Analysis of the Hypoxic Response in a Mouse Cortical Collecting Duct-Derived Cell Line Suggests That Esrra Is Partially Involved in Hif1α-Mediated Hypoxia-Inducible Gene Expression in mCCDcl1 Cells

Abstract

The kidney is strongly dependent on a continuous oxygen supply, and is conversely highly sensitive to hypoxia. Controlled oxygen gradients are essential for renal control of solutes and urine-concentrating mechanisms, which also depend on various hormones including aldosterone. The cortical collecting duct (CCD) is part of the aldosterone-sensitive distal nephron and possesses a key function in fine-tuned distal salt handling. It is well known that aldosterone is consistently decreased upon hypoxia. Furthermore, a recent study reported a hypoxia-dependent down-regulation of sodium currents within CCD cells. We thus investigated the possibility that cells from the cortical collecting duct are responsive to hypoxia, using the mouse cortical collecting duct cell line mCCDcl1 as a model. By analyzing the hypoxia-dependent transcriptome of mCCDcl1 cells, we found a large number of differentially-expressed genes (3086 in total logFC< −1 or >1) following 24 h of hypoxic conditions (0.2% O2). A gene ontology analysis of the differentially-regulated pathways revealed a strong decrease in oxygen-linked processes such as ATP metabolic functions, oxidative phosphorylation, and cellular and aerobic respiration, while pathways associated with hypoxic responses were robustly increased. The most pronounced regulated genes were confirmed by RT-qPCR. The low expression levels of Epas1 under both normoxic and hypoxic conditions suggest that Hif-1α, rather than Hif-2α, mediates the hypoxic response in mCCDcl1 cells. Accordingly, we generated shRNA-mediated Hif-1α knockdown cells and found Hif-1α to be responsible for the hypoxic induction of established hypoxically-induced genes. Interestingly, we could show that following shRNA-mediated knockdown of Esrra, Hif-1α protein levels were unaffected, but the gene expression levels of Egln3 and Serpine1 were significantly reduced, indicating that Esrra might contribute to the hypoxia-mediated expression of these and possibly other genes. Collectively, mCCDcl1 cells display a broad response to hypoxia and represent an adequate cellular model to study additional factors regulating the response to hypoxia.

Cascade amplification of tumor chemodynamic therapy and starvation with re-educated TAMs via Fe-MOF based functional nanosystem

“…For another thing, the hypoxia suppression induced by HFNP@GOX@PFC nanosystem was simultaneously investigated. Since the hypoxia level of tumor cells is positively associated with the expression of the typical hypoxia HIF-1α, CD47 and Glut-1 proteins [ 8 , 23 ], its expression was thus monitored for evaluation the hypoxia suppression in tumor cells using Western blot and Image J software. After treatment of the above groups for 24 h, there was no obvious difference in the expression of HIF-1α, Glut-1 and CD47 proteins of 4T1 cells between control (PBS) and HFNP group (Fig.…”

The Role of the Estrogen-Related Receptor Alpha (ERRa) in Hypoxia and Its Implications for Cancer Metabolism

“…Organs with high energy demand such as the brain, heart and kidney have low tolerance to hypoxia and are good models to evaluate ERRα's effects. Physiological oxygen gradients across the renal cortex and medulla participate in the mechanisms to concentrate urine [88,89]. The healthy human kidney cortex presents around 50 mmHg of oxygen pressure, while the medulla has much lower oxygen pressures between 10 and 20 mmHg [88].…”

“…The healthy human kidney cortex presents around 50 mmHg of oxygen pressure, while the medulla has much lower oxygen pressures between 10 and 20 mmHg [88]. Keppner et al recently evaluated the transcriptome during hypoxia (24 h at 0.2% O 2 ) of the cortical kidney murine cell line mCCD(cl1) [89]. They found over 3000 differentially expressed genes, many related to aerobic metabolism and ATP production through mitochondria, and the hypoxia response was mainly driven by HIF-1 and not HIF-2.…”

“…They found over 3000 differentially expressed genes, many related to aerobic metabolism and ATP production through mitochondria, and the hypoxia response was mainly driven by HIF-1 and not HIF-2. Interestingly, they knocked down ERRα and identified a reduced expression of some genes that typically function in hypoxia, such as EGLN3 (an alpha-ketoglutarate dependent hydroxylase that controls cell proliferation and transcription upon hypoxia) and SERPIN1 (plasminogen activator inhibitor-1, involved in the control of blood clotting) [89]. Since this regulation happened without a change in HIF-1α, the model suggests that ERRα controls the expression of specific genes important for the hypoxia response.…”