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Our understanding of how the course of opportunistic bacterial infection is influenced by the microenvironment is limited. We demonstrate that the pathogenicity of Pseudomonas aeruginosa strains derived from acute clinical infections is higher than that of strains derived from chronic infections, where tissues are hypoxic. Exposure to hypoxia attenuated the pathogenicity of strains from acute (but not chronic) infections, implicating a role for hypoxia in regulating bacterial virulence. Mass spectrometric analysis of the secretome of P. aeruginosa derived from an acute infection revealed hypoxia-induced repression of multiple virulence factors independent of altered bacterial growth. Pseudomonas aeruginosa lacking the Pseudomonas prolyl-hydroxylase domain–containing protein, which has been implicated in bacterial oxygen sensing, displays reduced virulence factor expression. Furthermore, pharmacological hydroxylase inhibition reduces virulence factor expression and pathogenicity in a murine model of pneumonia. We hypothesize that hypoxia reduces P. aeruginosa virulence at least in part through the regulation of bacterial hydroxylases.

Hypoxia plays a critical role during the evolution of malignant cells and tumour microenvironment (TME).Tumour-derived exosomes contain informative microRNAs involved in the interaction of cancer and stromal cells, thus contributing to tissue remodelling of tumour microenvironment. This study aims to clarify how hypoxia affects tumour angiogenesis through exosomes shed from lung cancer cells. Lung cancer cells produce more exosomes under hypoxic conditions than do parental cells under normoxic conditions. miR-23a was significantly upregulated in exosomes from lung cancer under hypoxic conditions. Exosomal miR-23a directly suppressed its target prolyl hydroxylase 1 and 2 (PHD1 and 2), leading to the accumulation of hypoxia-inducible factor-1 α (HIF-1 α) in endothelial cells. Consequently, hypoxic lung cancer cells enhanced angiogenesis by exosomes derived from hypoxic cancer under both normoxic and hypoxic conditions. In addition, exosomal miR-23a also inhibits tight junction protein ZO-1, thereby increasing vascular permeability and cancer transendothelial migration. Inhibition of miR-23a by inhibitor administration decreased angiogenesis and tumour growth in a mouse model. Furthermore, elevated levels of circulating miR-23a are found in the sera of lung cancer patients, and miR-23a levels are positively correlated with proangiogenic activities. Taken together, our study reveals the clinical relevance and prognostic value of cancer-derived exosomal miR-23a under hypoxic conditions, and investigates a unique intercellular communication, mediated by cancer-derived exosomes, which modulates tumour vasculature.

AbstractHypoxia inducible factor (HIF)-prolyl hydroxylase (PHD) inhibitors are shown to be protective in several models of inflammatory bowel disease (IBD). However, these non-selective inhibitors are known to inhibit all the three isoforms of PHD, i.e. PHD-1, PHD-2 and PHD-3. In the present report, we investigated the associated changes in levels of PHDs during the development and recovery of chemically induced colitis in mice. The results indicated that in the experimental model of murine colitis, levels of both, PHD-1 and PHD-2 were found to be increased with the progression of the disease; however, the level of PHD-3 remained the same in group of healthy controls and mice with colitis. Thus, the findings advocated that inhibitors, which inhibited all three isoforms of PHD could not be ideal therapeutics for IBD since PHD-3 is required for normal gut function. Hence, this necessitates the development of new compounds capable of selectively inhibiting PHD-1 and PHD-2 for effective treatment of IBD.

Mesenchymal stem cell (MSC) transplantation alone may be insufficient for treatment of liver fibrosis because of complicated histopathological changes in the liver. Given that miR‐122 plays an essential role in liver fibrosis by negatively regulating the proliferation and transactivation of hepatic stellate cells (HSCs), this study investigated whether miR‐122 modification can improve the therapeutic efficacy of adipose tissue‐derived MSCs in treating liver fibrosis. MiR‐122‐modified AMSCs (AMSC‐122) were constructed through lentivirus‐mediated transfer of pre‐miR‐122. MiR‐122‐modified AMSCs expressed high level of miR‐122, while they retained their phenotype and differentiation potential as naïve AMSCs. AMSC‐122 more effectively suppressed the proliferation of and collagen maturation in HSCs than scramble miRNA‐modified AMSCs. In addition, AMSC‐derived exosomes mediated the miR‐122 communication between AMSCs and HSCs, further affecting the expression levels of miR‐122 target genes, such as insulin‐like growth factor receptor 1 (IGF1R), Cyclin G(1) (CCNG1) and prolyl‐4‐hydroxylase α1 (P4HA1), which are involved in proliferation of and collagen maturation in HSCs. Moreover, miR‐122 modification enhanced the therapeutic efficacy of AMSCs in the treatment of carbon tetrachloride (CCl4)‐induced liver fibrosis by suppressing the activation of HSCs and alleviating collagen deposition. Results demonstrate that miR‐122 modification improves the therapeutic efficacy of AMSCs through exosome‐mediated miR‐122 communication; thus, miR‐122 modification is a new potential strategy for treatment of liver fibrosis.

A specific bone vessel subtype, strongly positive for CD31 and endomucin (CD31 hi Emcn hi ), is identified as coupling angiogenesis and osteogenesis. The abundance of type CD31 hi Emcn hi vessels decrease during ageing. Here we show that expression of the miR-497∼195 cluster is high in CD31 hi Emcn hi endothelium but gradually decreases during ageing. Mice with depletion of miR-497∼195 in endothelial cells show fewer CD31 hi Emcn hi vessels and lower bone mass. Conversely, transgenic overexpression of miR-497∼195 in murine endothelium alleviates age-related reduction of type CD31 hi Emcn hi vessels and bone loss. miR-497∼195 cluster maintains the endothelial Notch activity and HIF-1α stability via targeting F-box and WD-40 domain protein (Fbxw7) and Prolyl 4-hydroxylase possessing a transmembrane domain (P4HTM) respectively. Notably, endothelialium-specific activation of miR-195 by intravenous injection of aptamer-agomiR-195 stimulates CD31 hi Emcn hi vessel and bone formation in aged mice. Together, our study indicates that miR-497∼195 regulates angiogenesis coupled with osteogenesis and may represent a potential therapeutic target for age-related osteoporosis. H-type endothelium, defined by the high expression of CD31 and endomucin, is found in the bone where it promotes angiogenesis and osteogensis. Here Yang et al . show that the miR-497∼195 cluster regulates the generation and maintenance of the H-type endothelium by controlling the levels of Notch regulator Fbxw7 and the HIF regulator P4HTM.

Uncontrolled inflammation underpins a diverse range of diseases where effective therapy remains an unmet clinical need. Hypoxia is a prominent feature of the inflammatory microenvironment that regulates key transcription factors including HIF and NF-κB in both innate and adaptive immune cells. In turn, altered activity of the pathways controlled by these factors can affect the course of inflammation through the regulation of immune cell development and function. In this review, we will discuss these pathways and the oxygen sensors that confer hypoxic sensitivity in immune cells. Furthermore, we will describe how hypoxia-dependent pathways contribute to immunity and discuss their potential as therapeutic targets in inflammatory and infectious disease.

Key Points Hypoxia-inducible factors (HIFs) were originally described as transcription factors that promote transcriptional responses during hypoxia adaptation (for example, increased erythropoietin release following blood loss or during hypoxia). More recently, it has been appreciated that HIFs are also active during a wider range of disease conditions, including inflammation and ischaemia. During inflammatory or ischaemic conditions, HIFs are stabilized and are transcriptionally active. In many instances, they promote a gene programme that dampens acute inflammation and that promotes resolution of injury. During normoxic conditions, HIFs are inactive, as they are targeted for proteasomal degradation by prolyl hydroxylases (PHDs). During hypoxic conditions, PHDs are inactive, and HIFs become stabilized and transcriptionally active. Pharmacologically, HIFs can be stabilized by small molecules that function to inhibit PHDs. Such compounds have recently gained great interest. Using pharmacological HIF activators (mostly PHD inhibitors), a HIF-dependent gene expression programme can be activated with the goal to dampen inflammation while promoting the resolution of injury. Examples for diseases for which there is experimental evidence indicating a protective role for HIF activators include inflammatory bowel disease (IBD), acute lung injury (ALI), myocardial ischaemia and reperfusion injury, acute kidney injury and organ transplantation. Other studies have identified pharmacological approaches to inhibit HIFs. Such approaches could hold therapeutic potential in the treatment of cancer or fibrosis (for example, renal fibrosis). Several companies have developed orally available PHD inhibitors that promote HIF stabilization under normoxic conditions. Such compounds are currently examined in clinical trials, for example, for the treatment of renal anaemia or perioperative ischaemia and reperfusion injury. Although most clinical and experimental studies so far have found that short-term PHD inhibitor treatment is safe, careful monitoring for potentially detrimental side effects of PHD inhibitors will be crucial, particularly for their chronic application in patients. Potential side effects could include liver injury, sepsis, increased erythropoiesis or cancer. Challenges for the field include a better understanding of the specific roles for individual HIF isoforms (for example, HIF1α and HIF2α), the design of pharmacological approaches to specifically target individual PHDs or individual HIFs, and pharmacological delivery systems that allow tissue-specific delivery of HIF activators (for example, via inhalation to alveolar epithelial cells or approaches to specifically target intestinal epithelial cells). We anticipate that, in the near future, HIF activators will be used routinely in a clinical setting for organ protection in patients experiencing ischaemia and reperfusion injury, as well as to promote the resolution of inflammation during acute or chronic inflammatory disease states such as IBD or ALI. Hypoxia-inducible factors (HIFs) have important roles in ischaemic and inflammatory diseases and strategies aimed at therapeutically modulating hypoxia signalling pathways are gaining considerable attention. Here, Eltzschig and colleagues focus on a set of oxygen-sensing prolyl hydroxylases — which are responsible for marking HIFs for proteasomal degradation — and assess their emerging potential as therapeutic targets. Hypoxia-inducible factors (HIFs) are stabilized during adverse inflammatory processes associated with disorders such as inflammatory bowel disease, pathogen infection and acute lung injury, as well as during ischaemia–reperfusion injury. HIF stabilization and hypoxia-induced changes in gene expression have a profound impact on the inflamed tissue microenvironment and on disease outcomes. Although the mechanism that initiates HIF stabilization may vary, the final molecular steps that control HIF stabilization converge on a set of oxygen-sensing prolyl hydroxylases (PHDs) that mark HIFs for proteasomal degradation. PHDs are therefore promising therapeutic targets. In this Review, we discuss the emerging potential and associated challenges of targeting the PHD–HIF pathway for the treatment of inflammatory and ischaemic diseases.

Around birth, globin expression in human red blood cells (RBCs) shifts from γ-globin to β-globin, which results in fetal haemoglobin (HbF, α 2 γ 2 ) being gradually replaced by adult haemoglobin (HbA, α 2 β 2 ) 1 . This process has motivated the development of innovative approaches to treat sickle cell disease and β-thalassaemia by increasing HbF levels in postnatal RBCs 2 . Here we provide therapeutically relevant insights into globin gene switching obtained through a CRISPR–Cas9 screen for ubiquitin–proteasome components that regulate HbF expression. In RBC precursors, depletion of the von Hippel–Lindau (VHL) E3 ubiquitin ligase stabilized its ubiquitination target, hypoxia-inducible factor 1α (HIF1α) 3 , 4 , to induce γ-globin gene transcription. Mechanistically, HIF1α–HIF1β heterodimers bound cognate DNA elements in BGLT3 , a long noncoding RNA gene located 2.7 kb downstream of the tandem γ-globin genes HBG1 and HBG2 . This was followed by the recruitment of transcriptional activators, chromatin opening and increased long-range interactions between the γ-globin genes and their upstream enhancer. Similar induction of HbF occurred with hypoxia or with inhibition of prolyl hydroxylase domain enzymes that target HIF1α for ubiquitination by the VHL E3 ubiquitin ligase. Our findings link globin gene regulation with canonical hypoxia adaptation, provide a mechanism for HbF induction during stress erythropoiesis and suggest a new therapeutic approach for β-haemoglobinopathies. Detailed mechanistic insight into fetal globin gene induction during hypoxia-associated stress erythropoiesis provides new therapeutic approaches to treat β-haemoglobinopathies, such as sickle cell disease and β-thalassaemia.

Yes-associated protein 1 (YAP1), a key player in the Hippo pathway, has been shown to play a critical role in tumor progression. However, the role of YAP1 in prostate cancer cell invasion, migration, and metastasis is not well defined. Through functional, transcriptomic, epigenomic, and proteomic analyses, we showed that prolyl hydroxylation of YAP1 plays a critical role in the suppression of cell migration, invasion, and metastasis in prostate cancer. Knockdown (KD) or knockout (KO) of YAP1 led to an increase in cell migration, invasion, and metastasis in prostate cancer cells. Microarray analysis showed that the EMT pathway was activated in Yap1 -KD cells. ChIP-seq analysis showed that YAP1 target genes are enriched in pathways regulating cell migration. Mass spectrometry analysis identified P4H prolyl hydroxylase in the YAP1 complex and YAP1 was hydroxylated at multiple proline residues. Proline-to-alanine mutations of YAP1 isoform 3 identified proline 174 as a critical residue, and its hydroxylation suppressed cell migration, invasion, and metastasis. KO of P4ha2 led to an increase in cell migration and invasion, which was reversed upon Yap1 KD. Our study identified a novel regulatory mechanism of YAP1 by which P4HA2-dependent prolyl hydroxylation of YAP1 determines its transcriptional activities and its function in prostate cancer metastasis.

Hypoxia-inducible factor-prolyl hydroxylase (HIF-PHD) inhibitors are therapeutic agents for renal anemia that work through HIF2-mediated upregulation of erythropoietin (EPO) and have also been reported to suppress renal fibrosis. Group 2 innate lymphoid cells (ILC2s) have been proven to be involved in the pathogenesis of fibrosis in various organs, including the kidney. However, the relationship between the HIF pathway, renal fibrosis, and kidney ILC2s remains unclear. In the present study, we found that HIF activation by HIF-PHD inhibitors suppressed type 2 cytokine production from kidney ILC2s. The enhanced HIF pathway downregulated the IL-33 receptor ST2L on ILC2s, and phosphorylation of downstream p38 MAPK was attenuated. M2 macrophages that promote renal fibrosis were polarized by ILC2 supernatants, but reduced cytokine production from ILC2s treated with HIF-PHD inhibitors suppressed this polarization. Our findings suggest that HIF-PHD inhibitors are potential therapeutic agents for renal fibrosis that are mediated by the alteration of ILC2 function.