HIF-1alpha Gene Research Articles

The Effects of IGF1 and MGF on Neural Stem Cells in Hypoxic Conditions.

Ischemic stroke has high morbidity and mortality rates worldwide. Low oxygen (O2) levels detected in such conditions create a vulnerable environment for neural stem cells (NSC), altering neuronal function, and leading to neuronal injury or death. There are still no effective treatments for such consequences. This study investigates the molecular and functional effects of growth factors, namely, insulin-like growth factor 1 (IGF-I) and mechano growth factor (MGF), in NSC exposed to low O2 levels. An in vitro ischemia model was created by rat hippocampal NSC grown in culture that was exposed to varying oxygen levels, including 0%, 3%, and 20 % for the representation of anoxic, hypoxic, and normoxic conditions, respectively, during 24 h. NSC has investigated IGF-I, MGF, and HIF1-Alpha (HIF-1α) gene expressions by real-time reverse transcription polymerase chain reaction. The effects of external administration of growth factors (IGF-I and MGF) on NSC proliferation in such conditions were explored. Increased IGF-I and MGF gene expressions were detected in the samples exposed to low O2. Anoxia was the highest stimulant for IGF-I and MGF gene expressions. Meanwhile, HIF1-α that encodes hypoxia-inducible factor-1α revealed downregulation in relative gene expression fold change with IGF-I application in all conditions, whereas MGF application upregulated its change in an anoxic environment. Furthermore, MGF-induced NSC had more proliferationmigration rate in all oxygen conditions. IGF-I induced significant NSC proliferation in 0% and 20% O2. These findings suggest that IGF-I and MGF expressions were increased to reduce the damage in NSC exposed to low oxygen, and exogenous MGF and IGF-I application increased NSC proliferation at the time of injury. The results might imply the role of exogenous MGF and IGF-I in the treatment of ischemia for relieving the effect of neuronal damage due to their neuroprotective and proliferative effects.

The role of gene expression hypoxia-inducible factor-1a and serum level monocyte chemoattractant protein-1 in the incidence of chronic myeloid leukemia in Iraqi patients

Abstract: BACKGROUND: Chronic myeloid leukemia (CML) is a hematopoietic stem cell malignancy described by a translocation between chromosomes 9 and 22. There are many factors genetic or nongenetic effect on disease progression such as growth factors and transcription factors act as oncogenes or tumor suppressor genes. OBJECTIVES: The purpose of this research was to investigate the role of hypoxia-inducible factor (HIF1A) gene expression with CML, as well as the role of monocyte chemoattractant protein-1 (MCP-1) as a predictive biomarker on disease progression. MATERIALS AND METHODS: The current study consists of three groups: first group includes 50 newly diagnosed CML patients (females 22 and males 28), second group consists of 50 CML patients treated with tyrosine kinase inhibitor (TKI) with a complete molecular response (p210 BCR-ABL transcript levels ≤0.1% IS) (female 25 and male 25), and third group included another 50 apparently healthy volunteers (female 20 and male 30). The patients were admitted from the National Center of Hematology/Mustansiriyah University. All patients are diagnosed according to a complete blood count (CBC), a bone marrow examination, and a BCR-ABL gene test. RESULTS: Reverse transcription-quantitative polymerase chain reaction was applied to assess the expression levels of the HIF-1A gene and serum level of MCP1 by enzyme-linked immunosorbent assay. The results displayed downregulated of the HIF1A gene messenger RNA in CML patients in comparison to the controls group, as well as no statistically significant link was discovered when the fold of expression was correlated with the age and gender of CML patients. CONCLUSION: HIF1-alpha gene has an important role in pathological pathways such as angiogenesis. According to this study, HIF1-alpha gene is not an appropriate prognostic biomarker for detecting the risk of CML as well as MCP1 is thought to be a predictor of CML progression.

Oxydative stress in type 2 diabetic patients: involvement of HIF-1 alpha AND mTOR genes expression

Biochemical and genetic mechanisms of oxidative stress (OS) developing in blood of patients with type 2 Diabetes mellitus (T2DM) were studied. Twenty patients with T2DM and 10 healthy persons participated in this study. Lipid peroxidation, the content of protein carbonyls and H2O2 production were measured in blood plasma and erythrocytes as OS biomarkers. Activity of SOD, catalase, and GPx as well as reduced glutathionе (GSH) level in plasma and erythrocytes were estimated. The gene expression of key regulators of oxygen and metabolic homeostasis (HIF-1α and mTOR) in leukocytes was studied. It was found a significant rise of TBARS and protein carbonyls content in plasma as well as of H2O2 production in erythrocytes from patients with T2DM compared to control. The diabetic patients also demonstrated an increase in the SOD and catalase activity in plasma and significantly lower GSH content and GPx activity in erythrocytes compared to the healthy participants. The established marked inhibition of mTOR gene expression and the tendency to an increase in HIF-1α gene expression in leukocytes of patients with T2DM may serve as a protective mechanism which counteracts OS developing and oxidative cell damage. Keywords: HIF-1α, mTOR, oxidative stress, type 2 diabetes mellitus

Effects of genetic factors on high altitude training performance

High altitude is considered to be 1800-6000 meters. With the decrease of atmospheric pressure at this altitude, adequate oxygenation cannot be achieved in the tissues and hypoxia develops in the circulatory system. Athletes aim to provide superior performance by training in hypoxic conditions. Varying adaptations in hypobaric hypoxia environments by geographically separated populations represent well-trained specimens that may be relevant to endurance performance. While inhabitants of the Andes show higher levels of hemoglobin and saturation than Tibetans at similar altitude, Ethiopian climbers maintain oxygen delivery despite the hemoglobin levels and saturation typical in sea level ranges. It can also be predicted a significant relationship between the angiotensin converting enzyme (ACE) genotype, which affects metabolic efficiency and performance in hypoxic environments (high altitude). One of the genes that develop at high altitude and occur in response to hypoxia is the hypoxia inducible factor 1 alpha (HIF-1α) gene encoded by the hypoxia inducible factor (HIF-1A) gene. The vascular endothelial growth factor (VEGF-A) gene, which is another gene with angiogenetic factor produced in response to hypoxia, is revealed by the transcription of the HIF-1 alpha gene. Genetic heritage, environmental factors, and the character of exercise loads applied within the framework of lifestyle, neuromuscular development, balanced nutrition, and cultural differences that trigger athletic success may reveal individual changes or differences. Considering all these variables, monitoring and control of performance improvement and athletic achievement graph may become more predictable.

Molecular Evaluation of HIF-1α Gene Variation and Determination of Its Frequency and Association with Breast Cancer Susceptibility in Saudi Arabia.

Hypoxia-inducible factor 1 (HIF-1α) is responsible in regulating oxygen homeostasis in tissues and is a central effector of the hypoxic response besides its protein overexpression has been shown to have prognostic relevance in several cancers including breast cancer. Several reports indicated that HIF-1α gene variation C1772T (Pro582Ser) is associated with increased breast susceptibility but results remained controversial. Therefore, we performed the molecular evaluation of HIF-1α gene variation and determined its frequency and association with Breast Cancer susceptibility in Saudi Arabia. This study was conducted on histologically confirmed Breast cancer patients and gender matched healthy women. HIF-1α C1772T (Pro582Ser) genotyping was done by Amplification refractory mutation system PCR method. The HIF-1α gene genotypes were correlated with different clinicopathological characteristics of breast cancer patients. A significant difference was observed in genotype distribution of HIF-1α gene variation C1772T (Pro582Ser) between breast cancer cases and gender matched healthy controls (P=0.010). Our findings showed that the HIF- 1α variant was associated with an increased risk of Breast cancer for HIF-1α CC vs CT genotype OR = 2.20, 95% CI = (1.28 -3.77), P = 0.004) in codominant inheritance model. The significant association was reported for HIF1A for genotypes CC vs (CT+ TT) OR = 1.98, 95% CI = (1.17-3.34), P = 0.010) in dominant inheritance model tested. In case of recessive inheritance model, a non-significant association of HIF-1 alpha gene variants was reported for (CC+ CT) vs TT) OR = 1.03, 95% CI = (0. 064-16.79), P = 0.97). During the allelic comparison, a non-significant association was reported between A vs C allele among Breast cancer patients. A significant association of HIF- 1α polymorphism was reported with stage as well as distant metastasis of the disease. A significant difference was observed in genotype distribution of HIF-1α gene variation C1772T (Pro>Ser) between breast cancer cases and gene matched healthy controls (P=0.010). HIF-1α- CT heterozygosity and CC genotype increased the susceptibility .The HIF-1α polymorphism was reported to be significantly associated with the distant metastasis of Breast cancer. Further studies with larger data set and well-designed models are required to validate our findings.

HYPOXIC PRECONDITIONING AMELIORATES ISCHEMIA-INDUCED DAMAGE OF ISOLATED RAT ISLETS

Aim: Islet isolation results in disruption of the native islet vascular network until posttransplant islet revascularisation is completed. Prolonged exposure to a hypoxic and nutrient-depleted environment has detrimental effects on morphological and functional integrity of isolated islets. In solid organ transplantation it was shown that short sublethal periods of anoxia can induce adaptive mechanisms in the affected tissue offering protection against subsequent potentially lethal hypoxic insults. This study was aimed to explore the potential of hypoxic preconditioning for isolated islets. Methods: Rat islets, isolated from 20 non-heartbeating donors, were subjected either to 2 x 2 min-periods of anoxia, intermitted by a 2 min-period of oxygen supply, or to sham-treatment. Hypoxic preconditioning and sham-treatment were followed by 4 hours of recovery in normoxia prior to 24 hr-culture at 1.5% oxygen. Islet characterisation included yield of islet equivalents (IEQ) or islet particle number (IN), viability (FDA-PI), DNA (pico-green) and ATP (luciferase assay) content, intraislet reactive oxygen species (ROS) production (DCFH-DA), gene expression (qRT-PCR), and glucose stimulation index (2 vs 20 vs 2 mM). Data were normalised to islet DNA content and are presented as mean ± SEM. Results: Hypoxic preconditioning had no effect on islet survival when compared with sham-treatment (64 ± 7 vs 61 ± 7%). Compared with preconditioned islets, sham-treated controls were characterised by increased fragmentation when determined as IN/IEQ ratio (1.51 ± 0.18 vs 1.24 ± 0.13, p<0.05). Fragmentation corrrelated inversely with viability of sham-treated and preconditioned islets (61 ± 3 vs 66 ± 3 %, p<0.01). ATP (0.66 ± 0.14 vs 0.73 ± 0.20 pM/ng DNA) and insulin content (51.1 ± 12.5 vs 46.3 ± 11.7 ng/ng) were not altered by hypoxic preconditioning compared with sham-treatment. ROS production was reduced after hypoxic preconditioning compared with sham-treatment (645 ± 73 vs 858 ± 88 AU/ng, p<0.01). Islets of both experimental groups responded with insulin release after glucose challenge, but the stimulation index of preconditioned islets was significantly larger compared with sham-treated islets (2.39 ± 0.37 vs 2.09 ± 0.33, p<0.05). Whilst pro-apoptotic BAX and CHOP gene expression was reduced by 54 ± 15% (p<0.05) and 49 ± 8% (p<0.05), hypoxic preconditioning increased HIF-1 alpha and VEGF-A gene expression by 90 ± 41% (p<0.01) and 184 ± 65% (p<0.05), respectively. Conclusion: Although hypoxic preconditioning could not ameliorate ischaemia-induced loss of islets, islet morphology and function was significantly improved presumably as result of reduced intraislet ROS production. qRT-PCR-determined gene expression suggests that hypoxic preconditioning induces rescue mechanisms islets and mediates a significant anti-apoptotic effect. This study describes a new option for pretransplant treatment of isolated human islets. European Union’s Cooperation Programme FP7 (HEALTH-F2-2012-305746). Oxford NIHR Biomedical Research Centre.

PTP4A3, A Novel Target Gene of HIF-1alpha, Participates in Benzene-Induced Cell Proliferation Inhibition and Apoptosis through PI3K/AKT Pathway.

Benzene, a commonly used chemical, has been confirmed to specifically affect the hematopoietic system as well as overall human health. PTP4A3 is overexpressed in leukemia cells and is related to cell proliferation. We previously found that HIF-1alpha was involved in benzene toxicity and PTP4A3 may be the target gene of HIF-1alpha via ChIP-seq. The aim of this study is to confirm the relationship between HIF-1alpha and PTP4A3 in benzene toxicity, as well as the function of PTP4A3 on cell toxicity induced by 1,4-benzoquinone (1,4-BQ). Our results indicate that HIF-1alpha could regulate PTP4A3 with in vivo and in vitro experiments. A cell line with suppressed PTP4A3 was established to investigate the function of PTP4A3 in 1,4-BQ toxicity in vitro. The results revealed that cell proliferation inhibition was more aggravated in PTP4A3 low-expression cells than in the control cells after 1,4-BQ treatment. The relative oxygen species (ROS) significantly increased in cells with inhibited PTP4A3, while the rise was inferior to the control cells at the 20 μM 1,4-BQ group. An increase in DNA damage was seen in PTP4A3 down-regulated cells at the 10 μM 1,4-BQ group, whereas the results reversed at the concentration of 20 μM. Moreover, the apoptosis rate increased higher in down-regulated PTP4A3 cells after 1,4-BQ exposure. In addition, PI3K/AKT pathway was significantly restrained in cells with inhibited PTP4A3 after 1,4-BQ treatment. Our results indicate that HIF-1alpha may regulate PTP4A3 to be involved in benzene toxicity. Inhibition of PTP4A3 could aggravate cell proliferation suppression and apoptosis by regulating PI3K/AKT pathway after 1,4-BQ treatment.

Abstract B25: Meformin and LY294002 treatment regulate angiogenesis process through PI3K-MAPK/AKT/mTOR signaling pathways in mammary tumors

Abstract Introduction: The angiogenesis process is regulated by numerous factors, especially the hypoxia-inducible factor 1alpha (HIF-1 alpha) and vascular endothelial growth factor (VEGF). The HIF-1 is a central regulator of pathophysiologic response of cells to hypoxia conditions, able to activate transcription of the gene that promotes the induction of VEGF, which in turn promotes angiogenesis through its ability to stimulate growth, migration, and invasion of endothelial cells, leading to the formation of new blood vessels and subsequent tumor growth. Metformin has demonstrated its ability to reduce the incidence of cancer in diabetic patients with positive results, especially in breast cancer, inhibiting cell growth by AMPK/Akt/mTOR signaling pathway. Likewise, LY294002, a PI3K signaling pathway inhibitor, has antiangiogenic features. In this context, the aim of this study was to evaluate the effectiveness of metformin and LY294002 inhibitor treatment in angiogenesis as a therapeutic strategy in mammary tumors in an in vitro and in vivo study. Materials and Methods: In the in vitro study, canine mammary tumor cell line (CF41) was cultured in DMEM high-glucose culture medium, at 37°C in 5% CO2. Cell viability was measured by MTT assay after treatment with different concentrations of metformin and LY294002 inhibitor. Once established the concentration of 5 mM for metformin and 5 µM of LY294002, the protein and gene expression of HIF-1alpha and VEGF were detected by immunocytochemistry and real-time PCR, respectively. For an in vivo study, CF41 were injected in nude athymic female mice and treated with metformin (200 mg/kg i.p.) for 4 weeks and LY294002 (7.5 mg/kg intratumorally) every 3 days for three times. At the end, mice were euthanized and the tumors were collected to determine the protein and gene expression of HIF-1alpha and VEGF and microvessel density by immunohistochemistry for CD31. The immunostaining was quantified by optical densitometry technique, slides were analyzed and photographed in Nikon Eclipse E200 microscope at 40X objective, and proteins were quantified by the ImageJ software analysis. For real-time PCR, the relative expression of the genes of interest was determined by DataAssist v3.0 software, using Ct method. Results: There was a significant decrease of cell viability after treatment with different concentrations of metformin and LY294002 in 24 hours. Both HIF-1alpha and VEGFA protein and gene expression significantly decreased after treatment with metformin and LY294002 or in combination. Furthermore, in animals, there was a decrease in tumor size, protein and gene expression of HIF-1alpha and VEGFA, in addition to a decrease of CD31 expression after all treatments. Conclusion: Our results suggest the potential effectiveness of metformin and LY294002 acting in angiogenesis process of mammary tumors. Financial support: FAPESP. Citation Format: Marina Gobbe Moschetta, Camila Leonel, Larissa Bazela Maschio-Signorini, Thaiz Ferraz Borin, Gabriela Bottaro Gelaleti, Lívia Carvalho Ferreira, Bruna Victorasso Jardim-Perassi, Nathália Martins Sonehara, Debora Aparecida Pires de Campos Zuccari. Meformin and LY294002 treatment regulate angiogenesis process through PI3K-MAPK/AKT/mTOR signaling pathways in mammary tumors [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B25.

Acute hypoxic stress: Effect on blood parameters, antioxidant enzymes, and expression of HIF-1alpha and GLUT-1 genes in largemouth bass (Micropterus salmoides)

Dissolved oxygen (DO) plays a crucial role in survival, growth, and normal physiological functions of aquatic organisms. Nevertheless, the mechanisms involved in hypoxic stress and adaptation have not been fully elucidated in Largemouth bass (Micropterus salmoides). To reveal the effect of acute hypoxia on Largemouth bass, we simulated acute hypoxia (DO: 1.2 ± 0.2 mg/L) in the laboratory and analyzed physiological parameters (RBCs, Hb, SOD, CAT, NA+/K+-ATPase, GPx, and MDA) and gene expression (HIF-1alpha and GLUT-1) in Largemouth bass exposed to various durations of acute hypoxia (0, 1, 2, 4, 8, 12, and 24 h). Our results indicated that acute hypoxic exposure significantly increased RBCs but decreased Hb. In addition, antioxidant enzyme activity was enhanced significantly in the liver and muscles at the initial stage of acute hypoxic exposure, but decreased significantly in gills during the entire process of hypoxic exposure. Furthermore, the expression levels of HIF-1alpha and GLUT-1 mRNA were significantly up-regulated in Largemouth bass under acute hypoxic exposure. In conclusion, our study provides a valuable basis for further elucidation of hypoxic adaptation and facilitates husbandry for an economically valuable species.

Construction of recombinant adenovirus expressing combined human hypoxia inducible factor-1α and LIM mineralization protein-1 and its osteogenic effect

Objective This study was designed to construct a recombinant adenovirus vector expressing both hypoxia inducible factor-1α (HIF-1α) and LIM mineralization protein-1 (LMP-1) gene linked by internal ribozyme entry site (IRES), and to detect the osteogenesis in rat bone marrow stromal stem cells. Methods The full-length of human HIF-1α (2 481 bp) and LMP-1 (1 374 bp) genes and IRES (564 bp) sequence were acquired by polymerase chain reaction (PCR) and inserted into pShuttle-IRES-hrGFP-1 to construct recombinant adenovirus shuttle plasmid pS-HIF-LMP-1-GFP. Recombinant adenovirus vector pAd-HIF-LMP-1-GFP was obtained from homologous recombination in BJ5183 cells by pS-HIF-LMP-1-GFP and pAdEasy-1. Recombinant adenovirus Ad-HIF-LMP-1-GFP was obtained by packaging in AD293 cells. Ad-HIF-LMP-1-GFP was transduced to the rat bone marrow mesenchymal stem cells (BMSCs). The expression of HIF-1α and LMP-1 in BMSCs was observed by reverse transcription-polymerase chain reaction (RT-PCR). The alkaline phosphatase (ALP) activity, calcium formation, osteocalcin and related transcription factor-2 (Runx2) expression changes between experimental group and control group were detected to evaluate the osteogenitic capacity of combined HIF-1α and LMP-1. Results The recombinant adenovirus expressing combined HIF-1α and LMP-1 was successfully acquired. The expression of HIF-1α and LMP-1 was detected successfully in BMSCs. The ALP activity increased significantly (P=0.000). ALP activity was 0.145, 0.160, 0.170, 0.167 (0.160±0.011) ng/μg protein in experimental group and that was 0.045, 0.057, 0.056, 0.054 (0.053±0.005) ng/μg protein in control group, respectively. The calcium formation increased significantly (P=0.001). The quantification was 0.636, 0.544, 0.591, 0.515 (0.570±0.050) in experimental group and that was 0.121, 0.150, 0.144, 0.167 (0.145±0.019) in control group, respectively. The expression of osteocalcin and Runx2 was enhanced significantly (P=0.017 and 0.000). The expression of osteocalcin was 2.688, 2.997, 2.777 (2.820±0.159) in experimental group and that was 2.009, 1.916, 1.944 (1.956±0.047) in control group, respectively. The expression of Runx2 was 0.440, 0.465, 0.456 (0.454±0.010) in experimental group and that was 0.096, 0.108, 0.117 (0.107±0.010) in control group, respectively. Conclusion The recombinant adenovirus vector was successfully constructed by using IRES sequence to connect human HIF-1 alpha and LMP-1 gene. Combined HIF-1α and LMP-1 gene can promote the osteoblastic differentiation of BMSCs. Key words: Hypoxia inducible factor-1α; LIM mineralization protein-1; Mesenchymal stem cells; Osteogenic activity; Gene transfection

Association Between HIF-1 Alpha Gene Polymorphisms and Response in Patients Undergoing Neoadjuvant Chemotherapy for Locally Advanced Cervical Cancer

BackgroundThe aim of the study was to assess whether HIF-1α polymorphisms have an effect on the response to chemotherapy of locally advanced cervical cancer (LACC) patients treated with platinum-based neoadjuvant chemotherapy (NACT) and radical surgery.Material/MethodsWe conducted a retrospective study in 162 LACC patients. Hypoxia-inducible factor 1α C1772T and G1790A genetic polymorphisms were ascertained using direct sequencing methods.ResultsThe C1772T polymorphism was significantly related to response to chemotherapy (P=0.002), and there was an increased chance of treatment response in patients with the C/C genotype (OR=4.7; 95% CI: 1.67–13.49; P=0.004). The C1772T polymorphism was also associated with poor tumor grade (adjusted OR, 2.98; 95% CI: 1.08–8.13; P=0.037). However, The G1790A polymorphism was not associated with response (P>0.05).ConclusionsThe C1772T polymorphism was significantly related to response to chemotherapy and poor tumor grade. Our results may help to better manage individual patients and to improve clinical decision making regarding use of NACT.

GNG Motifs Can Replace a GGG Stretch during G-Quadruplex Formation in a Context Dependent Manner.

G-quadruplexes are one of the most commonly studied non-B DNA structures. Generally, these structures are formed using a minimum of 4, three guanine tracts, with connecting loops ranging from one to seven. Recent studies have reported deviation from this general convention. One such deviation is the involvement of bulges in the guanine tracts. In this study, guanines along with bulges, also referred to as GNG motifs have been extensively studied using recently reported HOX11 breakpoint fragile region I as a model template. By strategic mutagenesis approach we show that the contribution from continuous G-tracts may be dispensible during G-quadruplex formation when such motifs are flanked by GNGs. Importantly, the positioning and number of GNG/GNGNG can also influence the formation of G-quadruplexes. Further, we assessed three genomic regions from HIF1 alpha, VEGF and SHOX gene for G-quadruplex formation using GNG motifs. We show that HIF1 alpha sequence harbouring GNG motifs can fold into intramolecular G-quadruplex. In contrast, GNG motifs in mutant VEGF sequence could not participate in structure formation, suggesting that the usage of GNG is context dependent. Importantly, we show that when two continuous stretches of guanines are flanked by two independent GNG motifs in a naturally occurring sequence (SHOX), it can fold into an intramolecular G-quadruplex. Finally, we show the specific binding of G-quadruplex binding protein, Nucleolin and G-quadruplex antibody, BG4 to SHOX G-quadruplex. Overall, our study provides novel insights into the role of GNG motifs in G-quadruplex structure formation which may have both physiological and pathological implications.

Expression and clinical significance of ING4 and HIF-1 alpha in brain astrocytoma

To study the mRNA expression level of growth inhibition factor 4 (ING4) and hypoxia inducing factor 1 alpha (HIF-1 alpha), and their relationship with tumor malignant degree or the pathology classification in human brain astrocytoma. The mRNA expression levels of ING4 and HIF-1 alpha were detected by RT-PCR method in 45 cases of grade I-IV human brain astrocytoma and 11 cases of control brain tissues from January 2009 to June 2010 in the Second Affiliated Hospital of Zhengzhou University, and their correlation was also analyzed. In the non-tumor brain tissue, the expression level of ING4 mRNA was 1.19 ± 0.22, while they were 0.91 ± 0.19, 0.74 ± 0.28, 0.54 ± 0.33 and 0.22 ± 0.19 in I-IV grade astrocytoma, respectively. Compared with the non-tumor control, the mRNA expression level of ING4 gene decreased significantly in the astrocytoma (P<0.05). And the expression of ING4 gradually reduced with the increase of the pathological classification of the astrocytoma.In the non-tumor brain tissue, the expression level of HIF-1 alpha mRNA was 0.26 ± 0.16, and they were 0.34 ± 0.19, 0.50 ± 0.23, 0.96 ± 0.15 and 1.04 ± 0.15 in I-IV grade astrocytoma, respectively.For HIF-1 alpha gene, the mRNA expression level increased significantly in the astrocytoma. Meanwhile, the expression gradually increased with the increase of the pathological classification of the astrocytoma (P<0.05). The mRNA expression showed a negative correlation between ING4 and HIF-1 alpha with the increase of the tumor malignant degree. The ING4 and HIF-1 alpha genes play a role in the tumorigenesis and development of human brain astrocytoma, and closely associate with the malignant degree of astrocytoma.

Loss of Siah2 does not impact angiogenic potential of murine endothelial cells

Angiogenesis is triggered in response to hypoxia under many circumstances, from healthy cells and tissues during embryogenesis to pathological conditions like the formation of new blood vessels to supply tumours and promote invasive cancer. Siah2 has been shown to regulate the hypoxia pathway upstream of hypoxia-induced transcription factor subunit Hif-1alpha, and therefore may play an important role in angiogenesis in response to hypoxic stress in endothelial cells. This study aims to investigate the basic function of Siah2 in endothelial cells under hypoxia and to test the ability of Siah2 deficient cells to mount an angiogenic response when deprived of oxygen. We and others have previously shown that Siah2 is crucial for mediating the hypoxic response in many different cell types studied. In this study however, we describe that Siah2−/− endothelial cells have an intact hypoxic signalling pathway, including Hif-1alpha stabilisation and gene expression, the first report of a tissue or cell lineage in which the loss of Siah2 does not seem to impact hypoxic response signalling. In mice, the infiltration of Siah2−/− endothelial cells into a Matrigel plug containing a VEGF-A attractant was similar compared with wildtype endothelial cells. Ex vivo however, there was a reduced capacity of Siah2−/− aorta to form tubes or new vessels. Thus, we conclude that Siah2 is not essential for the hypoxic response of endothelial cells.

Strong hypoxia reduces leptin synthesis in purified primary human trophoblasts

IntroductionOxygen availability severely affects placental function. During placental hypoxia, stabilization of hypoxia inducible factors (HIFs) affects transcription, and leptin gene expression concomitantly increases in vivo and in vitro. However, a causal relationship is uncertain. MethodsWe investigated the effect of oxygen availability on HIF-1 alpha (HIF1A) and leptin regulation in primary human trophoblasts isolated from six normal term placentae cultured at 0.1%, 1%, 3%, and 8% oxygen for 6 h, 24 h and 48 h. Gene expressions of leptin (LEP), leptin receptors (LEPR), HIF1A, insulin receptor (INSR) and further genes relevant in hypoxia (VEGFA, EPO, NOS2) or apoptosis (BCL2, BAX, Tp53) were examined. Leptin, HIF1A, INSR, phospho-AKT/AKT (insulin receptor signaling), caspase 3 and cleaved caspase 3 (apoptosis) proteins were measured. ResultsA hypoxic reaction with stabilization of HIF1A protein as well as up-regulation of HIF1A and VEGFA gene expressions, but without any hint for apoptosis, was present at 0.1% and 1% oxygen. However, leptin protein concentration (cell supernatants) peaked at 8% oxygen (normoxia) and was significantly reduced at 0.1% oxygen. There was no significant correlation between leptin and HIF1A, neither on the gene nor on the protein level. DiscussionElevated leptin gene expression in hypoxic placentas may not originate from trophoblasts, but from other placental cells, or from interaction of trophoblasts with other cells. Not only fetal hyperleptinemia, but also fetal hypoleptinemia under hypoxic conditions is conceivable. Strategies to prevent leptin dysregulation during pregnancy should be elucidated to protect the offspring from fetal programming of leptin resistance and adiposity in later life.

Protective effect and action mechanism of petroleum ether extracts from Saussurea involucrate on brain tissues of hypoxia rats

To investigate the protective effect and action mechanism of petroleum ether extracts from Saussurea involucrate on brain tissues of hypoxia rats under constant pressure and closed conditions. The PESI dosage-dependent experiment for hypoxia rats was conducted under constant pressure and closed conditions by intraperitoneally injecting 125, 250, 500 mg x kg(-1) to finalize that the optimum dosage is the high dose of PESI. Afterwards, 90 Wistar rats were randomly divided into the hypoxic model group, the acetazolamide 250 mg x kg(-1) group and the PESI high dose group. Each group was further divided into three subgroups according to different hypoxia times, with 10 rats in each subgroup. Under the same hypoxia and administration conditions, the rats were sacrificed after 0, 3, 6 h respectively. Their brain samples were collected for common pathological observation and immunohistochemical staining of HIF-1alpha. Real-time RT-PCR was used to detect HIF-1alpha, EPO, HO-1 and Caspase-3 gene expressions. And the Western blot assay was adopted to detect HIF-1alpha protein expression. The brain tissues of the hypoxia model group were severely damaged with the increase in the hypoxia time. The acetazolamide group and the PESI high does group were damaged in a much lower degree. According to the gene expression and the Western blot assay, high dose of PESI could inhibit HIF-1alpha expression. According to the pure gene expression test, high dose of PESI could increase EPO and HO-1 mRNA expressions, but inhibit Caspase-3 mRNA expression. PESI's protective mechanism for brain tissues of hypoxia rats under constant pressure and closed conditions may be related to its effects in inhibiting HIF-1alpha expression, increasing EPO expression and resisting cell apoptosis.

Effect of Oxygen on Cardiac Differentiation in Mouse iPS Cells: Role of Hypoxia Inducible Factor-1 and Wnt/Beta-Catenin Signaling

BackgroundDisturbances in oxygen levels have been found to impair cardiac organogenesis. It is known that stem cells and differentiating cells may respond variably to hypoxic conditions, whereby hypoxia may enhance stem cell pluripotency, while differentiation of multiple cell types can be restricted or enhanced under hypoxia. Here we examined whether HIF-1alpha modulated Wnt signaling affected differentiation of iPS cells into beating cardiomyocytes.ObjectiveWe investigated whether transient and sustained hypoxia affects differentiation of cardiomyocytes derived from murine induced pluripotent stem (iPS) cells, assessed the involvement of HIF-1alpha (hypoxia-inducible factor-1alpha) and the canonical Wnt pathway in this process.MethodsEmbryoid bodies (EBs) derived from iPS cells were differentiated into cardiomyocytes and were exposed either to 24 h normoxia or transient hypoxia followed by a further 13 days of normoxic culture.ResultsAt 14 days of differentiation, 59±2% of normoxic EBs were beating, whilst transient hypoxia abolished beating at 14 days and EBs appeared immature. Hypoxia induced a significant increase in Brachyury and islet-1 mRNA expression, together with reduced troponin C expression. Collectively, these data suggest that transient and sustained hypoxia inhibits maturation of differentiating cardiomyocytes. Compared to normoxia, hypoxia increased HIF-1alpha, Wnt target and ligand genes in EBs, as well as accumulation of HIF-1alpha and beta-catenin in nuclear protein extracts, suggesting involvement of the Wnt/beta-catenin pathway.ConclusionHypoxia impairs cardiomyocyte differentiation and activates Wnt signaling in undifferentiated iPS cells. Taken together the study suggests that oxygenation levels play a critical role in cardiomyocyte differentiation and suggest that hypoxia may play a role in early cardiogenesis.

Фазовые изменения энергетического метаболизма при периодической гипоксии

Male Wistar rats were exposed to periodic hypobaric hypoxia (PHH), by "lifting" in barochamber at "altitude" 5600 m for 1 h every 3 days (6 séances). The dynamics of changes in oxygen consumption (VO2), and body temperature (Tm), as well as in HIF-1alpha and HIF-3alpha gene expression, and mitochondrial respiration in the ventricles of the heart was studied. On the basis of the data we identified four phases of the physiological changes. The first phase, hypometabolic (1-3 séances), is characterized by decrease in VO2 and Tm, induction of HIF-1alpha and HIF-3alpha with delayed transient stimulation of metabolism in response to each séance of hypoxia. In heart mitochondria, V3 and V4 are increased, but V3/V4 and ADP/O are reduced. During the second phase, transitional (3-4 séances), there is reorganization of metabolism and decrease its hypoxic reactivity. The third phase, hypermetabolic (4-5 séances), is characterized by intensification of metabolism and compensation of hypoxic disorders. The fourth phase (after 5 séance) - is a state of metabolic adaptation with normalization of VO2 and Tm, expression of HIF-1alpha and HIF-3alpha, mitochondrial respiration, increased NAD-dependent oxidation of carbohydrate and lipid substrates. Thus, during PHH consequent rebuilding of processes of oxygen transport, tissue respiration and thermogenesis occurs, mediated by induction of the HIF subunits.

Дослідження впливу екологічних факторів Антарктики на здатність людини до адаптації

A purpose of work is a study of influencing of ecological factors of Antarctic on the physiological functions of human and development of technologies of increase of adaptation stability of organism. Antarctic influences of heliophysicals factors are studied on the organism of human. It is set that natural hertzian waves with a frequency spectrum identical of brain of human biorhythm, straight influence on electric activity as a reaction of synchronization. It is shown that the reactions of the system of circulation of blood of human in Antarctic are closely associated with the changes of barometric pressure and humidity, have hypoxical genesises and depend on polymorphism of gene of HIF-1alpha. It is shown that the study of negative influence on the organism of ecological factors in the conditions of absence of factors of technogenidc origin is instrumental in development of new technologies of prophylaxis of morbidity and saving of capacity of human as in extreme terms so in the conditions of the modem status of ecological surroundings.

Hypoxia-Inducible Factor-1α Suppressed Hepatocellular Carcinoma Cell Apoptosis Through Influencing on Omi/HtrA2 Expression and Its Releasing From the Mitochondrion

Hypoxia-inducible factor-1alpha (HIF-1alpha) plays an important role in regulating hepatoma cell apoptosis. However, conclusions of different studies about the effects of HIF-1alpha expression on hepatoma cell apoptosis remain controversial. Omi/HtrA2 promotes cell apoptosis in some human cancer cells. Our previous experiments have demonstrated that primary hepatocellular carcinoma may need Omi/HtrA2 expression for cell apoptosis. Thus, we investigated the effect of HIF-1alpha on hepatocellular carcinoma cell apoptosis and Omi/ HtrA2 expression. In our study we found that HIF-1alpha gene could suppress hepatoma cell apoptosis, and Omi/HtrA2 mRNA and protein expression decreased with HIF-1alpha expression increase while Bcl-2 mRNA and protein expression increased with HIF-1alpha expression increase in HepG2 cells under normoxia condition. Meanwhile, Omi/HtrA2 protein expression increased with HIF-1alpha expression decrease in HepG2 cells under hypoxia culture. Taken together, these results demonstrated that HIF-1alpha suppressed hepatocellular carcinoma cell apoptosis through inhibiting Omi/HtrA2 expression and upregulating Bcl-2 expression to impede Omi/ HtrA2 releasing from the mitochondrion. The present finding further enriched and supported the role of HIF-1alpha expression on cell apoptosis of hepatoma cells.