Macrophage-specific promoter SP146-C1 enhances vascular endothelial growth factor C expression in atherosclerotic mice / 中国组织工程研究

BACKGROUND:The expression efficiency of recombinant adeno-associated virus serotype 9(rAAV9)carrying the macrophage-specific promoter synthetic promoter 146-C1(SP146-C1)and the exogenous gene vascular endothelial growth factor C(VEGFC)in atherosclerosis is uncertain. OBJECTIVE:To investigate the expression efficiency of rAAV9-SP146-C1-VEGFC in atherosclerotic mice and its effect on lymphangiogenesis. METHODS:Thirty ApoE-/-mice were fed high-fat diet for 12 weeks to establish atherosclerosis models and were randomly divided into six groups,five in each group:7-,14-,21-,28-,and 35-day transfection groups and control group.The mice in the transfection groups were transfected with 5.0×1011 vg rAAV9-SP146-C1-VEGFC by caudal vein injection.In the control group,the mice were injected with the same amount of control virus rAAV9-SP146-C1-Scramble.Animals in the first five groups were killed under anesthesia at 7,14,21,28 and 35 days after transfection,respectively,and those in the control group were killed under anesthesia at 7 days.Serum,femur,tibia,heart and aorta tissue samples were collected and retained in each group.The femur and tibia of mice in each group were used to extract bone marrow-derived macrophages.The gene expression of vascular endothelial growth factor C(VEGFC),vascular endothelial growth factor receptor 3(VEGFR3),Podoplanin and lymphatic vessel endothelial hyaluronan receptor-1(LYVE-1)in bone marrow-derived macrophages and the aorta were detected by RT-qPCR.VEGFC protein expression levels in bone marrow-derived macrophages and the aorta were detected by western blot,serum level of VEGFC was detected by ELISA,and VEGFC expression in the aortic sinus and LYVE-1 expression around the aorta and in the myocardium was detected by immunofluorescence. RESULTS AND CONCLUSION:The serum level of VEGFC,the mRNA expression of VEGFC,VEGFR3,Podoplanin,and LYVE-1 in bone marrow-derived macrophages and the aorta,the protein expression of VEGFC in bone marrow-derived macrophages,and the fluorescence intensity of VEGFC in aortic sinus plaques were significantly increased in the 7-day transfection group compared with the control group(P＜0.05,P＜0.01).Serum VEGFC level of mice transfected with rAAV9-SP146-C1-VEGFC gradually increased with time and began to decrease at 28 days.mRNA levels of VEGFC,VEGFR3,Podoplanin and LYVE-1 in mouse aorta and bone marrow-derived macrophages,VEGFC protein level in bone marrow-derived macrophages,VEGFC fluorescence intensity in aortic sinus plaques,LYVE-1 fluorescence intensity around the aortic sinus and in the myocardium gradually increased with time(P＜0.05).In addition,the mRNA level of LYVE-1 in the aorta and the fluorescence intensity of LYVE-1 around the aortic sinus and in the myocardium were the highest at 28 days(P＜0.05),and gradually decreased(P＜0.05).The expression of the other indicators reached the peak at 21 days.To conclude,rAAV9-SP146-C1-VEGFC could effectively transfect bone marrow-derived macrophages and promote lymphatic hyperplasia in atherosclerotic mice.

Inhibitory Effects of Ligustrazine Plus Emodin on Expression of VEGF-related Factors in Ascites Carcinoma Walker-256 Cells / 中国实验方剂学杂志

Objective:To investigate the effects of ligustrazine combined with emodin on angiogenesis of ascites carcinoma Walker-256 cells by observing their inhibition against nuclear factor-<italic>κ</italic>B （NF-<italic>κ</italic>B）， hypoxia-inducible factor-1<italic>α</italic> （HIF-1<italic>α</italic>）， and vascular endothelial growth factor-C （VEGF-C） in HIF signaling pathway. Method:Fifty SD rats were randomly divided into sham operation group， model group， ligustrazine group， emodin group and ligustrazine combined with emodin group. Following the in situ injection of rat ascites carcinoma Walker-256 cells into the liver of normal rats， they were grouped and administered with ligustrazine （10 mg·kg^-1）， emodin （10 mg·kg^-1）， and ligustrazine （10 mg·kg^-1） plus emodin （10 mg·kg^-1） for seven days. Afterwards， the tumor-inoculated liver tissue was sampled from the experimental group and prepared into pathological sections for investigating tumor cell survival and VEGF expression. The <italic>in vitro</italic> hypoxia and hypoglycemia model （oxygen-glucose deprivation model）， hypoxia model， and hypoglycemia model of Walker-256 cells were constructed respectively. In the ligustrazine group， emodin group， and ligustrazine combined with emodin group， three consecutive concentrations that did not affect the proliferation of Walker-256 cells were selected for investigation. The drugs were administered before modeling， and the model treatment lasted for 4 h. The levels of HIF-1<italic>α</italic>， VEGF-C， and NF-<italic>κ</italic>B in the cell culture supernatant of each group were tested. Result:After the rat liver was inoculated with Walker-256 cells， the total liver mass was significantly increased（<italic>P</italic><0.05）， higher than that in the ligustrazine group， the emodin group， or the ligustrazine combined with emodin group（<italic>P</italic><0.05）. Histopathological examination showed that the response of VEGF expression in the liver tissue of each administration group was lower than that of the model group. At the cellular level， the levels of HIF-1<italic>α</italic>， VEGF-C， and NF-<italic>κ</italic>B in oxygen-glucose deprivation model of the ligustrazine group and the ligustrazine combined with emodin group were significantly reduced（<italic>P</italic><0.05）， exhibiting a certain dose-dependent response， followed by the reduction in the hypoxia model. The levels of HIF-1<italic>α</italic> and NF-<italic>κ</italic>B in the oxygen-glucose deprivation model and the hypoglycemia model of the emodin（1×10^-2，1×10^-3 mol·L^-1） group and the ligustrazine combined with emodin（1×10^-2，1×10^-3 mol·L^-1） group were significantly reduced， but there was no significant change in VEGF-C level of the hypoxia model of all the administration groups. Conclusion:Ligustrazine or emodin alone or their combination inhibits the abnormal increase in the weight of rat liver after inoculation with Walker-256 cells and the expression of VEGF in the liver tissue. Ligustrazine and emodin inhibit the protein expression of NF-<italic>κ</italic>B and HIF-1<italic>α</italic>， thereby reducing the gene and protein expression of metastasis-related target VEGF-A activated by HIF-1<italic>α</italic> transcription， restricting tumor cell neovascularization， and inhibiting the invasion and spread of ascites carcinoma cells. Among them， ligustrazine has the most significant effect against hypoxia. Glucose interferes with the effect of ligustrazine. The combination of ligustrazine with emodin is conducive to diminishing the intervention of glucose and stabilizing the inhibition against tumor cells.

Efficacy and effect on serum VEGF-C of mild moxibustion plus functional exercise for upper-limb lymphedema after breast cancer surgery / 针灸推拿医学（英文版）

Objective: To observe the efficacy of mild moxibustion combined with functional exercise in the treatment of upper-limb lymphedema after breast cancer surgery and its effect on serum vascular endothelial growth factor C (VEGF-C). Methods: Seventy-eight patients were divided into a control group and an observation group by the random number table method, with 39 cases in each group. The control group received functional exercise and the observation group received mild moxibustion plus functional exercise. The differences in circumference between the two upper limbs, the lymphatic flow of the affected upper limb, the disability of arm, shoulder and hand (DASH) score, the functional assessment of cancer therapy-breast (FACT-B) score and serum VEGF-C level between the two groups were compared before and after treatment. Efficacy was evaluated after treatment. Results: The total effective rate was significantly higher in the observation group than in the control group (P<0.05). The difference in circumference between the two upper limbs of the two groups decreased significantly after treatment (both P<0.05), but it was significantly lower in the observation group than in the control group (P<0.05). The lymphatic flow of the affected upper limb of the two groups increased significantly after treatment (both P<0.05), but it was significantly greater in the observation group than in the control group (P<0.05). The DASH scores of the two groups decreased significantly after treatment (both P<0.05), but it was significantly lower in the observation group than in the control group (P<0.05). The FACT-B scores of the two groups increased significantly after treatment (both P<0.05), but it was significantly higher in the observation group than in the control group (P<0.05). After treatment, the serum VEGF-C level increased significantly in the observation group (P<0.05), whereas the control group did not show significant change (P>0.05). The post-treatment serum VEGF-C level was significantly higher in the observation group than in the control group (P<0.05). Conclusion: The efficacy of mild moxibustion combined with functional exercise for upper-limb lymphedema after breast cancer surgery is certain, which can reduce the difference in circumference between the two upper limbs, increase the lymphatic flow of the affected upper limb, improve the limb function and the quality of life, and regulate the serum VEGF-C level.

Increased expression of vascular endothelial growth factor-C and vascular endothelial growth factor receptor-3 after pilocarpine-induced status epilepticus in mice

Vascular endothelial growth factor (VEGF)-C and its receptor, vascular endothelial growth factor receptor (VEGFR)-3, are responsible for lymphangiogenesis in both embryos and adults. In epilepsy, the expression of VEGF-C and VEGFR-3 was significantly upregulated in the human brains affected with temporal lobe epilepsy. Moreover, pharmacologic inhibition of VEGF receptors after acute seizures could suppress the generation of spontaneous recurrent seizures, suggesting a critical role of VEGF-related signaling in epilepsy. Therefore, in the present study, the spatiotemporal expression of VEGF-C and VEGFR-3 against pilocarpine-induced status epilepticus (SE) was investigated in C57BL/6N mice using immunohistochemistry. At 1 day after SE, hippocampal astrocytes and microglia were activated. Pyramidal neuronal death was observed at 4 days after SE. In the subpyramidal zone, VEGF-C expression gradually increased and peaked at 7 days after SE, while VEGFR-3 was significantly upregulated at 4 days after SE and began to decrease at 7 days after SE. Most VEGF-C/VEGFR-3-expressing cells were pyramidal neurons, but VEGF-C was also observed in some astrocytes in sham-manipulated animals. However, at 4 days and 7 days after SE, both VEGFR-3 and VEGF-C immunoreactivities were observed mainly in astrocytes and in some microglia of the stratum radiatum and lacunosum-moleculare of the hippocampus, respectively. These data indicate that VEGF-C and VEGFR-3 can be upregulated in hippocampal astrocytes and microglia after pilocarpine-induced SE, providing basic information about VEGF-C and VEGFR-3 expression patterns following acute seizures.

Effects of three inducing factors on differentiation of bone marrow derived mesenchymal stem cells into lymphatic endothelial cells / 中华烧伤杂志

Objective@#To observe the effects of basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and vascular endothelial growth factor C (VEGF-C) on the differentiation of bone marrow mesenchymal stem cells (BMSCs) into lymphatic endothelial cells (LECs).@*Methods@#The third to the fifth passage of BMSCs of rats were collected for the following experiments. (1) BMSCs of rats were collected and divided into negative control group, CD90 group, CD44 group, and CD34 group according to the random number table (the same grouping method below), with 3 samples in each group. Phosphate buffer of 5 μL was added to cells in negative control group, and cells in the other 3 groups were added with 5 μL corresponding antibodies respectively. The positive expression of cell surface antigen was detected by flow cytometer. (2) BMSCs of rats in 3 batches were collected and divided into blank control group, VEGF-C group, HGF group, bFGF group, VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium, cells in VEGF-C group were added with 2 mL complete medium and 10 μL VEGF-C of 10 μg/mL, cells in HGF group were added with 2 mL complete medium and 16 μL HGF of 10 μg/mL, and cells in bFGF group were added with 2 mL complete medium and 20 μL bFGF of 1 μg/mL. Cells in VEGF-C+ HGF group, VEGF-C+ bFGF group, HGF+ bFGF group, and VEGF-C+ HGF+ bFGF group were added with 2 mL complete medium and induction factors with corresponding concentration and volume as above. On 10 d of culture, the morphology of the cells was observed by the inverted phase contrast microscope, and the protein and mRNA expressions of lymphatic vessel endothelial hyaluronic acid receptor 1 (LYVE-1), VEGF receptor 3 (VEGFR3), and integrin α9 were detected by Western blotting and real-time fluorescent quantitative reverse transcription polymerase chain reaction respectively. (3) BMSCs of rats were collected and divided into blank control group, HGF+ VEGF-C+ bFGF group, bFGF+ VEGF-C+ HGF group, and VEGF-C+ HGF+ bFGF group, with 3 samples in each group. Cells in blank control group were added with 2 mL complete medium. Cells in HGF+ VEGF-C+ bFGF group were added with 2 mL complete medium, 16 μL HGF of 10 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 20 μL bFGF of 1 μg/mL was added. Cells in bFGF+ VEGF-C+ HGF group were added with 2 mL complete medium, 20 μL bFGF of 1 μg/mL, and 10 μL VEGF-C of 10 μg/mL, after 6 hours, 16 μL HGF of 10 μg/mL was added. Cells in VEGF-C+ HGF+ bFGF group were simultaneously added with 2 mL complete medium and the same concentration and volume of three inducing factors as above. In addition, BMSCs of rats in another 2 batches were collected and grouped, and they were dealt with the same methods as above except that the interval time of 6 hours in HGF+ VEGF-C+ bFGF group and bFGF+ VEGF-C+ HGF group was adjusted to 12 and 24 hours. On 10 d of culture, protein expressions of LYVE-1, VEGFR3, and integrin α9 were detected by Western blotting. Data were processed with analysis of variance of factorial design, one-way analysis of variance, and least significant difference t test, and Bonferroni correction.@*Results@#(1) The positive expression rates of surface antigen of cells in negative control group, CD90 group, CD44 group, and CD34 group were 0.39%, 99.84%, 99.90%, and 0.57%, respectively. (2) On 10 d of culture, cells in blank control group, HGF group, bFGF group, and HGF+ bFGF group presented long fusiform, while cells in the other groups presented polygonal shape. (3) On 10 d of culture, there were no protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of blank control group, HGF group, bFGF group, and HGF+ bFGF group. On 10 d of culture, protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of VEGF-C+ HGF+ bFGF group were significantly higher than those in VEGF-C group (t=24.21, 11.04, 15.43, P<0.01), VEGF-C+ HGF group (t=10.81, 9.93, 10.20, P<0.01), and VEGF-C+ bFGF group (t=11.67, 6.32, 19.00, P<0.01). Protein expressions of LYVE-1 in cells of VEGF-C+ HGF group and VEGF-C+ bFGF group were significantly higher than the protein expression in VEGF-C group (t=8.69, 15.20, P<0.01). Protein expression of VEGFR3 in cells of VEGF-C+ bFGF group was obviously higher than the protein expressions in VEGF-C group and VEGF-C+ HGF group (t=8.67, 7.21, P<0.01). Protein expression of integrin α9 in cells of VEGF-C+ HGF group was obviously higher than the protein expressions in VEGF-C group and VEGF-C+ bFGF group (t=8.80, 8.83, P<0.01). (4) On 10 d of culture, there were no mRNA expressions of LYVE-1, VEGFR3, and integrin α9 in cells of blank control group, HGF group, bFGF group, and HGF+ bFGF group. On 10 d of culture, mRNA expressions of LYVE-1 and VEGFR3 in cells of VEGF-C group were significantly lower than those in VEGF-C+ bFGF group and VEGF-C+ HGF+ bFGF group (tLYVE-1=6.22, 18.01, tVEGFR3=8.49, 15.34, P<0.01), and mRNA expression of integrin α9 were significantly lower than that in VEGF-C+ HGF group and VEGF-C+ HGF+ bFGF group (t=13.24, 9.65, P<0.01). The mRNA expressions of LYVE-1, VEGFR3, and integrin α9 in cells of VEGF-C+ HGF+ bFGF group were obviously higher than those in VEGF-C+ HGF group and VEGF-C+ bFGF group (t=13.92, 11.95, 13.72, 5.27, 5.64, 9.10, P<0.01). Compared with those of VEGF-C+ bFGF group, the mRNA expression of VEGFR3 of cells in VEGF-C+ HGF group was significantly lower (t=6.91, P<0.01), while the mRNA expression of integrin α9 of cells in VEGF-C+ HGF group was significantly higher (t=11.69, P<0.01). (5) On 10 d of culture at interval time of 6, 12, 24 h, there were no protein expressions of LYVE-1, VEGFR3, or integrin α9 in cells of blank control group. On 10 d of culture at interval time of 6, 12, 24 h, the protein expressions of LYVE-1, VEGFR3, and integrin α9 in cells of HGF+ VEGF-C+ bFGF group, bFGF+ VEGF-C+ HGF group, and VEGF-C+ HGF+ bFGF group were close (F6 h=2.25, 2.47, 2.19, F12 h=2.93, 1.47, 3.25, F24 h=0.28, 0.20, 1.01, P>0.05).@*Conclusions@#VEGF-C is a necessary factor for inducing BMSCs to differentiate into LECs. HGF and bFGF may promote the differentiation by up-regulating the expressions of integrin α9 and VEGFR3 respectively. But the induction effects of the two factors may be independent. The combination of VEGF-C, HGF, and bFGF have the best effects of promoting differentiation.

Experimental research in athymic nude mice for autologous lymph nodes fragmentary transplantation combined with VEGF-C / 中华整形外科杂志

Objective@#Autologous lymph nodes fragmentary transplantation combined with vascular endothelial growth factor-C (VEGF-C) on athymic nude mice to explore the association between regeneration of lymphatic vessel and tumor cell migration.@*Methods@#A total of 45 nude mice were randomly divided into 3 groups: Group A, simple autologous lymph nodes fragmentary transplantation, n=15; Group B, autologous lymph nodes fragmentary transplantation together with VEGF-C, n=15; Group C, without any intervention, n=15. At 1 month, 2 months and 6months after surgeries, the axillary lymph nodes of 5 mice in each group were dynamically examined by in vivo indocyanine green (ICG) fluorescence imaging respectively. The regenerated lymph nodes and relevant skin were evaluated using hematoxylin-eosin (H&E) staining and the skin was quantitatively analyzed via immunofluorescence staining for lymphatic vessel endothelial hyaluronan receptor 1(LYVE-1) as well.@*Results@#One month after surgery, the right regenerated axillary lymph nodes in group B (5/5) were visible by in vivo ICG fluorescence imaging, whereas the same signals were not detected in group A (0/5). The results were the same at 2 and 6 months after surgery. HE staining showed that the cortical, paracortical, and medullary regions of the right axillary lymph nodes of the experimental group B were clear, and the lymphatic vessel structure was present, accompanied by lymphocyte infiltration. Immunofluorescence staining of the right upper limb showed that the expression of LYVE-1 in the lymphatic endothelium of the B group was significantly higher than that in group A (P<0.001) and the control group (both P<0.001).@*Conclusions@#Due to the promising consequence of regenerated lymph nodes, the procedure of autologous lymph nodes fragmentary transplantation combined with VEGF-C in athymic nude mice provides a reliable animal model for the next stage research.

Expressions of vascular endothelial growth factor C and cortactin in esophageal squamous cell carcinoma and their clinical significance / 医学研究生学报

Objective Vascular endothelial growth factor C (VEGFC) and cortactin (CTTN) have been found to be closely related to the growth of esophageal squamous cell carcinoma (ESCC), but their specific relationship has not been clearly defined up to the present time. This study aimed to investigate the effects of VEGFC and CTTN on the proliferation and apoptosis of ESCC cells. Methods Human ESCC TE1 cells were treated with normal culture medium (the blank control group), MATE transfection reagent ( the MATE group), negative control RNA and MATE reagent (the negative control group), positive control RNA and MATE reagent (the positive control group), VEGFC siRNA and MATE transfection reagent (the VEGFC siRNA group), and CTTN siRNA and MATE transfection reagent (the CTTN siRNA group). The proliferation of the ESCC TE1 cells in different groups was detected by CCK-8 assay and their apoptosis determined by flow cytometry. Results Compared with the blank control group, the ESCC cells of the VEGFC siRNA and CTTN siRNA groups showed significantly decreased expressions of VEGFC mRNA (1.00 ± 0.00 vs 0.13 ± 0.01, P < 0.05) and CTTN mRNA (1.00 ± 0.00 vs 0.29 ± 0.02, P < 0.05). The proliferation rate of the ESCC cells was remarkably lower in the VEGFC siRNA and CTTN siRNA than in the other groups (P < 0.05), and even lower in the VEGFC siRNA than in the CTTN siRNA group ([31.26 ± 5.25]% vs [46.99 ± 4.82]%, P < 0.05), but their apoptosis rate was markedly higher in the former than in the latter group ([48.41 ± 5.37]% vs [36.78 ± 4.29]%, P < 0.05). Conclusion Interfering with the expressions of VEGFC and CTTN can inhibit the proliferation and promote the apoptosis of ESCC cells, and VEGFC has an even better effect than CTTN.

Effects of miR-369-3p on the proliferation and apoptosis of bladder cancer cells by regulating the expression of VEGFC gene / 中国医师杂志

Objective To observe the effect of miR-369-3p on vascular endothelial growth factor C (VEGFC) gene in bladder cancer cells EJ and 5637 and observe its effect on cell proliferation and apoptosis.Methods Bladder cancer cell lines EJ and 5637 were transfected with miR-NC (control group) or transfected with miR-369-3p (experimental group).The target gene of miR-369-3p was predicted by Targets can target gene prediction software.Fluorescence real-time quantitative polymerase chain reaction (qPCR) was used to detect the changes of VEGFC at mRNA level.The changes of VEGFC,p-Raf-1 and phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) at the protein level were detected by Western blotting.The effect of miR-369-3p on the proliferation of bladder cancer cells was detected by cell counting kit (CCK-8) and clone formation experiment.The effect of miR-369-3p on apoptosis was detected by flow cytometry.Results After transfection with miR-369-3p,the expression of VEGFC at mRNA and protein levels was significantly decreased (P ＜ 0.05),the expression of p-Raf-1 and p-ERK1/2 protein was significantly decreased,the cell proliferation ability was significantly decreased (P ＜ 0.05),the number of clones formed was significantly decreased (P ＜ 0.05) and the apoptosis was significantly increased (P ＜ 0.01).Conclusions miR-369-3p can inhibit the proliferation and promote the apoptosis of bladder cancer cells by interfering with the expression of VEGFC gene,which may provide a new target for the biological therapy of bladder cancer.

Effects of Shuangdan Mingmu capsule on the expressions of VEGF in the retina of diabetic rat models / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To study the effects of Shuangdan Mingmu capsule on the expressions of vascular endothelial growth factor-a(VEGF-a), VEGF-b, VEGF-c in the retina of a diabetic rat model. <p>METHODS: Forty male SD rats were divided into Group A(normal group), Group B(model group), Group C(Shuangdan Mingmu group)and Group D(positive control group)10 rats(20 eyes)in each group. A rat model of diabetic retinopathy was established by one-time tail vein injection with STZ(50mg/kg). After modeling for 1wk, the rats were given medicine by gavage. After gavage for 4wk rats were sacrificed, and the expressions of VEGF-a, VEGF-b, VEGF-c in the retina tissues were detected by immunohistochemical method. <p>RESULTS: After gavage for 4wk the average gray values of VEGF-a, VEGF-b and VEGF-c protein in the retina of model group, Shuangdan Mingmu group and positive control group were lower than those of the normal group, and the average optical density were higher than those of the normal group. There was a significant difference between the model group and the normal group(<i>P</i><0.01). The average gray values of VEGF-a, VEGF-b and VEGF-c expression in Shuangdan Mingmu group and positive control group were higher than those in the model group(<i>P</i><0.05)and the average optical density value were lower than those in the model group.(<i>P</i><0.01). <p>CONCLUSION: Shuangdan Mingmu capsule could significantly reduce the expressions of VEGF-a, VEGF-b,VEGF-c in the retina and had a certain protective effect on the retina of rats in the diabetic retinopathy model.

Relationship between vascular endothelial growth factor C and lymphatic vessel density with clinicopathological features of the patients in stage Ⅱ colorectal cancer / 肿瘤研究与临床

Objective To investigate the relationship between vascular endothelial growth factor C (VEGF-C) and lymphatic vessel density (LVD) with clinicopathological features of the patients in stage Ⅱ colorectal cancer.Methods Fifty-seven tissues in stage Ⅱ colorectal cancer and 57 normal mucosal tissues in Shaanxi Provincial Cancer Hospital from January 2005 to December 2010 were collected.The expression of VEGF-C and LVD were detected by using immunohistochemical method.The relationship between VEGF-C and LVD with the clinicopathological features of colorectal cancer was analyzed.Results The expression of VEGF-C and LVD in cancer tissues was higher than that in normal tissues (6.80±1.24 vs.2.07±0.17,t =3.773,P =0.000;3.19±0.53 vs.1.84±0.26,t =2.276,P =0.025).There were statistical differences between VEGF-C and LVD in different tumor diameter (4.00±0.54 vs.11.26±2.89,t =-3.049,P =0.004;1.89±0.37 vs.5.27±1.12,t =-3.376,P =0.001) and relapse or metastasis (4.74±1.31 vs.10.62 ±2.39,t =-2.158,P =0.039;1.86±0.45 vs.5.65±1.06,t =-3.778,P =0.000).LVD in VEGF-C positive group was higher than that in VEGF-C negative group (4.16±0.64 vs.0.21±0.11,t =3.503,P =0.001).The expression of VEGF-C was positively correlated with LVD (r =0.892,P =0.000).Conclusions The expression of VEGF-C in stage Ⅱ colorectal cancer is correlated with LVD,as well as tumor size and relapse or metastasis.VEGF-C and LVD could also play important roles in the occurrence,development and metastasis in stage Ⅱ colorectal cancer.

VEGF-C Promotes Cervical Cancer Invasion via Regulation of microRNA-145/Smad Interacting Protein 1 / 中山大学学报(医学科学版)

[Objective]To investigate the role of microRNA-145/Smad interacting protein 1(SIP 1)in VEGF-C-enhanced cervical cancer metastasis.[Methods]Cervical cancer cell line SiHa cells were cultured and treated with VEGF-C to observe its effect on the expression of miR-145 and SIP1. After transfection with specific SIP1 siRNA,the invasion number of cultured cells were calculated by transwell chamber assay.[Results]Treatment with VEGF-C(100 ng/mL)for 12 h,24 h and 48 h all reduced miR-145 expression,with the expression abundance of(82.4±6.4)% (P<0.05),(72.5±7.2)%(P<0.01),and(60.6±9.6)%(P<0.001),respectively,when compared to control.Meanwhile, the same treatment with VEGF-C also increased SIP1 protein expression,with the expression abundance of(142.4 ± 16.5)%(P<0.05),(183.6 ± 11.4)%(P<0.01)and(220.8 ± 15.7)%(P<0.001),respectively. The transfection of miR-145 mimic significantly impaired VEGF-C effect on SIP1 expression. Finally,VEGF-C promoted SiHa cell invasion,which was largely inhibited by the tranfection of SIP siRNA with the inhibitory rate of(56.6±10.3)%(P<0.01).[Conclusion]VEGF-C downregulates miR-145,thus increases SIP1 expression and promotes cervical cancer cell invasion,which may contributes to cervical cancer malignant progression.

Gecko crude peptides inhibit migration and lymphangiogenesis by down regulating the expression of VEGF-C in human hepatocellular carcinoma cells and human lymphatic endothelial cells / 中国药理学与毒理学杂志

OBJECTIVE To explore the role of gecko crude peptides (GCPs) in the proliferation, apoptosis, migration and lymphangiogenesis of human hepatocellular carcinoma cells (HepG2) and human lymphaticendothelial cells (HLECs) in vitro. METHODS The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to evaluate the anti- proliferative effect of GCPs and siRNA-VEGF-C on HepG2 cells, Hoechst 33258 staining and flow cytometry were performed to analyze cycle and apoptosis. The migration and invasion ability of cells were assayed by transwell chamber experiment and wound-healing assay. The protein and mRNA expressions of vascular endo?thelial growth factor-C (VEGF-C) and CXC chemokine receptor-4 (CXCR4) were detected by q-PCR, immunofluorescence, Western blot. The protein expressions of the extracellular signal regulated kinase (ERKI/2), c-Jun N-terminal kinase (JNK), p38-mitogen activated protein kinases (p38 MAPK), serine/threonine kinase (Akt) and phosphatidylinositol- 3- kinase (PI3K) were detected by western blot. The anti-lymphangiogenesis effect of GCPs on the HLECs was analyzed using an in vitro tube-formation assay. The protein and mRNA expressions of vascular endothelial growth factor receptor-3 (VEGFR-3) and stromal cell-derived factor-1 (SDF-1) were detected by q-PCR, Western blot. RESULTS GCPs and siRNA-VEGF-C inhibited HepG2 proliferation, invasion and migration, and the most obvious inhibitory effect was both synergistic effects. Thus, GCPs suppressed HLECs proliferation, migration and tube-like structure formationin a dose- dependent manner, and had inhibitory effect of tumor- induced lymphangiogenesis in vitro. Additionally, we found that GCPs and siRNA- VEGF- C decreased the expressions of MMP-2, MMP-9, VEGF-C, CXCR4, phospho-ERK1/2, phospho-P38, phospho-JNK and PI3K in HepG2 cells. Moreover, GCPs had a dose-dependent depressive effecton the expressions of VEGFR- 3, SDF- 1 in HLECs. CONCLUSION The low expression of VEGF- C mediated by siRNA-VEGF-C and GCPs inhibit tumor proliferation, invasion and migrationby suppressing the MAPK signaling pathway through reduced levels of VEGF-C, and GCPs inhibit tumor lymphangiogenesis by suppressing the CXCR4/SDF-1 signaling pathway through suppressed VEGF-C/VEGFR-3.

Clinical value of the VEGF-C and VEGF-R3 in peripheral blood of patients with lung cancer / 国际肿瘤学杂志

Objective To investigate the relationship between the expression of vascular endothelial growth factor C (VEGF-C) and vascular endothelial growth factor receptor-3 (VEGF-R3) in peripheral blood of patients with lung cancer and the pathological characteristics, and to assess the ability to evaluate lymphatic and distant metastasis of the two marks.Methods VEGF-C and VEGF-R3 were detected by enzyme-linked immunosorbent assay (ELISA) in 124 patients with lung cancer and 30 normal controls, and used to analyze the relationship with the pathological characteristics of lung cancer.Results The serum levels [M(QR)] of VEGF-C and VEGF-R3 in patients with lung cancer were 283.57 (120.70) pg/ml and 62.72 (43.02) ng/ml, significantly higher than the control whose VEGF-C and VEGF-R3 were 234.62 (129.20) ng/ml and 43.08 (17.07) pg/ml, respectively (Z=-2.840, P=0.005;Z=-3.834, P<0.001).No correlation was found between the expression of VEGF-C and age, sex, primary tumor site, T stage (Z=-0.949, P=0.343;Z=-0.454, P=0.649;Z=-1.168, P=0.243;Z=-1.694, P=0.090).But the expression of VEGF-C was significantly related with pathologic type, N stage and M stage (χ2=8.829, P=0.012;χ2=27.148, P<0.001;Z=-2.221, P=0.026).However, the expression of VEGF-R3 was not correlated with age, sex, the site of the primary lesion, pathological type and T, N, M stage (Z=-0.558, P=0.577;Z=-0.599, P=0.549;Z=-0.703, P=0.482;χ2=1.166, P=0.558;Z=-0.680, P=0.496;χ2=0.353, P=0.950;Z=-1.523, P=0.128).Conclusion The expressions of VEGF-C and VEGF-R3 in patients with lung cancer are higher than those in normal control, and the expression of VEGF-C is related with patho-logic type, N stage and M stage.The detection of VEGF-C in peripheral blood of lung cancer is expected to be an assistant marker for the evaluation of lymph node metastasis and blood metastasis, but VEGF-R3 does not show its value.

Expression of interleukin-33 and vascular endothelial growth factor-C in gastric cancer and its clinical significance / 重庆医学

Objective To investigate the expression of interleukin-33(IL-33)and vascular endothelial growth factor C(VEGF-C)in gastric cancer tissues and serum,and to explore the relationship between these two indicators and gastric cancer lymph node metastasis.Methods The levels of IL-33 and VEGF-C in the tissues of gastric mucosa and serum were detected by immunohistochemical SP method and enzyme-linked immunosorbent assay(ELISA)in 98 patients with gastric cancer and 36 healthy subjects.Results The expression rates of IL-33 and VEGF-C in gastric cancer were 67.35％and 74.49％,which were significantly higher than the rates in normal gastric tissue(47.22％and 61.11％).The difference was statistically significant(P<0.01).The expression of IL-33 and VEGF-C was correlated with the degree of tumor differentiation,tissue infiltration,lymph node metastasis,distant metastasis and clinical stage(P<0.05).The positive rates of IL-33 and VEGF-C in gastric cancer lymph node metastasis group were higher than those in non-lymph node metastasis group(P<0.05).The serum concentrations of IL-33 and VEGF-C in patients with gastric cancer were(50.24±13.08)pg/mL and(210.73±58.35)pg/mL,respectively,which were higher than those in healthy control group(P<0.05);the expressions of serum concentration of IL-33 and VEGF-C in the cases with lymph node metastasis were higher than those without lymph node metastasis and the difference was statistically significant(P<0.05).Conclusion High levels of IL-33 in gastric carcinoma patients might induce the secretion of VEGF-C,promote lymph node metastasis,and be applied as an important index of the appraisal to the prognosis of gastric cancer.

Non small cell lung cancer patients with multiple brain metastases with erlotinib combined with whole brain radiation therapy clinical observation and effect of C on vascular endothelial growth factor level / 中国生化药物杂志

Objective To study the analysis of non-small cell lung cancer patients with multiple brain metastases with erlotinib combined with whole brain radiation therapy clinical observation and effect of C on vascular endothelial growth factor level. Methods 40 cases of non - small cell lung cancer patients with multiple brain metastases treated in Taizhou tumor hospital from January 2015 to April 2016 were selected and randomly divided into the control group and the experimental group, with 20 patients in each group. The control group and the experimental group patients were given clinical treatment of whole brain radiotherapy, the control group was given routine treatment, the experimental group received erlotinib. The clinical effects of the 2 groups were compared and analyzed. Results After the corresponding treatment, the experimental group of 20 patients, 8 cases of complete remission, 7 cases of partial remission, the number of effective treatment for 15 cases, the treatment rate was 75.0％. Of the patients in the control group, 6patients had complete remission, and 4 patients had partial remission. The effective rate was 50％. Available, the effective rate of the treatment group (75.0％) was significantly higher than that of the control group (50.0％), with statistical difference (P<0.05). The survival rate of the experimental group after one year (80.0％) was significantly higher than that of the control group (60.0％), with statistical difference (P<0.05). The level of vascular endothelial growth factor (C) in the experimental group was significantly lower than that in the control group (P<0.05). Conclusion Non small cell lung cancer patients with multiple brain metastases with erlotinib combined with radiotherapy in the clinical treatment effect of whole brain is better, can improve the survival rate in a large extent, improve the endothelial growth factor C levels, with the further promotion of the clinical significance.

Expression of CD147, vascular endothelial growth factor C in lung squamous cell carcinoma and adenocarcinoma and their clinical significances / 肿瘤研究与临床

Objective To discuss the relationships between expression levels of CD147 and vascular endothelial growth factor C (VEGF-C) in lung squamous cell carcinoma and adenocarcinoma and the clinicopathological features.Methods The expression of CD147 and VEGF-C in 57 lung cancer tissues was detected by immunohistochemical method.Results The positive expression rates of CD147 and VEGF-C in 57 lung cancer tissues were 63.16 ％ (36/57),66.67 ％ (38/57) respectively.The expression of CD147 and VEGF-C in lung squamous cell carcinoma and adenocarcinoma had no statistical difference (both P ＞ 0.05).The expression of CD147 and VEGF-C in lung squarnous cell carcinoma and adenocarcinoma had relationship with lymph node metastasis and tumor TNM stage (all P ＜ 0.05),however they had not correlation with the patient' s age,sex,smoking and tumor cell differentiation extent irrelevant (all P ＞ 0.05).The expression of CD147 was related positively with VEGF-C (P ＜ 0.01).Conclusions CD147 and VEGF-C play important roles in the progression and metastasis of lung squamous cell carcinoma and adenocarcinoma,which may have a synergistic effect.The joint detection of CD147 and VEGF-C can be used as a useful prognostic indicator in patients with lung cancer.

Effects of curcumin combined with cisplatin on growth and lymphatic metastasis of cervical cancer xeno-grafts in nude mice / 国际肿瘤学杂志

Objective To analyze the effects of curcumin combined with cisplatin on the growth and lymphatic metastasis of cervical cancer xenografts in nude mice.Methods The human cervical carcinoma Caski cells xenotransplanted tumor models were established.Inoculated mice were randomly divided into 4 groups according to random number table:control (normal saline 10 ml/kg),Cur (curcumin 100 mg/kg),Cis (cisplatin 3 mg/kg) and Cur + Cis group (100 mg/kg curcumin +3 mg/kg cisplatin).The tumor volume and anti-tumor rate were calculated.The mRNA levels of macrophage migration inhibitory factor (MIF) and vascular endothelial growth factor-C (VEGF-C) were analyzed by real-time fluorescent quantitative polymerase chain reaction (PCR).Results The inhibitory rates of Cur group,Cis group and Cur + Cis group were 40.8％,53.3％ and 60.0％.After 15 days treatment,the tumor volumes of the control group,Cur group,Cis group and Cur + Cis group were (123.44 ± 35.62),(71.72 ± 28.36),(65.47 ± 18.32),(53.44 ± 10.79) mm3.The growth rates of tumor volume in Cur group,Cis group and Cur + Cis group were slower,Cur + Cis group could more effectively inhibit tumor volume growth.The difference among the four groups had statistically significance (F =16.890,P =0.000).Real-time fluorescent quantitative PCR detection showed that compared with the control group (1.000),the MIF mRNA expressions in Cur group,Cis group and Cur + Cis group were 0.322 ±0.094,0.154 ± 0.006 and 0.136 ± 0.007,VEGF-C mRNA expressions in the three groups were 0.312 ± 0.068,0.263 ± 0.072 and 0.221 ± 0.041.The differences among groups had statistically significance (F =220.279,P =0.000;F =143.250,P =0.000).MIF mRNA was positively related with VEGF-C mRNA (r =0.815,P =0.001).Conclusion Curcumin combined with cisplatin can inhibit the growth of Carski cell xenografts in nude mice.Through the down-regulating expression of MIF mRNA and VEGF-C mRNA,cisplatin and curcumin can inhibit the lymphatic metastasis of cervical cancer,and it may be one of the important mechanisms of its anti-tumor effects.

The relationship among the expressions of vascular endothelial growth factor-C and its receptor and the cervical cancer growth and lymph node metastasis / 国际肿瘤学杂志

Objective To study the relationship between the expressions of vascular endothelial growth factor-C (VEGF-C) and vascular endothelial growth factor receptor-2 (KDR) in cervical carcinoma and the formation of cervical cancer and lymph node metastasis.Methods We selected 72 cervical carcinoma tissues,their corresponding adjacent tissues and 36 normal cervical tissues which have been resected in the Maternal and Child Health Care Hospital of Baoji of Shaanxi Province from January 2010 to December 2013.The mRNA and protein expressions of VEGF-C and KDR were examined by semi-quantitative PCR and enzyme linked immunosorbent assay in these tissues.The relationships between the expressions of VEGF-C and KDR and the formation of cervical cancer and lymph node metastasis were analyzed.Results The mRNA levels of VEGF-C in 72 cases of cervical cancer tissues and its corresponding adjacent tissues were 4.67 ± 1.05 and 2.02 ± 0.65,which were significantly higher than those in normal cervical tissues (0.36 ± 0.06),with significant differences (t =2.247,P =0.025;t =1.379,P =0.027).The protein levels of VEGF-C in 72 cases of cervical cancer tissues and their corresponding adjacent tissues were 68.30 ± 17.10 and 48.20 ± 12.70,which were significantly higher than those in normal cervical tissues (18.40 ± 10.70),with significant differences (t =4.357,P =0.016;t =6.337,P =0.012).The mRNA levels of KDR in 72 cases of cervical cancer tissues and their corresponding adjacent tissues were 3.52 ± 0.95 and 1.92 ± 0.87,which were significantly higher than those in normal cervical tissues (0.72 ±0.36),with significant differences (t =3.127,P =0.023;t =1.214,P =0.028).The protein levels of KDR in 72 cases of cervical cancer tissues and their corresponding adjacent tissues were 47.20 ± 15.60 and 38.60 ± 11.30,which were significantly higher than those in normal cervical tissues (16.40 ± 9.40),with significant differences (t =3.667,P =0.020;t =0.986,P =0.032).The expression level of VEGF-C protein in 72 cases of cervical cancer tissues was not correlated with age (x2 =0.54,P =0.17),tissue type (x2 =0.34,P =0.25),depth of invasion (x2 =5.39,P =0.08),pathological grade (x2 =0.78,P =0.11),but was correlated with tumor size (x2 =22.34,P =0.02),clinical stage (x2 =32.14,P =0.01) and lymph node metastasis (x2 =15.58,P =0.03).The expression level of its receptor KDR was correlated with tumor size (x2 =13.78,P =0.04),tissue type (x2 =32.74,P =0.01),pathological grade (x2 =13.72,P =0.04),depth of invasion (x2 =10.27,P =0.04),clinical staging (x2 =20.25,P =0.02) and lymph node metastasis (x2 =19.52,P =0.02),but was not correlated with age (x2 =4.17,P =0.09).Conclusion The expression levels of VEGF-C and KDR are correlated with the growth,invasion and metastasis of cervical cancer,which are good indicators of the lymph node metastasis.

Analysis of serum carbohydrate antigen (125,19-9),vascular endothelial growth factor C in detecting ovarian cancer lymph node metastasis / 中国基层医药

Objective To investigate the clinical value of serum carbohydrate antigen (CA125,CA199)and vascular endothelial growth factor C (VEGF-C)in early diagnosis of ovarian carcinoma.Methods 51 cases of ovarian cancer patients with retroperitoneal lymph node dissection were performed in the observation group,including 23 cases of ovarian cancer retroperitoneal lymph node metastasis,32 cases of ovarian benign tumor were selected as the control group.The serum levels of CA199 and CA125 were detected by the method of chemiluminescence detection,and the serum VEGF-C level was detected by enzyme-linked immunosorbent assay (ELISA).Results The serum level of CA125 in patients with ovarian cancer (1 682.5 ±261.5)u/mL was significantly higher than that in the control group (30.5 ±6.3)u/mL(P <0.01),serum VEGF-C level in ovarian cancer patients (2 125.6 ±96.7)pg/mL was signif-icantly higher than that in the control group (1 738.0 ±79.8)pg/mL (P <0.01 ).The serum CA199 level of 51 patients with newly diagnosed epithelial ovarian cancer was (72.5 ±30.6)u/mL,serum CA199 level of the control group was (17.4 ±8.5)u/mL,CA199 level of retroperitoneal lymph node metastasis was (134.9 ±72.5)u/mL, ovarian cancer retroperitoneal lymph node metastasis CAl99 serum level were significantly higher than those without lymph node metastasis and the control group (t =7.39,18.34,all P <0.01).The diagnostic sensitivity,specificity and accuracy of serum VEGF-C +CA199 +CA125 in detection of ovarian cancer retroperitoneal lymph node metasta-sis were 95.5%,96.5%,99.5%.Conclusion Combined detection of CA125,VEFG-C and CA199 in serum has important clinical value in predicting lymph node metastasis of ovarian cancer.

Expression of chemokine receptor CCR7 and VEGF-C is associated with prognosis of breast carcinoma / 中国病理生理杂志

AIM: To explore the protein levels of chemokine receptor 7 (CCR7) and vascular endothelial growth factor ( VEGF)-C in breast carcinoma , and to investigate the effects of CCR 7 and VEGF-C on prognosis of breast carcinoma.METHODS:The protein expression levels of CCR 7 and VEGF-C in the breast carcinoma tissues and normal breast tissues were detected by the method of immunohistochemistry .At the same time, the relationship between clinico-pathologic characteristics and the protein expression of CCR 7 and VEGF-C in the breast carcinoma tissues was analyzed . The relationship between the protein expression of CCR 7 and VEGF-C and survival time of the breast cancer patients was estimated by Kaplan-Meier method.RESULTS:The positive expression rates of CCR 7 and VEGF-C in the breast carcino-ma tissues were significantly higher than those in the normal breast tissues (P<0.01).A positive correlation was observed between the protein expression of CCR7 and the protein expression of VEGF-C in the breast carcinoma tissues (r=0.613, P<0.01).The protein expression of CCR7 and VEGF-C was correlated with lymph node metastasis and TNM stage (P<0.05), but both were not related to patients ’ age, primary tumor size, estrogen receptor and progesterone receptor .The survival time of the patients with CCR 7 and VEGF-C positive expression was significantly shorter than that of the patients without the expression (P<0.05).CONCLUSION:The positive expression of CCR7 and VEGF-C proteins is associated with the prognosis of breast cancer , and combined detection of CCR 7 and VEGF-C protein expression levels may be helpful to judge the prognosis of breast cancer .