Effect of Yiqi Huoxue Jiedu Fang on the growth,metastasis and angiogenesis of Lewis lung carcinomas

AIM: To study the effect of Yiqi Huoxue Jiedu Fang (YHJ), composed of ginsenoside, penex notogingseng and berberin, on tumor growth and metastasis and to explore its mechanism.METHODS: Murine Lewis lung carcinoma transplant model was established and mice were treated with YHJ by intraperitoneal injection. After 10 days, the inhibitory rate of tumor, pathology of tumor and PCNA of tumor cells were detected. After 20 days, numbers of metastatic foci on lung surface and microvessel density (MVD) were determined. Expression of VEGF in tumor and serum were also analyzed by immunohistochemical test and ELISA, respectively. RESULTS: YHJ reduced the weight of tumor and the amount of metastatic foci. The inhibitory rates of tumor at high and low dose of YHJ (24 mg·kg_^-1·d_^-1, 12 mg·kg_^-1·d_^-1) were 48.29% and 37.26%, and the number of metastatic foci was 1.67 and 3.50, while control was 6.44. Furthermore, PCNA of tumor cells, MVD of tumor and VEGF expression in serum and tumor were decreased in YHJ treatment goup as compared with control. CONCLUSION: YHJ remarkably inhibits Lewis lung carcinoma growth and metastasis in mice. Its mechanism may be related to inhibition of angiogenesis.     

Expressions of VEGF,ANG-1,ANG-2 and TSP-1 in cholangiocellular carcinoma and relationship with tumor angiogenesis,differentiation,invasion and metastasis

AIM: To investigate the angiogenesis status,the expression of vascular endothelial growth factor (VEGF),angiopoietin-1 (ANG-1),angiopoietin-2 (ANG-2),thrombospondin-1 (TSP-1) in cholangiocellular carcinoma (CCC) and relationship with tumor angiogenesis,differentiation,invasion and metastasis.METHODS: 33 specimen of surgically resected CCC were investigated.Immunohistochemical staining of CD34,VEGF,ANG-1,ANG-2 and TSP-1 was carried out.RESULTS: The mean MVD was (87.2±52.6)/mm2.VEGF positive expression was found in 75.6% cases;ANG-1 positive expression was observed in 36% cases;ANG-2 positive was detected in 57.6% cases and 45.5% cases exhibited positive TSP-1 expression.VEGF and ANG-2 expressions were found to be associated with significant higher level of MVD (P<0.01 and P<0.05,respectively).TSP-1 expression was found to be associated with significant low level of MVD (P<0.01).Positive TSP-1 expression was also found to be associated with higher level of intrahepatic metastasis (46.7% vs 5.6%,P<0.05).CONCLUSION: Considerable angiogenesis compared to other solid tumors can be observed in CCC.VEGF and ANG-2 might play a proangiogenic role and TSP-1 may play an inhibitory role.Although TSP-1 may increase the intrahepatic metastasis of CCC,neither MVD levels nor the expression of VEGF,ANG-1,or ANG-2 is associated with tumor differentiation,invasion and metastasis.     

Promoting angiogenesis of protein kinase D1 in vitro and in vivo

AIM: To explore the effect of protein kinase D1 (PKD1) on promoting angiogenesis in vitro and in vivo, and to furnish a new idea on targeting PKD1 for the treatment of ischemic heart disease such as myocardial infarction. METHODS: The culture, isolation and identification of endothelial progenitor cells (EPCs) were performed in vitro. The effects of PKD1 and its specific blocking agent CID755673 on expression levels of vascular endothelial growth factor (VEGF) and kinase insert domain receptor (KDR) in EPCs were determined. The rat model of myocardial infarction was established, the intervention effects of PKD1 and CID755673 on morphology, changes of microvessels and endothelial cells, and the expression of VEGF and KDR in the impaired myocardial tissue were analyzed. RESULTS: PKD1 significantly upregulated the expression of VEGF and KDR in EPCs in vitro. Meanwhile, the structure of myocardial tissue was more regular and clear, the cytomembrane of endothelial cells were more smooth and integrity, the pericytes were visible, and the expression of VEGF and KDR was significantly increased in PKD1 treatment group in vivo.CONCLUSION: PKD1 has the ability of angiogenesis obviously, which might be mediated by VEGF.     

Effects of phillyrin on VEGF and endostatin expression in Lewis lung carcinoma

AIM: To investigate the effects of phillyrin on vascular endothelial growth factor (VEGF) and endostatin expression in lung tumor tissues isolated from Lewis lung carcinoma. METHODS: The expression of VEGF and endostatin in control individuals and the patients with lung cancer was determined by immunohistochemistry. In the animal experiment, 5 groups of animals were examined: control, tumor model, and tumor model with 3 different concentrations of phillyrin treatments. For preparation of transplanted tumor model, Lewis cells were subcutaneously injected into the right limb armpit of the nude mice. After that, phillyrin was administered via oral gavage once daily for 20 d at dose of 5 or 10 g/kg, or twice daily at 10 g/kg. Lung tumor tissues isolated from each group were observed by hematoxylin-eosin staining. VEGF and endostatin expression were examined by immunohistochemistry. RESULTS: VEGF expression was increased in lung tumor tissues as compared with normal and pericarcinous tissues, while endostatin expression was decreased. Phillyrin significantly inhibited the tumor size and tumor tissue density dose-dependently, which was accompanied with a decrease in VEGF expression and an increase in endostatin expression. CONCLUSION: Phillyrin inhibits the development of lung tumor through reducing VEGF expression and increasing endostatin expression.     

Inhibition of 17-DMAG on VEGF expression in transplanted human gastric cancer in nude mice

AIM: To observe the anti-tumor effects of heat shock protein 90(HSP90) inhibitor 17-dimethylaminoethylamino-17 demethoxygeldanamycin (17-DMAG) on the tumor growth and angiogenesis in implanted gastric cancer nude mouse model. METHODS: Human gastric cancer cell HGC-27 was subcutaneous inoculation into the nude mice to develop a tumor model. Ten days later, 24 mice with implanted tumor were randomly divided into 3 groups: 17-DMAG group (receiving 17-DMAG at dose of 25 mg/kg), control group (treated with NS at dose of 10 mL/kg) and 5-fluorouracil(5-FU) group (treated with 5-FU at dose of 20 mg/kg). Four weeks after treatment, the tumor volume and weight, and the inhibitory rates of tumor growth were evaluated. In the meantime, the expression of CD31 was detected by immunohistochemical staining. The expression of vascular endothelial growth factor (VEGF) was determined by Western blotting. RESULTS: The size of xenografts in 17-DMAG treatment group was (288.10±23.32)mm³, and that in 5-FU treatment group was (366.37±26.42)mm³, both were significantly smaller than that in control group (957.66±117.51)mm³. The tumor weight in 17-DMAG treatment group was (0.41±0.02)g, significantly less than that in control group (1.12±0.08)g. The inhibitory rate of 17-DMAG was 63%. A significant decease of MVD in 17-DMAG group (21.72±1.24) was observed as compared to 5-FU group (36.70±1.51) and control group (37.78±1.68). The expression of VEGF in 17-DMAG group (15.39±4.37) was significantly lower than that in 5-FU group (26.11±6.26) and control group (36.45±7.45). CONCLUSION: HSP90 inhibitor 17-DMAG suppresses the expression of VEGF and the angiogenesis of the gastric cancer to inhibit the tumor growth.     

Inhibitory effect of survivin antisense oligodeoxyribonucleotide combined with DDP in nude mice bearing human osteosarcoma xenograft

AIM:To investigate the feasibility and its mechanisms of improving therapeutic effect by antisense gene therapy combined with chemotherapy in osteosarcoma. METHODS:The human osteosarcoma implanted tumor model in the nude mice was established. By intratumoral injection and abdominal cavity administration, the tumor bearing mice were treated with survivin ASODN in combination with diamminedichloroplatinum (DDP) for a week. Comparison with each single-agent therapy and control group was performed in aspects such as tumor growth condition, pathological changes of tumor tissues;survivin protein expression in tumor tissues by immunohistochemistry, survivin mRNA expression levels by RT-PCR method and tumor apoptosis by Tdt-mediated dUTP nick end labeling (TUNEL). RESULTS:All nude mice survived the therapy. As compared with the control group, the antisense gene therapy group presented synchronous decrease in survivin mRNA and protein expression;all therapy group displayed tumor growth inhibition and cell apoptosis with different extent;while in contrast to single-agent therapy group, the combined therapy group showed stronger inhibition of tumor growth and abundant tumor cell apoptosis with the highest apoptotic rate. CONCLUSION:Synergistic effect was achieved by combination of DDP with ASODN that may overcome drug resistant of DDP and the combined strategy may shed new light on the cancer therapy.     

Angiogenic effect of HIF-1α on extranodal nasal-type NK/T-cell lymphoma

AIM: To study the angiogenic effect of hypoxia inducible factor 1α(HIF-1α) and its significance on human extranodal nasal-type NK/T-cell lymphoma. METHODS: The protein levels of HIF-1α, vascular endothelial growth factor(VEGF) and VEGF receptor 2(VEGFR2) in human extranodal nasal-type NK/T-cell lymphoma were detected by immunohistochemistry. Microvessel density (MVD) of the tumor tissues was determined by labeling of microvessel endothelium with CD34 antibody. The correlation between the expression of HIF-1α, VEGF and VEGFR2 and MVD was analyzed with SPSS 13.0 statistical software. RESULTS: The positive expression of HIF-1α was observed in 39 cases (39/50, 78%) and the positive expression of VEGFR2 was 27 cases (27/50, 54%) of human extranodal nasal-type NK/T-cell lymphoma. A statistical difference of HIF-1α and VEGFR2 expression between tumor tissues and normal lymphocytes in lymph node was observed (P<0.05). In the tumor tissues, the co-expression of VEGF or VEGFR2 with HIF-1α was 72% and 64%, respectively, significantly higher than that without HIF-1α co-expression (P<0.05). The expression of HIF-1α, VEGFR and VEGFR2 was positively correlated with MVD of the tumor tissues (P<0.01). HIF-1α was expressed in all 15 cases of extranodal nasal-type NK/T-cell lymphoma with angiocentric infiltration.CONCLUSION: HIF-1α may promote angiogenesis of extranodal nasal-type NK/T-cell lymphoma through VEGF/VEGFR2 signaling pathway.     

PTEN,VEGF, COX-2 and angiogenesis in JAK2 V617F positive myeloroliferative neoplasms

AIM: To investigate the expression of PTEN, VEGF and COX-2, and the relationship with angiogenesis in the bone tissues of JAK2 V617F mutation positive myeloroliferative neoplasm(MPN)patients. METHODS: JAK2 V617F positive MPN patients (n=42) were selected in the First Hospital of Baoding including 25 cases of newly diagnosed group and 17 cases of treatment group with IFN-α 2b, and 10 cases of idiopathic thrombocytopenic purpura (ITP) patients served as controls. The ratio of mutant and wild type JAK2 was detected by real-time PCR. The protein levels of p-JAK2, PTEN, VEGF and COX-2, and microvascular density (MVD) marked with CD105 in pathological tissues of bone marrow were detected by immunohistochemistry. RESULTS: The levels of p-JAK2, VEGF, COX-2 and MVD in newly diagnosed group were significantly higher than those in control group. However, the expression level of PTEN was significantly lower than that in control group. The levels of p-JAK2, VEGF, COX-2 and MVD in treatment group were significantly lower than those in newly diagnosed group, while the expression level of PTEN was significantly higher than that in newly diagnosed group. JAK2 V617F mutation burden was positively correlated with VEGF, COX-2 and MVD (P<0.05). PTEN was negatively correlated with VEGF and MVD (P<0.05). The levels of p-JAK2, VEGF, COX-2 and MVD in JAK2 V617F/JAK2 ratio≥0.5 group were significantly higher than those in JAK2 V617F/JAK2 ratio<0.5 group, but PTEN showed the contrary to the above.CONCLUSION: PTEN, VEGF, COX-2 and JAK2 V617F are involved in the pathogenesis of angiogenesis in MPN patients.     

Significance of expression of thrombospondin-1 and receptor-CD36 in hepatocellular carcinoma

AIM: To study the expression of thrombospondin-1 (TSP-1) and receptor-CD36, and investigate the relationship between tumor invasive capability and microvessel density and thrombospondin-1. METHODS: 43 hepatocellular carcinoma (HCC) cases were under investigation. Tissues from tumor, corresponding adjacent non-HCC tissue were stained with CD34 to show the MVD. TSP-1 and CD36 were examined by immunohistochemistry (SP) and RT-PCR. Relationship between clinical pathological features and above parameters was analyed. RESULTS: The staining of TSP-1 in HCC tissue is significantly lower than that in corresponding adjacent non-HCC tissue. Expression of TSP-1 was correlated to tumor thrombi, capsule, tumor invasive capability and CD36. CD36 was also correlated to tumor thrombi and tumor invasive capability. MVD was significantly higher in TSP-1, CD36 positive group than that in negative group. CONCLUSION: TSP-1 inhibits the growth, invasion and angiogenesis in HCC. TSP-1 may take effect through CD36.     

MCP-1/CCR2 signaling pathway mediates alcohol-induced angiogenesis in breast cancer

AIM To investigate the role of monocyte chemoattractant protein-1 (MCP-1) and its receptor CC chemokine receptor 2 (CCR2) in ethanol-promoted breast cancer angiogenesis and the underlying mechanism. METH?ODS: A mouse model of transplanted breast tumor with moderate alcohol consumption was established. The correlations between the expression of MCP-1/CCR2 and the expression of angiogenesis markers [platelet endothelial cell adhesion molecule-1 (PECAM-1) and vascular endothelial growth factor (VEGF)] in tumor tissues were examined by immunohistochemistry. In vitro, a 3D tumor-endothelial co-culture system was established to observe tumor angiogenesis and the role of MCP-1/CCR2 signaling pathway in alcohol-mediated angiogenesis. The cell migration ability was detected to clarify whether MCP-1/CCR2 enhanced cell mobility to form new vessels. RESULTS MCP-1 and CCR2 were both highly expressed in the breast tumor tissues of tumor-bearing mice consuming alcohol, and their expression levels were consistent with the angiogenic markers PECAM-1 and VEGF (P<0.05). The interaction between mouse breast cancer E0771 cells and endothelial cells was observed to promote angiogenesis in the 3D tumor-endothelial co-culture system with or without alcohol stimulation. MCP-1 promoted this kind of tumor angiogenesis, while CCR2 antagonist effectively inhibited the tumor angiogenesis and especially blocked alcohol-induced angiogenesis. Activation of MCP-1/CCR2 signaling pathway enhanced the migration ability of endothelial cells. CONCLUSION The MCP-1/CCR2 signaling pathway plays an important role in promoting the angiogenesis of breast cancer stimulated by alcohol. The mechanism might be that MCP-1 improves the migration of endothelial cells and then promotes angiogenesis.     

Effect of insulin on the secretion of VEGF and its receptor in serum-free cultured bovine thoracic aortic endothelial cells

AIM: This study was designed to investigate the secretion of VEGF and its receptor (flt-1 or flk-1/KDR) protein by cultured bovine thoracic aortic endothelial cells treated with various insulin concentrations. METHODS: Endothelial cells was isolated from bovine thoracic aorta, and cultured in serum-free medium, then incubated with different insulin concentrations (30 mU/L, 300 mU/L, 3 000 mU/L). The level of VEGF and its receptor (flt-1 or flk-1/KDR) protein were detected by immunohistochemical staining. RESULTS: As compared with no insulin group, the expression of VEGF protein in low insulin concentration (30 mU/L and 300 mU/L) groups were significantly increased (P＜0.01). The expression of VEGF protein in high insulin concentration (3 000 mU/L) group was significantly decreased (P＜0.05). Howerer, no difference of the expression of VEGF receptor (flt-1 or flk-1/KDR) protein among all groups (P＞0.05) was observed. CONCLUSION: Low concentration insulin up-regulates the VEGF protein expression while high concentration insulin down-regulates the VEGF protein expression in bovine thoracic aortic endothelial cells, but insulin had no directly effect on the VEGF receptor (flt-1 or flk-1/KDR) protein expression in bovine thoracic aortic endothelial cells.     

The inhibitory effects of endostatin gene transfer on the growth of breast cancer cells in vivo and in vitro

AIM: To investigate the therapeutic action of secreted endostatin (ES) on breast cancer cells. METHODS: Retroviral-mediated endostatin gene was transferred to breast cancer cell line MDA-MB-231. The ES biological properties and function were evaluated by polymerase chain reaction (PCR), MTT and a murine xenograft model. RESULTS: After retroviral transduction, endostatin genetically modified breast tumor cells were confirmed by PCR, and the integration and durative expression of endostatin gene was successfully committed. Compared with controls, endostatin secreted by genetically modified cells markedly inhibited endothelial cell proliferation (P<0.05) while the influences on the growth of MDA-MB-231 cell line in vitro were not found (P>0.05). The results of the transplanted subcutaneous tumor model in nude mice suggested that the subcutaneous growth of MDA-MB-231 was significantly inhibited by the expression of endostatin gene (P<0.05). In experimental groups, the tumor microvascular density (MVD) and VEGF expression were decreased. CONCLUSION: Retroviral- mediated overexpression of endostatin inhibits the proliferation of vascular endothelial cells and angiogenesis that associated with tumor growth in vivo via the paracrine pathway, which has a potential effect in the angiostatic gene therapy for breast cancer.     

Expression of endostatin in ischemic myocardium of myocardial infarction rats

AIM: To study the expression of endostatin in ischemic myocardium of myocardial infarction (MI) rats in various periods and the correlation with VEGF expression and microvascular density (MVD).METHODS: Thirty-two male Sprague-Dawley rats after myocardial infarction were randomly divided into 7, 14, 21 and 28 days group.The sham group was normal control group (eight rats in each group).The expression of endostatin, VEGF and MVD in ischemic myocardium were observed by immunohistochemistry.RESULTS: The expression of endostatin significantly increased in the ischemic myocardium after MI, peaked at 7 days, then gradually decreased at 14, 21 and 28 days.The endostatin level at 28 days was the same as the shams.The changing trends of expression of endostatin in ischemic myocardium after MI were similar to that of VEGF and were significantly correlated with the MVD.CONCLUSION: The expression of endostatin increased in ischemic myocardium of myocardial infarction rats.The changing trends of endostatin were similar to that of VEGF and positively correlated with the MVD.These data suggest that endostatin may modulate ischemic myocardium angiogenesis after myocardial infarction.     

Microvessel density and expression of vascular endothelial growth factor in glomeruli of diabetic mice

AIM: To investigate the relationship between microvessel density (MVD) and expression of vascular endothelial growth factor (VEGF) in glomeruli of diabetic mice. METHODS: Streptozotocin-induced diabetic mice as well as the control mice were involved in this study for 6 weeks. The body weight and blood glucose level of the mice in each group were weekly measured at certain time point. The morphological changes of the kidney were observed under light microscope, and the diameter, perimeter and area of the glomeruli were detected by an image analysis system. The expression of CD34 and VEGF in glomeruli was examined by immunohistochemistry method, and MVD and VEGF index were also calculated. RESULTS: In comparison with the control mice, the blood glucose level was significantly increased,and the body weight was decreased in diabetic mice(P<0.01). The diameter, perimeter and area of glomeruli in diabetic mice were significant greater than those in control mice (P<0.05). Increased expression of CD34 and VEGF in the glomeruli of diabetic mice was observed. Glomerular MVD of diabetic mice was significantly higher than that of the controls (P<0.01), and was positively correlated with the VEGF index (r=0.9979, P<0.05). CONCLUSION: VEGF may promote the angiogenesis in glomeruli of diabetic mice. The increase in VEGF expression may play a role in the pathogenesis of diabetic nephropathy.     

Effect of sorafenib on liver regeneration after partial hepatoectomy in liver cirrhotic rats

AIM: To study the effect of sorafenib on the liver regeneration after partial hepatectomy (PH) in cirrhotic rats. METHODS: Thirty Wistar rats with liver cirrhosis induced successfully with diethylnitrosamine (DEN) underwent 30% PH and then were randomly divided into 2 groups (n=15). The rats in experimental group were fed with sorafenib at dose of 30 mg·kg^-1·d^-1 from the 1st day to the 10th day after PH, while those in control group were fed with vehicle by gavage. The blood and liver tissues of the rats were collected after PH and at the end of the experiment. Liver regeneration rate (LRR) and proliferating cell nuclear antigen (PCNA) expression were assessed for determining the hepatocyte proliferation. The content of alanine transaminase (ALT), albumin (ALB), total bilirubin (TBIL), direct bilirubin (DBIL), angiogenesis related factors including vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 2 (VEGFR-2), platelet-derived growth factor receptor β (PDGFR-β) and micro-vessel density (MVD) were measured in both groups. RESULTS: LRRs on day 10 after PH were 45.43%±3.36% and 44.21%±2.77% in experimental group and control group, respectively (P>0.05), and the expression of PCNA in hepatic tissues of the rats was not found by the method of immunohistochemistry in both groups. Liver function index had no significant difference between the 2 groups (P>0.05). However, other than VEGF, sorafenib resulted in inhibition of VEGFR-2 and PDGFR-β expression and reduction of MVD in experiment group, and significant difference between the 2 groups was observed (P<0.01). CONCLUSION: Sorafenib does not influence live regeneration after PH in liver cirrhotic rats.     

Establishing a three-dimensional angiogenesis model in vitro and secretion of MCP-1 and VEGF by tumor cells

AIM: To establish a three-dimensional angiogenesis model in vitro for observing the influence of tumor cells on angiogenesis, and to explore its possible molecular mechanism. METHODS: Human umbilical vein endothelial cells (HUVECs) and mouse endothelial cells SVEC4-10EE2 were coated on the surface of beads, and then mixed with fibrinogen and seeded in cell culture plates containing thrombin. The cultured endothelial cells coated on the beads grew into a vessel-like structure in a three-dimensional space containing fibrin condensate to establish an in vitro three-dimensional angiogenesis model. Tumor cells MDA-MB-231 and E0771 were co-cultured in the model to observe the effect of tumor cells on angiogenesis in vitro,respectively. The concentrations of monocyte chemoattractant protein-1 (MCP-1) and vascular endothelial growth factor (VEGF) in the tumor cells culture supernatant were measured by ELISA. An antagonist of CCR2 (receptor of MCP-1), along with SU5416 (an inhibitor of VEGF receptor), were added to the tumor cell co-culture system. The supernatant of the tumor cells was collected as a conditioned medium, which was then added to the angiogenesis system of HUVECs or SVEC4-10EE2 cultured individually. The effect of conditioned medium on angiogenesis was observed under the conditions of with or without SU5416, as well as with or without CCR2 antagonist. RESULTS: Under normal condition, HUVECs and SVEC4-10EE2 formed a multi-cellular vascular structure the three-dimensional angiogenesis model in vitro. The co-culture of MDA-MB-231 (E0771) cells significantly promoted in the formation of membrane-like structures. The ELISA results showed that the levels of MCP-1 and VEGF in the supernatant of tumor cells were significantly elevated (P<0.05). MCP-1 promoted the formation of membrane tube in three-dimensional culture system in vitro. After adding the antagonists of MCP-1 receptor CCR2 and VEGF (SU5416), the angiogenesis was significantly inhibited (P<0.05). At the same time, conditioned medium promoted the formation of extracorporeal membranes in the endothelial cells. The promoting effect was blocked by CCR2 antagonists and SU5416 (P<0.05). CONCLUSION: The in vitro three-dimensional angiogenesis model is successfully established. Tumor cells significantly promote the formation of membrane tubes. The effect of tumor cells might be related to MCP-1 and VEGF secretion.     

Effects of inhibition of Cripto gene siRNA on vascular endothelial growth factor of colon cancer cell line LS-174T

AIM:To study the effects of Cripto gene on vascular endothelial growth factor (VEGF) of colon carcinoma cells.METHODS:Cripto siRNA was designed and constructed. Colon cancer LS-174T cells were divided into 4 groups: control group and different dose (3.125, 6.25 and 12.5 nmol/L) of siRNA groups. After transfected for 24, 48 and 72 h, colon cancer cells were harvested to carry on the next tests. Expression of Cripto mRNA was determined with real-time PCR, and immunofluorescence isothiocyanate (FITC) labeling assay and Northern blotting were performed to examine the expression of protein and mRNA of VEGF, respectively. The cells in control group and cells transfected with 12.5 nmol/L siRNA were inoculated into nude mice respectively. 30 days after inoculated, the mice of two groups were executed, and immunohistochemical (ICH) assay was used to evaluate the VEGF protein of mice tumor. RESULTS:siRNA down-regulated the Cripto mRNA in a dose and time dependent manner. Protein and mRNA of VEGF in transfected cells reduced in a dose and time dependent manner. Compared to control, the expression of VEGF protein from ICH assay was lowered significantly (P<0.05).CONCLUSION:Cripto gene might contribute to the regulation of angiogenesis of colon carcinoma. The down-regulation of Cripto gene by siRNA can suppress angiogenesis of human colon cancer.     

Expression of EphA2 and ephrin-A1 in endometrial endometrioid adenocarcinoma and their relationship with angiogenesis

AIM: To investigate the expression of erythropoietin-producing hepatocellular receptor A2 (EphA2) and its ligand ephrin-A1 in endometrial endometrioid adenocarcinoma (EEA), and to analyze their relationship with angiogenesis of the tumor. METHODS: The CD34-stained microvessel density (MVD) and the expression of ephA2 and ephrin-A1 were detected by immunohistochemical assay in 56 cases of EEA, 20 cases of endometrial hyperplasia, 30 cases of normal proliferative endometrium and 30 cases of normal secretory endometrium. The correlations among the expression of EphA2 and ephrin-A1, MVD and clinicopathological features were analyzed. RESULTS: MVD and the expression of EphA2 and ephrin-A1 in EEA were significantly higher than those in the tissues from endometrial hyperplasia and normal endometrium (P<0.05). They were related to FIGO stage, histological differentiation, depth of myometrial invasion, lymphovascular invasion and progesterone receptor expression (P<0.05). A significant positive correlation between MVD and the expression of EphA2 and ephrin-A1 was observed by Spearman rank correlation test (r=0.476, P<0.05; r=0.501, P<0.05). CONCLUSION: Overexpression of EphA2 and its ligand ephrin-A1 in EEA may be involved in the angiogenesis and progesterone resistance.     

Relationship between changes of bone mineral density and bone marrow angiogenesis in ovariectomized rats

AIM: To explore the relationship between the changes of bone mineral density (BMD) and bone marrow angiogenesis in ovariectomized rats. METHODS: Thirty female Sprague-Dawley rats (3 month-age) were randomly divided into ovariectomized (OVX) groups and sham operated (sham) groups, and executed after 4 weeks, 8 weeks and 12 weeks respectively. The bone mineral density (BMD) of left femora was measured. The right distal femoral epiphysis was fixed in formalin, decalcificated by EDTA-Na2, dehydrated and embedded in paraffin. Four-μm-thick slices were obtained from the paraffin section and stained with haematoxylin-eosin (HE) for bone marrow pathological examination. The number of bone marrow microvessels was examined by means of immunohistochemical staining for CD34 to stain the endothelial cells of blood vessels to definite the bone marrow microvascular density (MVD). Plasma levels of vascular endothelial growth factor (VEGF) were examined by the method of ELISA. RESULTS: The BMD of femoral in 8-week OVX group were decreased significantly compared to the 8-week sham group (P<0.05), suggesting the establishment of osteoporosis model. Meanwhile, the area of hematopoietic tissue decreased and the area of adipose tissue increased. These changes became obviously in 12-week OVX group, and the area of trabecular bone and the bone marrow MVD significantly decreased compared to the 12-week sham group. A positive correlation between MVD and BMD, area of hematopoietic or trabecular bone as well as a negative correlation between MVD and area of adipose tissue were observed. The plasma levels of VEGF in OVX group were not significantly different from that in sham group, and had no correlation relationship with the indexes of bone marrow pathology. CONCLUSION: There has an increase in MVD companied with the bone mass loss and hematopoietic tissue decreased in ovarietomized rats, which provide experiment proof to treat osteoporosis with the means of promoting of MVD in bone marrow.