Identification of novel changes in skeletal muscle proteome after sildenafil treatment

Insulin Resistance (IR) is associated with increased matrix proteins, including collagens, fibronectins, and laminins in the endomysium surrounding skeletal muscle. The increased ECM deposition in skeletal muscle IR results in ECM stiffening that decreases muscle integrity and cellular signaling. Recent evidence has shown that Sildenafil, a phosphodiesterase 5a inhibitor, improves insulin sensitivity in humans, and in high fat diet mice, reduces collagen deposition in the matrix surrounding skeletal muscle. In addition, data shows that a short-term treatment of Sildenafil for 7 days increases protein synthesis and decreases muscle fatigue in humans, which may mitigate the effects of IR. However, very few studies have examined the molecular mechanisms and metabolic signaling directly associated with Sildenafil’s influence on skeletal muscle. The aim of our study was to perform an unbiased proteomic analysis on Sildenafil’s effects on skeletal muscle in C2C12 myotubes. We hypothesized that Sildenafil would alter the abundance of proteins in metabolic pathways that are involved in communicating with the ECM. We performed a dose response (n = 3 for each dose; 0.5 μM, 1 μM, 2.5 μM, 5 μM, and 10 μM) and time response (n = 3 for each time; 30 min, 1 hour, 4 hour, 6 hour) curve. Proteomic analyses revealed that the highest number of proteins significantly changing was at 1 μM for 30 min (n = 423). Analysis of the significant proteins revealed 396 and 27 proteins that increased and decreased, respectively. KEGG analysis revealed that 20% of the total changing proteins (n = 83) were in metabolic pathways. The top 10 metabolic protein increasers, represented as a fold change of abundance compared to vehicle, were ASCL3 (1.27 ± 0.08), GT251 (1.12 ± 0.10), ODPA (1.15 ± 0.03), PCKGM (1.08 ± 0.04), PCY1A (1.23 ± 0.04), PGM1 (1.06 ± 0.04), PTG1S (1.21 ± 0.05), PYR1 (1.37 ± 0.09). SAHH (1.19 + 0.07), and THIM (1.13 ± 0.08). Of the 83 metabolic proteins, only 6 were decreasing, P5CR2 (0.78 ± 0.04), ALD1 (0.82 ± 0.11), P4K2A (0.67 ± 0.05), DGAT1 (0.27 ± 0.19), SCLY (0.77 ± 0.15), and PLCD3 (0.61 ± 0.16). The results from this preliminary study conclude that Sildenafil affects proteins involved in metabolic pathways. Further analysis will be performed to confirm the roles of these proteins in relation to ECM signaling/communication in these metabolic pathways. University of Arizona's Research, Innovation & Impact Faculty Seed Grant This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.