骨碎补总黄酮对去卵巢大鼠RANKL/OPG平衡及MAPK信号通路的影响

Effects of drynaria total flavonoids on RANKL/OPG balance and MAPK signaling pathways in overiectomized rats

目的观察骨碎补总黄酮对去卵巢大鼠核因子κB受体活化因子配体(RANKL)/骨保护素(OPG)表达及丝裂原活化蛋白激酶(MAPK)信号通路的影响。方法选用雌性大鼠45只,分为假手术组(15只)和手术组(30只),手术组行双侧卵巢切除术,造模12周时测定BMD,确认OP复制成功后,将手术组大鼠又随机分为模型组和骨碎补总黄酮给药组,每组均15只;给药组大鼠给予骨碎补总黄酮灌胃,持续时间为12周,模型组和假手术组同时给予等量生理盐水灌胃;实验结束后测定大鼠的BMD,ELISA法检测大鼠血清RANKL、OPG含量,RT-PCR检测大鼠骨组织RANKL、OPG m RNA表达,Western blot法检测骨组织c-Jun氨基末端激酶(JNK)、p38MAPK(p38)、细胞外调节激酶(ERK)的磷酸化水平及c-Fos表达。结果干预治疗12周后:(1)与假手术组相比,模型组大鼠左、右股骨BMD降低(P<0.05),血清RANKL含量增加、OPG含量降低、RANKL-OPG比值增大(P均<0.05),骨组织RANKL m RNA表达增加、OPGm RNA表达减少(P均<0.05)。(2)与模型组比较,假手术组和骨碎补总黄酮治疗组左、右股骨BMD均增加(P均<0.05),血清RANKL含量降低、OPG含量增加、RANKL-OPG比值减小(P均<0.05),骨组织RANKL m RNA表达减少、OPG m RNA表达增加(P均<0.05),且假手术组和骨碎补总黄酮治疗组之间比较差异有统计学意义(P<0.05);(3)与模型组比较,假手术组和骨碎补总黄酮治疗组大鼠骨组织P-JNK、c-Fos表达降低(P均<0.05),但这两组间比较差异无统计学意义(P>0.05)。结论骨碎补总黄酮可增加去卵巢大鼠骨密度,其途径可能是通过调节RANKL-OPG平衡、抑制JNK磷酸化及c-Fos表达实现的。

Objective To explore the effect ofdrynaria total flavonoids on receptor activator of nuclear fac- tor kappa B ligand （RANKL）/Osteoprotegerin （OPG） expression and mitogen-activated protein kinase （MAPK） signal- ing pathways in ovariectomized rats. Methods Forty-five female rats were divided into sham operation group （15 rats） and operation group （30 rats）. The rats in operation group all underwent ovariectomy and had bone mineral densi- ty （BMD） detected 12 weeks later. After confirming the success of OP, the rats in operation group were divided into drynaria total fiavonoids group and model group, with 15 rats in each group. The drynaria total fiavonoids group was treated with drynaria total fiavonoids solution, while the model group and sham operation group received gastric perfu- sion of normal saline. The duration of treatment was 12 weeks. The level of BMD was checked. The expression of RANKL, OPG was detected by ELISA. The expression of RANKL and OPG mRNA of femur was detected by RT-PCR. Western blotting analysis was used to examine the phosphorylation of c-Jun Ntermingal kinase （JNK）, MAPK p38, extracellular-regulated kinase ERK, and c-FOS protei expression in bone tissues. Results （1） Compared with sham operation group, after 12-week treatment, the BMD in model group were significantly lower （P〈0.05）. The serum RANKL level increased significantly, serum OPG level decreased significantly, and the RANKL/OPG ratio in- creased significantly （all P〈0.05）. The RANKL mRNA expression significantly increased and OPG mRNA decreased significantly in bone tissues （all P〈0.05）. （2） Compared with model group, BMD was higher than that of rats in sham operation group and drynaria total flavonoids group （all P〈0.05）. The serum RANKL level decreased significantly, and the serum OPG level increased significantly, with the RANKL/OPG ratio decreased significantly （all P〈0.05）. The RANKL mRNA expression decreased significantly and OPG mRNA increased significantly in bone tissues （all P〈 0.05）. There were significant differences among sham operation group and drynaria total flavonoids group （P〈0.05）. （3） Compared with model group, bone tissue P-JNK and c-Fos expression in rats in sham operation group and drynaria total flavonoids group wcrc lower, but there were no significant differences among sham operation group and drynaria total flavonoids group （P〉0.05）. Conclusion Drynaria total flavonoids group can improve the BMD values in ovari- ectomizcd rats, which may be related to its function in regulating the balance of RANKL/OPG and inhibiting the expression of JNK phosphorylation and c-Fos.