富硒双歧杆菌缓解5-FU导致的化疗性肠粘膜炎的药效学研究

通过腹腔注射5-FU建立小鼠肠黏膜炎模型,探讨富硒长双歧杆菌(Selenium-enriched Bifidobacterium longum,Se-B.longum)能否改善5-FU所致的小鼠肠黏膜炎。将健康BALB/c小鼠随机分为对照组、5-FU组和Se-B.longum/5-FU组,分别灌胃生理盐水、生理盐水和Se-B.longum(Se0.3 mg/kg BW,1×106bacteria/只)6 d,然后5-FU组和Se-B.longum/5-FU组小鼠均腹腔注射5-FU(250 mg/kg),观察小鼠腹泻及死亡情况,5 d后处死小鼠,计算体重变化、脏器指数及考察肠道组织变化;将健康BALB/c小鼠随机分为对照组、5-FU组和Se-B.longum/5-FU组,分别灌胃生理盐水、生理盐水和Se-B.longum(Se 0.3 mg/kg BW,1×106bacteria/只)6 d,然后5-FU组和Se-B.longum/5-FU组小鼠均腹腔注射5-FU(300 mg/kg),观察小鼠死亡情况,绘制生存曲线。Se-B.longum能缓解5-FU导致的正常小鼠的肠粘膜炎、降低小鼠死亡率。     

Potential role of the NADPH oxidase NOX1 in the pathogenesis of 5-fluorouracil-induced intestinal mucositis in mice

Although NADPH oxidase 1 (NOX1) has been shown to be highly expressed in the gastrointestinal tract, the physiological and pathophysiological roles of this enzyme are not yet fully understood. In the present study, we investigated the role of NOX1 in the pathogenesis of intestinal mucositis induced by the cancer chemotherapeutic agent 5-fluorouracil (5-FU) in mice. Intestinal mucositis was induced in Nox1 knockout (Nox1KO) and littermate wild-type (WT) mice via single, daily administration of 5-FU for 5 days. In WT mice, 5-FU caused severe intestinal mucositis characterized by a shortening of villus height, a disruption of crypts, a loss of body weight, and diarrhea. In Nox1KO mice, however, the severity of mucositis was significantly reduced, particularly with respect to crypt disruption. The numbers of apoptotic caspase-3- and caspase-8-activated cells in the intestinal crypt increased 24 h after the first 5-FU administration but were overall significantly lower in Nox1KO than in WT mice. Furthermore, the 5-FU-mediated upregulation of TNF-α, IL-1β, and NOX1 and the production of reactive oxygen species were significantly attenuated in Nox1KO mice compared with that in WT mice. These findings suggest that NOX1 plays an important role in the pathogenesis of 5-FU-induced intestinal mucositis. NOX1-derived ROS production following administration of 5-FU may promote the apoptotic response through upregulation of inflammatory cytokines.     

Saireito (TJ-114), a Japanese Traditional Herbal Medicine,Reduces 5-Fluorouracil-Induced Intestinal Mucositis in Mice by Inhibiting Cytokine-Mediated Apoptosis in Intestinal Crypt Cells

Clinical chemotherapy frequently causes intestinal mucositis as a side effect, which is accompanied by severe diarrhea. We recently showed that the cytokine-mediated apoptotic pathway might be important for the development of intestinal mucositis induced by 5-fluorouracil (5-FU). Saireito, the traditional Japanese herbal (Kampo) medicine, is widely used to treat diarrhea and various inflammatory diseases in Japan. In the present study, we investigated the effect of saireito on 5-FU-induced intestinal mucositis in mice, especially in relation to apoptosis in the intestinal crypt. Male C57BL/6 mice were given 5-FU (50 mg/kg), i.p. once daily for 6 days. Intestinal mucositis was evaluated histochemically. Saireito (100–1000 mg/kg) was administered p.o. twice daily for 6 days. Repeated 5-FU treatment caused severe intestinal mucositis including morphological damage, which was accompanied by body weight loss and diarrhea. Daily administration of saireito reduced the severity of intestinal mucositis in a dose-dependent manner. Body weight loss and diarrhea during 5-FU treatment were also significantly attenuated by saireito administration. The number of apoptotic and caspase-3-activated cells in the intestinal crypt was increased, and was accompanied by up-regulated tumor necrosis factor (TNF)-α and interleukin (IL)-1β mRNA within 24 h of the first 5-FU injection. However, all of these measures were significantly lower after saireito administration. These results suggest that saireito attenuates 5-FU-induced intestinal mucositis. This action may come from the reduction of apoptosis in the intestinal crypt via suppression of the up-regulation of inflammatory cytokines. Therefore, saireito may be clinically useful for the prevention of intestinal mucositis during cancer chemotherapy.     

Amelioration of Chemotherapy-Induced Intestinal Mucositis by Orally Administered Probiotics in a Mouse Model

Background and Aims

Intestinal mucositis is a frequently encountered side effect in oncology patients undergoing chemotherapy. No well-established or up to date therapeutic strategies are available. To study a novel way to alleviate mucositis, we investigate the effects and safety of probiotic supplementation in ameliorating 5-FU-induced intestinal mucositis in a mouse model.

Methods

Seventy-two mice were injected saline or 5-Fluorouracil (5-FU) intraperitoneally daily. Mice were either orally administrated daily saline, probiotic suspension of Lactobacillus casei variety rhamnosus (Lcr35) or Lactobacillus acidophilus and Bifidobacterium bifidum (LaBi). Diarrhea score, pro-inflammatory cytokines serum levels, intestinal villus height and crypt depth and total RNA from tissue were assessed. Samples of blood, liver and spleen tissues were assessed for translocation.

Results

Marked diarrhea developed in the 5-FU groups but was attenuated after oral Lcr35 and LaBi administrations. Diarrhea scores decreased significantly from 2.64 to 1.45 and 0.80, respectively (P<0.001). Those mice in 5-FU groups had significantly higher proinflammatory cytokine levels (TNF-α: 234.80 vs. 29.10, P<0.001, IL-6: 25.13 vs. 7.43, P<0.001, IFN-γ: 22.07 vs. 17.06, P = 0.137). A repairing of damage in jejunal villi was observed following probiotics administration. We also found TNF-α, IL-1β and IL-6 mRNA expressions were up-regulated in intestinal mucositis tissues following 5-FU treatment (TNF-α: 4.35 vs. 1.18, IL-1β: 2.29 vs. 1.07, IL-6: 1.49 vs. 1.02) and that probiotics treatment suppressed this up-regulation (P<0.05). No bacterial translocation was found in this study.

Conclusions

In conclusion, our results show that oral administration of probiotics Lcr35 and LaBi can ameliorate chemotherapy-induced intestinal mucositis in a mouse model. This suggests probiotics may serve as an alternative therapeutic strategy for the prevention or management of chemotherapy-induced mucositis in the future.     

4th International Symposium on Autophagy: Exploiting the Frontiers of Autophagy Research

Minocycline has been shown to alleviate several neurological disorders. Unexpectedly, we found that minocycline had opposite effects on glioma cells: minocycline induced nonapoptotic cell death in glioma cells. The glioma cell death was associated with the presence of autophagic vacuoles in the cytoplasm. Minocycline induced autophagy was confirmed by acridine orange, monodansylcadaverine (MDC) stainings of vesicle formation and the conversion of microtubule-associated proteins light chain 3 (LC3-I) to LC3-II. Pretreatment with autophagy inhibitor 3-methyladenine (3-MA) suppressed the induction of acidic vesicular organelles and the accumulation of LC3-II to the autophagosome membrane in glioma cells treated with minocycline. Despite the pretreatment of 3-MA, minocycline induced cell death which could result from the activation of caspase-3. Minocycline effectively inhibited tumor growth and induced autophagy in the xenograft tumor model of C6 glioma cells. These results suggest that minocycline may kill glioma cells by inducing autophagic cell death. When autophagy was inhibited, minocycline still induced cell death through the activation of caspase-3. Thus, minocycline is a promising agent in the treatment of malignant gliomas.     

Minocycline inhibits the growth of glioma by inducing autophagy

Minocycline has been shown to alleviate several neurological disorders. Unexpectedly, we found that minocycline had opposite effects on glioma cells: minocycline induced nonapoptotic cell death in glioma cells. The glioma cell death was associated with the presence of autophagic vacuoles in the cytoplasm. Minocycline induced autophagy was confirmed by acridine orange, monodansylcadaverine (MDC) stainings of vesicle formation and the conversion of microtubule-associated proteins light chain 3 (LC3-I) to LC3-II. Pretreatment with autophagy inhibitor 3-methyladenine (3-MA) suppressed the induction of acidic vesicular organelles and the accumulation of LC3-II to the autophagosome membrane in glioma cells treated with minocycline. Despite the pretreatment of 3-MA, minocycline induced cell death which could result from the activation of caspase-3. Minocycline effectively inhibited tumor growth and induced autophagy in the xenograft tumor model of C6 glioma cells. These results suggest that minocycline may kill glioma cells by inducing autophagic cell death. When autophagy was inhibited, minocycline still induced cell death through the activation of caspase-3. Thus, minocycline is a promising agent in the treatment of malignant gliomas.     

Thymoquinone protects against 5-Fluorouracil-induced mucositis by NF-κβ and HIF-1 mechanisms in mice

Mucositis is among the most common side effects of 5-Fluorouracil (5-FU) and other cancer therapeutic drugs. Thymoquinone (TQ), a bioactive constituent extracted from Nigella sativa, has antioxidant and anti-inflammatory properties and can modify acute gastrointestinal injury. To investigate the effects of TQ on mucositis induced by 5-FU, studied animals were divided into four groups: control, 5-FU unit dose (300 mg/kg) to cause oral and intestinal mucositis (OM and IM), TQ (2.5 mg/kg) and TQ (2.5 mg/kg) plus 5-FU. Due to The molecular mechanisms, it was confirmed that the expression of NF-κβ and HIF-1 increases in OM. The serum levels of malondialdehyde (MDA), catalase (CAT), and superoxide dismutase (SOD), as well as pathological parameters, were assessed. Based on our results, the nuclear factor-kappa β gene expression in the tongue was downregulated significantly in the 5-FU + TQ compared to the 5-FU. TQ treatment can diminish MDA, and a reduction in oxidative stress was shown. TQ could also reduce the severity of tissue destruction and damaging effects induced by 5-FU on the tongue and intestine. We also observed lower villus length and width in the intestine of the 5-FU group compared to the control group. According to our research's pathological, biochemical, and molecular results, treatment with TQ as an anti-inflammatory and antioxidant compound may be the potential to improve and treat 5-FU-induced OM and IM, and TQ could be used against cancer treatment drugs and exhibit fewer adverse effects.     

Utidelone inhibits growth of colorectal cancer cells through ROS/JNK signaling pathway

Utidelone (UTD1), a novel microtubule stabilizing agent, is an epothilone B analogue which was produced by genetic engineering. UTD1 has exhibited broad antitumor activity in multiple solid tumors. However, its activity and mechanism in colorectal cancer (CRC) remain to be studied. In this study, UTD1 dramatically inhibited CRC cell proliferation (with 0.38 µg/ml, 0.77 µg/ml IC50 in RKO and HCT116, respectively) in vitro. Immunofluorescence staining showed that UTD1 induced the formation of microtubule bundling and asters in RKO cells. Flow cytometry analysis demonstrated that UTD1 induced cell cycle to arrest in G2/M phase, subsequent apoptosis. Significantly, UTD1 exhibited stronger effect on inducing apoptosis than paclitaxel and 5-FU, especially in HCT15 cells which is ABCB1 high-expression. UTD1 exposure cleaved caspase-3 and poly ADP-ribose polymerase (PARP), decreased mitochondrial membrane potential, released cytochrome c, increased the production of active oxygen and activated c-Jun N-terminal kinase (JNK), suggesting ROS/JNK pathway was involved in this process. Moreover, UTD1 inhibited tumor growth and was more effective and safer compared with paclitaxel and 5-FU in RKO xenograft in nude mice. Taken together, our findings first indicate that UDT1 inhibits tumor growth in CRC xenograft model and may be a promising agent for CRC treatment.Subject terms: Drug development, Preclinical research     

Inflammatory intestinal damage induced by 5-fluorouracil requires IL-4

Background5-Fluorouracil (5-FU) induces intestinal mucositis, which is characterized by epithelial ulcerations in the mucosa and clinical manifestations, such as pain and dyspeptic symptoms. Cytokines participate in the inflammatory and functional events of intestinal mucositis. IL-4 is an important mediator of intestinal inflammation, with either anti-inflammatory or pro-inflammatory functions, depending on the model of intestinal inflammation. This study aimed to evaluate the role of IL-4 in 5-FU-induced intestinal mucositis.MethodsIL-4^+/+ or IL-4^?/? mice (25–30 g) were intraperitoneally injected with 5-FU (450 mg/Kg) or saline (C). After 3 days, the mice were sacrificed and the duodenum was evaluated for epithelial damage, MPO activity and cytokine concentration.Results5-FU induced significant damage in the intestinal epithelium of IL-4^+/+ mice (reduction in the villus/crypt ratio: control = 3.31 ± 0.21 μm, 5-FU = 0.99 ± 0.10 μm). However, the same treatment did not induce significant damage in IL-4^?/? mice (5-FU = 2.87 ± 0.19 μm) compared to wild-type mice. 5-FU-induced epithelial damage increased the MPO activity (neutrophil number) and the level of pro-inflammatory cytokines (IL-4, TNF-α, IL-1β and CXCL-8) in the duodenum. These results were not observed in IL-4^?/? mice treated with 5-FU.ConclusionOur data suggest that IL-4 participates as a pro-inflammatory cytokine in a 5-FU-induced intestinal damage model and suggests that IL-4 antagonists may be novel therapeutics for this condition.     

Minocycline neuroprotects, reduces microglial activation, inhibits caspase 3 induction, and viral replication following Japanese encephalitis

Minocycline is broadly protective in neurological disease models featuring inflammation and cell death and is being evaluated in clinical trials. Japanese encephalitis virus (JEV) is one of the most important causes of viral encephalitis worldwide. There is no specific treatment for Japanese encephalitis (JE) and no effective antiviral drugs have been discovered. Studies indicate that JE involves profound neuronal loss as well as secondary inflammation caused because of cell death. Minocycline is a semisynthetic second-generation tetracycline that exerts anti-inflammatory and antiapoptotic effects that are completely separate from its antimicrobial action. Because tetracycline treatment is clinically well tolerated, we investigated whether minocycline protects against experimental model of JE. Intravenous inoculation of GP78 strain of JEV in adult mice results in lethal encephalitis and caused primarily because of neuronal death and secondary inflammation caused because of cell death. Minocycline confers complete protection in mice following JEV infection ( p  < 0.0001). Neuronal apoptosis, microglial activation, active caspase activity, proinflammatory mediators, and viral titer were markedly decreased in minocycline-treated JEV infected mice on ninth day post-infection. Treatment with minocycline may act directly on brain cells, because neuronal cell line Neuro2a were also salvaged from JEV-induced death. Our data suggest that minocycline may be a candidate to consider in human clinical trials for JE patients.     

A Novel Small Molecule FL118 That Selectively Inhibits Survivin,Mcl-1, XIAP and cIAP2 in a p53-Independent Manner,Shows Superior Antitumor Activity

Drug/radiation resistance to treatment and tumor relapse are major obstacles in identifying a cure for cancer. Development of novel agents that address these challenges would therefore be of the upmost importance in the fight against cancer. In this regard, studies show that the antiapoptotic protein survivin is a central molecule involved in both hurdles. Using cancer cell-based survivin-reporter systems (US 7,569,221 B2) via high throughput screening (HTS) of compound libraries, followed by in vitro and in vivo analyses of HTS-derived hit-lead compounds, we identified a novel anticancer compound (designated FL118). FL118 shows structural similarity to irinotecan. However, while the inhibition of DNA topoisomerase 1 activity by FL118 was no better than the active form of irinotecan, SN-38 at 1 µM, FL118 effectively inhibited cancer cell growth at less than nM levels in a p53 status-independent manner. Moreover, FL118 selectively inhibited survivin promoter activity and gene expression also in a p53 status-independent manner. Although the survivin promoter-reporter system was used for the identification of FL118, our studies revealed that FL118 not only inhibits survivin expression but also selectively and independently inhibits three additional cancer-associated survival genes (Mcl-1, XIAP and cIAP2) in a p53 status-independent manner, while showing no inhibitory effects on control genes. Genetic silencing or overexpression of FL118 targets demonstrated a role for these targets in FL118’s effects. Follow-up in vivo studies revealed that FL118 exhibits superior antitumor efficacy in human tumor xenograft models in comparison with irinotecan, topotecan, doxorubicin, 5-FU, gemcitabine, docetaxel, oxaliplatin, cytoxan and cisplatin, and a majority of mice treated with FL118 showed tumor regression with a weekly × 4 schedule. FL118 induced favorable body-weight-loss profiles (temporary and reversible) and was able to eliminate large tumors. Together, the molecular targeting features of FL118 plus its superior antitumor activity warrant its further development toward clinical trials.     

Minocycline causes widespread cell death and increases microglial labeling in the neonatal mouse brain

Minocycline, an antibiotic of the tetracycline family, inhibits microglia in many paradigms and is among the most commonly used tools for examining the role of microglia in physiological processes. Microglia may play an active role in triggering developmental neuronal cell death, although findings have been contradictory. To determine whether microglia influence developmental cell death, we treated perinatal mice with minocycline (45 mg/kg) and quantified effects on dying cells and microglial labeling using immunohistochemistry for activated caspase‐3 (AC3) and ionized calcium‐binding adapter molecule 1 (Iba1), respectively. Contrary to our expectations, minocycline treatment from embryonic day 18 to postnatal day (P)1 caused a > tenfold increase in cell death 8 h after the last injection in all brain regions examined, including the primary sensory cortex, septum, hippocampus and hypothalamus. Iba1 labeling was also increased in most regions. Similar effects, although of smaller magnitude, were seen when treatment was delayed to P3–P5. Minocycline treatment from P3 to P5 also decreased overall cell number in the septum at weaning, suggesting lasting effects of the neonatal exposure. When administered at lower doses (4.5 or 22.5 mg/kg), or at the same dose 1 week later (P10‐P12), minocycline no longer increased microglial markers or cell death. Taken together, the most commonly used microglial “inhibitor” increases cell death and Iba1 labeling in the neonatal mouse brain. Minocycline is used clinically in infant and pediatric populations; caution is warrented when using minocycline in developing animals, or extrapolating the effects of this drug across ages. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 753–766, 2017     

Chemotherapy-induced mucositis is associated with changes in proteolytic pathways

Mucositis, a common toxic side effect of chemotherapy, is characterized by an arrest of cell proliferation and a loss of gut barrier function, which may cause treatment reduction or withdrawal. Gut integrity depends on nutritional and metabolic factors, including the balance between protein synthesis and proteolysis. The effects of methotrexate (MTX; a frequently used chemotherapeutic agent) on intestinal proteolysis and gut barrier function were investigated in rats. Male Sprague-Dawley rats received 2.5 mg/kg of MTX subcutaneously during 3 days and were euthanized at Day 4 (D4) or Day 7 (D7). We observed at D4 that MTX induced mucosal damage and increased intestinal permeability (7-fold) and the mucosal concentration of interleukin (IL)-1beta and IL-6 (4- to 6-fold). In addition, villus height and glutathione content significantly decreased. Intestinal proteolysis was also affected by MTX as cathepsin D activity increased at D4, whereas chymotrypsin-like proteasome activity decreased and calpain activities remained unaffected. At D7, cathepsin D activity was restored to control levels, but proteasome activity remained reduced. This disruption of proteolysis pathways strongly contributed to mucositis and requires further study. Lysosomal proteolytic activity may be considered the main proteolytic pathway responsible for alteration of mucosal integrity and intestinal permeability during mucositis, as cathepsin D activity was found to be correlated with mucosal atrophy and intestinal permeability. Proteasome regulation could possibly be an adaptive process for survival. Future investigation is warranted to target proteolytic pathways with protective nutritional or pharmacological therapies during mucositis.     

Minocycline attenuates neuronal apoptosis and improves motor function after traumatic brain injury in rats

Minocycline is a type of tetracycline antibiotic with broad-spectrum antibacterial activity that has been demonstrated to protect the brain against a series of central nervous system diseases. However, the precise mechanisms of these neuroprotective actions remain unknown. In the present study, we found that minocycline treatment significantly reduced HT22 cell apoptosis in a mechanical cell injury model. In addition, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining confirmed the neuroprotective effects of minocycline in vivo through the inhibition of apoptosis in a rat model of controlled cortical impact (CCI) brain injury. The western blotting analysis revealed that minocycline treatment significantly downregulated the pro-apoptotic proteins BAX and cleaved caspase-3 and upregulated the anti-apoptotic protein BCL-2. Furthermore, the beam-walking test showed that the administration of minocycline ameliorated traumatic brain injury (TBI)-induced deficits in motor function. Taken together, these findings suggested that minocycline attenuated neuronal apoptosis and improved motor function following TBI.     

Tumor Grafting Induces Changes of Gut Microbiota in Athymic Nude Mice in the Presence and Absence of Medicinal Gynostemma Saponins

Recent findings have revealed that gut microbiota plays a substantial role in modulating diseases such as autism, rheumatoid arthritis, allergies, and cancer that occur at sites distant to the gut. Athymic nude mice have been employed for tumorigenic research for decades; however, the relationships between the gut microbiome and host’s response in drug treatment to the grafted tumors have not been explored. In this study, we analyzed the fecal microbiome of nonxenograft and xenograft nude mice treated with phytosaponins from a popular medicinal plant, Gynostemma pentaphyllum (Gp). Analysis of enterobacterial repetitive intergenic consensus (ERIC)-PCR data showed that the microbiota profile of xenograft mice departed from that of the nonxenograft mice. After ten days of treatment with Gp saponins (GpS), the microbiota of the treated mice was closer to the microbiota at Day 0 before the implantation of the tumor. Data obtained from 16S pyrosequencing of fecal samples reiterates the differences in microbiome between the nonxenograft and xenograft mice. GpS markedly increased the relative abundance of Clostridium cocleatum and Bacteroides acidifaciens, for which the beneficial effects on the host have been well documented. This study, for the first time, characterizes the properties of gut microbiome in nude mice responding to tumor implant and drug treatment. We also demonstrate that dietary saponins such as GpS can potentially regulate the gut microbial ecosystem by increasing the number of symbionts. Interestingly, this regulation of the gut ecosystem might, at least in part, be responsible for or contribute to the anticancer effect of GpS.     

Minocycline improves the functional recovery after traumatic brain injury via inhibition of aquaporin-4

Traumatic brain injury (TBI) is one of the main concerns worldwide as there is still no comprehensive therapeutic intervention. Astrocytic water channel aquaporin-4 (AQP-4) system is closely related to the brain edema, water transport at blood-brain barrier (BBB) and astrocyte function in the central nervous system (CNS). Minocycline, a broad-spectrum semisynthetic tetracycline antibiotic, has shown anti-inflammation, anti-apoptotic, vascular protection and neuroprotective effects on TBI models. Here, we tried to further explore the underlying mechanism of minocycline treatment for TBI, especially the relationship of minocycline and AQP4 during TBI treatment. In present study, we observed that minocycline efficaciously reduces the elevation of AQP4 in TBI mice. Furthermore, minocycline significantly reduced neuronal apoptosis, ameliorated brain edema and BBB disruption after TBI. In addition, the expressions of tight junction protein and astrocyte morphology alteration were optimized by minocycline administration. Similar results were found after treating with TGN-020 (an inhibitor of AQP4) in TBI mice. Moreover, these effects were reversed by cyanamide (CYA) treatment, which notably upregulated AQP4 expression level in vivo. In primary cultured astrocytes, small-interfering RNA (siRNA) AQP4 treatment prevented glutamate-induced astrocyte swelling. To sum up, our study suggests that minocycline improves the functional recovery of TBI through reducing AQP4 level to optimize BBB integrity and astrocyte function, and highlights that the AQP4 may be an important therapeutic target during minocycline treating for TBI.     

Subchronic methionine load induces oxidative stress and provokes biochemical and histological changes in the rat liver tissue

Melatonin has antitumor activity via several mechanisms including its anti-proliferative and pro-apoptotic effects. Moreover, it has been proven that melatonin in combination with chemotherapeutic agents enhances chemotherapy-triggered apoptosis in several types of cancer. Therefore, this study was intended to evaluate whether melatonin is able to strengthen the anti-cancer potential of different chemotherapeutic drugs in human colorectal adenocarcinoma HT–29 cells. We found that treatment with 20 µM cisplatin (CIS) or 1 mM 5-fluorouracil (5-FU) for 72 h induced a decrease in HT-29 cell viability. Furthermore, 1 mM melatonin significantly (P < 0.05) increased the cytotoxic effects of 5-FU. Likewise, simultaneous stimulation with 1 mM melatonin and 1 mM 5-FU significantly (P < 0.05) enhanced the ratio of cells with an overproduction of intracellular reactive oxygen species and substantially augmented the population of apoptotic cells compared to the treatment with 5-FU alone. Nonetheless, melatonin only displayed moderate chemosensitizing effects in CIS-treated HT-29 cells, as suggested by a slight increment in the fraction of early apoptotic cells that was observed only after 48 h. Consistently, co-stimulation of HT-29 cells with 20 µM CIS or 1 mM 5-FU in the presence of 1 mM melatonin further increased caspase-3 activation. Apart from this, the cytostatic activity displayed by CIS due to S phase arrest was not affected by concomitant stimulation with melatonin. Overall, our results indicate that melatonin increases the sensitivity of HT-29 cells to 5-FU treatment and, consequently, the indolamine could be potentially applied to colorectal adenocarcinoma treatment as a potent chemosensitizing agent.     

The effects of myeloablative or non-myeloablative total body irradiations on intestinal tract in mice

Acute radiation injury caused by high-dose radiation exposure severely impedes the application of radiotherapy in cancer management. To deeply understand the side effects of radiation on intestinal tract, an irradiation murine model was applied and evaluated. C57BL/6 mice were given 4 Gy non-myeloablative irradiation, 8 Gy myeloablative irradiation and non-irradiation (control), respectively. Results demonstrated that the 8 Gy myeloablative irradiations significantly damaged the gut barrier along with decreasing MECA32 and ZO-1. However, a slight increase in MECA32 and ZO-1 was detected in the 4 Gy non-myeloablative irradiations treatment from day 5 to day 10. Further, the irradiations affected the expression of P38 and JNK mitogen-activated protein kinase (MAPK) but not ERK1/2 MAPK signal pathway. Moreover, irradiation had adverse effects on hematopoietic system, altered the numbers and percentages of intestinal inflammatory cells. The IL-17/AhR had big increase in the gut of 4 Gy irradiation mice at day 10 compared with other groups. Both 8 Gy myeloablative and 4 Gy non-myeloablative irradiation disturbed the levels of short-chain fatty acids (SCFAs) in intestine. Meanwhile, high dosage of irradiation decreased the intestinal bacterial diversity and altered the community composition. Importantly, the fatty acids generating bacteria Bacteroidaceae and Ruminococcaceae played key roles in community distribution and SCFAs metabolism after irradiation. Collectively, the irradiation induced gut barrier damage with dosages dependent that led to the decreased p38 MAPK and increased JNK MAPK, unbalanced the mononuclear cells (MNCs) of gut, disturbed intestinal bacterial community and SCFAs level.