Is the hypoxia-inducible factor-1 alpha mRNA expression activated by ethanol-induced injury, the mechanism underlying alcoholic liver disease?

BACKGROUND: Excessive alcohol consumption can result in multiple organ injury, of which alcoholic liver disease (ALD) is the most common. With economic development and improvement of living standards, the incidence of diseases caused by alcohol abuse has been increasing in China, although its pathogenesis remains obscure. The aim of this study was to investigate the role of hypoxia in chronic ALD. METHODS: Twenty-eight male Sprague-Dawley rats were randomized into a control group (n=12) with a normal history and an experimental group (n=16) fed with 10 ml/ kg of 56% (vol/vol) ethanol once per day by gastric lavage for 24 weeks. At 24 weeks, blood samples were collected and then the rats were killed. Liver samples were frozen at -80 ℃ and used for RT-PCR; other liver samples were obtained for immunohistochemical staining. RESULTS: When the period of alcohol consumption increased, the positive rate of expression of hypoxia- inducible factor-1 alpha (HIF-1α) mRNA was more significantly elevated in the liver of the alcohol group than in the control group (P≤0.05). The HIF-1α protein located in the cytoplasm was seldom expressed in the control group, but significantly in the alcohol group (P≤0.01). CONCLUSION: HIF-1α mRNA expression was activated by ethanol-induced injury in this study, suggesting that hypoxia is involved in the underlying mechanism of ALD.

Clinicopathological and Prognostic Significance of Hypoxia-inducible Factor-1 alpha in Lung Cancer: a Systematic Review with Meta-analysis

Hypoxia-inducible factor-1 alpha（HIF-1α） plays a vital role in the initiation, evaluation and prognosis in lung cancer. The prognostic value of HIF-1α reported in diverse study remains disputable. Accordingly, a meta-analysis was implemented to further understand the prognostic role of HIF-1α in lung cancer. The relationship between HIF-1α and the clinicopathological characteristics and prognosis of lung cancer were investigated by a meta-analysis. Pub Med and Embase were searched from their inception to January 2015 for observational studies. Fixed-effects or random-effects meta-analyses were used to calculate odds ratios and 95% confidence intervals of different comparisons. A total of 20 studies met the criteria. The results showed that HIF-1α expression in lung cancer tissues was significantly higher than that in normal lung tissues. Expression of HIF-1α in patients with squamous cell carcinoma was significantly higher than that of patients with adenocarcinomas. Similarly, non-small cell lung cancer（NSCLC） patients had higher HIF-1α expression than small cell lung cancer（SCLC） patients. Moreover, lymph node metastasized tissues had higher HIF-1α expression than non-lymph node metastasized tissues. A high level HIF-1α expression was well correlated with the expression of vascular endothelial growth factor and epidermal growth factor receptor in the NSCLC. Notably, NSCLC or SCLC patients with positive HIF-1α expression in tumor tissues had lower overall survival rate than patients with negative HIF-1α expression. It was suggested that HIF-1α expression may be a prognostic biomarker and a potential therapeutic target for lung cancer.

Phosphatase and tensin homology deleted in chromosome 10,hypoxia-inducible factor-1 alpha gene expression in colorectal adenoma and adenocarcinoma and their relation to vascular endothelial growth factor protein expression

To examine phosphatase and tensin homology deleted in chromosome 10 (PTEN),hypoxia-inducible factor-1 alpha (HIF-1 alpha) gene expressions and their relation to vascular endothelial growth factor(VEGF) protein expression in the patients with human colorectal adenomas and adenocarcinomas.Methods The expression of PTEN,HIF-1 alpha gene was detected by using in situ hybridization,and the VEGF expression levels by immunohistochemistry in colorectal adenomas and primary colorectal adenocarcinoma.Results Strong expression of HIF-1 alpha was detectable in the majority of colorectal dadenocarcinoma,particularly surrounding areas of necrosis in adenocarcinoma.PTEN,HIF-1 alpha mRNA and VEGF protein were positive in 51.6%,67.7% and 59.7% respectively in 62 cases of adenocarcinomas,and 77.8%,44.4% and 33.3% respectively in 18 cases of adenomas.The positive rate of VEGF was higher in the patients with colorectal adenocarcinomas than that in those with adenomas,whereas that of PTEN mRNA was contrary.HIF-1 mRNA expression was correlated significantly with lymph node metastasis,liver metastasis,Duke’s stage and recurrence.During colorectal tumor progression,the expression of HIF-1 alpha mRNA was positively correlated with the VEGF protein expression (χ2= 4.751 ,P<0.05),but negatively with the PTEN mRNA expression(χ2=21.84,P<0.01).Conclusion The absence or low expression of PTEN and the increased levels of HIF-1α and VEGF may paly an important role in carcinogenesis and progression of colorectal carcinoma.These results suggest that VEGF upregulated by HIF-1 alpha gene may be involved in angiogenesis of colorectal adenocarcinoma.4 refs,1 tab.

Wortmannin influences hypoxia-inducible factor-1 alpha expression and glycolysis in esophageal carcinoma cells

AIM: To investigate the influence of phosphatidylinositol-3-kinase protein kinase B(PI3K/AKT)-HIF-1α signaling pathway on glycolysis in esophageal carcinoma cells under hypoxia. METHODS: Esophageal carcinoma cell lines Eca109 and TE13 were cultured under hypoxia environment, and the protein, m RNA and activity levels of hypoxia inducible factor-1 alpha(HIF-1α), glucose transporter 1, hexokinase-Ⅱ, phosphofructokinase 2 and lactate dehydrogenase-A were determined. Supernatant lactic acid concentrations were also detected. The PI3K/AKT signaling pathway was then inhibited with wortmannin, and the effects of hypoxia on the expression or activities of HIF-1α, associated glycolytic enzymes and lactic acid concentrations were observed. Esophageal carcinoma cells were then transfected with interference plasmid with HIF-1α-targeting si RNA to assess impact of the high expression of HIF-1α on glycolysis.RESULTS: HIF-1α is highly expressed in the esophageal carcinoma cell lines tested, and with decreasing levels of oxygen, the expression of HIF-1α and the associated glycolytic enzymes and the extracellular lactic acid concentration were enhanced in the esophageal carcinoma cell lines Eca109 and TE13. In both normoxia and hypoxic conditions, the level of glycolytic enzymesand the secretion of lactic acid were both reduced by wortmannin. The expression and activities of glycolytic enzymes and the lactic acid concentration in cells were reduced by inhibiting HIF-1α, especially the decreasing level of glycolysis was significant under hypoxic conditions.CONCLUSION: The PI3K/AKT pathway and HIF-1α are both involved in the process of glycolysis in esophageal cancer cells.

Inhibition of Expression of Hypoxia-inducible Factor-1α mRNA by Nitric Oxide in Hypoxic Pulmonary Hypertension Rats

In order to study the effect of nitric oxide (NO) on the expression of hypoxia inducible factor 1 alpha (HIF 1α) mRNA in hypoxic pulmonary hypertension (HPH) rats, 30 healthy male Wistar rats were randomly divided into normoxic control group, chronic hypoxic group and hypoxia plus L argine (L Arg) group. The animal model of HPH was developed. The mean pulmonary arterial pressure (mPAP) was measured by inserting a microcatheter into the pulmonary artery. The HIF 1α mRNA expression levels were detected by in situ hybridization (ISH) and semiquantitative RT PCR. It was found that after 14 days hypoxia, the mPAP in normoxic control group (17.6±2 7 mmHg,1 mmHg=0 133 kPa) was significantly lower than that in chronic hypoxic group(35.8±6.1 mmHg, t =0.2918, P <0.05) and mPAP in chronic hypoxic group was higher than that in hypoxia plus L argine group(24.4±3.8 mmHg, t =0.2563, P <0.05). ISH showed that the expression of HIF 1α mRNA in the intraacinar pulmonary arteriolae (IAPA) in normoxic control group (0.1076±0.0205) was markedly weaker than that in chronic hypoxic group (0.3317±0.0683, t =3.125, P <0.05) and that in chronic hypoxic group was stronger than that in hypoxia plus L argine group (0.1928±0.0381, t =2.844, P <0.05). RT PCR showed that the content of HIF 1α mRNA in chronic hypoxic group (2.5395±0.6449) was 2.16 times and 1.75 times higher than that in normoxic control group (1.1781±0.3628) and hypoxia plus L argine group (1.4511±0.3981), respectively. It is concluded that NO can reduce the mPAP by the inhibition of the expression of HIF 1α mRNA, which may be one of the mechanisms through which NO affects the pathogenesis of HPH.

Hippocampus hypoxia-inducible factor-1 alpha and heme oxygenase-1 expression in the delayed encephalopathy after acute carbon monoxide poisoning rat model

Objective To research the expression of hypoxia-inducible factor-1 alpha(HIF-1α)and heme oxygenase-1(HO-1)in hippocampus of rats with delayed encephalopathy after acute carbon monoxide poisoning(DEACMP)and its functions.Methods One hundred and fiftysix rats were selected and randomly divided into

Puerarin decreases hypoxia inducible factor-1 alpha in the hippocampus of vascular dementia rats

In this study, a rat vascular dementia model was established by permanent bilateral common carotid arterial occlusion. Rats were intraperitoneally injected with puerarin 3 days before modeling, for 45 successive days. Results demonstrated that in treated animals hippocampal structures were clear, nerve cells arranged neatly, and cytoplasm was rich in Nissl bodies. The number of cells positive for hypoxia inducible factor-1 alpha, erythropoietin and endothelial nitric oxide synthase was reduced; and the learning and memory abilities of rats were significantly improved. Our experimental findings indicate that puerarin can significantly improve learning and memory in a vascular dementia model, and that the underlying mechanism may be associated with the regulation of the expression of hypoxia inducible factor-1 alpha.

Downregulation of miR-491-5p promotes neovascularization after traumatic brain injury

Micro RNA-491-5 p(miR-491-5 p) plays an important role in regulating cell proliferation and migration;however,the effect of miR-491-5 p on neovascularization after traumatic brain injury remains poorly understood.In this study,a controlled cortical injury model in C57 BL/6 mice and an oxygen-glucose deprivation model in microvascular endothelial cells derived from mouse brain were established to simulate traumatic brain injury in vivo and in vitro,respectively.In the in vivo model,quantitative real-time-polymerase chain reaction results showed that the expression of miR-491-5 p increased or decreased following the intracerebroventricular injection of an miR-491-5 p agomir or antagomir,respectively,and the expression of miR-491-5 p decreased slightly after traumatic brain injury.To detect the neuroprotective effects of miR-491-p,neurological severity scores,Morris water maze test,laser speckle techniques,and immunofluorescence staining were assessed,and the results revealed that miR-491-5 p downregulation alleviated neurological dysfunction,promoted the recovery of regional cerebral blood flow,increased the number of lectin-stained microvessels,and increased the survival of neurons after traumatic brain injury.During the in vitro experiments,the potential mechanism of miR-491-5 p on neovascularization was explored through quantitative real-time-polymerase chain reaction,which showed that miR-491-5 p expression increased or decreased in brain microvascular endothelial cells after transfection with an miR-491-5 p mimic or inhibitor,respectively.Dual-luciferase reporter and western blot assays verified that metallothionein-2 was a target gene for miR-491-5 p.Cell counting kit 8(CCK-8) assay,flow cytometry,and 2′,7′-dichlorofluorescein diacetate(DCFH-DA) assay results confirmed that the downregulation of miR-491-5 p increased brain microvascular endothelial cell viability,reduced cell apoptosis,and alleviated oxidative stress under oxygen-glucose deprivation conditions.Cell scratch assay,Transwell assay,tube formation assay,and western blot assay results demonstrated that miR-491-5 p downregulation promoted the migration,proliferation,and tube formation of brain microvascular endothelial cells through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor pathway.These findings confirmed that miR-491-5 p downregulation promotes neovascularization,restores cerebral blood flow,and improves the recovery of neurological function after traumatic brain injury.The mechanism may be mediated through a metallothionein-2-dependent hypoxia-inducible factor-1α/vascular endothelial growth factor signaling pathway and the alleviation of oxidative stress.All procedures were approved by Ethics Committee of the First Affiliated Hospital of Chongqing Medical University,China(approval No.2020-304) on June 22,2020.

Hypoxia-inducible factor-lαand semaphorin4D genes involved with tumor-associated macrophage-induced metastatic behavior and clinical significance in colon cancer

Background Hypoxia promotes tumor angiogenesis and hypoxia-inducible factor-1 alpha （HIF-lg） plays a pivotal role in this process. Recently identified pro-angiogenic factor, semaphorin4D （Sema4D） also promotes angiogenesis and enhances invasive proliferation in some tumors. Furthermore, tumor-associated macrophages （TAMs） can increase the expression of HIF-la and Sema4D in cancer cells and thus influence tumor growth and progression. The purpose of this study was to evaluate the effect of TAMs on the expression of Sema4D and HIF-la and the impact of biologic behavior in colon cancer cells. Methods Immunohistochemistry was used to analyze HIF-la and Sema4D expression in 86 curatively resected colon cancer samples and 52 normal colon tissues samples. The relationship between their expression and clinicopathological factors was analyzed. Furthermore, macrophage-tumor cell interactions, such as metastasis, angiogenesis, were also studied using in vitro co-culture systems. Statistical analysis was performed using SPSS 17.0 software （SPSS Inc., USA）. Differences between two groups were analyzed with Student＇s t test. Results HIF-la （58%） and Sema4D （60%） were expressed at a significantly higher level in tumors than in normal tissues （P 〈0.01, for both）. Furthermore, HIF-la and Sema4D expression was significantly correlated with lymphatic metastasis, specific histological types and TNM stages （P 〈0.05）, but not with age and tumor size （P 〉0.05）. Sema4D expression was correlated with that of HIF-la （r=0.567, P 〈0.01）. TAMs markedly induced HIF-la and Sema4D expression in colon cancer calls and subsequently increased their migration and invasion. Conclusions HIF-la and Sema4D expression are closely related to lymphatic metastasis, specific histological types and TNM stages in colon cancer. Furthermore, TAMs promote migration and invasion of colon cancer cells and endothelial tube formation, possibly through up-regulation of HIF-la and Sema4D.

HIF-1α signaling: Essential roles in tumorigenesis and implications in targeted therapies

The hypoxic microenvironment is an essential characteristic of most malignant tu-mors.Notably,hypoxia-inducible factor-1 alpha(HIF-1a)is a key regulatory factor of cellular adaptation to hypoxia,and many critical pathways are correlated with the biological activity of organisms via HIF-1a.In the intra-tumoral hypoxic environment,HIF-1αis highly expressed and contributes to the malignant progression of tumors,which in turn results in a poor prog-nosis in patients.Recently,it has been indicated that HiF-1αinvolves in various critical pro-cesses of life events and tumor development via regulating the expression of HiF-1a target genes,such as cell proliferation and apoptosis,angiogenesis,glucose metabolism,immune response,therapeutic resistance,etc.Apart from solid tumors,accumulating evidence has re-vealed that HiF-1αis also closely associated with the development and progression of hemato-logical malignancies,such as leukemia,lymphoma,and multiple myeloma.Targeted inhibition of HiF-1a can facilitate an increased sensitivity of patients with malignancies to relevant ther-apeutic agents.In the review,we elaborated on the basic structure and biological functions of HIF-1a and summarized their current role in various malignancies.It is expected that they will have future potential fortargeted therapy.

Efficacy of bioactive compounds of Chaihu(Radix Bupleuri Chinensis)on glaucomatous optic atrophy through interleukin-6/hypoxia inducible factor-1αsignal pathway

OBJECTIVE:To investigate the bioactive compounds of Chaihu(Radix Bupleuri Chinensis)(RB)on glaucomatous optic atrophy(GOA),and to study the pharmacological mechanism.METHODS:We collected information on the bioactive compounds of RB from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP).Targets related to bioactive compounds and GOA were also obtained.Gene Ontology(GO),Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway and network analyses were performed to investigate the potential mechanism of RB against GOA.Subsequently,the main bioactive compounds of RB and targets of GOA were docked by Autodock software.Moreover,a GOA model of retinal ganglion cells(RGCs)induced by cobalt chloride was established to verify the effect of RB on GOA.RESULTS:There were 17 main bioactive compounds and 46 key targets were screened as potential players in GOA.The compound-target network mainly contained 17 compounds and 46 corresponding targets,and the key targets consisted of interleukin-6(IL-6),hypoxia inducible factor-1α(HIF1A),Caspase-3,estrogen receptor alpha(ESR1),MYC proto-oncogene(MYC),and vascular endothelial growth factor A(VEGFA).Forty-nine significantly enriched GO terms,and 134 KEGG signaling pathways were identified(P<0.05),including HIF-1,tumor necrosis factor,VEGF,prolaction,and other signaling pathways.Molecular docking results showed that the main bioactive compounds of RB exhibited the strongest binding activity with IL-6.Furthermore,experimental validation showed that the RB extract inhibited the activity and promoted apoptosis of RGCs in a dose-dependent manner.The RB extract also suppressed the expression of Bax,Caspase-3,and Caspase-9 and regulated malonaldehyde,superoxide dismutase,and glutathione peroxide by inhibiting the IL-6/HIF-1αsignaling pathway.CONCLUSIONS:The present study provided insights into the mechanism of RB on GOA.RB mainly reverses GOA by inhibiting the IL-6/HIF-1αsignaling pathway.

HIF-1α and MIF enhance neutrophil-driven type 3 immunity and chondrogenesis in a murine spondyloarthritis model

The hallmarks of spondyloarthritis(SpA)are type 3 immunity-driven inflammation and new bone formation(NBF).Macrophage migration inhibitory factor(MIF)was found to be a key driver of the pathogenesis of SpA by amplifying type 3 immunity,yet MIF-interacting molecules and networks remain elusive.Herein,we identified hypoxia-inducible factor-1 alpha(HIF1A)as an interacting partner molecule of MIF that drives SpA pathologies,including inflammation and NBF.HIF1A expression was increased in the joint tissues and synovial fluid of SpA patients and curdlan-injected SKG(curdlan-SKG)mice compared to the respective controls.Under hypoxic conditions in which HIF1A was stabilized,human and mouse neutrophils exhibited substantially increased expression of MIF and IL-23,an upstream type 3 immunity-related cytokine.Similar to MIF,systemic overexpression of IL-23 induced SpA pathology in SKG mice,while the injection of a HIF1A-selective inhibitor(PX-478)into curdlan-SKG mice prevented or attenuated SpA pathology,as indicated by a marked reduction in the expression of MIF and IL-23.Furthermore,genetic deletion of MIF or HIF1A inhibition with PX-478 in IL-23-overexpressing SKG mice did not induce evident arthritis or NBF,despite the presence of psoriasis-like dermatitis and blepharitis.We also found that MIF-and IL-23-expressing neutrophils infiltrated areas of the NBF in curdlan-SKG mice.These neutrophils potentially increased chondrogenesis and cell proliferation via the upregulation of STAT3 in periosteal cells and ligamental cells during endochondral ossification.Together,these results provide supporting evidence for an MIF/HIF1A regulatory network,and inhibition of HIF1A may be a novel therapeutic approach for SpA by suppressing type 3 immunity-mediated inflammation and NBF.

External use of Ruyanneixiao cream efficiently blocks precancerous mammary lesions by interfering with glycolysis induced by inhibition of hypoxia inducible factor-1α, hexokinase 2, phosphofructokinase, and pyruvate kinase M2 expression

OBJECTIVE:To investigate the effect of Ruyanneixiao cream(RYNX) on the expression of hypoxia inducible factor-1α(HIF-1α), hexokinase 2(HK2),phosphofructokinase(PFK), and pyruvate kinase M2(PKM2) mRNA and protein in MCF-10 AT cells and in an animal model of precancerous mammary lesions.METHODS:Following treatment of MCF-10 AT cells with RYNX, tamoxifen(TAM) and YC-1 for 48 h,HIF-1α, HK2, PFK, PKM2 mRNA and protein expression was analyzed.Fifty female SD rats were randomly divided into control, model, TAM, and highand low-dose RYNX groups, with 10 rats in each group.A precancerous mammary lesion model was established for all groups except the control group.High-and low-dose RYNX cream containing TAM was coated on the breasts of animals in the corresponding groups.The rat mammary tissue was removed in the 10 th week and HIF-1α, HK2, PFK,PKM2 mRNA and protein was analyzed.RESULTS:In vitro analyses demonstrated that, compared with the matrix group, HIF-1α, HK2, PFK,PKM2 mRNA and protein expression was significantly decreased in the RYNX group(P < 0.05).Compared with the YC-1 + RYNX group, HK2, PFK,and PKM2 protein expression was significantly reduced in the RYNX group.HIF-1α, HK2, PFK, and PKM2 protein expression was increased significantly in the model group(P < 0.05) compared with the control group, while HIF-1α, HK2, PFK, and PKM2 mRNA and protein expression was significantly decreased in both the high-and low-dose RYNX groups(P < 0.05), with the effect being greater in the high-dose group.CONCLUSION:RYNX can block precancerous breast lesions by decreasing the expression of HK2,PFK, and PKM2 mRNA and protein via inhibition of HIF-1α mRNA and protein overexpression in a dose-dependent manner.

A Correlative Study between CT Perfusion Parameters and Angiogenesis in Rabbit VX2 Liver Tumors

Objective: The purpose of this study was to evaluate the correlation between CT perfusion parameters and the hypoxia-inducible factor-1 alpha (HIF-1α), vascular en-dothelial growth factor (VEGF), matrix metalloproteinase-2 (MMP-2) and microvessel density (MVD) marked by CD34 molecular of rabbit VX2 liver tumors and to investigate the value of CT perfusion imaging in evaluating tumor angiogenesis. Material and methods: Twenty-four cases of rabbit VX2 liver tumor were performed by CT perfusion scanning. Hepatic artery perfusion (HAP), portal vein perfusion (PVP), total hepatic blood flow (THBF) and hepatic perfusion index (HPI) were measured by perfusion software. HIF-1α, VEGF and MMP-2 expression and MVD were detected in the 24 rabbit VX2 liver tumor tissue samples using immunohistochemical method. The correlation between the HIF-1α, VEGF, MMP-2 expression and MVD and CT perfusion parameters were analyzed. Results: Correlation analysis revealed that the expression of HIF-1α, MMP-2, MVD were positively related to the HAP, THBF, HPI (p < 0.01), but no relations with PVP (p > 0.05);and correlation analysis revealed that the expression of VEGF was positively related to the HAP, HPI (p 0.05). There was a positive relationship between the expression of HIF-1α, VEGF, MMP-2 and MVD (p < 0.01). Conclusions: CT perfusion imaging can reflect the blood perfusion of the rabbit VX2 liver tumors and evaluate the information of angiogenesis about tumors.

Hypoxia and cytokines regulate carbonic anhydrase 9 expression in hepatocellular carcinoma cells in vitro

AIM: To study the expression of carbonic anhydrase(CA) 9 in human hepatocellular carcinoma(HCC) cells.METHODS: We studied CA9 protein, CA9 m RNA and hypoxia-inducible factor-1 alpha(HIF-1α) protein levels in Hep3 B cells exposed in different parallel approaches. In one of these approaches, HCC cells were exposed to extreme in vitro hypoxia(24 h 0.1% O2) without or with interleukin(IL)-1, IL-6, tumor necrosis factoralpha(TNF-α) and transforming growth factor-beta(TGF-β) stimulation for the same hypoxic exposure time or exposed to normoxic oxygenation conditions without or with cytokine stimulation.RESULTS: The tumour cell line analysed showed a strong hypoxic CA9 m RNA expression pattern in response to prolonged severe hypoxia with cell-line specific patterns and a marked induction of CA9 protein in response to severe hypoxia. These results were paralleled by the results for HIF-1α protein under identical oxygenation conditions with a similar expression tendency to that displayed during the CA9 protein expression experimental series. Continuous stimulation with the cytokines, IL-1, IL-6, TNF-α and TGF-β, under normoxic conditions significantly increased the carbonic anhydrase 9 expression level at both the protein and m RNA level, almost doubling the CA9 m RNA and CA9 and HIF-1α protein expression levels found under hypoxia. The findings from these experiments indicated that hypoxia is a positive regulator of CA9 expression in HCC, and the four signal transduction pathways, IL-1, IL-6, TNF-α and TGF-β, positively influence CA9 expression under both normoxic and hypoxic conditions.CONCLUSION: These findings may potentially be considered in the design of anti- cancer therapeutic approaches involving hypoxia-induced or cytokine stimulatory effects on expression. In addition, they provideevidence of the stimulatory role of the examined cytokine families resulting in an increase in CA9 expression under different oxygenation conditions in human cancer, especially HCC, and on the role of the CA9 gene as a positive disease regulator in human cancer.

Transforming Growth Factor-beta I Involved in the Pathogenesis of Endometriosis through Regulating Expression of Vascular Endothelial Growth Factor under Hypoxia

Background： Endometriosis （EMs） is a common gynecological disorder characterized by endometrial-like tissue outside the uterus. Hypoxia induces the expression of many important downstream genes to regulate the implantation, survival, and maintenance ofectopic endometriotic lesions. Transtbrming growth factor-beta I （TGF-β1） plays a major role in the etiology of EMs. We aimed to determine whether TGF-β1 affects EMs development and progression and its related mechanisms in hypoxic conditions. Methods： Endometrial tissue was obtained from women with or without EMs undergoing surgery from October, 2015 to October, 2016. Endometrial cells were cultured and then exposed to hypoxia and TGF-β1 or TGF-β1 inhibitors. The messenger RNA （mRNA） and protein expression levels ofTGF-β1, vascular endothelial growth fhctor （VEGF）, and hypoxia-inducible fhctor-Ic~ （HIF-β1） were measured. A DuaI-Luciferase Reporter Assay was used to examine the effect ofTGF-[31 and hypoxia on a VEGF promoter construct. Student＇s t-test was pertbrmed/br comparison among groups （one-sided or two-sided） and a value ofP 〈 0.05 was considered statistically significant. Results： TGF-β1, VEGF, HIF-β1 mRNA, and protein expression were significantly higher in EMs tissue than that in normal endometrial tissue （t = 2.16, P = 0.042）. EMs primary cultured cells exposed to hypoxia expressed 43.8% higher VEGF mRNA and protein （t = 6.84, P - 0.023）. VEGF mRNA levels increased 12.5% in response to TGF-β1, whereas the combined treatment of hypoxia/TGF-β1 resulted in a much higher production （87.5% increases） of VEGF. The luciferase activity of the VEGF promoter construct was increased in the presence of either TGF-β1 （2.6-fold, t = 6.08, P = 0.032） or hypoxia （11.2-fold, t = 32.70, P 〈 0.001 ）, whereas the simultaneous presence of both stimuli resulted in a significant cooperative effect （ 18.5-fold, t = 33.50, P 〈 0.001 ）. Conclusions： The data support the hypothesis that TGF-β1 is involved in the pathogenesis of EMs through regulating VEGF expression. An additive effect of TGF-[31 and hypoxia is taking place at the transcriptional level.

Efficacy of Shouzhangshen(Rhizoma Gymnadeniae Crassinervidis)extract against acute high altitude hypoxia-induced brain injury in mice

OBJECTIVE:To evaluate the protective effect of Shouzhangshen(Rhizoma Gymnadeniae Crassinervidis)extract against acute high altitude hypoxia-induced brain injury in mice.METHODS:Sixty C57BL/6J mice were selected and assigned to six groups(n=10):normal control group,low-pressure hypoxia group,positive control group(dexamethasone 500 mg/kg),and three groups treated with Shouzhangshen extract(250,500,and 750 mg/kg,respectively).The Morris water maze test was performed to evaluate alterations in spatial learning and memory deficits.Nissl staining was performed to detect Nissl bodies and neuron damage.Hypoxia-inducible factor(HIF)-1α,interleukin(IL)-1β,tumor necrosis factor(TNF)-α,vascular endothelial growth factor(VEGF),and malondialdehyde(MDA)expression in brain tissue and serum,as well as superoxide dismutase(SOD)and glutathione(GSH)activity in brain tissues were measured by enzyme-linked immunosorbent assays,quantitative real-time-polymerase chain reaction and western blots.RESULTS:The Morris water maze test results showed that Shouzhangshen extract can significantly reduce the latency and swimming distance to escape onto a visible platform,increase neuron density and hierarchy and the number of pyramidal neurons,and decrease the expression of HIF-1α,IL-1β,TNF-α,and VEGF mRNAs and proteins in both brain tissue and serum(P<0.05).Furthermore,significantly lower MDA expression and higher GSH activity were detected in the three groups treated with Shouzhangshen compared with the low-pressure hypoxia group(P<0.05).However,no significant alteration was observed for SOD activity(P>0.05).CONCLUSION:Our findings suggest that Shouzhangshen extract may have a significant effect on acute high altitude hypoxia-induced brain injury in mice.

Forensic Significance of Messenger RNA and Protein Expression of Genes Downstream of Hypoxia Inducible Factor 2 in Myocardial Tissue for Death Discrimination

Background: As a heterodimeric transcription factor, hypoxia-inducible factor 2 alpha subunit (HIF2A), is an important member of the HIF family. It plays a significant role in the hypoxia adaptation process by regulating the different types of downstream transcription factors and auxiliary regulatory factors. HIF2A-related factors are believed to participate in the progression of myocardial injury or myocardial ischemia, support the protection of ischemic myocardium, and provide guiding significance for the diagnosis and discrimination of sudden cardiac death in forensic pathology. Aim and Objectives: This study aimed to explore the discriminability and applicability of HIF2A-related factors in myocardial infarction cases compared with other causes of death, provide further insights for the forensic diagnosis of heart failure (HF) cases with myocardial infarction, and support the clinical treatment of patients with HF after myocardial infarction. Materials and Methods: The relative expression levels of HIF2A, amphiregulin (AREG), potassium large conductance calcium-activated channel subfamily M β1 (KCNMB1), peroxisome proliferator-activated receptor α (PPARA), vascular endothelial growth factor (VEGF), and VEGFR2 messenger RNAs (mRNAs) in myocardial tissue samples were performed using quantitative reverse transcriptase-polymerase chain reaction. A partial least squares-discriminant analysis model was constructed to select the indicators with better identification effects for myocardial infarction cases. The protein levels of HIF2A, AREG, KCNMB1, and PPARA were further detected by immunohistochemistry. The forensic autopsy cases (27 cases in total, postmortem interval <72 h) included seven cases of acute myocardial infarction and ten cases of myocardial ischemia. There were ten cases in the control group, including four cases of traffic injury, one case of injury by fall from height, and five cases of blunt force injury. Results: Characteristic results were observed in the myocardial ischemia/infarction samples. Compared with the control group, the relative mRNA expression levels of AREG, KCNMB1, and PPARA were significantly increased during the progression of myocardial ischemia, but this was not observed for HIF2A, VEGF, or VEGFR2 mRNA. Immunohistochemistry assays further verified the expression levels of the related factors at the protein level, and H and E staining showed signs of angiogenesis and inflammation in the ischemia/infarction group. Conclusions: By controlling the expression of downstream target genes (AREG, KCNMB1, and PPARA) during myocardial cell hypoxia adaptation, HIF2A has a potential significance in the diagnosis of myocardial infarction in forensic medicine. We believe that HIF2A, AREG, KCNMB1, and PPARA can be used as molecular pathological biomarkers for the discrimination of causes of death in myocardial infarction cases.