Jian Jin

MiR-489 regulates chemoresistance in breast cancer via epithelial mesenchymal transition pathway.

To investigate the role of microRNAs in the development of chemoresistance and related epithelial–mesenchymal transition (EMT), we examined the effect of miR‐489 in adriamycin (ADM)‐resistant human breast cancer cells (MCF‐7/ADM). MiR‐489 was significantly suppressed in MCF‐7/ADM cells compared with chemosensitive parental control MCF‐7/WT cells. Forced‐expression of miR‐489 reversed chemoresistance. Furthermore, Smad3 was identified as the target of miR‐489 and is highly expressed in MCF‐7/ADM cells. Forced expression of miR‐489 both inhibited Smad3 expression and Smad3 related EMT properties. Finally, the interactions between Smad3, miR‐489 and EMT were confirmed in chemoresistant tumor xenografts and clinical samples, indicating their potential implication for treatment of chemoresistance.

A Methylation-Based Regulatory Network for MicroRNA 320a in Chemoresistant Breast Cancer

We previously demonstrated that the overexpression of transient receptor potential channel C5 (TRPC5) and nuclear factor of activated T-cells isoform c3 (NFATC3) are essential for cancer chemoresistance, but how TRPC5 and NFATC3 are regulated was still unclear. In this study, microRNA 320a (miR-320a) was found to be down-regulated in chemoresistant cancer cells. MiR-320a directly targeted TRPC5 and NFATC3, and down-regulation of miR-320a triggered TRPC5 and NFATC3 overexpression. In chemoresistant cells, down-regulation of miR-320a was associated with regulation by methylation, which implicated promoter methylation of the miR-320a coding sequence. Furthermore, the transcription factor v-ets erythroblastosis virus E26 oncogene homolog 1 (ETS-1), which inhibited miR-320a expression, was activated in chemoresistant cancer cells; such activation was associated with hypomethylation of the ETS-1 promoter. Lastly, the down-regulation of miR-320a and high expression of TRPC5, NFATC3, and ETS-1 were verified in clinically chemoresistant samples. Low expression of MiR-320a was also found to be a significant unfavorable predictor for clinic outcome. In conclusion, miR-320a is a mediator of chemoresistance by targeting TRPC5 and NFATC3. Expression of miR-320a is regulated by methylation of its promoter and that of ETS-1.

Methylation-regulated miR-149 modulates chemoresistance by targeting GlcNAc N-deacetylase/N-sulfotransferase-1 in human breast cancer

Dysregulation of microRNA is strongly implicated in the chemoresistance of cancer. In this study, we found that miR‐149 was downregulated and involved in chemoresistance in adriamycin (ADM)‐resistant human breast cancer cells (MCF‐7/ADM). Downregulation of miR‐149 was related to hypermethylation of its 5′‐UTR; this methylation also affected the expression of the glypican 1 gene, which is both the host and the target gene of miR‐149. Furthermore, we found that miR‐149 modulated chemoresistance through targeting the expression of GlcNAc N‐deacetylase/N‐sulfotransferase‐1 (NDST1). With downregulated miR‐149, NDST1 expression was increased in chemoresistant MCF‐7/ADM cells versus control MCF‐7 wild‐type cells. The increased NDST1 then activated a heparan sulfate‐related pathway involving activation of heparanase. Finally, expression of miR‐149 and NDST1 was confirmed in clinical chemoresistant samples of breast cancers receiving anthracycline/taxane‐based chemotherapies. The high expression of NDST1 was also an unfavorable predictor for distant relapse‐free survival in Her2 and basal breast cancers. Taken together, our findings demonstrate that miR‐149 is regulated by methylation, and is a modulator of cancer chemoresistance by targeting NDST1.

Acquisition of paclitaxel resistance via PI3K‑dependent epithelial‑mesenchymal transition in A2780 human ovarian cancer cells.

Epithelial ovarian cancer is a major cause of mortality among women with gynecological malignancies. Paclitaxel is commonly used for chemotherapy of ovarian cancer, yet its efficacy is limited by chemoresistance. Generally, drug resistance is associated with acquisition of the epithelial-mesenchymal transition (EMT) in cancer. The aim of the present study was to determine whether the EMT is involved in acquired resistance to paclitaxel in A2780 human ovarian cancer cells. Using the paclitaxel-resistant A2780/PTX cell line, we examined the cellular morphology, molecular changes, migration and proliferation consistent with the EMT. Furthermore, we found that inhibition of phosphatidylinositol 3-kinase (PI3K) activity reduced the proliferation and migration and restored their sensitivity to paclitaxel. Our study provides new insights into EMT-like phenotypic changes that are linked to paclitaxel resistance in A2780 cells. We believe that inhibition of the PI3K signaling pathway could provide a novel therapeutic approach to overcome chemoresistance and prevent metastasis during paclitaxel chemotherapy.

Apigenin, a plant‐derived flavone, activates transient receptor potential vanilloid 4 cation channel

BACKGROUND AND PURPOSE Transient receptor potential vanilloid 4 (TRPV4) is a Ca2+‐permeable channel with multiple modes of activation. Apigenin is a plant‐derived flavone, which has potential preventive effects on the development of cardiovascular disease. We set out to explore the effects of apigenin on TRPV4 channel activity and its role in vasodilatation.

Tumor endothelial expression of P-glycoprotein upon microvesicular transfer of TrpC5 derived from adriamycin-resistant breast cancer cells

Treatment of carcinoma commonly fails due to chemoresistance. Studies have shown that endothelial cells acquire resistance via the tumor microenvironment. Microvesicle (MV) shedding from the cell membrane to the microenvironment plays an important role in communication between cells. The aim of the present study was to determine whether MCF-7 adriamycin-resistant cells (MCF-7/ADM) shed MVs that alter the characteristics of human microvessel endothelial cells (HMECs). MVs from tumor cells transferred a Ca(2+)-permeable channel TrpC5 to HMECs, inducing the expression of P-glycoprotein (P-gp) by activation of the transcription factor NFATc3 (nuclear factor of activated T cells isoform c3). Expression of the mdr1 gene was blocked by the TrpC5-blocking antibody T5E3, and the production of P-gp in HMECs was reduced by blockade of TrpC5. Thus, we postulate that endothelial cells acquire the resistant protein upon exposure to TrpC5-containg MVs in the microenvironment, and express P-gp in the TrpC5-NFATc3 signal pathway.

Inhibition of Transient Receptor Potential Channel 5 Reverses 5-Fluorouracil Resistance in Human Colorectal Cancer Cells

Background: Resistance to 5-fluorouracil leads to the failure of chemotherapy for colorectal cancer. Results: Suppressing TrpC5 expression decreased nuclear β-catenin accumulation, reduced the induction of ABCB1, and reversed 5-fluorouracil resistance. Conclusion: TrpC5 is essential in ABCB1 induction and drug resistance in human colorectal cancer cells. Significance: These findings may help develop a novel target for overcoming resistance to chemotherapy in colorectal cancer. 5-Fluorouracil (5-Fu) is commonly used in the chemotherapy of colorectal cancer (CRC), but resistance to 5-Fu occurs in most cases, allowing cancer progression. Suppressing ABCB1 (ATP-binding cassette, subfamily B, member 1), which is a pump overproduced in cancer cells to export cytotoxic drugs, is an attractive strategy to overcome drug resistance. In the present study, transient receptor potential channel TrpC5 was found to be overproduced at the mRNA and protein levels together with ABCB1 in 5-Fu-resistant human CRC HCT-8 (HCT-8/5-Fu) and LoVo (LoVo/5-Fu) cells. More nuclear-stabilized β-catenin accumulation was found in HCT-8/5-Fu and LoVo/5-Fu cells than in HCT-8 and LoVo cells. Suppressing TrpC5 expression with TrpC5-specific siRNA inhibited the canonical Wnt/β-catenin signal pathway, reduced the induction of ABCB1, weakened the ABCB1 efflux pump, and caused a remarkable reversal of 5-Fu resistance in HCT-8/5-Fu and LoVo/5-Fu cells. On the contrary, enforcing TrpC5 expression resulted in an activated Wnt/β-catenin signal pathway and up-regulation of ABCB1. Taken together, we demonstrated an essential role of TrpC5 in ABCB1 induction and drug resistance in human CRC cells via promoting nuclear β-catenin accumulation.

Total flavonoids of Flos Chrysanthemi protect arterial endothelial cells against oxidative stress

ETHNOPHARMACOLOGICAL RELEVANCE Total flavonoids of Flos Chrysanthemi (TFFC) are known to modulate vascular functions, but their effect on endothelial cells injured by oxidative stress is unknown. Our objective was to investigate the vasoprotective effect and mechanism of action of TFFC on rat mesenteric artery exposed to superoxide anions produced by pyrogallol. MATERIALS AND METHODS The vasoprotective effect and mechanism of action of TFFC on primary cultured rat mesenteric arterial endothelial cells and small mesenteric arteries was investigated using small-vessel myography, fluorescent Ca(2+) measurement, fluorescent membrane potential measurement and oxidative fluorescent studies. RESULTS Experiments using small-vessel myography of third-order rat mesenteric arterial rings showed that pretreatment with pyrogallol (10-1000μM), an auto-oxidizing source of superoxide anions, dose-dependently decreased ACh-induced endothelium-dependent relaxation. TFFC (2.5-320mg/L) evoked a concentration-dependent dilation (pD(2): 29.6±0.276mg/L), which was weakened by ChTX plus apamin. TFFC markedly attenuated the inhibition of vasorelaxation induced by pyrogallol (E(max) elevated from 50.4±7.36% to 86.2±3.61%, and pD(2) increased from 6.74±0.06 to 7.28±0.12). Furthermore, in primary cultured endothelial cells, fluorescent Ca(2+) measurement, fluorescent membrane potential measurement and oxidative fluorescent studies demonstrated that ACh-induced endothelial Ca(2+) influx and hyperpolarization were significantly weakened by the increased basal superoxide level induced by pyrogallol. When the endothelial cells were concurrently exposed to TFFC, the impairment effect of oxidative stress on ACh-induced Ca(2+) influx, hyperpolarization and vasorelaxation were attenuated due to its superoxide-lowering activity. CONCLUSION This study shows that oxidative stress has a pronounced deleterious effect on EDHF-mediated vasorelaxation to ACh in rat mesenteric artery. TFFC has vasodilating effect and protects EDHF-mediated vasodilator reactivity from oxidative stress. Thus, our experiments suggest that TFFC is potentially useful for the development of therapeutic treatments for cardiovascular diseases associated with oxidative stress.

Enhancement of vascular endothelial growth factor release in long-term drug-treated breast cancer via transient receptor potential channel 5-Ca2+-hypoxia-inducible factor 1α pathway

Chemotherapy targeting anti-angiogenesis in tumors may have insufficient efficacy, but little is known about the underlying mechanisms. Here, we showed that the Ca(2+)-permeable channel, TrpC5, is highly expressed in human breast cancer after long-term chemotherapy drug-treatment. It mediates downstream hypoxia-inducible factor 1α accumulation in the nucleus, and then activates the transcription of vascular endothelial growth factor which promotes tumor angiogenesis, leading to a poor chemotherapeutic outcome. We verified this mechanism at both the cellular and xenograft levels. Moreover, in samples from patients, high TrpC5 expression was correlated with enhanced tumor vasculature after chemotherapy. Taken together, our research demonstrated the essential role of TrpC5 in tumor angiogenesis when facing the challenge of chemotherapy and presents a new potential target for overcoming the high vasculature of human breast cancer after chemotherapy.

A2780 human ovarian cancer cells with acquired paclitaxel resistance display cancer stem cell properties

The use of chemotherapy to treat cancer is effective, but chemoresistance reduces this efficacy. Chemotherapy resistance involves several mechanisms, including the cancer stem cell (CSC) concept. The aim of the present study was to assess whether paclitaxel-resistant epithelial ovarian carcinoma is capable of generating cells with CSC-like properties. Using the paclitaxel-resistant A2780/PTX cell line, it was demonstrated that high aldehyde dehydrogenase 1 (ALDH1) activity identifies CSCs from diverse sources. Furthermore, the A2780/PTX cells had a strong ability to form colonies in soft agar assays. Notably, it was demonstrated that the inhibition of the PI3K signaling pathway abolished colony formation. These data suggest that there is a link between paclitaxel resistance and CSC enrichment. It is possible that therapeutic benefits, such as the restoration of chemosensitivity or the suppression of tumorigenicity, may be enabled by gaining further insights into the mechanisms underlying chemoresistance and the generation of CSCs.