Expression And Significance Of COL3A1 In Gastric Cancer

Objective: To detect the mRNA and protein expression levels of COL3A1 in gastric cancer and its normal tissues,and to preliminarily investigate the relationship between COL3A1 and various clinical and pathological features of gastric cancer.To investigate the role of COL3A1 in the proliferation,migration and invasion of gastric cancer cells.Methods: Ten fresh specimens and 50 paraffin specimens were collected from cancerous and normal tissues of gastric cancer patients,and the mRNA and protein expression levels of COL3A1 in the two tissues were detected by quantitative real-time PCR and immunohistochemistry,respectively.Meanwhile,clinical data of the examined patients were collected to analyse the relationship between COL3A1 expression and the clinicopathological characteristics of gastric cancer(gender,age,degree of differentiation,lymphatic metastasis,TNM stage,depth of tumour infiltration and tumour size).The gastric cancer cell line MGC803 was transfected with the interference lentivirus targeting COL3A1(3 targets)and the negative control lentivirus,and the stable cell line was screened for purinomycin resistance.The mRNA expression of COL3A1 in MGC803 cells after transfection was verified by q RT-PCR,and two cell lines with the highest transfection efficiency were screened out.The protein expression of COL3A1 in gastric cancer cells after transfection was further detected by Western blot.MTT and Transwell assay were used to detect the proliferation,invasion and migration phenotypes of gastric cancer cells in the COL3A1 interference group and the control group,and the changes in the biological characteristics of gastric cancer cells before and after COL3A1 knockdown were observed.Results:(1)The expression level of COL3A1 mRNA in gastric cancer tissues was6.86±3.07,while that in normal tissues was only 2.48±1.25,the difference was statistically significant(t=4.181,P = 0.0013).By comparing the protein levels of COL3A1 in gastric cancer tissues and their normal tissues,the former was significantly higher than the latter,and the difference was statistically significant(P = 0.001).(2)The expression level of COL3A1 protein was correlated with TNM stage(P < 0.001)and depth of invasion(P=0.012),but not with gender(P=0.529),age(P=0.880),degree of differentiation(P=0.869),tumor size(P=0.145)and lymph node metastasis(P=0.416).(3)mRNA expression level of COL3A1 was significantly decreased after gastric cancer cells were transfected with sh RNA-COL3A1 lentivirus,and the inhibitory effect was best in sh RNA 1 group(P < 0.01)and sh RNA 3 group(P <0.001).Western blot analysis showed that the relative expression of COL3A1 protein in NC group was 2.80±0.08,that in sh RNA 1 group was 1.31±0.38(P=0.0031),and that in sh RNA 2 group was 0.45±0.28(P=0.0013).The inhibitory effect of sh RNA 2 group was more significant(P < 0.01).(4)MTT results showed that 48 h after sh RNACOL3A1 lentivirus transfection,the growth of gastric cancer cells was significantly inhibited,and the absorption values at all time points were lower than those in NC group(P < 0.05),and sh RNA 2 group was the most obvious(P < 0.0001).(5)Transwell chamber was used to detect the migration ability of sh RNA-COL3A1 gastric cancer cells.It was found that after sh RNA-COL3A1 transfection,the number of model penetrating cells in the NC group was 236±13.5,that in the sh RNA 1 group was176±19.0,and that in the sh RNA 2 group was 83±12.1.The migration ability of gastric cancer cells in sh RNA 1 group(P=0.0029)and sh RNA 2 group(P < 0.0001)was significantly decreased,and sh RNA 2 group was the most significant(P < 0.0001).(6)The invasion ability of sh RNA-COL3A1 gastric cancer cells was detected by Transwell chamber,and the number of model penetrating cells in NC group was 427±39.0,the number of sh RNA 1 group was 316±28.6,and the number of sh RNA 2 group was169±23.0.The invasion ability of gastric cancer cells in sh RNA 1 group(P=0.0046)and sh RNA 2 group(P=0.0002)were significantly decreased,and sh RNA 2 group was the most significant(P < 0.001).Conclusion:(1)The expression of COL3A1 at both mRNA and protein levels in gastric cancer tissues was significantly higher than that in normal tissues;(2)The high expression of COL3A1 was associated with tumor stage and infiltration depth,but not with gender,age,differentiation,tumor size and lymph node metastasis;(3)Knockdown of COL3 Al gene can effectively inhibit the proliferation,invasion and migration of gastric cancer cells,and COL3 Al plays a role in played an important role in the development of gastric cancer.