Project description:Hypoxia leads to the aggregation of PD-L1 on the cell membrane without altering the overall protein expression level of PD-L1. Identify differentially expressed genes in the hypoxic transcriptome to reveal the key proteins involved in PD-L1 membrane trafficking, using samples of SCC25 cells treated with normoxia and hypoxia for 12 hours.
Project description:Lysates of Mycobacterium tuberculosis (H37Rv auxotroph mc(2)6020) grown under various conditions (normoxia, hypoxia, reactivation from hypoxia) probed the serine hydrolase probe with ActivX-desthiobiotin FP.
Project description:Transient hypoxia in pregnancy stimulates a physiological reflex response that redistributes blood flow and defends oxygen delivery to the fetal brain. The chemoreceptor reflex that is responsible for this physiological response is dependent on glutamatergic neurotransmission which, in times of vigorous activity, could produce cell death secondary to calcium uptake. We designed the present experiment to test the hypotheses that transient hypoxia produces damage of the cerebral cortex and that ketamine, an antagonist of NMDA receptors, reduces the damage. Late-gestation, chronically catheterized fetal sheep were subjected to a 30 min period of ventilatory hypoxia that decreased fetal PaO2 from 17±1 to 10±1 mm Hg, or normoxia (PaO2 17±1 mm Hg), with or without pretreatment (10 min before hypoxia/normoxia) with ketamine (3 mg/kg, iv). One day (24 h) after hypoxia/normoxia, fetal cerebral cortex was removed and mRNA extracted for transcriptomics and systems biology analysis. Hypoxia stimulated a transcriptomics response consistent with a reduction in cellular metabolism and an increase in inflammation. Ketamine pretreatment reduced both of these responses. The inflammation response modeled with transcriptomic system biology was validated by immunohistochemistry and showed increased abundance of microglia/macrophages after hypoxia in the cerebral cortical tissue that ketamine significantly reduced. We conclude that transient hypoxia produces inflammation of the fetal cerebral cortex and that ketamine, in a standard clinical dose, reduces the inflammation response. 4 groups: hypoxia, hypoxia plus ketamine, normoxia, normoxia plus ketamine. Hypoxia produced by low PO2 in maternal inspired gas for 30 min, followed by normoxia recovery for 23.5 hours. Control fetuses maintained at normoxia for 30 min, followed by another 23.5 h of normoxia. Fetal frontal cerebral cortex collected for mRNA at end of 23.5 h recovery period.
Project description:Estrogen receptor alpha plays a critical role in breast cancer and is a major target in endocrine therapy. HIF-1 alpha have been associated with ER alpha and predict a worse outcome. Recent studies indicate that histone demethylase JMJD2B is a HIF-1 alpha target. However, little is known about the biological functions of JMJD2B, especially in breast cancer. To elucidate the mechanism by which JMJD2B reguates gene expression in normoxia and hypoxia, MCF-7 breast cancer cells were depleted forJMJD2B in normoxia and hypoxia. Our results provide insight into JMJD2B regulation of gene expression in breast cancer cells in normoxia and hypoxia.
Project description:Estrogen receptor alpha plays a critical role in breast cancer and is a major target in endocrine therapy. HIF-1 alpha have been associated with ER alpha and predict a worse outcome. Recent studies indicate that histone demethylase JMJD2B is a HIF-1 alpha target. However, little is known about the biological functions of JMJD2B, especially in breast cancer. To elucidate the mechanism by which JMJD2B reguates gene expression in normoxia and hypoxia, MCF-7 breast cancer cells were depleted forJMJD2B in normoxia and hypoxia. Our results provide insight into JMJD2B regulation of gene expression in breast cancer cells in normoxia and hypoxia. MCF7 cells were subjected to transfection with siRNA controls and two different siRNA oligos against JMJD2B for 24 hours. Cells were treated in normoxia and hypoxia for another 16 hours.
Project description:Extracellular vesicles (EVs) represent a diverse group of small membrane-encapsulated particles and provide a universal molecular cell-cell communication system. Hypoxia is a typical condition in solid tumors, and cancer-derived EVs support growth and invasion of tissues by tumor cells. EVs were purified from cell culture medium by ultracentrifugation followed by size exclusion chromatography with Exo-spin™ columns (Cell Guidance Systems Ltd). We performed proteomic analysis of five hypoxia/normoxia pairs of EV samples; each of these replicates for either hypoxia or normoxia was analyzed in duplicates. Obtained data were compared with proteomics analysis of corresponding cell culture media supernatants (SN) after EV removal with 100 000 g ultracentrifugation. We found that exposure of tumor cells to hypoxia (1% oxygen) induced EV secretion, altered EV sizes, and led to notable changes in the EV protein cargo in comparison to normoxia.
Project description:We used miRNA array to study differential expression of miRNA under hypoxia exposure in cancer cells. Cells cultured under normoxia were used as controls.
Project description:To investigate sex differences at the transcriptome level in human pulmonary microvascular endothelial cells (HPMECs) from healthy male and female donors basally (in normoxia) and in hypoxic conditions. RNA-seq was performed on male (n=3) and female (n=4) HPMECs that were cultured in conditions of physiological shear stress (PMID: 36730645) in normoxia (21% O2) or in hypoxia (1% O2) for either 24 or 48 hours.
Project description:Hypoxia inducible factors (HIF), specifically, HIF1A, is continuously synthesized and degraded rapidly in normoxia. During hypoxia, HIF1A stabilization limits oxygen utilization and inhibits protein synthesis, but there are limited data on HIF1A function(s) in normoxia. Given the high energy needs of contraction and maintenance of the large protein mass in skeletal muscle, we determined HIF1A function in normoxia in differentiated murine myotubes with loss/gain of function and skeletal muscle from mice with Hif1afl/f or post-natal muscle-specific deletion (Hif1amsd). Integration of transcriptomics and proteomics in myotubes and muscle from mice with Hif1amsd showed enrichment of mitochondrial/metabolic regulatory molecules including TCA cycle and electron transport chain components. Mitochondrial oxidative functions and ATP content were higher with less free radical generation with Hif1a deletion. Untargeted metabolomics showed enrichment in TCA cycle components and senescence regulation. Targeted metabolomics showed higher concentrations of most TCA cycle intermediates and expression of sirtuin 3, with less abundance of markers of post-mitotic senescence. Under normoxia, regulation of mitochondrial oxidation and post-mitotic senescence in skeletal muscle by the transient expression of HIF1A may have relevance in other tissues also.
Project description:Hypoxia inducible factors (HIF), specifically, HIF1A, is continuously synthesized and degraded rapidly in normoxia. During hypoxia, HIF1A stabilization limits oxygen utilization and inhibits protein synthesis, but there are limited data on HIF1A function(s) in normoxia. Given the high energy needs of contraction and maintenance of the large protein mass in skeletal muscle, we determined HIF1A function in normoxia in differentiated murine myotubes with loss/gain of function and skeletal muscle from mice with Hif1afl/f or post-natal muscle-specific deletion (Hif1amsd). Integration of transcriptomics and proteomics in myotubes and muscle from mice with Hif1amsd showed enrichment of mitochondrial/metabolic regulatory molecules including TCA cycle and electron transport chain components. Mitochondrial oxidative functions and ATP content were higher with less free radical generation with Hif1a deletion. Untargeted metabolomics showed enrichment in TCA cycle components and senescence regulation. Targeted metabolomics showed higher concentrations of most TCA cycle intermediates and expression of sirtuin 3, with less abundance of markers of post-mitotic senescence. Under normoxia, regulation of mitochondrial oxidation and post-mitotic senescence in skeletal muscle by the transient expression of HIF1A may have relevance in other tissues also.