| Objective:To establish a pancreatic cancer PANC-1 subcutaneous xenograft model;To investigate the effect of the tumorigenic ability of silencing KLK7 gene on pancreatic cancer PANC-1 cells in SCID mice.To explore the drug toxicity of KLK7 small molecule inhibitors and its pharmacological effect on SCID mouse PANC-1 cell xenograft models.Methods:(1)Pancreatic cancer PANC-1(primordial cells),S4(cells silencing KLK7 gene in PANC-1 cells by gene knockout technique)and NC(PANC-1 was transfected with empty lentivirus)were cultured and three kinds of subcutaneous xenograft models of SCID mice were established respectively.(2)SCID mice were divided into five groups to construct a toxicity test mouse model according to the concentration of KLK7 small molecule inhibitor: control group(DMSO)、6.25 m M group、12.5m M group、25m M group and 50mmol/L group;After continuous administration for 21 days,the symptoms of poisoning and death of mice in each group were observed,and the changes of body weight of mice were monitored.The pathological changes of heart,liver,spleen,lung and kidney were observed.At the end of the experiment,the mice were killed and the pathological changes of important organs(heart,liver,spleen,lung,kidney)was observed by using HE staining.(3)After the subcutaneous xenograft models models of PANC-1,S4 and NC were established,the general conditions(such as spirit,diet,activity,etc.)and the tumorigenesis rate of mice in each group were observed and recorded.The body weight and tumor volume of mice were monitored regularly.Two months after inoculation,the mice were killed and the tumor weight was measured,and the metastasis of important organs(heart,liver,spleen,lung,kidney)was observed by using HE staining.(4)The subcutaneous xenograft model of PANC-1 mice were established and divided into three groups(control group,10 m M group and 50 m M group),the corresponding concentrations of KLK7 small molecule inhibitor were administered respectively,and the control group was given the same volume of DMSO.The general situation(such as spirit,diet,activity,etc.)and the tumorigenesis rate of mice in each group were observed and recorded.The body weight and tumor volume of mice were monitored regularly.At the end of the experiment,the mice were killed and the tumor weight was measured;the metastasis of important organs(heart,liver,spleen,lung,kidney)was observed by using HE staining.Results:(1)A subcutaneous xenograft model of pancreatic cancer PANC-1 cells was successfully established.On the 11 th day after inoculation,induration can be reached at theinoculation site,and the overall tumor formation rate was 81.25%.(2)Toxicity experiments:Through experimental observation,no mice died in the control group and 6.25 m M group during the whole experiment,and One female mice in 12.5m M,25 m M and 50 m M groups died on the 11 th,10th and 20 th day respectively;Through weight monitoring,it was found that compared with the control group,the female mice in the experimental group fluctuated greatly and had different degrees of decline.In the experimental group,Pathological sections of important organs showed partial necrosis of nephron in the kidneys and at the same time,the cell density becomes thinner and the white pulp becomes less in the spleen.It shows that KLK7 small molecule inhibitors have certain toxic effects on kidney and spleen of SCID mice,but no obvious toxic effects on heart,liver and lung.(3)The subcutaneous xenograft models of PANC-1,NC and S4 mice were successfully established.The results showed that the subcutaneous transplantation tumors of the gene knockdown group grew slowly in the whole experimental process,and the tumor volume was significantly smaller than that of the blank control group.It was also smaller than the blank control group.The blank control composition rate was 100%,the tumor mass was(0.57±0.20)g,the negative control composition rate was 85.71%,the tumor mass was(0.30±0.20)g,and the gene knockdown composition rate was 64.29%.(0.15±0.15)g,it can be seen that the tumor quality of the gene knockdown group and the negative control group was significantly lower than that of the blank control group,the difference was statistically significant(P<0.05);HE staining showed that there were no obvious signs of metastasis in heart,liver,spleen,lung and kidney.(4)The efficacy test showed that the subcutaneous transplantation tumors in the 50 m M group grew most slowly,and the tumor volume was significantly smaller than that of the control group.The growth rate of subcutaneous transplantation tumors in the 10 m M group was faster than that in the 50 m M group,but it was also smaller than the control group.The control tumor formation rate was78.57%,the tumor mass was(0.031±0.030)g,the 10 m M tumor formation rate was64.29%,the tumor mass was(0.017±0.017)g,the 50 m M tumor formation rate was31.43%,and the tumor mass was(0.004±).0.008)g,it can be seen that the tumor quality of the 50 m M group was significantly lower than that of the control group,and the difference was statistically significant(P<0.05);HE staining showed that there were no obvious signs of metastasis in heart,liver,spleen,lung and kidney.Conclusion:In our study,we confirmed that silencing KLK7 gene could inhibit the tumorigenesis and proliferation of PANC-1 subcutaneous xenografts model.At the sametime,it was confirmed that the pre-synthesized KLK7 small molecule inhibitor inhibited the biological malignant behavior of PANC-1 cells subcutaneously transplanted tumor,which provided a basis for drug development of targeted therapy for pancreatic cancer. |
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