共查询到20条相似文献，搜索用时 15 毫秒
1.
AIM: To study the effect of hypoxia inducible factor-1 alpha (HIF-1α) on tumor necrosis factor alpha (TNF-α) production in rat alveolar macrophages cultured under hypoxic condition. METHODS: Using HIF-1α decoy inhibiting its function, Immunohistochemistry, Western blot, semiquantitative RT-PCR and ELISA were used to determine the expression of HIF-1α protein and mRNA and the production of TNF-α in rat alveolar macrophages cultured under hypoxic condition (3% O2, 5% CO2, 92% N2), respectively. RESULTS: Expression of HIF-1α was positive in cultured macrophage nucleoli in hypoxia group and HIF-1α decoy group but it was negative in nomoxic control group. The content of HIF-1α protein in hypoxia group and HIF-1α decoy group were significantly higher than that in nomoxic control group (P<0.05). The content of HIF-1α mRNA in hypoxia group and HIF-1α decoy group were markedly higher than that in nomoxic control group (P<0.05), respectively. The content of TNF-α in hypoxia group (115±17 ng/L) was higher than that in control group ［（69±13） ng/L, P<0.05］ and HIF-1α decoy group ［（81±15） ng/L, P<0.05］. CONCLUSION: Hypoxia can increase significantly expression and activity of HIF-1α， which can promote the production of TNF-α in rat alveolar macrophages. It suggests that HIF-1α plays an important role in the pathogenesis of chronic inflammation-related diseases that can give rise to lung hypoxia such as COPD. 相似文献
2.
AIM: To investigate the cell cycle arrest induced by hypoxia, hypoxia inducible factor-1 and their possible mechanism in human ovarian cancer cell line SW626. METHODS: CoCl2, a chemical inducer of hypoxia and hypoxic cell culture chamber were used to induce chemical and physical hypoxia in human ovarian cancer cell line SW626. The method of ‘decoy’ was used to block the function of HIF-1α because it acts as the core sequence of the target gene as a competitor combined to the HIF-1α. The cells were divided into group A1 (normal oxygen), A2 (normal oxygen plus HIF-1α decoy), B1 (CoCl2), B2 (CoCl2 plus HIF-1α decoy), C1 (hypoxia) and C2 (hypoxia plus HIF-1α). The expression of the HIF-1α protein, mRNA and cell cycle analysis were detected by Western blotting, RT-PCR and flow cytometry (FCM). RESULTS: The expression level of HIF-1α protein in group B1 (3.75±1.31) and group C1 (3.48±1.01) was significantly higher than that in group A1 (0.97±0.31) (P<0.05). The expression levels of HIF-1α mRNA in group A1 (0.65±0.32) and group B1 (0.64±0.34) were significantly lower than that in group C1 (1.28±0.62) (P<0.05). Decoy had no effect in the expression of HIF-1α protein and mRNA level (P>0.05). FCM showed that the G0/G1 phase was markedly increased in group B1 (81.78±24.33) and group C1 (77.62±22.76) and was significantly higher than that in group A1 (49.49±18.54) (P<0.05), group B2 (61.54±20.84) was lower than that in group B1 with statistical significance (P<0.05) and group C2 (56.03±21.42) was lower than that in group C1 with statistical significance (P<0.05) , but the difference between group A1 and group A2 (51.77±16.45) had no statistical significance (P>0.05). CONCLUSION: Both CoCl2 and physical hypoxia could distinctly induce cell cycle arrest in G0/G1 phase and the expression of HIF-1α in human ovarian cancer cell line SW626. HIF-1α plays an important role in cell cycle arrest induced by hypoxia in human ovarian cancer cell line SW626. 相似文献
3.
AIM:To investigate relationship among phosphatidylinositol 3-kinase (PI3K) and hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) in lung of rats with hypoxia-inducible pulmonary hypertension. METHODS:Forty male adult Wistar rats were randomly divided into five groups (eight rats in each group):control group (C group) and groups with hypoxia for 3, 7, 14 and 21 days (H3, H7, H14 and H21 group). Mean pulmonary arterial pressure (mPAP), right ventric hypertrophy index (RVHI) and vessel morphometry were measured. The levels of HIF-1α mRNA expression in lung tissue was measured by in siteu hybridization (ISH). The protein expression of HIF-1α，VEGF and phosphorylated protein kinase β (P-AKT) were observed by immunohistochemistry or Western blotting. RESULTS:mPAP increased significantly 7 days after hypoxia ［(23.53±1.78) mmHg］, peaked 14 days after hypoxia, then remained on the high level. Pulmonary artery remodeling index (extern diameter 100 μm) and RVIH became evident 14 days after hypoxia. Expression of P-AKT protein in control group was poorly positive, but was up-regulated in pulmonary arterial tunica intima and tunica media in all hypoxia rats. HIF-1α mRNA staining was poorly positive in control，hypoxia for 3 days and hypoxia for 7 days, but began to increase significantly 14 days after hypoxia (0.305±0.104, P<0.05), then remained stable. Expression of HIF-1α protein in control group was poorly positive, but was up-regulated in pulmonary arterial tunica intima in all hypoxic rats. In pulmonary arterial tunica media, the levels of HIF-1α protein was markedly up-regulated after 3 days (0.029±0.019, P<0.05 ), reached its peak 7 days after hypoxia (0.232±0.008, P<0.05), then tended to decline 14 days and 21 days after hypoxia. Expression of VEGF protein began to increase 7 days after hypoxia (0.188±0.018, P<0.05), reached its peak 14 days after hypoxia (0.238±0.017, P<0.05), then remained on the high level in pulmonary arterial tunica intima. Linear correlation analysis showed that P-AKT, HIF-1α mRNA, VEGF and mPAP were correlated with vessel the morphometry and RVHI (P<0.01). P-AKT was positively correlated with HIF-1α and VEGF (tunica intima). CONCLUSION:P-AKT, HIF-1α and VEGF are all involved in the pathogenesis of hypoxia-induced pulmonary hypertension in rats. 相似文献
4.
AIM: To investigate the effect of hypoxia on the proliferation and apoptosis of pulmonary artery smooth muscle cells (PASMC); and to evaluate the role of hypoxia-inducible factor-1α(HIF-1α), iNOS, P-ERK1/2 protein expression in hypoxic pulmonary hypertension (HPH) pathogenesis.METHODS: Cultured rat PASMC were divided into normoxic group; hypoxic group; hypoxia+ADM(adrenomedulin) group, hypoxia+L-NAME(iNOS inhibitor) group; hypoxia+PD98059 group. Proliferation was investigated by MTT and PCNA. Apoptosis was examined by flow-cytometry. Westen blotting was used to measure protein expression of HIF-1α, P-ERK1/2 and iNOS. RESULTS: (1) A value of 24 h-hypoxia was significantly higher than that in the normoxic group (P<0.01). In the hypoxia+PD98059 group, ADM was significantly lower than that in hypoxia group, whereas A value of the hypoxia+L-NAME was significantly higher than that in hypoxic group and normoxic group (P<0.01). (2) PCNA was positive in PASMC after 24 h hypoxia (P<0.01). PD98059, ADM inhibited the expression of PCNA significantly (P<0.01), whereas L-NAME increased the expression of PCNA significantly (P<0.01). (3) Apoptosis index was not significantly difference among the different groups (P>0.05). (4) HIF-1α, iNOS and P-ERK1/2 expression was poorly positive in normoxic group, positive after hypoxia for 4h (P<0.01), reaching its peak at 8 h hypoxia (P<0.01), HIF-1α, P-ERK1/2 expression declined after 24 h hypoxia. L-NAME promoted the expression of HIF-1α, PD98059 inhibited the expression of HIF-1α, iNOS and P-ERK1/2 partly. ADM inhibited the expression of HIF-1α partly, promoted the expression of iNOS. CONCLUSION: (1) Hypoxia stimulates the proliferation of PASMC, and has no obvious effects on the apoptosis of PASMC. (2) HIF-1 plays an importent role in the proliferation of hypoxic PASMC. 相似文献
5.
AIM: To explore the differences in the effects of 2 representative hypoxia models on human breast cancer MDA-MB-231 cells. METHODS: To establish a chemical hypoxia model, MDA-MB-231 cells were treated with CoCl2 at different concentrations (0~300 μmol/L) for different time (24 h, 48 h and 72 h). On the other hand, MDA-MB-231 cells were exposed to hypoxia with different volume fractions (1%, 2% and 5%) of oxygen for different time (4 h, 16 h and 24 h) to establish a physical hypoxia model. The viability of the cells in the 2 hypoxia models was measured by CCK-8 assay. The protein levels of hypoxia-inducible factor-1α (HIF-1α), Bcl-2 and matrix metalloproteinase-2 (MMP-2) were determined by Western blot. The apoptosis of the cells was analyzed by flow cytometry. The cell invasion and migration were examined by Transwell assay and wound-healing assay, respectively. RESULTS: The HIF-1α was expressed in time- and dose-dependent manners. These results indicated that treatment of MDA-MB-231 cells with CoCl2 at 100 μmol/L for 24 h served as the final chemical hypoxia model, and exposure of MDA-MB-231 cells to hypoxia with 2% oxygen for 24 h served as the final physical hypoxia model. Compared with the cells with chemical hypoxia, the protein le-vel of HIF-1α was significantly increased in the cells with physical hypoxia. In addition, reoxygenation from hypoxia caused a rapidly degraded HIF-1α expression level in physical hypoxia model. Flow cytometry results showed that apoptosis was increased and the protein expression level of anti-apoptotic Bcl-2 was decreased in the cells with physical hypoxia, while no significant change was observed in the cells with chemical hypoxia. The protein expression level of MMP-2 in both hypoxia models was increased, and the invasion and migration capabilities of the cells with chemical hypoxia were enhanced. CONCLUSION: Two hypoxia models were successfully constructed. There were differences in the expression and stability of HIF-1α protein, and the viability, apoptosis, invasion and migration of the cells in the 2 different hypoxia models, which provide references for the selection of hypoxia models and the further study of characteristics of tumor cells in hypoxia environment. 相似文献
6.
Effects of HIF-1α/SDF-1 signaling axis on hypoxia-induced migration and adhesion of progenitor cells
AIM: To observe the expression of hypoxia-inducible factor-1alpha (HIF-1α) and stromal cell-derived factor 1 (SDF-1) in pulmonary artery endothelial cells (PAECs) of SD rats and to investigate the role of HIF-1α/SDF-1 signaling axis on hypoxia-induced migration and adhesion of progenitor cells to PAECs. METHODS: Immunomagnetic beads were used to separate and purify the CD34+/CXCR4+ progenitor cells derived from the peripheral circulation of SD rats. The expression of HIF-1α and SDF-1 in PAECs exposed to hypoxia (1% O2, 5% CO2 and 94% N2) was detected by immunofluorescence, Western blotting and ELISA. The migration index and adhesion rate were measured in the progenitor cells, which were subjected to the following different treatments: (1) normoxia (21% O2); (2) hypoxia 12 h; (3) hypoxia 12 h +HIF-1α inhibitor (2ME2); (4) hypoxia 12 h+SDF-1 neutralizing antibody; (5) hypoxia 12 h+2ME2+SDF-1 neutralizing antibody.RESULTS: The expression of HIF-1α and SDF-1 in PAECs was effectively induced by the hypoxic exposure, and both of them reached the peak levels after 12 h of hypoxic treatment (P<0.01), while administration of 2ME2 decreased the hypoxia-induced SDF-1 expression (P<0.05). Treatment of the PAECs with 2ME2 or SDF-1 neutralizing antibody attenuated the migration index and adhesion rate of progenitor cells to the PAECs (P<0.05). CONCLUSION: There is a HIF-1α/SDF-1 signaling axis in hypoxia-exposed PAECs, which may play a crucial role in the migration and adhesion of progenitor cells to PAECs. 相似文献
7.
8.
AIM: To investigate the expression of hypoxia inducible factor-1alpha (HIF-1α) and the role of HIF-1α in tumor necrosis factor alpha (TNF-α) production in rat alveolar macrophages activated by lipopolysaccharide (LPS). METHODS: HIF-1α function was inhibited by using the method of HIF-1α decoy. Western blotting and semiquantitative RT-PCR were applied to determine the expression of HIF-1α protein and mRNA, respectively. The production of TNF-α was determined with ELISA. RESULTS: The content of HIF-1α protein in LPS group (1.95±0.57) and HIF-1α decoy group (1.89±0.59) were 4.8 times and 4.6 times higher than that in control group (0.41±0.14), respectively. The expression of HIF-1α mRNA showed no difference among three groups (F=3.14，P>0.05). The production of TNF-α in LPS group was higher than that in control group (61 ng/L vs 156 ng/L, q=5.12, P<0.05) and HIF-1α decoy group (90 ng/L vs 156 ng/L, q=4.63, P<0.05), respectively. However, the content of TNF-α in HIF-1α decoy group was still higher than that in control group (61 ng/L vs 94 ng/L, q=4.47, P<0.05). CONCLUSION: The enhanced stability of HIF-1α protein results in the marked upregulation of its protein and HIF-1α is contributed to the production of TNF-α in LPS-stimulating rat alveolar macrophages. It is indicated that HIF-1α plays important role in the pathogenesis of chronic inflammation involved in diseases such as COPD. 相似文献
9.
AIM: To observe the effects of Tongxinluo (TXL), a Chinese medicine, on hypoxic tolerance and expression of Bcl-associated X (Bax) and B-cell leukemia/lymphoma 2 (Bcl-2) in hypoxia-preconditioned mice. METHODS: The mice were randomly divided into groups of hypoxia preconditioning without (control group) or with TXL treatment (TXL group). The mice in TXL group were administered with TXL at dose of 1.52 g·kg-1·d-1 crude drug for 5 days. The mouse was exposed to normoxia (0 run, H0) and acute repetitive hypoxia for 1-5 runs (H1-H5) by placing the animal in an air-sealed jar. The hypoxic environment was established in the jar through consumption of the oxygen by the respiration of the mouse. A gasp breath was regarded as the hypoxic tolerant limit of the mouse and the animal was then transferred to another new jar. The mouse was exposed to hypoxia in this way for 5 times. In each run of hypoxic exposure, the time of hypoxic tolerance was measured. Western blotting was used to measure the protein levels of hypoxia inducible factor-1α (HIF-1α), Bax and Bcl-2 in the cerebral cortex. RESULTS: The hypoxic tolerance time in control and TXL groups was gradually increased run by run (P<0.01 or P<0.05). The protein levels of HIF-1α and Bcl-2 in the two groups were gradually increased (P<0.01 or P<0.05). Bax in control and TXL groups was significantly increased in H1. After H1, Bax was decreased run by run (P<0.01 or P<0.05). Compared with control group, the tolerance time, the expression of HIF-1α and Bcl-2 in the cerebral cortex in TXL group were increased in H1,H3 and H5. However, the expression of Bax was lower than that in control group in every run. CONCLUSION: Hypoxia preconditioning makes the organism produce a strong adaptive response. The increase in Bcl-2 and the decrease in Bax may be involved in the mechanism of adaptation. TXL obviously increases the adaptive ability of the mice to hypoxia. 相似文献
10.
11.
AIM: To investigate the relationship between the expression of hypoxia-inducible factor-1α (HIF-1α) and myocardial angiogenesis induced by hypoxic preconditioning (HPC). METHODS: Male Sprague-Dawley rats were randomly divided into control group and hypoxic-preconditioned group. Rats in preconditioned group were subjected to hypoxic exposure (10% O2 and 90% N2) for 4 h, these rats were maintained in a normal environment respectively for 1 d, 7 d or 21 d. Then, vascular density was observed by the method of immunohistochemical staining with the antibody against factor Ⅷ-related antigen in rat myocardial tissue. Protein extracts of rat myocardial tissue were prepared for Western blotting analysis to measure the activities of extracellular signal-regulated protein kinases (ERKs) and HIF-1α. RESULTS: In hypoxic-preconditioned group, microvascular density significantly increased in myocardial tissue. The expression of ERK1/2 increased by 18.67% at 1 d after HPC. HPC induced the expression of HIF-1α, and the peak of expression was at first day after HPC. CONCLUSION: HPC promotes myocardial angiogenesis. Activation of ERKs and expression of HIF-1α might be involved in myocardial angiogenesis induced by hypoxic preconditioning. 相似文献
12.
AIM: To observe the expression of hypoxia inducible factor-1α (HIF-1α) gene and heme oxygenase-1 (HO-1) gene in pulmonary arteries in hypoxic rats. METHODS: Forty male Wistar rats were exposed to hypoxia for 0, 3, 7, 14 or 21 days. Mean pulmonary pressure (mPAP), vessel morphometry, right ventricle hypertrophy index (RVHI) were measured. Lungs were either inflation fixed for immunohistochemistry and in situ hybridization or frozen for later measurement of HO-1 enzyme activity. RESULTS: During hypoxia, mPAP increased to significantly higher values than the control values after 7-day of hypoxia，reaching its peak after 14-day of hypoxia, then remained on the high level. Pulmonary artery remodeling developed significantly after 14-day of hypoxia. Expression of HIF-1α protein in control was poorly positive, but was up-regulated in pulmonary arterial tunica intimae of all hypoxic rats. In pulmonary arterial tunica media, the levels of HIF-1α protein were markedly up-regulated after 3-day and 7-day of hypoxia, then tended to decline after 14-day and 21-day of hypoxia. HIF-1α mRNA staining was poorly positive in control, hypoxia for 3 days and hypoxia for 7 days, but began to enhance significantly after 14-day of hypoxia, then remained stable. Expression of HO-1 protein began to increase after 7-day of hypoxia, reaching its peak after 14-day of hypoxia, then remained stable. Expression of HO-1 mRNA began to increase after 3-day of hypoxia, reaching its peak after 7-day of hypoxia, then declined. CONCLUSION: HIF-1α and HO-1 are both involved in the pathogenesis of hypoxia-induced pulmonary hypertension in rats. Furthermore, HIF-1α may inter-regulate with HO-1 gene in this process. 相似文献
13.
HUANG Hai-chang YU Ying LIANG Yan MIN Ya-li HU Zhao CHEN Min LI Jing-zi WANG Hai-yan 《园艺学报》2005,21(3):432-435
AIM: To study the expression of connective tissue growth factor (CTGF) induced by hypoxia, and the role and mechanism of hypoxia on promoting renal interstitial fibrosis. METHODS: Renal interstitial fibrosis was induced by unilateral ureteral obstruction (UUO) in rat animal model for 9 days as in vivo studies; marker of hypoxia-HIF-1α mRNA and protein, the expression of CTGF in the obstructed kidneys were assessed by RT-PCR, immunohistochemistry and Western blotting respectively. In vitro, normal rat kidney interstitial fibroblast cells (NRK-49F) were exposed to hypoxia (1%O2) for up to 6 hours, hypoxia was confirmed by detecting the expression of HIF-1α protein in cells, cellular level of CTGF mRNA and protein were assessed by RT-PCR and Western blotting respectively. RESULTS: Neither HIF-1α mRNA nor HIF-1α protein was expressed in the kidney from sham-operated group of rats. High level of HIF-1α mRNA were occurred, and strongly HIF-1α positive immunostaining were seen in the tubular and interstitial cells in kidney from UUO rats. Expression and location of CTGF protein were paralleled and relevant with the expression of HIF-1α protein in kidney of UUO rats. In cultured NRK-49F cell line, subjected to hypoxia even for 6 hours stimulated the expression of CTGF mRNA and protein. CONCLUSION: Our results indicated that hypoxia could stimulate the expression of CTGF mRNA and protein in kidney from UUO rats, which may in turn contribute to renal interstitial fibrosis. 相似文献
14.
AIM: To study the effect of hypoxia-inducible factor 1α(HIF-1α) silencing on the expression of p27 and Ki67 in rat hepatoma cell line CBRH-7919 under hypoxia. METHODS: Hypoxic condition was induced by CoCl2 and the expression of HIF-1α was silenced by small interference RNA. HIF-1α-specific RNAi lentiviral vector was constructed. Real-time RT-PCR and Western blotting were used to detect the expression of HIF-1α at mRNA and protein levels in CBRH-7919 cells under hypoxia. The expression of p27 and Ki67 was observed by Western blotting after HIF-1α silencing was performed. The cell cycle of hepatoma cells was detected by flow cytometry. RESULTS: Under hypoxic condition, the expression of HIF-1α at mRNA and protein levels in CBRH-7919 cells increased significantly (P<0.05). HIF-1α silencing significantly reduced the expression of Ki67 but increased the expression of p27 (P<0.05) in CBRH-7919 cells. In transfected cells, the number of cells in G0/G1 phase was much higher and that in S phase was much lower than those in the control cells. CONCLUSION: Hypoxia induces the expression of HIF-1α. HIF-1α silencing can regulate the proliferation of hepatoma cells through reducing the expression of Ki67 and increasing the expression of p27. 相似文献
15.
CAI Xiao-hong HONG Fang-fang CHEN Li-ya MEI Hong-fang LIN Jing WANG Hong-xia LI Xiu-cui LIANG Dong-shi WEN Zheng-wang 《园艺学报》2016,32(1):187-192
AIM: To establish and validate a novel model of cultured cells for imitating intermittent hypoxia. METHODS: In a chamber with experiment cabin and simulated air control cabin, the frequency and duration of the intermittent hypoxia model according to the time of hypoxia and reoxygenation were evaluated. The A549 cells were randomly divided into 7 groups, named as control (Con) group, 6 h intermittent hypoxia (6IH) group, 9 h intermittent hypoxia (9IH) group, 6 h simulated air control (6AC) group, 9 h simulated air control (9AC) group, 4 h sustained hypoxia (4SH) group, 6 h sustained hypoxia (6SH) group, respectively. When the model was established, the cellular morphology was observed under inverted microscope. The mRNA expression of hypoxia-inducible factor (HIF)-1α was detected by real-time PCR. The protein expression of HIF-1α was analyzed by immunohistochemistry. RESULTS: The intermittent hypoxia cycle (5% O2 60 min-20% O2 30 min for 6 cycles) was established. The damaged A549 cells were observed in 6IH group, 9IH group and 6SH group. Compared with 6IH group, the expression of HIF-1α at mRNA and protein levels was significantly increased in 9IH group (P<0.05). The expression of HIF-1α at mRNA and protein levels in 6IH group and 9IH group was higher than that in 4SH group and 6SH group, respectively (P<0.05). No significant difference among the control group, 6AC group and 9AC group was found. CONCLUSION: The model (5% O2 60 min-20% O2 30 min for 6 cycles) can simulate the pathological process of obstructive sleep apnea hypopnea syndrome. This model is suitable for studying intermittent hypoxia in adherent cells. 相似文献
16.
17.
LIU Li-xuan WU Ling-fei DENG Wei ZHOU Xiao-tao CHEN Rui-pei XIANG Meng-qi GUO Yi-tian PU Ze-jin LI Guo-ping 《园艺学报》2014,30(12):2155-2160
AIM:To investigate the effects of tanshinone IIA (Tan IIA) on proliferation, apoptosis and its molecular mechanism in human hepatoma HepG2 cells under hypoxic condition. METHODS:Hypoxia model was established by treatment with cobalt chloride (CoCl2). The cells were divided into normoxia control group, hypoxia control group and hypoxia combined at different concentrations of Tan IIA groups. After HepG2 cells were incubated with different concentrations of Tan IIA (0.5, 1.0, 2.0, 5.0 and 10.0 mg/L) for 24 h, 48 h and 72 h under hypoxic condition, the cell proliferation was determined by MTT assay. After Tan IIA was added to the media at different concentrations for 24 h and 48 h, the apoptotic cells were observed by Hoechst 33258 staining. The protein levels of hypoxia-inducible factor 1 alpha (HIF-1α), vascular endothelial growth factor (VEGF) and wild-type P53 were detected by Western blotting after cultured with different concentrations of Tan IIA for 48 h. RESULTS:Tan IIA inhibited the proliferation of HepG2 cells in a dose- and time-dependent manner. Tan IIA induced the typical morphology of apoptotic cells and increased the apoptotic rate in a dose- and time-dependent manner after treatment with 1.0 mg/L~5.0 mg/L for 24 h and 48 h under hypoxic condition. The protein levels of HIF-1α and VEGF were weakly expressed in HepG2 cells under normoxia but up-regulated after incubated under hypoxia for 48 h. The protein expression of HIF-1α and VEGF were decreased with the increase in the concentration of Tan IIA under hypoxia. The protein expression of wild-type P53 was increased with the increase in the concentrations of Tan IIA under hypoxia. CONCLUSION: Tan IIA significantly inhibits the proliferation and induces the apoptosis of human hepatoma HepG2 cells under hypoxia, which may be related to the down-regulation of HIF-1α and VEGF and up-regulation of wild-type P53. 相似文献
18.
LIANG Jun-qing XU Hai-bo CHEN Xiao-juan XING Zhi-jun YIN Bei-pei WU Kun ZHANG Zi-qian ZHOU Li-tao WU Yi-ling 《园艺学报》2012,28(5):846-851
AIM: To investigate the effects of PI-3K/Akt/HIF pathway on anti-hypoxia ability of vascular endothelial cells influenced by Tongxinluo under hypoxic condition. METHODS: Human umbilical vein endothelial cells (HUVECs) were divided into the following groups: control group, Tongxinluo (100 mg/L) group, hypoxia group and hypoxia+Tongxinluo (100 mg/L) group. The CCK-8 assay were used to detect the viability and proliferation rate of the cells in each group. The protein levels of HIF-1α, Bcl-2, Mcl-1, Bax and phosphorylated Akt were studied by immunoblotting analysis. The HUVECs were transiently transfected with the dominant negative mutant of HIF-1α (DN-HIF). The apoptotic rates were analyzed by flow cytometry (FCM). The HUVECs were transiently transfected with the dominant negative mutant of PI-3K (Δp85) or Akt (DN-Akt) to investigate the role of PI-3K/Akt signal pathway in the anti-hypoxia ability of Tongxinluo on endothelial cells. RESULTS: Under hypoxic condition, although the proliferation rate increased significantly in Tongxinluo group compared with hypoxia group, the degree was notably weak compared with control group. The protein levels of HIF-1α, Bcl-2, Mcl-1 and phosphorylated Akt were up-regulated by Tongxinluo. Meanwhile, the expression of Bax was down-regulated. Inhibition of HIF-1α activation by DN-HIF and inhibition of the PI-3K/Akt pathway by Δp85 or DN-Akt attenuated the increase in HIF-1α expression and HUVEC viability induced by Tongxinluo. The percentage of apoptotic HUVECs was down-regulated to a certain extent by Tongxinluo. CONCLUSION: Tongxinluo improves the anti-hypoxia ability of vascular endothelial cells by up-regulating the protein level of HIF-1α, promoting the expression of anti-apoptotic factors, improving the cell viability and eventually reducing the apoptotic rate.These effects of Tongxinluo depend on PI-3K/Akt signal pathway. 相似文献
19.
AIM: To study the effects of glucose-6-phosphate dehydrogenase (G6PD) silencing by small interference RNA(siRNA) on the levels of reduced nicotinamide-adenine dinucleotide phosphate (NADPH) and hypoxia-inducible factor 1α(HIF-1α) under hypoxia in human colon cancer cell line LoVo.METHODS: Specific siRNA expression vector targeting G6PD gene was constructed. The recombinant plasmid was identified by restriction endonuclease and DNA sequencing, and then transfected into LoVo cells. The effects of G6PD silencing were evaluated by detecting the activity and mRNA expression of G6PD. LoVo cells were cultured in vitro under hypoxic condition. NADPH levels were determined.The mRNA and protein levels of HIF-1α were analyzed by RT-PCR and Western blotting,respectively. RESULTS: The recombinant plasmid for G6PD silencing by siRNA was successfully constructed and transfected into LoVo cells. Compared with untransfected cells,the mRNA expression of G6PD in transfected cells was decreased by 43% and G6PD activity was decreased by 63.5%. Under hypoxic condition, the level of NADPH in transfected cells was significantly decreased (41% vs 100%, P<0.05).HIF-1α protein was also decreased significantly but its mRNA expression had no change as compared with the control cells. CONCLUSION: G6PD silencing by siRNA decreases NADPH level, resulting in the decline of HIF-1α stability in cancer cells under hypoxic condition. By this mechanism, G6PD silencing can influence the hypoxic responses in cancer. 相似文献