Mechanisms involved in the cytotoxic effects of berberine on human colon cancer HCT-8 cells

Berberine,a constituent of some traditional Chinese medicinal plants,has been reported to have cytotoxicity effects on different human cancer cell lines.There is no available information about the effects and mechanism of action of berberine on human colon cancer cell line HCT-8.In this paper,the cytotoxicity of berberine on HCT-8 cancer cells was investigated by MTT assay,fluorescence microscopy and flow cytometry analysis.Our data revealed that berberine could significantly inhibit the growth of HCT-8 cells in a dose-and time-dependent manner.Morphology of apoptotic cells was studied with acridine orange/ethidium bromide staining.The concentrations of lactate dehydrogenase and both acid and alkaline phos-phatases were significantly increased in cell supernatants after berberine treatment,suggesting cell death.Furthermore,flow cytometry analysis showed that berberine could arrest HCT-8 cells at S phase in a time-dependent manner.To further investigate the apoptotic molecular mechanism,reverse transcription-poly-merase chain reaction(RT-PCR)and western blotting methods were used.The up-regulated mRNA and/or protein expressions of Fas,FasL,TNF-α,caspase-3 and down-regulation of pro-caspase-3 suggest that the death receptor pathway may be involved in the apoptotic pathway induced by berberine.Decrease of Bcl-2 and increase of Bax in mRNA and/or protein expressions showed that the Bcl-2 family proteins were involved in berberine-induced apoptosis.We also found that berberine-induced apoptosis was associated with an up-regulated expressions of p53 and prohibitin(PHB),and decreased vimentin expression.These results suggest that berberine can suppress cell growth and reduce cell survival by arresting the cell-cycle and by inducing apoptosis of HCT-8 cells.

The antioxidant activity and hypolipidemic activity of the total flavonoids from the fruit of Rosa laevigata Michx

In the present work, the antioxidant activity in vitro and hypolipidemic activity of the total fla-vonoids (TFs) from the Rosa laevigata Michx fruit was evaluated, and the antioxidant effect in vivo was also discussed. The TFs exhibited a high scavenging effect on 2, 2-diphenyl-1- picrylhydrazyl (?DPPH) with IC50 values 0.01 mg/mL, and a stong reduce power in the test. Hyperli-pemic mice were intragastric administrated with TFs (25, 50 mg/kg/day) for 4 weeks, and fenofi-brate was used as the positive reference sub-stance. After the experiment, the levels of TC (total cholesterol), TG (triglyceride), LDL- C (low density lipoprotein-cholesterol) of the mice ad-ministrated with high-dose of TFs were mark-edly declined by 45.02%, 33.86% and 73.68%, re-spectively, while HDL-C (high density lipopro-tein-cholesterol) was significantly increased com-pared with model group. To investigate the hepatoprotective effect, histopathological assay, ALT (alanine aminotransferase), AST (aspartate aminotransefrase) and ALP (alkaline phosphatase) were also studied, and the results showed that TFs exhibited a favorable effect on liver protec-tion, of which the levels of ALT, AST and ALP were elevated by 55.85%, 29.15% and 25.68%, respectively. Furthermore, the TFs could sig-nificantly decrease the MDA (malondialdehyde) level and improve the levels of CAT (Catalas), SOD (superoxide dismutase), GSH (reduced glutathione), and GPX (glutathione peroxidase) compared with hyperlipemia mice. Our results suggested that TFs has a high antioxidant ac-tivity and hypolipidemic activity, which can be used as a potential medicine for cardiovascular diseases.

LncRNA AFAP1-AS1/miR-27b-3p/VEGF-C axis modulates stemness characteristics in cervical cancer cells

Background:Long non-coding RNA(lncRNA)actin filament-associated protein 1 antisense RNA 1(AFAP1-AS1)functions as a competing endogenous RNA to regulate target genes expression by sponging microRNAs(miRs)to play cancer-promoting roles in cancer stem cells.However,the regulatory mechanism of AFAP1-AS1 in cervical cancer(CC)stem cells is unknown.The present study aimed to provide a new therapeutic target for the clinical treatment of CC.Methods:Hyaluronic acid receptor cluster of differentiation 44 variant exon 6(CD44v6)(+)CC cells were isolated by flow cytometry(FCM).Small interfering RNAs of AFAP1-AS1(siAFAP1-AS1)were transfected into the(CD44v6)(+)cells.The levels of AFAP1-AS1 were measured by quantitative real-time PCR(qRT-PCR).Sphere formation assay,cell cycle analysis,and Western blotting were used to detect the effect of siAFAP1-AS1.RNA pull-down and luciferase reporter assay were used to verify the relationship between miR-27b-3p and AFAP1-AS1 or vascular endothelial growth factor(VEGF)-C.Results:CD44v6(+)CCcells had remarkable stemness and a high level ofAFAP1-AS1.However,AFAP1-AS1knockdownwithsiAFAP1-AS1suppressed the cell cycle transitionofG(1)/S phase and inhibited self-renewal ofCD44v6(+)CCcells,the levels of the stemnessmarkers octamer-binding transcription factor 4(OCT4),osteopontin(OPN),and cluster of differentiation 133(CD133),and the epithelialmesenchymal transition(EMT)-related proteins Twist1,matrix metalloprotease(MMP)-9,and VEGF-C.In the mechanism study,miR-27b-3p/VEGF-C signaling was demonstrated to be a key downstream of AFAP1-AS1 in the CD44v6(+)CC cells.Conclusions:LncRNA AFAP1-AS1 knockdown inhibits the CC cell stemness by upregulating miR-27b-3p to suppress VEGF-C.

Hypoglycemic effects of Tibetan medicine Huidouba in STZ-induced diabetic mice and db/db mice

Objective:Huidouba(HDB) is a Chinese folk medicine used to treat diabetes in Sichuan Province,China.Therefore,we investigated the anti-diabetic effects of HDB and its underlying mechanisms.We hypothesized that HDB treatment could enhance glucose tolerance and insulin sensitivity,and thus prevent a hyperglycemia state.Methods:To test the hypothesis,streptozotocin(STZ)-induced diabetic mice and db/db mice,widely used models of hyperglycemia and insulin-resistant diabetes,were either treated with HDB,metformin,or acarbose.Blood glucose,oral glucose tolerance test,insulin tolerance test,pancreatic histopathology and serum biochemistry were detected to assess the hypoglycemic effect of HDB.Results:HDB treatments were found to show the effect in reducing glucose levels.HDB also resulted in a significant reduction in body weight and food intake in the STZ-induced diabetic mouse model.Furthermore,it significantly improved glucose and insulin tolerance in the two diabetic mouse models.Importantly,insulin,glucagon,pancreatic polypeptide,and somatostatin immunohistochemistry revealed that HDB treatment improved the function and the location of the cells in the islets compared with the other two treatments.HDB treatment resulted in significant restoration of islet function.Our results illustrated the underlying mechanism of HDB in the progression of diabetes,and HDB can be an effective agent for the treatment of diabetes.Conclusion:The results of this study suggested that HDB can reduce blood glucose levels in STZ-induced hyperglycemic mice and db/db mice.