The Adjustment Of Male Rats' Bone Remodeling After The Intervention Of NFATc1 Blocker By Electropuncture At Foot-shaoyang Meridian

Objective: In this study,bone tissue sections of FK506(signal blocker)rats were further observed after the electroacupuncture at foot-yangming meridian,and the expression of bone remodeling-related transcriptional genes was detected,so as to explore the effect of electroacupuncture on bone remodeling-related signaling molecules,transduction pathways and their upstream and downstream in osteoporotic rats.Methods: Forty male clean-grade SD rats(age,10 months;body weight,310 ± 10 g)were provided by the Laboratory Animal Center of Southwest Medical University.The rats were divided into blank group(n = 10)and model group(n = 30)using the random number table generated by searching Excel after the numbering of Arabic numerals.The blank group received any treatment.The model group was further randomly divided into model control group(FK506 group)ad model + pre-electroacupuncture group(pre-electroacupuncture + FK506 group)and model + electroacupuncture group(electroacupuncture + FK506 group).The model group was treated with subcutaneous injection of FK506 solution(injected dose,0.3ml/kg;FK506 concentration,1 mg/ml)for 7 days.The pre-electroacupuncture + FK506 group was treated with electroacupuncture at foot-yangming meridian before injection.Three days after the establishment of FK506 model,the electroacupuncture + FK506 group and the pre-electroacupuncture + FK506 group were intervened with electroacupuncture at foot-yangming meridian(Huantiao point,Yanglingquan point,Xuanzhong point).The electroacupuncture was performed with dilatational wave at 20 Hz(1 time/d,15 min/time,interruption for 2 d after 5 times,a total of 6 courses of treatment).HE staining of bone tissue sections and the ratio of trabecular bone area were observed under a light microscope after the rats were put to death,and the mRNA expression of key genes involved in bone remodeling,such as OPG,RANKL,NFATc1 and CBFa1 was detected using RT-PCR.Results:(1)Trabecular bone in the bank group presented close arrangement in lumps and patches,and was connected into reticulation with good continuity.No obvious changes were found in the bone marrow cavity.Compared with the blank group,trabecular bone in the FK506 group showed disordered arrangement,significantly reduced number,poor continuity,narrowed width and obviously enlarged spacing between trabecular bones,and the bone marrow cavity was enlarged significantly.Compared with the FK506 group,trabecular bone in the electroacupuncture + FK506 group presented regular arrangement,good continuity,significantly increased number,widened width and narrowed spacing between trabecular bones,and the bone marrow cavity was diminished significantly,which showed no obvious differences with the pre-electroacupuncture + FK506 group.Compared with the FK506 group,trabecular bone in the pre-electroacupuncture + FK506 group showed regular arrangement,good continuity,significantly increased number,widened width and narrowed spacing between trabecular bones,and the bone marrow cavity was diminished significantly.(2)The ratio of trabecular bone area showed statistically altitudinal differences among the blank group,the pre-electroacupuncture + FK506 group,the electroacupuncture + FK506 group and the FK506 group(P = 0.006).Compared with the blank group,the ratio of trabecular bone increased in the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group,without statistically significant differences(P > 0.05).Comparison between groups revealed no obvious difference between the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group.(3)CBFa1 mRNA expression in bone in the FK506 group was higher than that in the blank group,without statistical significance(P > 0.05).Compared with the FK506 group,CBFa1 mRNA expression decreased in the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group,without statistical significance(P > 0.05).Compared with the pre-electroacupuncture + FK506 group,CBFa1 m RNA expression showed no statistical significance in the electroacupuncture + FK506 group(P > 0.05).(4)RANKL mRNA expression in bone in the FK506 group was obviously higher than that in the blank group,with statistical significance(P < 0.05).Compared with the FK506 group,RANKL m RNA expression decreased significantly in both the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group,with statistical significance(P < 0.05).Compared with the pre-electroacupuncture + FK506 group,RANKL mRNA expression showed no statistical significance in the electroacupuncture + FK506 group(P > 0.05).(5)OPG mRNA expression in bone showed no obvious different between the FK506 group and the blank group(P > 0.05).Compared with the FK506 group,OPG mRNA expression presented no obvious differences in the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group(P > 0.05).Compared with the pre-electroacupuncture + FK506 group,OPG mRNA expression showed no statistical significance in the electroacupuncture + FK506 group(P > 0.05).(6)NFATc1 mRNA expression in bone in the FK506 group was slightly higher than that in the blank group,without statistical significance(P > 0.05).Compared with the FK506 group,NFATc1 mRNA expression presented no obvious differences in the pre-electroacupuncture + FK506 group and the electroacupuncture + FK506 group(P > 0.05).Compared with the pre-electroacupuncture + FK506 group,NFATc1 mRNA expression showed no statistical significance in the electroacupuncture + FK506 group(P > 0.05).Conclusion: Electroacupuncture at foot-yangming meridian can regulate the coupling of bone remodeling,and thereby balance bone formation and bone resorption.The mechanism may be explained by regulating body metabolism in the whole body to produce and regulate bone remodeling and reconstruct the net effect of normal coupling through two main transcriptional factors RANKL and NFATc1.The subcutaneous injection of FK506 solution with a high concentration can block the mRNA expression of NFATc1,and over-express CBFa1 mRNA,resulting in the high mRNA expression of RANKL,which can induce the establishment of Bone remodeling decoupled model in animals successfully.Pre-acupuncture and electroacupuncture can not reverse the blocking of FK506 on NFATc1 signaling,and their prevention and treatment effect shows no significant differences;however,they can prevent the high expression of CBFa1 and down-regulate the mRNA expression of RANKL.