The Effect of Nutrient Deprivation on Markers of Oxidative Stress, Inflammation, and Transcriptome in Normal and Type-2 Diabetic Human Skeletal Muscle Myoblasts.

Abstract

Background: Intermittent fasting has become a new fad diet that may promote an environment to facilitate muscle atrophy, placing aging, and diabetic populations at risk for muscle loss due to nutrient deprivation. The purpose of this study was to investigate how nutrient availability and serum environment influence Type 2 diabetic myoblast density and viability, autophagy-related oxidative and inflammatory markers, and upstream gene expression signaling relevant to proteostasis. Methods: To explore this outcome in human skeletal muscle myoblast (HSMM) and diabetic human skeletal muscle myoblast (D-HSMM), myoblast lines were cultured per standard protocol and were incubated for 12 or 24 h with either human serum (HS) or fetal bovine serum (FBS) at varying serum media concentrations: 5%, 10%, and 15%. Cell viability and density were determined; ELISAs were used to assess SOD1 and TNFα; TaqMan gene array plates were used to explore mRNA gene expression related to growth and atrophy. Results: Cell viability (%) analysis showed that 0% concentration, 12 h incubation, and FBS media have lower viability (p ≤ 0.0001); cell density analysis showed lower values in 0% concentration and in the FBS media (p ≤ 0.0001); SOD1 analysis showed a scaled effect (p ≤ 0.05) and higher concentration in HS (12,795.07 ± 677.87 pg/mL; p ≤ 0.0001); TNFα concentration was higher in HSMM compared to D-HSMM (61 ± 0.82 vs. 2.52 ± 0.94 pg/mL; p=0.017), higher at 12 h (6.07 ± 0.88 vs. 2.50 ± 0.88 pg/mL; p=0.006), and higher in FBS (6.05 ± 0.88 vs. 2.08 ± 0.88 pg/mL; p=0.002); no meaningful increase in fold change was seen in mRNA. Conclusions: Myoblast viability and density were lower in the nutrient-deprived conditions and in the FBS compared to the HS serum. The biomarker of oxidative stress was lower in the serum concentration in a scaled effect, yet higher in HS. The biomarker of inflammation was higher in the HSMM cell line, shorter incubation time period, and in FBS. HS used as a media may supply nutrients and hormonal confounders that may support or stress myoblast growth.