ABSTRACT
Objective@#To study the effect and mechanism of angiotensin (Ang II) on the proliferation of human hepatocellular carcinoma HepG2 cells.@*Methods@#The effects of different concentrations of Ang II's (10-8-10-4 mol/L) on proliferated hepatocellular carcinoma HepG2 cells were detected by CCK-8 assay. The expression of angiotensin II type 1 receptor (AT1) protein and activation of ERK1/2 protein in hepatocellular carcinoma HepG2 cells after processing with Ang II were assayed by Western blot. The cells were pretreated with candesartan (AT1 receptor antagonist), sorafenib (Raf kinase inhibitor) and PD98059 (ERK1/2 inhibitor) for 1.5 h and then Ang II (10-6 mol/L) was added. CCK-8 assay was used to determine whether it could reverse the proliferation of Ang II, and ERK phosphorylation levels were detected by Western blot. The changes in Bcl-2 and c-myc gene expression before and after Ang II processing were detected by Rt-PCR. According to different data, t-test, one-way analysis of variance or SNK method were used for statistical analysis.@*Results@#HepG2 cells treated with different concentrations of Ang II promoted cell proliferation after 24h and 48h. After 24 h, cell vitality was strongest with Ang II concentration 10-5 mol/L and the absorbance value was 0.990 8±0.097 8; and again after 48 h, the cell viability was strongest with Ang II concentration 10-6 mol/L and the absorbance value was 1.302 7 ± 0.030 9. Moreover, the pro-proliferation effect of Ang II on HepG2 cells blocked candesartan, sorafenib and ERK1/2 isolated inhibitors. After treatment with 10-6 mol/L Ang II, Western blot showed that Ang II significantly promoted AT1 receptor expression and phosphorylation of ERK1/2 protein confirmed that Ang II activated the AT1/RAF/ERK1/2 signaling pathway. In addition, Rt-PCR detection showed that the downstream of Bcl-2 and c-myc genes expressions rose significantly when the concentration of Ang II ranged from 10-8 to 10-6 mol/L.@*Conclusion@#Ang II can promote the proliferation of HepG2 cells by activating AT1/Raf /ERK1/2 signaling pathway and enhance the downstream of Bcl-2 and c-myc gene expression.
ABSTRACT
Objective To observe the effect of human adipose mesenchymal stem cells (hADMSCs) on pancreatic tissue repair and inflammatory reaction of acute necrotic pancreatitis (ANP) in rat, and explore the possible mechanism.Methods Isolation and purification of hADMSCs and flow cytometry to detect the the surface markers including CD90, CD29, CD34 and CD45 were performed.Eighty SD male rats with the body weight of 170~210 g were randomly divided into 4 groups.There were 8 rats in the control group, 24 rats in other group.Control group underwent no treatment;sham operation group underwent intestinal wall stirring and then abdominal closure;ANP model group was established by open abdominal retrograde injection of sodium taurocholate into bile duct;and in hADMSCs group, DAPI labeled hADMSCs were injected by tail vein into the rat at 12 h after sodium taurocholate injection.The survival of the rats, and gross morphological and pathological changes of the pancreas was observed at 12, 24, and 48 h, and the serum TNF-α, IL-6, IL-10 and amylase were detected.The distribution of hADMSCs in the pancreas, liver and lung was examined in hADMSCs group.Results Rats in control group and sham operation group were all alive.In ANP group, 5 and 11 rats were dead at 24 and 48 h, respectively, and in hADMSCs group 12 rats were dead at 48 h.Compared with ANP group, the difference was not statistically significant (P>0.05).The pathological changes of the pancreas were significantly less severe in hADMSCs group than in ANP group.In hADMSCs group, the amylase at 12, 24 and 48 h was(999±110 )、(1 831±110)、(3 991±130 )U/L;TNF-α level was (62.40±2.35), (80.51±4.51) and (93.46±6.60)ng /L;IL-6 was (60.46±7.34), (80.61±8.40) and(100.58±9.49)ng /L;and these were all significantly lower than those in ANP model group [amylase (2 402±146), (3 292±137) and (5 632±112)U/L;TNF-α(87.13±3.39), (105.41±10.06), (114.57±3.06)ng/L;IL-6 (70.67±10.90)、(107.61±10.53)、(145.34±10.48)U/L], and the differences were all statistically significant (all P<0.05).IL-10 in hADMSCs group was (56.63±6.35), (81.32±5.96), (100.26±6.51)ng/L, which were increased compared with those in ANP model group [(45.26±8.04), (68.25±8.42), (80.38±5.71)ng/L], and the difference was statistically significant (all P<0.05).hADMSCs can migrate to the pancreas, liver, lungs and other damaged tissue, with most in pancreatic tissue, less in lung tissue, and least in liver tissue.Conclusions The mechanism of hADMSCs in repairing pancreatic tissue injury was associated with inhibiting TNF-α and IL-6 secreting and increasing IL-10, thus reducing inflammatory reaction.