Ursodeoxycholic acid alleviates experimental liver fibrosis involving inhibition of autophagy.

AIMS: Ursodeoxycholic acid (UDCA) has been widely used in the treatment of primary biliary cholangitis (PBC) with chronic liver fibrosis, but its detailed mechanism remains unclear. This study was aimed to determine whether autophagic signaling is involved in the therapeutic effect of UDCA on liver fibrosis. METHODS: By using hepatic stellate cell (HSC) line LX2 and CCl4-induced fibrotic rat model, autophagy signaling was investigated by western blotting and mRFP-EGFP-LC3 tandem fluorescent tagged plasmid (ptfLC3) transfection technique. Anti-fibrotic profile was determined by western blotting, qRT-PCR, MTT assay, trypan blue, hydroxyproline assay and Masson staining. KEY FINDINGS: TGFß1 treatment decreased P62 accumulation and increased both autophagosomes and autolysosomes in LX2 cells, thereby elevated autophagic flux. Hydroxychloroquine (HCQ), antagonist of autophagy, was found to dramatically inhibit COL1A2 mRNA expression and cell proliferation in a dose-dependent manner. This coincides with the effect of UDCA intervention on collagen aggradation and cell viability. Meanwhile, UDCA inhibited TGFß1-induced autophagy flux. And rapamycin, agonist of autophagy, was found to impair the anti-fibrotic effect of UDCA. Moreover, study in vivo showed that UDCA alone or in combination with HCQ restored the CCl4-induced liver fibrosis in rodent models with autophagy inhibited profile. SIGNIFICANCE: Taken together, our study revealed that UDCA displays anti-fibrotic role by protecting HSC against production of collagen and inhibiting cellular viability involving autophagy inhibition and provide a new insight into the pharmacological basis of UDCA treatment for hepatic fibrosis.

Association between NCF4 rs4821544T/C polymorphism and inflammatory bowel disease risk in Caucasian: a meta-analysis.

OBJECTIVE: Published studies on the association between NCF4 rs4821544T/C polymorphism and inflammatory bowel disease (IBD) risk in Caucasian have yielded conflicting results. The present study aimed to provide more reliable conclusions by conducting a meta-analysis. METHODS: All eligible studies were extracted from Wiley Online Library, Chinese National Knowledge Infrastructure and PubMed databases. Odds ratios (ORs) with 95 % confidence intervals (CIs) were used to assess the associations between rs4821544T/C polymorphism and IBD risk in Caucasian. RESULTS: A total of 13 case-control studies comprising 7441 Crohn's disease (CD) patients, 2565 ulcerative colitis (UC) patients and 8315 controls were included in this meta-analysis. Significant associations were found between CD and the rs4821544T/C polymorphism in three genetic models (C vs T: OR = 1.11, 95 % CI: 1.06, 1.16, P = 0.000; CC vs TT: OR = 1.31, 95 % CI: 1.18, 1.45, P = 0.000; CC/TC vs TT: OR = 1.07, 95 % CI: 1.01, 1.13, P = 0.014; CC vs TC/TT: OR = 1.28, 95 % CI: 1.16, 1.42, P = 0.000). However, significant associations were not found in UC under any genetic models (C vs T: OR = 1.04, 95 % CI: 0.97, 1.11, P = 0.264; CC vs TT: OR = 1.10, 95 % CI: 0.93, 1.30, P = 0.284; TC vs TT: OR = 1.04, 95 % CI: 0.95, 1.13, P = 0.429; CC/TC vs TT: OR = 1.04, 95 % CI: 0.95, 1.13, P = 0.390; CC vs TC/TT: OR = 1.07, 95 % CI: 0.91, 1.26, P = 0.409). CONCLUSION: This meta-analysis suggested that the rs4821544T/C polymorphism was associated with CD, but not UC in Caucasian.

Autophagy inhibition contributes to the synergistic interaction between EGCG and doxorubicin to kill the hepatoma Hep3B cells.

(-)-Epigallocatechin-3-O-gallate(EGCG), the highest catechins from green tea, has promisingly been found to sensitize the efficacy of several chemotherapy agents like doxorubicin (DOX) in hepatocellular carcinoma (HCC) treatment. However, the detailed mechanisms by which EGCG augments the chemotherapeutic efficacy remain unclear. Herein, this study was designed to determine the synergistic impacts of EGCG and DOX on hepatoma cells and particularly to reveal whether the autophagic flux is involved in this combination strategy for the HCC. Electron microscopy and fluorescent microscopy confirmed that DOX significantly increased autophagic vesicles in hepatoma Hep3B cells. Western blot and trypan blue assay showed that the increasing autophagy flux by DOX impaired about 45% of DOX-induced cell death in these cells. Conversely, both qRT-PCR and western blotting showed that EGCG played dose-dependently inhibitory role in autophagy signaling, and that markedly promoted cellular growth inhibition. Amazingly, the combined treatment caused a synergistic effect with 40 to 60% increment on cell death and about 45% augmentation on apoptosis versus monotherapy pattern. The DOX-induced autophagy was abolished by this combination therapy. Rapamycin, an autophagic agonist, substantially impaired the anticancer effect of either DOX or combination with EGCG treatment. On the other hand, using small interference RNA targeting chloroquine autophagy-related gene Atg5 and beclin1 to inhibit autophagy signal, hepatoma cell death was dramatically enhanced. Furthermore, in the established subcutaneous Hep3B cells xenograft tumor model, about 25% reduction in tumor growth as well as 50% increment of apoptotic cells were found in combination therapy compared with DOX alone. In addition, immunohistochemistry analysis indicated that the suppressed tendency of autophagic hallmark microtubule-associated protein light chain 3 (LC3) expressions was consistent with thus combined usage in vitro. Taken together, the current study suggested that EGCG emerges as a chemotherapeutic augmenter and synergistically enhances DOX anticancer effects involving autophagy inhibition in HCC.

Expression of the Pokemon proto-oncogene in nasopharyngeal carcinoma cell lines and tissues.

To study the differentiated expression of the proto-oncogene Pokemon in nasopharyngeal carcinoma (NPC) cell lines and tissues, mRNA and protein expression levels of CNE1, CNE2, CNE3 and C666-1 were detected separately by reverse transcription polymerase chain reaction (RT-PCR), real-time PCR and Western-blotting. The immortalized nasopharyngeal epithelial cell line NP69 was used as a control. The Pokemon protein expression level in biopsy specimens from chronic rhinitis patients and undifferentiated non keratinizing NPC patients was determined by Western-blotting and arranged from high to low: C666-1>CNE1>CNE2> CNE3>NP69. The Pokemon mRNA expression level was also arranged from high to low: CNE1>CNE2>NP69>C666-1>CNE3. Pokemon expression of NP69 and C666-1 obviously varied from mRNA to protein. The Pokemon protein level of NPC biopsy specimens was obviously higher than in chronic rhinitis. The data suggest that high Pokemon protein expression is closely associated with undifferentiated non-keratinizing NPC and may provide useful information for NPC molecular target therapy.