摘要目的:探讨猪脱细胞真皮基质(pADM)在体外培养中对毛囊角质形成细胞增殖的影响及其机制。方法:体外实验培养毛囊角质形成细胞，按照干预措施不同分为6组，A组为单纯毛囊角质形成细胞组；B组为毛囊角质形成细胞+pADM组(20 mg/L)；C组为毛囊角质形成细胞+肿瘤坏死因子-α(TNF-α)组(10 μg/L)；D组为毛囊角质形成细胞+TNF-α抑制剂组(Lenalidomide)；E组为毛囊角质形成细胞+TNF-α(10 μg/L)+pADM组(20 mg/L)；F组为毛囊角质形成细胞+TNF-α抑制剂+pADM组(20 mg/L)。用实时荧光定量聚合酶链反应(RT-PCR)法分别测定各组的淋巴增强因子1(LEF-1) mRNA及细胞核增殖抗原(Ki-67) mRNA的表达。免疫荧光标记法检测B、E、F组的毛囊角质形成细胞，观察pADM对毛囊角质形成细胞增殖的影响。两组间比较采用独立样本 t检验。 结果:A组(LEF-1，1.093±0.121，Ki-67，1.100±0.089)毛囊角质形成细胞中LEF-1 mRNA及Ki-67 mRNA的表达低于B组(LEF-1，2.973±0.234，Ki-67，2.577±0.067)和D组(LEF-1，1.867±0.045，Ki-67，1.967±0.136)，但高于C组(LEF-1，0.450±0.090，Ki-67，0.490±0.026)。A组与B组、A组与C组、A组与D组之间的差异均有统计学意义(A比B， tLEF-1＝14.280， P<0.05； tKi-67＝15.390， P<0.05；A比C， tLEF-1＝4.887， P<0.05； tKi-67＝ 6.356， P<0.05；A比D， tLEF-1＝5.875， P<0.05； tKi-67＝9.030， P<0.05)；B组(LEF-1，2.973±0.234，Ki-67，2.577±0.067)毛囊角质形成细胞中LEF-1 mRNA及Ki-67 mRNA的表达高于E组(LEF-1，1.793±0.078，Ki-67，1.940±0.165)，而低于F组(LEF-1，4.257±0.266，Ki-67，3.193±0.155)，B组与E组，B组与F组的差异均有统计学意义(B比E， tLEF-1＝8.964， P<0.05； tKi-67＝6.634， P<0.05；B比F， tLEF-1＝9.749， P<0.05； tKi-67＝-6.425， P<0.05)；免疫荧光结果显示，B组毛囊角质形成细胞数量高于E组而低于F组。 结论:在体外培养中，pADM可能通过下调TNF-α促进LEF-1和Ki-67的表达，从而促进毛囊角质形成细胞的增殖。

abstractsObjective:To investigate the effect of porcine acellular dermal matrix (pADM) on the proliferation of hair follicle keratinocytes in vitro and its possible mechanism. Methods:Hair follicle keratinocytes were cultured in vitro and divided into 6 groups according to different intervention measures. group A, hair follicle keratinocytes only; group B, hair follicle keratinocytes treated with the pADM (20 mg/L); group C, hair follicle keratinocytes treated with tumor necrosis factor-α (TNF-α, 10 μg/L); group D, hair follicle keratinocytes treated with TNF-α inhibitor (Lenalidomide); group E, hair follicle keratinocytes treated with TNF-α (10 μg/L) and pADM (20 mg/L); group F, hair follicle keratinocytes treated with TNF-α inhibitor and pADM (20 mg/L). The real-time polymerase chain reaction (RT-PCR) method was used to detect the expression of LEF-1 mRNA and Ki-67 mRNA in each group. Immunofluorescence labeling method was used to detect hair follicle keratinocytes in groups B, E and F, and the effects of pADM on the proliferation of hair follicle keratinocytes were observed. Measurement data were expressed as mean±standard deviation ( mean± SD). The independent sample t test was used for comparison between the two groups, and P<0.05 indicated that the difference was statistically significant. Results:By RT-PCR analysis, the expressions of LEF-1 mRNA and Ki-67 mRNA in hair follicle keratinocytes of group A (LEF-1, 1.093±0.121, Ki-67, 1.100±0.089) were lower than those of group B (LEF-1, 2.973±0.234), Ki-67, 2.577±0.067) and group D (LEF-1, 1.867±0.045, Ki-67, 1.967±0.136), but higher than group C (LEF-1, 0.450±0.090, Ki-67, 0.490±0.026). The differences between group A and group B, group A and group C, group A and group D were statistically significant (A vs. B, tLEF-1=14.280, P<0.05; tKi-67=15.390, P<0.05; A vs. C, tLEF-1=4.887, P<0.05; tKi-67= 6.356, P<0.05; A vs. D, tLEF-1=5.875, P<0.05; tKi-67=9.030, P<0.05); Group B (LEF-1, 2.973±0.234, Ki-67, 2.577±0.067) The expression of LEF-1 mRNA and Ki-67 mRNA in hair follicle keratinocytes was higher than that of group E (LEF-1, 1.793±0.078, Ki-67, 1.940±0.165), but lower than group F (LEF-1, 4.257±0.266, Ki-67, 3.193±0.155), the differences between group B and group E, group B and group F were statistically significant (B vs. E, tLEF-1=8.964, P<0.05; tKi-67=6.634, P<0.05; B vs. F, tLEF-1=9.749, P<0.05; tKi-67=-6.425, P<0.05); immunofluorescence results showed that the number of hair follicle keratinocytes in group B was higher than that in group E but lower in Group F. Conclusion:In vitro, the pADM may up-regulate the expression of LEF-1 and Ki-67 by down-regulating TNF-α, and then promote the proliferation of hair follicle keratinocytes.