Effects Of Liuwei Dihuang Pills On Intervertebral Disc Degeneration In Rats With Diabetes Mellitus

Objective:To study the effect of diabetes on intervertebral disc degeneration,and to explore the mechanism of Liuwei Dihuang Pill in the prevention and treatment of diabetes intervertebral disc degeneration,so as to provide basis for Liuwei Dihuang Pill in the treatment of diabetes low back pain.Methods:The cases of patients with lumbago and leg pain in the orthopedic outpatient department of our hospital were collected and divided into control group and diabetes group.The height of each lumbar intervertebral space was measured based on CT reconstruction images,and the lumbar intervertebral disc degeneration score was performed based on lumbar MRI.In the topic,Liuwei Dihuang Pills was selected as the intervention drug in DM model group.The models of DM rats were constructed.A total of 60 SD rats were divided into TCM group(n=20),observation group(n=20)and control group(n=20).The observation group and TCM group were intraperitoneally injected with STZ sodium citrate solution(30 mg/kg),while control group was intraperitoneally injected with the same dose of sodium citrate solution as placebo.After successful modeling,TCM group was given intragastric administration of Liuwei Dihuang Capsule suspension(26 mg/kg),while control and experimental groups were given the same volume of normal saline(once/d for 8 weeks and 16 weeks).Half of the specimens were collected at 8 weeks and the others were 16 weeks after intragastric administration,and L5/6 intervertebral disc was fixed with paraformaldehyde.L5/6 intervertebral discs and bilateral vertebrae were scanned by CT to collect data.L5/6 intervertebral disc tissues were stained by hematoxylin-eosin(HE),their morphological changes were observed and scored under optical microscope.The changes in expressions of silencing signal regulator protein 6(SIRT6),type I collagen(Col-1),type Ⅱcollagen(Col-2),proteoglycan,matrix metalloproteinase-3(MMP3)and matrix metalloproteinase-13(MMP13)proteins in L5/6 intervertebral disc tissues were detected by immunohistochemistry.The changes in expressions of SIRT6,Col-1,Col-2,proteoglycan,MMP3 and MMP13 mRNA in L5/6 intervertebral disc tissues were detected by real-time fluorescent quantitative PCR.The effects of Liuwei Dihuang Pills on the pathology of L5/6 intervertebral discs in DM rats,and the changes in expression of SIRT6,Col-1,Col-2,proteoglycans,MMP3 and MMP13 were explored.Results:Compared with control group,the L2/3,L3/4,L4/5,and L5/S1 intervertebral space heights of the diabetic patients were significantly decreased;the L2/3,L3/4,L4/5,and L5/S1 intervertebral disc degeneration scores were significantly increased.The effects of Liuwei Dihuang Pills on the pathology of intervertebral disc in DM rats were observed by HE staining.And the results showed that after 8 weeks and 16 weeks of intragastric administration of Liuwei Dihuang Pills,there was no significant change in L5/6 intervertebral disc tissues in control group,and there were significant degeneration characteristics in observation group,indicating that DM could cause intervertebral disc degeneration and modeling was successful.After intervention with Liuwei Dihuang Pills,degeneration pathology of L5/6 intervertebral disc tissues was significantly improved in TCM group compared with observation group.After 8 weeks and 16 weeks of intragastric administration,histology scores of L5/6 intervertebral disc morphology were increased in observation group compared with control group,which were decreased in TCM group compared with observation group(P>0.05).The factors affecting the normal growth and differentiation of L5/6 intervertebral disc tissues were detected by immunohistochemistry and real-time fluorescence quantitative PCR.And the results showed that expression levels of SIRT6 and proteoglycan mRNA and proteins were significantly decreased,while expression levels of MMP13 mRNA and proteins were significantly increased in observation group compared with control group after 8 weeks and 16 weeks of intragastric administration(P<0.05).Compared with observation group,expression levels of SIRT6 and proteoglycan mRNA and proteins were significantly increased,while expression levels of MMP13 mRNA and proteins were significantly decreased in TCM group(P<0.05).Conclusions:Through clinical research,it can be found that the intervertebral disc degeneration of diabetes patients is significantly more severe than that of non diabetes patients with similar age.The experimental results showed that after the intervention of Liuwei Dihuang pill,the morphology and score of the intervertebral disc of DM rats were significantly improved by up regulating the expression of SIRT6,COL-1,col-2,proteoglycan protein,and down regulating the expression of MMP3 and MMP13,suggesting that Liuwei Dihuang pill can effectively alleviate the intervertebral disc degeneration of DM rats,which is expected to be used as a drug for the clinical treatment of intervertebral disc degeneration in DM patients,and provide a theoretical basis for the clinical treatment of intervertebral disc.