Effect of protease activated receptor 2 on colonic motility in diabetes mice / 上海交通大学学报（医学版）

Objective • To observe the effect of protease activated receptor 2 (PAR2) on the colonic motility in diabetic mice and investigate the mechanism. Methods • The mouse model of type 1 diabetes mellitus was established by intraperitoneal injection of streptozotocin. The smooth muscle strips and segments of colons were isolated. The effects of PAR2 agonist on colonic motility were observed by muscle strip tension contraction and colonic migrating motor complex experiments. The effect of small conductance calcium-activated potassium channel (SK3 channel) antagonist on it was also observed. Results • PAR2 agonist inhibited colonic motility and colonic smooth muscle was more sensitive to PAR2 agonist in diabetic mice. PAR2 agonist-induced inhibition was inhibited by SK3 channel antagonist. Conclusion • PAR2 activity in diabetic mice colons is significantly enhanced, which may inhibit colonic motility through SK3 channel.

Mechanism of CCT2, a new downstream substrate of PDGFRα, on proliferation of tumor cells / 上海交通大学学报（医学版）

Objective: To study the molecular mechanism of chaperonin containing TCP1 subunit 2(CCT2), a new downstream substrate of platelet derived growth factor receptor α(PDGFRα), in tumorigenesis. Methods: Non-small cell lung cancer cell line H1703 was used. Western blotting was used to measure the phosphorylation of CCT2 upon PDGFRα inhibitor Gleevec treatment and PDGF stimulation. H1703 cells were divided into siCon group, siPDGFRα group and siCCT2 group; 48 h later, cell number counting was used to test the effect of CCT2 on cell growth after siRNA transfection. H1703 cells were divided into siCon group, siPDGFRα group, siAKT group and siCCT2 group; Western blotting was used to measure the protein level of PDGFRα and PARP. Cell fractionation was used to detect the cellular localization of CCT2 and co-immunoprecipitation was used to test the interaction between CCT2 and PDGFRα. Results: CCT2 phosphorylation was inhibited by Gleevec and induced by PDGF. Compared to the control group, the number of cells transfected by siCCT2 reduced by 30% (P=0.006). The protein level of PDGFRα was also decreased in siCCT2 transfected cells, whereas the cleavage of PARP was increased. CCT2 was localized in both cytoplasmic and membrane fractions and interacted with PDGFRα directly. Conclusion: CCT2 is a new downstream substrate of PDGFRα. CCT2 can promote tumor cells growth by interacting and stabilizing PDGFRα.

Effects of PDGFRα on melanocyte apoptosis induced by hydrogen perox-ide / 中国病理生理杂志

AIM:To investigate the effects of platelet-derived growth factor receptor α(PDGFRα) on melano-cyte apoptosis induced by hydrogen peroxide(H2O2). METHODS:Melanocyte PIGI was used as the research object. Af-ter exposed to H2O2at different concentrations,the cell viability was detected by MTT assay. The PIGI cells were transfec-ted with empty vector pCMV6 or PDGFRα over-expression vector pCMV6-PDGFRα. The transfection efficiency was deter-mined by RT-qPCR and Western blot. The effect of H2O2on the viability of the PIGI cells after over-expression of PDGFRα was measured by MTT assay. The cell apoptosis was analyzed by flow cytometry. The protein levels of p38, p-p38 and cleaved caspase-3 in the cells were detected by Western blot. DCDHF-DA was used to estemate the generation of reactive oxygen species (ROS) in the cells. RESULTS:The viability of PIGI cells decreased after exposed to H2O2(P<0.05), and the half maximal inhibitory concentration of H2O2was 0.7 mmol/L. Transfection with PDGFRα over-expression vector successfully induced high expression of PDGFRα at mRNA and protein levels in the PIGI cells,and increased the viability of the cells with H2O2treatment(P<0.05). Over-expression of PDGFRα decreased the apoptotic rate of PIGI cells trea-ted with H2O2(P<0.05),and the level of ROS in the cells(P<0.05). The protein levels of cleaved caspase-3 and p-p38 were also decreased (P <0.05). CONCLUSION:PDGFRα inhibits the apoptosis of melanocytes induced by H2O2,partially reverses the growth inhibition of melanocytes by H2O2,and decreases the ROS level. The mechanism may be related to regulating the protein levels of p-p38 and cleaved caspase-3 in the cells.

Gene mutation and prognosis of extragastrointestinal stromal tumor / 中华普通外科杂志

Objective To evaluate prognostic significance of c-kit and PDGFR-α gene mutation in extragastrointestinal stromal tumors(EGIST). Methods Paraffin embedded tissue specimens from 23 EGISTs were tested for CD117,CD34 and Ki-67 expression by immunohistochemical method.EGIST cases were also tested for the presence of c-kit exons 9,11,13,17 mutations and PDGFR-α exons 12,18 mutations.Kaplan-meier survival rate was used to evaluate the prognostic factors. Results Of 23 cases of EGIST,23(100%)were positive for CD117,17(74%)were positive for CD34.For Ki-67 labeling index(Ki-67 LI):30%were＜1%,44%were between 1%-5%,26%were＞5%.C-kit mutations were detected in 44% of EGIST patients and all were of exon 11 mutations.PDGFR-α mutations were found in 13%of all the 23 cases and all were of exon 18 mutations(The commonest type of mutation D842V).Survival analysis indicated that mitotic count and Ki-67 index were significant predictors for survival.Conclusion The pattern of c-kit and PDGFR-α mutation in EGIST was essentially similar to that in GIST.But the mutation frequency of PDGFR-α was slightly higher in EGIST than in GIST.EGIST could be a special subtype of GIST.The results of this study also show combination of mitotic counts and Ki-67 labeling index may be useful for predicting the prognosis of EGIST.