AIMTo study the effects of ginkgolides (Gin) on the expression of hypoxia-inducible factor-1alpha (H1F-1alpha) in primary cultured cortical neurons treated with CoCl2 and the relationship with ERK signal pathway.

METHODSWe observed the effects of Gin (37.5 mg/L) on morphology and viability on primary cultured cortical neurons with treatment of CoCl2 (125 micromol/L). The expression of HIF-1alpha and p-ERK of neurons induced by CoCl2 pretreated with Gin were assessed by Western-blot. We analyzed the relationship between HIF-1alpha expression activated by Gin and ERK signal pathway with treatment of PD98059 (100 micromol/L), a selective inhibitor of ERK.

RESULTSIt was shown that Gin had protective effects on CoCl2 damaged neurons by raising the neuronal viability. Some basic expression of HIF-1alpha and p-ERK were observed in normal cultured cortical neurons. The expression of HIF-1alpha and p-ERK increased strikingly when treated with CoCl2 for 4 h. The levels of HIF-1alpha and p-ERK increased even more in the neurons pretreated with Gin for 24 h before CoCl2. The levels of HIF-1alpha and p-ERK were notably inhibited with pretreatment of PD98059, while Gin could prevent this inhibition.

CONCLUSIONGin has protective effects on neurons damaged by CoCl2 which might be related to the increase of the level of HIF-1alpha and the activation of ERK signal pathway.

Animals ; Cell Hypoxia ; drug effects ; Cells, Cultured ; Ginkgolides ; pharmacology ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; MAP Kinase Signaling System ; Mice ; Mice, Inbred ICR ; Neurons ; drug effects ; metabolism

Animals ; Animals, Newborn ; Cerebral Cortex ; drug effects ; metabolism ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Hypoxia-Ischemia, Brain ; metabolism ; Progesterone ; pharmacology ; Rats ; Rats, Sprague-Dawley

AIMTo study the effects of ginkgolides (Gin) on the expression of hypoxia inducible factor-1 (HIF-1alpha) in hypoxic/ischemic neurons.

METHODSThe gene expression of HIF-1alpha pretreated with or without Gin (37.5 microg/ml) was observed by RT-PCR on primary cultured cortical neurons in the condition of hypoxia and oxygen-glucose deprivation.

RESULTSSome basic expression of HIF-1alpha mRNA were observed in cultured cortical neurons. The expression of HIF-1alpha mRNA increased after 24 h treatment with Gin. The level of HIF-la mRNA increased also after 1 h hypoxia and further enhanced after the pretreatment with Gin. The expression of HIF-1alpha mRNA decreased with the deprivation of both oxygen and glucose, which reversed after the pretreatment of Gin.

CONCLUSIONGin could increase the expression of HIF-1alpha mRNA in hypoxic/ischemic cortical neurons.

Animals ; Cell Hypoxia ; Gene Expression ; Ginkgolides ; pharmacology ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; metabolism ; Mice ; Mice, Inbred ICR ; Neurons ; drug effects ; metabolism ; RNA, Messenger ; genetics

OBJECTIVETo investigate the effect of hypoxia on HIF-1alpha activation in intestinal epithelial cells.

METHODSIntestinal epithelial cells were randomly divided into normal control group, hypoxia group and hypoxia plus oligomycin group (oligomycin group). In hypoxia group, the cells were exposed to hypoxia for 1, 2, 6, 12 and 24 h. In oligomycin group, the cells were treated with oligomycin in concentration of 5, 10, 20 and 40 microg/mL for 1 h prior to 6-hour hypoxic exposure. HIF-1alpha protein expression was assayed by western blot method. Nuclear translocation of HIF-1alpha was detected by immunofluorescence analysis.

RESULTSCompared with that in control group (0.08 +/- 0.07), HIF-1alpha protein expression in hypoxia group increased significantly at 1 h (0.52 +/- 0.30, P < 0.05), and reached the peak value (2.37 +/- 1.08, P < 0.05) at 6 h. Nuclear translocation of HIF-1alpha was also induced by hypoxia. HIF-1alpha protein expression in oligomycin group in the concentration of 5, 10, 20 and 40 microg/mL of oligomycin was 1.62 +/- 0.96, 1.48 +/- 0.56, 1.08 +/- 0.36 and 0.58 +/- 0.11 respectively, which was significantly lower than that only after exposure to hypoxia for 6 h (2.67 +/- 1.38, P < 0.05). The nuclear translocation of HIF-1alpha induced by hypoxia was also obviously inhibited by oligomycin pretreatment.

CONCLUSIONOligomycin, a specific inhibitor of respiratory chain, inhibits HIF-1alpha activation, which suggests that mitochondria respiratory chain may play an important role in aforementioned process.

Blotting, Western ; Cell Hypoxia ; Cells, Cultured ; Epithelial Cells ; drug effects ; metabolism ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Intestines ; cytology ; Oligomycins ; pharmacology

OBJECTIVETo explore the effects of stanniocalcin-1 (STC-1) and hypoxia-inducible factor-1&alpha; (HIF-1&alpha;) on the calcium and thus on the mitochondrial membrane potential (Δψm) in renal carcinoma cells.

METHODSWe successfully established the renal carcinoma cell models with high HIF-1&alpha; gene expression. After various concentrations of STC-1 solutions were added to the culture medium, the proliferation of cells, expressions of HIF-1&alpha; and STC-1, levels of Ca(2+), Δψm, and mPTP were detected by MTT, RT-PCR, ELISA, fluorescence spectrophotometry, and ultraviolet spectrophotometry, respectively.

RESULTSThe proliferation of renal carcinoma cells and Δψm were improved after HIF-1&alpha; gene transfection, STC-1 protein intervention, and STC-1 protein intervention after gene transfection. While the intracellular Ca(2+) level and mPTP were decreased significantly (P<0.05), all the changes were intensified with the gradual increase of STC-1. However, the increasing trend of cell proliferation gradually declined.

CONCLUSIONHIF-1&alpha; may participate in malignant proliferation of renal carcinoma cells by promoting STC-1 proliferation or down-regulating Ca(2+); however, such an effect may be gradually attenuated due to the inhibitory effect of STC-1 on HIF-1&alpha;.

Calcium ; metabolism ; Carcinoma, Renal Cell ; pathology ; Glycoproteins ; pharmacology ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Membrane Potential, Mitochondrial ; drug effects ; Tumor Cells, Cultured

The effects of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor apocynin on the enhanced hypoxia induced factor-1&alpha; (HIF-1&alpha;) and endothelin-1 (ET-1) expression, elevated systolic blood pressure under chronic intermittent hypoxia (CIH) condition and its action mechanism were investigated. Thirty healthy 8-week old Sprague-Dawley (SD) male rats were randomly divided into three groups (n=10 each): sham group, CIH group, and apocynin-treated CIH group. Tail artery systolic blood pressure was measured by tail-cuff method. Real-time fluorescence quantitative polymerase chain reaction (PCR) was used to detect the mRNA expression of HIF-1&alpha; and ET-1 in the carotid body, and the HIF-1&alpha; protein expression was examined by using Western blotting. The levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were determined by using colorimetric method. In addition, the plasma ET-1 and HIF-1&alpha; levels were measured by using enzyme-linked immunosorbent assay. It was found that CIH exposure was associated with increased MDA levels, and apocynin-treated CIH animals showed reduction in MDA levels. Apocynin treatment prevented CIH-induced hypertension as well as CIH-induced decrease in SOD. The increases of HIF-1&alpha; and ET-1 mRNA along with HIF-1&alpha; protein expression in the carotid body, and elevated circulating HIF-1&alpha; and ET-1 levels were observed in CIH-exposed animals. Treatment with apocynin significantly decreased the ET-1 mRNA, HIF-1&alpha; protein expression and circulating HIF-1&alpha; level in CIH-exposed animals, and there was no statistically significant difference in the HIF-1&alpha; mRNA expression between CIH group and apocynin-treated group. These results indicated that apocynin alleviated CIH-induced hypertension by inhibiting NADPH oxidase, further leading to the reduced vasoconstrictor ET-1 level and oxidative stress. HIF-1&alpha;/ET-1 system signal pathway may interact with CIH-induced NADPH oxidase-dependent oxidative stress. Inhibition of NADPH oxidase activity may hopefully serve as a useful strategy for prevention and treatment of obstructive sleep apnea hypopnea syndrome-induced hypertension.
Acetophenones ; administration & dosage ; Animals ; Antioxidants ; administration & dosage ; Carotid Body ; drug effects ; metabolism ; Endothelin-1 ; metabolism ; Hypoxia ; drug therapy ; metabolism ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Male ; NADP ; antagonists & inhibitors ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species ; metabolism ; Treatment Outcome

OBJECTIVETo study the effect of hypoxia on hypothalamus-adenohypophysis-testis axis hormone levels, germ cell apoptosis and hypoxia-inducible factor-1alpha (HIF-1alpha) expression in testis of adolescent mice, and explore HIF-1alpha regulation on the reproductive function of male mice.

METHODSEighty SPF grade adolescent C57BL/6 mice were randomly divided into normoxia group, hypoxia 3, 7, 14 and 28 d groups. The level of serum testosterone (T), free testosterone (FT), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) was analyzed by ELISA. Detected the sperm count, motility rate and abnormal sperm rate of epididymal sperm suspension. The apoptosis cells in testis were determined using TUNEL method. The expression of HIF-1alpha was analyzed using Western blot.

RESULTSCompared with corresponding normoxia group, serum T, FT, FSH and LH concentrations in hypoxia 3 d group were significantly higher (P < 0.05); T and LH concentrations in hypoxia 14 d group were significantly lower (P < 0.05). Sperm count and motility rate in hypoxia 7 and 14 d groups significantly declined (P < 0.05); abnormal sperm rate in all hypoxia groups significantly increased (P < 0.05). The apoptosis index (AI) of germ cells in hypoxia 7, 14 and 28 d groups significantly increased (P < 0.05), and the levels of HIF-1alpha protein expression were significantly higher (P < 0.05).

CONCLUSIONHIF-1alpha protein highly expressed in mice testis could induce germ cell apoptosis increased in chronic hypoxia environment.

Animals ; Apoptosis ; drug effects ; Germ Cells ; drug effects ; metabolism ; Gonadal Steroid Hormones ; metabolism ; Hypoxia ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Testis ; cytology ; metabolism

OBJECTIVETo investigate the effect of amphotericinB (AmB) on migration and invasion of esophageal carcinoma Eca109 cells exposed to hypoxia and explore the molecular mechanisms.

METHODSRoutinely cultured esophageal carcinoma Eca109 cells were treated with 0, 1.25, 2.5, or 5 µg/ml AmB in hypoxic condition (3% O2, 5% CO2, and 92% N2) for 24 h. The cell migration and invasion were assessed by cell scratch test and Transwell chamber assay, respectively. Real-time quantitative PCR and Western blotting were used to detect the mRNA and protein expressions of hypoxia-inducible factor-1&alpha; (HIF-1&alpha;), matrix metalloproteinase-2 (MMP-2), and E-cadherin in the cells, respectively.

RESULTSCompared with the control cells, the cells treated with different doses of AmB showed attenuated ability of migration and invasion (P<0.05). AmB treatment resulted in significantly lowered mRNA and protein expressions of MMP-2 (P<0.05) and increased expressions of E-cadherin (P<0.05); the protein expression of HIF-1&alpha; decreased significantly in cells after AmB treatment (P<0.05) but its mRNA levels showed no significant changes (P>0.05).

CONCLUSIONAmB can suppress the migration and invasion of esophageal carcinoma Eca109 cells in hypoxic microenvironment possibly by regulating the expressions of HIF-1&alpha;, MMP-2 and E-cadherin.

Amphotericin B ; pharmacology ; Cadherins ; metabolism ; Cell Hypoxia ; Cell Line, Tumor ; drug effects ; Cell Movement ; drug effects ; Down-Regulation ; Esophageal Neoplasms ; metabolism ; pathology ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; RNA, Messenger

OBJECTIVETo study the expression of HIF-1alpha and SDF-1/CXCR4 in repopulating H22 tumor tissue and the mechanism of angiogenesis of polypeptide extract from scorpion venom (PESV) during chemotherapy treatment.

METHODThe expression of HIF-1alpha and SDF-1/CXCR4 in H22 tumor tissue was monitored by immunohistochemistry, and the expression level was determined by Qwin V3 image analyzing software. The correlation between HIF-1alpha and SDF-1 was analyzed. SDF-1 content was detected by ELISA.

RESULTHIF-1alpha expression was found no difference in model group between 14 d and 21 d, and up-regulated in 28 d. There was no change of HIF-1alpha expression was observed in low-dose PESV group. In high-dose PESV group, the level of HIF-1alpha expression was high in 14 d and low in 21 d. ELISA detecting showed SDF-1 content increased slowly from 14 d to 21 d, highly from 21 d to 28 d. But in high-dose PESV groups, the content increased slowly all the time. The immunohitochemistry method got the same result with ELISA. Correlation analysis showed r = 0.805. CXCR4 expression down-regulated in two PESV treated groups, and no difference was found between these two groups.

CONCLUSIONHIF-1alpha and SDF-1 participated in VEGF expression and angiogenesis in tumor tissue during chemotherapy, while PESV could inhibit the expression of HIF-1alpha and SDF-I.

Animals ; Cell Line, Tumor ; Chemokine CXCL12 ; drug effects ; metabolism ; Down-Regulation ; drug effects ; Hypoxia-Inducible Factor 1, alpha Subunit ; drug effects ; metabolism ; Mice ; Peptides ; pharmacology ; Receptors, CXCR4 ; drug effects ; metabolism ; Scorpion Venoms ; chemistry ; pharmacology ; Scorpions ; chemistry ; Time Factors

The present study was designed to determine the effects of a traditional Chinese medicine, called Qishen Yiqi Dropping Pill on chronic hypoxia-induced myocardial injury. To establish a rat chronic hypoxia model to be used in the evaluation of the therapeutic effects of the Qishen Yiqi Dropping Pill, Sprague-Dawley (SD) rats were randomly divided into three groups: the control, model, and treatment groups (n = 10 per group). The animals were housed in a plexiglass container. The control animals were under normal oxygen concentration and the model and treatment groups were exposed to air and nitrogen for 5 weeks. The rats in the treatment group were orally administered the Qishen Yiqi Dropping pill (35 mg·kg(-1)·d(-1)) for 5 weeks. After the treatment, the cardiac function and morphology were analyzed, and the expression levels of hypoxia-inducible factor 1α (HIF-1α) were determined using Western blotting. Our results indicated that the cardiac function was impaired, cell apoptosis was enhanced, and HIF-1α expression was up-regulated in the model group, compared to the control group. These changes were ameliorated by the treatment with the Qishen Yiqi Dropping Pill. In conclusion, Qishen Yiqi Dropping pill can ameliorate myocardial injury induced by chronic hypoxia, improve cardiac function, and decrease myocardial cell apoptosis, which may provide a basis for its clinical use for the treatment of chronic cardiovascular diseases.
Animals ; Apoptosis ; Cardiomyopathies ; drug therapy ; etiology ; metabolism ; pathology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Heart ; drug effects ; physiopathology ; Hypoxia ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Myocardium ; metabolism ; pathology ; Oxygen ; metabolism ; Phytotherapy ; Rats, Sprague-Dawley