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  • br receptor positive MCF breast cancer cells Chen

    2020-08-28

    
    receptor-positive MCF-7 breast cancer cells (Chen and Chien, 2014). BBP and DBP were also reported to promote the proliferation and in-vasiveness of estrogen receptor-negative MDA-231 breast cancer cells through AhR/HDAC6/c-Myc signaling pathway (Hsieh et al., 2012). In our study we found that low dose of BBP increased the cell viability in both androgen receptor-positive LNCaP and androgen receptor-nega-tive PC-3 prostate cancer cells. BBP regulated the mRNA and protein levels of cell cycle related genes (cylinD1, PCNA and p21) to induce cell proliferation of LNCaP and PC-3 cells. These findings provided new insight into the promoting effect of BBP on prostate cancer cells.
    miR-34a is a tumor suppressor miRNA. Low expression of miR-34a was found in various types of solid and hematological malignancies including prostate, breast, 3Methyladenine and brain cancers (Li et al., 2014). Recent studies have shown that miR-34a may be a potential therapeutic target and predictive biomarker in prostate cancer progression (Corcoran et al., 2014). Since miR-34a may play a key role in the de-velopment of prostate cancer, we determined if miR-34a was involved in BBP-induced cell proliferation. We examined the expression of miR-34a in LNCaP and PC-3 cells after BBP treatment. We found that low dose of BBP significantly down-regulated the level of miR-34a and in-creased cell proliferation in two prostate cancer cell lines. Moreover, c-myc, an oncogene which plays a critical role in regulating cell cycle genes like cyclinD1, PCNA and p21 (Jackstadt and Hermeking, 2015), was reported to be a target of miR-34a in prostate cancer (Yamamura et al., 2012). Then we investigated the expression level of c-myc after BBP treatment. The results indicated that BBP treatment increased the mRNA and protein level of c-myc along with downregulation of miR-34a. Furthermore, we found that transfection of miR-34a mimic altered
    Fig. 3. BBP downregualted miR-34a ex-pression in LNCaP and PC-3 cells. Cells was treated with 10−7 and 10−6 mol/L of BBP for 6 days, and quantitative real-time PCR was used to detected level of miR-34a(A); the mRNA level of c-myc was detected by quantitative real-time PCR (B); Western blotting was used to analyze protein level of miR-34a target gene c-myc (C). Data are expressed as mean ± SD. *p < 0.05, **p < 0.01; compared with control group. All experiments were performed in tripli-cate.
    Fig. 4. miR-34a inhibited cell growth of prostate cancer cells. PC-3 cells were seeded on dishes, and the cells were transfected with miR-34a mimic, miR-34a inhibitor or negative control 24 h later. After incubated for 4 h, the medium was replaced with fresh medium and cultured for72h. The effect of miR-34a transfection was analyzed by quantitative real-time PCR (A); the effect of miR-34a on cell viability was detected by MTT assay(B); Western blotting was used to analyze the protein levels of c-myc, cylinD1, PCNA and p21 (C). Data are expressed as mean ± SD. *p < 0.05, **p < 0.01; compared with control mimic or control inhibitor group. All experiments were performed in triplicate.
    Fig. 5. BBP promoted cell growth via regulating miR-34a. PC-3 cells were seeded on dishes, and the cells were transfected with miR-34a mimic, miR-34a inhibitor or negative control 24 h later. After incubated for 4 h, the medium was replaced with culture medium contain 10−7 mol/L BBP and cultured for 6 days. The effect of BBP on cell viability after miR-34a transfection was detected by MTT assay (A) and (B). Data are expressed as mean ± SD. **p < 0.01; compared with control mimic or control inhibitor group. All experiments were performed in triplicate.
    Fig. 6. miR-34a/c-myc axis mediated BBP-induced cell proliferation of prostate cancer cells. Cells were seeded on dishes, and the cells were transfected with miR-34a mimic, miR-34a inhibitor or negative control 24 h later. After incubated for 4 h, the medium was replaced with culture medium contain 10−7 mol/L BBP and cultured for 6 days. The expression levels of cell cycle related proteins and miR-34a target gene c-myc were detected by Western blotting. All experiments were performed in triplicate.
    the expression of cell cycle related genes (c-myc, cyclinD1, PCNA and p21) and inhibited cell growth of prostate cancer cells. The effect of miR-34a inhibitor also indicated the crucial regulation of miR-34a in the proliferation of prostate cancer cells. In order to verify the role of miR-34a/c-myc axis in BBP-induced cell proliferation, miR-34a mimic and inhibitor were transfected in combination with BBP treatment. The results showed that BBP treatment alone decreased miR-34a expression level, increased c-myc level and promoted cell growth; while transfec-tion of miR-34a mimic diminished BBP-induced c-myc upregulation and cell cycle promotion. Moreover, the opposite effects were observed in cells transfected with miR-34a inhibitor Together, these findings revealed the key role of miR-34a/c-myc axis in BBP-induced cell pro-liferation of prostate cancer cells.