目的 通过复制人肝癌细胞株HepG2裸鼠皮下移植瘤模型，观察绿茶提取物表没食子儿茶素没食子酸酯（EGCG）干预对HepG2移植瘤新生血管生成的影响。 方法 瘤体接种复制HepG2移植瘤模型，荷瘤裸鼠20只随机分组，实验组给予EGCG溶液每日20 mg/（kg·只），腹腔注射3周，对照组给予等量灭菌注射用水3周，末次用药24 h，后处死裸鼠，剥离移植瘤。常规病理切片观察移植瘤组织结构；逆转录-聚合酶链式反应和免疫组织化学法检测移植瘤缺氧诱导因子-1α（HIF-1α）、血管内皮生长因子（VEGF）mRNA及蛋白表达，并通过检测CD34表达计数瘤组织微血管密度（MVD）。 结果 组织病理学观察实验组移植瘤见大量坏死区，瘤体内血管数量明显少于对照组；实验组HIF-1α、VEGF mRNA及蛋白表达水平比对照组均明显下调（P＜0.05），实验组MVD比对照组明显下降（P＜0.05）。 结论 EGCG可抑制荷瘤裸鼠HepG2移植瘤新生血管生成。
Objective To investigate the effects and mechanism of 17β-estradiol on the retinal neovasularization in rats with oxygen-induced retinopathy (OIR). MethodsA total of 48 SD rats were randomly divided into control group A, control group B, experimental group A and experimental group B with 12 rats in each group. The rats in control group A and experimental group A received a hypodermic injection of 0.1 ml PBS, and the rats in control group B and experimental group B group received an a hypodermic injection of 0.1 ml 17β-estradiol. At postnatal day 7 (P7) and P14, the mRNA expression of vascular endothelial growth factor (VEGF) and Hypoxia-inducible factor (HIF) -1α in the retina were measured by real-time polymerase chain reaction (RTPCR). At P14, endothelial cell nuclei breaking through the internal limiting membrane were counted after staining with hematoxylin and eosin (HE), and the protein expression of VEGF was measured after immunohistochemical staining. The changes of retinal ultrastructure were observed by transmission electron microscopy. ResultsAt P14, the difference of the number of endothelial cell nuclei among four groups was statistically significant(F=10.7, P＜0.05). The number of endothelial cell nuclei in experimental group A was increased greater than that in control group A (P＜0.05) and experimental group B(q=5.16，P＜0.05). But there was no difference between control group A and experimental group B (q=0.25，P＞0.05). The difference of VEGF protein expression among the four groups was statistically significant (P＜0.05). Comparing experimental group A with control group A, B and experimental group B, the difference was statistically significant (P＜0.05). In experimental group A there was ganglion cell swelling, pale staining cytoplasm and mitochondria vacuolizationin, while these were normal in other three groups. At P7 and P14, the differences of VEGF and HIF-1 mRNA expression among four groups were statistically significant(F=14.7,16.1, 13.4, 17.5; P=0.001, 0.005, 0.003, 0.009). At P7, the VEGF mRNA expression in control group B was more than that in control group A (q=5.22, P＜0.05). The VEGF mRNA expression in experimental group B was more than that in experimental group A (q=4.32, P＜0.05). At P14, the VEGF mRNA expression in control group B was more than that in control group A (q=3.72, P＜0.05), but there was no difference of HIF-1 mRNA expression between two groups. The VEGF and HIF-1 mRNA expression in experimental group B were both decreased more than those in experimental group A (q=5.12, 4.08;P＜0.05). Conclusions 17β-estradiol has the effect of two way regulation in VEGF mRNA, which increases VEGF expression in retina under hyperoxic conditions so as to develop the vascular system; which reduces VEGF and HIF-1α expression so as to prevent pathologic neovascularization under hypoxic conditions. It provides some protection from the damage of retinal neovascularization.
ObjectiveTo review the development and applications of hypoxia-inducible factor 1α (HIF-1α) in the strategy of tissue engineered angiogenesis and osteogenesis. MethodThe literature about HIF-1α in tissue engineering technology was reviewed, analyzed, and summarized. ResultsHIF-1α plays a key role in angiogenic-osteogenic coupling, and as an upstream regulator, HIF-1α can regulate the expressions of its target genes related with angiogenesis and osteogenesis. In addition, HIF-1α not only can control and improve the angiogenesis, but also has important significance in proliferation and differentiation of seed cells, especially stem cells, which is the foundation for bone healing. ConclusionsWith the development of tissue engineering technology, the problems in the applications of HIF-1α, such as the effective dose of targeting controlled-release, pro-inflammatory effect, and carcinogenicity, will be explored and solved in the future, so it can be used better in clinical.
ObjectiveTo observe and investigate the effect of HIF-2α in the process of neovascularization of proliferative diabetic retinopathy (PDR).MethodsRetrospective clinical study. From July 2014 to July 2015, 60 eyes of 57 PDR patients diagnosed in Ophthalmology Department of Affiliated Hospital of Qingdao University were included in the study. Twenty-eight eyes of 27 patients received intravitreal injections of 0.5 mg ranibizumab (0.05 ml) at 2-7 days before surgery (ranibizumab group) and other 32 eyes of 30 patients did not (group without ranibizumab). Eighteen eyes of 18 patients with epiretinal membranes were included as controls. Pathological specimens of PDR fibrovascular membrane and premacular membrane were obtained during vitrectomy. The immunohistochemical staining and real-time PCR (RT-PCR) were used to detecting the expression of HIF-2α, Dll4 and VEGF. Kruskal-wallis test was used to compare the expression differences of correlation factors between groups. Spearman correlation analysis was used to analyze the relationship between the two variables.ResultsThe immunohistochemical staining revealed that there were positive expression of HIF-2α, Dll4 and VEGF in all PDR membranes, regardless of the injection of the ranibizumab. The levels of HIF-2α, Dll4 and VEGF protein in the group without ranibizumab were higher than those of the ranibizumab group (t=4.36, 6.01, 4.82; P=0.000, 0.008, 0.016). RT-PCR showed that the differences of the mRNA expression of HIF-2α, Dll4 and VEGF were all statistically significant among the PDR patients and controls (H=18.81,19.60, 20.50; P=0.000, 0.000, 0.000). The expression of HIF-2α, Dll4 and VEGF in the PDR membranes was higher than that of epiretinal membranes from non-diabetic patients. In the PDR patients,the expression of HIF-2α, Dll4 and VEGF of the group without ranibizumab was higher than that of the ranibizumab group. The spearman correlation analysis showed that the expression of mRNA between HIF-2α and Dll4, HIF-2α and VEGF were both significantly correlated (r=0.95, 0.87; P＜0.05).ConclusionsThe expression of HIF-2α in the PDR membranes was higher than that of the controls. It is positively correlated with the expression of the DLL4 and VEGF.
To review the role of hypoxia inducible factor 1α (HIF-1α) in hypoxic-ischemic injury and its repair, and to analyze the possible mechanisms. Methods Recent l iterature on HIF-1α and its role in hypoxic-ischemic injury was reviewed and analyzed. Results HIF-1α was involved in the hypoxic-ischemic injury of various organs or tissues and their repair processes. Conclusion HIF-1α has a potential to treat common cl inical hypoxic-ischemic injuries and has a promisingfuture for appl ication.
The intervention therapy targeting vascular endothelial growth factor (VEGF) has become a specific and effective method for the treatment of diabetic retinopathy (DR). However, some patients did not respond or responded poorly to anti-VEGF therapy, and its effects of eliminating edema and improving vision appear to be unstable in the same patient. Hypoxia-inducible factor-1α (HIF-1α), an important upstream transcriptional regulator of VEGF, is an oxygen concentration-sensitive protein expressed in tissues under hypoxia. It can simultaneously target many downstream target genes except VEGF, such as placental growth factor and angiopoietin-like protein 4, to cause blood-retinal barrier damage and neovascularization, and thus participate in various pathological changes of DR to promote the occurrence and development of DR. Therefore, direct intervention of HIF-1α or targeting one or more downstream target genes regulated by HIF-1α to treat DR may have better efficacy. In the future, the development of effective and safe HIF inhibitors or anti-VEGF with HIF-1α other target gene inhibitors may have broader clinical application prospects.
Objective Ginsenoside Rg1 could increase the tolerance of neural hypoxia and ischemia under stress, and play an anti-apoptotic effect in hypoxia ischemia brain damage (HIBD). To investigate the effects of ginsenoside Rg1 on neural apoptosis and recovery of neurological function in neonatal rats with HIBD, and to explore the possible mechanism. Methods Fifty-four 10-day-old SD rats (weighing 16-22 g) were randomly allocated into sham-operation group (Sham group, n=6), HIBD model group (HIBD group, n=24), and ginsenoside Rg1 treatment group (Rg1 group, n=24). SDrats in HIBD group and Rg1 group were made the models of HIBD by l igation of the right common carotid artery (CCA) and subsequently hypoxic ventilation (8%O2 plus 92%N2) for 2.5 hours; and in Sham group, the right CCA was only exposed without l igation of CCA and hypoxic ventilation. Intraperitoneal injection of 0.1 mL normal sal ine (NS) containing 40 mg/kg Rg1 was given immediately after operation in Rg1 group, intraperitoneal injection of 0.1 mL pure NS was given in both HIBD group and Sham group and was repeated every 24 hours. The general state of SD rats was monitored after operation, and Longa scores were recorded to evaluate the neurological function at 4, 8, 24, and 72 hours after HIBD. Western blot and immunohistochemistry staining were used to detect protein expressions of both hypoxia inducible factor 1α (HIF-1α) and cleaved caspase 3 (CC3). TUNEL staining was used to evaluate neural apoptosis in situ. Results All rats survived to the end of the experiment. Neurological dysfunction was observed in both HIBD group and Rg1 group, showing significant difference in Longa score when compared with that in Sham group (P lt; 0.05). There was significant difference in Longa score between Rg1 group and HIBD group at 72 hours after HIBD (P lt; 0.05). Western blot showed that the protein expressions of both HIF-1α and CC3 were observed at every time point in every group. The expressions of HIF-1α protein in HIBD group and Rg1 group were significantly higher than those in Sham group at 4, 8, 24, and 72 hours (P lt; 0.05), and the expressions in Rg1 group were significantly higher than those in HIBD group (P lt; 0.05). The expressions of CC3 protein in HIBD group were significantly higher than those in Sham group at 4, 8, 24, and 72 hours (P lt; 0.05), and significant difference was found between Rg1 group and Sham group only at 4 hours (P lt; 0.05). Immunohistochemistry staining demonstrated that HIF-1α and CC3 protein mainly distributed in nucleusand cytoplasma, the results of HIF-1α and CC3 protein expression were similar to the results by Western blot. TUNEL staining showed that the positive cells were characterized by yellow or brown particle confined within nucleus. The number of apoptotic cells at every time point in HIBD group was significantly higher when compared with that in Sham group (P lt; 0.05), and the number of apoptotic cells in Rg1 group was significantly lower when compared with that in HIBD group at 8, 24, and 72 hours (P lt; 0.05). Conclusion Rg1 could inhibit Caspase 3 activation by strengthening and stabil izing HIF-1α signal pathway, and plays a role of anti-apoptosis in neonatal rats with HIBD.
Objective To investigate the expression of hypoxia inducible factor 1(HIF1alpha;) in ratsprime; retinae during the embryonic and earlier postnatal period. Methods The retinal expression patterns of HIF-1alpha; protein and mRNA of embryonic day 12 (E12), E16, E20, and postnatal day 1(P1) and P5 rats were determined by immunohistochemical staining and reverse transcriptionpolymerase chain reaction (RT-PCR). Results HIF-1alpha; protein was detected in the neural epithelial layer and the pigment epithelial layer at all those 5 timepoints, with higher expression in the ganglion cell layer and the inner plexiform layer, and seems limited to the ganglion cell layer when re tina became more mature. Embryonic rat retina had higher expression of HIF-1alpha; protein and mRNA than postnatal retina, the difference was significant (P＜0.01）. Conclusion The expression of HIF1alpha; in ratsprime;retina e differs from embryonic to earlier postnatal stages.