Padhy, Gayatri, Niroj Kumar Sethy, Lilly Ganju, and Kalpana Bhargava. Abundance of plasma antioxidant proteins confers tolerance to acute hypobaric hypoxia exposure. High Alt Med Biol 14:289-297, 2013-Systematic identification of molecular signatures for hypobaric hypoxia can aid in better understanding of human adaptation to high altitude. In an attempt to identify proteins promoting hypoxia tolerance during acute exposure to high altitude, we screened and identified hypoxia tolerant and susceptible rats based on hyperventilation time to a simulated altitude of 32,000 ft (9754 m). The hypoxia tolerance was further validated by estimating 8-isoprotane levels and protein carbonyls, which revealed that hypoxia tolerant rats possessed significant lower plasma levels as compared to susceptible rats. We used a comparative plasma proteome profiling approach using 2-dimensional gel electrophoresis (2-DGE) combined with MALDI TOF/TOF for both groups, along with an hypoxic control group. This resulted in the identification of 19 differentially expressed proteins. Seven proteins (TTR, GPx-3, PON1, Rab-3D, CLC11, CRP, and Hp) were upregulated in hypoxia tolerant rats, while apolipoprotein A-I (APOA1) was upregulated in hypoxia susceptible rats. We further confirmed the consistent higher expression levels of three antioxidant proteins (PON1, TTR, and GPx-3) in hypoxia-tolerant animals using ELISA and immunoblotting. Collectively, these proteomics-based results highlight the role of antioxidant enzymes in conferring hypoxia tolerance during acute hypobaric hypoxia. The expression of these antioxidant enzymes could be used as putative biomarkers for screening altitude adaptation as well as aiding in better management of altered oxygen pathophysiologies.
GNE myopathy is a rare neuromuscular genetic disorder characterized by early adult onset and muscle weakness due to mutation in sialic acid biosynthetic enzyme, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). More than 180 different GNE mutations are known all over the world with unclear pathomechanism. Although hyposialylation of glycoproteins is speculated to be the major cause, but cellular mechanism leading to loss of muscle mass has not yet been deciphered. Besides sialic acid biosynthesis, GNE affects other cellular functions such as cell adhesion and apoptosis. In order to understand the effect of mutant GNE protein on cellular functions, differential proteome profile of HEK293 cells overexpressing pathologically relevant recombinant mutant GNE protein (D207V and V603L) was analyzed. These cells, along with vector control and wild-type GNE-overexpressing cells, were subjected to two-dimensional gel electrophoresis coupled with mass spectrometry (MALDI-TOF/TOF MS/MS). In the study, 10 differentially expressed proteins were identified. Progenesis same spots software revealed downregulation of peroxiredoxin IV (PrdxIV), an ER-resident HO sensor that regulates neurogenesis. Significant reduction in mRNA and protein levels of PrdxIV was observed in GNE mutant cell lines compared with vector control. However, neither total reactive oxygen species was altered nor HO accumulation was observed in GNE mutant cell lines. Interestingly, ER redox state was significantly affected due to reduced normal GNE enzyme activity. Our study indicates that downregulation of PrdxIV affects ER redox state that may contribute to misfolding and aggregation of proteins in GNE myopathy.
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