Introduction
Low pO2 conditions, also termed environmental hypoxia, are expressly encountered as one ascends to altitude. In a previous study, we delineated the effects of varying levels of pO2 during acute exposure of 24 h on the lung and plasma proteomes. We observed that the lung’s redox homeostasis, cytoskeletal integrity and energy homeostasis was greatly perturbed at very high altitude zone (simulated) when the ascent rate was very high. In addition, we also observed global proteome responding to these perturbations and affirming systemic redox and energy homeostasis (Paul, Gangwar, Bhargava, & Ahmad, 2018). However, the effects of extended exposure to low pO2 conditions at varying time points remain unknown. Hence, we needed to investigate the effects of increasing the duration of low pO2. We chose the lowest survivable pO2 (8.19 kPa) corresponding to PB 40 kPa which is the atmospheric pressure encountered at 25,000 ft (7620 m; beginning of death zone altitude zone).
In this study, we focus on the effects of increasing durations of low pO2 exposure (8.19 kPa) on the lung and plasma proteomes. Previous studies regarding extended low pO2(environmental hypoxia) exposure have elucidated multiple facets of exposure to chronic environmental hypoxia at high altitude. A pioneering human study on arterial pH and hemoglobin affinity during extended environmental hypoxia exposure was conducted by Chiodi in 1957 (Chiodi, 1957). Mazzeo and co-workers have reported a higher arterial norepinephrine and epinephrine concentration after 21 days at Pikes Peak (~PB 61 kPa; pO2 12.81 kPa) in human subjects (Mazzeo et al., 1991). Other studies on endocrine system during extended low pO2 exposure report a suppressive effect of endocrine mediators to facilitate maximal energy efficiency even at the cost of better oxygen delivery (Barnholt et al., 2006; Chaiban, Bitar, & Azar, 2008). Weight loss is a highly studied phenomenon during chronic low pO2 exposure and is attributed in part to hormones like leptin (Boyer & Blume, 1984; Butterfield et al., 1992; Hamad & Travis, 2006; Surks, Chinn, & Matoush, 1966; Tschöp & Morrison, 2001; Tschöp, Strasburger, Hartmann, Biollaz, & Bärtsch, 1998). Physiological features like heartbeat interval distributions were reported to be affected by chronic environmental hypoxia (Meyer et al., 1998). Ostadal and Kolar have extensively reviewed the cardiac adaptations during extended low pO2 exposures, particularly the underlying molecular mechanisms (Ostadal & Kolar, 2007). An exquisite review focused on cardiac metabolic perturbations during extended exposure to low pO2 was authored by Essop (Essop, 2007). Even in the early 80s, researchers had implicated energy efficiency with survival during chronic low pO2 exposures owing to increased energy expenditure and reduced caloric intake (Durand, 1982). A greater emphasis was laid on the muscle tissue due to their energy intensive nature during extended exposure to low pO2. Cerretelli and Hoppeler provide an early review of metabolic consequences of extended environmental hypoxia exposure on the muscle (Cerretelli & Hoppeler, 1996). Early reports stated changes in muscle capillary geometry during chronic environmental hypoxia in rats (Poole & Mathieu-Costello, 1989). The same authors also elucidate the effects of chronic environmental hypoxia on deer mouse muscle fiber size (Mathieu-Costello, Poole, & Logemann, 1989). Human muscle studies revealed a lower mitochondrial density and fiber size with unchanged capillary network during mountaineering expeditions which involved low pO2 exposures for extended time periods (Hoppeler et al., 1990). Similar studies revealed an overall shift to anaerobic metabolism in muscles (Howald et al., 1990). Muscle metabolism selectively utilizing free fatty acids instead of glycogen reserves after extended exposure to low pO2 was reported by Young and colleagues (Young, Evans, Cymerman, Pandolf, & Knapik, 1981). The influence of exercise on muscle during high altitude ascent and extended environmental hypoxia exposure has also been discussed by Kayser and co-workers in context of muscle fatigue where they showed that the nervous system had a bigger role to play rather than metabolic intermediates like lactate in causing exhaustion (Kayser, Narici, Binzoni, Grassi, & Cerretelli, 1994). A more recent study focused on the muscle mitochondrial processes during low pO2exposure (pO2 9.87 kPa) of up to 66 days and stated that muscle energy utilization and oxidative stress were modulated towards efficient ATP generation (Levett et al., 2011). Radak et al have provided a detailed review of oxidative stress during environmental hypoxia (Dosek, Ohno, Acs, Taylor, & Radak, 2007) and linked DNA repair with fatty acid metabolism in human skeletal muscle exposed to environmental hypoxia (Acs et al., 2014). Another study during Caudwell Xtreme Everest expedition concluded that chronic environmental hypoxia exposure caused significant muscle atrophy, however, muscle function remained constant (Edwards et al., 2010).
Lung (Pulmonary) system has also been studied during extended exposures to low pO2. Few of the earliest publications detail carotid body insensitivity during extended low pO2exposure in man (Severinghaus, Bainton, & Carcelen, 1966; Weil, Byrne-Quinn, Sodal, Filley, & Grover, 1971). In 1975, Tucker and co-workers implicated lung vascular smooth muscle as determinant of pulmonary hypertension, common during extended low pO2exposures (Tucker et al., 1975). Later on, mechanical properties of lung as an organ were elucidated (Weil, 1986). Decreased pressor response and increased mast cell density have been reported in rats exposed to longer durations of low pO2 (McMurtry, Petrun, & Reeves, 1978; Tucker, McMurtry, Alexander, Reeves, & Grover, 1977). Pulmonary hypertension during low pO2 exposure at high altitude has been reported in numerous studies and said to be a causative factor of HAPE and CMS (Aldashev et al., 2002; Antezana et al., 1998; Ge & Helun, 2001; Maggiorini & Leon-Velarde, 2003; Naeije, De Backer, Vachiery, & De Vuyst, 1996; Rabinovitch, Gamble, Miettinen, & Reid, 1981; Scherrer et al., 1999; Vender, 1994; Yagi, Yamada, Kobayashi, & Sekiguchi, 1990). Molecular aspects of the hypoxic lung have also been investigated. HO-1, VEGF and Rho kinase based signaling events have been implicated in chronic hypoxic lung (Gao et al., 2007; Tuder, Flook, & Voelkel, 1995). However, rat lung proteome analysis focused on extended exposure to low pO2 are rare (Ohata et al., 2013). Only recently, studies on the effects of acute low pO2exposure on rat lung proteome have been reported (Ahmad et al., 2015). Thus, this study focusing on extended low pO2 exposure induced lung proteome alterations in rat was explorative in nature. Furthermore, we also investigated changes in the plasma proteome of same animals so as to find the interlinks between the two proteomes. This was done to further facilitate their translational application during later investigations. This study elucidated many crucial aspects of lung and plasma proteome during extended exposure to low pO2. One of the key findings was that unlike redox homeostasis being a central process during acute exposure (Paul et al., 2018), extended exposure caused cytoskeletal re-arrangements to dominate in lungs while plasma proteome was dominated by proteins involved in metabolic and immune processes. An overall view that emerges from this study is the cytoskeletal remodeling that occurs in the lung signals the systemic response manifested in energy metabolism and inflammation signaling processes. A common low pO2 mediator between lung and plasma is STAT3 (Signal transducer and activator of transcription 3) (Paul et al., 2018). In an attempt to gather qualitative insights in humans undergoing extended exposure to low pO2conditions, we performed ELISA based validations of Alpha-1-antitrypsin, cofilin-1 and S100A8 in humans exposed to 14,000 ft (~pO2 12.81 kPa) for 7, 30 and 120 days. The selection of these proteins was from the protein datasets of rats, with the focus being that each of the proteins represent the processes found perturbed in rat, i.e. lung cytoskeletal re-modeling, and its derived innate immune/inflammatory signaling.
Overall, we observed that extended exposure to low pO2perturbed lipid metabolism due to redox stress in pursuit of redox homeostasis. The disturbed lipid metabolism led to inflammatory processes being activated. STAT3 played an important upstream regulatory role which may be specifically evaluated in future investigations.