Inhibitor of Differentiation-2 Protein Ameliorates DSS-Induced Ulcerative Colitis by Inhibiting NF-κB Activation in Neutrophils.

The loss of inhibitor of differentiation-2 (ID2) could lead to the development of colitis in mice, supplementation with exogenous ID2 protein might be a potential strategy to ameliorate colitis. In this study, the effects of ID2 protein supplementation on Dextran sodium sulfate (DSS)-induced colitis were investigated. Firstly, we confirmed that the expression of ID2 was reduced in the colon tissues of DSS-induced colitis mice and patients with ulcerative colitis (UC). Then, we constructed a recombinant plasmid containing the human Id2 gene and expressed it in Escherichia coli (E. coli) successfully. After purification and identification, purified hID2 could ameliorate DSS-induced colitis efficiently in mice by improving disease symptoms, decreasing the levels of proinflammatory cytokines in colon tissues, maintaining the integrity of intestinal barrier and reducing the infiltration of neutrophils and macrophages in the colon. Further study showed that hID2 could be endocytosed efficiently by neutrophils and macrophages, and hID2 lost its protection function against colitis when neutrophils were depleted with an anti-Gr-1 antibody. hID2 decreased the mRNA levels of IL-6, IL-1ß and TNF-α in lipopolysaccharides (LPS)-stimulated neutrophils and efficiently inhibited the activation of NF-κB signalling pathway in neutrophils. Interestingly, hID2 showed a synergistic role in inhibition of NF-κB activation with pyrrolidine dithiocarbamic acid (PDTC), an inhibitor of NF-κB activation. Therefore, this study demonstrated the potential use of hID2 to treat UC, and hID2 protein might be a promising anti-inflammatory agent that targets the NF-κB signalling pathway in neutrophils.

Saikosaponin-d ameliorates dextran sulfate sodium-induced colitis by suppressing NF-κB activation and modulating the gut microbiota in mice.

Saikosaponin-d (SSd), extracts from Bupleurum falcatum L, exhibits anti-inflammatory and anti-infectious activities. However, the effect of SSd on intestinal inflammation has not been investigated. The aim of this study was to evaluate the effect of SSd on dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) mice, and to elucidate the underlying mechanisms. UC was induced in mice by administrating 3% DSS in drinking water for 7 days. SSd (4 mg/kg and 8 mg/kg) was administered by gavage every day during the experimental process. The results showed that SSd treatment (8 mg/kg) significantly ameliorated UC mice by decreasing disease activity index (DAI), increasing colon length and improving pathological characteristics. SSd treatment (8 mg/kg) significantly suppressed the mRNA levels of pro-inflammatory cytokines including TNF-α, IL-6 and IL-1ß, increased that of anti-inflammatory cytokine IL-10. Furthermore, SSd (8 mg/kg) suppressed the activation of NF-κB by decreasing the degradation and phosphorylation of IκB. SSd (8 mg/kg) also protected the intestinal barrier by increasing the mRNA levels of mucin (Muc1 and Muc2) and the protein levels of zonula occludens-1 (ZO-1) and Claudin-1. The 16S rDNA gene high-throughput sequencing revealed that SSd treatment (8 mg/kg) increased the alpha diversity and regulated the structure of gut microbiota in UC mice. Taken together, our findings demonstrated that SSd (8 mg/kg) improved DSS-induced intestinal inflammation by inhibiting NF-κB activation and regulated the gut microbiota.

Chitooligosaccharides Prevents the Development of Colitis-Associated Colorectal Cancer by Modulating the Intestinal Microbiota and Mycobiota.

Gut microbes play a crucial role in the development of colorectal cancer. Chitooligosaccharides (COS), are oligomer that are depolymerized from chitosan and possess a wide range of biological activities. In this study, the effects of COS on colorectal cancer (CRC) development were evaluated using azoxymethane and dextran sulfate sodium (AOM/DSS) induced mouse model of CRC (CACM). In the COS-treated CRC group (CMCOS), COS protected mice from CRC by decreasing the disease activity index, tumor incidences and multiplicity, and the mRNA levels of COX-2, IL-6, TNF-α, IL-1ß, IL-10, and IKK-ß mRNA in colonic epithelial cells. The results of a cage-exchanged experiment, in which mice from the CACMe and CMCOSe treatments exchanged cages every day to interact with microbes, showed that gut microbes play an important role in preventing CAC by COS. The abundances of fecal bacteria (total bacteria, Lactobacillus, Enterococcus, Fusobacterium nucleatum and butyrate-producing bacteria) were detected by qPCR on the 0th, 1st, 3rd, 6th, 9th, and 10th weekends. Furthermore, microbiota and mycobiota were analyzed by high-throughput sequencing on an Illumina MiSeq PE300 system. COS protected mice from CRC by reversing the imbalance of bacteria and fungi, especially by reducing the abundance of Escherichia-Shigella, Enterococcus, and Turicibacter, and increasing the levels of Akkermansia, butyrate-producing bacteria and Cladosporium.

Expression of lncRNA PVT1 in colorectal cancer tissues and cells and its effect on chemo-sensitivity to cisplatin and the possible mechanisms / 中国肿瘤生物治疗杂志

@#Objective: To study the regulatory effects and possible mechanism of long non-coding RNA plasmacytoma variant translocation 1 (lncRNA PVT1) on chemotherapy sensitivity to cisplatin (DDP) of colorectal cancer (CRC)．Methods: A total of 112 pairs of matched cancer and adjacent non-cancerous tissues were obtained from the CRC patients who underwent surgical resection in the First Affiliated Hospital of Xinxiang Medical University betweenApril 2006 and March 2011.All specimens were confirmed by pathological examinations. Tumor tissues and corresponding adjacent non-cancerous tissues from 30 cisplatin-sensitive CRC patients and 30 cisplatin-resistant patients were selected. Human CRC cell lines (HT29, SW480, HCT116, RKO and LoVo) and normal colonic epithelial cell line NCM460 were also collected for this study; and DDP-resistant RKO/DDP and LoVo/DDP cell lines were constructed. siPVT1, siNC, LV-PVT1 and LV-NC were transfected into LoVo and RKO cells or LoVo/DDP and RKO/DDP cells using lipofectamineTM2000. The expression of lncRNA PVT1 in CRC tissues and cells was tested by Real-time qPCR. CCK-8 assay, flow cytometry and WB were performed to test the effect of PTV1 knockout or enforcement on cell proliferation, apoptosis and expressions of apoptosis-related proteins, respectively. The CRC subcutaneous transplanted xenograft model was established on athymic nude mice to study the effect of PVT1 over-expression on tumor growth and DDP resistance. Results: PVT1 was highly expressed in the cancer tissues and CRC cells, and its expression was positively associated with cisplatin resistance of CRC. After knockdown of PVT1, the proliferation of cisplatinresistant CRC cells was significantly suppressed, while the apoptosis was significantly enhanced (P<0.05 or P<0.01); Mechanically, the levels of drug resistance-associated molecules, including MDR1 and MRP1, as well as the expression of anti-apoptotic Bcl-2 were significantly downregulated whereas the levels of pro-apoptotic Bax and cleaved caspase-3 were increased in PVT1-silenced DDP-resistant CRC cells. Over-expression of PVT1 reversely increased proliferation and decreased apoptosis of CRC cells (P<0.05 or P<0.01). In addition, PVT1 over-expression in CRC cells significantly promoted DDP-resistance in vivo (P<0.05). Conclusion: Collectively, knockdown of PVT1 expression can significantly suppress cell proliferation and promote apoptosis of DDP-resistant CRC cells. Overexpression of PVT1 can significantly promote the growth of CRC cells in vitro and transplanted xenograft in vivo. PVT1 regulates endogenous apoptosis pathways and further promotes the sensitivity of CRC cells to cisplatin chemotherapy via inhibiting the expressions of MDR1 and MRP1.

Interplay between long noncoding RNA ZEB1-AS1 and miR-101/ZEB1 axis regulates proliferation and migration of colorectal cancer cells.

Long noncoding RNAs (lncRNAs) are dysregulated in many diseases. MicroRNA-101 (miR-101) functions as a tumor suppressor by directly targeting ZEB1 in various cancers. However, the potential mechanism of lncRNA ZEB1-AS1 and miR-101/ZEB1 axis in CRC remains unknown. In this study, we further investigated the potential interplay between miR-101/ZEB1 axis and lncRNA ZEB1-AS1 in colorectal cancer (CRC). Results showed that ZEB1-AS1 was upregulated in CRC tissues and cells. MiR-101 was downregulated in CRC tissues and negatively correlated with ZEB1-AS1 and ZEB1 expression levels in CRC. Functional experiments showed that, consistent with ZEB1-AS1 depletion, miR-101 overexpression and ZEB1 depletion inhibited the proliferation and migration of CRC cells. Overexpression of miR-101 partially abolished the effects of ZEB1-AS1 on the proliferation and migration of these cells. Moreover, combined ZEB1-AS1 depletion and miR-101 overexpression significantly inhibited cell proliferation and migration of the CRC cells. Hence, ZEB1-AS1 functioned as a molecular sponge for miR-101 and relieved the inhibition of ZEB1 caused by miR-101. This study revealed a novel regulatory mechanism between ZEB1-AS1 and miR-101/ZEB1 axis. The interplay between ZEB1-AS1 and miR-101/ZEB1 axis contributed to the proliferation and migration of CRC cells, and targeting this interplay could be a promising strategy for CRC treatment.

Silencing long noncoding RNA PVT1 inhibits tumorigenesis and cisplatin resistance of colorectal cancer.

Long noncoding RNA plasmacytoma variant translocation 1 (PVT1) plays pivotal roles in tumorigenesis of many cancers, including colorectal cancer (CRC). However, the clinical significance and the biological functions of PVT1 in CRC remain largely unknown. In this study, we found that PVT1 was highly expressed in CRC tissues and cell lines compared with the corresponding non-cancerous samples and normal colon epithelial cells. Clinically, increased expression of PVT1 was positively correlated with tumor size, advanced histological stages, metastases, poor prognosis, and cisplatin resistance of CRC patients. In vitro studies showed that PVT1 silencing inhibited the proliferation, migration, invasion, and apoptosis escape of CRC cells. Knockdown of PVT1 in cisplatin-resistant CRC cells induced proliferation inhibition and apoptosis, whereas overexpression of PVT1 increased proliferation and decreased apoptosis of CRC cells. Mechanically, the levels of drug resistance-associated molecules, including multidrug resistance 1 and multidrug resistance protein 1, as well as the expression of anti-apoptotic Bcl-2 were significantly downregulated whereas the levels of pro-apoptotic Bax and cleaved caspase-3 were increased in PVT1-silenced cisplatin-resistant CRC cells. However, ectopic expression of PVT1 in CRC cells reversed the expressions of the molecules mentioned above. In addition, PVT1 overexpression in CRC cells significantly promoted cisplatin resistance in vivo. Collectively, these results demonstrated that PVT1 is a significant regulator in tumorigenesis and cisplatin resistance of CRC and provided evidence that PVT1 may be a promising target for CRC therapy.

Suppression of microRNA-144-3p attenuates oxygen-glucose deprivation/reoxygenation-induced neuronal injury by promoting Brg1/Nrf2/ARE signaling.

Accumulating evidence has reported that microRNA-144-3p (miR-144-3p) is highly related to oxidative stress and apoptosis. However, little is known regarding its role in cerebral ischemia/reperfusion-induced neuronal injury. Herein, our results showed that miR-144-3p expression was significantly downregulated in neurons following oxygen-glucose deprivation and reoxygenation (OGD/R) treatment. Overexpression of miR-144-3p markedly reduced cell viability, promoted cell apoptosis, and increased oxidative stress in neurons with OGD/R treatment, whereas downregulation of miR-144-3p protected neurons against OGD/R-induced injury. Brahma-related gene 1 (Brg1) was identified as a potential target gene of miR-144-3p. Moreover, downregulation of miR-144-3p promoted the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and increased antioxidant response element (ARE) activity. However, knockdown of Brg1 significantly abrogated the neuroprotective effects of miR-144-3p downregulation. Overall, our results suggest that miR-144-3p contributes to OGD/R-induced neuronal injury in vitro through negatively regulating Brg1/Nrf2/ARE signaling.

microRNA-9 functions as a tumor suppressor in colorectal cancer by targeting CXCR4.

MicroRNAs (miRs) dysregulation has been proven to play a crucial role in the initiation and progression of colorectal cancer (CRC). miR-9 functions as a tumor suppressor in many cancer types, including CRC. However, the precise role of miR-9 and the underlying molecular mechanisms that miR-9 involves in CRC progression remain largely unknown. In this study, it was reported that miR-9 had lower expression in CRC tissue samples than in those matched adjacent non-tumor tissues. Deregulated miR-9 expression was inverse correlated with the TNM stage, lymph node metastasis, and prognosis of CRC patients. Ectopic miR-9 expression suppressed CRC cell proliferation, migration, and invasion. Dual-Luciferase Reporter Assay confirmed that C-X-C Motif Chemokine Receptor 4 (CXCR4) was a direct miR-9 target, and the effects of miR-9 were mimicked through CXCR4 depletion in vitro. CXCR4 rescue experiments further verified that CXCR4 is a functional target of miR-9. Animal xenograft assays also provided evidence that miR-9 functions as a tumor suppressor via targeting CXCR4 in vivo. Mechanistically, miR-9 overexpression or CXCR4 knockdown influenced cell proliferation and epithelial-mesenchymal transition (EMT). Results suggest that miR-9 acts as a tumor suppressor in CRC progression by regulating CXCR4.

TXNIP overexpression suppresses proliferation and induces apoptosis in SMMC7221 cells through ROS generation and MAPK pathway activation.

Thioredoxin binding protein (thioredoxin-interacting protein, TXNIP), known as vitamin D3 increase protein 1, has been identified as a tumor suppressor in various cancers such as pancreatic, breast, lung and thyroid cancer. However, the role of TXNIP in hepatocellular carcinoma cell proliferation and apoptosis remains unknown. In this study, we first used qRT-PCR, western blotting and immunohistochemistry to compare the expression of TXNIP between hepatocellular carcinoma tissues and tumor-adjacent normal liver tissues. In vitro, we explored the role of TXNIP in hepatocellular carcinoma progression via transfection of the pcDNA-3.1-TXNIP plasmid into SMMC7221 cells. Our results showed that the expression of TXNIP was significantly decreased in hepatocellular carcinoma tissues. Moreover, TXNIP over-expression inhibited hepatocellular carcinoma cell proliferation and induced apoptosis by triggering mitochondrial-mediated ROS generation and activating MAPK pathways. This study provides insight into the molecular mechanisms of TXNIP overexpression in liver cancer cell survival and apoptosis and indicated that TXNIP may be a novel promising agent for liver cancer treatment.

NOR1 promotes hepatocellular carcinoma cell proliferation and migration through modulating the Notch signaling pathway.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide. Previous studies have reported that the oxidored-nitro domain containing protein 1 (NOR1) is a novel tumor suppressor in several tumors. Recent evidence suggests that NOR1 is strongly expressed in HCC cells. However, its role and mechanism in HCC are unclear. In the current study, Western blot and qPCR detected strong NOR1 mRNA and protein expression in HepG2 and Hep3B cells. After transfection with NOR1 siRNA or pcDNA3.1-myc-his-NOR1, the proliferation and migration of HepG2 and Hep3B cells were analyzed in vitro. HepG2 or Hep3B cells overexpressing NOR1 showed an increased proliferation and migration, whereas siRNA-mediated silencing of NOR1 showed the opposite effect. Furthermore, NOR1 activated the Notch signaling pathway, indicated by increased levels of Notch1, NICD, Hes1, and Hey1 in protein. Importantly, the Notch inhibitor DAPT downregulated Notch activation and further enhanced siNOR1-induced reduction of cell proliferation and migration in HepG2 and Hep3B cells, whereas DAPT reversed the effect of NOR1 overexpression on cell proliferation and migration. In conclusion, these results indicate that NOR1 may be involved in the progression of HCC and thus may be a potential target for the treatment of liver cancer.