids ingested in Western diets.
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In this study, the content, structure, anti-tyrosinase activity and mechanism of longan bark condensed tannins were evaluated. The findings obtained from mass spectrometry demonstrated that longan bark condensed tannins were mixtures of procyanidins, propelargonidins, prodelphinidins, and their acyl derivatives (galloyl and p-hydroxybenzoate). The enzyme analysis indicated that these mixtures were efficient, reversible, and mixed (competitive is dominant) inhibitor of tyrosinase. What’s more, the mixtures showed good inhibitions on proliferation, intracellular enzyme activity and melanogenesis of mouse melanoma cells (B16 ). From molecular docking, the results showed the interactions between inhibitors and tyrosinase were driven by hydrogen bond, electrostatic and hydrophobic interactions. In addition, high levels of total phenolic and extractable condensed tannins suggested that longan bark might be a good source of tyrosinase inhibitor. This study would offer theoretical basis for the development of longan bark condensed tannins as novel food preservative and medicine of skin diseases.
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5-Fluorouracil (5-FU) is a commonly used chemotherapeutic agent for breast cancer. However, its use often leads to drug resistance and mucositis. This study aimed to investigate whether proanthocyanidins from Ficus virens possessed anti-breast cancer and anti-mucositis activities. The results showed that the cytotoxic effects of the proanthocyanidins against MDA-MB-231 and MCF-7 breast cancer cells were in the order of stem barks proanthocyanidins (SPAs) > leaves proanthocyanidins > fruits proanthocyanidins. Moreover, SPAs induced apoptosis in both cell lines which were accompanied with an increase in loss of mitochondrial membrane potential, production of reactive oxygen species, Bax to Bcl-2 protein expression ratio, and activated caspase 3. Furthermore, intraperitoneal injection of 5-FU (150 mg/kg body weight) resulted in body weight loss and jejunal injury in the rats while administration of SPAs (100 mg/kg body weight) counteracted these changes. Collectively, our study demonstrated that SPAs induced apoptosis cell death in breast cancer cells while ameliorating the symptoms of intestinal mucositis in rats.Therefore, SPAs merits further exploration as a potential therapeutic agent for breast cancer and chemotherapy-induced mucositis.
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Breast cancer is the most frequently diagnosed cancer and cause of cancer death in women worldwide. Current treatments often result in systematic toxicity and drug resistance. Combinational use of non-toxic phytochemicals with chemotherapy agents to enhance the efficacy and reduce toxicity would be one promising approach. In this study, bioactive proanthocyanidins from Uncaria rhynchophylla (UPAs) were isolated and their anti-breast cancer effects alone and in combination with 5- fluorouracil (5-FU) were investigated in MDA-MB-231 breast cancer cells. The results showed that UPAs significantly inhibited cell viability and migration ability in a dose-dependent manner. Moreover, UPAs induced apoptosis in a dose-dependent manner which was associated with increased cellular reactive oxygen species production, loss of mitochondrial membrane potential, increases of Bax/Bcl-2 ratio and levels of cleaved caspase 3. Treatments of the cells with UPAs resulted in an increase in G2/M cell cycle arrest. Cytotoxic effects of 5-FU against MDA-MB-231 cells were enhanced by UPAs. The combination treatment of UPAs and 5-FU for 48 h elicited a synergistic cytotoxic effect on MDA-MB-231 cells. Altogether, these data suggest that UPAs are potential therapeutic agents for breast cancer.
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Objectives: The optimal time to initiate adjuvant chemotherapy after surgery in patients with colon cancer is not clear. We investigated the benefit of combined intraportal chemotherapy administered during radical surgery with adjuvant chemotherapy for treating stage II and III colon cancer. Methods: Patients were randomly assigned to OCTREE arm (intraportal chemotherapy plus mFOLFOX6) or a standard adjuvant chemotherapy arm (mFOLFOX6). The primary study endpoint was disease-free survival. The secondary endpoints included metastasis-free survival, overall survival, and safety. Results: The intent-to-treat population comprised 237 patients. With a median follow-up of 44 months, the hazard ratio (OCTREE vs mFOLFOX6) was 0.66 (95% confidence interval, 0.43-0.90), a 34% risk reduction in favor of OCTREE (P = 0.016). The 3-year disease-free survival rate was 85.2% for OCTREE and 75.6% for mFOLFOX6 alone (P = 0.030). The 3-year metastasis-free survival rates were 87.6% for OCTREE and 78.0% for mFOLFOX6 (P = 0.035). Patients had lower distant metastatic rate in the OCTREE arm (12.7% vs 22.7%; P = 0.044), when compared with the mFOLFOX6 arm. The 3-year overall survival was no significant difference between 2 arms (P = 0.178). Neutropenia occurred in 12.7% of the patients receiving OCTREE and in 2.5% of the patients receiving mFOLFOX6 (P = 0.003) within 2 weeks of surgery, and grade 3 or 4 toxicity event was no difference between 2 regimens. Conclusions: Combination of intraoperative intraportal chemotherapy with mFOLFOX6 reduced the occurrence of distant metastases and improved disease-free survival in patients with stage II and stage III colon cancer.
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Background and purpose: Chemotherapeutic agents, including 5-fluorouracil (5-FU), frequently cause intestinal mucositis resulting in severe diarrhoea and morphological mucosal damage. 5-HT₃ receptor antagonists are clinically effective in the treatment of nausea and emesis during cancer chemotherapy. Therefore we here have examined the effects of 5-HT₃ receptor antagonists on 5-FU-induced intestinal mucositis in mice. Experimental approach: Intestinal mucositis was induced in male C57BL/6 mice by daily administration of 5-FU (50 mg·kg⁻¹) for 5 days. Effects of 5-HT₃ receptor antagonists, ramosetron (0.01-0.1 mg·kg⁻¹) and ondansetron (5 mg·kg⁻¹), on the accompanying histology, cytokine production and apoptosis were assessed. Key results: Continuous administration of 5-FU to mice caused severe intestinal mucositis, which was histologically characterized by the shortening of villi and destruction of intestinal crypts, accompanied by body weight loss and diarrhoea. Daily ramosetron administration dose-dependently reduced the severity of intestinal mucositis, body weight loss and diarrhoea. Similar beneficial effects were observed with ondansetron. The number of apoptotic, caspase-3- and caspase-8-activated cells increased 24 h after the first 5-FU administration, and these responses were reduced by ramosetron. The up-regulation of TNF-α, IL-1β and IL-6 following 5-FU treatment was also attenuated by ramosetron. Conclusions and implications: 5-HT₃ receptor antagonists ameliorated 5-FU-induced intestinal mucositis in mice, and this action could result from suppression of apoptotic responses in the intestinal crypt cells via inhibition of cytokine expression. Thus, 5-HT₃ receptor antagonists may be useful for preventing not only nausea and emesis but also intestinal mucositis during 5-FU chemotherapy.
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Pomegranate (Punica granatum L.) fruits are widely consumed fresh and in processed forms as juice, jams and wine. Pomegranate fruit husk/peel is a rich source of hydrolyzable tannins called ellagitannins (ETs). In the commercial pomegranate juice (PJ) industry, these ETs are extracted from the husk in significant quantities into the juice due to their hydrophilic properties. Pomegranate husk, a by-product of the PJ industry, is therefore an inexpensive and abundant source of ETs. Previous methods to isolate pomegranate ETs included labor intensive and time-consuming solid phase extractions by column chromatography (C-18, polyamides, cellulose, Sephadex Lipophilic LH-20, Diaion HP20) and/or use of specialized instruments such as preparative-high performance liquid chromatography (HPLC). We have used an Amberlite XAD-16 resin vacuum-aspirated column to rapidly purify an aqueous extract of pomegranate husk to afford total pomegranate tannins (TPT) in substantial yields (58–60 g TPT/kg husk; time <1 h). Using analytical HPLC and tandem LC-ES/MS, evaluation of TPT showed that it contains the major fruit husk ET, punicalagin (80–85% w/w) and ellagic acid (EA; 1.3% w/w) and unquantified amounts of punicalin and EA-glycosides (hexoside, rhamnoside and pentoside). Since pomegranate ETs are reported to show potent antioxidant, antiatherosclerotic and anticancer activities, this method can be used for the large-scale production of TPT for future in vitro and in vivo biological studies. This method is practical for industrial applications and could provide a low-cost means to use a currently underutilized food by-product to develop phytoceuticals with potential health benefits or to develop products for use in the cosmetic and food biopreservative industries.
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The aim of the present review is to discuss the cumulative evidence that suggests that pomegranate consumption possesses a diverse array of biological actions and may be helpful in the prevention of some inflammatory-mediated diseases including cancer. The pomegranate fruit can be divided into at least three parts-seeds, peel, and juice. All these components have been studied for their antioxidant properties in a chemoprevention approach. Pomegranate exerts antiproliferative, anti-invasive, and antimetastatic effects, induces apoptosis through modulation of Bcl-2 proteins, increases p21 and p27, and downregulates cyclin-cdk network. In addition, pomegranate inhibits the activation of inflammatory pathways including, but not limited to, the NFκ-B pathway. Anti-cancer effects with the most impressive data have been demonstrated so far in prostate cancer.
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Fas is expressed constitutively in colonic epithelial cells and is also expressed in colon carcinomas and in cultured colon carcinoma cell lines. However, the potential role of Fas signaling in mediating apoptosis in cells of this type remains unknown. We have developed human colon carcinoma cell models deficient in thymidylate synthase that demonstrate acute (TS- cells) or delayed (Thy4 cells) apoptosis following DNA damage induced by thymineless stress. Complete protection of cells from acute apoptosis and prolongation of delayed apoptosis was obtained following exposure to the NOK-1 monoclonal antibody (inhibitory to Fas signaling) during the period of dThd deprivation. These results suggested that apoptosis induced by thymineless stress was regulated by autocrine signaling via Fas-FasL interactions. Fas expression was high in both TS- and Thy4 cells. However, FasL, undetectable in synchronous cultures, was up-regulated in TS- cells at 48 hr, when cells were undergoing acute apoptosis, and in Thy4 cells at 96 hr, correlating with the delayed onset of thymineless death. FasL expression also correlated with acute apoptosis induced in parental GC3/cl cells, commencing at 48 hr, following thymidylate synthase inhibition by 5-fluorouracil/leucovorin exposure. Fas-mediated apoptosis induced by the cytotoxic anti-Fas monoclonal antibody CH-11 was inhibited following adenoviral delivery of a Bcl-2 cDNA, and Bcl-2 also protected cells from acute apoptosis induced by dThd deprivation. Taken together, these data demonstrate a functional Fas system in these cultured colon carcinoma cell models, and they demonstrate that Fas-FasL interactions can link DNA damage induced by thymineless stress to the apoptotic machinery of colon carcinoma cells.
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To elucidate the relationship between 5-fluorouracil (5-FU) distribution and 5-FU-induced apoptosis and/or cell cycle arrest, microautoradiography was applied to murine intestinal crypts exposed to [14C] 5-FU by intravenous infusion. The histologic location of apoptotic cells in the crypt did not correlate to that of 5-FU. The temporal profiles of apoptotic and/or mitotic indexes corresponded to those of orally administered 5-FU in a previous study. Two cell cycle-related proteins, p21(WAF/Cip1) and bax, were also investigated in the present study. With time, p21(WAF/Cip1)-positive nuclei apparently migrated up the crypt. Bax-positive cytoplasm was observed throughout the crypt epithelial cells, accompanied by the occurrence of apoptosis, and remained until 48 h when the control level recovered. The findings demonstrate that 5-FU mainly exerts an apoptotic effect and/or cell cycle arrest by systemic exposure, and p21 and bax expression determine an individual cell's fate.
Article
Necrotizing enterocolitis (NEC) is a devastating intestinal disease of premature infants. Although end-stage NEC is characterized histopathologically as extensive necrosis, apoptosis may account for the initial loss of epithelium before full development of disease. We have previously shown that epidermal growth factor (EGF) reduces the incidence of NEC in a rat model. Although EGF has been shown to protect intestinal enterocytes from apoptosis, the mechanism of EGF-mediated protection against NEC is not known. The aim of this study was to investigate if EGF treatment elicits changes in expression of apoptotic markers in the ileum during the development of NEC. With the use of a well-established neonatal rat model of NEC, rats were divided into the following three experimental groups: dam fed (DF), milk formula fed (NEC), or fed with formula supplemented with 500 ng/ml EGF (NEC+EGF). Changes in ileal morphology, gene and protein expression, and histological localization of apoptotic regulators were evaluated. Anti-apoptotic Bcl-2 mRNA levels were markedly reduced and pro-apoptotic Bax mRNA levels were markedly elevated in the NEC group compared with DF controls. Supplementation of EGF into formula significantly increased anti-apoptotic Bcl-2 mRNA, whereas pro-apoptotic Bax was significantly decreased. The Bax-to-Bcl-2 ratio for mRNA and protein was markedly decreased in NEC+EGF animals compared with the NEC group. The presence of caspase-3-positive epithelial cells was markedly reduced in EGF-treated rats. These data suggest that alteration of the balance between pro-and anti-apoptotic proteins in the site of injury is a possible mechanism by which EGF maintains intestinal integrity and protects intestinal epithelium against NEC injury.
Article
Polyphenol-rich dietary foodstuffs have attracted attention due to their cancer chemopreventive and chemotherapeutic properties. Ellagitannins (ETs) belong to the so-called hydrolysable tannins found in strawberries, raspberries, walnuts, pomegranate, oak-aged red wine, etc. Both ETs and their hydrolysis product, ellagic acid (EA), have been reported to induce apoptosis in tumour cells. Ellagitannins are not absorbed in vivo but reach the colon and release EA that is metabolised by the human microflora. Our aim was to investigate the effect of a dietary ET [pomegranate punicalagin (PUNI)] and EA on human colon cancer Caco-2 and colon normal CCD-112CoN cells. Both PUNI and EA provoked the same effects on Caco-2 cells: down-regulation of cyclins A and B1 and upregulation of cyclin E, cell-cycle arrest in S phase, induction of apoptosis via intrinsic pathway (FAS-independent, caspase 8-independent) through bcl-XL down-regulation with mitochondrial release of cytochrome c into the cytosol, activation of initiator caspase 9 and effector caspase 3. Neither EA nor PUNI induced apoptosis in normal colon CCD-112CoN cells (no chromatin condensation and no activation of caspases 3 and 9 were detected). In the case of Caco-2 cells, no specific effect can be attributed to PUNI since it was hydrolysed in the medium to yield EA, which entered into the cells and was metabolised to produce dimethyl-EA derivatives. Our study suggests that the anticarcinogenic effect of dietary ETs could be mainly due to their hydrolysis product, EA, which induced apoptosis via mitochondrial pathway in colon cancer Caco-2 cells but not in normal colon cells.
Article
Phytochemicals from fruits such as the pomegranate (Punica granatum L) may inhibit cancer cell proliferation and apoptosis through the modulation of cellular transcription factors and signaling proteins. In previous studies, pomegranate juice (PJ) and its ellagitannins inhibited proliferation and induced apoptosis in HT-29 colon cancer cells. The present study examined the effects of PJ on inflammatory cell signaling proteins in the HT-29 human colon cancer cell line. At a concentration of 50 mg/L PJ significantly suppressed TNFalpha-induced COX-2 protein expression by 79% (SE = 0.042), total pomegranate tannin extract (TPT) 55% (SE = 0.049), and punicalagin 48% (SE = 0.022). Additionally, PJ reduced phosphorylation of the p65 subunit and binding to the NFkappaB response element 6.4-fold. TPT suppressed NFkappaB binding 10-fold, punicalagin 3.6-fold, whereas ellagic acid (EA) (another pomegranate polyphenol) was ineffective. PJ also abolished TNFalpha-induced AKT activation, needed for NFkappaB activity. Therefore, the polyphenolic phytochemicals in the pomegranate can play an important role in the modulation of inflammatory cell signaling in colon cancer cells.
Article
Mucositis is a clinically important and sometimes dose-limiting complication of cancer therapy. Mucositis lesions can be painful, affect nutrition and quality of life, lead to sepsis, and have significant economic impact. Recent modeling of the toxicity has been based on the continuum of clinical signs and symptoms of mucositis involving the alimentary tract, including both oral and gastrointestinal sites. The pathogenesis of oral and gastrointestinal mucositis is multifactorial and complex. In recent years, there has been a substantial increase in both basic and clinical research related to mucosal injury in cancer patients. Since most of this research has been directed to oral mucositis, the present review principally addresses this component of the toxicity. Morbidity, economic impact, pathogenesis and clinical course of mucositis are discussed. In addition, several agents in clinical development for mucositis are discussed in the context of the current pathobiologic model as well as the recently updated evidence-based clinical management guidelines.
World cancer report; International Agency for Research on Cancer
  • B W Stewart
  • C Wild
Stewart, B. W.; Wild, C. World cancer report; International Agency for Research on Cancer, WHO Press: Lyon, France, and Geneva, Switzerland, 2014.