β-联蛋白促进大鼠外周血间充质干细胞存活及扩增

本研究旨在探讨β-联蛋白在促进外周血间充质干细胞（PBMSCs）的存活和扩增中的作用。采取SD大鼠外周血，分离培养得到PBMSCs，随机将细胞分3组：未处理组（CON）、β-联蛋白激动剂组（WAY-262611，WAY)和β-联蛋白抑制剂组（XAV-939，XAV）。体外培养90 d，观察各组细胞增生和凋亡情况。应用Western 印迹和ELISA方法观察细胞衰老标记性蛋白质p16和p21表达情况，以及β-联蛋白信号通路分子表达情况。外周血经过3代培养，可获得纯度较高的PBMSCs，体外培养40 d即发生衰老和凋亡，但WAY-262611可延长细胞存活时间至90 d以上，增加其体外增殖率1.08倍（P<0.05），明显提高细胞增殖能力（P<0.05）；使WAY组p21和p16蛋白含量较CON组减少72.1%和68.4%（两者均P<0.05），较XAV组减低92.9%和93.3%（两者均P<0.05）。与CON组比较，WAY组衰老细胞数减少61.4%（P<0.05），而XAV组则增加53.3%（P<0.05）；WAY组细胞凋亡率减少50.2%（P<0.05），而XAV组细胞凋亡率增加35.7%（P<0.05）；WAY组β-联蛋白表达水平增加2.2倍（P<0.05），而XAV组则降低77.7%（P<0.05）。WAY-262611可升高WAY组β-联蛋白磷酸化水平（包括p-Ser552 和p-Ser675）97%和79%（两者均P<0.05），升高抗凋亡蛋白质Bcl2表达水平1.39倍（P<0.05），降低凋亡蛋白Bax和胱天蛋白酶3表达67.6%和83.3%（两者均P<0.05）；而XAV-939则降低Bcl2蛋白表达水平69.1%（P<0.05），升高Bax和胱天蛋白酶3表达水平1.27倍和1.02倍（两者均P<0.05）。β-联蛋白可能是影响MSCs存活和增殖的重要信号途径。这些发现可能有助于提高用于组织工程的PB-MSCs的体外生产。

β-Catenin Promotes the Survival and Amplification of Rat Peripheral Blood Mesenchymal Stem Cells

The purpose of this study was to investigate the role of β-catenin in promoting the survival and amplification of peripheral blood mesenchymal stem cells (PBMSCs). PBMSCs were isolated and collected from peripheral blood of SD rats, and were randomly divided into three groups by receiving different treatment: no treatment (served as control, CON group), β-catenin agonist (WAY-262611, WAY group), and β-catenin inhibitor (XAV-939，XAV group). The proliferation and apoptosis of PBMSCs were observed after in vitro culture for 90 d. Western blotting and ELISA were used to evaluate the expression of cellular aging proteins including p16 and p21, and the expression of β-catenin signaling molecules. Pure PBMSCs were obtained from peripheral blood after three culture passages. However, these cells underwent aging and apoptosis after 40 days of in vitro culture. WAY-262611 prolonged PBMSC survival beyond 90 d, promoted their in vitro proliferation rate by 1.08 fold (P<0.05), and significantly increased the number of cumulative population doublings (P<0.05). The protein levels of p21 and p16 in the WAY group were 72.1% and 68.4% lower than those in the CON group (both P <0.05), and 92.9% and 93.3% lower than those in the XAV group (both P <0.05). Compared with the CON group, the aging number of PBMSCs was decreased by 61.4% in the WAY group (P <0.05) and was increased by 53.3% in the XAV group (P <0.05); the cell apoptosis rate was decreased by 50.2% in the WAY group (P <0.05), and was increased by 35.7% in the XAV group (P <0.05); the expression of β-catenin was increased by 2.2-fold in the WAY group, and was reduced by 77.7% in the XAV group (P <0.05). WAY-262611 increased the phosphorylation level of β-catenin (including p-Ser552 and p-Ser675) by 97% and 79% (both P <0.05), and increased the expression level of anti-apoptotic protein Bcl2 by 1.39 fold (P < 0.05), reduced the expression of apoptotic proteins Bax and Caspase3 by 67.6% and 83.3% (both P <0.05). However, XAV-939 reduced Bcl2 protein expression by 69.1% (P <0.05) and increased the protein expression levels of Bax and Caspase 3 by 1.27 and 1.02 (both P <0.05). β-Catenin may be an important signaling pathway affecting the survival and proliferation of PBMSCs. These findings may help to improve the production of PBMSCs for tissue engineering in vitro.