| Purpose:By observing the effects of Eechinops latifolius on bone mineral density,bone morphology and bone metabolism markers of postmenopausal osteoporosis（PMOP）model rats,to clarify the effect of Eechinops latifolius on PMOP.By observing the changes in the expression of inflammatory factors and related factors in PI3K/AKT/m TOR signaling pathway in postmenopausal osteoporosis model rats,the possible mechanism of preventing and treating postmenopausal osteoporosis was investigated.To provide more biological mechanism for clinical application of Eechinops latifolius to prevent and treat PMOP.Method:1.Experimental animals:4-month-old SPF SD rats,72 healthy females without pregnancy,weighing 250±20g.2.Experimental grouping:The method of random number table was used to divide 6groups,which were respectively sham group（sham group）,ovariectomized osteoporosis model group（OVX group）,Eechinops latifolius high-dose group（Echinops-H group）,medium-dose group（Echinops-M group）,low-dose group（Echinops-L group）and Herba Epimedium group（HEP group）.3.Modeling:The postmenopausal osteoporosis model was prepared by bilateral ovary extraction[1].Smear of exfoliated vaginal cells to verify successful modeling.4.Index detection:The contents of amino terminal peptide of type 1 procollagen（P1NP）,β-CTX,tumor necrosis factor-α（TNF-α）,interleukin1β（IL-1β）and interleukin6（IL-6）in serum bone metabolism index of rats were detected by ELISA.Bone mineral density was measured by Micro-CT,and morphological changes of bone tissue were observed after HE staining.Protein expressions of PI3K,AKT and m TOR were detected by Western blot and m RNA expressions of PI3K,AKT and m TOR were detected by RT-PCR.Results:1.Bone mineral density test results of rats:Compared with sham group,the proximal femur BMD of rats in OVX group was significantly decreased（P<0.05）.Compared with OVX group,BMD in Echinops-H,Echinops-M,Echinops-L and HEP groups was increased（P<0.05）.Compared with Echinops-M and Echinops-L groups,the BMD of proximal femur in Echinops-H group was different（P<0.05）.There was no significant difference in proximal femur BMD between Echinops-M group and Echinops-L group（P>0.05）.There was no significant difference in BMD between HEP group,Echinops-H group and Echinops-M group（P>0.05）.2.Observation results of bone tissue morphology of rats（HE staining）:The bone tissue structure of the sham group was basically normal,with closely arranged cells and abundant granulocytes in the bone marrow cavity visible in the visual field,and the structure was normal.There are fewer fat cells in the tissue.Compared with sham group,OVX group showed moderate abnormal bone structure,loose arrangement of cells in the bone marrow cavity,significantly reduced number of granulocytes,and a large number of fat cells in the tissue.The bone structure was basically normal and the number of adipocytes in the tissue decreased after the treatment of Eechinops latifolius and Herba Epimedium.The improvement of bone structure was most obvious with the medium dose of Eechinops latifolius.3.Compared with sham group,P1NP andβ-ctx in OVX group were significantly increased（P<0.01）.Compared with OVX group,P1NP andβ-ctx in Echinops-H,Echinops-M,Echinops-L and HEP groups were significantly decreased（P<0.01）.Compared with Echinops-M group and Echinops-L group,the Echinops-H group was significantly lower than Echinops-M group（P<0.05）.There was a significant difference between the Echinops-M group and the Echinops-L group（P<0.01）,and the Echinops-M group was lower.There was no significant difference between HEP group and Echinops-L group（P>0.05）.4.Results of serum inflammatory markers:Compared with sham group,the contents of TNF-α,IL-6and IL-1βin serum of OVX group were significantly increased（P<0.01）.Compared with OVX group,TNF-α,IL-6 and IL-1βin Echinops-H group and Echinops-M group were significantly decreased（P<0.01）,and TNF-α,IL-6 and IL-1βin Echinops-L group were significantly decreased（P<0.05）.There were no significant differences in TNF-α,IL-6 and IL-1βbetween Echinops-L group and HEP group（P>0.05）.5.Detection results of PI3K,AKT and m TOR protein in bone tissue:Compared with sham group,m RNA expressions of PI3K,AKT and m TOR in bone tissue of OVX rats were significantly increased（P<0.01）.Compared with OVX group,the expressions of PI3K,AKT and m TOR in bone tissue of Echinops-H and Echinops-M groups were significantly decreased（P<0.01）.Compared with HEP group,the expression of PI3K,AKT and m TOR proteins in Echinops-L group were different（P<0.05）.6.The detection results of PI3K,AKT and m TOR m RNA in bone tissue:compared with sham group,the expressions of PI3K,AKT and m TOR m RNA in OVX rats were significantly increased（P<0.01）.Compared with OVX group,m RNA expressions of PI3K,AKT and m TOR in bone tissue of Echinops-H and Echinops-M groups were significantly decreased（P<0.01）.The m RNA expressions of PI3K,AKT and m TOR in bone tissue of Echinops-L and HEP groups were decreased（P<0.05）.Conclusion:1.Eechinops latifolius can improve the BMD and bone morphology of PMOP model rats to achieve anti-osteoporosis effect.2.Eechinops latifolius can regulate osteocoupling,reduce bone conversion rate and increase bone strength by reducing the contents of P1NP andβ-ctx in serum of PMOP model rats,thus preventing and treating PMOP.3.Eechinops latifolius can reduce the concentrations of TNF-α,IL-6 and IL-1βin serum of PMOP model rats,and then improve the inflammatory state of PMOP model rats,alleviate bone injury,inhibit osteoclast differentiation,reduce bone resorption,and achieve the effect of anti-osteoporosis.4.Eechinops latifolius can inhibit the activation of PI3K/AKT/m TOR signaling pathway and prevent PMOP.Its mechanism is speculated to be related to inhibiting the activation of PI3K/AKT/m TOR signaling pathway,thus improving autophagy level,preventing the formation of osteoclasts,improving the activity of osteoblasts,inhibiting bone resorption,promoting bone formation,and increasing bone strength.Whether it is mediated by inflammatory factors needs to be further studied in vitro. |
PDF Full Text Request