Contributions of cardiotrophin-1 to cardiac fibroblast proliferation

AIM: To investigated the role of CT-1 in the hypertensive ventricular remodeling. METHODS: The cardiac fibroblasts in 3-4 passages were cultured in vitro, and divided into eight groups according to the different intervention factors: group C (control); group DM: dimethyl sulphoxide (DMSO); group P: cultured under high hydrostatic pressure (160 mmHg); group ASODN: interfered with CT-1 antisense oligodeoxynucleotides (10 μmol/L); group SODN: incubated with CT-1 sense oligodeoxynucleotides (10 μmol/L); group AG: interfered with AG490 (25 μmol/L); group PD: interfered with PD98059 (20 μmol/L); group LY: interfered with LY 294002 (10 μmol/L). Western blotting was employed to assess the expression of STAT3, ERK1/2 and PI3-K respectively at protein level. Cell proliferation was quantified by MTT. RESULTS: High hydrostatic pressure stimulated the proliferation of cardiac fibroblasts, upregulated the expression of CT-1. CT-1ASODN inhibited the proliferation of cardiac fibroblasts (0.132±0.013 vs 0.154±0.011, P<0.05). ASODN extensively inhibited the expression of STAT3, ERK1/2 and PI3-k respectively at protein level (2.09±0.25 vs 2.47±0.28, P<0.05), (1.13±0.19 vs 1.61±0.22, P<0.05), (1.25±0.23 vs1.71±0.25, P<0.05). AG490, a JAK-STAT3 inhibitor, reversed the increase in the proliferation of cardiac fibroblasts stimulated by high hydrostatic pressure and the expression of ERK1 phosphorylation (0.118±0.018 vs 0.155±0.010, P<0.05). PD98059, a MAPK-ERK1/2 inhibitor, increased the proliferation of cardiac fibroblasts stimulated by high hydrostatic pressure (0.185±0.011 vs 0.155±0.010, P<0.05) and the expression of STAT3 phosphorylation (1.83±0.23 vs 1.58±0.22, P<0.05). LY294002, a PI3-K inhibitor, had no effect on the proliferation of cardiac fibroblasts stimulated by high hydrostatic pressure (0.157±0.015 vs 0.155±0.010, P>0.05). No difference of the expression of the above factors was observed in SOND (0.151±0.010 vs 0.154±0.011, P>0.05) and DMSO (0.141±0.017 vs 0.155±0.010, P>0.05) groups as compared with the control group. CONCLUSION: Under high hydrostatic pressure, the proliferation of cardiac fibroblasts is essentially mediated by STAT3, independent of PI3-K and the action is negatively regulated by ERK1/2 via inhibiting STAT3.The interaction between STAT3 and ERK1/2 may assist CT-1 in developing adequate cardiac fibroblast proliferation.     

Effect of Qingyi decoction on p-STAT3 expression in pancreas of mice with severe acute pancreatitis induced by L-arginine

AIM: To explore the role of STAT3 signaling pathway in acute pancreatitis induced by L-arginine, and the mechanisms of Qingyi decoction(QYD) treatment on severe acute pancreatitis (SAP). METHODS: The Kunming mice were randomly divided into 3 groups (n=10): control group, SAP group and QYD treatment group (SAP+QYD). The mice in SAP group and SAP+QYD group were intraperitoneally injected with 20% L-arginine (3 g/kg, bid). The mice in SAP+QYD group were also administered intragastrically with QYD(10 mL/kg) 30 min after the second injection of 20% L-arginine and twice a day for the following 2 days. The mice were anesthetized and sacrificed 72 h after SAP induction. The activity of amylase was measured in serum, the relative pancreatic weight was assayed, and the activity of myeloperoxidase (MPO) was analyzed to evaluate the neutrophil infiltration in lung tissues. The morphology of pancreas and lung was observed. The protein of pancreas was extracted to detect the expression of p-STAT3 by Western blotting. The mRNA expression of monocyte chemoattractant protein-1(MCP-1) was determined by real-time PCR. RESULTS: Compared with control group at 72 h, L-arginine induced SAP with increased serum amylase activity, pancreatic wet weight ratio and MPO activity in lung tissues (P<0.05). In SAP+QYD group, the activity of amylase, pancreatic wet weight ratio and MPO levels were significantly lower than those in SAP group (P<0.01). Compared with control group at 72 h, the pancreas and lung were obvious injured, the protein level of p-STAT3 and mRNA expression of MCP-1 in pancreas tissues increased significantly in SAP group. Compared with SAP group, the pathologic damage of the pancreas and lung tissues, the protein level of p-STAT3 and mRNA expression of MCP-1 in pancreas were significantly reduced in SAP+QYD group. CONCLUSION: The expression of p-STAT3 in pancreas increases in the mice with SAP induced by L-arginine. The activation of STAT3 may take part in the development of SAP. Inhibition of STAT3 activation in pancreas is one of the mechanisms of QYD treatment for SAP.     

Growth inhibition of colorectal cancer HCT116 cells by lincRNA-p21 through STAT3 signaling pathway

AIM: To study the effect of long intergenic non-coding RNA-p21 (lincRNA-p21) on the growth inhibition of colorectal cancer HCT116 cells via STAT3 signaling pathway. METHODS: The human colorectal cancer cell line HCT116 was used to construct the cells with over-expression of lincRNA-p21 by transfection of pcDNA-lincRNA-p21, and negative control cells were also set up. After transfection, the expression level of lincRNA-p21 was detected by RT-qPCR. The cell viability and proliferation were examined by MTT assay and plate colony formation assay, respectively. The protein levels of STAT3 and phosphorylated STAT3 (p-STAT3) were determined by Western blot. After STAT3 signaling pathway activator SD19 was used to treat the colorectal cancer HCT116 cells with over-expression of lincRNA-p21, Western blot was used to detect the protein levels of STAT3 and p-STAT3, MTT assay was used to measure the viability of the cells, and flow cytometry analysis was used to determine the cell apoptosis. RESULTS: Compared with control group and pcDNA group, the expression of lincRNA-p21 in pcDNA-lincRNA-p21 group was significantly up-regulated, the cell proliferation was inhibited, and the protein levels of STAT3 and p-STAT3 were significantly decreased (P<0.05). After treatment with STAT3 activator SD19, the protein levels of STAT3 and p-STAT3 in pcDNA-lincRNA-p21+SD19 group were higher than those in pcDNA-lincRNA-p21 group, the cell viability was increased, and the apoptotic rate was decreased significantly (P<0.05). CONCLUSION: Over-expression of lincRNA-p21 inhibits the growth of colorectal cancer HCT116 cells. STAT3 signaling pathway activator abolishes the growth inhibitory effect of lincRNA-p21 over-expression. lincRNA-p21 inhibits the growth of colorectal cancer cells by inhibiting the activation of STAT3 signaling.     

Effect of high mobility group box protein 1 on expression of STAT/SOCS/cyclin D1 in rat class fibroblast-like synovial cells

AIM: To investigate the regulatory effect of high mobility group box protein 1 (HMGB1) on STAT/SOCS signal transduction in RSC-364 synoviocytes. METHODS: RSC-364 cells induced by 10 μg/L human recombinant HMGB1 were collected at 6 h, 12 h or 24 h after treatment, respectively. The cells without any treatment were also collected at same time points for normal control. The mRNA expression of STAT 1/3 was analyzed by RT-PCR. The phospho-STAT 1/3 (p-STAT 1/3) and STAT 1/3 proteins were detected by Western blotting. The flow cytometric analysis (FCM) was applied to detect the protein expression of p-STAT 1/3, SOCS 1/3 and cyclin D1. RESULTS: HMGB1 at concentration of 10 μg/L significantly up-regulated the mRNA and protein levels of STAT1 expression, as well as cyclin D1 protein, and markedly increased the protein expression of p-STAT1 (P<0.01). Compared to control group, no significant difference of STAT3 and p-STAT3 expression was observed. HMGB1 elevated the protein expression of SOCS1, while the protein level of SOCS3 was only up-regulated at 6 h. CONCLUSION: HMGB1 may activate the process of STAT1 signal transduction and up-regulate the gene expression of cyclin D1, suggesting the role of HMGB1 in promoting the proliferation of synoviocytes.