论文部分内容阅读
目的:探讨钙敏感受体(CaSR)过表达对缺血后未成熟脑白质祖细胞增殖分化的促进作用。方法:从5 d龄SD大鼠脑室周围白质组织中提取、培养脑白质祖细胞，并分为对照组、氧糖剥夺(OGD)组、OGD+三氯化钆(GdCln 3)组和OGD+n CaSR沉默组，其中利用GdCln 3特异性激动n CaSR表达，利用基因沉默方式抑制n CaSR表达。OGD后24 h、48 h、72 h、7 d和14 d，采用实时荧光定量逆转录PCR(RT-qPCR)检测各组细胞中n CaSR mRNA水平，采用免疫荧光双标染色检测各组细胞的分化情况；OGD后48 h细胞球形成时，利用倒置显微镜检测细胞增殖情况。n 结果:(1)n CaSR mRNA表达情况：OGD后48 h、72 h、7 d, OGD组细胞内n CaSR mRNA表达量均明显高于对照组，差异有统计学意义(n P<0.05)。OGD后48 h、72 h、7 d及14 d，OGD+GdCln 3组n CaSR mRNA表达量均明显高于对照组及OGD组，OGD+n CaSR沉默组n CaSR mRNA表达量均明显低于对照组及OGD组，差异均有统计学意义(n P<0.05)。(2)细胞增殖分化情况：OGD后48 h时，OGD组细胞球直径[(75.26±26.07) μm]较对照组[(57.96±18.92) μm]明显增大，OGD+GdCln 3组细胞球直径[(91.92±21.82) μm]较对照组及OGD组明显增大，而OGD+n CaSR沉默组细胞球直径[(24.09±8.34) μm]较对照组及OGD组明显减小，差异均有统计学意义(n P<0.05)。OGD后48 h、72 h时，OGD组中O4n +/CaSRn +少突胶质细胞前体细胞(OPCs)数目明显高于对照组，OGD+GdCln 3组中O4n +/CaSRn +OPCs数目明显高于对照组及OGD组，OGD+n CaSR沉默组中O4n +/CaSRn +OPCs数目明显低于对照组及OGD组，差异均有统计学意义(n P<0.05)。n 结论:CaSR过表达可进一步促进脑白质祖细胞增殖及分化为OPCs。“,”Objective:To explore the promoted role of calcium-sensing receptor (CaSR) overexpression in proliferation and differentiation of immature white matter progenitor cells after ischemia n in vitro.n Methods:Periventricular white matter cells from 5-d-old rats were cultured n in vitro and divided into control group, oxygen-glucose deprivation (OGD) group, OGD+gadolinium chloride (GdCln 3) group, and OGD+n CaSR silenced group. n CaSR expression was agitated by GdCln 3, and n CaSR gene expression was inhibited by gene silencing. n CaSR mRNA levels 24, 48 and 72 h, and 7 and 14 d after OGD were detected by real-time fluorescence quantification-PCR (RT-qPCR); cell proliferation 48 h after OGD was detected by inverted microscope, and cell differentiation 24, 48, and 72 h, and 7 and 14 d was detected by double immunofluorescence staining.n Results:(1) n CaSR mRNA expressions: n CaSR mRNA expressions in OGD group 48 and 72 h and 7 d after OGD were statistically higher than those in control group (n P<0.05). Forty-eight and 72 h, and 7 and 14 d after OGD,n CaSR mRNA expressions in OGD+GDCLn 3 group were statistically higher than those in control group and OGD group (n P<0.05); then CaSR mRNA expressions in the OGD+CaSR silenced group were significantly lower than those in the control group (n P<0.05). (2) Cell proliferation and differentiation: 48 h after OGD, the cell sphere diameter of OGD group ([75.26±26.07] μm) was significantly increased as compared with that of control group ([57.96±18.92] μm,n P<0.05); the cell sphere diameter of OGD+GdCln 3 group ([91.92±21.82] μm) was significantly increased as compared with that of control group and OGD group ( n P<0.05); and the cell sphere diameter of OGD+n CaSR silenced group ([24.09±8.34] μm) was significantly shorter than that of control group and OGD group ( n P<0.05). At 48 and 72 h after OGD, the number of O4n +/CaSRn + olidoendrocyte precursor cells (OPCs) in OGD group was significantly larger than that in control group, that in OGD+GdCln 3 group was significantly larger than that in control group and OGD group, and that in OGD+n CaSR silenced group was significantly smaller than that in control group and OGD group (n P<0.05).n Conclusion:CaSR overexpression could promote the proliferation and differentiation of progenitor cells to OPCs.