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Hypoxia is an important feature of multiple diseases like cancer and obesity and also an environmental stressor to high altitude travelers. Emerging research suggests the importance of redox signaling in physiological responses transforming the notion of oxidative stress into eustress and distress. However, the behavior of redox protein post-translational modifications (PTMs), and their correlation with stress acclimatization in humans remains sketchy. Scant information exists about modifications in redoxome during physiological exposure to environmental hypoxia. In this study, we investigated redox PTMs, nitrosylation and carbonylation, in context of extended environmental hypoxia exposure.The volunteers were confirmed to be free of any medical conditions and matched for age and weight. The human global redoxome and the affected networks were investigated using TMT-labeled quantitative proteo-bioinformatics and biochemical assays. The percolator PSM algorithm was used for peptide-spectrum match (PSM) validation in database searches. The FDR for peptide matches was set to 0.01. 1-way ANOVA and Tukey's Multiple Comparison test were used for biochemical assays. p-value0.05 was considered statistically significant. Three independent experiments (biological replicates) were performed. Results were presented as Mean ± standard error of mean (SEM).This investigation revealed direct and indirect interplay between nitrosylation and carbonylation especially within coagulation and inflammation networks; interlinked redox signaling (via nitrosylation‑carbonylation); and novel nitrosylation and carbonylation sites in individual proteins.This study elucidates the role of redox PTMs in hypoxia signaling favoring tolerance and survival. Also, we demonstrated direct and indirect interplay between nitrosylation and carbonylation is crucial to extended hypoxia tolerance.

Exposure to high altitude above 3000 m leads to two outcomes-acclimation or high-altitude maladies. To reach a particular outcome, the plasma proteome is modified differentially, either in context of an acclimation response or mal-acclimation response leading to disease. This ensures that hypoxia-responsive plasma protein trends reflect acclimation in acclimated individuals when compared with their levels prior to acclimation. Such protein trends could be used to assess acclimation in an individual and any significant deviation from this trend may indicate non-acclimation, thereby preventing high-altitude illnesses before they manifest. In this study, we investigate and statistically evaluate the trendlines of various hypoxia-responsive plasma protein levels, reported significantly perturbed in our previous studies, in individuals (male; n = 20) exposed to 3520 m at high-altitude day 1 (HAD1), HAD4, and HAD7L and to 4420 m at HAD7H, HAD30, and HAD120. We observe that thioredoxin (Trx), glutathione peroxidase 3 (GPx-3), and apolipoprotein AI (Apo-AI) are statistically robust markers to assess acclimation across the exposure duration while sulfotransferase 1A1 (ST1A1) is a capable negative control whose levels increase only in cases of HAPE. We also observe exposure day-specific and resident altitude-specific proteins capable of accurately assessing acclimation when compared with baseline levels or the lower altitude zone.

Background: Hypoxia is a pathophysiological condition which arises due to low oxygen concentration in conditions like cardiovascular diseases, inflammation, ascent to higher altitude, malignancies, deep sea diving, prenatal birth, etc. A number of microRNAs (miRNAs), Transcription Factors (TFs) and genes have been studied separately for their role in hypoxic adaptation and controlling cell-cycle progression and apoptosis during this stress. Objective: We hypothesize that miRNAs and TFs may act in conjunction to regulate a multitude of genes and play a crucial and combinatorial role during hypoxia-stress-responses and associated cellcycle control mechanisms. Method: We collected a comprehensive and non-redundant list of human hypoxia-responsive miRNAs (also known as hypoxiamiRs). Their experimentally validated gene-targets were retrieved from various databases and a comprehensive hypoxiamiR-gene regulatory network was built. Results: Functional characterization and pathway enrichment of genes identified phospho-proteins as enriched nodes. The phospho-proteins which were localized both in the nucleus and cytoplasm and could potentially play important role as signaling molecules were selected; and further pathway enrichment revealed that most of them were involved in NFkB signaling. Topological analysis identified several critical hypoxiamiRs and network perturbations confirmed their importance in the network. Feed Forward Loops (FFLs) were identified in the subnetwork of enriched genes, miRNAs and TFs. Statistically significant FFLs consisted of four miRNAs (hsa-miR-182-5p, hsa- miR-146b-5p, hsa-miR-96, hsa-miR-20a) and three TFs (SMAD4, FOXO1, HIF1A) both regulating two genes (NFkB1A and CDKN1A). Conclusion: Detailed BioCarta pathway analysis identified that these miRNAs and TFs together play a critical and combinatorial role in regulating cell-cycle under hypoxia, by controlling mechanisms that activate cell-cycle checkpoint protein, CDKN1A. These modules work synergistically to regulate cell-proliferation, cell-growth, cell-differentiation and apoptosis during hypoxia. A detailed mechanistic molecular model of how these co-regulatory FFLs may regulate the cell-cycle transitions during hypoxic stress conditions is also put forth. These biomolecules may play a crucial and deterministic role in deciding the fate of the cell under hypoxic-stress.

Objective This study explores whether the prognosis of interstitial lung disease in rheumatoid arthritis (RA-ILD) has improved over time and assesses the potential influence of drug therapy in a large multicentre UK network. Methods We analysed data from 18 UK centres on patients meeting criteria for both RA and ILD diagnosed over a 25-year period. Data included age, disease duration, outcome and cause of death. We compared all cause and respiratory mortality between RA controls and RA-ILD patients, assessing the influence of specific drugs on mortality in four quartiles based on year of diagnosis. Results A total of 290 RA-ILD patients were identified. All cause (respiratory) mortality was increased at 30% (18%) compared with controls 21% (7%) (P =0.02). Overall, prognosis improved over quartiles with median age at death rising from 63 years to 78 years (P =0.01). No effect on mortality was detected as a result of DMARD use in RA-ILD. Relative risk (RR) of death from any cause was increased among patients who had received anti-TNF therapy [2.09 (1.1–4.0)] P =0.03, while RR was lower in those treated with rituximab [0.52(0.1–2.1)] or mycophenolate [0.65 (0.2–2.0)]. Patients receiving rituximab as their first biologic had longer three (92%), five (82%) and seven year (80%) survival than those whose first biologic was an anti-TNF agent (82%, 76% and 64%, respectively) (P =0.037). Discussion This large retrospective multicentre study demonstrates survival of patients with RA-ILD has improved. This may relate to the increasing use of specific immunosuppressive and biologic agents.

Lack of zero side-effect, prescription-less prophylactics and diagnostic markers of acclimatization status lead to many suffering from high altitude illnesses. Although not fully translated to the clinical setting, many strategies and interventions are being developed that are aimed at providing an objective and tangible answer regarding the acclimatization status of an individual as well as zero side-effect prophylaxis that is cost-effective and does not require medical supervision. This short review brings together the twin problems associated with high-altitude acclimatization, i.e. acclimatization status and zero side-effect, easy-to-use prophylaxis, for the reader to comprehend as cogs of the same phenomenon. We describe current research aimed at preventing all the high-altitude illnesses by considering them an assault on redox and energy homeostasis at the molecular level. This review also entails some proteins capable of diagnosing either acclimatization or high-altitude illnesses. The future strategies based on bioinformatics and systems biology is also discussed.

Hypobaric hypoxia elicits several patho-physiological manifestations, some of which are known to be lethal. Among various molecular mechanisms proposed so far, perturbation in redox state due to imbalance between radical generation and antioxidant defence is promising. These molecular events are also related to hypoxic status of cancer cells and therefore its understanding has extended clinical advantage beyond high altitude hypoxia. In present study, however, the focus was to understand and propose a model for rapid acclimatization of high altitude visitors to enhance their performance based on molecular changes. We considered using simulated hypobaric hypoxia at some established thresholds of high altitude stratification based on known physiological effects. Previous studies have focused on the temporal aspect while overlooking the effects of varying pO2 levels during exposure to hypobaric hypoxia. The pO2 levels, indicative of altitude, are crucial to redox homeostasis and can be the limiting factor during acclimatization to hypobaric hypoxia. In this study we present the effects of acute (24 h) exposure to high (3049 m; pO2: 71 kPa), very high (4573 m; pO2: 59 kPa) and extreme altitude (7620 m; pO2: 40 kPa) zones on lung and plasma using semi-quantitative redox specific transcripts and quantitative proteo-bioinformatics workflow in conjunction with redox stress assays. It was observed that direct exposure to extreme altitude caused 100% mortality, which turned into high survival rate after pre-exposure to 59 kPa, for which molecular explanation were also found. The pO2 of 59 kPa (very high altitude zone) elicits systemic energy and redox homeostatic processes by modulating the STAT3-RXR-Nrf2 trio. Finally we posit the various processes downstream of STAT3-RXR-Nrf2 and the plasma proteins that can be used to ascertain the redox status of an individual., Graphical abstract fx1, Highlights • A comparison of effects on redox and energy homeostasis between three altitude zones. • Exposure at 59 kPa (10 h) followed by normoxia (1 h) rescues homeostasis at 40 kPa. • Redox and energy homeostasis dependent on STAT-3-RXR-Nrf interplay. • Lung redox and energy homeostasis fails at 15,000 ft. • Systemic redox and energy homeostasis activated on failure of lung homeostatic mechanisms.

Intermittent hypoxia, initially associated with adverse effects of sleep apnea, has now metamorphosed into a module for improved sports performance. The regimen followed for improved sports performance is milder intermittent hypoxic training (IHT) as compared to chronic and severe intermittent hypoxia observed in sleep apnea. Although several studies have indicated the mechanism and enough data on physiological parameters altered by IH is available, proteome perturbations remain largely unknown. Altitude induced hypobaric hypoxia is known to require acclimatization as it causes systemic redox stress and inflammation in humans. In the present study, a short IHT regimen consisting of previously reported physiologically beneficial FIO2 levels of 13.5% and 12% was administered to human subjects. These subjects were then airlifted to altitude of 3500 m and their plasma proteome along with associated redox parameters were analyzed on days 4 and 7 of high altitude stay. We observed that redox stress and associated post-translational modifications, perturbed lipid metabolism and inflammatory signaling were induced by IHT exposure at Baseline. However, this caused activation of antioxidants, energy homeostasis mechanisms and anti-inflammatory responses during subsequent high-altitude exposure. Thus, we propose IHT as a beneficial non-pharmacological intervention that benefits individuals venturing to high altitude areas.

Aims HAPE remains the most common lethal high-altitude disease. Although its pathophysiology and other associated causal factors have been partially uncovered along with some potential biomarker proteins, it has not been completely elucidated. A major hindrance to improving the understanding of HAPE pathophysiology and associated molecular events has been the absence of a quick, reliable and definitive animal model of HAPE. This study is aimed at development of a rapid and reliable SD rat model of high altitude pulmonary edema (HAPE) that can be roentgenographically confirmed and be used to study protein markers of HAPE. Main methods In this study, we detail the process of rapidly inducing HAPE in male SD rats within 18 h of simulated high-altitude exposure without causing high rates of mortality. Thereafter, we confirmed HAPE using roentgenography. We assessed Sulfotransferase 1A1 (SULT1A1), IL-1 beta, TNF- alpha and IFN-gamma using ELISA. Finally, H&E staining of lung tissues was also performed. Key findings A roentgenographically confirmed HAPE model was demonstrated. SULT 1A1 levels are found to be highest in rats suffering HAPE, as previously confirmed in human patients. Inflammation was also assessed based on levels of inflammatory proteins like IL-1b, TNF-a, and IFN-g in addition to H&E staining of lung tissues. Inflammation and HAPE were observed to be synergistic events and not cause and effect of each other. Significance This rat model of HAPE will help researchers and clinicians in evaluating performance of therapies, potential biomarker and also further elucidate underlying molecular processes causing HAPE.

Hypobaric Hypoxia (HH) is an established risk factor for various neuro-physiological perturbations including cognitive impairment. The origin and mechanistic basis of such responses however remain elusive. We here combined systems level analysis with classical neuro-physiological approaches, in a rat model system, to understand pathological responses of brain to HH. Unbiased ‘statistical co-expression networks’ generated utilizing temporal, differential transcriptome signatures of hippocampus—centrally involved in regulating cognition—implicated perturbation of Glio-Vascular homeostasis during early responses to HH, with concurrent modulation of vasomodulatory, hemostatic and proteolytic processes. Further, multiple lines of experimental evidence from ultra-structural, immuno-histological, substrate-zymography and barrier function studies unambiguously supported this proposition. Interestingly, we show a significant lowering of H2S levels in the brain, under chronic HH conditions. This phenomenon functionally impacted hypoxia-induced modulation of cerebral blood flow (hypoxic autoregulation) besides perturbing the strength of functional hyperemia responses. The augmentation of H2S levels, during HH conditions, remarkably preserved Glio-Vascular homeostasis and key neuro-physiological functions (cerebral blood flow, functional hyperemia and spatial memory) besides curtailing HH-induced neuronal apoptosis in hippocampus. Our data thus revealed causal role of H2S during HH-induced early Glio-Vascular dysfunction and consequent cognitive impairment., Highlights • Glio-Vascular dysfunction temporally precedes Hypobaric Hypoxia (HH) induced neuro-pathological effects. • Exposure to HH significantly lowers the levels of H2S in brain. • Augmentation of H2S, utilizing its donor, preserves Glio-Vascular homeostasis and curtails HH-induced memory impairment. The exposure to Hypobaric Hypoxia (HH) environment (such as that encountered by humans at high altitude) culminates in cognitive impairment in an altitude- and duration-dependent manner. The mechanistic basis for such effects, however, remains elusive. Our present study showed that HH-induced neuro-pathological perturbations are temporally preceded by Glio-Vascular dysfunction and are concomitant with lowered levels of gaseous messenger, H2S, in brain. The maintenance of H2S levels (utilizing a specific donor, NaHS) during hypoxia curtailed HH-induced brain-vascular dysfunction and ensuing neuro-pathological effects (on spatial memory). Interestingly, identification of origin of disease in the present study effectively revealed a possible interventional strategy.

Hypoxia is a complex pathophysiological condition. The physiological and molecular responses to this stress have been extensively studied. However, the management of its ill effects still poses a challenge to clinicians. MicroRNAs (miRNAs) are short non-coding RNA molecules that control post-transcriptional gene expression. The regulatory role of miRNAs in hypoxic environments has been studied in many hypoxia-related disorders, however a comprehensive compilation and analysis of all data and the significance of miRNAs in hypoxia adaption is still lacking. This review summarizes the miRNAs related to various hypoxia-related disorders and highlights the computational approaches to study them. This would help in designing novel strategies toward efficient management of hypoxia-related disorders.

Objective Identification of molecular signatures having key roles in hypobaric hypoxia by analysing the salivary proteome. Saliva holds a promising future in the search for new clinical biomarkers that are easily accessible, less complex, accurate, and cost effective as well as being non-invasive. Methodology We employed qualitative proteomics approach to develop discriminatory biomarker signatures from human saliva exposed to hypobaric hypoxia. Salivary proteins were analyzed and compared between age-matched healthy subjects exposed to high altitude (∼13700 ft) for seven days (HAD7) with control subjects at sea level (Normoxia) by using 2-Dimensional gel electrophoresis/Mass Spectrometry approach. Results Several proteins with significant differential expression were found. The up-regulated proteins were apoptosis inducing factor-2, cystatin S, cystatin SN and carbonic anhydrase 6. The down regulated proteins were polymeric immunoglobulin receptor, alpha-enolase and prolactin-inducible protein. Further confirmation of the altered proteins such as alpha enolase, carbonic anhydrase 6, prolactin-inducible protein, apoptosis inducing factor 2, cystatin S and cystatin SN were performed using immunoblotting. The expression patterns of the selected proteins observed by immunoblot were in concurrence with 2-Dimesional gel electrophoresis results, therefore affirming the authenticity of the proteomic investigation. Conclusion This study provides the proof of concept of salivary biomarkers for the non-invasive detection of hypobaric hypoxia induced effects. It is highly feasible to turn these biomarkers into an applicable clinical test after large scale validation.

Oxygen-compromised environments, such as high altitude, air travel, and sports, and pathological conditions, such as solid tumors, have been suggested to be prothrombotic. Despite the indispensable role of platelets in thrombus formation, the studies linking hypoxia, platelet reactivity, and thrombus formation are limited. In the present study, platelet proteome/reactivity was analyzed to elucidate the acute hypoxia-induced prothrombotic phenotype. Rats exposed to acute simulated hypoxia (282 torr/8% oxygen) demonstrated a decreased bleeding propensity and increased platelet reactivity. Proteomic analysis of hypoxic platelets revealed 27 differentially expressed proteins, including those involved in coagulation. Among these proteins, calpain small subunit 1, a 28-kDa regulatory component for calpain function, was significantly upregulated under hypoxic conditions. Moreover, intraplatelet Ca(2+) level and platelet calpain activity were also found to be in accordance with calpain small subunit 1 expression. The inhibition of calpain activity demonstrated reversal of hypoxia-induced platelet hyperreactivity. The prothrombotic role for calpain was further confirmed by an in vivo model of hypoxia-induced thrombosis. Interestingly, patients who developed thrombosis while at extreme altitude had elevated plasma calpain activities and increased soluble P-selectin level. In summary, this study suggests that augmented calpain activity is associated with increased incidence of thrombosis under hypoxic environments.

Despite its extraordinary antioxidant capacity, the clinical usage of silymarin has remained restricted to amelioration of hepatic pathology. Perhaps its low bioavailability and uneven bio-distribution, owing to its poor aqueous solubility, are two main causes that have dampened the clinical applicability and scope of this preparation. We took these two challenges and suggested an unexplored application of silymarin. Apart from liver, two of the most susceptible vital organs at the highest risk of oxidative stress are brain and lung, especially during reduced oxygen saturation (hypoxia) at anatomical level. Hypoxia causes excess generation of radicals primarily in the lungs as it is the first organ at the interphase of atmosphere and organism making it the most prone and vulnerable to oxidative stress and the first responder against hypobaric hypoxia. As our first objective, we improved the silymarin formulation by restricting its size to the lower threshold and then successfully tested the prophylactic and therapeutic action in rat lung challenged with simulated hypobaric hypoxia. After dose optimization, we observed that 50mg/kg BW silymarin as size restricted and homogenous aqueous suspension successfully minimized the reactive oxygen species and augmented the antioxidant defense by significant upregulation of catalase and superoxide dismutase and reduced glutathione. Moreover, the well-established hypoxia markers and proteins related to hypoxia adaptability, hif1a and VEGF were differentially regulated conferring significant reduction in the inflammation caused by hypobaric hypoxia. We therefore report,the unexplored potential benefits of silymarin for preventing high altitude associated pathophysiology further paving its road to clinical trials.

We have investigated the plasma proteome using 2D gel electrophoresis and matrix-assisted laser desorption/ionization tandem time of flight from patients with high altitude pulmonary edema (HAPE). A complete proteomic analysis was performed on 20 patients with HAPE and ten healthy sea level controls. In total, we have identified 25 protein spots in human plasma and found that 14 of them showed altered changes in HAPE patients, which mainly were acute phase proteins (APPs), compliment components, and apolipoproteins among others. Among the APPs, haptoglobin α2 chain, haptoglobin β chain, transthyretin, and plasma retinol binding precursor showed overexpression in HAPE patients as compared to controls. To validate the result of proteomic analysis, two proteins were selected for enzyme-linked immunosorbent assay and Western blotting analysis. Our data conclusively shows that two proteins, haptoglobin and apolipoprotein A-I are upregulated in plasma of HAPE patients. These proteins may provide a fast and effective control of inflammatory damage until the subsequent mechanisms can begin to operate. Taken together, our findings further support the hypothesis that inflammatory response system is linked to the pathophysiology of HAPE.

Exposure to high altitude induces physiological responses due to hypoxia. Lungs being at the first level to face the alterations in oxygen levels are critical to counter and balance these changes. Studies have been done analysing pulmonary proteome alterations in response to exposure to hypobaric hypoxia. However, such studies have reported the alterations at specific time points and do not reflect the gradual proteomic changes. These studies also identify the various biochemical pathways and responses induced after immediate exposure and the resolution of these effects in challenge to hypobaric hypoxia. In the present study, using 2-DE/MS approach, we attempt to resolve these shortcomings by analysing the proteome alterations in lungs in response to different durations of exposure to hypobaric hypoxia. Our study thus highlights the gradual and dynamic changes in pulmonary proteome following hypobaric hypoxia. For the first time, we also report the possible consideration of SULT1A1, as a biomarker for the diagnosis of high altitude pulmonary edema (HAPE). Higher SULT1A1 levels were observed in rats as well as in humans exposed to high altitude, when compared to sea-level controls. This study can thus form the basis for identifying biomarkers for diagnostic and prognostic purposes in responses to hypobaric hypoxia.

A technique i.e., Soil Solarization and Amendments (neem, chicken farmyard manure, farmyard manure and biokhad viz synthetic bio fertilizer), towards the natural cropping system has been evaluated for its effectiveness and practicability at the National Agricultural Research Center Islamabad Pakistan. Soil solarization and amendments were analyzed as a control measure against soil born fungi and nematodes. Eight weeks of solarization resulted in about 11 degrees C increase in the soil temperature. This increase in soil temperature caused a reduction of about 70 to 80% in the fungal population and about 99% in nematode population at various depths. Neem and Biokhad amendments were proved synergistic for solarization and also improved the properties of soil in the benefit of crop plants. Fusarium sp., Macrophomina phyaseolina and Verticillium sp. of fungi and Tylenchus sp., Haplolaimus sp., Xiphenema sp. and almost all of the parasitic nematodes were significantly (p

Purpose Computational biology has opened a gateway to omics data analysis and shifted the focus from molecules to systemic molecular networks in the domain of hypobaric hypoxia (HH). Yet there are no meta-analytical investigations circumventing constraints such as organism (rat/human), HH exposure conditions (acute/chronic), and the tissues that can be investigated simultaneously in the realm of wet lab experiments. Experimental design We analyzed 154 differentially expressed proteins upon HH exposure using Ingenuity Pathway Analysis (IPA) tool, without the constraint of using a single organism or tissue type, to determine the most significant pathways and networks that are perturbed across a range of HH conditions. Results We found acute phase response signaling, farsenoid X receptor/retinoid X receptor activation, liver X receptor/retinoid X receptor activation, clathrin-mediated endocytosis signaling, mitochondrial dysfunction, production of nitric oxide and ROS in macrophages, and integrin signaling to be the most significant universally perturbed pathways. Unique protein–function relationships have also been highlighted. Conclusion and clinical relevance This meta-analysis provides a list of specific pathways and networks across two model organisms that are perturbed due to HH exposure irrespective of its duration/intensity. Thus, it will be a map of important pathways and proteins to look at when exploring effects of HH exposure irrespective of tissue/organism chosen, particularly in the context of prophylactic/therapeutic targets.

Adaptation to hypobaric hypoxia is required by animals and human in several physiological and pathological situations. Hypobaric hypoxia is a pathophysiological condition triggering redox status disturbances of cell organization leading, via oxidative stress, to proteins, lipids, and DNA damage. Identifying the molecular variables playing key roles in this process would be of paramount importance to shed light on the mechanisms known to counteract the negative effects of oxygen lack. To obtain a molecular signature, changes in the plasma proteome were studied by using proteomic approach. To enrich the low-abundance proteins in human plasma, two highly abundant proteins, albumin and IgG, were first removed. By comparing the plasma proteins of high altitude natives with those of a normal control group, several proteins with a significant alteration were found. The up-regulated proteins were identified as vitamin D-binding protein, hemopexin, alpha-1–antitrypsin, haptoglobin β-chain, apolipoprotein A1, transthyretin and hemoglobin beta chain. The down-regulated proteins were transferrin, complement C3, serum amyloid, complement component 4A and plasma retinol binding protein. Among these proteins, the alterations of transthyretin and transferrin were further confirmed by ELISA and Western blotting analysis. Since all the up- and down- regulated proteins identified above are well-known inflammation inhibitors and play a positive anti-inflammatory role, these results show that there is some adaptive mechanism that sustains the inflammation balance in high altitude natives exposed to hypobaric hypoxia.