calpain and Carcinoma--Ovarian-Epithelial

Circular RNA ABCB10 (circ‑ABCB10) modulates cellular functions and microRNA (miR)‑1271 in epithelial ovarian cancer (EOC). The present study aimed to investigate the interaction between circ‑ABCB10 and miR‑1271 in regulating EOC cellular function and the calpain small subunit 1 (Capn4)/Wnt/β‑catenin signaling pathway. circ‑ABCB10 and miR‑1271 expression levels were detected in EOC cells (OVCAR3, UWB1.289, SKOV3 and CAOV3) and normal ovarian epithelial cells (IOSE80) via reverse‑transcription quantitative PCR. SKOV3 cells were transfected with control short hairpin (sh)RNA plasmids, control inhibitor, circ‑ABCB10 shRNA plasmids and miR‑1271 inhibitor. UWB1.289 cells were transfected with control overexpression plasmids, control mimic, circ‑ABCB10 overexpression plasmids and miR‑1271 mimic. Subsequently, cell proliferation, apoptosis, invasion and the Capn4/Wnt/β‑catenin signaling pathway were assessed. In addition, a luciferase activity assay was performed. circ‑ABCB10 expression was significantly increased in OVCAR3, SKOV3 and CAOV3 cells compared with IOSE80 cells, but was not significantly altered in UWB1.289 cells. miR‑1271 expression was significantly decreased in OVCAR3, UWB1.289, SKOV3 and CAOV3 cells compared with IOSE80 cells. In both SKOV3 and UWB1.289 cells, circ‑ABCB10 negatively regulated miR‑1271, whereas miR‑1271 did not affect circ‑ABCB10. Furthermore, circ‑ABCB10 enhanced cell proliferation, invasion and the Capn4/Wnt/β‑catenin signaling pathway, but inhibited cell apoptosis, whereas miR‑1271 suppressed cell proliferation, invasion and the Capn4/Wnt/β‑catenin signaling pathway, but facilitated cell apoptosis. Moreover, miR‑1271 attenuated the proproliferative, proinvasive and antiapoptotic effects of circ‑ABCB10, and reversed the positive regulation of circ‑ABCB10 on the Capn4/Wnt/β‑catenin signaling pathway. Besides, the luciferase activity assay indicated that circ‑ABCB10 directly bound to miR‑1271. In conclusion, the present study indicated that circ‑ABCB10 promoted cell proliferation and invasion, and suppressed apoptosis by regulating the miR‑1271‑mediated Capn4/Wnt/β‑catenin signaling pathway in EOC.

Topics: Apoptosis; ATP-Binding Cassette Transporters; Calpain; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Neoplasm Invasiveness; RNA, Circular; RNA, Long Noncoding; RNA, Small Interfering; Wnt Signaling Pathway

Increasing evidence shows that the calpain regulatory subunit Capn4 can modulate the proliferation and metastasis of cancer cells, and plays an important role in the development of malignant tumors. However, there is no information on the clinical significance of Capn4 in epithelial ovarian carcinoma (EOC) or the molecular mechanisms by which Capn4 promotes the growth and metastasis of EOC. Therefore, the aim of this study was to clarify the role of Capn4 in EOC.. We evaluated Capn4 and osteopontin (OPN) expression in EOC cell lines and tissues from patients with ovarian cancer by western blotting and immunohistochemical analysis. We then created cell lines with downregulated and upregulated Capn4 expression, using Capn4-targeting small interfering RNA and a pcDNA3.1-Capn4 overexpression vector, respectively, to investigate its function in EOC in vitro. In addition, we investigated the potential mechanism underlying the function of Capn4 by examining the effect of modifying Capn4 expression on Wnt/β-catenin signaling pathway-related genes by western blotting.. Capn4 was overexpressed in clinical EOC tissues compared with that in normal ovarian epithelial tissue, and was associated with poor clinical outcomes. Upon silencing or overexpressing Capn4 in EOC cells, we concluded that Capn4 promotes cell proliferation and migration in vitro. Furthermore, Capn4 promoted EOC metastasis by interacting with the Wnt/β-catenin signaling pathway to upregulate OPN expression.. Our study indicates that Capn4 plays a critical role in the progression and metastasis of EOC, and could be a potential therapeutic target for EOC management.

Topics: beta Catenin; Calpain; Carcinoma, Ovarian Epithelial; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Movement; Cell Proliferation; Down-Regulation; Female; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Middle Aged; Neoplasm Metastasis; Neoplasms, Glandular and Epithelial; Osteopontin; Ovarian Neoplasms; Plasmids; RNA Interference; RNA, Small Interfering; Up-Regulation; Wnt Proteins; Wnt Signaling Pathway

Peritoneal dissemination and retroperitoneal lymph node involvement are main routes for tumour spread of epithelial ovarian cancer (EOC), possibly determined by the intercellular connecting protein E-Cadherin (E-Cad) and its fragments.. Tumour tissue of 105 advanced EOC patients was evaluated for protein expression of E-Cad, β-Catenin and Calpain by western blotting and immunohistochemistry. Expression patterns were compared between tumours with solely intraperitoneal (pT3c, pN0; n=41) and tumours with retroperitoneal metastases (pT1a-3c, pN1; n=64). Lysates of the EOC cell line SKOV3 and tumour tissue from the intraperitoneal group were tested for E-Cad expression following Calpain treatment.. E-Cad full-length (E-Cad-FL, 120 kDa) and two major fragments at 85 kDa (E-Cad-85) and 23 kDa (E-Cad-23) were detected by western blotting. E-Cad-85 expression was significantly higher in tumours with solely intraperitoneal metastases and correlated strongly with E-Cad-23 and the protease Calpain. Calpain-mediated cleavage was identified as a potential mechanism to generate E-Cad-85 from E-Cad-FL by treating lysates from SKOV3 cells and tumour tissue with this enzyme. Increased cytoplasmic localisation of β-Catenin in tumours with high E-Cad-85 expression corroborates that E-Cad-85 loses the binding site for β-Catenin after fragmentation, enabling tumour cluster formation and peritoneal dissemination.. Calpain-mediated E-Cad fragmentation appears to promote intraperitoneal EOC progression. Understanding these mechanisms might eventually lead to new tailored subtype-specific diagnostic and therapeutic interventions.

Topics: Adult; Aged; Aged, 80 and over; beta Catenin; Blotting, Western; Cadherins; Calpain; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Female; Humans; Immunohistochemistry; Lymph Nodes; Middle Aged; Neoplasms, Glandular and Epithelial; Ovarian Neoplasms; Peptide Fragments; Peritoneal Neoplasms; Retroperitoneal Neoplasms; Retroperitoneal Space; Retrospective Studies