Acquisition of resistance to anti-cancer drugs is a significant obstacle to effective cancer treatment. Although several efforts have been made to overcome drug resistance in cancer cells, the detailed mechanisms have not been fully elucidated. Here, we investigated whether microRNAs (miRNAs) function as pivotal regulators in the acquisition of anti-cancer drug resistance to 5-fluorouracil (5-FU). A survey using a lentivirus library containing 572 precursor miRNAs revealed that five miRNAs promoted cell survival after 5-FU treatment in human hepatocellular carcinoma Hep3B cells. Among the five different clones, the clone expressing miR-200a-3p (Hep3B-miR-200a-3p) was further characterized as a 5-FU-resistant cell line. The cell viability and growth rate of Hep3B-miR-200a-3p cells were higher than those of control cells after 5-FU treatment. Ectopic expression of a miR-200a-3p mimic increased, while inhibition of miR-200a-3p downregulated, cell viability in response to 5-FU, doxorubicin, and CDDP (cisplatin). We also showed that dual-specificity phosphatase 6 (DUSP6) is a novel target of miR-200a-3p and regulates resistance to 5-FU. Ectopic expression of DUSP6 mitigated the pro-survival effects of miR-200a-3p. Taken together, these results lead us to propose that miR-200a-3p enhances anti-cancer drug resistance by decreasing DUSP6 expression.
Carcinoma, Hepatocellular ; Cell Line ; Cell Survival ; Clone Cells ; Doxorubicin ; Drug Resistance ; Dual Specificity Phosphatase 6* ; Dual-Specificity Phosphatases* ; Ectopic Gene Expression ; Fluorouracil* ; Humans ; Lentivirus ; MicroRNAs

BACKGROUNDThe MAPK phosphatases (MKPs) are a family of dual-specificity phosphatases (DUSPs) that can dephosphorylate both phosphothreonine and phosphotyrosine residues, thus inactivating MAPK signaling. DUSP6 is a cytoplasmic MKP that can inactivate ERK. DUSP6 has been implicated in the development of some tumors. The aim of this research was to investigate the expression of DUSP6 in hepatocellular carcinoma (HCC) and the correlation of DUSP6 with mitogen-activated protein kinases (MAPKs), clinicopathological characteristics, and prognosis.

METHODSTissues from 305 patients who had undergone hepatectomy for HCC was used in this study. The expression of DUSP6, p-ERK, p-JNK, and p-p38α was determined using tissue microarrays for immunohistochemical analysis. The prognostic value of DUSP6 and other clinicopathological factors were evaluated.

RESULTSThe expression of DUSP6 was significantly higher in the tumor tissue when compared to the peritumor or normal liver tissue (P < 0.001). Tumor DUSP6 expression was significantly associated with disease-free survival (DFS) (P = 0.013). Tumor DUSP6 expression was an independent prognostic factor for DFS (Hazard ratio = 1.635, P = 0.006).

CONCLUSIONSDUSP6 is over expressed in tumor tissue compared to peritumor or normal liver tissue. Higher expression of DUSP6 in tumor tissue, than in peritumor tissue, is associated with the recurrence after curative resection of HCC, and the relative tumor DUSP6 expression has good power to predict the recurrence of HCC.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Hepatocellular ; metabolism ; Dual Specificity Phosphatase 6 ; metabolism ; Female ; Humans ; Immunohistochemistry ; Liver Neoplasms ; metabolism ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; metabolism ; Tissue Array Analysis ; Young Adult

12(S)-Hydroxyheptadeca-5Z,8E,10E-trienoic acid (12-HHT) is an enzymatic product of prostaglandin H2 (PGH2) derived from cyclooxygenase (COX)-mediated arachidonic acid metabolism. Despite the high level of 12-HHT present in tissues and bodily fluids, its precise function remains largely unknown. In this study, we found that 12-HHT treatment in HaCaT cells remarkably down-regulated the ultraviolet B (UVB) irradiation-induced synthesis of interleukin-6 (IL-6), a pro-inflammatory cytokine associated with cutaneous inflammation. In an approach to identify the down-stream signaling mechanism by which 12-HHT down-regulates UVB-induced IL-6 synthesis in keratinocytes, we observed that 12-HHT inhibits the UVB-stimulated activation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-kappaB). In addition, we found that 12-HHT markedly up-regulates MAPK phosphatase-1 (MKP-1), a critical negative regulator of p38 MAPK. When MKP-1 was suppressed by siRNA knock-down, the 12-HHT-mediated inhibitory effects on the UVB-stimulated activation of p38 MAPK and NF-kappaB, as well as the production of IL-6, were attenuated in HaCaT cells. Taken together, our results suggest that 12-HHT exerts anti-inflammatory effect via up-regulation of MKP-1, which negatively regulates p38 MAPK and NF-kappaB, thus attenuating IL-6 production in UVB-irradiated HaCaT cells. Considering the critical role of IL-6 in cutaneous inflammation, our findings provide the basis for the application of 12-HHT as a potential anti-inflammatory therapeutic agent in UV-induced skin diseases.
Anti-Inflammatory Agents, Non-Steroidal/pharmacology ; Cell Line ; Dual Specificity Phosphatase 1/biosynthesis/genetics ; Enzyme Activation ; Fatty Acids, Unsaturated/*pharmacology ; Humans ; Interleukin-6/*biosynthesis ; Keratinocytes/*metabolism/radiation effects ; NF-kappa B/metabolism ; RNA Interference ; RNA, Small Interfering ; Receptors, Leukotriene B4/genetics ; Signal Transduction/drug effects ; Skin Diseases/drug therapy ; *Ultraviolet Rays ; Up-Regulation ; p38 Mitogen-Activated Protein Kinases/metabolism