Experimental study of Qianlongtong Capsule on mediating miR-216a-5p/TPT1/mTORC1 pathway to regulate benign prostate hyperplasia
Objective To investigate whether Qianlongtong Capsule(QLTC)can inhibit benign prostatic hyperplasia(BPH)by regulating the signaling pathway of miR-216a-5p/tumor protein translationally controlled 1/mammalian target of rapamycin complex 1(miR-216a-5p/TPT1/mTORC1)through cell experiments.Methods A total of 25 rats were randomized into control(equal volume of normal saline),low-(56.25 mg/mL),medium-(112.50 mg/mL),and high-dose(225.00 mg/mL)QLTC,and LBSC groups(168.75 mg/mL),with five rats in each group.The rats in each group were given corresponding drug 1 mL by gavage,twice a day,for continual five days.Drug-containing serum was prepared from rats in each group after anesthesia.According to different experimental purposes,CP-H022 cells were treated experimentally by five steps,and each part of the experiment was grouped independently.The comparative study on overexpression and silencing expression of miR-216a-5p,and TPT1 overexpression was performed;RT-qPCR was used to determine the expression levels of miR-216a-5p in normal and BPH model CP-H022 cells,and observe the differences of miR-216a-5p expression levels in BPH cells treated with different concentrations of QLTC;cell colony formation assay was used to examine cell proliferation;CCK-8 was used to test the proliferation of BPH model cells;RT-qPCR was used to determine the expression levels of miR-216a-5p and TPT1mRNA;cell apoptosis was checked by flow cytometry;bioinformatics analysis and dual luciferase assay were used to verify the targeting relationship between miR-216a-5p and TPT1;after overexpression of TPT1,Western blot was used to examine the expression of TPT1/mTORC1 signaling pathway related molecules in BPH cells.Results Compared with the control group,the expression level of miR-216a-5p in CP-H022 cells in the model group 1 downregulated(P<0.05);different concentrations of QLTC can upregulate the expression of miR-216a-5p;based on the results of this experiment,QLTC(high-dose)group CP-H022 cells will be selected for subsequent experiments in this study.Compared with the model group 2,cell proliferation decreased and apoptosis increased in QLTC group 2(P<0.05),Bcl-2 expression decreased(P<0.05),and expressions of Bax and cleaved Caspase-3 increased(P<0.05);compared with the model group 4,after knocking down miR-216a-5p,the QLTC group 4 showed higher cell proliferation and lower apoptosis(P<0.05),increased expression of Bcl-2(P<0.05),and decreased expressions of Bax and cleaved Caspase-3(P<0.05).Compared with the mimic-NC group,the expression of TPT1 of miR-216a-5p mimic group decreased(P<0.05);after QLTC treatment,the expressions of TPT1 and p-mTORC1 in cells decreased(P<0.05);after overexpression of TPT1,the proliferation function of BPH cells was higher(P<0.05),apoptosis was lower(P<0.05),Bcl-2 expression increased(P<0.05),and expressions of Bax and Cleaved Caspase-3 decreased(P<0.05).Conclusion QLTC can inhibit benign prostatic hyperplasia by mediating miR-216a-5p to downregulate TPT1/mTORC1 pathway.
Qianlongtong Capsulebenign prostatic hyperplasiacell experimentmiR-216a-5ptumor protein transla-tionally controlled 1mammalian target of rapamycin complex 1signaling pathway
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维普
