Levels Of Hypoxia-inducible Factor Research Articles

Hypoxic preconditioning accelerates the healing of ischemic intestinal injury by activating HIF-1α/PPARα pathway-mediated fatty acid oxidation

Hypoxic preconditioning (HPC) has been shown to improve organ tolerance to subsequent severe hypoxia or ischemia. However, its impact on intestinal ischemic injury has not been well studied. In this study, we evaluated the effects of HPC on intestinal ischemia in rats. Intestinal rehabilitation, levels of fatty acid oxidation (FAO) by-products, intestinal stem cells (ISCs), levels of hypoxia-inducible factor 1 subunit α (HIF-1α) and its downstream genes such as peroxisome proliferator-activated receptor α (PPARα), and carnitine palmitoyltransferase 1a (CPT1A) were assessed at distinct time intervals following intestinal ischemia with or without the interference of HIF-1α. Our data showed that HPC facilitates the restoration of the intestinal structure and enhances the FAO, by boosting intestinal stem cells. Additionally, HIF-1α, PPARα, and CPT1A mRNA and their protein levels were generally up-regulated in the small intestine of HPC rats as compared to the control group. Our vitro experiment also shows low-oxygen induces highly levels of HIF-1α and its downstream genes, with a concurrent increase in FAO products in IEC-6 cells. Furthermore, the above phenomenon could be reversed by silencing HIF-1α. In conclusion, we hypothesize that HPC can stimulate the activation of intestinal stem cells via HIF-1α/PPARα pathway-mediated FAO, thereby accelerating the healing process post ischemic intestinal injury.

Roxadustat ameliorates experimental colitis in mice by regulating macrophage polarization through increasing HIF level

BackgroundGastro-intestinal (GI) tract inflammation is as a result of inflammatory hypoxia which is also induced by long-standing group of disorders like inflammatory-bowel disease (IBD). Regulation of GI immune homeostasis by macrophage involves hypoxia-inducible factor (HIF). As inhibitor of HIF prolyl hydroxylase, roxadustat (ROX) increases the levels of HIF. MethodsWe induced experimental colitis (EC) model in mice via dextran-sulfate sodium (DSS) to evaluate ROX role in above-mentioned disease. ResultsROX ameliorated EC in mice by blocking colonic length shorten and loss of body weight, thereby reducing scores of disease-activity index (DAI) and histopathology. ROX significantly reduced inflammatory cytokines levels, suppressed M1 and increased M2 macrophage polarization in colonic tissues. Besides, ROX blocked declining hematocrit (HCT) level in blood and increased HIF-1-α and HIF-2-α level in colonic tissues. The inhibitor of HIF-1- α, KC7F2 decreased body weight and colonic length in ROX-treated DSS mice. Meanwhile, DAI scores and histopathology in KC7F2 treated DSS mice were markedly higher than that of treatment with ROX alone. KC7F2 treatments also significantly increased inflammatory cytokines levels, respectively promoted and reduced polarization of M1 and M2 macrophages in colonic tissue from ROX treated mice. Further, KC7F2 treatments inhibited ROX induced HCT level increasing in blood and decreased HIF-1-α and HIF-2-α level in colonic tissue. ConclusionCollectively, we discovered that ROX ameliorated EC in mice by regulating macrophage polarization through promotion of HIF expression. General significanceTaken together, we developed a new application of ROX, which provides new ideas and a scientific basis for IBD treatment.

Controlled hypoxia-hyperoxytherapy as a component of a targeted approach in the rehabilitation of patients with multimorbidity: a single-center, randomized, placebo-controlled, prospective study

BACKGROUND: Multimorbidity is one of the most important public health problems, leading not only to a deterioration in the functional abilities of an individual, but also to an increase in the disability of the population, which determines the need to use effective measures in the treatment and rehabilitation of patients with multimorbid pathology.
 AIM: The aim of this study was to evaluate the effect of a two-week medical rehabilitation program including the respiratory therapy method based on short-term intermittent exposure to hypoxia and hyperoxia on the dynamics of hypoxia-inducible factor-1 (HIF-1) and C-reactive protein in patients with multiple chronic diseases. The novelty of this study is due to the peculiarities of the study group — patients with multimorbid pathology.
 MATERIALS AND METHODS: A single-center, randomized, placebo-controlled, prospective study (NCT05053672, ClinicalTrials.gov) included 42 patients aged 43 to 68 years with multimorbid pathology: generalized osteoarthritis — 61.9%, knee OA — 21.4%, hip OA — 16.7%, controlled arterial hypertension stage I–II — 90.5%, post-Covid syndrome — 100%. Patients were randomized into 2 groups by the method of numbered identical containers. According to clinical and demographic characteristics of patients, concomitant pathology, C-reactive protein and HIF-1 levels, the studied groups were comparable. The main group (27 patients) received a course (8–10 sessions) of individually dosed hypoxy-hyperoxytherapy on the basis of biofeedback (reoxytherapy) against the background of the standard complex of medical rehabilitation. The comparison group (15 patients) received sham reoxy therapy (placebo treatment) along with standard rehabilitation.
 RESULTS: The data obtained as a result of the study in multimorbid patients using reoxy-therapy clearly demonstrated reduction of clinical symptoms, improvement of general well-being according to visual analogue scale and decrease of C-reactive protein as a classical marker of inflammation against the background of stabilization of HIF-1 dynamics.
 CONCLUSION: The use of biofeedback-based short-term intermittent exposure to hypoxia and hyperoxia (reoxy therapy) in patients with different initial characteristics in terms of clinical health status and multimorbid pathology is reasonable taking into account modern personalized approaches to medical rehabilitation.

A Genome Scale CRISPR Screen Identifies Novel Genes That Regulate Erythropoietin Production

Erythropoietin (EPO) is a plasma glycoprotein that binds bone marrow erythroid progenitors, stimulating their proliferation and differentiation. EPO is secreted into the circulation by hepatocytes and specialized kidney peritubular fibroblasts, which contribute ~20% and ~80% of the total plasma EPO level, respectively. EPO production is induced during hypoxia, due to enhanced stability of the transcription factor hypoxia-inducible factor (HIF). However, increased HIF activity results in enhanced expression of angiogenesis and oncogenesis promoting genes, raising concerns about clinical strategies that promote EPO production via increasing HIF levels. To define novel regulators of EPO production, we developed a genome-wide functional screen that provides a quantifiable and selectable readout of EPO production. In order to perform such a screen, we generated a reporter HEP3B cell line with homozygous insertion of coding sequences for p2A and eGFP at the endogenous EPO locus. This HEP3B EPO-p2A-eGFP (HEG) cell line expresses equivalent levels of EPO and eGFP proteins, both translated from the same mRNA molecule. Treatment of HEG cells with the HIF stabilizing molecule DMOG results in increased EPO expression, which is abolished by deletion of the hypoxia-responsive-element in the EPO locus. These findings demonstrate the utility of this reporter cell line to study the regulation of EPO production. To identify novel genes that regulate EPO production, we mutagenized the HEG cell line with the GeCKO-v2 CRISPR/Cas9 knock-out library, which delivers a puromycin resistance cassette, SpCas9, and a pooled collection of 123,411 single guide RNAs (sgRNAs) that include 6 independent sgRNAs targeting every human gene. Transduction was performed at a low multiplicity of infection (MOI ~0.3), so that most infected cells receive 1 sgRNA to delete 1 gene only. Puromycin selection was applied from days 1-4 post-transduction and cells were subsequently recovered for 8 days. Cells were then treated with DMOG for 24 hours or left untreated. Cells with increased (top ~10%) or decreased (bottom ~10%) eGFP fluorescence were isolated. This screen was done in biological triplicates. Genes for which sgRNAs are enriched/depleted in eGFP high versus eGFP low DMOG-treated cells likely impact EPO expression independently of the HIF pathway. In contrast, genes for which sgRNAs are enriched/depleted in eGFP high versus eGFP low untreated cells may impact EPO expression in either a HIF dependent or independent manner. As expected, sgRNAs targeting VHL were found to be among the most significantly enriched sgRNAs in eGFP-high versus eGFP-low cells under no treatment conditions. Also as anticipated, sgRNAs targeting genes known to promote EPO production ( HNF4A, WT1, RXRA, and CREBBP) were enriched in eGFP-low versus eGFP-high cells under either DMOG-treated or untreated conditions. To refine our findings, we generated a custom library targeting the top 1,200 candidate regulators of EPO production nominated by the genome-scale screen. Using this library, we performed a secondary CRISPR screen (in biological triplicates) as above. Owning to the 10-fold smaller size of the custom library, the secondary screen was performed with average coverage of >150x throughout the entire screen (including the sorted cell populations). Indeed, the statistical confidence obtained with the secondary screen was superior than that achieved in the genome scale screen. Using a false discovery rate (FDR) cutoff of 1%, gene-level analysis identified 100 and 79 candidate genes whose targeting was associated with increased or decreased EPO production respectively, under either untreated or DMOG-treated conditions. A high degree of concordance was observed for regulators of EPO expression between both screens, performed with and without DMOG treatment. To identify genes that regulate EPO production independent of HIF, we evaluated several of the top genes for which sgRNAs were enriched in eGFP high vs. eGFP low cells treated with DMOG. Using 3 independent sgRNAs targeting each of these candidates, we found 2 genes ( ZNF574 and RBM18), that when deleted, result in increased EPO mRNA, even following DMOG treatment. Additional validation experiments as well as studies aimed to define the role of these 2 genes in the regulation of EPO production are ongoing.

Chronic Intermittent Hypoxia-Induced Dysmetabolism Is Associated with Hepatic Oxidative Stress, Mitochondrial Dysfunction and Inflammation.

The association between obstructive sleep apnea (OSA) and metabolic disorders is well-established; however, the underlying mechanisms that elucidate this relationship remain incompletely understood. Since the liver is a major organ in the maintenance of metabolic homeostasis, we hypothesize that liver dysfunction plays a crucial role in the pathogenesis of metabolic dysfunction associated with obstructive sleep apnea (OSA). Herein, we explored the underlying mechanisms of this association within the liver. Experiments were performed in male Wistar rats fed with a control or high fat (HF) diet (60% lipid-rich) for 12 weeks. Half of the groups were exposed to chronic intermittent hypoxia (CIH) (30 hypoxic (5% O2) cycles, 8 h/day) that mimics OSA, in the last 15 days. Insulin sensitivity and glucose tolerance were assessed. Liver samples were collected for evaluation of lipid deposition, insulin signaling, glucose homeostasis, hypoxia, oxidative stress, antioxidant defenses, mitochondrial biogenesis and inflammation. Both the CIH and HF diet induced dysmetabolism, a state not aggravated in animals submitted to HF plus CIH. CIH aggravates hepatic lipid deposition in obese animals. Hypoxia-inducible factors levels were altered by these stimuli. CIH decreased the levels of oxidative phosphorylation complexes in both groups and the levels of SOD-1. The HF diet reduced mitochondrial density and hepatic antioxidant capacity. The CIH and HF diet produced alterations in cysteine-related thiols and pro-inflammatory markers. The results obtained suggest that hepatic mitochondrial dysfunction and oxidative stress, leading to inflammation, may be significant factors contributing to the development of dysmetabolism associated with OSA.

Estrogen receptor-related receptor γ uppresses hypoxia-induced angiogenesis by regulating VEGFA in endometrial cancer

Objective Estrogen receptor-related receptor γ (ERRγ), is implicated in cancer cell proliferation and metastasis. The function of ERRγ in tumor angiogenesis, however, is to be revealed. This study was designed to elaborate the regulatory effect of ERRγ on angiogenesis in endometrial cancer (EC). Methods Immunohistochemistry (IHC) was adopted to determine the protein expression of ERRγ, VEGFA, CD31 and hypoxia-inducible factor-1 (HIF-1) in tumor tissues. HEC-1A cells stably expressing ERRγ were established bytransfection, and then an endothelial cell tube formation assay was performed. CCK-8 assay was employed for cell viability, and wound healing assay for cell migration ability. Besides, western blot, ELISA and qRT-PCR were used to examine the VEGFA expression. After hypoxia treatment of ERRγ overexpressing HEC-1A cells, the ERRγ expression and VEGFA expression were determined by western blot. Finally, EC xenografts in nude mice were constructed by subcutaneous injection of ERRγ stably expressing HEC-1A cells and control HEC-1A cells. Results IHC results revealed a negative correlation between the expression of ERRγ and VEGFA in EC tissues. ERRγ overexpression significantly decreased the level of HIF-1 in tumor tissue of nude mice. ERRγ overexpression down-regulated inhibited angiogenesis capability and inhibited the proliferation and migration of HEC-1A cells. Furthermore, ERRγ expression was suppressed under the condition of hypoxia while restoration of ERRγ partially inhibited hypoxia-induced VEGFA expression in HEC-1A cells. Conclusions ERRγ is an angiogenesis suppressor and involved in hypoxia-induced VEGFA expression in EC. Hence, ERRγ might be a promising antiangiogenic target for human EC.

Differential expression of angiogenesis-related genes in goat uterus and corpus luteum during pregnancy.

Vascularization and the control of luteal and endometrial development are regulated by hypoxia-inducible factors (HIFs) and vascular endothelial growth factor (VEGF) during pregnancy. In this study, the mRNA and protein expression levels of HIFs (HIF1A, HIF2A and HIF3A) and VEGF in goat uterine and ovarian tissues during various stages of pregnancy were evaluated. A total of 42 Hair goats were used and were allocated into six groups, namely embryo-positive (G1), early pregnancy (G2), mid-term pregnancy (G3), late pregnancy (G4), oocyte-positive group (G5) and diestrus group (G6). The mRNA expression of the examined genes was evaluated by RT-qPCR, and protein expression was evaluated by immunohistochemistry (IHC). In caruncles, HIF1A mRNA expression was greater in G1, G2 and G4 than in G3 (p < .05). HIF1A and HIF2A expression was greater in G1 than in G5 (p < .05). In cotyledons, HIF1A, HIF2A and HIF3A mRNA expression was greater in G2 and G3 compared to G4 (p < .05). In luteal tissue, HIF1A mRNA expression was greater in G1 and G2 than in G3 and G4 (p < .05). In the immunohistochemical examination, HIF1A, HIF2A, HIF3A and VEGF immunoreactions were detected in uterine and luteal tissues. Findings suggest that HIFs and VEGF are involved in the regulation of ovarian functions as well as the processes of implantation and placentation.

Qirong Tablets inhibits apoptosis of ovarian granulosa cells via PI3K/Akt/ HIF-1 signaling pathway

This study aims to observe the effect and explore the mechanism of Qirong Tablets in the treatment of premature ovarian insufficiency(POI) in mice via the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/hypoxia inducible factor 1(HIF-1) signaling pathway. Sixty SPF female BALB/c mice were randomly divided into normal group, model group, positive control group, Qirong Tablets low-, medium-and high-dose group. The normal group was intraperitoneally injected with the same amount of normal saline, and the other groups were intraperitoneally injected with cyclophosphamide 120 mg·kg~(-1)·d~(-1) once to establish a POI animal model. After the model was successfully established, the low-, medium-and high-dose groups of Qirong Tablets were administered orally with 0.6, 1.2, 2.4 mg·kg~(-1)·d~(-1) respectively. The positive control group was given 0.22 mg·kg~(-1)·d~(-1) Clementine Tablets by intragastric administration, and the normal group and model group were given intragastric administration with the same amount of normal saline, and the treatment was 28 d as a course of treatment. After drug intervention, enzyme-linked immunosorbent assay(ELISA) was employed to measure the levels of estradiol(E_2), follicle-stimulating hormone(FSH), luteinizing hormone(LH), and anti-mullerian hormone(AMH) in peripheral blood, and hematoxylin-eosin(HE) staining to observe the ovarian tissue. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling(TUNEL) assay was used to detect the apoptosis of granulosa cells, and Western blot to determine the expression levels of B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), caspase-3, PI3K, Akt, and HIF-1. Compared with the normal group, the modeling of POI caused loose or destroyed ovarian tissue with vacuolar structures, edema and fibrosis in the ovarian interstitium, disordered or loose arrangement of granulosa cells, and reduced normal follicles. Compared with the model group, drug interventions restored the ovarian tissue and follicles at all the development stages and reduced atretic follicles. Compared with the normal group, the modeling of POI lowered the serum level of E_2 and AMH(P&lt;0.01), and elevated the level of FSH and LH(P&lt;0.01). Compared with the model group, high-dose Qirong Tablets elevated the levels of E_2 and AMH(P&lt;0.05), and lowered the levels of FSH and LH(P&lt;0.05). Compared with the normal group, the modeling of POI up-regulated the protein levels of PI3K, Akt, HIF-1, Bax, and caspase-3 and down-regulated the protein level of Bcl-2 in the ovarian tissue(P&lt;0.01). Compared with the model group, low-, medium-, and high-dose Qirong Tablets down-regulated the protein levels of PI3K, Akt, HIF-1, Bax, and caspase-3 proteins and up-regulated the protein level of Bcl-2 in the ovarian tissue(P&lt;0.05). In conclusion, Qirong Tablets can up-regulate the expression Bcl-2, down-regulate the expression of Bax and caspase-3 in POI mice. Qirong Tablets may inhibit the apoptosis of follicular granulosa cells in mice, thereby delaying ovarian aging, improving reproductive axis function, and strengthening ovarian reserve capacity, which may be associated with the inhibition of PI3K/Akt/HIF-1 pathway.

Examining the functions of the vascular endothelial growth factor/hypoxia-inducible factor signaling pathway in psoriatic arthritis.

The present study aimed to examine the roles of the vascular endothelial growth factor (VEGF), hypoxia-inducible factor (HIF), and heme oxygenase-1 (HO-1) in psoriatic arthritis (PsA). In this cross-sectional study conducted between November 2020 and May 2021, 64 patients (43 female, 21 male; mean age: 43.2±10.4 years; range, 22 to 60 years) with active PsA were included in the patient group, and 64 healthy volunteers (43 female, 21 male; mean age: 42.8±10.5 years; range, 23 to 61 years) were included in the control group. The demographic features of all cases were recorded. The following indices were used to assess the activity of PsA: Bath Ankylosing Spondylitis Disease Activity Index, Disease Activity Score in 28 joints (DAS28), and Visual Analog Scale. Additionally, Disease Activity in Psoriatic Arthritis (DAPSA) and Psoriasis Area and Severity Index (PASI) were used to evaluate the patients. The biochemical parameters of the patients were calculated. The serum levels of VEGF, HIF, and HO-1 were determined using an enzyme-linked immunosorbent assay. When the molecule levels and clinical features of the groups were evaluated, it was found that the VEGF and HIF-1 levels were higher in the patient group compared to the control group (p<0.05). No difference was observed in the comparison of the HO-1 levels of the patient group and the control group (p<0.05). A positive correlation was found between VEGF, HIF-1, and HO-1 (p<0.05). A positive relationship was found between VEGF and HIF-1 and erythrocyte sedimentation rate, C-reactive protein, DAPSA score, and PASI score (p<0.05). It was also determined that there was a positive relationship between the HIF molecule and DAS28 (p<0.05). According to the results obtained in the present study, VEGF and HIF play a role in the etiology of PsA, and the observation of intermolecular correlation suggests that these molecules move together in pathogenesis.

HIF-1 regulates pathogenic cytotoxic T cells in lupus skin disease.

Cutaneous lupus erythematosus (CLE) is a disfiguring autoimmune skin disease characterized by an inflammatory infiltrate rich in T cells, which are strongly implicated in tissue damage. How these cells adapt to the skin environment and promote tissue inflammation and damage is not known. In lupus nephritis, we previously identified an inflammatory gene program in kidney-infiltrating T cells that is dependent on HIF-1, a transcription factor critical for the cellular and developmental response to hypoxia as well as inflammation-associated signals. In our present studies using a mouse model of lupus skin disease, we find that skin-infiltrating CD4+ and CD8+ T cells also express high levels of HIF-1. Skin-infiltrating T cells demonstrated a strong cytotoxic signature at the transcript and protein levels, and HIF-1 inhibition abrogated skin and systemic diseases in association with decreased T cell cytotoxic activity. We also demonstrate in human CLE tissue that the T cell-rich inflammatory infiltrate exhibited increased amounts of HIF-1 and a cytotoxic signature. Granzyme B-expressing T cells were concentrated at sites of skin tissue damage in CLE, suggesting relevance of this pathway to human disease.

Hypoxia-Inducible Factors-Based Single Nucleotide Polymorphism in Breast Cancer with More Cancer Susceptibility.

Hypoxia-inducible factors (HIFs) are a collection of transcriptional factors that engage in the regulation of oxygen homeostasis. They are hypoxia-responsive stress factors whose expression is linked to tumor growth and angiogenesis. HIF is a crucial player in the progression of breast cancer. Patients with high levels of hypoxia-inducible HIFs in their primary tumor biopsies had a higher chance of metastasis, the leading cause of breast cancer-related death. HIF polymorphisms have been shown in several epidemiological studies to influence breast cancer susceptibility. In the oxygendependent degradation domain, several short nucleotide polymorphisms (SNPs) of the HIF gene have been connected with higher HIF activity. To find SNP that make up the genetic diversity that underpins the phenotypic difference found between individuals in their susceptibility to cancer and the course of their disease, researchers used a variety of potential pathway-based approaches.

Elucidation of the mechanisms underlying tumor aggravation by the activation of stress-related neurons in the paraventricular nucleus of the hypothalamus

A growing body of evidence suggests that excess stress could aggravate tumor progression. The paraventricular nucleus (PVN) of the hypothalamus plays an important role in the adaptation to stress because the hypothalamic–pituitary–adrenal (HPA) axis can be activated by inducing the release of corticotropin-releasing hormone (CRH) from the PVN. In this study, we used pharmacogenetic techniques to investigate whether concomitant activation of CRHPVN neurons could directly contribute to tumor progression. Tumor growth was significantly promoted by repeated activation of CRHPVN neurons, which was followed by an increase in the plasma levels of corticosterone. Consistent with these results, chronic administration of glucocorticoids induced tumor progression. Under the concomitant activation of CRHPVN neurons, the number of cytotoxic CD8+ T cells in the tumor microenvironment was dramatically decreased, and the mRNA expression levels of hypoxia inducible factor 1 subunit α (HIF1α), glucocorticoid receptor (GR) and Tsc22d3 were upregulated in inhibitory lymphocytes, tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs). Furthermore, the mRNA levels of various kinds of driver molecules related to tumor progression and tumor metastasis were prominently elevated in cancer cells by concomitant activation of CRHPVN neurons. These findings suggest that repeated activation of the PVN-CRHergic system may aggravate tumor growth through a central–peripheral-associated tumor immune system.

Stroke by inducing HDAC9-dependent deacetylation of HIF-1 and Sp1, promotes TfR1 transcription and GPX4 reduction, thus determining ferroptotic neuronal death.

Background: The inhibition of histone deacetylase 9 (HDAC9) represents a promising druggable target for stroke intervention. Indeed, HDAC9 is overexpressed in neurons after brain ischemia where exerts a neurodetrimental role. However, mechanisms of HDAC9-dependent neuronal cell death are not yet well established. Methods: Brain ischemia was obtained in vitro by primary cortical neurons exposed to glucose deprivation plus reoxygenation (OGD/Rx) and in vivo by transient middle cerebral artery occlusion. Western blot and quantitative real-time polymerase chain reaction were used to evaluate transcript and protein levels. Chromatin immunoprecipitation was used to evaluate the binding of transcription factors to the promoter of target genes. Cell viability was measured by MTT and LDH assays. Ferroptosis was evaluated by iron overload and 4-hydroxynonenal (4-HNE) release. Results: Our results showed that HDAC9 binds to hypoxia-inducible factor 1 (HIF-1) and specificity protein 1 (Sp1), two transcription activators of transferrin 1 receptor (TfR1) and glutathione peroxidase 4 (GPX4) genes, respectively, in neuronal cells exposed to OGD/Rx. Consequently, HDAC9 induced: (1) an increase in protein level of HIF-1 by deacetylation and deubiquitination, thus promoting the transcription of the pro-ferroptotic TfR1 gene; and (2) a reduction in Sp1 protein levels by deacetylation and ubiquitination, thus resulting in a down-regulation of the anti-ferroptotic GPX4 gene. Supporting these results, the silencing of HDAC9 partially prevented either HIF-1 increase and Sp1 reduction after OGD/Rx. Interestingly, silencing of the neurodetrimental factors, HDAC9, HIF-1, or TfR1 or the overexpression of the prosurvival factors Sp1 or GPX4 significantly reduced a well-known marker of ferroptosis 4-HNE after OGD/Rx. More important, in vivo, intracerebroventricular injection of siHDAC9 reduced 4-HNE levels after stroke by preventing: (1) HIF-1 and TfR1 increase and thus the augmented intracellular iron overload; and (2) a reduction of Sp1 and its target gene GPX4. Conclusions: Collectively, results obtained suggest that HDAC9 mediates post-traslational modifications of HIF-1 and Sp1 that, in turn, increases TfR1 and decreases GPX4 expression, thus promoting neuronal ferroptosis in in vitro and in vivo models of stroke.

The Novel Lysosomal Autophagy Inhibitor (ROC-325) Ameliorates Experimental Pulmonary Hypertension.

Autophagy plays an important role in the pathogenesis of pulmonary hypertension (PH). ROC-325 is a novel small molecule lysosomal autophagy inhibitor that has more potent anticancer activity than the antimalarial drug hydroxychloroquine, the latter has been prevalently used to inhibit autophagy. Here, we sought to determine the therapeutic benefit and mechanism of action of ROC-325 in experimental PH models. Hemodynamics, echocardiography, and histology measurement showed that ROC-325 treatment prevented the development of PH, right ventricular hypertrophy, fibrosis, dysfunction, and vascular remodeling after monocrotaline and Sugen5416/hypoxia administration. ROC-325 attenuated high K+ or alveolar hypoxia-induced pulmonary vasoconstriction and enhanced endothelial-dependent relaxation in isolated pulmonary artery rings. ROC-325 treatment inhibited autophagy and enhanced endothelial nitric oxide synthase activity in lung tissues of monocrotaline-PH rats. In cultured human and rat pulmonary arterial smooth muscle cell and pulmonary arterial endothelial cell under hypoxia exposure, ROC-325 increased LC3B (light chain 3 beta) and p62 accumulation, endothelial cell nitric oxide production via phosphorylation of endothelial nitric oxide synthase (Ser1177) and dephosphorylation of endothelial nitric oxide synthase (Thr495) as well as decreased HIF (hypoxia-inducible factor)-1α and HIF-2α stabilization. These data indicate that ROC-325 is a promising novel agent for the treatment of PH that inhibits autophagy, downregulates HIF levels, and increases nitric oxide production.

Hypoxia-inducible factor expression is related to apoptosis and cartilage degradation in temporomandibular joint osteoarthritis

BackgroundIt remains unclear etiology of cartilaginous tissues in osteoarthritis (OA) lesions. In this study, we hypothesized the accumulation of hypoxia-inducible factor (HIF) and activated apoptosis relate to condylar cartilage degeneration in vivo.MethodsMalocclusion stress was applied for 2 weeks, 4 weeks and 8 weeks to induce an OA-like lesion animal model in rats. Histological analysis was performed by H&E staining and Safranin O/fast green staining. The expression levels of protein in condylar cartilage were examined by immunostaining to evaluate cartilage degeneration.ResultsWe found apparent histological phenotypes associated with degeneration in the occlusion disorder (OD) stress group. The OD group at 4 weeks and 8 weeks had obviously reduced expression of Aggrecan (Acan) and type II collagen (Col II) in cartilage. In contrast, the OD groups had higher levels of ADAM metallopeptidase with thrombospondin type 5 (ADAMTS5) and matrix metallopeptidase 13 (MMP13) in the condylar cartilage than the control group. Moreover, the OD group cartilage had prominent degenerative changes with reduced levels of hypoxia inducible factor 1 alpha (HIF1α) and increased levels of hypoxia inducible factor 2 alpha (HIF2α) and the apoptosis factor Caspase3 in condylar cartilage at 8 weeks.ConclusionThus, abnormal hypoxic conditions inducing Occlusion disorder stress results in cartilage degeneration. opposite expression patterns of HIF1α and HIF2α could be involved in the pathogenesis of condylar cartilage degeneration and chondrocyte apoptosis. HIF2α may provide a potential negative feedback mechanism for HIF1α during cartilage damage.

The Effect of Hypothermic Machine Perfusion to Ameliorate Ischemia-Reperfusion Injury in Donor Organs.

Hypothermic machine perfusion (HMP) has become the new gold standard in clinical donor kidney preservation and a promising novel strategy in higher risk donor livers in several countries. As shown by meta-analysis for the kidney, HMP decreases the risk of delayed graft function (DGF) and improves graft survival. For the liver, HMP immediately prior to transplantation may reduce the chance of early allograft dysfunction (EAD) and reduce ischemic sequelae in the biliary tract. Ischemia-reperfusion injury (IRI), unavoidable during transplantation, can lead to massive cell death and is one of the main causes for DGF, EAD or longer term impact. Molecular mechanisms that are affected in IRI include levels of hypoxia inducible factor (HIF), induction of cell death, endothelial dysfunction and immune responses. In this review we have summarized and discussed mechanisms on how HMP can ameliorate IRI. Better insight into how HMP influences IRI in kidney and liver transplantation may lead to new therapies and improved transplant outcomes.

Human amniotic fluid stem cells can alleviate detrusor dysfunction caused by bladder outlet obstruction in rats

The present study examined whether bladder detrusor dysfunction due to partial bladder outlet obstruction (pBOO) could be improved after the treatment of human amniotic fluid stem cells (hAFSCs). 72 female rats were grouped into sham operation, pBOO, and pBOO with hAFSCs treatment (pBOO + hAFSCs) for in vitro and in vivo studies. Bladder weight, bladder wall thickness, the ratio of collagen to smooth muscle and the levels of positive CD11b/c and HIS48 cells was significantly increased after pBOO but improved after hAFSCs treatment. Cystometries showed impaired bladder function after pBOO. Protein and mRNA levels of hypoxia inducible factor-1α, CCL2, interleukin-1β, transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF), α-smooth muscle actin, collagen I and collagen III were increased at 2 and/or 6 weeks, but proteins and mRNA expressions of protein gene product 9.5 were decreased at 2 and 6 weeks after pBOO. These abnormalities were improved after hAFSCs treatment. The expressions of TGF-β1 and CTGF in cultured detrusor cells of pBOO rats were increased but were improved after hAFSCs treatment. The present results showed hAFSCs treatment could improve bladder detrusor dysfunction in pBOO rats, which may be related to the reduction of inflammatory and pro-fibrotic markers in detrusor muscle cells.

Periodontal ligament cells under mechanical force regulate local immune homeostasis by modulating Th17/Treg cell differentiation.

Improper orthodontic force often causes root resorption or destructive bone resorption. There is evidence that T helper 17 (Th17) cells and regulatory T (Treg) cells may be actively involved in bone remodeling during tooth movement. In a combination of in vitro and in vivo studies, we investigated the effect of human periodontal ligament cells (hPDLCs) on Th17/Treg cells under different orthodontic forces and corticotomy. hPDLCs were cultured in vitro and subjected to different mechanical forces. The expression of interleukin (IL)-6 and transforming growth factor (TGF)-β in the supernatant and the mRNA levels of hypoxia inducible factor (HIF)-1α, Notch1, and TGF-β in hPDLCs were investigated. Supernatants were collected and co-cultured with activated CD4+T cells, and the differentiation of Th17/Treg cells was analyzed by flow cytometry. We also established an animal model of tooth movement with or without corticotomy. The tooth movement distance, alveolar bone height, and root resorption were analyzed using micro-computed tomography. Expression of interleukin (IL)-17A, forkhead Box P3 (Foxp3), and IL-6 were analyzed using immunohistochemistry, while osteoclasts were evaluated by tartrate-resistant acid phosphatase (TRAP) staining. The mRNA levels of IL-17A, IL-6, Foxp3, IL-10, HIF-1α, notch1, and C-X-C motif chemokine ligand 12 (CXCL12) in alveolar bone and gingiva were investigated. Heavy force repressed cell viability and increased the mortality rate of hPDLCs; it also improved the expression of IL-6, declined the expression of TGF-β, and promoted the mRNA expression level of HIF-1α. The expression of TGF-β and Notch1 mRNA decreased and then increased. The supernatant of hPDLCs under heavy force promotes the polarization of Th17 cells. The heavy force caused root resorption and decreased alveolar bone height and increased the positive area of IL-17A immunohistochemical staining and the expression of IL-17A, IL-6, HIF-1α, and Notch1 mRNA. Corticotomy accelerated tooth movement, increased the proportion of Foxp3-positive cells, and up-regulated the expression of Foxp3, IL-10, and CXCL12 mRNA. During orthodontic tooth movement, the heavy force causes root resorption and inflammatory bone destruction, which could be associated with increased expression of Th17 cells and IL-6. Corticotomy can accelerate tooth movement without causing root resorption and periodontal bone loss, which may be related to the increased expression of Treg cells. Altogether, this report provides a new perspective on the prevention of inflammatory injury via the regulation of Th17/Treg cells in orthodontics.

The Effect of Hypoxia Inducible Factor -1 Alpha and Vascular Endothelial Growth Factor Level in Type 2 Diabetes Microvascular Complications and Development.

Background:Angiogenesis in diabetic patients is often caused by hyperglycemia induced by hypoxiaObjective:The aim of this study was to analyze the serum level of Hypoxia Inducible Factor -1α (HIF-1α) and Vascular Endothelial Growth Factor (VEGF) between March until Desember 2020.Methods:This is a cross-sectional analytic methods, 135 patients with Type 2 Diabetes 48 samples with Microvascular complication and 87 samples with non-microvascular complication were recruited from the various primary health care centers in Medan city and surrounding areas in North Sumatera. VEGF levels and HIF-1α tested were done with ELISA methods in the laboratory of Medical Faculty, Universitas Sumatera Utara. Statistical analysis was performed using the IBM SPSS Statistics version 24. The significance level was set up to 0.005.Results:The median HIF-1 levels in patients with microvascular complications were lower than those without microvascular complications, with a range of HIF-1α values in non-complicated samples (0.02-13.96) ng/ml and a range of HIF-1α values in vascular complications (0.52- 8.87) mg/dL. There was a significant difference in HIF-1α levels in patients with Type-2 DM with complications compared to those without complications (p<0.05). Median VEGF levels were higher in complicated Type-2 DM. There was no difference in VEGF levels in patients with Type-2 DM with complications compared to those without complications (p > 0.005).Conclusion:HIF-1α and VEGF levels showed the development in vascularity. With the higher level of HIF-1α, an increase in VEGF levels were found, indicating the angiogenesis is occurring. Although complications have not yet occurred, it is predicted that high VEGF values will cause vascular complications in the future.

Research Progress of Traditional Chinese Medicine Intervention on HIF-2α in the Prevention and Treatment of Knee Osteoarthritis

Knee osteoarthritis (KOA) is the most common chronic, degenerative and progressive osteoarthritis in clinical practice, which mainly manifests as knee pain, deformity and even joint function loss. With the advent of the global aging era, its incidence continues to increase, seriously occupying the medical resources of various countries. Its pathogenic factors are complex and its pathogenesis is not clear. How to effectively prevent and cure it has always been a hot topic in the medical field. Recent studies have found that the expression level of hypoxia inducible factor (HIIF) -2α plays an important role in regulating chondrocyte homeostasis, which is involved in the degradation of chondrocyte extracellular matrix and the generation of subchondral bone vessels, and accelerates cartilage degeneration. Studies have shown that the active ingredients of traditional Chinese medicine can regulate the expression of HIF-2α and slow down the development of knee osteoarthritis. This article reviews the related research on the prevention and treatment of knee osteoarthritis by the intervention of traditional Chinese medicine on HIF-2α expression, so as to provide scientific clinical basis and treatment plan for the later research on the treatment of KOA with traditional Chinese medicine.