探討腫瘤分泌的胞外體的前胸腺素在肌肉細胞經由TLR4- NF-κB- E3 ubiquitin ligase訊息傳遞引發癌症惡病質的肌肉萎縮
謝嘉興a、蕭璦莉c、吳昭良b
衛生福利部 台南醫院 泌尿科a, 國立成功大學醫學院 生化所b，國立成功大學醫學院 微免所c
Transfer of prothymosin-α in cancer-derived exosome triggers muscle atrophy in cancer cachexia via TLR4-NF-κB-E3 ubiquitin ligase signaling pathway
Gia-Shing Shieha, Ai-Li Shiauc, and Chao-Liang Wub
a Department of Urology, Tainan Hospital, Ministry of Health and Welfare, Taiwan,
b Department of Biochemistry and Molecular Biology, National Cheng Kung University Medical College, Tainan, Taiwan,
c Department of Microbiology and Immunology, National Cheng Kung University Medical College, Tainan, Taiwan
Purpose: Cachexia is a metabolic disease with the wasting of skeletal muscle and the decrease in body fat. Prothymosin-α (ProT) involved in cell proliferation, apoptosis, oxidative stress, and immunomodulation. In previous study, a statistically significant overexpression of ProT was found in human bladder carcinoma. The cancer-derived exosomes, which contained proteins, mRNA, miRNAs, and DNAs, were reported for the connection between cancers and distant organs, and supported survival of incoming metastatic cells. Recently, tumor cells secreted exosomes to induce muscle wasting in mice and muscle cells. In this study, we will evaluate whether the cancer-derived exosomal ProT induces cachexia-related muscle wasting.
Methods: Murine MBT-2 bladder cancer and C2C12 muscle cells were used in the study. Exosome from bladder cancer cells was extracted by ultracentrifugation. ProT and NF-ĸB were detected western blotting and immunofluorescence staining. The Q-PCR assays were performed for ProT and cytokine expressions. The expressions of ProT and TLR-4 were knocked down by lentivirus-mediated shRNA interference targeting ProT and TLR-4. MBT-2 cells were subcutaneously inoculated into C3H /HeN mice, followed by intratumoral injection with lentiviral vectors carrying shProT or shLuc. Cisplatin was intraperitoneally injected. Group differences were determined by Student t test. All statistical tests were two-sided.
Result: In sera from bladder cancer patients, our results show that cachexia groups express higher levels of ProT and IL-6 than non-cachexia counterparts. In MBT-2 cells, the ProT level is higher in exosome than in cellular extracts. C2C12 muscle cells treated with MBT-2 cell-derived exosome induce TGF-β, IL-1β, and IL-6 overexpressions. In contrast, C2C12 cells treated with exosome from ProT- knockdown MBT-2 cells reduce TGF-β, IL-1β, and IL-6 expressions. Furthermore, our results show exosomal ProT activates nuclear translocation of NF-ĸB via TLR-4 and induces overexpressions of cachexia factors. In a chemotherapy-induced cachexia mouse model, lentivirus-mediated knockdown of ProT alleviated the body weight decline in tumor-bearing mice. In addition, the muscle atrophy and cachexia-related E3 ubiquitin ligase are improved in the ProT-knockdown groups.
Conclusion: Our results suggested that exosomal ProT- cytokine-cell death axis played a pivotal role in muscle wasting of cancer cachexia. The inhibition of cancer-derived exosomal ProT and muscle cell interaction provided the therapeutic potential for cancer cachexia.