Comparative proteomic analysis of the insulin-induced L6 myotube secretome.
Emerging evidence has revealed an endocrine function for skeletal muscle; in fact, certain anti-inflammatory cytokines are secreted only from contractile skeletal muscle. However, the skeletal muscle secretome as a whole is poorly characterized, as is how it changes in response to extracellular stimuli. Herein, we sought to identify and characterize the members of the skeletal muscle secretome, and to determine which protein secretion levels were modulated in response to insulin stimulation. To conduct these studies, we treated differentiated L6 rat skeletal muscle cells with insulin or left them untreated, and we comparatively analyzed the proteins secreted into the media. We fractionated this conditioned media using offline RP HPLC, digested the fractionated proteins, and analyzed the resulting peptides with LC-ESI-MS/MS. We identified a total of 254 proteins, and by using three different filtering methods, we identified 153 of these as secretory proteins. Fourteen proteins were secreted at higher levels under insulin stimulation, including several proteins known to be highly secreted in metabolic diseases; 19 proteins were secreted at lower levels under insulin stimulation. These result not only pinpointed several previously unknown, insulin induced, secretory proteins of skeletal muscle, it also described a novel approach for conditioned secretome analysis.
Yoon
Jong Hyuk
JH
Division of Molecular and Life Sciences, Pohang University of Science and Technology, Pohang, Republic of Korea.
Yea
Kyungmoo
K
Kim
Jaeyoon
J
Choi
Yoon Sup
YS
Park
Sehoon
S
Lee
Hyeongji
H
Lee
Chang Sup
CS
Suh
Pann-Ghill
PG
Ryu
Sung Ho
SH
D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media / Q000737:chemistry; D007328:Insulin / Q000378:metabolism*; D013058:Mass Spectrometry; D053858:Metabolic Networks and Pathways; D008954:Models, Biological; D018485:Muscle Fibers, Skeletal / Q000737:chemistry* / Q000378:metabolism*; D009124:Muscle Proteins / Q000032:analysis*; D040901:Proteomics; D051381:Rats
D003470:Culture Media; D007328:Insulin; D009124:Muscle Proteins