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variability in hypoxia sensing and response pathways in Prakriti…

- - - - Variability towards hypoxia response at cellular level in different pathological conditions is widely reported
        
        But as basal level in healthy individuals we do not know the actual state
        
        What if we have basal level differences among different healthy individuals
        
        Q1 - How this variation going to affect the hypoxia response in healthy individuals or in disease conditions ?
        Q2 - How we are going to study hypoxia response in healthy individuals ?
  - - - Like cancerous stable cell lines
    - - Prakriti system of Ayurveda provides us a system to classify individuals on the basis of different Physiological, behavioural, and Anatomical features
        
        We can harness the existing knowledge of Ayurvedic system of classification to address our question
        
        Cellular model (Lymphoblastoid cell lines )of extreme Prakriti individuals from their PBMCs allow us to look inter-individual variability in healthy individuals
  - - - Prakriti component and its role in cellular machinery is already known
      - But when we observe hypoxia response without Prakriti or any other stratification method, we might loose several molecular players which has important role in its sensing and response pathways
  - - - EGLN1-HIF1a axis was observed in Prakriti specific manner (Kapha-Pitta) towards hypoxia responsiveness
        
        rs480902 overrepresented in Pitta and Natives of high altitude
        
        rs479200 overrepresented in Kapha and HAPE patients
      - Genetic link between EGLN1 and VWF in a Prakriti specific manner which could modulate the thrombosis / bleeding susceptibility and outcomes of hypoxia
    - - We are trying to find molecular correlates to study inter-individual variability with response to hypoxia sensing and its response pathways
- - - - MTT / SRB assay
    - - PHD2, HIF-1a, BNIP3, BNIP3L, and LC3
  - - - Affymetrix HTA 2.0
  - - - Toppfun
      - DAVID
  - - - CRISPR Cas9
      - Si RNA
- - - - Chamber mediated
        
        Chronic Hypoxia
        
        0.2% Oxygen concentration
        
        Selected for hypoxia generation and gene expression analysis
      - Chemically induced
        
        Chronic Hypoxia
        
        50 to 600 micromolar conc. of CoCl2 and DFO
        
        Act as a surrogate for hypoxia stress in normoxic conditions
    - - Selective dose of hypoxia given at different time points
        
        0 to 72 hrs
        
        24 and 48 hrs of hypoxia treatment is selected for gene expression profiling
        
        We observed maximum difference in known hypoxia markers after 24 hr of hypoxia exposure
        
        Massive cell death due to cellular acidity and nutrient deprivation observed after 48 hrs of Hypoxia stress
  - - - Compare Hypoxia treated samples in Prakriti specific manner
        
        Classify deferentially expressed genes into groups
        
        Common in any two Prakriti types
        
        Share common but some alternative approach to initiate hypoxia response
        
        Glycolysis and carbohydrate mechanism activated in Pitta kapha Prakriti types
        
        Exclusively expressed
        
        Specific to Prakriti types, provide insight at cellular levels to understand interindividual variability
        
        Common in all groups
        
        Core hypoxia responsive genes, essential for hypoxia mediated survival response
        
        HIF-1 and HIF-2 transcription factor network
        
        Promotes cellular response to hypoxia
        
        Activate kinase activity to enhance cellular signalling to promote hypoxia response
        
        Initiate Oxidoreductase and other mitochondrial mediated response to minimise the oxygen consumption and promotes hypoxia responsive adaptive machinery
        
        Among each Prakriti types
        
        Upregulated genes
        
        Common in 24 and 48 both
        
        Candidate genes which remains activated from initial to prolonged hypoxia
        
        Exclusive in 24 or 48
        
        24 hrs DEG provide hypoxia mediated early survival response
        
        48 hrs DEG provide late hypoxia responsive genes
        
        Downregulated genes
        
        Common in 24 and 48 both
        
        Genes which required Oxygen for its activation and regulation
        
        Energy consuming processes
        
        TCA cycle
        
        Ribosome biogenesis
        
        Global reduction of Translation initiation and protein folding
        
        RNA binding / ATPase activity / Nucleotide synthesis
        
        Exclusive in 24 or 48
        
        Oxidoreductase activity
        
        Helicase activity
      - Compare all hypoxia treated samples with control samples without Prakriti labelling
        
        To know the core hypoxia response with known hypoxia markers
        
        HIF-1 /HIF-2 Transcription factor network
  - - - Related to COVID19