Hypoxia-inducible factor 1alpha and vascular endothelial growth factor in Glioblastoma Multiforme:a systematic review going beyond pathologic implications

Glioblastoma multiforme(GBM)is an aggressive primary brain tumor characterized by extensive heterogeneity and vascular proliferation.Hypoxic conditions in the tissue microenvironment are considered a pivotal player leading tumor progression.Specifically,hypoxia is known to activate inducible factors,such as hypoxia-inducible factor 1alpha(HIF-1α),which in turn can stimulate tumor neo-angiogenesis through activation of various downward mediators,such as the vascular endothelial growth factor(VEGF).Here,we aimed to explore the role of HIF-1α/VEGF immunophenotypes alone and in combination with other prognostic markers or clinical and image analysis data,as potential biomarkers of GBM prognosis and treatment efficacy.We performed a systematic review(Medline/Embase,and Pubmed database search was completed by 16th of April 2024 by two independent teams;PRISMA 2020).We evaluated methods of immunoassays,cell viability,or animal or patient survival methods of the retrieved studies to assess unbiased data.We used inclusion criteria,such as the evaluation of GBM prognosis based on HIF-1α/VEGF expression,other biomarkers or clinical and imaging manifestations in GBM related to HIF-1α/VEGF expression,application of immunoassays for protein expression,and evaluation of the effectiveness of GBM therapeutic strategies based on HIF-1α/VEGF expression.We used exclusion criteria,such as data not reporting both HIF-1αand VEGF or prognosis.We included 50 studies investigating in total 1319 GBM human specimens,18 different cell lines or GBM-derived stem cells,and 6 different animal models,to identify the association of HIF-1α/VEGF immunophenotypes,and with other prognostic factors,clinical and macroscopic data in GBM prognosis and therapeutic approaches.We found that increased HIF-1α/VEGF expression in GBM correlates with oncogenic factors,such as miR-210-3p,Oct4,AKT,COX-2,PDGF-C,PLDO3,M2 polarization,or ALK,leading to unfavorable survival.Reduced HIF-1α/VEGF expression correlates with FIH-1,ADNP,or STAT1 upregulation,as well as with clinical manifestations,like epileptogenicity,and a favorable prognosis of GBM.Based on our data,HIF-1αor VEGF immunophenotypes may be a useful tool to clarify MRI-PET imaging data distinguishing between GBM tumor progression and pseudoprogression.Finally,HIF-1α/VEGF immunophenotypes can reflect GBM treatment efficacy,including combined first-line treatment with histone deacetylase inhibitors,thimerosal,or an active metabolite of irinotecan,as well as STAT3 inhibitors alone,and resulting in a favorable tumor prognosis and patient survival.These data were supported by a combination of variable methods used to evaluate HIF-1α/VEGF immunophenotypes.Data limitations may include the use of less sensitive detection methods in some cases.Overall,our data support HIF-1α/VEGF’s role as biomarkers of GBM prognosis and treatment efficacy.

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.

Modified Shenlingbaizhu decoction reduces intestinal adenoma formation in adenomatous polyposis coli multiple intestinal neoplasia mice by suppression of hypoxia-inducible factor 1α-induced CD4+CD25+forkhead box P3 regulatory T cells

OBJECTIVE： To test the hypothesis that modified Shenlingbaizhu decoction （MSD） attenuates the for- mation of intestinal adenomas by regulating activa- tion of CD4＋CD25＋ forkhead box P3 （FoxP3） regu- latory T cells （Tregs） by downregulation of hypox- ia-inducible factor la （HIF-la）. METHODS： Chemical fingerprints of ginsenoside Rbl, ginsenoside Rc, paeoniflorin, and dioscin in standard extractions were used as material bases of MSD. Adenomatous polyposis coli multiple intesti- nal neoplasia （ApcM＇n/＋） mice, which harbor a muta- tion in adenomatous polyposis coil, were used to host intestinal adenomas. Peripheral blood and spleen Tregs were analyzed by flow cytometry. Pro- tein expression was analyzed by immunohisto- chemistry and Western blotting. RESULTS： The number and size of intestinal adenomas were significantly reduced by MSD treatment. Mucosal thickening and the spleen size were also substantially decreased by MSD. The carcinogenesis process in Apc^min/＋ mice resembled that of human colorectal cancer. Molecular markers of neoplasms, such as 13-catenin, cyclooxygenase-2, prolif- erating cell nuclear antigen, and p53, were substantially ameliorated by MSD treatment. Moreover, MSD downregulated peripheral and spleen CD4＋ CD25＋FoxP3＋ Tregs and reduced in situ expression of CD4, CD25, and FoxP3 in intestinal adenomas. MSD also suppressed HIF-la expression in the intestinal adenomas, and HIF-la inhibition decreased expression of FoxP3 in Jurkat T cells under hypoxic conditions. CONCLUSION： MSD is a valid prescription to control the formation of intestinal adenomas in Apc^min/＋mice. It exerts anti-cancer effects partially through suppression of HIF-la that induced activation of CD4＋CD25＋FoxP3＋ Tregs in vivo and in vitro.

Extracellular vesicles from hypoxia-preconditioned mesenchymal stem cells alleviates myocardial injury by targeting thioredoxininteracting protein-mediated hypoxia-inducible factor-1αpathway

BACKGROUND Extracellular vesicles(EVs)derived from hypoxia-preconditioned(HP)mesenchymal stem cells(MSCs)have better cardioprotective effects against myocardial infarction(MI)in the early stage than EVs isolated from normoxic(NC)-MSCs.However,the cardioprotective mechanisms of HP-EVs are not fully understood.AIM To explore the cardioprotective mechanism of EVs derived from HP MSCs.METHODS We evaluated the cardioprotective effects of HP-EVs or NC-EVs from mouse adipose-derived MSCs(ADSCs)following hypoxia in vitro or MI in vivo,in order to improve the survival of cardiomyocytes(CMs)and restore cardiac function.The degree of CM apoptosis in each group was assessed by the terminal deoxynucleotidyl transferase dUTP nick end-labeling and Annexin V/PI assays.MicroRNA(miRNA)sequencing was used to investigate the functional RNA diversity between HP-EVs and NC-EVs from mouse ADSCs.The molecular mechanism of EVs in mediating thioredoxin-interacting protein(TXNIP)was verified by the dual-luciferase reporter assay.Co-immunoprecipitation,western blotting,and immunofluorescence were performed to determine if TXNIP is involved in hypoxia-inducible factor-1 alpha(HIF-1α)ubiquitination and degradation via the chromosomal region maintenance-1(CRM-1)-dependent nuclear transport pathway.RESULTS HP-EVs derived from MSCs reduced both infarct size(necrosis area)and apoptotic degree to a greater extent than NC-EVs from CMs subjected to hypoxia in vitro and mice with MI in vivo.Sequencing of EV-associated miRNAs showed the upregulation of 10 miRNAs predicted to bind TXNIP,an oxidative stress-associated protein.We showed miRNA224-5p,the most upregulated miRNA in HP-EVs,directly combined the 3’untranslated region of TXNIP and demonstrated its critical protective role against hypoxia-mediated CM injury.Our results demonstrated that MI triggered TXNIP-mediated HIF-1αubiquitination and degradation in the CRM-1-mediated nuclear transport pathway in CMs,which led to aggravated injury and hypoxia tolerance in CMs in the early stage of MI.CONCLUSION The anti-apoptotic effects of HP-EVs in alleviating MI and the hypoxic conditions of CMs until reperfusion therapy may partly result from EV miR-224-5p targeting TXNIP.

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.

Expression of hypoxia-inducible factor la and vascular endothelial growth factor in hepatocellular carcinoma: Impact on neovascularization and survival

AIM: To study the expression of hypoxia-inducible factor 1α(HIF-1α) and vascular endothelial growth factor (VEGF) in hepatocellular carcinoma (HCC) and the impact on neovascularization and survival. METHODS: Expressions of HIF-1α, VEGF and microvessel density (MVD) are studied through immunohistochemistry in 36 cases of HCC and the corresponding paraneoplastic tissue and 6 cases of normal liver tissue. The relationship of the expressions of HIF-1α and VEGF with the clinicopathological data and survival are analyzed. RESULTS: The positive rate of VEGF in HCC was 32/36, which is significantly higher than that in paraneoplastic tissue and normal liver tissue (P<0.05). The expression of HIF-1aaaaaa in HCC tissue is 24/36, also higher than that in paraneoplastic tissue and normal liver tissue (P<0.05). The expression of VEGF and HIF-1α in HCC with microscopic venous invasion is significantly higher than that in HCC without microscopic venous invasion (P<0.05). Spearman correlation analysis does not only show the expression of HIF-1α as correlated with the expression of VEGF (rs = 0.459, P<0.01), but it also shows the expression of HIF-1α and VEGF as correlated with MVD (rs=0.412 and 0.336, respectively, P<0.05). The differences of the survival rates among VEGF positive group and VEGF negative group are significant (P<0.05), whereas the differences of the survival rates among the HIF-1α negative group and positive group are not significant (P>0.05). CONCLUSION: HIF-1α plays important roles in neovascularization in HCC possibly through regulation of VEGF transcription.

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.

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.

乙醇对胃癌细胞缺氧与抗凋亡蛋白的影响

目的探讨乙醇对胃癌细胞缺氧诱导因子与抗凋亡蛋白Survivin的影响。方法将人胃癌细胞BGC823分组培养,加入5%-30%终浓度的乙醇培养1-4h,并设空白对照组,提取蛋白后采用免疫印迹法检测HIF-1α蛋白及Survivin蛋白的表达。结果在BGC823细胞中HIF-1α蛋白的表达随着乙醇浓度和时间的加大而增加;Survivin蛋白表达在乙醇处理1h后随着乙醇浓度的增加表现为先增加后减少,在乙醇处理4h后随着乙醇浓度的增加反而减少。结论在胃癌细胞BGC823中乙醇通过HIF-1α途径引起细胞缺氧,通过抑制抗凋亡蛋白Survivin促进细胞凋亡。

Melatonin combined with exercise cannot alleviate cerebral injury in a rat model of focal cerebral ischemia/reperfusion injury

Previous studies have demonstrated that melatonin combined with exercise can alleviate secondary damage after spinal cord injury in rats. Therefore, it is hypothesized that melatonin combined with exercise can also alleviate ischemic brain damage. In this study, adult rats were subjected to right middle cerebral artery occlusion after receiving 10 mg/kg melatonin or vehicle subcutaneously twice daily for 14 days. Forced exercise using an animal treadmill was performed at 20 m/min for 30 minutes per day for 6 days prior to middle cerebral artery occlusion. After middle cerebral artery occlusion, each rat received melatonin combined with exercise, melatonin or exercise alone equally for 7 days until sacrifice. Interestingly, rats receiving melatonin combined with exercise exhibited more severe neurological deficits than those receiving melatonin or exercise alone. Hypoxia-inducible factor la mRNA in the brain tissue was upregulated in rats receiving melatonin combined with exercise. Similarly, microtubule associated protein-2 mRNA expression was significantly upregulated in rats receiving melatonin alone. Chondroitin sulfate proteoglycan 4 （NG2） mRNA expression was significantly decreased in rats receiving melatonin combined with exercise as well as in rats receiving exercise alone. Furthermore, neural cell loss in the primary motor cortex was significantly reduced in rats receiving melatonin or exercise alone, but the change was not observed in rats receiving melatonin combined with exercise. These findings suggest that excessive intervention with melatonin, exercise or their combination may lead to negative effects on ischemia/reperfusion-induced brain damage.

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.

JMJD2C和HIF-1α在胃癌组织中的表达及其临床意义

目的 探讨含十字形结构域蛋白2C （JMJD2C） 和缺氧诱导因子1α（HIF-1α）在胃癌组织中的表达及其与临床病理特征的关系.方法 回顾性分析2005至2007年浙江省人民医院胃肠外科收治的110例胃癌患者的临床资料,其中男78例,女32例,平均年龄57（32～79） 岁,术前未经任何放化疗.通过免疫组织化学法检测110例原发性胃癌组织及80例癌旁正常胃黏膜组织中JMJD2C和HIF-1α蛋白表达情况.结果 JMJD2C约在69.1%（76例）的胃癌组织中呈高表达,正常胃黏膜组织中表达阴性（P〈0.05）;胃癌组HIF-1α的阳性表达率为73.6%（81/110）,而正常胃黏膜组织中无表达（P〈0.05）.胃癌组织中JMJD2C的表达与肿瘤的分化程度、浸润深度、临床TNM分期和淋巴结转移、远处转移密切均相关（均P〈0.05）,而与患者性别、年龄、肿瘤大小等因素均无关（均P＞0.05）.HIF-1α的表达与胃癌的TNM分期、浸润深度、淋巴结转移、远处转移等均有关（均P〈0.05）,而与年龄、性别、肿瘤大小、分化程度等因素均无关（均P＞0.05）.JMJD2C和HIF-1α表达呈正相关（r=0.219,P〈0.05）.Kaplan-Meier生存分析显示JMJD2C阳性组和HIF-1α阳性组平均生存时间均分别低于其阴性组[（38±4）比（56±6）个月,（38±4）比（60±6）个月,χ2=8.006、7.218,均P〈0.01];JMJD2C和HIF-1α双阳性组平均生存时间显著低于单阳性组和双阴性组（χ2=10.425,P〈0.01）.结论 JMJD2C和HIF-1α在胃癌组织中呈高表达,二者的表达呈正相关;JMJD2C和HIF-1α共同参与胃癌的发生,浸润,转移等生物学行为,可作为评价胃癌恶性程度和预后分析的新指标.

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.

缺氧诱导因子1α表达对裸鼠肝癌皮下移植瘤生长的促进作用

目的探讨缺氧诱导因子1α表达对肝癌移植瘤生长的影响。方法建立强力霉素诱导缺氧诱导因子1α表达的裸鼠肝癌HepG2^Tet-on-HIF-1α细胞皮下移植瘤模型；荷瘤裸鼠口服强力霉素（Dox）后，观察上调缺氧诱导因子1α对皮下移植瘤生长的影响。结果荷瘤裸鼠口服强力霉素可以上调裸鼠皮下移植瘤中缺氧诱导因子1α mRNA和蛋白表达水平；肿瘤体积Dox（＋）组vs．Dox（-）组为（513．545±276．229）mm^3 vs．（166．506±110．142）mm^3（P〈0．05），肿瘤重量（1．251±0．438）g vs．（0．640±0．296）g（P〈0．05），肿瘤生长速度Dox（＋）组明显超过Dox（-）组，肿瘤内面积坏死率明显小于Dox（-）组（31．360％±2．728％）vs．（36．640±3．804％）（P〈0．05）；同Dox（-）组相比，Dox（＋）组裸鼠体重下降更为明显（P〈0．01）。两组荷瘤鼠均无肝、肺转移发生。结论强力霉素可以诱导裸鼠肝癌HepG2^Tet-on-HIF-1α细胞皮下移植瘤模型中缺氧诱导因子1α的表达，促进肿瘤的生长。

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.

贝伐单抗联合SN-38抑制结肠癌细胞增殖的机制

目的探讨贝伐单抗联合伊立替康（CPT-11）的活性代谢产物SN-38对结肠癌细胞增殖的抑制作用及其机制。方法在缺氧条件下，将贝伐单抗和sN-38作用于人结肠腺癌细胞LoVo，应用四甲基偶氮唑蓝（MTT）法，检测药物对细胞增殖的抑制作用。分别采用逆转录聚合酶链反应（RT—PCR）、Western blot和酶联免疫吸附试验（ELISA），分析药物作用后缺氧诱导因子1α（HIF-1α）及血管内皮生长因子（VEGF）在基因和蛋白表达水平的变化；通过药物对丝裂原激活蛋白激酶（MAPK）和磷脂酰三磷酸肌醇-丝苏氨酸蛋白激酶（P13K-AKT）通路的影响，分析贝伐单抗和sN-38的联合作用机制。结果贝伐单抗联合sN-38序贯给药方案较同时给药对细胞增殖的抑制更强，且以先sN-38后贝伐单抗的给药顺序作用最强，IC50值为（0．057±0．009）μmol／L，较相反给药顺序强3．8倍，较同时给药强8．9倍。HIF—1α的表达水平随着缺氧时间的延长而升高，贝伐单抗和sN-38同时作用可显著抑制其表达；VEGF的表达也随着缺氧时间的增加逐渐增加，SN-38作用12h以上可使VEGF表达下降59．4％，200μg/ml的贝伐单抗作用8h几乎可完全抑制VEGF表达，此时加入SN-38仍可使VEGF表达进一步降低。MAPK通路的p-ERK表达随缺氧时间的增加而升高，sN-38和贝伐单抗同时作用可使p-ERK的表达完全阻断，这与两药对HIF-1α的抑制是一致的。ERK的选择性抑制剂PD98059能够增强贝伐单抗对p-ERK的抑制，同时也增加了贝伐单抗对HIF-1α的抑制。p-AKT的表达虽然随着缺氧时间的延长有所升高，但贝伐单抗和sN-38对其影响并不显著。结论VEGF靶向药物联合化疗药物对细胞增殖的抑制是方案依赖性的，贝伐单抗和sN-38通过对HIF-1α和MAPK通路的调控发挥协同作用。