Objective To Study The Inhibitory Effect Of Resveratrol On The Angiogenesis Of Glioma By Targeting Pericytes

Objective The purpose of this study was to investigate the effect of resveratrol on the survival of human glioma xenografts in nude mice and to figure out how resveratrol exerts inhibitory effects on glioma angiogenesis by targeting pericytes,which can provide experimental basis for further development and clinical application of resveratrol.Methods1.Stable U87 glioma cells were cultured and the orthotopic transplantation model of human glioma was established.The patients were randomly divided into two groups:(1)RES drug treatment group;(2)CMC solvent control group,12 only.After the establishment of the model,the control group was given RES: 40 mg /(kg · d).The CMC group was given 0.5% CMC by 0.2ml /(20g · d).The above two groups were administered continuously for 4 weeks.2.We measured the weight of nude mice every 5 days,and observed the vital signs and the living status of the model animals.Until we found nude mice appeared cachectic state,killed it as soon as possible then collected the tumor specimens.After all the specimens were collected,the animal survival curve was drawn.3.To calculate the size of orthotopic transplanted tumor in nude mice,and to observe the pathological morphology of tumor cells.4.Tumor microvessel endothelial cells and pericytes were labeled with CD34 and ?-SMA respectively.The microvessel density(MVD)of the transplanted tumor was analysised,which was CD34+-MVD.Microvascular pericyte number density(MPND)were also detected and analyzed by immunohistochemical method.5.The CD34+and ?-SMA+ immunohistochemical staining scores(IRS)were calculated and the differences between the two groups were compared.6.The morphological changes of pericytes were observed by transmission electron microscopy(TEM).The volume density(Vv),specific surface(?),specific membrane(?m)of mitochondria in pericytes were measured and analyzed by using the international common stereotaxic point analysis method.Compare the differences between the two groups.Results1.The orthotopic transplantation model of human glioma cells in nude mice was established successfully.2.Weight change map and survival curve The results showed that the body weight of the control group showed a steady increase in body weight during resveratrol treatment and after treatment,and the body weight of the solvent control group showed a slight fluctuation in the early stage of the experiment,but the overall presentation Down trend.SPSS statistical analysis showed that the treatment group(16.62 ± 1.59),the control group(16.74 ± 1.49),p> 0.05,there was no difference in body weight change between the two groups.The survival time of two groups of experimental animals was analyzed statistically.The results showed that the survival time of the two groups of glioma orthotopic transplantation was significantly different after resveratrol treatment(53.41 ± 11.62),veratrum(71.67 ± 20.35),the difference was statistically significant(P<0.05).Resveratrol can increase the survival time of tumor-bearing nude mice,but its effect on body weight is not obvious.3.Tumor volume and cell morphology Calculated two groups of tumor volume found that resveratrol treatment group significantly reduced tumor volume,or even disappeared.The results of statistical analysis showed that the solvent control group(67.60 ± 34.22)and resveratrol group were(18.27 ± 22.45),the difference was statistically significant(P <0.05).Pathological sections showed that the control group showed bulging white matter,invading white matter and brain bone,the tumor occupying the phenomenon of obvious,the internal position of the brain to the contralateral side of the serious,inoculated tumor contralateral brain tissue compression serious deformation,loss of the original structure Location;treatment group brain tissue normal,normal structure,most of the tumor decreased,or disappeared,did not cause normal brain tissue caused by oppression or destruction of the original brain tissue structure.HE staining showed that glioma cells were infiltrating,and glioma cells had cell dysplasia and structural atypia.The cells were different in size and shape.There were spindle shape,oval shape,caterpillar shape and so on.Showing a mixed staining;nuclear abnormalities,large nuclear deep dye,increased the proportion of nuclear and nuclear,mitotic like,can be seen giant nucleus,multi-core or strange nuclear and other pathological mitotic like.Part of the resveratrol treatment group tumor HE staining can be seen fence-like tumor necrosis zone,see a large number of fragmented nuclei.Plant antitumor agent Polygonum cuspidatum extract resveratrol can reduce nude mice orthotopic transplantation of glioma volume,causing fence-like necrosis.4.MVD and MPND The results of microvessel density statistics showed that the results of CD34 +-MVD(34.00 ± 2.70)and resveratrol treatment group were(25.43 ± 1.54)in the solvent control group,the difference was statistically significant(P <0.05).The number of MPND(14.40 ± 0.95)in the solvent control group was(16.12 ± 1.60)in the resveratrol group,and the difference was statistically significant(P <0.05).Plant antitumor agent Polygonum cuspidatum extract resveratrol can reduce tumor microvessel density and increase pericyte cell perivascular cell coverage.5.Immunoreactive score IRS and ?-SMA staining were(8.36 ± 1.90)and(6.91 ± 1.63)respectively in the solvent control group,and the results of resveratrol treatment group were significantly decreased(5.55 ± 1.50)And(4.58 ± 1.14),the difference was statistically significant(P<0.05).Plant antitumor agent Polygonum cuspidatum extract resveratrol can reduce tumor immune response to vascular endothelial cells and pericytes.6.The ultrastructure of pericytesObserved and compared the two groups of transmission electron microscopy results found that the treatment group of peritoneal cytoskeleton disorders,peritoneal fake small,less protrusion,cytoplasm limitations,cell space expansion;chromatin condensation,marginal phenomenon;collagen fibers decreased,;The number of mitochondria is reduced,the structure is irregular in swelling,vacuolization,irregularities of the ridge and membranous form become shorter and even disappear;the surrounding cells Of the endoplasmic reticulum shorter,part of the ribosome shedding or deletion;autophagosome formation.In the control group,the periplasm of the control group was more abundant,and the protrusions were abundant and overlapped with each other.The cytoplasm was abundant and diffuse around the vascular endothelial cells.The mitochondria were abundant,the collagen fibers were abundant,the intercellular cells were closely connected,,Gap junctions,desmosomes and fusion connections,and other complex connections;Golgi rich,rich vesicles rich,long endoplasmic reticulum,rich ribosomes.The results showed that the Vv,? and ?m of the treatment group were(22.00 ± 1.68),(2.41 ± 0.50)and(40.10 ± 2.26),respectively.The Vv,? and ?m of the control group were(37.70 ± 3.03),(1.20 ± 0.60)and(43.75 ± 1.19),respectively.The difference of Vv and ?m in resveratrol treatment group was significantly decreased and ? was significantly increased(P <0.05),the difference was statistically significant(P <0.05).Plant antitumor agent Polygonum cuspidatum extract resveratrol can cause peritoneal cytoskeleton disorder,single connection;pericardium pseudopodia,collagen fibers,Golgi apparatus and secretion of vesicles to reduce;mitochondrial swelling,vacuolization,mitochondrial reduction with the function reduce.Conclusion1.The oral plant antitumor agent Polygonum cuspidatum extract resveratrol has the effect of increasing the survival time of nude mice bearing human glioma orthotopic transplantation,reducing the tumor volume and inhibiting the growth of glioma.2.Polygonum cuspidatum extract resveratrol can induce intercellular mitochondria and other ultrastructural disorders and functional reduction;to promote autophagy formation of pericytes;to improve cell coverage,reduce microvascular density and endothelial cell and peripheral cell immune response scores,which can play a role in inhibiting the angiogenesis of glioma in nude mice in situ and then produce anti-glioma effects.