Long-Term Structural Changes in the Osteochondral Unit in Patients with Osteoarthritis Undergoing Corrective Osteotomy with Platelet-Rich Plasma or Stromal Vascular Fraction Post-Treatment.

This pilot study examined the long-term structural changes in the osteochondral unit of 20 patients with knee osteoarthritis (KOA) who underwent high tibial osteotomy (HTO) and received post-treatment with either platelet-rich plasma (PRP) or stromal vascular fraction (SVF). Ten patients were injected with autologous PRP (PRP subgroup), while another ten patients received autologous SVF (SVF subgroup) six weeks after surgery and were monitored for 18 months. Histological samples of bone and cartilage (2 mm in diameter and 2 cm long) were taken from tibial and femoral sites during surgery and 18-month post-HTO, and morphometric analyses were conducted using Mega-Morf12 software. Both post-treatment resulted in an increase in articular cartilage height at both sites (p < 0.001 in the tibia and femur), indicating positive outcomes. Significant improvements in subchondral and trabecular bone architecture were also observed, with SVF injection showing higher reparative capacity in terms of bone volume (p < 0.001 for the tibia and p = 0.004 for the femur), subchondral bone height (p < 0.001 for the tibia and p = 0.014 for the femur), trabecular bone volume (p < 0.001 for the femur), and intertrabecular space (p = 0.009 for the tibia and p = 0.007 for the femur). This pilot study, for the first time, demonstrates that HTO surgery combined with PRP and SVF post-treatments can lead to significant enhancements in knee articular cartilage and bone architecture in KOA patients, with SVF showing higher regenerative potential. These findings may contribute to improving treatment strategies for better clinical outcomes in HTO therapy for patients with KOA.

Metabolic Dysregulation and Its Role in Postoperative Pain among Knee Osteoarthritis Patients.

Knee osteoarthritis (KOA) is characterized by low-grade inflammation, loss of articular cartilage, subchondral bone remodeling, synovitis, osteophyte formation, and pain. Strong, continuous pain may indicate the need for joint replacement in patients with end-stage OA, although postoperative pain (POP) of at least a two-month duration persists in 10-40% of patients with OA. STUDY PURPOSE: The inflammation observed in joint tissues is linked to pain caused by the production of proinflammatory cytokines. Since the biosynthesis of cytokines requires energy, their production is supported by extensive metabolic conversions of carbohydrates and fatty acids, which could lead to a disruption in cellular homeostasis. Therefore, this study aimed to investigate the association between POP development and disturbances in energy metabolic conversions, focusing on carbohydrate and fatty acid metabolism. METHODS: Peripheral blood samples were collected from 26 healthy subjects and 50 patients with end-stage OA before joint replacement surgery. All implants were validated by orthopedic surgeons, and patients with OA demonstrated no inherent abnormalities to cause pain from other reasons than OA disease, such as malalignment, aseptic loosening, or excessive bleeding. Pain levels were assessed before surgery using the visual analogue scale (VAS) and neuropathic pain questionnaires, DN4 and PainDETECT. Functional activity was evaluated using the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC). Three and six months after surgery, pain indices according to a VAS of 30 mm or higher were considered. Total RNA isolated from whole blood was analyzed using quantitative real-time RT-PCR (qRT-PCR) for the expression of genes related to carbohydrate and fatty acid metabolism. Protein levels of the examined genes were measured using an ELISA in the peripheral blood mononuclear cells (PBMCs). We used qRT-PCR because it is the most sensitive and reliable method for gene expression analysis, while an ELISA was used to confirm our qRT-PCR results. KEY FINDINGS: Among the study cohort, 17 patients who reported POP demonstrated significantly higher (p < 0.05) expressions of the genes PKM2, LDH, SDH, UCP2, CPT1A, and ACLY compared to pain-free patients with KOA. Receiver-operating characteristic (ROC) curve analyses confirmed the association between these gene expressions and pain development post-arthroplasty. A principle component analysis identified the prognostic values of ACLY, CPT1A, AMPK, SDHB, Caspase 3, and IL-1ß gene expressions for POP development in the examined subjects. CONCLUSION: These findings suggest that the disturbances in energy metabolism, as observed in the PBMCs of patients with end-stage KOA before arthroplasty, may contribute to POP development. An understanding of these metabolic processes could provide insights into the pathogenesis of KOA. Additionally, our findings can be used in a clinical setting to predict POP development in end-stage patients with KOA before arthroplasty.

Cathepsin S Upregulation Measured in the Peripheral Blood Mononuclear Cells Prior to Surgery Points to Postoperative Pain Development in Patients with Hip Osteoarthritis.

Disability caused by hip osteoarthritis has increased due to population aging, obesity, and lifestyle behaviors. Joint failure after conservative therapies results in total hip replacement, which is considered to be one of the most successful interventions. However, some patients experience long-term postoperative pain. Presently, there are no reliable clinical biomarkers for the prognosis of postoperative pain prior to surgery. Molecular biomarkers can be considered as intrinsic indicators of pathological processes and as links between clinical status and disease pathology, while recent innovative and sensitive approaches such as RT-PCR have extended the prognostic value of clinical traits. In light of this, we examined the importance of cathepsin S and proinflammatory cytokine gene expression in peripheral blood in addition to the clinical traits of patients with end-stage hip osteoarthritis (HOA) to predict postoperative pain development prior to surgery. This study included 31 patients with radiographic Kellgren and Lawrence grade III-IV HOA who underwent total hip arthroplasty (THA) and 26 healthy volunteers. Before surgery, a visual analog scale (VAS), DN4, PainDETECT, and the Western Ontario and McMaster Universities osteoarthritis index scores were used for pain and function assessment. Three and six months post-surgery, VAS pain scores of 30 mm and higher were reported. The intracellular protein levels of cathepsin S were measured using ELISA. The expression of the cathepsin S, tumor necrosis factor α, interleukin-1ß, and cyclooxygenase-2 genes in peripheral blood mononuclear cells (PBMCs) was assessed using quantitative real-time RT-PCR. Pain persisted in 12 (38.7%) patients after THA. Patients who developed postoperative pain demonstrated significantly higher cathepsin S gene expression in the PBMCs and higher rates of neuropathic pain based on the DN4 testing compared to the other HOA subjects that were examined. No significant differences in proinflammatory cytokine gene expression were noted in either patient cohort prior to THA. The development of postoperative pain in patients with hip osteoarthritis might be associated with disturbances in pain perception, while increased expression of cathepsin S in the peripheral blood prior to surgery may serve as its prognostic biomarker and could be used in clinical settings to improve medical service for patients with end-stage hip OA.

Differences in Synovial Cytokine Profile Associated with Long-Term Clinical Outcomes in Patients with Knee Osteoarthritis Undergoing Corrective Osteotomy with Platelet-Rich Plasma or Stromal Vascular Fraction Post-Treatments.

Functional outcomes and synovial fluid (SF) cytokine concentrations in response to platelet-rich plasma (PRP) or stromal vascular fraction (SVF) post-treatments following open wedge high tibial osteotomy (HTO) in 20 patients with knee osteoarthritis (OA) were examined. Six weeks after surgery, the knees of 10 patients were injected with autologous PRP (PRP subgroup), while another 10 patients were injected with autologous SVF (SVF subgroup) and monitored for 1.5 years. Pain assessment (VAS score) and functional activity (KOOS, KSS, Outerbridge, and Koshino scores) were applied. PRP subgroup performed better compared with the SVF subgroup according to KOOS, KSS, and VAS scores, while the SVF subgroup demonstrated better results according to Outerbridge and Koshino testing and produced more pronounced cartilage regeneration in the medial condyle and slowed down cartilage destruction in its lateral counterpart. SF was collected before and one week after PRP or SVF injections and tested for concentrations of 41 cytokines (Multiplex Assay). In the PRP subgroup, a significant decrease in IL-6 and CXCL10 synovial concentrations was accompanied by an increase in IL-15, sCD40L, and PDGF-AB/BB amounts. The SVF subgroup demonstrated a significant decrease in synovial TNFα, FLT-3L, MIP-1ß, RANTES, and VEGF concentrations while SF concentrations of MCP-1 and FGF2 increased. Both post-treatments have a potential for increased tissue regeneration, presumably due to the downregulation of inflammation and augmentation of synovial growth factor concentrations.

Downregulation of Tumour Necrosis Factor α Gene Expression in Peripheral Blood Mononuclear Cells Cultured in the Presence of Tofacitinib Prior to Therapy Is Associated with Clinical Remission in Patients with Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by pain, synovial hyperplasia, mononuclear cell infiltration, bone erosion and joint destruction. Efficacy of personalized therapy in RA is associated with correct choice of therapeutic agent and a possibility to predict its effect prior to treatment. Our objective was to examine the association of baseline expression of metalloproteinase (MMP)-9 and cathepsin K, which are involved in cartilage and bone degradation, as well as proinflammatory cytokines tumour necrosis factor (TNF)α and interleukin (IL)-1ß in the peripheral blood mononuclear cells (PBMCs) obtained from patients with RA cultured with tofacitinib (TFCN) and remission achievement. We examined 12 tofacitinib-naïve patients with RA, with a median age of 51 years and disease duration of 37.6 months. After three months of TFCN therapy, six of these patients reached clinical remission criteria while others preserved high and moderate disease activity. PBMCs were tested prior to therapy followed by their isolation in Ficoll density gradient and cultured with 100 nM TFCN for 48 h. Gene expression analysis for MMP-9, cathepsin K, IL-1ß, and TNFα was performed with quantitative real-time RT-PCR using total RNA isolated from and cultured with TFCN PBMCs compared with untreated cells. Expression of all the examined genes was significantly upregulated in those cultured with TFCN PBMCs from patients who maintained high and moderate disease activity after TFCN therapy while TNFα gene expression was significantly downregulated in patients who gained remission compared with untreated counterparts. Downregulation of TNFα gene expression in PBMCs from TFCN-naïve patients with RA cultured with TFCN prior to therapy compared with untreated counterparts might serve a prognostic biomarker for remission attainment in response to tofacitinib therapy.

Putative Association between Low Baseline Gene Expression in the Peripheral Blood and Clinical Remission in Rheumatoid Arthritis Patients Treated with Tofacitinib.

We investigated the importance of the baseline expression of genes involved in energy generation, as prognostic biomarkers of the treatment response to tofacitinib in patients with rheumatoid arthritis (RA). Peripheral blood samples were obtained from 28 patients with RA who received 3 months of tofacitinib therapy from 26 healthy controls. Clinical response was evaluated based on the disease activity score, the erythrocyte sedimentation rate (DAS28-ESR), and the serum levels of ACPA, RF, CRP, and ESR. Clinical remission was assessed based on DAS28 score <2.6. Protein concentrations were measured using ELISA. Total RNA isolated from whole blood was used for gene expression analysis using quantitative RT-PCR. All patients were diagnosed with Steinbrocker's radiographic stage II-III at baseline, and most showed erosive arthritis with ACPA and RF positivity. Tofacitinib treatment significantly decreased the disease activity. Upon study completion, seven patients showed remission. Before and after TOFA therapy, a significantly higher expression of succinate dehydrogenase and pyruvate kinase genes was observed in all the examined patients compared to healthy subjects. However, the pre-therapy expression of these genes and corresponding proteins was significantly (p ≤ 0.05) lower in patients who showed remission than in other patients with RA. Moreover, we observed that, during follow-up, patients who developed remission showed an increasing trend in the expression of the examined genes, whereas the others showed some decreases in gene expression, although this was not statistically significant. We concluded that, compared with RA patients maintaining persistent moderate or high disease activity, those with clinical remission following tofacitinib treatment showed a significantly lower baseline expression of genes involved in energy generation.

Development of Postoperative Pain in Patients with End-Stage Knee Osteoarthritis Is Associated with Upregulation of Genes Related to Extracellular Matrix Degradation, Inflammation, and Apoptosis Measured in the Peripheral Blood before Knee Surgery.

Osteoarthritis (OA) pain implies an indication for joint replacement in patients with end-stage OA. However, chronic postoperative pain is observed in 10-40% of patients with OA. Here, we identified genes whose expression in the peripheral blood before surgery could denote the risk of postoperative pain development. We examined the peripheral blood of 26 healthy subjects and 50 patients with end-stage OA prior to joint replacement surgery. Pain was evaluated before surgery using the visual analog scale (VAS) index and neuropathic pain questionnaires, Douleur Neuropathique 4 Questions (DN4) and PainDETECT questionnaires. Functional activity was assessed using the Western Ontario and McMaster Universities osteoarthritis index (WOMAC). Three and six months after surgery, pain indices according to VAS of 30% and higher were considered. Metalloproteinase (MMP)-9 and tissue inhibitor of metalloproteinase (TIMP)1 protein levels were measured using ELISA in the peripheral blood mononuclear cells (PBMCs). Total RNA isolated from whole blood was analysed using quantitative real-time RT-PCR for caspase-3, MMP-9, TIMP1, cathepsins K and S, tumour necrosis factor (TNF)α, interleukin (IL)-1ß, and cyclooxygenase (COX)-2 gene expression. Seventeen patients reported post-surgical pain. Expression of cathepsins K and S, caspase-3, TIMP1, IL-1ß, and TNFα genes before surgery was significantly higher in these patients compared to pain-free patients with OA. Receiver-operating characteristic (ROC) curve analyses confirmed significant associations between these gene expressions and the likelihood of pain development after arthroplasty. High baseline expression of genes associated with extracellular matrix destruction (cathepsins S and K, TIMP1), inflammation (IL-1ß, TNFα), and apoptosis (caspase-3) measured in the peripheral blood of patients with end-stage OA before knee arthroplasty might serve as an important biomarker of postoperative pain development.

Insulin Resistance in Osteoarthritis: Similar Mechanisms to Type 2 Diabetes Mellitus.

Osteoarthritis (OA) and type 2 diabetes mellitus (T2D) are two of the most widespread chronic diseases. OA and T2D have common epidemiologic traits, are considered heterogenic multifactorial pathologies that develop through the interaction of genetic and environmental factors, and have common risk factors. In addition, both of these diseases often manifest in a single patient. Despite differences in clinical manifestations, both diseases are characterized by disturbances in cellular metabolism and by an insulin-resistant state primarily associated with the production and utilization of energy. However, currently, the primary cause of OA development and progression is not clear. In addition, although OA is manifested as a joint disease, evidence has accumulated that it affects the whole body. As pathological insulin resistance is viewed as a driving force of T2D development, now, we present evidence that the molecular and cellular metabolic disturbances associated with OA are linked to an insulin-resistant state similar to T2D. Moreover, the alterations in cellular energy requirements associated with insulin resistance could affect many metabolic changes in the body that eventually result in pathology and could serve as a unified mechanism that also functions in many metabolic diseases. However, these issues have not been comprehensively described. Therefore, here, we discuss the basic molecular mechanisms underlying the pathological processes associated with the development of insulin resistance; the major inducers, regulators, and metabolic consequences of insulin resistance; and instruments for controlling insulin resistance as a new approach to therapy.

Regulation of energy metabolism in the growth plate and osteoarthritic chondrocytes.

Osteoarthritis (OA) is a chronic disorder associated mainly with pain, limited range of motion, stiffness, low-grade systemic inflammation, and articular cartilage destruction. Recent studies have demonstrated the involvement of chondrocyte differentiation (hypertrophy) as one of the mechanisms in cartilage degradation in OA. This implicates the involvement of principal changes in the regulation of cellular function associated with profound alterations in chondrocyte energy metabolism in the course of cartilage resorption. Therefore, this review describes the major energy-generating pathways and their regulatory molecules used by the growth plate chondrocytes during endochondral ossification and by articular chondrocytes in OA. These regulatory molecules facilitate either the glycolytic pathway of energy generation, which controls cell proliferation, or mitochondrial oxidative phosphorylation promoted by AMPK and sirtuins and responsible for tissue regeneration. Consideration of the disturbances in energy metabolic pathways associated with OA might provide an approach to disclose the primary causes of the disease's development and progression. Medline/PubMed was searched for publications in English using key words: osteoarthritis, epiphyseal growth plate, articular cartilage, glycolysis, oxidative phosphorylation, and regulation of energy metabolism.

Increased baseline RUNX2, caspase 3 and p21 gene expressions in the peripheral blood of disease-modifying anti-rheumatic drug-naïve rheumatoid arthritis patients are associated with improved clinical response to methotrexate therapy.

OBJECTIVE: To investigate the potential of the baseline gene expression in the whole blood of disease-modifying anti-rheumatic drug-naïve rheumatoid arthritis (RA) patients for predicting the response to methotrexate (MTX) treatment. METHODS: Twenty-six control subjects and 40 RA patients were examined. Clinical, immunological and radiographic parameters were assessed before and after 24 months of follow-up. The gene expressions in the whole blood were measured using real-time reverse transcription polymerase chain reaction. The protein concentrations in peripheral blood mononuclear cells were quantified using enzyme-linked immunosorbent assay. Receiver operating characteristic curve analyses were used to suggest thresholds that were associated with the prediction of the response. RESULTS: Decreases in the disease activity at the end of the study were accompanied by significant increases in joint space narrowing score (JSN). Positive correlations between the expressions of the Unc-51-like kinase 1 (ULK1) and matrix metalloproteinase 9 (MMP-9) genes with the level of C-reactive protein and MMP-9 expression with Disease Activity Score of 28 joints (DAS28) and swollen joint count were noted at baseline. The baseline tumor necrosis factor (TNF)α gene expression was positively correlated with JSN at the end of the follow-up, whereas p21, caspase 3, and runt-related transcription factor (RUNX)2 were correlated with the ΔDAS28 values. CONCLUSIONS: Our results suggest that the expressions of MMP-9 and ULK1 might be associated with disease activity. Increased baseline gene expressions of RUNX2, p21 and caspase 3 in the peripheral blood might predict better responses to MTX therapy.

The clinical utility of gene expression examination in rheumatology.

Rheumatoid arthritis (RA) is a chronic inflammatory disease with unknown etiology that affects various pathways within the immune system, involves many other tissues and is associated with pain and joint destruction. Current treatments fail to address pathophysiological and biochemical mechanisms involved in joint degeneration and the induction of pain. Moreover, RA patients are extremely heterogeneous and require specific treatments, the choice of which is complicated by the fact that not all patients equally respond to therapy. Gene expression analysis offer tools for patient management and personalization of patient's care to meet individual needs in controlling inflammation and pain and delaying joint destruction.

Current and future trends in Russian Rheumatology Care and Research.

This short article provides a description of the present state of rheumatology care and research in Russia and discusses opportunities for development and co-operation.

Rituximab Downregulates Gene Expression Associated with Cell Proliferation, Survival, and Proteolysis in the Peripheral Blood from Rheumatoid Arthritis Patients: A Link between High Baseline Autophagy-Related ULK1 Expression and Improved Pain Control.

Objective. To clarify molecular mechanisms for the response to rituximab in a longitudinal study. Methods. Peripheral blood from 16 RA patients treated with rituximab for a single treatment course and 26 healthy controls, blood and knee articular cartilages from 18 patients with long-standing RA, and cartilages from 14 healthy subjects were examined. Clinical response was assessed using ESR, ACPA, CRP, RF, DAS28 levels, CD19+ B-cell counts, bone erosion, and joint space narrowing scores. Protein expression in PBMCs was quantified using ELISA. Gene expression was performed with quantitative real-time PCR. Results. A decrease (p < 0.05) in DAS28, ESR, and CRP values after rituximab treatment was associated with the downregulation of MTOR, p21, caspase 3, ULK1, TNFα, IL-1ß, and cathepsin K gene expression in the peripheral blood to levels found in healthy subjects. MMP-9 expression remained significantly higher compared to controls although decreased (p < 0.05) versus baseline. A negative correlation between baseline ULK1 gene expression and the number of tender joints at the end of follow-up was observed. Conclusions. The response to rituximab was associated with decreased MTOR, p21, caspase 3, ULK1, TNFα, IL-1ß, and cathepsin K gene expression compared to healthy subjects. Residual increased expression in MMP-9, IFNα, and COX2 might account for remaining inflammation and pain. High baseline ULK1 gene expression indicates a good response in respect to pain.

Deferoxamine Suppresses Collagen Cleavage and Protease, Cytokine, and COL10A1 Expression and Upregulates AMPK and Krebs Cycle Genes in Human Osteoarthritic Cartilage.

This study reports the effects of the iron chelator deferoxamine (DFO) on collagen cleavage, inflammation, and chondrocyte hypertrophy in relation to energy metabolism-related gene expression in osteoarthritic (OA) articular cartilage. Full-depth explants of human OA knee articular cartilage from arthroplasty were cultured with exogenous DFO (1-50 µM). Type II collagen cleavage and phospho-adenosine monophosphate-activated protein kinase (pAMPK) concentrations were measured using ELISAs. Gene expression studies employed real-time PCR and included AMPK analyses in PBMCs. In OA explants collagen cleavage was frequently downregulated by 10-50 µM DFO. PCR analysis of 7 OA patient cartilages revealed that 10 µM DFO suppressed expression of MMP-1, MMP-13, IL-1ß, and TNFα and a marker of chondrocyte hypertrophy, COL10A1. No changes were observed in the expression of glycolysis-related genes. In contrast, expressions of genes associated with the mitochondrial Krebs cycle (TCA), AMPK, HIF1α, and COL2A1 were upregulated. AMPK gene expression was reduced in OA cartilage and increased in PBMCs from the same patients compared to healthy controls. Our studies demonstrate that DFO is capable of suppressing excessive collagenase-mediated type II collagen cleavage in OA cartilage and reversing phenotypic changes. The concomitant upregulation of proanabolic TCA-related gene expressions points to a potential for availability of energy generating substrates required for matrix repair by end-stage OA chondrocytes. This might normally be prevented by high whole-body energy requirements indicated by elevated AMPK expression in PBMCs of OA patients.

Association of bone loss with the upregulation of survival-related genes and concomitant downregulation of Mammalian target of rapamycin and osteoblast differentiation-related genes in the peripheral blood of late postmenopausal osteoporotic women.

We aimed to identify bone related markers in the peripheral blood of osteoporotic (OP) patients that pointed toward molecular mechanisms underlying late postmenopausal bone loss. Whole blood from 22 late postmenopausal OP patients and 26 healthy subjects was examined. Bone mineral density (BMD) was measured by DXA. Protein levels of p70-S6K, p21, MMP-9, TGFß1, and caspase-3 were quantified by ELISA. Gene expression was measured using real-time RT-PCR. OP registered by low BMD indices in late postmenopausal patients was associated with a significant upregulation of autophagy protein ULK1, cyclin-dependent kinase inhibitor p21, and metalloproteinase MMP-9 gene expression in the blood compared to the healthy controls and in a significant downregulation of mTOR (mammalian target of rapamycin), RUNX2, and ALPL gene expression, while expression of cathepsin K, caspase-3, transforming growth factor (TGF) ß1, interleukin- (IL-) 1ß, and tumor necrosis factor α (TNFα) was not significantly affected. We also observed a positive correlation between TGFß1 and RUNX2 expression and BMD at femoral sites in these patients. Therefore, bone loss in late postmenopausal OP patients is associated with a significant upregulation of survival-related genes (ULK1 and p21) and MMP-9, as well as the downregulation of mTOR and osteoblast differentiation-related genes (RUNX2 and ALPL) in the peripheral blood compared to the healthy controls.

Rheumatoid factor positivity is associated with increased joint destruction and upregulation of matrix metalloproteinase 9 and cathepsin k gene expression in the peripheral blood in rheumatoid arthritic patients treated with methotrexate.

We evaluated changes in gene expression of mTOR, p21, caspase-3, ULK1, TNF α , matrix metalloproteinase (MMP)-9, and cathepsin K in the whole blood of rheumatoid arthritic (RA) patients treated with methotrexate (MTX) in relation to their rheumatoid factor status, clinical, immunological, and radiological parameters, and therapeutic response after a 24-month follow-up. The study group consisted of 35 control subjects and 33 RA patients without previous history of MTX treatment. Gene expression was measured using real-time RT-PCR. Decreased disease activity in patients at the end of the study was associated with significant downregulation of TNF α expression. Downregulation of mTOR was observed in seronegative patients, while no significant changes in the expression of p21, ULK1, or caspase-3 were noted in any RA patients at the end of the study. The increase in erosion numbers observed in the seropositive patients at the end of the follow-up was accompanied by upregulation of MMP-9 and cathepsin K, while seronegative patients demonstrated an absence of significant changes in MMP-9 and cathepsin K expression and no increase in the erosion score. Our results suggest that increased expression of MMP-9 and cathepsin K genes in the peripheral blood might indicate higher bone tissue destruction activity in RA patients treated with methotrexate. The clinical study registration number is 0120.0810610.