A Study On The Analgesic Effect And Mechanism Of Platelet-rich Plasma On Monosodium Iodoacetate Induced Knee Osteoarthritis

Objective:To observe the effect of platelet rich plasma(PRP)injection into knee joint on the pain behavior of neuropathic pain induced by monosodium iodoacetate(MIA)in rats.As well as ionized calcium-binding adapter molecule-1(Iba-1),tumor necrosis factor-α(TNF-α)at the central level of the spinal cord,and activating transcription factor 3(ATF3)expression in dorsal root ganglion(DRG)to investigate the therapeutic effect of PRP on neuropathic pain caused by knee arthritis and its possible mechanism.Methods:Experiment 1: PRP preparation and preservationThe whole blood of 4 male rats was collected by abdominal aortic puncture before operation and injected into a catheter containing 3.8% sodium citrate.PRP was obtained from anticoagulated blood by centrifugation at 800 rpm for 15 min at 25 ° C.Platelets in whole blood and PRP were counted by automatic blood cell analyzer.PRP achieved platelet concentrations that were 3-4 times higher than baseline levels.PRP was activated by freezing at-80 ° C for 24 h and incubated at 37 ° C for 1 h.Following incubation,activated PRPs were centrifuged at 12000 g for 2 min to separate fragments.The supernatants were collected and stored at-80 ° C until use.Experiment 2: Induction and treatment of arthritis and behavioral measurementIn this study,40 male SD rats were randomly selected and injected 60μL 80 mg/m L monosodium iodoacetate(MIA)into the left knee joint to induce joint degeneration.Another20 SD rats were injected with the same volume of normal saline into the left knee joint as the control group.The mechanical pain threshold(PWT)and thermal pain threshold(PWL)were measured at 1 day before modeling and 1,3,7,14 days after modeling,and the pathological changes of knee joints were observed by HE staining to evaluate the establishment of MIA model.On the 14 th day after MIA injection(after successful modeling),54 male SD rats were randomly divided into 3 groups(n = 18 each): sham operation group(control group,injected with 60μl normal saline),MIA group(MIA group,injected with 60μl normal saline),and MIA group(MIA group,injected with 60μl normal saline).In group MIA,60μl PRP or the same volume of normal saline was injected into the unilateral knee joint of rats at 15.17 and 19 days after modeling.All injections were performed under isoflurane anesthesia.The mechanical pain threshold and thermal pain threshold were measured at 21,28 and 42 days after MIA injection.The rats were sacrificed at 28 and 42 days after MIA injection(10 rats per group at each time point),and the left knee joint was taken for histological examination.Experiment 3: Molecular biology experimentImmunohistochemical staining was used to observe ionized calcium-binding adapter molecule-1(Iba-1)and activating ATF3(activating)in the L3-L5 spinal cord and dorsal root ganglion of rats in the three groups at 28 days and 42 days after MIA injection The expression of TNFα in the spinal cord was detected by ELISA.The knee joint was observed under a light microscope and the Osteoarthritis Research Society International(OARSI)score was used to evaluate the knee joint degeneration.Results:1.Effects of MIA and PRP on Mechanical Allodynia and Thermal HyperalgesiaNo significant differences were observed in the PWT and PWL among all groups before the MIA injection.The MIA group showed prolonged allodynia and a significant decrease in PWL and PWT compared to the comparison group on days 1–14 after the MIA injection.Fourteen days after MIA injection,rats with OA and NP received PRP or NS treatment.The results showed a significant increase in both PWL and PWT at days 9 and 23 after PRP administration compared with the MIA group,indicating the beneficial effects of PRP in alleviating NP.2.PRP Treatment Decreased Expression for ATF3 and Iba-1The expression of Iba-1 in the spinal dorsal horn and ATF-3 protein in the DRG of rats in each group were observed using immunohistochemical staining on days 9 and 23 after administering PRP.The PRP group showed a significant decrease in Iba-1 expression on day23.Similarly,after 9 days of PRP administration,the expression of ATF3 started decreasing compared with that in the control group,and the effects lasted for 23 days after administration.3.PRP downregulates the expression of TNF-α in the spinal cordNo significant differences were observed in the expression of TNF-α in the L4-L6 spinal cord between the MIA+PRP and MIA+NS groups on day 28.The expression of TNF-α was downregulated in the PRP group compared to that in the MIA group on day 42.4.PRP Reduces the Progression of Cartilage DegenerationThe OARSI scores were used to quantify the histological features of OA.The scores for knees injected with PRP were significantly lower than those for control knees on day 28.However,no significant difference was observed between the groups on day 42.Conclusions:In summary,PRP may be a potential therapeutic agent for relieving chronic NP in KOA by inhibiting the activation of microglial cells and the release of microglia-derived TNF-α.This effect is likely to be mediated by a reduction in nerve injury.This might contribute to a better understanding of knee arthritis patients with NP pathophysiology and further development of PRP use for pain management.Further investigation is needed to clarify the detailed mechanisms for relieving pain and alleviating nerve injury in knee arthritis patients with NP.