Icariin regulates acidic microenvironment to alleviate pain caused by postmenopausal osteoporosis in the elderly
Xue Chunyang, Wang Xiuhui
Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
Abstract: BACKGROUND: Previous studies have demonstrated that icariin has important roles in promoting bone formation and inhibiting bone resorption, but its effects on osteoporosis-mediated bone pain have not been reported.
OBJECTIVE: To investigate the possible mechanism of icariin alleviating bone pain in postmenopausal senile osteoporosis.
METHODS: (1) Animal experiment: 200 C57BL/6 mice were randomly divided into 4 groups: sham group (n=50), model group (n=50), icariin treatment group (n=50), and carbonic anhydrase II inhibitor (Brinzolamide) treatment group (n=50). Ovariectomy was performed on C57BL/6 mice to establish a postmenopausal osteoporosis model in all groups except the sham group. The icariin group was given icariin on the second day after modeling, and pain behavior tests (Von Frey, Hot Plate, and Tail Flick tests) were performed every 2 weeks for 20 weeks. After sampling, bone mass was detected by microCT, osteoclast activity was detected by hematoxylin-eosin and tartrate-resistant acid phosphatase staining, and neuronal morphology and related ion channel expression were detected by tissue immunofluorescence staining. (2) Cell experiment: Osteoclast precursor cells derived from mouse bone marrow were extracted and induced to differentiate into osteoclasts using the RANKL/M-CSF system in vitro and supplemented with icariin of different concentrations (1 and 10 μmol/L). Tartrate-resistant acid phosphatase staining was used to detect osteoclast differentiation, ghost pen cyclic peptide staining was used to detect osteoclast actin ring, bone plate absorption assay was used to detect osteoclast osteophagy function, pH value of the system was detected by pH meter, and expression of osteoclast differentiation-related proteins was detected by western blot. In addition, mouse dorsal root ganglion-derived nerve cells were extracted and treated with icariin. Cell counting kit-8 was used to detect neuronal activity and CGRP staining was used to detect neuronal morphology.
RESULTS AND CONCLUSION: Compared with the model group, the icariin treatment group had higher bone mineral density, fewer tartrate-resistant acid phosphatase-positive osteoclasts in bone tissue, decreased neuronal activity, and decreased neuronal transient receptor potential vanilloid subtype 1 channel and carbonic anhydrase II expression. Behavioral results showed that the icariin treatment group was less sensitive to pain than the model group. Icariin inhibited osteoclast differentiation and bone phagocytosis in vitro. Icariin enhanced the activity of dorsal root ganglion neurons and inhibited the expression of calcitonin gene-related peptide and transient receptor potential vanilloid subtype 1 channels in dorsal root ganglia in a relatively non-toxic pH range. To conclude, icariin alleviates bone pain caused by postmenopausal osteoporosis by regulating the acidic microenvironment through its effect on osteoclasts.
Key words: icariin, osteoporosis, pain, acidic microenvironment, osteoclast differentiation, neuron