Effects And Mechanism Reaserch Of Yaotu Keli On Protrusion Of Lumbar Intervertebral Disc Based On The Signal Path Of NF-κB

1 Objective This study intervention the human nucleus pulposus cells by serum which containing Yao Tu Ke Li firstly, and the purpose was to investigated mechanism of traditional Chinese medicine Yao Tu Ke Li for treatment lumbar disc herniation, which by means of detected the changes in key protein expression included the Type II collagen, TypeⅠcollagen, proteoglycans, inflammatory cytokines interleukin-8, tumor necrosis factor and p-P65, P50, IΚB-α, IKK-β in NF-κB signaling pathway etc.2 Methods The eight New Zealand rabbits were randomly divided into high-dose group,middle dose group, low dose group, normal saline group and two rabbits in each group.Each group feded with decoction of Yao Tu Ke Li, dose were as follows: high dose group feded with 40 ml, middle dose group 20 ml, low dose group 10 ml. Saline group fed with 10 ml of normal saline.Manufacture the containing pharmaceutical ingredients serum collected blood from carotid artery in 2h after the last gavage when continuous gavage 7 days. Obtain human nucleus pulposus cells in surgery and cultured primary, than identified the cells by Toluidine blue staining, safranin O staining and immunity histochemical staining. At the third generation of nucleus pulposus cells were cultured in serum containing 10% pharmaceutical ingredients 24 h,nucleus pulposus cells in each group were extracted RNA,and reverse transcription,detected Type II collagen, TypeⅠcollagen, proteoglycans, IL-8, TNF-α, P50, IΚB-α,IKK-β expression level by Real-time PCR, WB carried out to detected the expression of p-P65, P50, IΚB-α, IKK-β in NF-κB signaling pathway behind cellular proteins in each group extracted, finally, observed the effects of Yao Tu Ke Li on its expression.3 Result3.1 The culture and identification for primary generation nucleus pulposus cells of humanThe form of primary generationnucleus pulposus cells of human was mostly rotundity and fusiform after adherent, and with abundant cytoplasm, the nuclei of the nucleus pulposus cells was clear. After passage, cells were polygonal, large nuclei,nucleoli mostly 2-3 gradually. Cells were sky-blue color under toluidine blue staining and red under the safranin O staining, cytoplasm was brown and blue nucleus in type II collagen immunohistochemical staining.3.2 The expression of human nucleus pulposus cells Type II collagen, TypeⅠcollagen,proteoglycans after intervention by containing pharmaceutical ingredients serumCompared with normal saline group, the type II collagen and proteoglycans of low dose group, middle dose group, high dose group, expression levels increased relatively,and expression levels of type I collagen was decreases. And with the increase dose of fed Yao Tu Ke Li, type II collagen and proteoglycan expression gradually increased and TypeⅠcollagen gradually decreased, the difference was statistically significant(P<0.05).3.3 Human nucleus pulposus cells, IL-8, TNF-α expression after intervention by containing pharmaceutical ingredients serumThe IL-8, TNF-α relative expression levels decreased in low dose group, middle dose group and high dose group compared with normal saline group. And with the dose increased of Yao Tu Ke Li, the expression level decreased gradually, the difference was statistically significant(P <0.05).3.4 Human nucleus pulposus cells Ip-P65 、 P50 、 IΚB-α 、 IKK-β expression after intervention by containing pharmaceutical ingredients serumLow dose group, middle dose group, high dose group p-P65, P50, IΚB-α, IKK-βrelative expression levels decreased, and with the increase of fed Yao Tu Ke Li dose,the expression level decreased gradually compared with normal saline group, the difference was statistically significant(P <0.05).4 Conclusion4.1 The effect of containing pharmaceutical ingredients serum of Yao Tu Ke Li for human nucleus pulposus cells.4.1.1 The containing pharmaceutical ingredients serum of Yao Tu Ke Li increased expression of type II collagen and proteoglycan, decrease the expression of type I collagen;4.1.2 The containing pharmaceutical ingredientsserum of Yao Tu Ke Li reduced expression of IL-8, TNF-α and inflammatory factors.4.1.3 The containing pharmaceutical ingredientsserum of Yao Tu Ke Li reduced expression of P50、p-P65、IΚB-α、IKK-βand suppressed start of NF-κB signaling pathway.4.2 The mechanism of Yao Tu Ke Li for lumbar disc herniation4.2.1 Yao Tu Ke Li slow down the nucleus pulposus cell aging, degeneration of nucleus pulposus;4.2.2 Adjust cytokines and suppressed start of local inflammatory reaction, enhanced the anti-inflammatory effect;4.2.3 Inhibit the activation of NF-κB signaling pathway, and reducing the local inflammatory reaction disc.