ObjectiveTo find the role of oncogene cmet and suppressor gene p53 in the process of tumor angiogenesis and their clinical significance. MethodsBy immunohistochemical method and computer image analysis technique, microvessel count and cmet, p53 protein expression were quantitatively determined in 80 cases of breast carcinoma and 20 cases of breast fibroadenoma. ResultsThe high microvessel count and the positive expression of cmet, p53 were significantly correlated with histologic grade, lymph node metastasis and the stage of the tumor (P＜0.01). The high microvessel count was significantly correlated with the positive expression of cmet and p53 (P＜0.01).ConclusionBoth oncogene cmet and suppressor gene p53 modulate tumor angiogenesis of breast carcinoma.
ObjectiveTo discuss the feasibility of treating noumenon tumor by antiangiogenesis.MethodsRelated literatures of recent 5 years was reviewed.ResultsTumor angiogenesis were related closely with growth, development, metastasis and prognosis of noumenon tumor. It was possible to inhibit the growth and metastasis of noumenon tumor with antiangiogenesis in vitro and vivo.ConclusionAntiangiogenesis will be a new therapy of treating noumenon tumor.
Objective To study the effect of dimethyloxalylglycine (DMOG) on angiogenesis in Choke Ⅱ zone of rats cross-zone perforator flaps and its mechanism. Methods One hundred and twenty-six adult male Sprague Dawley rats were randomly divided into DMOG group, YC-1 group, and control group, with 42 rats in each group. Cross-zone perforator flap model with size of 12 cm×3 cm was made on the back of rats in the three groups. DMOG group was intraperitoneally injected with DMOG (40 mg/kg) at 1 day before operation, 2 hours before operation, and 1, 2, and 3 days after operation; YC-1 group and control group were intraperitoneally injected with YC-1 (10 mg/kg) and the same amount of normal saline at the same time points, respectively. The survival of flap was observed after operation. At 7 days after operation, the survival area of flap in each group was measured and the survival rate of flap was calculated. Flap transmittance test, gelatin-lead oxide angiography, and HE staining were used to observed the angiogenesis in the Choke Ⅱ zone of flaps in each group. Immunohistochemical staining and Western blot were used to detect the expressions of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1α (HIF-1α) in Choke Ⅱ zone of flaps in each group. The expressions of VEGF and HIF-1α were also determined by ELISA at 3, 5, and 7 days. Results At 7 days after operation, there was no obvious necrosis at the distal end of the flap in DMOG group, while necrosis occurred in both the control group and YC-1 group, mainly located at the distal end. The flap survival rate of DMOG group was 90.28%±1.37%, which was significantly higher than that of YC-1 group (84.28%±1.45%) and control group (85.83%±1.60%) (P<0.05). DMOG group had more angiogenesis in Choke Ⅱ zone and the vascular structure was clear and complete. In YC-1 group and control group, the vessels in Choke Ⅱ zone was less and the vascular structure was disordered. The number of vessels was (25.56±1.29)/field in the DMOG group, which was significantly higher than that in the YC-1 group [(7.38±0.54)/field] and the control group [(14.48±0.91)/field] (P<0.05). At 3, 5, and 7 days after operation, HIF-1α and VEGF expressions in ChokeⅡzone of DMOG group were significantly higher than those in YC-1 group and control group (P<0.05). ConclusionDMOG can promote angiogenesis in Choke Ⅱ zone, accelerate the early angiogenesis of the flap, improve the microcirculation and blood supply in the potential zone of the flap, reduce the injury of flap ischemia and hypoxia, and increase the survival rate of the flap.
Objective To explore the effects of adipose-derived stem cell released exosomes (ADSC-Exos) on the proliferation, migration, and tube-like differentiation of human umbilical vein endothelial cells (HUVECs). Methods Adipose tissue voluntarily donated by liposuction patients was obtained. The ADSCs were harvested by enzyme digestion and identified by flow cytometry and adipogenic induction. The ADSC-Exos were extracted from the supernatant of the 3rd generation ADSCs and the morphology was observed by transmission electron microscopy. The surface proteins (Alix and CD63) were detected by Western blot. The nanoparticle tracking analyzer NanoSight was used to analyze the size distribution of ADSC-Exos. After co-culture of PKH26 fluorescently labeled ADSC-Exos with HUVECs, confocal microscopy had been used to observe whether ADSC-Exos could absorbed by HUVECs. ADSC-Exos and HUVECs were co-cultured for 1, 2, 3, 4, and 5 days. The effect of ADSC-Exos on the proliferation of HUVECs was detected by cell counting kit 8 (CCK-8) assay. The expression of VEGF protein in the supernatant of HUVECs with or without ADSC-Exos had been detected by ELISA after 12 hours. Transwell migration assay was used to detect the effect of ADSC-Exos on the migration ability of HUVECs. The effect of ADSC-Exos on the tubular structure formation of HUVECs was observed by Matrigel experiments in vitro. The formation of subcutaneous tubular structure in vivo was observed in BALB/c male nude mice via the injection of HUVECs and Matrigel with or without ADSC-Exos. After 2 weeks, the neovascularization in Matrigel was measured and mean blood vessel density (MVD) was calculated. The above experiments were all controlled by the same amount of PBS. Results After identification, the cultured cells were consistent with the characteristics of ADSCs. ADSC-Exos were circular or elliptical membranous vesicle with uniform morphology under transmission electron microscopy, and expresses the signature proteins Alix and CD63 with particle size ranging from 30 to 200 nm. Confocal microscopy results showed that ADSC-Exos could be absorbed by HUVECs. The CCK-8 analysis showed that the cell proliferation of the experimental group was better than that of the control group at each time point (P<0.05). The result of Transwell showed that the trans-membrane migration cells in the experimental group were significantly more than that in the control group (t=9.534, P=0.000). In vitro, Matrigel tube-forming experiment showed that the number of tube-like structures in the experimental group was significantly higher than that of the control group (t=15.910, P=0.000). In vivo, the MVD of the experimental group was significantly higher than that of the control group (t=16.710, P=0.000). The ELISA assay showed that the expression of VEGF protein in the supernatant of the experimental group was significantly higher than that of the control group (t=21.470, P=0.000). Conclusion ADSC-Exos can promote proliferation, migration, and tube-like structure formation of HUVECs, suggesting that ADSC-Exos can promote angiogenesisin vitro and in vivo.
Objective To explore the effect of natural hirudin on proliferation of human microvascular endothelial cells (HMVECs) and its preliminary mechanism of promoting angiogenesis. Methods Three-dimensional culture models of HMVECs were established in vitro and observed by inverted phase contrast microscopy after 24 hours of culturing. Then, the three-dimensional culture models of HMVECs were treated with different concentrations (1, 4, and 7 ATU/mL) of the natural hirudin, respectively, and Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum as control. The cell proliferations of 4 groups were detected by cell counting kit 8 (CCK-8) method at 24, 48, and 72 hours; the angiogenesis of 4 groups were observed by tube formation assay at 24 hours; the expressions of vascular endothelial growth factor (VEGF) and Notch1 of HMVECs in 4 groups were observed by immunofluorescence staining at 24 hours. Results The observation of cells in three-dimensional culture models showed that HMVECs attached to Matrigel well, and the cells formed tube structure completely after 24 hours. The results of CCK-8 test showed that the absorbance (A) value of 1 and 4 ATU/mL groups were higher than that of control group at each time point (P<0.05), andA value of 4 ATU/mL group was the highest. The A value of 7 ATU/mL group was significantly lower than those of 1 and 4 ATU/mL groups and control group (P<0.05). The tube formation assay showed that the tube structure was more in 1 and 4 ATU/mL groups than in 7 ATU/mL group and control group, and in 4 ATU/mL group than in 1 ATU/mL group, showing significant differences (P<0.05). There was no significant difference between 7 ATU/mL group and control group (P>0.05). The results of immunofluorescence staining showed that compared with control group, the Notch1 expression was higher in 1 and 4 ATU/mL groups and lower in 7 ATU/mL group; and there was significant difference between 4 and 7 ATU/mL groups and control group (P<0.05). The VEGF expression was higher in 1, 4, and 7 ATU/mL groups than in control group, in 4 ATU/mL group than in 1 and 7 ATU/mL groups, showing significant differences (P<0.05). Conclusion Natural hirudin can promote angiogenesis at low and medium concentrations, but suppress angiogenesis at high concentrations. Its mechanism may be related to the VEGF-Notch signal pathway.
Objective To review the research progress of P75 neurotrophin receptor (P75NTR) so as to clarify its mechanism, and to explore its relationship with nonunion so as to provide a new idea for the treatment of nonunion. Methods The related domestic and foreign literature of P75NTR in recent years was extensively reviewed, summarized, and analyzed to find out the mechanism of action of P75NTR and the pathological factors of nonunion formation. Results P75NTR can express in nonunion tissues and lead to defect of fibrin degradation and inhibition of angiogenesis, which play an important role in the pathogenesis of nonunion. Conclusion It needs to be confirmed by further study whether the purpose of treating nonunion can be achieved by blocking the effects described above of P75NTR.
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.
This research is to explore the perfusion time-intensity curve parameters of a lung adenocarcinoma xenograft into nude mouse model with contrast enhanced ultrasonography (CEUS); and to investigate the angiogenesis features of tumor at different growth time. Twenty one lung adenocarcinoma xenografted nude mice were divided into three groups and inculcated with human lung adenocarcinoa. Time window for examining CEUS were respectively in 7-day, 14-day and 28-day. The perfusion parameters including rise time (RT), peak intensity (PI), area under the curve (AUC) of lung tumor were obtained on CEUS images by using off-line software Q lab. Immunohistochemically staining for CD34 was used to observe the microvessel density (MVD).The 7-day group had the highest AUC and PI; AUC and PI of 14-day and 28-day group decreased gradually (P < 0.05). RT was increased as tumor growth. In tumor with necrosis, AUC and PI of non-necrosis part were also larger than necrosis part (P < 0.05). Immunohistochemically staining for CD34 of all tumors reflected that the density of microvessels in necrosis tumor was significantly higher than those without necrosis (7.50±3.44 vs.12.44±5.74, P=0.034). Pearson correlation indicated that PI was positively related with MVD (r=0.668, P=0.008). Lung adenocarcinoma perfusion characteristic can be accessed from time-intensity curve parameters by using noninvasively and non-radiative contrast enhanced ultrasonography. Time-intensity curve parameters including AUC, PI and RT may reflect tumor angiogenesis.
【Abstract】Objective To understand the features of lymphatic vessel, and to summarize the foundation and mechanism of the promotion and inhibition of tumor lymphangiogenesis recorded on the current studies of animal experiments and clinical researches. Methods The related literatures of the structural features of lymphatic vessel, lymphatic endothelial molecular markers, the origin of lymphatic tumors, the molecular mechanisms and regulatory factors were reviewed, and the relationship between tumor lymphangiogenesis and lymphatic metastasis, the treatment targeting at the formation of the anti-tumor lymphatic vessel and its existing problems were also analyzed. Results Hyperplasia of lymphatic vessels occurred during the process of tumor formation and progression. The structural features of the lymphatic vessels in the tumor were conducive to tumor lymphatic metastasis. In recent years, methods of anti-lymphangiogenesis and inhibition of tumor lymphatic metastasis had achieved considerable success in animal experiments. However, there were still a lot of problems need to be solved. Conclusion Tumor lymphangiogenesis has a significantly positive correlation between tumor lymphatic metastasis and patients’ prognosis, which may indicate that treatment against the formation of tumor lymphatic vessel maybe effective.
Objective To summarize the regulatory effect of non-coding RNA (ncRNA) on type H vessels angiogenesis of bone. Methods Recent domestic and foreign related literature about the regulation of ncRNA in type H vessels angiogenesis was widely reviewed and summarized. ResultsType H vessels is a special subtype of bone vessels with the ability to couple bone formation. At present, the research on ncRNA regulating type H vessels angiogenesis in bone diseases mainly focuses on microRNA, long ncRNA, and small interfering RNA, which can affect the expressions of hypoxia inducible factor 1α, platelet derived growth factor BB, slit guidance ligand 3, and other factors through their own unique ways of action, thus regulating type H vessels angiogenesis and participating in the occurrence and development of bone diseases. ConclusionAt present, the mechanism of ncRNA regulating bone type H vessels angiogenesis has been preliminarily explored. With the deepening of research, ncRNA is expected to be a new target for the diagnosis and treatment of vascular related bone diseases.