| Objective: Through the different groups of rabbits to retain the self-ligamentum flavum,do not retain the self-ligamentum flavum and self-fat reposition of the laminectomy,the number density of fibroblasts,the percentage of collagen area,the optical density of TGF-β1 and Smad3 protein,the expression of protein and the corresponding m RNA expression level of this protein were compared at 3 and 6 weeks after operation,to explore the possible molecular biological mechanism of the inhibitory effect of self-ligamentum flavum on the formation of epidural scar after lumbar surgery,and to provide theoretical basis for better prevention of the formation of epidural scar after lumbar surgery.Methods:A total of 48 6-8 month-old Japanese white rabbits,weighing 3.1±0.4kg,were provided by the Inner Mongolia Medical University Animal Experimental Center,the rabbits which were in bad condition,could not cooperate with the feeding and could not bear the operation were excluded,and then added to 48 rabbits.The rabbits were randomly divided into three groups: the ligamentum flavum preservation group(n=16),the fat reposition group(n=16)and without ligamentum flavum preservation group(n=16).The operation models were made for different groups.The animals in each group were killed after 3 and 6 weeks respectively(8 animals in each group were killed after 3 and 6 weeks respectively).HE staining and VG staining were used to observe and calculate the number density of fibroblasts and the area percentage of collagen fibers in the epidural scar tissue of each group Immunohistochemistry,western-blot Quantitative analysis and RT-PCR were used to detect the expression of TGF-β1 and Smad3 protein,the protein content and m RNA expression level,and the above data were compared between groups to analyze the differences.Result:(1)By HE staining and light microscopic observation,it was found that the fibroblasts in the retained group were few and loosely arranged,while the fibroblasts in the non-preservation group and the fat group were more numerous and closely arranged.After cell counting,the number density of fibroblasts in 3 and 6 weeks group was lower than that in non-preservation group and fat group,and the difference was significant(p<0.05),there was no significant difference between the fat group and the non-preservation group;(2)VG staining showed that the collagen fibers in the retained group were sparse and unevenly distributed,while a lot of red collagen fibers were gathered in the non-preservation group and the fat group.After calculating the area percentage of collagen fibers in different groups,it was found that the area percentage of collagen fibers in the preservation group was lower than that in the non-preservation group and the fat group at 3 and 6 weeks(p<0.05),but there was no significant difference between the non-preservation group and the fat group.(3)The results of immunohistochemistry showed that the degree of positive staining of retained histone was significantly lower than that of the other two groups.The optical density of TGF-β1 and Smad3 in the preservation group was significantly lower than that in the other two groups at 3and 6 weeks(p<0.05),but there was no significant difference between the non-preservation group and the fat group.(4)The results of Western blot showed that the expression levels of TGF-β1 and Smad3 in the 3-and 6-week group were lower than those in the non-preservation group and the fat group(p<0.05),but there was no significant difference between the non-preservation group and the fat group.(5)The results of RT-PCR showed that the m RNA expression levels of TGF-β1 and Smad3 in the preserved group at 3 and 6 weeks were significantly lower than those in the non-preserved group and the fat group(p<0.05),but there was no significant difference between the non-preserved group and the fat group.Conclusion: There are different degrees of epidural fibrous scar formation after lumbar surgery.If the ligamentum flavum is preserved during the operation,the number of fibroblasts and the formation of collagen fibers can be reduced,thus preventing the adhesion between the scar and the Dura Sac and nerve root.The mechanism is not only mechanical blockade,but also the interference of TGF-β1/Smad3 signal transduction pathway to reduce the formation of fibrous scar. |
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