ObjectiveTo investigate the effects of hypoxic three-dimensional culture microenvironment on the proliferation of bone marrow mesenchymal stem cells and its mechanism. MethodsP5 generation mouse bone marrow mesenchymal stem cells and P (3HB-co-4HB) were co-cultured under normoxic three-dimensional (20%) and hypoxic three-dimensional microenvironment (4%) respectively. After 24 hours, the proliferation of the two groups was determined by CCK-8 method. The expression of HIF-1α gene was detected by real-time quantitative PCR after 12 hours. Western blotting was used to detect the expression of HIF-1α protein after 24 hours. ResultsAfter 24 hours, the CCK-8 method showed that the OD value of the hypoxia group was significantly higher than that of the normoxia group (0.455±0.027 vs. 0.352±0.090, n=12, P<0.05). After 12 hours of hypoxic culture, the expression level of HIF-1α mRNA in the hypoxia group was significantly higher than that in the normoxia group (P<0.05). Compared with the normoxia group (0.47± 0.05), the relative expression level of HIF-1α protein in the hypoxia group (0.63±0.06) significantly increased in the Western blotting after 24 hours (n=3, P<0.05). ConclusionThe hypoxic three-dimensional microenvironment can promote the proliferation of bone marrow mesenchymal stem cells, which may be related to the activation of HIF-1α signaling pathway.
ObjectiveTo investigate the expression and correlation of hypoxia inducible factor 1α (HIF-1α) and autophagy related molecules (Beclin1 and LC3B) in rat nucleus pulposus cells under hypoxia in vitro.MethodsThe nucleus pulposus cells were extracted from the nucleus pulposus of healthy adult Sprague Dawley rats and passaged. The 3rd generation cells were identified by HE staining and collagenase type Ⅱ immunofluorescence staining and randomly divided into 4 groups. The cells in group A were cultured for 8 hours under normal oxygen condition (37℃, 5%CO2, 20%O2); the cells in group B were cultured for 8 hours under hypoxia condition (37℃, 5%CO2, 1%O2); the cells in group C were transfected with HIF-1α-small interfering RNA and cultured for 8 hours under hypoxia condition; and the cells in group D were cultured with autophagy inhibitor 3-MA for 8 hours under hypoxia condition. Western blot and real-time fluorescence quantitative PCR (qRT-PCR) were used to detect the expressions of HIF-1α and autophagy related molecules (Beclin1 and LC3B) in all groups.ResultsHE staining of the 3rd generation nucleus pulposus cells showed that the cytoplasm was light pink and the nucleus was blue black, and the collagenase type Ⅱ immunofluorescence staining was positive. Western blot and qRT-PCR results showed that the relative expressions of HIF-1α, Beclin1, and LC3B proteins and genes in group B were significantly higher than those in group A (P<0.05); the relative expressions of HIF-1α, Beclin1, and LC3B proteins and genes in group C were significantly lower than those in group B (P<0.05). There was no significant difference in the relative expression of HIF-1α protein and gene between groups B and D (P>0.05); while the relative expressions of Beclin1 and LC3B proteins and genes in group D were significant lower than those in group B (P<0.05).ConclusionHypoxia can induce the expressions of HIF-1α and autophagy related molecules (Beclin1 and LC3B) in rat nucleus pulposus cells, and HIF-1α in rat nucleus pulposus cells under hypoxia is related to the expression of autophagy related molecules, that is, down-regulation of HIF-1α can significantly reduce the expression of autophagy related molecules, while the down-regulation of autophagy levels under hypoxia has no or little effect on the expression of HIF-1α.
Objective To evaluate the effects of Shengji Yuhong collagen on promoting angiogenesis of the ischemia tissues and probe the possible mechanisms. Methods Forty-eight Wistar rats were divided by random method of paired into blank group， control group （collagen），and experimental group （Shengjiyuhong collagen）. After made the rats hind limb ischemia model， collagens with or without the extracts of Shengji Yuhong Gao were randomly paired implanted locally in hind limb ischemia tissues of rats in experimental group or control group. The samples of collagens and tissues about 0.5 cm large surrounding the collagen were explanted respectively on day 3，7， 14， and 28 for detected the hemog-lobin contents in colagen， microvascular counting by using CD34 immunohistochemical markers， and the expressions of HIF-1α mRNA and VEGF mRNA by using real-time fluorescent quantitative RT-PCR. The blood perfusion of the ischemic tissues at each time were determined by using laser speckle imaging system of Moor-FLPI. Results The results of Moor-FLPI showed that the obvious ischemia condition after model made， the blood perfusion was significantly lower than that before operation （P＜0.01）. On day 3 after operation it showed obvious congestion in the ischemic tissues， and from day 7 to day 14， it showed the ischemia state locally till day 28 after operation which showed improved situation of ischemic. Except for the day 3， the blood perfusion of experimental group were higher than those of blank group （P＜0.05）. There was no statistical significance between the blank group and control group （P＞0.05）. The blood perfusion on day 7 and day 14 after operation of experimental group were higher than those of control group （P＜0.05）. The hemoglobin contentsof each time point in the experimental group were higher than those in the control group （P＜0.01）. The microvascular counting on day 7 and day 14 in experimental group were higher than those of control group （P＜0.05）. The expressions of HIF-1α mRNA and VEGF mRNA at each time point of experimental group were higher than those of control group and blank group （P＜0.05）， and there was no significant differences between the control group and blank group （P＞0.05）. Conclusion The effects on promoting angiogenesis of rat hind limb ischemia tissues with Shengji Yuhong collagen may though inducing the expressions of HIF-1 α mRNA and VEGF mRNA locally.
ObjectiveTo systematically review the correlation between the expression of hypoxia inducible factor-1α (HIF-1α) protein and different clinical pathological features of renal cell cancer.
MethodsWe electronically searched databases including The Cochrane Library, PubMed, EMbase, CNKI, VIP, CBM and WanFang Data from inception to June 2015 to collect case-control studies investigating the correlation between HIF-1α protein expression and different clinical pathological features of renal cell cancer. Two reviewers independently screened literature, extracted data, and assessed the risk of bias of included studies. Then meta-analysis was performed using RevMan 5.3 software.
ResultsA total of 8 case-control studies involving 429 cases of renal cell cancer and 130 cases of normal renal tissue were included. The results of meta-analysis showed that:HIF-1α protein expression was higher in the renal cell cancer group than that in the normal renal tissue group (OR=16.76, 95%CI 8.53 to 32.92, P<0.000 01); HIF-1α protein expression was higher in the lymph node metastasis group than that in the non-lymphnode metastasis group (OR=4.33, 95%CI 2.53 to 7.39, P<0.000 01); HIF-1α protein expression was higher in the TNM Ⅲ-IV group than that in the TNM I-Ⅱ group (OR=0.30, 95%CI 0.18 to 0.51, P<0.000 1); HIF-1α protein expression was higher in the Fuhrman pathology classification G3+G4 group than that in the G1+G2 group (OR=0.54, 95%CI 0.29 to 0.98, P=0.04). However, there were no significant differences in HIF-1α protein expression between the age≥50 group and the age <50 group (OR=1.09, 95%CI 0.54 to 2.19, P=0.82), and between the male group and the female group (OR=0.77, 95%CI 0.48 to 1.25, P=0.29).
ConclusionHIF-1α protein expression is significantly correlated to the clinical stage and pathological grading of renal cell cancer. It is possibly involved in the initiation and development of renal cell cancer. Due to the limited quantity and quality of included studies, the above conclusion needs to be further verified by more high quality studies.
