Renal hypoxia contributes to chronic kidney disease (CKD) progression, as validated in experimental and human CKD. In the early stages, increased oxygen consumption causes oxygen demand/supply mismatch, leading to hypoxia. Hence, early targeting of the determinants and regulators of oxygen consumption in CKD may alter the disease course before permanent damage ensues. Here, we focus on hypoxia inducible factor-1α (HIF-1α) and AMP-activated protein kinase (AMPK) and on the mechanisms by which they may facilitate cellular hypoxia adaptation. We found that HIF-1α activation in the subtotal nephrectomy (STN) model of CKD limits protein synthesis, inhibits apoptosis, and activates autophagy, presumably for improved cell survival. AMPK activation was diminished in the STN kidney and was remarkably restored by HIF-1α activation, demonstrating a novel role for HIF-1α in the regulation of AMPK activity. We also investigated the independent and combined effects of HIF-1α and AMPK on cell survival and death pathways by utilizing pharmacological and knockdown approaches in cell culture models. We found that the effect of HIF-1α activation on autophagy is independent of AMPK, but on apoptosis it is partially AMPK dependent. The effects of HIF-1α and AMPK activation on inhibiting protein synthesis via the mTOR pathway appear to be additive. These various effects were also observed under hypoxic conditions. In conclusion, HIF-1α and AMPK appear to be linked at a molecular level and may act as components of a concerted cellular response to hypoxic stress in the pathophysiology of CKD.

The pathophysiology of chronic kidney disease (CKD) is driven by alterations in surviving nephrons to sustain renal function with ongoing nephron loss. Oxygen supply-demand mismatch, due to hemodynamic adaptations, with resultant hypoxia, plays an important role in the pathophysiology in early CKD. We sought to investigate the underlying mechanisms of this mismatch. We utilized the subtotal nephrectomy (STN) model of CKD to investigate the alterations in renal oxygenation linked to sodium (Na) transport and mitochondrial function in the surviving nephrons. Oxygen delivery was significantly reduced in STN kidneys because of lower renal blood flow. Fractional oxygen extraction was significantly higher in STN. Tubular Na reabsorption was significantly lower per mole of oxygen consumed in STN. We hypothesized that decreased mitochondrial bioenergetic capacity may account for this and uncovered significant mitochondrial dysfunction in the early STN kidney: higher oxidative metabolism without an attendant increase in ATP levels, elevated superoxide levels, and alterations in mitochondrial morphology. We further investigated the effect of activation of hypoxia-inducible factor-1α (HIF-1α), a master regulator of cellular hypoxia response. We observed significant improvement in renal blood flow, glomerular filtration rate, and tubular Na reabsorption per mole of oxygen consumed with HIF-1α activation. Importantly, HIF-1α activation significantly lowered mitochondrial oxygen consumption and superoxide production and increased mitochondrial volume density. In conclusion, we report significant impairment of renal oxygenation and mitochondrial function at the early stages of CKD and demonstrate the beneficial role of HIF-1α activation on renal function and metabolism.

Cytotoxic T lymphocyte antigen 4 (CTLA4) and programmed cell death protein 1 (PD-1) are immune checkpoint proteins expressed in T cells. Although CTLA4 expression was found in multiple tumours including non-small cell lung cancer (NSCLC) tissues and cells, its function in tumour cells is unknown. Recently, PD-1 was found to be expressed in melanoma cells and to promote tumorigenesis. We found that CTLA4 was expressed in a subset of NSCLC cell lines and in a subgroup of cancer cells within the lung cancer tissues. We further found that in NSCLC cells, anti-CTLA4 antibody can induce PD-L1 expression, which is mediated by CTLA4 and the EGFR pathway involving phosphorylation of MEK and ERK. In CTLA4 knockout cells, EGFR knockout cells or in the presence of an EGFR tyrosine kinase inhibitor, anti-CTLA4 antibody was not able to induce PD-L1 expression in NSCLC cells. Moreover, anti-CTLA4 antibody promoted NSCLC cell proliferation in vitro and tumour growth in vivo in the absence of adaptive immunity. These results suggest that tumour cell-intrinsic CTLA4 can regulate PD-L1 expression and cell proliferation, and that anti-CTLA4 antibody, by binding to the tumour cell-intrinsic CTLA4, may result in the activation of the EGFR pathway in cancer cells.

IL-33 and its receptor ST2 play important roles in airway inflammation and contribute to asthma onset and exacerbation. The IL-33/ST2 signaling pathway recruits adapter protein myeloid differentiation primary response 88 (MyD88) to transduce intracellular signaling. MyD88 forms a complex with IL-R-associated kinases (IRAKs), IRAK4 and IRAK2, called the Myddosome (MyD88-IRAK4-IRAK2). The myddosome subsequently activates downstream NF-κB and MAPKs p38 and JNK. We established an asthma-like mouse model by intratracheal administration of IL-33. The IL-33 model has a very similar phenotype compared with the OVA-induced mouse asthma model. The importance of MyD88 in the IL-33/ST2 signaling transduction was demonstrated by the MyD88 knockout mice, which were protected from the IL-33-induced asthma. We synthesized small molecule mimetics of the α-helical domain of IRAK2 with drug-like characteristics based on the recent advances in the designing of α-helix compounds. The mimetics can competitively interfere in the protein-protein interaction between IRAK2 and IRAK4, leading to disruption of Myddosome formation. A series of small molecules were screened using an NF-κB promoter assay in vitro. The lead compound, 7004, was further studied in the IL-33-induced and OVA-induced asthma mouse models in vivo. Compound 7004 can inhibit the IL-33-induced NF-κB activity, disrupt Myddosome formation, and attenuate the proinflammatory effects in asthma-like models. Our data indicate that the Myddosome may represent a novel intracellular therapeutic target for diseases in which IL-33/ST2 plays important roles, such as asthma and other inflammatory diseases.

No abstract