| 【Objective】The proportion of patients diagnosed with advanced prostate cancer in my country is significantly higher than that in European and American countries,and the treatment of advanced prostate cancer is still the focus of clinical treatment.The standard chemotherapy regimen represented by Docetaxel(DTX)is an important treatment for advanced prostate cancer,while chemotherapy resistance is a difficulty in clinical treatment,and the relevant mechanism has not been fully elucidated.Studies have shown that tumor microenvironment is closely related to tumor chemotherapy resistance.This study links the symbiotic relationship between cancer-associated fibroblasts(CAF)and prostate cancer cells,and explores the potential mechanism of docetaxel chemotherapy resistance in advanced prostate cancer.【Methods】 We use miR-145-5p mimics and inhibitors to transfect prostate cancer PC3 cells to observe the effects of miR-145-5p on PC3 cell migration,invasion and DTX resistance,and use Western blot and q RT-PCR to detect PC3 in each group cell SP1,USP8 and CX43 expression.The cytokine TGFβ1 was used to induce fibroblasts to become cancer-related fibroblasts,the CAF was identified and cultured,and the exosomes in the CAF supernatant were separated by ultracentrifugation and identification experiments was performed.The experiment is divided into 3 groups: CON group(basal medium),CAF-medium(CAF supernatant),CAF-exo(basic medium containing exosomes).After the 3 groups of PC3 cells were cultured in different medium for 24 hours,they were cultured with10 n M DTX medium for 36 hours.Cell viability was detected by CCK8 method to observe the effect of CAF-exo on the DTX resistance of prostate cancer PC3 cells,q RT-PCR used to investigate the expression of miR-145-5p in each group.The changes of cell migration and invasion of each group were observed,and the expression of SP1,USP8 and CX43 of PC3 cells in each group was detected by Western blot and q RT-PCR.SP1 protein si RNA was used to knock down the expression of SP1,Western blot,q RT-PCR and CCK8 method were used to observe the expression of USP8,CX43 and the effect of DTX resistance on PC3 cells after SP1 knockdown【Results】miR-145-5p inhibits the invasion and migration of prostate cancer PC3 cells,and improves the drug resistance to docetaxel significantly.Western blot and q RT-PCR showed that miR-145-5p can reduce the expression of SP1,USP8,CX43 protein and m RNA in PC3 cells.The results of the grouping experiment showed that the DTX sensitivity of the CAF-medium group and CAF-exo group was significantly lower than that of the CON group(P<0.01).At the same time,the migration and invasion ability of CAF-medium group and CAF-exo group was significantly inhibited compared with CON group.The q RT-PCR results showed that the expression of miR-145-5p in prostate cancer PC3 cells treated with CAFconditioned medium or CAF exosomes was significantly up-regulated(P<0.01).Western blot and q RT-PCR detection of PC3 cell gene expression found that the protein and m RNA expression of SP1,USP8 and CX43 in CAF-medium group and CAF-exo group were down-regulated compared with the CON group,significantly.After knocking down the expression of SP1 with transcriptional regulator SP1 specific si RNA,the expression of USP8,CX43 protein and m RNA in PC3 cells was significantly down-regulated,with statistical differences.At the same time,after SP1 knockdown,the sensitivity of prostate cancer PC3 cells to docetaxel was significantly reduced(P<0.05).【Conclusions】(1)miR-145-5p reduce the drug sensitivity of prostate cancer PC3 cells to docetaxel;(2)CAF-exo promotes docetaxel resistance through the SP1 /USP8 / CX43 pathway by up-regulating the expression of miR-145-5p in prostate cancer PC3 cells. |
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