A Comparative Study On The Regulatory Mechanism Of Notch Signaling Pathway In Rats With Postmenopausal Osteoporosis

Objective: 1. The ancient literature in atrophic debility of bones "Gubi",“G uwei” and other related problems of int erpretation, discussion on osteoporosis of ancient TCM disease name and the treatment were analyzed an d summarized. 2.Notch signaling is involved in bone metabolism by directly effecting the differentiation of osteoblast and osteoclast, so that it widely relates to human develop‐ment, wounding repairmen and cell differentiation. In this research, we intended to ex‐plain the pathogenesis osteoporosis(OP) development basing on Notch signaling and todemonstrate the mechanism of Zuogui Decoction and Yougui Decoction in treating OP by observing the changes in Notch signaling induced by the above two decoctions.Material and Method: 1. Analyzing the ancient literatures on the pathogenesis and clinical manifestation of ’’Guwei’, "Gubi" and related diseases, and also the latest research literatures on the Chinese medicine treatment to osteoporosis to summarize the major formulas, especially the rules of correcting kidney in treating bone loss. 2. 84 female 3‐month old SPF taken ovariectomy to establish OP rat model. The rats were divided in to 6 groups: control group, sham group, model control group, Zuogui group, Yougui group and Fosamax group. After 12 weeks intragastric administration with Zuogui decoction, yougui decoction and Fosamax respectively, serum, kidney and femur were harvested. bone density of femur( left hind) by X‐ray examination and ALP and TRAP protein expression in serum were for evaluating OP model establishment and the efficiency of all the above medicines. Gene and protein expression of Notch1, Jagged and HES in kidney and bone were detected to analyze the involvement of Notch signaling. Analyzed the data statistically by SPSS software to compare the difference in between the groups to demonstrate OP development and the mechanism of Zuogui decoction and Youugui decoction in preventing and treating OP.Results: Article 1 1.Since no exact term for OP in ancient medical records, ‘bone atrophy’and ‘bone rheumatis ‐m’ are the best two descriptions on OP. 2.The pathogenesis of OP is blamed on kidney disorder. The syndrome differentiation and treatment is mai nly foncusing on corecting kidney, including benefiting kidney yang, nouris‐ hing kidney yin and tonifying ki dney essence. Zhongjing’ Jinguishenqi Pill and Qian YI’ s Liu‐ weidihuang Pill are the best two representitiv es. Jingyue in M‐ing Dynasty advocated ‘drawing yang from yin, drawing yin from yang’ and created Yougu i Pill and Zuogui Pill. Totally 12 herbs are included in these 2 formulas. 6 herbs are invovled in both which c ompose the major function of benefitting kidney. however, the 2 formulas are focusing on different targets. Zuogui Pill is mainly nourishing Kidney yin and essence, while Yougui Pill is mainly warming up kidney yan g and driving off the coldness. Comparing with control group, no difference in sham group(P>0.05), decreasing in model group(P<0.01). Comparing with model group, all the other groups are significantly increased(P<0.01). There’s no difference between Yougui group, Zuogui group and Fosamax group. 2. Serum ALP result of each group Comparing with control group, sham group and model group show significantly decreased(P<0.01). Article 2 1. BMD result of each group comparing with model group, all all the other groups are significantly increased(P<0.01).There’s no difference between Yougui group and Zuogui group. 3. Serum TRAP result of each group Comparing with control group, sham group and model group show significantly decreased(P<0.01). comparing with model group, all the other groups are significantly decreased(P<0.01). comparing with Zuogui group, Yougui group is significantly decreased(P<0.01).comparing with Fosamax group, no difference in Yougui group, while significant increase in Zuogui group. Article 3 1. Result of Notch1 protein expression in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01). comparing Yougui group, Zuogui group and Fosamax group are dramatically increased(P<0.01). 2. Result of Jagged1 protein expression in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).Zuogui group is higher than yougui group. 3. Result of HES protein expression in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01). Comparing with Zuogui group, Yougui group and Fosamax group are signifigantly increased(P<0.01), but no difference between Yougui group and Fosamax group. 4. m RNA expression of Notch1 in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).comparing with Fosamax group, no statistical difference in Zuogui group and Yougui group.5. m RNA expression of Jagged1 in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).comparing with Fosamax group, no statistical difference in Zuogui group and Yougui group. 6. m RNA expression of HES in bone Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).comparing with Fosamax group, no statistical difference in Zuogui group and Yougui group. Article 4 1. Result of Notch1 protein expression in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01). comparing zougui group, yuogui group and Fosamax group are dramatically increased(P<0.01). 2. Result of Jagged1 protein expression in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).Zuogui group is higher than yougui group. 3. Result of HES protein expression in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01). Comparing with Zuogui group, no difference in Yougui group. Fosamax group is higher than Zuogui and Yougui group(P<0.01). 4. m RNA expression of Notch1 in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). compar ing with model group, Zuogui group, Yougui group, and Fosa max group are signif icantly incr eased(P<0.01).comparing with Fosa max group, no statistical difference in Zuogui group and Yougui group.5. m RNA expression of Jagged1 in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).comparing with Fosamax group, no statistical difference in Zuogui group and Yougui group. 6. m RNA expression of HES in kidney Comparing with control group, no difference in sham group, decreasing in model group(P<0.01). comparing with model group, Zuogui group, Yougui group, and Fosamax group are significantly increased(P<0.01).comparing with Fosamax group, no statistical difference in Zuogui group and Yougui group.Conclusion: 1. In TCM, osteoporosis is considered referred to "Guwei" and "Gubi", and the key pathogenesis closely related to kidney. Therefore, in treating OP, correcting kidney is major method. Zuogui Pill and Yougui Pill are usually applied, half herbs of which are the same, which indicates they are working potentially on the same material foundation and mechanism. 2. Zuogui and You gui decoction have a similar drmatic efficiency in reversing bone density and decreasing ALP and TRAP in serum, which proves that benefiting kidney can cure osteoporosis. Yougui has a better eff ect in decreasing TRAP in serum, wh‐ich indicates Yougui decoction may inhibit bone asorbtion stronger th an Zuogui deco‐ction. 3. Notch signaling related indicators Notch1, Jagged1 and HES gene and protein exp‐ression in bone tissue were upregulated by Zuogui and Yougui Decoction, which mi‐ght be the mechanism of the above 2 decoc tions in preventing and cureing OP. 4. Notch signaling related indicators Notch1, Jagged1 and HES gene and protein exp‐ression in kidney tiss ue were upregulated by Zuogui and Yougui Decoction, which m‐ight be the mechanism of the above 2 dec octions in preventing and cureing OP. 5. Notch signaling related indicators Notch1, Jagged1 and HES gene and protein expr‐ession in kidney and bone were upregulated by Zuogui and Yougui Decoction, which indicates that is bone development is closl y related to kidney. The consistent chan‐ge of Notching signaling in bone and kidney reveals Notch signali ng is one of the bi‐ological evidences to support ‘kidney governs bone’ theory.