同种异体富血小板血浆提取液对脂肪干细胞的体外促增殖作用

Proliferation promotion of allogenic platelet-rich plasma extraction solution on adipose-derived stem cells in vitro

目的:观察不同浓度同种异体富血小板血浆(PRP)提取液对脂肪干细胞(ADSCs)体外增殖的影响,为将同种异体PRP提取液应用于创面修复提供实验依据。方法:取SD大鼠腹股沟脂肪组织进行原代及传代培养;通过将获得的细胞向脂肪、骨和神经方向诱导分化,鉴定其干细胞特性;三次离心法制备同种异体PRP提取液,加入DMEM/F12配制成6.67%、3.35%和1.67%的培养液;将第3代ADSCs分为4组:实验组(A、B和C组)分别以含体积分数为6.67%、3.35%和1.67%PRP提取液的培养液培养,对照组(D组)ADSCs用仅含10%胎牛血清、DMEM/F12的基础培养液培养;在培养24和48h后采用MTT法观察ADSCs生长状态。结果:原代培养成功获得贴壁生长细胞,呈成纤维细胞样,并成功向脂肪细胞、骨细胞和神经细胞方向诱导分化,即具有多向分化潜能,证明所培养的细胞为ADSCs;酶标仪测定,A、B、C、D组在培养24h后吸光度(A)值分别为0.434 3±0.084 3、0.351 6±0.076 6、0.258 1±0.060 0和0.259 2±0.073 6,在培养48h后A值分别为1.016 8±0.443 6、0.741 7±0.109 7、0.367 8±0.034 2和0.395 7±0.106 2;A组和B组的A值均高于C组和D组(均P<0.05);A组的A值高于B组(P<0.05);C组A值与D组比较差异无统计学意义(P>0.05)。A和B组细胞增殖率明显升高,而C组细胞增殖率呈负增长。结论:适当浓度的同种异体PRP提取液可显著促进ADSCs体外增殖,有望应用于创面修复。

Objective To explore the effect of allogenic platelet-rich plasma（PRP） extraction solution on the proliferation of adipose-derived stem cells（ADSCs） in vitro and to provide an experimental evidence for wound healing by allogenic PRP extracting solution.Methods The adipose tissue derived from SD rats was isolated,cultured and passaged.The cells harvested above were cultured respectively in adipose-,osteo-and neuro-lineages inducing culture solution.The rats＇ blood was extracted and prepared by three times＇ centrifugation for making PRP and different concentrations of allogenic PRP extraction solution（6.67%,3.35%,and 1.67% volume fractions） were prepared by DMEM/F12.The third passage of ADSCs were divided into 4 groups.The ADSCs in experimental groups（A,B and C groups） were respectively cultured with 6.67%,3.35% and 1.67% volume fractions of PRP extraction solution.The cells in control group（D group） were cultured with 10% fetal bovine serum and DMEM/F12.The proliferative activities of the ADSCs were detected by MTT after 24 and 48 h culture.Results The harvested cells appeared to be adherent and fibroblast-like,which were successfully obtained and culured from rat adipose tissue.The cells could differentiate into adipocytes,osteocytes and neurocytes by adipose-inducing,osteo-inducing,and neuro-inducing.It was proved that the cells had the potential of differentiating into adipose-,osteo-and neuro-lineage cells and the harvested cells were ADSCs.The A values assayed by MTT after 24 h in A,B,C and D groups were 0.434 3±0.084 3,0.351 6±0.076 6,0.258 1±0.060 0,and 0.259 2±0.073 6,respectively;and the values after 48 h were 1.018 6±0.443 6,0.741 7±0.109 7,0.367 8±0.034 2,and 0.395 7±0.106 2,respectively.The A values in A group and B group were significantly higher than those in C group and D group（P0.05）,but there was no statistically significant difference between C and D groups（P0.05）.Compared with D group,the proliferation rates in A group and B group were significantly increased,but it showed negative increase in C group.Conclusion Allogenetic PRP extraction solution with suitable concentration can enhance the proliferation of ADSCs in vitro,which is expected to accelerate wound repaire.