Hypoxia Pathways Research Articles

Lactylation Modification as a Promoter of Bladder Cancer: Insights from Multi-Omics Analysis

Bladder cancer (BLAC) is a malignant tumor with high morbidity and mortality. The establishment of a prognostic model for BLAC is of great significance for clinical prognosis prediction and treatment guidance. Lactylation modification is a newly discovered post-transcriptional modification of proteins, which is closely related to the occurrence and development of tumors. Multiple omics data of BLAC were obtained from the GEO database and TCGA database. The Lasso algorithm was used to establish a prognostic model related to lactylation modification, and its predictive ability was tested with a validation cohort. Functional enrichment analysis, tumor microenvironment analysis, and treatment response evaluation were performed on the high- and low-risk groups. Single-cell and spatial transcriptome data were used to analyze the distribution characteristics of model genes and their changes during epithelial carcinogenesis. A prognostic model consisting of 12 genes was constructed. The survival rate of the high-risk group was significantly lower than that of the low-risk group. The multiple ROC curve showed that the prediction efficiency of the model was higher than that of the traditional clinical tumor grading. Functional enrichment analysis showed that glycolysis and hypoxia pathways were significantly upregulated in the high-risk group. The high-risk group was more sensitive to most first-line chemotherapy drugs, while the low-risk group had a better response to immunotherapy. Single-cell sequencing analysis revealed the dynamic changes of model genes during the transition of epithelial cells to squamous-differentiated cells. Spatial transcriptome analysis showed the spatial distribution characteristics of the model genes. The lactylation-related models have a satisfactory predictive ability and the potential to guide the clinical treatment of BLAC. This model has significant biological implications at the single-cell level as well as at the spatial level.

Abstract 4136204: Clearing Senescent Cells Improves Mouse Survival Rate Post Myocardial Infarction through Alteration of Cardiomyocyte and Immune Cell Subpopulations

Introduction: Cellular senescence often involves a p16-pathway, and p16 overexpression is a hallmark of senescent cells. The role of cellular senescence in myocardial infarction (MI) and any mediating mechanisms remain unclear. Aims: To investigate the effect of p16 + cell clearance on survival post MI and elucidate underlying mechanisms. Methods: We utilized INK-ATTAC transgenic mice, in which p16 + cells undergo targeted apoptosis upon exposure to AP20187 (AP). Sham and MI mice were treated with AP or vehicle (V) twice-weekly for one month, starting 3-4 hours post-MI. Survival rate improvement post MI in the AP group (Fig A, P<0.001) prompted us to evaluate effects of senescent cell clearance at the cardiac cellular level and its involvement in MI-associated pathways at 3 days post-MI. The infarct and peri-infarct areas were collected and snap-frozen. Nuclei were extracted with modified gentleMACS methods; libraries were constructed using 10X Genomics. Standard software (CellRanger, Seurat) were employed for single-nuclei-RNA-seq data analysis. Results: Cardiomyocytes (CMs) showed 5 distinct subsets. CM subclusters 1 and 2 showed enrichments of genes characteristic of infarct boarder zones, while most sham CMs were found in subcluster 0, reflecting healthy-CMs. AP-treated mice exhibited a higher proportion of healthy CMs (Fig B). Differentially expressed genes (DEGs) between V and AP group were enriched in the healthy CMs, suggesting that clearance of p16 + cells improves the status of CMs post MI (Fig C). Immune-cell profiling showed that by far the largest number of DEG for AP vs vehicle were expressed in the (arginase-1) Arg1 macrophage subgroup (Fig D). The percentage of Arg1 macrophages decreased in the AP group (FDR=0.059) (Fig E). We noted significant enrichment in the hypoxia and cardiac rupture pathways in Arg1 macrophages (Fig F), correlating with the high death rate from myocardial rupture in V-mice. Conclusion: These results suggest a significant role of p16 + senescent cells in post-MI mortality, potentially through effects on cardiomyocytes and a specific macrophage subpopulation, Arg1 macrophages.

TMIC-09. DENDRITIC CELL DYSFUNCTION RESTRICTS TUMOR ANTIGEN SPREADING IN ADOPTIVE CELLULAR TRANSFER THERAPY-ESCAPED GLIOMAS: IMPLICATIONS FOR TUMOR IMMUNE EVASION

Abstract Dendritic cells (DCs) play a pivotal role in initiating anti-tumor immune responses. Our group has developed an adoptive cellular transfer therapy (ACT) that significantly increases the infiltration of DCs and T cells into tumors, markedly enhancing survival in murine brain tumor models. However, tumors develop mechanisms to restrict the immune surveillance and the efficacy of immunotherapy, leading to tumor escape. In this study, we identified a mechanism involving DC dysfunction in ACT-escaped brain tumors. The finding that DCs are enriched after ACT treatment prompts an investigation into whether DC function is impaired in ACT-escaped tumors. Our results showed that DCs from both primary and ACT-escaped tumors were dysfunctional in their ability to activate T cells when co-cultured with tumor-reactive T cells or OT1 T cells. RNA-seq data revealed that, compared to primary tumor DCs, ACT-escaped tumor DCs exhibited a more pronounced reduction in conventional DC markers and antigen-presenting genes, alongside increased tolerance markers. Gene set enrichment analysis indicated that epithelial-mesenchymal transition (EMT) and hypoxia pathways were significantly enriched in ACT-escaped tumor DCs, suggesting divergent DC dysfunction mechanisms in primary versus escaped tumors. The anti-tumor role of DCs is primarily exerted through the activation of tumor-reactive T cells. Therefore, we assessed the cytotoxicity and exhaustion status of T cells in tumors. Surprisingly, we found that T cells from ACT-escaped tumors exhibited significantly higher levels of cytotoxicity and lower levels of T cell exhaustion compared to primary tumors. This suggests a T cell exhaustion-independent mechanism in ACT-escaped tumors. We further demonstrated that T cells in ACT-escaped tumor are mainly sourced from the adoptive transferred T cells, which recognize primary tumor antigens but not escaped tumor antigens. These results suggest that a combined mechanism of tumor antigen shifting and impaired DC function leads to a failure of antigen spreading and tumor eradication.

Molecular docking and molecular dynamics of hypoxia-inducible factor (HIF-1alpha): towards potential inhibitors

HIF-1α is a primary regulator in the adaptation of cancer cells to hypoxia. The aim was to find out new inhibitors of the HIF-1α. A molecular dynamic (MD) simulation performed on HIF-1α showed stable dynamic features. Virtual screening of 217 anticancer drugs was performed along with a positive control (2-Methoxyestradiolm, 2-ME2) on an optimized HIF-1α and dynamically simulated structure. Docking results produced two compounds namely pycnidione and nilotinib of high binding affinity −9.34 kcal/mol and −9.04 kcal/mol respectively, whereas 2-ME2 displayed a relatively lower affinity (-6.68 kcal/mol). For the three complexes, MD of 200 ns simulation was run. Data analysis showed that the three medications behaved similarly in the MD simulation. Nilotinib had a lower RMSD and higher SASA than the other complexes. In addition, the Nilotinib-HIF-1α combination had a lower RMSF value, a flatter Rg, and a number of hydrogen bonds similar to other complexes. MM-GBSA analysis revealed that nilotinib, pycnidione and 2-ME2 compounds had free binding energy of −23.77 ± 5.29, −21.85 ± 4.24 and −7.53 ± 6.62 kcal/mol respectively. Nilotinib and pycnidione bind competitively to HIF-1α, with nilotinib showing consistent molecular-dynamic properties. They relatively pass the blood-brain barrier, non-carcinogenic, and have IV-category acute oral toxicity. They have low CYP inhibitory characteristics. Further investigations are therefore warranted to elucidate their implications in hypoxia pathways, cell proliferation, apoptosis, survival, and metastatic potential.

Onvansertib in Combination With Chemotherapy and Bevacizumab in Second-Line Treatment of KRAS-Mutant Metastatic Colorectal Cancer: A Single-Arm, Phase II Trial.

This phase II study evaluated the efficacy and tolerability of onvansertib, a polo-like kinase 1 (PLK1) inhibitor, in combination with fluorouracil, leucovorin, and irinotecan (FOLFIRI) + bevacizumab for the second-line treatment of KRAS-mutant metastatic colorectal cancer (mCRC). This multicenter, open-label, single-arm study enrolled patients with KRAS-mutated mCRC previously treated with oxaliplatin and fluorouracil with or without bevacizumab. Patients received onvansertib (15 mg/m2 once daily on days 1-5 and 15-19 of a 28-day cycle) and FOLFIRI + bevacizumab (days 1 and 15). The primary end point was the objective response rate (ORR), and secondary endpoints included progression-free survival (PFS), duration of response (DOR), and tolerability. Translational and preclinical studies were conducted in KRAS-mutant CRC. Among the 53 patients treated, the confirmed ORR was 26.4% (95% CI, 15.3 to 40.3). The median DOR was 11.7 months (95% CI, 9.4 to not reached). Grade 3/4 adverse events were reported in 62% of patients. A post hoc analysis revealed that patients with no prior bevacizumab treatment had a significantly higher ORR and longer PFS compared with patients with prior bevacizumab treatment: ORR of 76.9% versus 10.0% (odds ratio of 30.0, P < .001) and median PFS of 14.9 months versus 6.6 months (hazard ratio of 0.16, P < .001). Our translational findings support that prior bevacizumab exposure contributes to onvansertib resistance. Preclinically, we showed that onvansertib inhibited the hypoxia pathway and exhibited robust antitumor activity in combination with bevacizumab through the inhibition of angiogenesis. Onvansertib in combination with FOLFIRI + bevacizumab showed significant activity in the second-line treatment of patients with KRAS-mutant mCRC, particularly in patients with no prior bevacizumab treatment. These findings led to the evaluation of the combination in the first-line setting (ClinicalTrails.gov identifier: NCT06106308).

Over-expression of Transmembrane Protein 158 Predicts Aggressive Tumor Behavior and Poor Prognosis in Lung Cancer.

Transmembrane protein 158 (TMEM158) has emerged as a potential contributor to cancer progression. While TMEM158 has been studied in various cancer types, its role in lung cancer remains unclear. Kaplan-Meier curves were used to evaluate the association between TMEM158 expression and overall survival rates. Gene set enrichment analysis (GSEA) was performed to explore pathways related to TMEM158, and sequence analysis was conducted to identify putative hypoxia-responsive element (HRE) sites in the TMEM158 promoter region. Hypoxic conditions were induced, and TMEM158 expression levels were measured by qPCR. The effect of hypoxia-inducible factor-1α (HIF-1α) depletion on TMEM158 expression was also examined. Additionally, lung cancer cells with either over-expressed or reduced TMEM158 levels were analyzed for epithelial-mesenchymal transition (EMT) and cell migration. Analysis of clinical datasets revealed elevated TMEM158 expression in tumors, particularly in patients with advanced stages. High TMEM158 expression was correlated with lower overall survival. TMEM158 was associated with EMT, hypoxia, and other tumor-promoting pathways. Under hypoxic conditions, TMEM158 expression was at least partially induced in a HIF-1α-dependent manner. Functional studies showed that over-expression of TMEM158 promoted EMT and increased lung cancer cell migration, while depletion of TMEM158 reduced these effects, indicating its role in aggressive tumor behavior. TMEM158 is highly expressed in lung cancer, is associated with hypoxia, and promotes EMT and cell migration. These findings suggest TMEM158 as a potential target for lung cancer therapies.

Novel bioassays based on 3D-printed device for sensing of hypoxia and p53 pathway in 3D cell models.

Cell-based assays are widely exploited for drug screening and biosensing, providing useful information about bioactivity of target analytes and complex biological samples. It is well recognized that 3D cell models are required to achieve highly valuable information, also from the perspective of replacing animal models. However, bioassays relying on 3D cell models are generally highly demanding in terms of facilities, equipment, and skilled personnel requirements. To reduce cost, increase sustainability, and provide a flexible 3D cell-based platform for bioassays, we here report a novel approach based on a 3D-printed microtissue device. To assess the suitability of this strategy for reporter gene technology, we selected to monitor two molecular pathways which were of interest in several applications, hypoxia signaling and the p53 pathway. The investigation of such pathways is highly relevant in fields spanning from drug screening to bioactivity monitoring for industrial by-product valorization. Microtissues of human hepatocarcinoma (HepG2) and human embryonic kidney (Hek293T) cell lines were obtained with a low-cost and sustainable chip platform and bioassays were developed to monitor the hypoxia-inducible factors (HIFs) and the p53 tumor suppressor pathway. HepG2 and Hek293T 3D cell models were genetically engineered to express the Luc2P from Photinus pyralis firefly either under the regulation of p53 or HIF response elements. The bioassays allowed quantitative assessment of hypoxia and tumoral activity with 1,10-phenanthroline for HIF and with doxorubicin for p53 pathway activation, respectively, showing good potential for applications of this sustainable and low-cost 3D-printed microfluidic platform for bioactivity analyses, drug screening, and precision medicine.

Molecular mechanism of hypoxia and alpha-ketoglutaric acid on collagen expression in scleral fibroblasts.

To investigate the molecular mechanisms underlying the influence of hypoxia and alpha-ketoglutaric acid (α-KG) on scleral collagen expression. Meta-analysis and clinical statistics were used to prove the changes in choroidal thickness (ChT) during myopia. The establishment of a hypoxic myopia model (HYP) for rabbit scleral fibroblasts through hypoxic culture and the effects of hypoxia and α-KG on collagen expression were demonstrated by Sirius red staining. Transcriptome analysis was used to verify the genes and pathways that hypoxia and α-KG affect collagen expression. Finally, real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was used for reverse verification. Meta-analysis results aligned with clinical statistics, revealing a thinning of ChT, leading to scleral hypoxia. Sirius red staining indicated lower collagen expression in the HYP group and higher collagen expression in the HYP+α-KG group, showed that hypoxia reduced collagen expression in scleral fibroblasts, while α-KG can elevated collagen expression under HYP conditions. Transcriptome analysis unveiled the related genes and signaling pathways of hypoxia and α-KG affect scleral collagen expression and the results were verified by RT-qPCR. The potential molecular mechanisms through which hypoxia and α-KG influencing myopia is unraveled and three novel genes TLCD4, TBC1D4, and EPHX3 are identified. These findings provide a new perspective on the prevention and treatment of myopia via regulating collagen expression.

Gene expression profiles in placenta and their association with anesthesia, delivery mode and maternal diabetes

IntroductionFetal development is dependent on placenta and affected by multiple factors including maternal diabetes. Here we aimed to identify maternal diabetes-associated changes in placentas and analyzed placental gene expression to understand its modulation by maternal diabetes and birth mode. MethodsPlacental RNAseq transcriptome analyses were performed on maternally-derived decidua and fetal-derived villous tissue from pregnancies of mothers with type 1 diabetes (n = 14), gestational diabetes (n = 6) and without diabetes (n = 14). Information on delivery mode and anesthesia were included as covariables. Analyses were performed separately for decidua and fetal tissues and adjusted for sex. ResultsSubstantial placenta gene expression variation was associated with factors other than maternal diabetes, including site, sex, anesthesia type and delivery mode. Two dominant gene expression clusters aligned to anesthesia and delivery mode were observed for decidua and villous tissue. Upregulation of genes within pathways related to organ morphogenesis and downregulation of immune response to steroid- and hypoxia pathway genes was characteristic of placentas from primary cesarean section deliveries with spinal anesthesia. Opposite profiles were observed for placentas from secondary cesarean and epidural anesthesia deliveries. Placentas from vaginal delivery had intermediate gene expression profiles. More subtle changes were associated with maternal diabetes: upregulation of ribosome activity, down-regulation of maternally-derived decidua chemokine signaling pathways and for gestational diabetes, alteration in hypoxia response genes. DiscussionThe findings reveal suppression of immune pathways and upregulation of ribosome activity in the placenta by maternal diabetes highlighting the importance of confounding factors when examining cell and tissue expression profiles. Further studies should determine whether the observed gene expression differences are related to underlying causes for cesarean section deliveries.

Role of Sodium-Dependent Vitamin C Transporter-2 and Ascorbate in Regulating the Hypoxic Pathway in Cultured Glioblastoma Cells.

The most common and aggressive brain cancer, glioblastoma, is characterized by hypoxia and poor survival. The pro-tumour transcription factor, hypoxia-inducible factor (HIF), is regulated via HIF-hydroxylases that require ascorbate as cofactor. Decreased HIF-hydroxylase activity triggers the hypoxic pathway driving cancer progression. Tissue ascorbate accumulates via the sodium-dependent vitamin C transporter-2 (SVCT2). We hypothesize that glioblastoma cells rely on SVCT2 for ascorbate accumulation, and that knockout of this transporter would disrupt the regulation of the hypoxic pathway by ascorbate. Ascorbate uptake was measured in glioblastoma cell lines (U87MG, U251MG, T98G) by high-performance liquid chromatography. CRISPR/Cas9 was used to knockout SVCT2. Cells were treated with cobalt chloride, desferrioxamine or 5% oxygen, with/without ascorbate, and key hypoxic pathway proteins were measured using Western blot analysis. Ascorbate uptake was cell line dependent, ranging from 1.7 to 11.0 nmol/106 cells. SVCT2-knockout cells accumulated 90%-95% less intracellular ascorbate than parental cells. The hypoxic pathway was induced by all three stimuli, and ascorbate reduced this induction. In the SVCT2-knockout cells, ascorbate had limited effect on the hypoxic pathway. This study verifies that intracellular ascorbate is required to suppress the hypoxic pathway. As patient survival is related to an activated hypoxic pathway, increasing intra-tumoral ascorbate may be of clinical interest.

Deletion of Transmembrane protein 184b leads to retina degeneration in mice.

Transmembrane protein 184b (Tmem184b) has been implicated in axon degeneration and neuromuscular junction dysfunction. Notably, Tmem184b exhibits high expression levels in the retina; however, its specific function within this tissue remains poorly understood. To elucidate the role of Tmem184b in the mammalian visual system, we developed a Tmem184b knockout (KO) model for further investigation. Loss of Tmem184b led to significant decreases in both a and b wave amplitudes of scotopic electroretinogram (ERG) and reduced b wave amplitudes of photopic ERG, respectively, reflecting damage to both the photoreceptors and secondary neuronal cells of the retina. Histologic analyses showed a progressive retinal thinning accompanied by the significantly loss of retinal cells including cone, rod, bipolar, horizontal and retinal ganglion cells. The expression levels of photo-transduction-related proteins were down-regulated in KO retina. TUNEL (terminal deoxynucleotidyl transferase-mediated biotinylated Uridine-5'-triphosphate [UTP]nick end labelling) and glial fibrillary acidic protein (GFAP)-labelling results suggested the increased cell death and inflammation in the KO mice. RNA-sequencing analysis and GO enrichment analysis revealed that Tmem184b deletion resulted in down-regulated genes involved in various biological processes such as visual perception, response to hypoxia, regulation of transmembrane transporter activity. Taken together, our study revealed essential roles of Tmem184b in the mammalian retina and confirmed the underlying mechanisms including cell death, inflammation and hypoxia pathway in the absence of Tmem184b, providing a potential target for therapeutic and diagnostic development.

Sex differences in the molecular profile of adult diffuse glioma are shaped by IDH status and tumor microenvironment.

Sex differences in adult diffuse glioma (ADG) are well-established clinically, yet the underlying molecular mechanisms remain inadequately understood. Here, we aim to reveal molecular features and cellular compositions unique to each sex in ADG to comprehend the role of sex in disease etiology. We quantified sex differences in transcriptome of ADG using multiple independent glioma patient datasets. Next, we delved into the single-cell landscape to examine sex differences in gene expression and cellular composition. To explore how sex influences disease progression, we analyzed paired samples from primary and recurrent ADG cases, aiming to identify sex-specific differences in molecular and cellular features. Our analysis revealed that mutations in isocitrate dehydrogenase (IDH) genes and the tumor microenvironment emerged as primary influencers of sex-differential molecular enrichments. In IDHwt tumors, genes in neuronal signaling pathway are found to be enriched in male tumors, while genes in hypoxia and inflammatory response pathways are enriched in female tumors. This pattern was reversed in IDHmut gliomas. We hypothesized that these distinctions could be attributed to heterogeneous cellular composition between sexes. Using single-cell data, we observed distinctive patterns of sex differences in cell states, cell composition and cell-cell interaction in IDHwt and IDHmut tumors separately. Further, by comparing molecular changes in paired primary and recurrent ADG samples, we identified sex-specific differences in molecular characteristics and cellular compositions of recurrent tumors. Our results provide a comprehensive multi-level characterization of sex differences in ADG, such findings provide novel insights into glioma disease progression in each sex.

7068 Paraganglioma in Cyanotic Congenital Heart Disease

Abstract Disclosure: R. Lis: None. V. Hamidi: None. Paragangliomas arise from the non-chromaffin cells of the parasympathetic nervous system, which are composed of glomus cells. These function as chemoreceptors in response to hypoxia. Carotid body paragangliomas have been linked to conditions with chronic hypoxia. This was first seen in residents of high altitude locations in a case report presented by Saldana et. al and regression of a paraganglioma with resolution of hypoxia was reported by Gruber et. al. There have been few additional case reports of Paraganglioma in other hypoxic states. We present a rare case of a functional Paraganglioma in a patient with Cyanotic Congenital Heart Disease (CCHD).This is a 19 year old female with a history of CCHD due to double inlet left ventricle and transposition of the great arteries. Soon after birth, she had a fontan procedure. She subsequently developed fontan associated liver disease as well as chronic kidney disease, hypertension, sinus node dysfunction, and complete heart block. Her baseline oxygen saturation was 78-85%. Her heart rate was 40-49 beats per minute with upcoming plans for pacemaker placement. Her blood pressure ranged 90/50 to 150/80 mmHg on Amlodipine 5 mg and Lisinopril 2.5 mg. She routinely received abdominal imaging to monitor her liver disease. A liver MRI incidentally revealed a retroperitoneal mass concerning for a paraganglioma. Serum Normetanephrines were found to be 26.60 nmol/L (reference range less than 0.89). 24 hr Urine Norepinephrine was 1,187 ug/d (reference range 14-20). Further imaging was negative for additional sites of disease. Given her complete heart block, she was started on alpha blockade only prior to pacemaker placement. After the pacemaker was placed, she was started on beta blockade and underwent surgical resection of her paraganglioma. The pathology confirmed paraganglioma and mutation testing was negative for germline mutations including SDH, VHL, RET, NF1 mutations. Paragangliomas in patients with CCHD have been scarcely described in the literature given the rarity of both of these conditions. However, there is a relatively increased incidence of paragangliomas in the CCHD population due to chronic hypoxia triggering the hypoxia pathway. EPAS1 mutated HIF2alpha is often found to be the culprit in these patients. In addition, case reports have often described these patients to either be asymptomatic or have potential symptoms masked by their cardiac disease. Therefore, CCHD patients are at increased risk for paragangliomas and warrant additional awareness and regular screening. Screening has been proposed to start after 10 years of hypoxia in a case review by Agarwal et. al. as paraganglioma incidence in CCHD populations has not been seen before age 11. Incidence of recurrence is not yet widely studied, but significant concern for recurrence is warranted if the etiology of hypoxia is not reversed. Presentation: 6/3/2024

In-depth proteomics and Phosphoproteomics reveal biomarkers and molecular pathways of chronic intermittent hypoxia in mice

Obstructive sleep apnea (OSA) syndrome is characterized by Chronic Intermittent Hypoxia (CIH). In this study, we employed Data-independent acquisition (DIA) Mass Spectrometry to conduct comprehensive proteomic and phosphoproteomic profiling of a murine model subjected to Chronic Intermittent Hypoxia (CIH), a model we had previously established. Utilizing three CIH and three normal control genioglossus samples, we gathered valuable insights into the molecular alterations associated with CIH. Our analyses identified a total of 4576 protein groups and 13,867 phosphosites. Differential analysis of the proteomic data highlighted a significant upregulation of Ras signaling (Egf, Ngf, and Fyb1) and calcium signaling (Tnn, Thbs4, and Ppp2r2d) in CIH samples, contrasting with a notable decrease in oxidative phosphorylation (Atp5mf, Atp5me, and Atp5mg). Additionally, we observed a substantial increase in the phosphorylation of PI3K-AKT signaling (Ptk2_Y861, Mapk3_T203, and Eif4b_S230) and HIF-1 signaling (Gapdh_S208, Eno3_T229, and Camk2b_T382) in CIH samples. These findings prompted a deeper investigation into the association of the characterized proteins and phosphoproteins with Obstructive Sleep Apnea (OSA). The comprehensive profiling revealed molecular signatures that may serve as valuable insights into the pathophysiology of chronic intermittent hypoxia and its link to obstructive sleep apnea. Our observations provide a foundation for future research endeavors, offering potential avenues for advancing our understanding and treatment strategies for these conditions. SignificanceThe significance of this study lies in its comprehensive exploration of the molecular mechanisms underpinning Chronic Intermittent Hypoxia (CIH), a key feature of Obstructive Sleep Apnea (OSA). By employing Data-independent acquisition (DIA) Mass Spectrometry, this research provides an in-depth proteomic and phosphoproteomic analysis, uncovering critical signaling pathways and molecular alterations associated with CIH. The identification of significant changes in Ras and calcium signaling pathways, along with increased phosphorylation in PI3K-AKT and HIF-1 signaling, offers novel insights into the pathophysiological processes involved in CIH and OSA. These findings not only enhance our understanding of the molecular basis of OSA but also pave the way for the development of targeted therapeutic strategies, ultimately contributing to better management and treatment of OSA and related conditions.

PYGL regulation of glycolysis and apoptosis in glioma cells under hypoxic conditions via HIF1α-dependent mechanisms.

Gliomas are highly aggressive brain tumors with complex metabolic and molecular alterations. The role of glycolysis in glioma progression and its regulation by hypoxia remain poorly understood. This study investigated the function of glycogen phosphorylase L (PYGL) in glioma and its interaction with glycolytic pathways under hypoxic conditions. Differential expression analysis was conducted using The Cancer Genome Atlas (TCGA) glioma and GSE67089 datasets, revealing significant changes in the expression of genes. A prognostic risk model incorporating PYGL was built by univariate and multivariate Cox regression analyses. The impacts of PYGL on glioma cell proliferation, glycolysis, apoptosis, and metabolic activities were evaluated by in vitro assays. Additionally, the influences of hypoxia and hypoxia-inducible factor 1-alpha (HIF1α) on PYGL expression were evaluated. Our prognostic prediction model showed a C-index of 0.76 [95% confidence interval (CI): 0.70-0.82], indicating a good predictive accuracy of the model. In addition, genetic predictors included in the nomogram included PYGL, HIF1α, and other genes associated with the glycolytic pathway. Differential expression analysis identified PYGL as a key gene associated with glioma survival. PYGL expression was significantly upregulated in glioma cells. PYGL knockdown inhibited cell invasion, proliferation, migration, and colony formation and enhanced apoptosis via modulation of Bcl-2, caspase-3, and Bax. Glycolysis was impaired in PYGL-knockdown cells, as indicated by increased glycogen levels and a reduced extracellular acidification rate (ECAR), adenosine triphosphate (ATP) levels, lactate levels, and PKM2 and LDHA expression. PYGL overexpression promoted glycolysis and cell viability, which was counteracted by 2-deoxy-D-glucose (2-DG). Hypoxia-induced PYGL expression was regulated by HIF1α, underscoring the interplay between the hypoxia and glycolysis pathways. PYGL is a crucial regulator of glycolysis in gliomas and contributes to tumor progression under hypoxic conditions. Targeting PYGL and its associated metabolic pathways may offer new therapeutic strategies for glioma treatment.

Inflammatory Status in Deceased Kidney Donors Led by Impaired Mitochondrial Metabolism through the HIF1α Hypoxia Pathway

Inflammatory Status in Deceased Kidney Donors Led by Impaired Mitochondrial Metabolism through the HIF1α Hypoxia Pathway

Hypoxia Pathways in Parkinson’s Disease: From Pathogenesis to Therapeutic Targets

The human brain is highly dependent on oxygen, utilizing approximately 20% of the body’s oxygen at rest. Oxygen deprivation to the brain can lead to loss of consciousness within seconds and death within minutes. Recent studies have identified regions of the brain with spontaneous episodic hypoxia, referred to as “hypoxic pockets”. Hypoxia can also result from impaired blood flow due to conditions such as heart disease, blood clots, stroke, or hemorrhage, as well as from reduced oxygen intake or excessive oxygen consumption caused by factors like low ambient oxygen, pulmonary diseases, infections, inflammation, and cancer. Severe hypoxia in the brain can manifest symptoms similar to Parkinson’s disease (PD), including cerebral edema, mood disturbances, and cognitive impairments. Additionally, the development of PD appears to be closely associated with hypoxia and hypoxic pathways. This review seeks to investigate the molecular interactions between hypoxia and PD, emphasizing the pathological role of hypoxic pathways in PD and exploring their potential as therapeutic targets.

Unveiling the impact of estrogen exposure on ovarian cancer: a comprehensive risk model and immune landscape analysis

This study examines the impact of estrogenic compounds like bisphenol A (BPA), estradiol (E2), and zearalenone (ZEA) on human ovarian cancer, focusing on constructing a risk model, conducting gene set variation analysis (GSVA), and evaluating immune infiltration. Differential gene expression analysis identified 980 shared differentially expressed genes (DEGs) in human ovarian cells exposed to BPA, E2, and ZEA, indicating disruptions in ribosome biogenesis and RNA processing. Using the cancer genome atlas ovarian cancer (TCGA-OV) dataset, a least absolute shrinkage and selection operator (LASSO)-based risk model was developed incorporating prognostic genes 4-hydroxyphenylpyruvate dioxygenase like (HPDL), Thy-1 cell surface antigen (THY1), and peptidase inhibitor 3 (PI3). This model effectively stratified ovarian cancer patients into high-risk and low-risk categories, showing significant differences in overall survival, disease-specific survival, and progression-free survival. GSVA analysis linked HPDL expression to pathways related to the cell cycle, DNA damage, and repair, while THY1 and PI3 were associated with apoptosis, hypoxia, and proliferation pathways. Immune infiltration analysis revealed distinct immune cell profiles for high and low-expression groups of HPDL, THY1, and PI3, indicating their influence on the tumor microenvironment. The findings demonstrate that estrogenic compounds significantly alter gene expression and oncogenic pathways in ovarian cancer. The risk model integrating HPDL, THY1, and PI3 offers a strong prognostic tool, with GSVA and immune infiltration analyses providing insights into the interplay between these genes and the tumor microenvironment, suggesting potential targets for personalized therapies.

Missense mutations in the CITED2 gene may contribute to congenital heart disease

BackgroundCongenital heart disease (CHD) is a lifelong abnormality present from birth. Multiple studies have shown that mutations in genes involved in heart development could cause congenital heart disease. The CITED2 gene works as a transcription factor in the hypoxic pathway for the development of the heart. Therefore, five CHD types, ventricular septal defect, atrial septal defect, atrioventricular septal defect, tetralogy of fallot, and patent ductus arteriosus, were evaluated by conducting a targeted single nucleotide polymorphism (SNP) analysis of the CITED2 gene variant rs375393125 (T > C). This study aimed to identify the association of CITED2 gene mutations in CHD patients.MethodsThree hundred fifty samples, 250 from patients and 100 from controls, were collected for this genetic analysis. Allele-specific PCR and gel electrophoresis were used to identify the target missense mutations. The genotypic results of the CHDs were further validated through Sanger sequencing.ResultsThe frequency of the homozygous mutant (CC) in CHD patients was 48.4%, and of the heterozygous mutant (TC) genotype was 11.4%; these percentages are higher than controls (1%). The control samples had only one heterozygous TC and no homozygous CC genotype. The chi-square value was obtained at 103.9 with a probability of 0.05, more significant than the significance value of 21.03. The odds ratio was 43.7, which is > 1. The calculated value of ANOVA was 11.6, which was more significant than the F critical value of 3.7. As a result of sequencing, the mutant sample of each selected CHD type was found heterozygous or homozygous, and the results were like those obtained through conventional PCR.ConclusionThe samples of CHD patients showed mutations. Therefore, the CITED2 gene SNP might be associated with CHD.

Single-cell transcriptomics reveal distinctive patterns of fibroblast activation in heart failure with preserved ejection fraction

AbstractInflammation, fibrosis and metabolic stress critically promote heart failure with preserved ejection fraction (HFpEF). Exposure to high-fat diet and nitric oxide synthase inhibitor N[w]-nitro-l-arginine methyl ester (L-NAME) recapitulate features of HFpEF in mice. To identify disease-specific traits during adverse remodeling, we profiled interstitial cells in early murine HFpEF using single-cell RNAseq (scRNAseq). Diastolic dysfunction and perivascular fibrosis were accompanied by an activation of cardiac fibroblast and macrophage subsets. Integration of fibroblasts from HFpEF with two murine models for heart failure with reduced ejection fraction (HFrEF) identified a catalog of conserved fibroblast phenotypes across mouse models. Moreover, HFpEF-specific characteristics included induced metabolic, hypoxic and inflammatory transcription factors and pathways, including enhanced expression of Angiopoietin-like 4 (Angptl4) next to basement membrane compounds, such as collagen IV (Col4a1). Fibroblast activation was further dissected into transcriptional and compositional shifts and thereby highly responsive cell states for each HF model were identified. In contrast to HFrEF, where myofibroblast and matrifibrocyte activation were crucial features, we found that these cell states played a subsidiary role in early HFpEF. These disease-specific fibroblast signatures were corroborated in human myocardial bulk transcriptomes. Furthermore, we identified a potential cross-talk between macrophages and fibroblasts via SPP1 and TNFɑ with estimated fibroblast target genes including Col4a1 and Angptl4. Treatment with recombinant ANGPTL4 ameliorated the murine HFpEF phenotype and diastolic dysfunction by reducing collagen IV deposition from fibroblasts in vivo and in vitro. In line, ANGPTL4, was elevated in plasma samples of HFpEF patients and particularly high levels associated with a preserved global-longitudinal strain. Taken together, our study provides a comprehensive characterization of molecular fibroblast activation patterns in murine HFpEF, as well as the identification of Angiopoietin-like 4 as central mechanistic regulator with protective effects.