| Objective:To observe the effect of ShengyangYiwei Decoction on the microvessel density?MVD?and the expression of tumor suppressor gene P16,wild-type P53 gene and protein in rats with PLGC model.To explore its molecular biology mechanism for the intervention and treatment of precancerous lesions,and to provide experimental and theoretical basis for the clinical application of Shengyang Yiwei Decoction in related fields.Methods:Ninety SPF Wistar rats were divided into blank group,model group vitamins group,and the high,middle,low-dose groups of Shengyang Yiwei Decoction according to the random number table.15 in each group,except blank group.The remaining rats of each group were exposed to a concentration of 150?g·ml-11 MNNG solution for free drinking,while ranitidine was dissolved in saline at a concentration of 2.25g·L-11 at 60?once daily at a dose of 1mL·100g-1.For 12 consecutive weeks,the PLGC model was successfully replicated.In the twelfth week,the high-,middle-,and low-dose ShengyangYiwei Decoction groups and vitamins group were started to be subdivided with different concentrations of Shengyang Yiwei Decoction andvitamins,once daily,blank group and model group were intragastrically administrated with equal volume of normal saline,once daily for 6 weeks,and18 weeks fasted for 24 hours.The animals were sacrificed by chloral hydrate anesthesia and were stained with hematoxylin and eosin?HE staining?.The pathological status of gastric mucosa was observed.CD34 was detected by immunohistochemical method to observe the microvessel density of gastric mucosa.The expression of P16 and wild-type P53 protein in gastric mucosa was detected by immunohistochemistry.The level of P16 in gastric mucosa was detected by fluorescence quantitative PCR.Transcription of wild-type P53 using immunohistochemistry to detect gastric mucosa The relative expression of P16 and P53.Results:1.The general state of change in each group of rats.The rats in the blank group had normal diet,a good mental state,shiny skin,normal body weight,and grayish-white granules in the stool.Compared with the blank group rats,the rats in the model group had a reduced diet,poor mental status,heavier piles,rough coats,slower body weight gain,smaller body size,and unformed stool.The litter was heavier than the blank group.The rats in the drug group improved their diets compared with those in the model group,increased the activity,showed stools,and reduced the odor of the litter.2.The pathological changes of gastric mucosa in rats in each group.In the blank group,the gastric mucosa thickness was normal,and the gastric mucosal epithelial cells showed a single columnar column with clear or small vacuoles in the cytoplasm.The boundaries with the adenomatous ducts were clearly defined,and the glands were arranged neatly with uniform size and shape.Compared with the blank group,the model In the group,the gastric epithelial atrophy became thinner,the number of glands decreased,the inflammatory cells infiltrated in the interstitium,the number of red blood cells increased,the structure of the glands was irregular,the proportion of nucleus pulposus increased,and the cystic dilatation of individual glands appeared.Compared with the model group The treatment of the gland structure is more regular,inflammatory cell infiltration is reduced,the number of glands gradually increased,the pathological state of gastric mucosa tends to trend,of which the high dose group of the sun Yang Yiwei soup is most obvious.3.Changes in MVD of rats in each group.Compared with the blank group,the number of microvessel density in the model group was significantly reduced?P<0.01?.Compared with the model group rats,the number of microvessels in the vitamin group and the middle-dose group of Shengyang Yiwei Decoction was significantly different.The difference was significant?P<0.05?.The number of microvessels in the high dose group of Shengyang Yiwei Decoction increased significantly?P<0.01?.There was no significant difference in the number of microvessels in the low-dose group?P>0.05?.4.Immunohistochemical detection of P16 and wild-type P53 protein expression in each groupCompared with the blank group,the expression of P16 and P53 protein in the model group significantly decreased,and the difference was extremely significant?P<0.01?.Compared with the model group,the expression of P16 protein was significantly increased in the Vitrin group and the middle-dose group of Shengyang Yiwei Decoction,and the difference was significant?P<0.05?.The expression of P16 protein in high dose group of Shengyang Yiwei Decoction was significantly increased?P<0.01?.There was no significant difference in the expression of P16 protein in low dose group of Shengyang Yiwei Decoction?P>0.05?.Compared with the model group,the expression of P53 protein was significantly increased in the Vitrin group and the low dose group of Shengyang Yiwei Decoction?P<0.05?.The expression of P53 protein in the high-and middle-dose group of Shengyang Yiwei Decoction was significantly increased,and the difference was extremely significant?P<0.01?.4.Expression of P16 and P53 mRNA in gastric mucosa of rats in each group by fluorescence quantitative PCRCompared with the blank group,the expression of P16 and P53 mRNA in the gastric mucosa of the model group were all decreased,and the difference was statistically significant?P<0.05?.Compared with the model group,the expression of P16 mRNA in the high-,middle-dose group and the retinulin group of Shengyang Yiwei Decoction significantly increased,and the difference was statistically significant?P<0.05?.The expression of P16 mRNA in the low dose group of Shengyang Yiwei Decoction did not increase significantly?P>0.05?.Compared with the model group,the expression of P53 m RNA in gastric mucosa of each treatment group was significantly increased,and the difference was statistically significant?P<0.05?.5.Western blot analysis of relative expression of P16 and P53 protein in ratsCompared with the blank group,the expression of P16 and P53 protein in the gastric mucosa of the model group was significantly decreased,and the difference was statistically significant?P<0.05?.Compared with the model group,the expression levels of P16 and P53 in the vitamine group and the high dose group of Shengyang Yiwei Decoction significantly increased,and the difference was statistically significant?P<0.05?.Although the expression of P16 and P53protein in the low dose group of Shengyang Yiwei Decoction increased,there was no significant difference?P>0.05?.ConclusionFirst of all,in this study,the PLGC model was duplicated by the MNNG-based complex modeling method.For 6 consecutive weeks,combined with pathological mucosal status analysis,the PLGC model was successfully replicated,greatly shortening the previous modeling time.After intervention of Shengyang Yiwei Decoction,the MVD in gastric mucosa and wild-type P53 and P16 were significantly increased.It is suggested that Shengyang Yiwei Decoction may improve the expression of tumor suppressor gene P16 and wild-type P53 by improving the local blood circulation of gastric mucosa,and achieve the purpose of intervention and treatment of PLGC. |
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