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目的:探讨体外培养的人早孕绒毛膜滋养细胞在抗子宫内膜抗体(EmAb)环境中,低分子肝素对其增殖及凋亡能力的调节作用。方法:胰蛋白酶/DNA酶Ⅰ联合消化,通过Percoll细胞分离液纯化得到绒毛滋养细胞。按照EmAb(+)血清未加低分子肝素(A1组)、EmAb(+)血清加低分子肝素(A2组)、EmAb(-)血清未加低分子肝素(B1组)、EmAb(-)血清加低分子肝素(B2组)4个分组进行培养24h后,四甲基偶氮唑蓝比色法(MTT)检测滋养细胞细胞增殖情况,以490nm处的平均吸光度(A)值表示;采用免疫荧光化学法检测增殖细胞核抗原(PCNA)的表达水平,以平均荧光灰度值表示;采用流式细胞术检测细胞凋亡,以细胞凋亡率(%)表示。结果:(1)A1组细胞490nm处的A值为0.1007±0.0125,A2组为0.1982±0.0135,两者比较,差异有统计学意义(P<0.05);B1组细胞490 nm处的A值为0.2117±0.0106,B2组为0.2211±0.0080,两者比较,差异无统计学意义(P>0.05);(2)A1组细胞PCNA的表达水平为9.77±0.61,A2组为20.96±1.31,两者比较,差异有统计学意义(P<0.05);B1组细胞PCNA的表达水平为21.40±1.93,B2组为22.42±1.70,两者比较差异无统计学意义(P>0.05);(3)A1组细胞凋亡率为(2.64±0.10)%,A2组为(0.82±0.02)%,两者差异有统计学意义(P<0.05);B1组细胞凋亡率为(0.79±0.03)%,B2组为(0.76±0.03)%,两者差异无统计学意义(P>0.05)。结论:EmAb能抑制人早孕绒毛膜滋养细胞增殖,并促进其凋亡,这可能是导致自然流产的原因之一,而低分子肝素能够逆转这一过程。
Objective: To investigate the effect of low molecular weight heparin on the proliferation and apoptosis of human chorionic trophoblast cultured in vitro in the anti-endometrial antibody (EmAb) environment. Methods: Combined trypsin / DNase Ⅰ digestion, vill trophoblast cells were obtained by Percoll cell separation. EmAb (+) serum and low molecular weight heparin (Group A1), EmAb (+) serum plus low molecular weight heparin (Group A2) and EmAb (- Addition of low molecular weight heparin (B2 group) 4 groups after 24h, tetracycline blue cell colorimetry (MTT) detection of trophoblast cell proliferation, 490nm at the average absorbance (A) value; using immunization Fluorescence staining was used to detect the expression of proliferating cell nuclear antigen (PCNA), which was expressed as mean fluorescence gray value. The apoptosis rate was measured by flow cytometry (%). Results: (1) The value of A at 490 nm in group A1 was 0.1007 ± 0.0125 and that in group A2 was 0.1982 ± 0.0135, the difference was statistically significant (P <0.05). The value of A at 490 nm in group B1 was 0.2117 ± 0.0106 in B2 group and 0.2211 ± 0.0080 in B2 group (P> 0.05). (2) The expression of PCNA in group A1 was 9.77 ± 0.61 and in group A2 was 20.96 ± 1.31 (P <0.05). The expression level of PCNA in group B1 was 21.40 ± 1.93 and in group B2 was 22.42 ± 1.70, there was no significant difference between the two groups (P> 0.05). (3) The expression of A1 The apoptosis rate of group B1 was (2.64 ± 0.10)%, that of group A2 was (0.82 ± 0.02)%, the difference was statistically significant (P <0.05) B2 group was (0.76 ± 0.03)%, the difference was not statistically significant (P> 0.05). CONCLUSION: EmAb can inhibit the proliferation and promote the apoptosis of human chorionic trophoblast in early pregnancy, which may be one of the reasons leading to spontaneous abortion. However, low molecular weight heparin can reverse this process.