Orthopeadic Surgery
The inhibitor of cyclin-dependent kinase 4a signaling pathway induces aging in human skeletal muscle myoblasts and decreases the mitochondrial membrane potential
Shuo Tang, Hong Gao, Dong Yan, Juan Yang, Zhuolun Li, Pengtao Hu, Donglin Xiong, Xiang Liao
Published 2017-06-08
Cite as Chin J Exp Surg, 2017, 34(6): 924-926. DOI: 10.3760/cma.j.issn.1001-9030.2017.06.007
Abstract
ObjectiveTo study the effect of the inhibitor of cyclin-dependent kinase 4a (INK4a) signaling pathway on myoblastic aging.
MethodsWe transfected human skeletal muscle myoblasts with a recombinant lentiviral vector, pLVX-p16INK4a, encoding the p16INK4a gene, and real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) and Western blotting were used to identify p16INK4a gene transcription and protein expression. Senescence-associated β-galactosidase staining was used to assess the degree of cell senescence. The mitochondrial membrane potential was analyzed by JC-1 staining and flow cytometry.
ResultsWe demonstrated that a senescence phenotype was evident in myoblasts transfected with p16INK4a, the analysis of mitochondrial membrane potential after JC-1 staining by flow cytometry showed a marked decrease in p16 group (-40.287±12.663) mV, while it was (-9.267±1.332) mV in CV group and (-6.967±2.031) mV in NC group. The relative amount of p16 protein and mRNA in p16 group was 45.25±8.21 and 33.89±7.87, which was more than that in CV (10.18±2.69 and 5.26±1.92)and NC (9.41±1.70 and 7.16±1.86) groups (P=0.000).
ConclusionThese findings indicated that upregulation of the INK4a signaling pathway directly induced aging in human skeletal muscle myoblasts. Moreover, INK4a signaling pathway activates the mitochondrial pro-aging pathway by reducing the mitochondrial membrane potential, which indirectly accelerates aging in myoblasts.
Key words:
Aging; Human skeletal muscle myoblast; Inhibitor of cyclin-dependent kinase 4a; Mitochondrial membrane potential
Contributor Information
Shuo Tang
Department of Pain Medicine, the Nanshan District People’s Hospital, Shenzhen 518052, China
Hong Gao
Dong Yan
Juan Yang
Zhuolun Li
Pengtao Hu
Donglin Xiong
Xiang Liao