Hypoxia-inducible factor-1αin myocardial infarction

Hypoxia-inducible factor 1(HIF1)has a crucial function in the regulation of oxygen levels in mammalian cells,especially under hypoxic conditions.Its importance in cardiovascular diseases,particularly in cardiac ischemia,is because of its ability to alleviate cardiac dysfunction.The oxygen-responsive subunit,HIF1α,plays a crucial role in this process,as it has been shown to have cardioprotective effects in myocardial infarction through regulating the expression of genes affecting cellular survival,angiogenesis,and metabolism.Furthermore,HIF1αexpression induced reperfusion in the ischemic skeletal muscle,and hypoxic skin wounds in diabetic animal models showed reduced HIF1αexpression.Increased expression of HIF1αhas been shown to reduce apoptosis and oxidative stress in cardiomyocytes during acute myocardial infarction.Genetic variations in HIF1αhave also been found to correlate with altered responses to ischemic cardiovascular disease.In addition,a link has been established between the circadian rhythm and hypoxic molecular signaling pathways,with HIF1αfunctioning as an oxygen sensor and circadian genes such as period circadian regulator 2 responding to changes in light.This editorial analyzes the relationship between HIF1αand the circadian rhythm and highlights its significance in myocardial adaptation to hypoxia.Understanding the changes in molecular signaling pathways associated with diseases,specifically cardiovascular diseases,provides the opportunity for innovative therapeutic interventions,especially in low-oxygen environments such as myocardial infarction.

DI-3-n-butylphthalide exerts neuroprotective effects by modulating hypoxia-inducible factor 1-alpha ubiquitination to attenuate oxidative stress-induced apoptosis

DI-3-n-butylphthalide is used to treat mild and moderate acute ischemic stroke.However,the precise underlying mechanism requires further investigation.In this study,we investigated the molecular mechanism of DI-3-n-butylphthalide action by various means.We used hydrogen peroxide to induce injury to PC12cells and RAW264.7 cells to mimic neuronal oxidative stress injury in stroke in vitro and examined the effects of DI-3-n-butylphthalide.We found that DI-3-nbutylphthalide pretreatment markedly inhibited the reduction in viability and reactive oxygen species production in PC12 cells caused by hydrogen peroxide and inhibited cell apoptosis.Furthermore,DI-3-n-butylphthalide pretreatment inhibited the expression of the pro-apoptotic genes Bax and Bnip3.DI-3-nbutylphthalide also promoted ubiquitination and degradation of hypoxia inducible factor 1α,the key transcription factor that regulates Bax and Bnip3 genes.These findings suggest that DI-3-n-butylphthalide exhibits a neuroprotective effect on stroke by promoting hypoxia inducible factor-1α ubiquitination and degradation and inhibiting cell apoptosis.

Quantitative analysis of hepatic hypoxia-inducible factor-1α and its abnormal gene expression during the formation of hepatocellular carcinoma

BACKGROUND:Hepatic hypoxia-inducible factor-1(HIF-1) is activated in the progression of hepatocellular carcinoma (HCC).This study aimed to investigate the dynamic alterations of HIF-1αand its gene expression so as to explore the relationship between HIF-1αexpression and hepatocarcinogenesis at the early stage of HCC. METHODS:A hepatoma model was made with 2-fluorenyl- acetamide(2-FAA)in male Sprague-Dawley rats.Morphological changes of rat hepatocytes were assessed pathologically (HE staining).The dynamic expression of hepatic and circulating HIF-1αwas quantitatively analyzed by ELISA. The gene fragments of hepatic HIF-1αmRNA were amplified by RT-PCR and confirmed by sequencing.The cellular distribution of hepatic HIF-1αexpression was confirmed by immunohistochemistry. RESULTS:Histological examination confirmed granulelike degeneration to atypical hyperplasia and HCC development in rat hepatocytes and progressive increases in the levels of hepatic and circulating HIF-1αand its gene expression during the course.The levels of HIF-1α expression in the liver and blood of rats with hepatoma were significantly higher than those in normal ratsand those with degeneration.Immunohistochemical analysis confirmed the positive expression and hepatocyte distribution of HIF-1αin the development of rat hepatoma. A positive relationship was found between HIF-1α expression in the liver and blood(P<0.01). CONCLUSIONS:The above observations support the hypothesis that the overexpression of HIF-1αand its gene are closely associated with the malignant transformation of hepatocytes and play an important role at the stage of hepatocarcinogenesis.

Hypoxia inducible factor-1αaccumulation in steatotic liver preservation:Role of nitric oxide

AIM:To examine the relevance of hypoxia inducible factor(HIF-1)and nitric oxide(NO)on the preservation of fatty liver against cold ischemia-reperfusion injury(IRI). METHODS:We used an isolated perfused rat liver model and we evaluated HIF-1αin steatotic and non-steatotic livers preserved for 24 h at 4℃in University of Wisconsin and IGL-1 solutions,and then subjected to 2 h of normothermic reperfusion.After normoxic reperfusion,liver enzymes,bile production,bromosulfophthalein clearance,as well as HIF-1αand NO[endothelial NO synthase(eNOS)activity and nitrites/nitrates]were also measured.Other factors associated with the higher susceptibility of steatotic livers to IRI,such as mitochondrial damage and vascular resistance were evaluated. RESULTS:A significant increase in HIF-1αwas found in steatotic and non-steatotic livers preserved in IGL-1 after cold storage.Livers preserved in IGL-1 showed a significant attenuation of liver injury and improvement in liver function parameters.These benefits were enhanced by the addition of trimetazidine(an antiischemic drug),which induces NO and eNOS activation, to IGL-1 solution.In normoxic reperfusion,the presence of NO favors HIF-1αaccumulation,promoting also the activation of other cytoprotective genes,such as hemeoxygenase-1. CONCLUSION:We found evidence for the role of the HIF-1α/NO system in fatty liver preservation,especially when IGL-1 solution is used.

Prognostic value of hypoxia-inducible factor-1 alpha and prolyl 4-hydroxylase beta polypeptide overexpression in gastric cancer

AIM To investigate the relationship between hypoxia-inducible factor-1α(HIF-1α), prolyl 4-hydroxylase beta(P4 HB) expression, and clinicopathologic parameters, as well as the prognostic value of these genes for patients with gastric cancer(Gc).METHODS Hypoxia is a critical factor that shapes the Gc microenvironment. In previous reports, we have demonstrated that P4 HB is a potential target of HIF-1α. In the present study, gene expression profiling interactive analysis(GEPIA) was used to analyze the relationship between P4 HB and hypoxia-associated genes. To this end, 428 Gc tissue samples were used to analyze the expression of HIF-1α and P4 HB via immunohistochemical staining. Patient samples were classified as having weak-expression or over-expression both in terms of HIF-1α and P4 HB. Correlations between biomarkers and clinicopathological factors were analyzed to predict survival. RESULTS P4 HB demonstrated a positive correlation with hypoxiaassociated genes(P < 0.05). HIF-1α and P4 HB overexpression have a significant correlation with TNM staging(χ2 = 23.32, P = 0.00; χ2 = 65.64, P = 0.00) and peritoneum cavity metastasis(χ2 = 12.67, P = 0.00; χ2 = 39.29, P = 0.00). In univariate analysis, patients with a high HIF-1α expression trend had a shorter disease-free survival(DFS: 44.80 mo vs 22.06 mo) and overall survival(OS: 49.58 mo vs 39.92 mo). P4 HB overexpression reflected similar results: patients with over-expression of P4 HB had a shorter survival time than those with weak-expression(DFS: 48.03 mo vs 29.64 mo, OS: 52.48 mo vs 36.87 mo). Furthermore, HIF-1α is also a clinicopathological predictor of dismal prognosis according to multivariate analysis(DFS, 95%c I: 0.52-0.88, P < 0.00; OS, 95%c I: 0.50-0.85, P < 0.00). However, P4 HB was meaningful in DFS(95%c I: 0.58-1.00, P < 0.05) but not in OS(95%c I: 0.72-1.23, P > 0.05).CONCLUSION Overexpression of HIF-1α and P4 HB is associated with poor prognosis in patients with Gc. Thus, these genes may be potential prognostic biomarker candidates in GC.

Expression of Nerve Growth Factor and Hypoxia Inducible Factor-1α and Its Correlation with Angiogenesis in Non-Small Cell Lung Cancer

Summary： In order to investigate the expression of nerve growth factor （NGF） and hypoxia inducible factor-1α （HIF-1α） and its correlation with angiogenesis in non-small cell lung cancer （NSCLC）, paraffin-embedded tissue blocks from 20 patients with NSCLC were examined. Twenty corresponding para-cancerous lung tissue specimens were obtained to serve as a control. The expression of NGF, HIF-1α, and vascular endothelial growth factor （VEGF） in the NSCLC tissues was detected by using immunohistochemistry. The microvascular density （MVD） was determined by CD31 staining. The resuits showed that the expression levels ofNGF, HIF-1α and VEGF in the NSCLC tissues were remarkably higher than those in the para-cancerous lung tissues （P〈0.05）. There was significant difference in the MVD between the NSCLC tissues （9.19±1.43） and para-cancerous lung tissues （2.23±1.19） （P〈0.05）. There were positive correlations between NGF and VEGF, between HIF-1α and VEGF, and between NGF and HIF-1α in NSCLC tissues, with the spearman correlation coefficient being 0.588, 0.519 and 0.588, respectively. In NSCLC tissues, the MVD had a positive correlation with the three factors （P〈0.05）. Theses results suggest that NGF and HIF-1α are synergically involved in the angiogenesis of NSCLC.

Metabolic shift in liver: Correlation between perfusion temperature and hypoxia inducible factor-1α

AIM: To study at what temperature the oxygen carried by the perfusate meets liver requirements in a model of organ perfusion. METHODS: in this study, we correlated hypoxia induciblefactor(Hi F)-1α expression to the perfusion temperature and the hepatic oxygen uptake in a model of isolated perfused rat liver. Livers from Wistar rats were perfused for 6 h with an oxygenated medium at 10, 20, 30 and 37 ℃. Oxygen uptake was measured by an oxygen probe; lactate dehydrogenase activity, lactate release and glycogen were measured spectrophotometrically; bile flow was gravitationally determined; p H of the perfusate was also evaluated; Hi F-1α m RNA and protein expression were analyzed by real time-polymerase chain reaction and ELi SA, respectively. RESULTS: Livers perfused at 10 and 20 ℃ showed no difference in lactate dehydrogenase release after 6 h of perfusion(0.96 ± 0.23 vs 0.93 ± 0.09 m U/min per g) and had lower hepatic damage as compared to 30 and 37 ℃(5.63 ± 0.76 vs 527.69 ± 45.27 m U/min per g, respectively, P s < 0.01). After 6 h, tissue ATP was significantly higher in livers perfused at 10 and 20 ℃than in livers perfused at 30 and 37 ℃(0.89 ± 0.06 and 1.16 ± 0.05 vs 0.57 ± 0.09 and 0.33 ± 0.08 nmol/mg, respectively, P s < 0.01). No sign of hypoxia was observed at 10 and 20 ℃, as highlighted by low lactate release respect to livers perfused at 30 and 37 ℃(121.4 ± 12.6 and 146.3 ± 7.3 vs 281.8 ± 45.3 and 1094.5 ± 71.7 nmol/m L, respectively, P s < 0.02), and low relative Hi F-1α m RNA(0.40 ± 0.08 and 0.20 ± 0.03 vs 0.60 ± 0.20 and 1.47 ± 0.30, respectively, P s < 0.05) and protein(3.72 ± 0.16 and 3.65 ± 0.06 vs 4.43 ± 0.41 and 6.44 ± 0.82, respectively, P s < 0.05) expression.CONCLUSION: Livers perfused at 10 and 20 ℃ show no sign of liver injury or anaerobiosis, in contrast to livers perfused at 30 and 37 ℃.

Hypoxia inducible factor-1 alpha stabilization for regenerative therapy in traumatic brain injury

Mild traumatic brain injury（TBI）, also called concussion, initiates sequelae leading to motor deficits, cognitive impairments and subtly compromised neurobehaviors. While the acute phase of TBI is associated with neuroinflammation and nitroxidative burst, the chronic phase shows a lack of stimulation of the neurorepair process and regeneration. The deficiency of nitric oxide（NO）, the consequent disturbed NO metabolome, and imbalanced mechanisms of S-nitrosylation are implicated in blocking the mechanisms of neurorepair processes and functional recovery in the both phases. Hypoxia inducible factor-1 alpha（HIF-1α）, a master regulator of hypoxia/ischemia, stimulates the process of neurorepair and thus aids in functional recovery after brain trauma. The activity of HIF-1α is regulated by NO via the mechanism of S-nitrosylation of HIF-1α. S-nitrosylation is dynamically regulated by NO metabolites such as S-nitrosoglutathione（GSNO） and peroxynitrite. GSNO stabilizes, and peroxynitrite destabilizes HIF-1α. Exogenously administered GSNO was found not only to stabilize HIF-1α and to induce HIF-1α-dependent genes but also to stimulate the regeneration process and to aid in functional recovery in TBI animals.

Meta-analysis of immunohistochemical expression of hypoxia inducible factor-1α as a prognostic role in gastric cancer

AIM:To conduct a meta-analysis to evaluate the prognostic role of hypoxia inducible factor-1α(HIF-1α)expression in gastric cancer.METHODS:The PubMed,EMBASE,and Web of Science databases were searched systematically for all articles published in English before August,2013.Pooled effect was calculated from the available data to evaluate the association between HIF-1αexpression and 5-year overall survival and tumor clinicopathological features in gastric cancer patients.Pooled odds ratios(ORs)with95%CIs were calculated using either a fixed-effects or a random-effects model.RESULTS:Nine studies matched the selection criteria,which reported on 1103 subjects,548 of whom had HIF-1αpositive expression(50%).This meta-analysis indicated that HIF-1αpositive expression in gastric cancer correlated with lower 5-year overall survival(OR=0.36;95%CI:0.21-0.64),worse tumor differentiation(OR=0.38;95%CI:0.23-0.64),deeper invasion(OR=0.42;95%CI:0.32-0.57),higher rates of lymph node metastasis(OR=2.23;95%CI:1.46-3.40),lymphatic invasion(OR=2.50;95%CI:1.46-4.28),and vascular invasion(OR=1.80;95%CI:1.29-2.51),and higher TNM stage(Ⅲ+Ⅳ)(OR=0.31;95%CI:0.15-0.60).CONCLUSION:HIF-1αpositive expression indicates a poor prognosis for patients with gastric cancer.Further studies are required to confirm these results.

Expression of caspase-3 and hypoxia inducible factor 1αin hepatocellular carcinoma complicated by hemorrhage and necrosis

BACKGROUND Hepatocellular carcinoma(HCC)is a malignant tumor that occurs in the liver.Its onset is latent,and it shows high heterogeneity and can readily experience intrahepatic metastasis or systemic metastasis,which seriously affects patients’quality of life.Numerous studies have shown that hypoxia inducible factor1α(HIF-1α)plays a significant role in the occurrence and development of tumors,as it promotes the formation of intratumoral vessels and plays a key role in their metastasis and invasion.Some studies have reported that caspase-3,which is induced by various factors,is involved in the apoptosis of tumor cells.AIM To investigate the expression of caspase-3 and HIF-1αand their relationship to the prognosis of patients with primary HCC complicated by pathological changes of hemorrhage and necrosis.METHODS A total of 88 patients with HCC complicated by pathological changes of hemorrhage and necrosis who were treated at our hospital from January 2017 to December 2019 were selected.The expression of caspase-3 and HIF-1αin HCC and paracancerous tissues from these patients was assessed.RESULTS The positive expression rate of caspase-3 in HCC tissues was 27.27%,which was significantly lower than that in the paracancerous tissues(P<0.05),while the positive expression rate of HIF-1αwas 72.73%,which was significantly higher than that in the paracancerous tissues(P<0.05).The positive expression rates for caspase-3 in tumor node metastasis(TNM)stage III and lymph node metastasis tissues were 2.78%and 2.50%,respectively,which were significantly lower than those in TNM stage I-II and non-lymph node metastasis tissues(P<0.05).The positive expression rates of HIF-1αin TNM stage III,lymph node metastasis,and portal vein tumor thrombus tissues were 86.11%,87.50%,and 88.00%,respectively,and these values were significantly higher than those in TNM stage I-II,non-lymph node metastasis,and portal vein tumor thrombus tissues(P<0.05).The expression of caspase-3 and HIF-1αin HCC tissues were negatively correlated(rs=−0.426,P<0.05).The median overall survival time of HCC patients was 18.90 mo(95%CI:17.20–19.91).The results of the Cox proportional risk regression model analysis showed that TNM stage,portal vein tumor thrombus,lymph node metastasis,caspase-3 expression,and HIF-1αexpression were the factors influencing patient prognosis(P<0.05).CONCLUSION The expression of caspase-3 decreases and HIF-1αincreases in HCC tissues complicated by pathological changes of hemorrhage and necrosis,and these are related to clinicopathological features and prognosis.

Hypoxia inducible factor-1alpha mediates protection of DL-3-n-butylphthalide in brain microvascular endothelial cells against oxygen glucose deprivation-induced injury

Studies have demonstrated that DL-3-n-butylphthalide can significantly alleviate oxygen glucose deprivation-induced injury of human umbilical vein endothelial cells at least partly associated with its enhancement on oxygen glucose deprivation-induced hypoxia inducible factor-1α expression.In this study,we hypothesized that DL-3-n-butylphthalide can protect against oxygen glucose deprivation-induced injury of newborn rat brain microvascular endothelial cells by means of upregulating hypoxia inducible factor-1α expression.MTT assay and Hoechst staining results showed that DL-3-n-butylphthalide protected brain microvascular endothelial cells against oxygen glucose deprivation-induced injury in a dose-dependent manner.Western blot and immunofluorescent staining results further confirmed that the protective effect was related to upregulation of hypoxia inducible factor-1α.Real-time RT-PCR reaction results showed that DL-3-n-butylphthalide reduced apoptosis by inhibiting downregulation of pro-apoptotic gene caspase-3 mRNA expression and upregulation of apoptosis-executive protease bcl-2 mRNA expression;however,DL-3-n-butylphthalide had no protective effects on brain microvascular endothelial cells after knockdown of hypoxia inducible factor-1α by small interfering RNA.These findings suggest that DL-3-n-butylphthalide can protect brain microvascular endothelial cells against oxygen glucose deprivation-induced injury by upregulating bcl-2 expression and downregulating caspase-3 expression though hypoxia inducible factor-1α pathway.

Effect of lentiviral vector-mediated overexpression of hypoxia-inducible factor 1 alpha delivered by pluronic F-127 hydrogel on brachial plexus avulsion in rats

Brachial plexus avulsion often results in massive motor neuron death and severe functional deficits of target muscles. However, no satisfactory treatment is currently available. Hypoxia-inducible factor 1α is a critical molecule targeting several genes associated with ischemia-hypoxia damage and angiogenesis. In this study, a rat model of brachial plexus avulsion-reimplantation was established, in which C5–7 ventral nerve roots were avulsed and only the C6 root reimplanted. Different implants were immediately injected using a microsyringe into the avulsion-reimplantation site of the C6 root post-brachial plexus avulsion. Rats were randomly divided into five groups: phosphate-buffered saline, negative control of lentivirus, hypoxia-inducible factor 1α(hypoxia-inducible factor 1α overexpression lentivirus), gel(pluronic F-127 hydrogel), and gel + hypoxia-inducible factor 1α(pluronic F-127 hydrogel + hypoxia-inducible factor 1α overexpression lentivirus). The Terzis grooming test was performed to assess recovery of motor function. Scores were higher in the hypoxia-inducible factor 1α and gel +hypoxia-inducible factor 1α groups(in particular the gel + hypoxia-inducible factor 1α group) compared with the phosphate-buffered saline group. Electrophysiology, fluorogold retrograde tracing, and immunofluorescent staining were further performed to investigate neural pathway reconstruction and changes of neurons, motor endplates, and angiogenesis. Compared with the phosphate-buffered saline group, action potential latency of musculocutaneous nerves was markedly shortened in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor1α groups. Meanwhile, the number of fluorogold-positive cells and ChAT-positive neurons, neovascular area(labeled by CD31 around av ulsed sites in ipsilateral spinal cord segments), and the number of motor endplates in biceps brachii(identified by α-bungarotoxin) were all visibly increased, as well as the morphology of motor endplate in biceps brachil was clear in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Taken together, delivery of hypoxia-inducible factor 1α overexpression lentiviral vectors mediated by pluronic F-127 effectively promotes spinal root regeneration and functional recovery post-brachial plexus avulsion. All animal procedures were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.

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.

Hypoxia inducible factor 1α promotes interleukin-1 receptor antagonist expression during hepatic ischemia-reperfusion injury

BACKGROUND Ischemia-reperfusion injury(IRI) is a major risk associated with liver surgery and transplantation,and its pathological mechanism is complex.Interleukin-1 receptor antagonist(IL-1ra) can protect the liver from IRI.However,the regulatory mechanism of IL-1ra expression is still unclear.AIM To identify the mechanism that could protect the liver in the early stage of IRI.METHODS To screen the key genes in hepatic IRI,we performed RNA sequencing and gene enrichment analysis on liver tissue from mice with hepatic IRI.Subsequently,we verified the expression and effect of IL-1ra in hepatic IRI.We also used promoter mutagenesis and chromatin immunoprecipitation assay to search for the transcriptional regulatory sites of hypoxia-inducible factor(HIF)-1α.Finally,to explore the protective mechanism of ischemic preconditioning(IP),we examined the expression of HIF-1α and IL-1ra after IP.RESULTS We identified IL-1ra as a key regulator in hepatic IRI.The expression of IL-1ra was significantly upregulated after hepatic IRI both in vivo and in vitro.Furthermore,we found that HIF-1αregulated Il-1ra transcription in response to hypoxia.Increased HIF-1α accumulation promoted IL-1ra expression,whereas inhibition of HIF-1α exhibited the opposite effect.We also confirmed a predominant role for hypoxia response element in the regulation of Il1ra transcription by HIF-1αactivation.Of note,we demonstrated that IP protects against hepatic IRI by inducing IL-1ra expression,which is mediated through HIF-1α.CONCLUSION We demonstrated that ischemia or hypoxia leads to increased expression of IL-1ra through HIF-1α.Importantly,IP protects the liver from IRI via the HIF-1α–IL-1ra pathway.

Antisense Oligonucleotide of Hypoxia-inducible Factor-1alpha Suppresses Growth and Tumorigenicity of Lung Cancer Cells A549

Summary： To study the role and mechanisms of hypoxia-inducible factor-lalpha （HIF-1α on the growth and tumorigenicity of lung cancer cells A549, the antisense oligonucleotide of HIF-1α was transfected to A549 cells. The effect of the antisense oligonucleotide on tumor growth in vitro and in vivo was evaluated by the growth rate suppression of A549 cells and subcutaneous implanted tumor in nude mice, and the effect on tumorigenicity was evaluated by the expression inhibition of angiogenic factors, the microvessel density （MVD）and vascular endothelial growth factor （VEGF） protein expression which were detected by immohistochemistry and western blot respectively. This study revealed that in vitro the growth rate of antisense oligonucleotide group was significantly decreased as compared with that of control group, sense oligonucleotide group and false-sense oligonucleotide group; in vivo the weight of implanted tumors in nude mice of antisense oligonucleotide group was 1.51±0.40 g, which was significantly lower than that of control group （2.79±0.33 g）, sense oligonucleotide group （2.81±0.45g） and false-sense oligonucleotide group （2.89±0.39 g） and the inhibitory rate was 47 %. Both MVD and VEGF protein expression were significantly inhibited in antisense oligonucleotide group compared with those in other groups. These results indicated that antisense oligonucleotide of HIF-1α could inhibit lung cancer cells A549 growth in vitro and in vivo, and the mechanism may be due to the inhibition of vascular growth and VEGF protein expression.

Expression and clinical significance of CD73 and hypoxia-inducible factor-1α in gastric carcinoma

AIM: To investigate the expression of CD73 and hypoxia-inducible factor-1α (HIF-1α) in human gastric carcinoma, and explore their clinical significance and prognostic value. METHODS: CD73 and HIF-1α expressions were detected by immunohistochemistry in consecutive sections of tissue samples from 68 gastric carcinoma patients. The peritumor tissues 2 cm away from the tumor were obtained and served as controls. The presence of CD73 and HIF-1α was analyzed by immunohis-tochemistry using the Envision technique. RESULTS: CD73 and HIF-1α expressions in gastric carcinoma were significantly higher than those in gastric mucosal tissues as control (P < 0.001) and showed a close correlation (Spearman r = 0.390, P = 0.001). Overexpression of CD73 was positively correlated with differentiation of tumor (P = 0.000), histopathology (P = 0.041), depth of invasion (P < 0.001), nodal status (P = 0.003), metastasis (P = 0.013), and the American Joint Committee on Cancer (AJCC) stage (P < 0.001). High expression of HIF-1α was positively correlated with tumor diameter (P = 0.031), depth of invasion (P = 0.022), and AJCC stage (P = 0.035). The overall survival rate was low in the patients with high expression of CD73 (P < 0.001). Moreover, CD73+/HIF-1α+ patients had the worst prognosis (P < 0.001). CD73 expression was proven to be an independent predictor for patients with gastric carcinoma by both multivariate Cox regression analysis (P = 0.021) and receiver operating characteristic curves (P = 0.001).CONCLUSION: CD73 expression correlates closely with HIF-1α expression in gastric carcinoma. CD73 could be an independent prognostic indicator for gastric carcinoma.

Hypoxia upregulates hypoxia inducible factor(HIF)-3α expression in lung epithelial cells: characterization and comparison with HIF-1α

The role of the hypoxia-inducible factor(HIF)subunits 1α and 2α in response to hypoxia is well established in lungepithelial cells,whereas little is known about HIF-3α with respect to transcriptional and translational regulation by hy-poxia.HIF-3α and HIF-1α are two similar but distinct basic helix-loop-helix-PAS proteins,which have been postulatedto activate hypoxia responsive genes in response to hypoxia.Here,we used quantitative real time RT-PCR and immu-noblotting to determine the activation of HIF-3α vs.HIF-1α by hypoxia.HIF-3α was strongly induced by hypoxia(1%O_2)both at the level of protein and mRNA due to an increase in protein stability and transcriptional activation,whereasHIF-1α protein and mRNA levels enhanced transiently and then decreased because of a reduction in its mRNA stabilityin A549 cells,as measured on mRNA and protein levels.Interestingly,HIF-3α and HIF-1α exhibited strikingly similarresponses to a variety of activating or inhibitory pharmacological agents.These results demonstrate that HIF-3α is ex-pressed abundantly in lung epithelial cells,and that the transcriptional induction of HIF-3α plays an important role in theresponse to hypoxia in vitro.Our findings suggest that HIF-3α,as a member of the HIF system,is complementary ratherthan redundant to HIF-1α induction in protection against hypoxic damage in alveolar epithelial cells.

Activation of hypoxia-inducible factor 1 attenuates periapical inflammation and bone loss

Hypoxia（low oxygen level） is an important feature during infections and affects the host defence mechanisms. The host has evolved specific responses to address hypoxia, which are strongly dependent on the activation of hypoxia-inducible factor 1（HIF-1）.Hypoxia interferes degradation of HIF-1 alpha subunit（HIF-1α）, leading to stabilisation of HIF-1α, heterodimerization with HIF-1 beta subunit（HIF-1β） and subsequent activation of HIF-1 pathway. Apical periodontitis（periapical lesion） is a consequence of endodontic infection and ultimately results in destruction of tooth-supporting tissue, including alveolar bone. Thus far, the role of HIF-1 in periapical lesions has not been systematically examined. In the present study, we determined the role of HIF-1 in a wellcharacterised mouse periapical lesion model using two HIF-1α-activating strategies, dimethyloxalylglycine（DMOG） and adenovirusinduced constitutively active HIF-1α（CA-HIF1 A）. Both DMOG and CA-HIF1 A attenuated periapical inflammation and tissue destruction. The attenuation in vivo was associated with downregulation of nuclear factor-κappa B（NF-κB） and osteoclastic gene expressions. These two agents also suppressed NF-κB activation and subsequent production of proinflammatory cytokines by macrophages. Furthermore, activation of HIF-1α by DMOG specifically suppressed lipopolysaccharide-stimulated macrophage differentiation into M1 cells, increasing the ratio of M2 macrophages against M1 cells. Taken together, our data indicated that activation of HIF-1 plays a protective role in the development of apical periodontitis via downregulation of NF-κB, proinflammatory cytokines, M1 macrophages and osteoclastogenesis.

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.

Hypoxia-inducible factor-1 modulates upregulation of mutT homolog-1 in colorectal cancer

AIM: To investigate the roles and interactions of mut T homolog(MTH)-1 and hypoxia-inducible factor(HIF)-1α in human colorectal cancer(CRC).METHODS: The expression and distribution of HIF-1α and MTH-1 proteins were detected in human CRC tissues by immunohistochemistry and quantitative realtime polymerase chain reaction(q RT-PCR). SW480 and HT-29 cells were exposed to normoxia or hypoxia. Protein and m RNA levels of HIF-1α and MTH-1 were analyzed by western blotting and q RT-PCR, respectively. In order to determine the effect of HIF-1α on the expression of MTH-1 and the amount of 8-oxodeoxyguanosine triphosphate(d GTP) in SW480 and HT-29 cells, HIF-1α was silenced with small interfering RNA(si RNA). Growth studies were conducted on cells with HIF-1α inhibition using a xenograft tumor model. Finally, MTH-1 protein was detected by western blotting in vivo.RESULTS: High MTH-1 m RNA expression was detected in 64.2% of cases(54/84), and this was significantly correlated with tumor stage(P = 0.023) and size(P = 0.043). HIF-1α protein expression was correlated significantly with MTH-1 expression(R = 0.640; P < 0.01) in human CRC tissues. Hypoxic stress induced m RNA and protein expression of MTH-1 in SW480 and HT-29 cells. Inhibition of HIF-1α by si RNA decreased the expression of MTH-1 and led to the accumulation of 8-oxo-d GTP in SW480 and HT-29 cells. In the in vivo xenograft tumor model, expression of MTH-1 was decreased in the HIF-1α si RNA group, and the tumor volume was much smaller than that in the mock si RNA group.CONCLUSION: MTH-1 expression in CRC cells was upregulated via HIF-1α in response to hypoxic stress, emphasizing the crucial role of HIF-1α-induced MTH-1 in tumor growth.