Tie-2在口腔鳞状细胞癌侵犯颌骨过程中的作用机制研究

目的:研究酪氨酸激酶受体-2（tyrosinekinase receptor-2，Tie-2）在口腔鳞状细胞癌侵犯颌骨过程中的作用机制。方法:免疫组化SP法比较Tie-2在颌骨侵犯前沿的口腔鳞状细胞癌与其他部位的口腔鳞癌中的表达差异；构建3种针对Tie-2的siRNA慢病毒载体（662、663、664）为实验组、空载病毒429和生理盐水（NS）为对照组，并将各组转染至人舌鳞癌细胞系（Cal-27）中，应用qRT-PCR筛选出抑制效果最好的siRNA；CCK-8法检测细胞活性；将抑制效果最好的慢病毒载体转染至Cal-27后，将其培养基上清添加至人单核细胞系（THP-1）培养基中，qRT-PCR和Western blotting检测THP-1中的破骨细胞功能相关基因抗酒石酸酸性磷酸酶（tartrate resistant acid phosphatase，TRAP）和组织蛋白酶K（cathepsin K，CTSK）的mRNA和蛋白量在各组中的表达差异。结果:颌骨侵犯前沿的骨旁鳞癌组织染色比其他部位鳞癌组织显著加深，骨旁鳞癌组织MOD值显著增大（P＜0.000 1）；siRNA-664对Cal-27中Tie-2表达的抑制效果最佳（P＜0.000 1）；用慢病毒转染Cal-27和THP-1后，细胞活性无明显变化，差异无统计学意义；沉默Cal-27中Tie-2的表达后，THP-1的破骨细胞功能相关基因和蛋白表达显著降低（P＜0.000 1）。结论:下调Tie-2表达后，THP-1向破骨细胞分化的潜力降低，表现为破骨细胞功能相关基因和蛋白表达下调。OSCC可能通过激活Ang/Tie-2信号体系释放某些破骨细胞分化因子，促进破骨细胞分化，从而导致局部骨侵犯发生。

The mechanism of Tie-2 in the process of oral squamous cell carcinoma invading the jaw

Objective:To study the mechanism of Tie-2(tyrosinekinase receptor-2) in the process of oral squamous cell carcinoma invading the jaw.Methods:Immunohistochemistry was used to compare the expression of Tie-2 in oral squamous cell carcinoma at the front of jaw invasion and other oral squamous cell carcinoma.Three siRNA lentiviral vectors(662,663,664) against Tie-2 were constructed as the experimental group,no-load virus 429 and normal saline（NS) as the control group,and each group was transfected into the hunan tongue squamous carcinoma cell line(Cal-27),the siRNA with the best inhibitory effect was screened by qRT-PCR.The cell viability was detected by CCK-8 method.The lentiviral vector with the best inhibitory effect was transfected into Cal-27,the culture supernatant was added to the human monocytic cell line(THP-1) medium,the expression of mRNA and protein in osteoclast function related genes TRAP(tartrate resistant acid phosphatase) and CTSL(cathepsin K) in THP-1 were detected by qRT-PCR and Western blotting.Results:The staining of the parosteal OSCC tissue at the invasion edge of the jaw was significantly deeper than that at other parts of the OSCC tissue，and the MOD value of parosteal OSCC tissue was significantly increased(P＜0.000 1).siRNA-664 had the best inhibitory effect on the expression of Tie-2 in Cal-27.After transfecting Cal-27 and THP-1 with lentivirus,the cell viability did not change significantly,and the difference was not statistically significant.After silencing the expression of Tie-2 in Cal-27,theosteoclast function-related genes and proteins of THP-1 significantly reduced expression.Conclusion:After down-regulating the expression of Tie-2,the potential of THP-1 to differentiate into osteoclasts was reduced,which is manifested as the down-regulation of osteoclast function-related genes and proteins.OSCC may release some osteoclast differentiation factors by activating the Ang/Tie-2 signaling system to promote osteoclast differentiation,thereby leading to local bone invasion.