Comparison of bioactive substances in novel-developed freeze-dried platelet-rich plasma (PRP) and activated normal PRP, and investigation of bioactive substance levels after long-term storage.

Introduction: In recent years, biotherapy in orthopedics has become widespread, and platelet-rich plasma (PRP) has been readily used to treat sports injuries and osteoarthritis. Production of freeze-dried PRP (PRP-FD) results in PRP that is in powder form, allowing it to be stored for long periods at room temperature. Using this technology, we have developed Valuable Platelet-Derived Factor Concentrate Freeze Dry (VFD). However, whether VFD contains sufficient levels of bioactive substances (BS) remains unknown and retains the same levels of BS during long-term storage. In this study, we examined whether VFD contains sufficient amounts of BS and whether they retain these BS levels during long-term storage. Methods: Peripheral blood was collected from 10 healthy men (mean ± SD: 46.5 ± 15 years old) and various BS, including transforming growth factor ß (TGF-ß), basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), tissue inhibitors of metalloproteinases-1 (TIMP-1), interleukin-1 receptor antagonist (IL-1ra), matrix metallopeptidase-9 (MMP-9), and interleukin-6 (IL-6), were compared between VFD and normal PRP samples, including both leukocyte-rich PRP (LR-PRP) and leukocyte-poor PRP (LP-PRP). VFD was prepared using two rounds of centrifugation. LP-PRP and LR-PRP were activated by freezing and thawing before measurement. To evaluate the effects of long-term storage, the BS of VFD purified from five professional football players was compared between baseline and 1 year after storage. Results: In terms of the growth factors, the TGF-ß and EGF levels were higher in LR-PRP than in VFD and LP-PRP (p < 0.05), while the bFGF levels were higher in VFD than in the LR-PRP and LP-PRP groups (p < 0.01). In terms of anti-inflammatory cytokines, the TIMP-1 level was lower in VFD than that in the other groups (p < 0.01), whereas the IL-1ra levels were higher in VFD than those in LP-PRP (p < 0.05) and lower than those in LR-PRP (p < 0.01). In terms of inflammatory enzymes and cytokines, the IL-1ra level was higher in VFD than that in LP-PRP (p < 0.05) and lower than that in LR-PRP (p < 0.01), whereas the IL-6 levels did not differ among the groups. Furthermore, the TGF-ß, bFGF, TIMP-1, and IL-1ra levels were 5.61 â†’ 3.38 (x103 pg/µL), 61.0 â†’ 63.0 (pg/µL), 3.4 â†’ 2.7 (x105 pg/µL), and 14.9 â†’ 14.5 (x103 pg/µL) at baseline and 1 year later, respectively. No significant differences in the BS levels were observed between baseline and 1 year after storage. Conclusions: The VFD samples prepared in this study exhibited higher levels of anti-inflammatory cytokines than LP-PRP and contained growth factor levels similar to LP-PRP and LR-PRP. In addition, the BS levels in VFD samples were maintained after one year of storage. These results suggest that VFD can be prepared and stored and may serve as a novel treatment strategy for sports injuries in high-risk groups, such as athletes.

Safety and feasibility of locoregional platelet-rich plasma injection for iliopsoas impingement after total hip arthroplasty: A phase 1 prospective observational study.

Introduction: Although numerous studies have reported outcomes with various conservative approaches for the iliopsoas impingement after total hip arthroplasty (THA), reports on the use of locoregional autologous platelet rich plasma (PRP) injections for the iliopsoas impingement after THA are lacking. This phase 1 study therefore aimed to investigate the safety and feasibility of locoregional PRP injection for iliopsoas impingement after THA. Materials and methods: Patients diagnosed with iliopsoas impingement after THA who met the criteria for participation (symptoms persisting for more than 3 months, aged 20 years or older, and unable to receive non-steroidal analgesic or anti-inflammatory drugs) were eligible to participate in this clinical study. The primary endpoint was observed adverse events including procedure-related pain, and the secondary endpoints included pain and functionality of the hip joint, that were assessed using the Western Ontario and McMaster Universities Arthritis Index, Japanese Hip Disease Evaluation Questionnaire, and Forgotten Joint Score-12. Results: Three patients were screened for eligibility, and 3 patients were finally included in this study. Two participants (patients 1 and 2; aged 66 and 65 years, respectively) were female. The third participant (patient 3; age 73 years) was male. All patients experienced adverse events; however, none were found to be serious. None of the patients experienced any infections, or intra- or post-operative symptoms of nerve damage, or subcutaneous haemorrhage owing to the administration of locoregional PRP. Although patient 2 showed almost complete resolution of the symptom, patient 1 and 3 demonstrated persistent groin pain after the injection. Conclusion: We demonstrated the results of preliminary phase 1 prospective observational clinical study that administration of locoregional PRP injections for iliopsoas impingement following THA is both, safe and feasible.

Study protocol for a randomised placebo controlled trial of platelet-rich plasma injection to prevent post-traumatic knee osteoarthritis after anterior cruciate ligament reconstruction.

INTRODUCTION: The elevated cytokine levels in patients suffering from anterior cruciate ligament (ACL) rupture may lead to acute post-traumatic arthritis (APTA) and post-traumatic osteoarthritis (PTOA). Due to its chondrogenerative and anti-inflammatory effect, platelet-rich plasma (PRP) therapy is expected to show a positive outcome in APTA and PTOA. The proposed trial aims to quantitatively measure the efficacy of PRP injection in arresting post-traumatic cartilage degeneration among patients after ACL reconstruction. METHODS AND ANALYSIS: This will be a single-blind, randomised, prospective, controlled clinical trial designed following the Consolidated Standards of Reporting Trials guidelines. After ACL reconstruction, 80 patients will be randomised to receive either leucocyte-poor PRP injection after joint aspiration or a placebo control group receiving only joint aspiration. Participants (age 20-49 years) will be those who have undergone ACL reconstruction within the past 2 weeks with a body mass index<35 and Kellgren Lawrence osteoarthritis grade<2. The primary outcome will include MRI-T2 values of knee cartilage at 6 months. The secondary outcomes will include pain assessment by Visual Analogue Scale, Knee injury and Osteoarthritis Outcome Score, blood and urine test, physical findings, measurements for muscle strength and joint stability. ETHICS AND DISSEMINATION: The study was approved by The Independent Ethics Committee for Clinical Trials of the Japanese Association for the Promotion of State-of-the-Art Medicine. Results of the trial and each of the outcomes will be shared via conferences and publication in a peer-reviewed journal. TRIAL REGISTRATION NUMBER: jRCTb030200391.

Intra-Articular Injections of the Adipose-Derived Mesenchymal Stem Cells Suppress Progression of a Mouse Traumatic Knee Osteoarthritis Model.

OBJECTIVE: This study aimed to determine whether the intra-articular injection of human adipose-derived mesenchymal stem cells (ADSCs) protects against the progression of murine post-traumatic osteoarthritis. DESIGN: ADSCs were isolated from human abdomen or buttock adipose tissues. In in vitro study, ADSCs conditioned medium was added to human chondrocytes pre-treated with interleukin-1ß (IL-1ß), and resultant gene expression of target inflammatory genes was measured by real-time quantitative polymerase chain reaction. A mouse model of knee osteoarthritis was generated by unilaterally transecting the medial meniscus in the right hind limb of 20 female C57BL/6 mice. Mice were randomly assigned to 2 treatment groups that received 6 µl intra-articular injections of either phosphate-buffered saline (control) or 2 × 104 cells/µl of ADSCs 14, 28, and 42 days post-surgery. Mice were euthanized 84 days post-surgery and histological and micro-computed tomography evaluation of knee joints were analyzed. Hind limb weight-bearing distribution was measured pre-surgery and 28 and 84 days post-surgery. RESULTS: Conditioned medium from cultured human adipose-derived mesenchymal stem cells suppressed the expression of target inflammatory genes in chondrocytes pre-treated with IL-1ß, suggesting anti-inflammatory properties (P < 0.01). Histological analyses indicated that the progression of destabilization of medial meniscus-induced knee osteoarthritis was suppressed by the administration of ADSCs compared with control group at medial femorotibial joint in vivo. This protective effect was related to a reduction in articular cartilage loss. CONCLUSION: The intra-articular injection of ADSCs suppressed articular cartilage loss in a mouse model of knee osteoarthritis, possible through anti-inflammatory mechanisms.

Predictors of Effectiveness of Platelet-Rich Plasma Therapy for Knee Osteoarthritis: A Retrospective Cohort Study.

There has recently been growing interest worldwide in biological therapies such as platelet-rich plasma injection for the treatment of knee osteoarthritis. However, predicting the effectiveness of platelet-rich plasma therapy remains uncertain. Therefore, this retrospective cohort study was performed to assess a range of predictors for the effectiveness of platelet-rich plasma therapy in treating knee osteoarthritis. The study included 517 consecutive patients who underwent three injections of leucocyte-poor platelet-rich plasma therapy from 2016 to 2019 at a single institution. The treatment outcomes, including patient-oriented outcomes (visual analogue scale score and Knee Injury and Osteoarthritis Outcome Score), were analyzed and compared according to the severity of knee osteoarthritis based on Kellgren-Lawrence (KL) grading using standing plain radiographs. Fisher's exact test, univariate regression, and multivariate regression were used for data analysis. Patient-oriented outcomes were significantly improved 6 and 12 months after platelet-rich plasma therapy. The overall responder rate in patients who met the Outcome Measures in Rheumatology (OMERACT)-Osteoarthritis Research Society International (OARSI) responder criteria was 62.1%. The responder rate was significantly lower in patients with severe knee osteoarthritis (KL4, 50.9%) than in those with mild (KL2, 75.2%) and moderate (KL3, 66.5%) knee osteoarthritis. The multivariate logistic regression analysis revealed that deterioration of the knee osteoarthritis grade (increased KL grade) was a significant predictor of a worse clinical outcome (odds ratio, 0.58; 95% confidence interval, 0.45-0.75; p < 0.001). The relative risk for non-responders in severe (KL4) KOA was 2.1 (95% CI, 1.5-3.0) at 6 months and 2.3 (1.6-3.2) at 12 months compared with mild-to-moderate (KL2-3) KOA. The efficacy of platelet-rich plasma therapy was not affected by age, sex, body weight, or platelet count. This study revealed that the effectiveness of platelet-rich plasma therapy for the treatment of knee osteoarthritis is approximately 60% and that the effectiveness depends on the severity of knee osteoarthritis. This observation is useful not only for physicians but also for patients with knee osteoarthritis.

The immature platelet fraction affects the efficacy of platelet rich plasma therapy for knee osteoarthritis.

INTRODUCTION: Platelet-rich plasma (PRP) therapy is used to treat pathological conditions such as degenerative inflammatory diseases including osteoarthritis (OA) by enhancing tissue repair and promoting anti-inflammatory effects. Although PRP therapy for patients with knee OA improved pain and functional scores, the association of clinical outcomes and quality of PRP including cell composition and concentration is unclear. METHODS: Therefore, this study analyzed blood cell counts, including the immature platelet fraction (IPF), in peripheral blood and PRP of 144 patients with knee OA who underwent PRP therapy. The mean leukocyte and platelet concentrations in whole blood and PRP were analyzed using an XN-1000 automated hematology analyzer. Visual analogue scale (VAS) scores and knee injury and osteoarthritis outcome scores (KOOS) before and 1 month after a single PRP injection were also determined. RESULTS: Higher platelet and lower leukocyte concentration rates were observed in PRP compared with whole blood. The platelet concentration in whole blood was negatively correlated with VAS improvement. The percentage of IPF (IPF%) in whole blood was positively correlated with VAS improvement and KOOS (pain) improvement, whereas the IPF% in PRP tended to correlate with VAS improvement. Furthermore, multivariate logistic regression demonstrated the high IPF% in whole blood was significantly associated with VAS improvement. The low percentage of neutrophil (neutrophil%) in PRP was significantly associated with the VAS improvement and KOOS (ADL) improvement. CONCLUSIONS: Therefore, PRP efficacy for OA might depend on the patient's biological status.

Quality comparison between two different types of platelet-rich plasma for knee osteoarthritis.

INTRODUCTION: Knee osteoarthritis (KOA), the most common form of osteoarthritis (OA) is a considerable health concern worldwide. Platelet-rich plasma (PRP) is a common therapeutic option for KOA. Different types of PRPs have varying efficacies. However, a comparative analysis of the qualities of these PRPs is lacking. METHODS: Two types of PRPs, including autologous protein solution (APS), and leukocyte-poor PRP (LP-PRP) along with whole blood (WB) and platelet-poor plasma (PPP) were characterized for platelet content, leukocyte content, and composition in 10 healthy volunteers (HV) (the controlled laboratory study) and 16 KOA patients (a retrospective observational study). Additionally, the levels of the platelet-derived growth factor (PDGF)-BB, and different cytokines were estimated in HV. RESULTS: In HV, the concentrations of platelets and leukocytes, levels of different cytokines, including interleukin 1 receptor antagonist (IL-1Ra), soluble TNF receptor type II (sTNF-RII), and IL-1ß, and the ratio of IL-1Ra/IL-1ß were significantly higher in APS, whereas the PDGF-BB was higher in LP-PRP than APS. In KOA patients, a higher concentration of platelets was observed in LP-PRP, and a higher concentration of leukocytes was observed in APS than LP-PRP. Following the PAW classification system, LP-PRP was classified as P2-B type in HV (51.3 × 104/µl) and KOA (53.4 × 104/µl), whereas APS was classified as P3-A type in HV (110.1 × 104/µl) and P2-A type in KOA (29.0 × 104/µl). In a retrospective observational study, the KOA patients who underwent APS injection had a higher incidence of arthralgia, and this arthralgia lasted for a longer time than LP-PRP injection in the same individual. DISCUSSION: The quality of the two PRPs differed distinctively depending on their preparation methods, which might affect their clinical efficacies and adverse events. Therefore, the characterization of these parameters should be prioritized while choosing PRP.

Platelet-rich plasma promotes recruitment of macrophages in the process of tendon healing.

INTRODUCTION: Researchers have investigated the use of platelet-rich plasma (PRP) therapy. However, the mechanisms through which PRP affects tissue repair remain unclear. We hypothesize that PRP promotes tissue repair through not only via direct manner on the local cells but also via indirect manner that encourage the recruitment of reparative cells such as macrophages (MPs), and it depends on the quality of PRP including the concentration of leukocytes. The aim of this study is to elucidate the actions of the MPs in the mechanisms of PRP on tissue repair processes. METHODS: Leukocyte-rich (LR) PRP and leukocyte-poor (LP) PRP were prepared from 12-week-old C57BL6 mice. Full-thickness defects were created in central third of patellar tendons of 12-week-old C57BL/6 mice for histologic analysis (n = 36) and 12-week-old B6.129P-Cx3cr1tm1Litt/J mice for flow cytometry analysis (n = 108). B6.129P-Cx3cr1tm1Litt/J mouse is GFP-positive only in the MP-linage cells thus MPs recruited to the repair tissue can be distinguished whether it had originated from administrated PRP or recruited from host mouse. Mice were treated either with LR-PRP, LP-PRP, or without PRP (control group). Histological analyses were performed to evaluate the tendon healing using Bonar score as semi-quantitative histological scoring system. Flow cytometric analyses were performed to count the number of GFP-positive cells around repaired patellar tendon. In addition, the ratio of pro-inflammatory MPs (M1)/anti-inflammatory MPs (M2) were analyzed in those GFP-positive cells. The statistical analysis was performed using GraphPad Prism ver6. P values < 0.05 were considered statistically significant. RESULTS: In LR-PRP and LP-PRP groups, all variables in Bonar score such as cell morphology, cellularity, vascularity, and collagen arrangement were significantly improved in comparison with control group, indicating that both PRPs promote tendon hearing. LP-PRP promoted the tendon healing significantly faster than that of LR-PRP on postoperative day 28 (P < 0.001). LR-PRP enhanced angiogenesis (vascularity: P < 0.001), while LP-PRP improved the collagen arrangement on postoperative day 28 (collagen arrangement: P < 0.01). In other variables such as cell morphology and cellularity score, there were no significant differences between LR-PRP and LP-PRP groups in any time points. Flow cytometric findings showed that recruitment of GFP-positive MPs in the LR and LP-PRP groups were significantly increased from postoperative day 4 compared with control group without PRP treatment (P < 0.001). The majority of GFP-positive MPs were M1 at the initiation of tendon healing phase, and M2 were gradually increased from postoperative day 4. The number of M1 was significantly high both in the LP- and LR-PRP groups (day 4 and 7, p < 0.001), but the number of M2 was high only in the LP-PRP group (day 7 and 14, P < 0.05) when it compared with control group. The M1/M2 ratio on postoperative day 7 was significantly lower in the LP-PRP group than those in the control group (P < 0.05). CONCLUSIONS: This study demonstrated that PRP enhanced the tendon healing and promoted the recruitment of MPs to the injured tissue. The subtypes of MPs were different depends on the types of PRPs, suggesting that leukocytes in PRP influence the effect of PRP therapy.

Bilateral non-osteochondroma-related proximal tibiofibular synostosis.

We report the case of a 67-year-old male with bilateral proximal tibiofibular synostosis, presenting with unilateral symptoms. The patient complained of pain around the left fibular head, which was attributed to incomplete bone bridging between the proximal tibia and fibula; he underwent proximal fibular head resection, which alleviated the pain and improved knee mobility. Eleven months later, the patient continued to be pain-free and did not experience any adverse effects. An examination of this case and a review of similar cases revealed that participation in sport activities such as long-distance running may be one of the causes of proximal tibiofibular synostosis. In this report, we have also reconsidered the classification of proximal tibiofibular synostosis and provided information for a better understanding of this unusual condition.