神经节苷脂对神经母细胞瘤细胞SK-N-SH黏附过程中pp125焦点黏附激酶表达的调节作用

Effect of Endogeneous Gangliosides on pp125 Focal Adhesion Kinase Expression during Adhesion of SK-N-SH Neuroblastoma Cells to Collagen

目的探讨内生肿瘤神经节苷脂(Gls)及整合素α2β1对神经母细胞瘤(NB)细胞SK-N-SH与胶原蛋白(Col)黏附反应中pp125焦点黏附激酶(pp125FAK)表达的影响。方法用葡萄糖苷神经酰氨合成酶抑制剂(D-PDMP)抑制SK-N-SH细胞株Gls合成,采用免疫沉淀和Western印迹技术,观察肿瘤内生Gls和抗整合素α2及β1单克隆抗体后对该细胞黏附于胶原蛋白后酪氨酸磷酸化蛋白表达变化。结果暴露于D-PDMP 6 d后,细胞Gls几乎完全清除,Mg2+为1 mmol/L条件下,SK-N-SH细胞与胶原蛋白黏附后,肿瘤细胞Gls清除组较未清除Gls组pp125FAK酪氨酸磷酸化蛋白的表达明显下降。肿瘤细胞神经节苷脂主要成分GD2一定程度能增加细胞黏附后酪氨酸磷酸化蛋白的表达。采用anti-α2单克隆抗体6F1和anti-β1单克隆抗体可降低肿瘤细胞黏附于Col后pp125FAK酪氨酸磷酸化蛋白的表达。结论内生肿瘤Gls调节肿瘤细胞对Col的黏附作用中pp125FAK酪氨酸磷酸化蛋白的表达,其调节作用可能是通过影响整合素α2β1的功能而发挥作用。Gls增强肿瘤细胞整合素α2β1的信号传导可能通过促进pp125FAK的酪氨酸磷酸化而起作用。

Objective To explore effect of endogeneous gangliosides（Gls） and integrin α2β1 on protein phosphotyrosine expression of pp125 focal adhesion kinase （pp125FAK） after adhesion of SK- N - SH neuroblsstoma cells to collagen（Col）. Methods SK - N - SH cell line with high expression of integrin α2β1 was cultured in presence of D threo- 1 - phenyl - 2 - decanolamino - 3 - morphinoline - 1 - propanol （D - PDMP）. Effect of endogeneous Gls, anti - α2 and anti - β1 monoelonal antibody on protein phosphotyrosine expression of pp125FAK during adhesion of SK - N - SH cells to Col were determined by immunoprecipitate and Western blotting. Results After 6 days, endogenous Gls in cells were almost depleted. Gls - depletion, anti - α2 and anti - β1 monoelonal antibody were able to decrease pp125FAK expression of SK - N - SH cells adherent to Col respectively. GD2, the major component of neuroblastoma cell Gls could recover pp125FAK expression to a certain degree. Conclusions Endogenous tumor Gls regulate protein phosphotyrosine expression of pp125FAK during adhesion of neuroblastoma cells to Col. It is suggested that tumor Gls may increase signal transduction of tumor cell integrin α2β1 by increasing tyrosine phosphorylation of pp125FAK.