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宣城市人民医院肛肠科,宣城,242000
修回日期:2020-09-04,
录用日期:2020-09-24,
纸质出版日期:2023
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彭胜建. 过表达细胞因子信号转导抑制因子2调控JAK2/STAT3通路抑制直肠癌细胞活性和移植瘤生长[J]. 重庆医科大学学报, 2023,48(1):37-42.
Peng Shengjian. Overexpression of SOCS2 on inhibition of rectal cancer cell growth activity and xenograft tumor growth by JAK2/STAT3 pathway[J]. Journal of chongqing medical university, 2023, 48(1): 37-42.
目的: 探究细胞因子信号转导抑制因子2(suppressor of cytokine signaling 2,SOCS2)过表达对直肠癌细胞生长、运动和JAK2/STAT3通路的影响。方法: HCT8细胞分为对照组、pcDNA组和pcDNA-SOCS2组,根据组别转染pcDNA空载体或pcDNA-SOCS2重组表达载体,RT-PCR检测SOCS2基因表达,BrdU染色检测细胞增殖,流式细胞术检测细胞凋亡,Transwell检测细胞侵袭,Western blot检测细胞SOCS2、上皮钙黏蛋白、波形蛋白、纤连蛋白表达以及JAK2和STAT3的磷酸化。另设BALB/c nu/nu裸鼠分为对照组、pcDNA-SOCS2组,建立HCT8皮下移植瘤模型,称取移植瘤质量,免疫组化检测Ki-67和血管内皮生长因子(vascular endothelial growth factor,VEGF)平均光密度,Western blot检测瘤组织JAK2和DTAT3磷酸化。结果: 离体实验中,RT-PCR结果显示,与对照组(1.00±0.00)比较,pcDNA-SOCS2组细胞SOCS2 mRNA水平(4.70±0.27)明显升高(P=0.000)。Western blot结果显示,pcDNA-SOCS2组细胞SOCS2蛋白表达(0.130±0.020)较对照组(0.010±0.005)明显上调(P=0.000)。BrdU阳性百分比明显降低(P=0.000);流式细胞术结果显示,pcDNA-SOCS2组[(35.0±5.0)%]HCT8细胞凋亡率较对照组[(6.0±3.4)%]明显升高(P=0.000)。Transwell分析结果显示,pcDNA-SOCS2组(89.0±10.0)HCT8细胞侵袭数目较对照组(87.0±13.0)明显减少(P=0.000)。Western blot结果显示,pcDNA-SOCS2组HCT8细胞中上皮钙黏蛋白水平(0.120±0.030)较对照组(0.010±0.004)明显升高(P=0.000),同时波形蛋白水平(0.008±0.004)和纤连蛋白水平(0.010±0.005)较对照组(0.170±0.024,0.070±0.020)明显降低(P=0.000),JAK2和STAT3磷酸化水平降低(P=0.000)。在体实验中,pcDNA-SOCS2组瘤体质量(0.14±0.06)较对照组(0.45±0.08)明显减轻(P=0.000)。免疫组化结果显示,pcDNA-SOCS2组Ki-67和VEGF平均光密度(17.0±6.0,7.0±6.0)较对照组(52.0±9.0,43.0±9.0)明显降低(P=0.000),瘤体JAK2和STAT3磷酸化水平明显下调(P=0.000)。结论: SOCS2过表达可抑制直肠癌HCT8细胞的增殖和运动能力并促进其凋亡,这一作用与JAK2/STAT3通路有关。
Objective: To investigate the effect of suppressor of cytokine signaling 2 (SOCS2) overexpression on the growth
movement and JAK2/STAT3 pathway of rectal cancer cells. Methods: HCT8 cells were divided into control group
pcDNA group and pcDNA-SOCS2 group
transfected with pcDNA empty vector or pcDNA-SOCS2 group recombinant expression vector according to different groups. SOCS2 gene expression was detected by RT-PCR
cell proliferation was detected by BrdU staining
flow cytometry was used to detect apoptosis
Transwell method was used to detect cell invasion
and Western blot was used to detect SOCS2
E-cadherin
vimentin
fibronectin protein expression
and phosphorylation of JAK2 and STAT3. In addition
BALB/c nu/nu nude mice were divided into con-trol group and pcDNA-SOCS2 group. HCT8 subcutaneous xenograft model was established
and the weight of xenograft tumor was weighed. The average optical density of Ki-67 and vascular endothelial growth factor (VEGF) was measured by immunohistochemistry
and the Western blot was used to detect the phosphorylation of JAK2 and STAT3 in tumor tissues. Results: In the ex-vivo experiment
the RT-PCR results showed that compared with the control group (1.00±0.00)
the SOCS2 mRNA level (4.70±0.27) of the pcDNA-SOCS2 group was significantly increased (P=0.000). Western blot results showed that the expression of SOCS2 protein in the pcDNA-SOCS2 group (0.130±0.020) was significantly up-regulated (P=0.000) compared with that in the control group (0.010±0.005). The percentage of BrdU positive was signifi-cantly reduced (P=0.000); flow cytometry results showed that the apoptosis rate of HCT8 cells in the pcDNA-SOCS2 group [(35.0±5.0) %] was significantly higher than that in the control group [(6.0±3.4) %
P=0.000]. Transwell analysis results showed that the number of HCT8 cell invasions in the pcDNA-SOCS2 group (89.0±10.0) was significantly lower than that of the control group (87.0±13.0
P=0.000). Western blot results showed that the E-cadherin protein level (0.120±0.030) of HCT8 cells in the pcDNA-SOCS2 group was significantly increased compared to the con-trol group (0.010±0.004
P=0.000)
while the Vimentin protein level (0.008±0.004) and Fibronectin protein level (0.010±0.005) were significantly reduced compared to the control group (0.170±0.024
0.070±0.020)
P=0.000
the phosphorylation level of JAK2 and STAT3 decreased (P=0.000). In the in-vivo experiment
the tumor weight in the pcDNA-SOCS2 group (0.14±0.06) was signifi-cantly less than that in the control group (0.45±0.08
P=0.000). The results of immunohistochemistry showed that the average optical density of Ki-67 and VEGF in the pcDNA-SOCS2 group (17.0±6.0
7.0±6.0) was significantly lower than that in the control group (52.0±9.0
43.0±9.0
P=0.000)
and the phosphorylation levels of JAK2 and STAT3 tumor bodies were significantly down-regulated (P=0.000). Conclusion: The overexpression of SOCS2 can inhibit the proliferation and exercise ability of rectal cancer HCT8 cells and promote the apoptosis
which is related to JAK2/STAT3 pathway.
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