Alteration Of Expression And Distribution Of Pericellular Matrix In Intervertebral Disc Degeneration

Objective: In this study,a rat model of tail intervertebral disc degeneration was established to study the expression and distribution of pericellular matrix in the process of intervertebral disc degeneration,which provided a new idea for the study of the mechanism of intervertebral disc degeneration.Methods: The rat tail intervertebral disc degeneration model was established by acupuncture.The nucleus pulposus tissues of the rat tail intervertebral disc were collected at the 3rd,6th,9th and 12 th weeks after the establishment of the model to observe the expression and distribution changes of the pericellular matrix around the nucleus pulposus cells at the gene level and protein level at different time points with the degeneration of the intervertebral disc.Real-Time PCR was used to detect the gene expression levels of extracellular matrix(ECM)components(Collagen Ⅱ and aggrecan)at different time points.Toluidine blue staining was used to observe the degeneration of the intervertebral disc nucleus pulposus at corresponding time points,so as to evaluate the effectiveness of the rat tail intervertebral disc degeneration model.Then Real-Time PCR was used to detect the gene expressions of Perlecan,Laminin,Nidogen and Collagen Ⅳ of pericellular matrix components around nucleus pulposus cells at different time points.Immunohistochemistry was used to observe the expression and distribution of pericellular matrix components around nucleus pulposus cells in nucleus pulposus tissues at different time points.Results: At the gene level,Real-Time PCR results showed that the Perlecan gene expression level in the control group had no significant difference at 3 weeks,6 weeks,9 weeks and 12 weeks,while in the model group,the Perlecan gene expression level at9 weeks and 12 weeks was significantly decreased compared with that at 3 weeks(P<0.01).In the control group,the gene expression levels of Nidogen-1 and Nidogen-2 were not significantly different at 3 weeks,6 weeks,9 weeks and 12 weeks.In the model group,the gene expression levels of Nidogen-1 and Nidogen-2 were significantly decreased at 9 weeks and 12 weeks compared with 3 weeks,with statistical significance(P<0.01).In the control group,there was no significant difference in Laminin gene expression at 3,6,9 and 12 weeks,but in the model group,Laminin gene expression at 9 and 12 weeks was significantly increased compared with 3 weeks,with statistical significance(P<0.01).At 3 weeks,6 weeks,9 weeks and12 weeks in the control group,there was no significant difference in gene expression levels of Col4a1,Col4a2,Col4a3,Col4a4,Col4a5 and Col4a6.In the model group,The gene expressions of Col4a1,Col4a2,Col4a3,Col4a4,Col4a5 and Col4a6 at 9 and12 weeks were significantly decreased compared with those at 3 weeks,with statistical significance(P<0.01).At the protein level,immunohistochemical section results showed that with disc degeneration,the distribution of Collagen Ⅳ,Nidogen and Perlecan proteins in the pericellular matrix of the model group was significantly reduced compared with the control group at 3,6,9 and 12 weeks at different time points,while the distribution of Laminin proteins increased significantly.According to statistical analysis,there was no significant difference in the expressions of Collagen Ⅳ,Nidogen,Laminin and Perlecan between the control group and the model group at3 and 6 weeks.Collagen Ⅳ,Nidogen and Perlecan in the model group decreased significantly compared with the control group at 9 and 12 weeks,with statistical significance(P<0.01).Compared with control group,Laminin expression level in model group was significantly increased,with statistical significance(P<0.01).Conclusion: In this study,a rat tail intervertebral disc degeneration model was established by acupuncture.It was found that the expression of Perlecan,Nidogen and Collagen Ⅳ in nucleus pulposus cells PCM was significantly decreased at the gene and protein levels,while the expression of Laminin was significantly increased with the degeneration of intervertebral disc.Further research on the mechanism of intervertebral disc degeneration may be helpful.