Objective To investigate the osteogenesis effects of angiopoietin 1 (Ang-1) gene transfected bone marrow mesenchymal stem cells (BMSCs) seeded on β tricalcium phosphate (β-TCP) scaffolds (tissue engineered bone) with platelet-rich plasma (PRP). Methods BMSCs were isolated from bone marrow tissue of rabbits. The Ang-1 gene was transfected into the BMSCs at passage 2 by lentivector, which were seeded on β-TCP scaffolds with PRP (0.5 mL) after 48 hours of transfection. Bilateral radial segmental bone defects (15 mm in length) were created in 20 3-month-old New Zealand rabbits. Then the tissue engineered bone with the Ang-1 gene transfected BMSCs (experimental group) and untransfected BMSCs (control group) were implanted into the defects in the right and left radius, respectively. X-ray, histology, immunohistochemistry, and biomechanics observations were done at 2, 4, 8, and 12 weeks after operation. Results In vitro, the transfected rate was over 90% and RT-PCR showed that the Ang-1 expression were significantly increased after transfection. The X-ray films showed that some callus formed at 4 weeks, partial bony union was observed at 8 weeks, and complete union at 12 weeks in experimental group; and bone union was not observed at 12 weeks in control group. HE staining showed that capillary appeared at 8 weeks and more capillaries were observed in new bone at 12 weeks in experimental group; only a few capillaries were observed at 12 weeks in control group. At 8 and 12 weeks, the microvascular density were (50.1 ± 7.8) /mm2 and (66.1 ± 3.5) /mm2 in experimental group and were 0 and (30.3 ± 7.2)/mm2 in control group, showing significant differences between 2 groups at 12 weeks (Z= —2.107, P=0.031). Immunohistochemistry examination showed that the positive cells can be found at 8 weeks in experimental group. And the biomechanical analysis showed that maximum loads of experimental group were significantly higher than those of control group in three-point bending test and compression test at 12 weeks (P lt; 0.05). Conclusion The tissue engineered bone with PRP and Ang-1 can increase the osteogenic properties by enhancing capillary regeneration, thus it can be used to repair radial segmental bone defects of rabbit.
Objective To investigate the possibility of constructing eukaryoticexpression vector for human angiopoietin 1（hAng-1），transfecting it to bonemarrow mesenchymal stem cells (MSCs) so as to repair bone defect. Methods The eukaryotic expression vector pcDNA3-hAng-1 was constructed by recombinant DNA technique, transfected into MSCs by liposome DOTAP, and selected with G418. The hAng-1 expression of mRNA and protein was detected by reverse transcript-PCR and Western Blot. Results After the recombinant eukaryotic expressionvector for hAng-1 was digested with Xho-I and BamH-I, electrophoresis revealed 1.4 kb fragment for hAng-1 gene and 5.4 kb fragment for pcDNA3 vector. In the transfected MSCs, the mRNA and protein expression of hAng-1 gene were detected with reverse transcriptPCR and Western Blot. Conclusion The constructed eukaryotic expression vector hAng-1 could be expressed in the transfected MSCs, thus to provide the basis for bone repair with tissue engineering.
Objective To study the relationship between angiopoietin and the angiogenesis of gastric carcinoma, and to investigate the inter-regulation effect between different vascular growth factors which exist in microenvironment of gastric carcinoma. Methods Literatures about angiopoietin and angiogenesis of gastric carcinoma were collected and reviewed. Results There is a relationship between the expression of angiopoietin in microcirculation and the angiogenesis of gastric carcinoma. Yet, it is still a disputable issue on the inter-regulation effect of different vascular growth factors. Conclusion Angiopoietin may play an important role in the processes of sprouting, development and apoptosis of the microvessel of gastric carcinoma.
【Abstract】ObjectiveTo observe the chemotactic role on umbilical vein endothelial cells of SMMC7721 hepatic carcinoma cells with angiopoietin gene expression in order to study the effects of angiopoietin on hepatocellular carcinoma angiogenesis. MethodsAngiopoietin gene 1 (Ang-1) fragment and Ang-2 fragment was transfected into SMMC7721 liver carcinoma cell line by Lipofectamine induced gene transfection technique. The chemotactic role of SMMC7721 liver carcinoma cell line on umbilical vein endothelial cells was observed through microchemotaxis analysis. ResultsThe chemotactic response of the Umbilical vein endothelial cells was obviously improved with Ang1 expression (P＜0.05). This effect seemed to be inhibited by Ang-1 antibody (P＜0.05). However, there was no difference of the chemotactic effects with or without Ang-2 expression (Pgt;0.05). ConclusionAng-1 is a chemotactic factor for vascular endothelial cell and a promoter for angiogenesis, whereas Ang-2 does not show obvious chemotactic role.
【Abstract】 Objective To study the expressions of cyclooxygenase-2 (Cox-2) and angiopoietin-2 (Ang-2) in colorectal cancer tissues, cancer adjacent tissues and normal colorectal tissues, and the relationship between these expressions and the clinicopathologic features of colorectal cancer. Methods Forty-five excised samples of colorectal adenocarcinoma were confirmed pathologically and 39 of them were of well or moderately differentiated and 6 of poorly differentiated. Lymph nodes metastasis developed in 30 patients. And 15 cases were in stage of A or B and the rest were in the stage of C or D according to the Dukes stage. Taken PBS as the negative control and the verified Cox-2 or Ang-2 positive sections as positive controls, this study detected the expressions of Cox-2 and Ang-2 protein in 45 colorectal cancer tissues, 45 cancer adjacent tissues and 15 normal colorectal tissues by using immunohistochemical SP technique method. Results Cox-2 and Ang-2 were expressed in colorectal cancer tissues and cancer adjacent tissues, but were not expressed in normal colorectal tissues. In 45 colorectal cancer tissues, the positive expression rates of Cox-2 and Ang-2 were 80.0% and 66.7%; in 45 cancer adjacent tissues, the positive expression rates of Cox-2 and Ang-2 were 35.6% and 11.1%, respectively. The positive expression rates of both Cox-2 and Ang-2 in colorectal cancer tissues were significantly higher than those in cancer adjacent and normal colorectal tissues. There were close correlations between the expressions of Cox-2 and Ang-2 and some pathologic features, such as lymph node metastasis and Dukes stage; whereas there were no significant association between the expressions and gender, histological type and position of tumor. There was also a close correlation between the expressions of Cox-2 and Ang-2 themselves. Conclusion Cox-2 and Ang-2 play an important role in the occurrence and development of colorectal cancer. The use of specific inhibitor of Cox-2 as a treatment for colorectal cancer may become feasible and necessary.
ObjectiveTo investigate the mechanism of early vascularization of the tissue engineered bone in the treatment of rabbit radial bone defect by local injection of angiopoietin 2 (Ang-2).MethodsForty-eight New Zealand white rabbits were established unilateral 1.5 cm long radius defect models. After implantation of hydroxyapatite/collagen scaffolds in bone defects, the rabbits were randomly divided into 2 groups: control group (group A) and Ang-2 group (group B) were daily injected with 1 mL normal saline and 1 mL saline-soluble 400 ng/mL Ang-2 at the bone defect within 2 weeks after operation, respectively. Western blot was used to detect the expressions of autophagy related protein [microtubule associated protein 1 light chain 3 (LC3), Beclin-1], angiogenesis related protein [vascular endothelial growth factor (VEGF)], and autophagy degradable substrate protein (SQSTMl/p62) in callus. X-ray films examination and Lane-Sandhu X-ray scoring were performed to evaluate the bone defect repair at 4, 8, and 12 weeks after operation. The rabbits were sacrificed at 12 weeks after operation for gross observation, and the angiogenesis of bone defect area was observed by HE staining.ResultsWestern blot assay showed that the relative expressions of LC3-Ⅱ/LC3-Ⅰ, Beclin-1, and VEGF in group B were significantly higher than those in group A, and the relative expression of SQSTMl/p62 was significantly lower than that in group A (P<0.05). Radiographic and gross observation of specimens showed that only a few callus were formed in group A, the bone defect was not repaired; more callus were formed and complete repair of bone defect was observed in group B. The Lane-Sandhu scores in group B were significantly higher than those in group A at 4, 8, and 12 weeks after operation (P<0.05). HE staining showed that the Harvard tubes in group B were well arranged and the number of new vessels was significantly higher than that in group A (t=–11.879, P=0.000).ConclusionLocal injection of appropriate concentration of Ang-2 may promote early vascularization and bone defect repair of tissue engineered bone in rabbits by enhancing autophagy.
Diabetic retinopathy (DR) is one of the most common microvascular complications of diabetes, and it is the main cause of vision loss in diabetic patients. Angiopoietin (Ang), a superfamily of secreted proteins, is a vascular growth factor that regulates the stability of vascular environment, participates in angiogenesis and repair, and lipid metabolism. It plays an important role in the development of DR and has become a new target for the treatment of diabetic retinopathy. With the in-depth study of Ang and the research and development of various drugs for Ang, it is expected to bring new ideas and strategies for the treatment of DR in the future.
Angiopoietin-like protein (ANGPTL), a group of secreted glycoproteins, is widely expressed in vivo and is involved in many pathophysiological processes such as glycolipid metabolism, stem cell growth, local inflammation, vascular leakage and angiogenesis. Many kinds of ANGPTL are closely related to the occurrence and development of diabetic retinopathy (DR), especially ANGPTL4, which has gradually become a new hotspot in the field of DR Research. ANGPTL is involved in glucose metabolism and lipid metabolism, promotes increased vascular permeability, pathological angiogenesis, and participates in intraocular inflammation. ANGPTL is a promising molecular target. It can not only be used as a biomarker to predict the occurrence and progression of DR, but also provide new ideas for the treatment of DR by making antibody drugs to interfere with this molecule.