Zhe Cai1, Yu Feng2, Chentian Li1, Kedi Yang1, Tianhao Sun1, Lei Xu1, Yan Chen3, Chun-Hoi Yan1, William Weijia Lu4, Kwong-Yuen Chiu5. 1. Department of Orthopaedics and Traumatology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong. 2. Department of Traumatology, General Hospital of Ningxia Medical University, Yinchuan, Ningxia Hui Autonomous Region, China. 3. Department of Bone and Joint Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi, China. 4. Department of Orthopaedics and Traumatology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong; Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, China. Electronic address: wwlu@hku.hk. 5. Department of Orthopaedics and Traumatology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong. Electronic address: pkychiu@hku.hk.
Abstract
OBJECTIVE: To investigate efficacy of Chinese medicine magnoflorine combined with hyaluronic acid (HA)-gel in promoting subchondral bone (SCB) regeneration and attenuating cartilage degeneration in early osteoarthritis (OA). METHODS: MC3T3-E1 under magnoflorine treatment was assayed by XTT to determine cell viability. Cell proliferation was reflected through cell cycle. Osteoblast mineralization was stained by Alizarin Red. Standardized bone canal of 1 mm in diameter and 4 mm in depth was made on tibial medial plateau of 4-month-old Dunkin-Hartley spontaneous knee OA guinea pigs. Guinea pigs (n = 5/group) were treated once intra-bone canal injection of 2 μl HA-gel, 2 μl HA-gel+50 ng magnoflorine and null (Defect) respectively, except sham group. The left hind limbs were harvested for μCT scan and histopathological staining 2-month post-surgery. RESULTS: 25 μg/ml magnoflorine treatment significantly increased cell viability, S-phase and mineralization of MC3T3-E1 cells. In vivo, HA-gel + magnoflorine treatment significantly altered SCB microstructure; changes included increase in bone volume fraction (BV/TV), trabecular number (Tb.N), connectivity density (Conn.Dn), and decrease in degree of anisotropy (DA), which implied trabecular bone regeneration. Treatment also resulted in a decrease in modified Mankin's scores, and an increase in volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and fractal dimension (FD, roughness indicator of osteochondral conjunction), compared to Defect and HA groups. Furthermore, FD was positively associated with volume ratio of HC/CC and negatively associated with modified Mankin's scores. Finally, histological results showed that due to a faster regeneration of SCB with the HA-gel + magnoflorine treatment, the reduction of cartilage matrix and the decreased expression of chondrogenic signals were attenuated. CONCLUSION: Our study elucidated the potential benefits of HA-gel + magnoflorine in promoting SCB regeneration and revealed a protective effect of stimulating recovery of the SCB integrity on attenuating cartilage degradation to prevent OA progression.
OBJECTIVE: To investigate efficacy of Chinese medicine magnoflorine combined with hyaluronic acid (HA)-gel in promoting subchondral bone (SCB) regeneration and attenuating cartilage degeneration in early osteoarthritis (OA). METHODS: MC3T3-E1 under magnoflorine treatment was assayed by XTT to determine cell viability. Cell proliferation was reflected through cell cycle. Osteoblast mineralization was stained by Alizarin Red. Standardized bone canal of 1 mm in diameter and 4 mm in depth was made on tibial medial plateau of 4-month-old Dunkin-Hartley spontaneous knee OA guinea pigs. Guinea pigs (n = 5/group) were treated once intra-bone canal injection of 2 μl HA-gel, 2 μl HA-gel+50 ng magnoflorine and null (Defect) respectively, except sham group. The left hind limbs were harvested for μCT scan and histopathological staining 2-month post-surgery. RESULTS: 25 μg/ml magnoflorine treatment significantly increased cell viability, S-phase and mineralization of MC3T3-E1 cells. In vivo, HA-gel + magnoflorine treatment significantly altered SCB microstructure; changes included increase in bone volume fraction (BV/TV), trabecular number (Tb.N), connectivity density (Conn.Dn), and decrease in degree of anisotropy (DA), which implied trabecular bone regeneration. Treatment also resulted in a decrease in modified Mankin's scores, and an increase in volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and fractal dimension (FD, roughness indicator of osteochondral conjunction), compared to Defect and HA groups. Furthermore, FD was positively associated with volume ratio of HC/CC and negatively associated with modified Mankin's scores. Finally, histological results showed that due to a faster regeneration of SCB with the HA-gel + magnoflorine treatment, the reduction of cartilage matrix and the decreased expression of chondrogenic signals were attenuated. CONCLUSION: Our study elucidated the potential benefits of HA-gel + magnoflorine in promoting SCB regeneration and revealed a protective effect of stimulating recovery of the SCB integrity on attenuating cartilage degradation to prevent OA progression.