Effect Of Bujingyishipian On Scleral Fibroblast In Myopia Mice Based On Single-cell RNA Sequencing Technology

Research ObjectiveTo explore the improvement of the biological function of scleral fibroblasts in myopic mice by Bujingyishi,and to provide a theoretical basis for the early intervention of myopia by Bujingyishi.Research methodsNormal and healthy C57BL/6J mice were randomly divided into the negative control group,the myopia model group and the traditional Chinese medicine intervention group.Except for the negative control group,the anterior segment of the eyes was not covered,and the myopia model group and the traditional Chinese medicine intervention group all used translucent eye masks.Right eye form deprivation was performed,and the animal eye A-ultrasound was used to measure the axis of the eye at the beginning of the experiment,three weeks and six weeks.At the third week of the experiment(after the success of myopia modeling was proved),the traditional Chinese medicine intervention group was suspended with Bujingyi optic tablets The liquid was intragastrically administered for three weeks,and the other two groups were intragastrically administered with distilled water;after the experiment,the right eye was taken,and scleral fibroblasts were captured by single cells for RNA-seq,differentially expressed genes were screened for GO and KEGG analysis.Research result1.A total of 169 different genes were identified between myopia model group and negative control group,among which 112 genes were up-regulated and 57 genes were down-regulated.GO functional analysis showed 727 GO items,466 of which were statistically significant.Among them,16 GO items related to hypoxia,23 GO items related to lipids,17 GO items related to inflammation,and 38 GO items related to protein.A total of 192 pathways were detected by KEGG analysis,of which 39 pathways were statistically significant.Pathways with high expression were: PPAR signal pathway and HIF-1 signal pathway associated with hypoxia,PI3K/ Akt signal pathway,leukocyte transendothelial transport,TNF signal pathway,MAPK signal pathway associated with inflammation,protein digestion and absorption associated with ER stress,Huntington’s disease,Parkinson’s disease,etc.2.A total of 176 different genes were identified in the TCM intervention group compared with the myopia model,among which 75 genes were up-regulated and 101 genes were down-regulated.GO functional analysis showed 581 GO items;454 GO items were statistically significant;There were 14 GO items related to oxidative phosphorylation,16 GO items related to lipid metabolism,and 48 GO items related to protein regulation.A total of 192 pathways were detected by KEGG pathway analysis,32 of which were statistically significant.Among them,PPAR signaling pathway,peroxisome,glutathione metabolism and other oxidation channels were continuously expressed,MAPK,TNF-α and NF-κB signaling pathways related to inflammation,as well as various amino acid metabolism and degradation,TGF-β signaling pathways were involved in protein synthesis.3.A total of 88 different genes were analyzed and significantly expressed in the pairwise comparison of the three groups(P > 0.05).Through GO and KEGG analysis,it was found that Hsp90b1,Lrp6,Col12a1,COL1a2 and Apoa2 showed a downregulated trend in the myopia model group compared with the negative control group,and up-regulated trend in the Chinese medicine intervention group compared with the myopia model group.Analysis conclusion1.The dysfunction of scleral fibroblasts appeared in the development of myopia,which is related to the regulation of gene expression and pathways related to hypoxia,inflammation,lipid metabolism and abnormal protein regulation.2.Bujingyishi tablet can slow down the oxidative stress and inflammatory damage of scleral fibroblasts in myopic mice,balance endoplasmic reticulum stress and autophagy,so as to maintain the normal activities of scleral fibroblasts.