A method for assessing an effect of hypoxia on a tissue includes providing a sample of the tissue in a hermetically sealed container, determining a first amount of a reaction substrate (e.g., protocatechuic acid) to be introduced into the sealed container and determining a second amount of a reaction enzyme (e.g., protocatechuate dioxygenase) to be introduced into the sealed container. The method further includes introducing the reaction substrate and the reaction enzyme into the sealed container. At least one of the first amount of the reaction substrate and the second amount of the reaction enzyme is selected to induce at least one of a predetermined amount of hypoxia less than anoxia and a predetermined rate of hypoxia in the tissue during a reaction between the reaction substrate and the reaction enzyme. Values of properties of the tissue can be measured before and after the reaction to assess effects of hypoxia.

The present invention provides biomarkers for sensitive, specific, accurate and quantitative diagnosis and assessment of chronic hypoxia. In particular, the present invention provides 2-hydroxyglutarate as a biomarker that is differentially produced in chronic hypoxia. Furthermore, embodiments of the invention are able to differentiate between chronic and acute hypoxia. Assays for levels of 2-hydroxyglutarate may be used alone or in conjunction with additional biomarkers of hypoxia to increase the precision of analysis. In particular embodiments of the invention, the level of 2-hydroxyglutarate and at least one second biomarker may be assayed to generate a hypoxic profile that can be compared to a reference or control profile, thereby diagnosing a subject as normoxic, chronically hypoxic, or acutely hypoxic.

Hypoxia induced mitogenic factor (HIMF) is a member of the “found in inflammatory zone” (FIZZ)/resistin family of proteins and has potent mitogenic, angiogenic, and vasoconstrictive effects in the lung vasculature. The receptor/binding partners for this family of proteins have been largely unknown. We identified Bruton's tyrosine kinase (BTK) as a functional HIMF binding partner through GST-HIMF pull-downs and mass spectrometry. Using primary cultured HIMF-stimulated murine bone marrow cells, we demonstrated that BTK was recruited to the leading edge of the cells. We also demonstrated that BTK and the closely related tyrosine kinase Fyn, colocalized at the growth cone process in these cells. HIMF stimulation induced BTK autophosphorylation, which peaked at 2.5 minutes. A transwell migration assay showed that treatment with recombinant murine HIMF induced migration of primary cultured bone marrow cells, which was completely blocked by the BTK inhibitor, LFM-A13. In vivo studies, using the rat hindlimb ischemia model, revealed that HIMF can stimulate angiogenesis in the hypoxic tissue probably through inducing the migration of endothelial progenitor cells (EPCs) to areas of active angiogenesis. Our results indicate that HIMF may acts as a chemotactic molecule in stimulating the migration of leukocytes/EPCs from bone marrow to targeted tissues through activation of the BTK pathway.

The instant disclosure describes a method for measuring and reporting vascularity in a biological tissue sample. The method generally includes: within a digital image of a tissue section, (i) identifying endothelial cells, lymphatic cells, or a combination thereof; (ii) mapping one or more proximity regions, each of the proximity regions defining an area between detected vessels and a first distance outwardly therefrom; and (iii) calculating one or more of: a vessel proximity score or a hypoxia score, wherein the vessel proximity score relates a composition of objects within the proximity regions, and wherein the hypoxia score relates a composition of tissue within or outside of the proximity regions, respectively.

A method of generating human mature oligodendrocytes is disclosed. The method comprises contacting a cell population which comprises human pre-oligodendrocytes with an inhibitor of the MAPK/ERK pathway under conditions that allow the pre-oligodendrocytes to differentiate into mature oligodendrocytes. Use of the MAPK/ERK pathway inhibitor for treating diseases is also disclosed.

A method for assessing an effect of hypoxia on a tissue includes providing a sample of the tissue in a hermetically sealed container, determining a first amount of a reaction substrate (e.g., protocatechuic acid) to be introduced into the sealed container and determining a second amount of a reaction enzyme (e.g., protocatechuate dioxygenase) to be introduced into the sealed container. The method further includes introducing the reaction substrate and the reaction enzyme into the sealed container. At least one of the first amount of the reaction substrate and the second amount of the reaction enzyme is selected to induce at least one of a predetermined amount of hypoxia less than anoxia and a predetermined rate of hypoxia in the tissue during a reaction between the reaction substrate and the reaction enzyme. Values of properties of the tissue can be measured before and after the reaction to assess effects of hypoxia.

Systems, devices, and methods to measure interstitial concentration of selected compounds over time to provide early detection of shock are described. In an example method, interstitial fluid is obtained from a subject. Analytes are detected in the interstitial fluid. At least one of the analytes corresponds to hypoxia of a tissue of the subject and at least one other analyte corresponds to vascular permeability of the subject. The example method further includes determining whether the subject is in shock based on a computing model and the detected analytes and generating an alert based on whether the subject is in shock. Treatments can be administered following the alert generation.

Detection of under-expression of selected genes in NK cells is used to confirm a hypoxic tumor microenvironment that is ordinarily suppressive with respect to ADCC and cytotoxic cell killing of NK cells. Most notably, while hypoxia is known to upregulate HIF-1α and genes under the control of HIF-1α, hypoxia in a tumor microenvironment led to under-expression of selected genes, including HIF-1α. Thus, gene expression analysis of certain genes on NK cells can be used to detect conditions in a tumor that would indicate use of haNK cells.

The present disclosure provides, among other things, methods and compositions for diagnosing and treating stress-induced injuries, hypoxia in particular. The present invention is based, in part, on the novel discovery that a metabolite, L-2-hydroxyglutarate, and certain enzymes and substrates regulating its metabolism, mediate stress-induced cellular mechanisms. In some embodiments, provided methods and compositions are used to diagnose and treat diseases with hypoxia-induced injuries. In some embodiments, provided methods and compositions are used to modulate cell pluripotency or differentiation in vivo or in vitro.

The invention provides peptides capable of inhibiting the individual activity and inter-pathway communication of BNIP3, BAX and mitochondria. The peptides can be used in methods of treating a disease or condition in a subject in which it is desirable to prevent cell damage and cell death.