Therapeutic Effects And Related Mechanism Of Slow-release Melatonin Targeted Delivery System In Osteoarthritis

Osteoarthritis(OA)is one of the most common forms of arthritis,usually involving the joints of the knees,hips and hands.OA is a complex disease that affects the whole joint.Its pathological manifestations include progressive synovial destruction,subchondral bone remodeling and synovial inflammation.At present,the pathogenesis of OA is not completely clear,in which the vicious circle of innate immune response and reactive oxygen species(ROS)can not be ignored in the course of OA.Melatonin,as an endogenous hormone,has been proved to be effective in the treatment of OA,but the related mechanism has not been fully explored.In addition,because of the non-vascular nature of cartilage,how to improve the utilization rate of therapeutic drugs in vivo is also a major difficulty in OA treatment.The development of targeted nano-drug delivery system is expected to be one of the means to solve this difficultyPurpose:To explore whether melatonin can treat OA by inhibiting the innate immune response of articular chondrocytes and scavenging reactive oxygen species,and on this basis,a nano drug delivery system with cartilage-targeting function was prepared to improve the utilization rate of melatonin in vivo.Method:Firstly,the primary chondrocytes were extracted from the knee joint of C57BL/6 mice,and the effects of different concentrations of melatonin on the proliferation and secretion of chondrocytes were tested.Then the OA cell model induced by interleukin-1β(IL-1β)was established.The oxidative stress level of OA cell model was detected by DCFH-DA probe,and the ROS scavenging ability of melatonin was detected by flow cytometry and fluorescence microscope.Western blotting and toluidine blue staining were used to identify the effect of melatonin on TLR2/4-MyD88-NFκB pathway in OA chondrocytes and RAW264.7 cells,and the effects of melatonin on catabolism and anabolism of cartilage matrix.Finally,C57BL/6 mice were treated with anterior cruciate ligament transection(ACLT)to establish animal OA model to verify the therapeutic effect of intra-knee injection of melatonin on animal OA model.On this basis,a nano-drug delivery system(MT@PLGA-COLBP)targeting cartilage matrix was prepared and characterized by poly(lactic-co-glycolicacid)(PLGA)and collagen targeting peptide COLBP.Then the targeting ability of MT@PLGA-COLBP was verified by the endocytosis of nano drug delivery system,the infiltration of knee joint in vitro and the accumulation of knee joint in vivo.Finally,the therapeutic effect of nanodrug delivery system in animal OA model was verified by histology and imaging examination.Results:Melatonin at a concentration of 100 μM is safe for chondrocytes.Melatonin can effectively eliminate ROS produced by OA cell model,and this effect is receptor-dependent.Melatonin can significantly inhibit the activated TLR2/4-MyD88-NFκB pathway in OA chondrocytes and RAW264.7 cells induced by LPS.Melatonin can also significantly reduce matrix catabolism and increase matrix anabolism in OA chondrocytes.Knee joint injection of melatonin can effectively delay the development of OA animal.In addition,nano-drug delivery system has good ability of endocytosis and cartilage targeting,and accumulated melatonin in the knee joint cavity of OA mice for 14 days.The establishment of nano drug delivery system can reduce the frequency of knee joint injection and has a good therapeutic effect on OA animal.ConclusionsThis study confirmed the new mechanism of melatonin in the treatment of OA:inhibiting the activation of innate immune response in the course of OA and scavenging reactive oxygen species at the same time.The nano drug delivery system prepared in this study can improve the utilization rate of melatonin and provide a new method for O A therapy.