Vitaly Olegovich Omelchenko

Monocytic myeloid-derived suppressor cells (M-MDSCs) expand in axial spondyloarthritis (axSpA), although their pathogenetic role and functional state are still unclear. The aim of this study was to investigate checkpoint receptors PD-1/Tim-3 expression on M-MDSCs in axSpA, assessing their relationship with clinical disease parameters and M-MDSC suppressor potential. The study included 19 healthy donors and 32 axSpA patients. As markers of the suppressor potential of M-MDSCs, suppressor molecules arginase 1 (Arg-1) and tyrosine kinase Mer (MerTK) were evaluated. M-MDSC count and their expression of PD-1, Tim-3, Arg-1, and MerTK were evaluated using flow cytometry. M-MDSC frequency directly correlated with the level of C-reactive protein, whereas PD-1 expression on M-MDSCs inversely correlated with the erythrocyte sedimentation rate. Thus, high axSpA activity was associated with an increased M-MDSC content with unchanged PD-1 expression, whereas low activity was linked to an increased PD-1 expression without changing M-MDSC frequency. Additionally, the PD-1⁺ M-MDSC count directly correlated with the percentage of MerTK⁺ M-MDSCs. Tim-3 showed no correlation with the clinical disease parameters; however, an inverse correlation was observed between Tim-3 and Arg-1 expression in M-MDSCs. In turn, the Arg-1⁺ M-MDSCs count in patients receiving non-steroidal anti-inflammatory drugs and having higher disease activity was reduced, while in the patient group receiving tumor necrosis factor alpha-inhibitors and having lower activity, it was increased. The obtained results may indicate a positive role of PD-1 in preventing inflammation in axSpA, whereas Tim-3 presumably weakens the anti-inflammatory M-MDSC potential in axSpA.

Rheumatoid arthritis (RA) is a chronic autoimmune disease that leads to irreversible structural joint damage and disability. Even with modern therapeutic approaches, a subset of patients continues to exhibit persistent inflammatory activity, which underscores the importance of identifying prognostic markers of treatment efficacy. Increasing attention is currently being paid to the morphological features of synovial membrane remodeling and to the subpopulation composition of lymphocytes in the peripheral blood of RA patients. The aim of our study was to investigate associations between the subpopulation profile of immunocompetent cells in the synovial membrane and in the peripheral blood of patients with rheumatoid arthritis of varying disease activity. Material and methods . The study included 52 patients, including 40 with RA and 12 with osteoarthritis as a comparison group. All patients underwent standard clinical evaluation, disease activity assessment (ESR, C-reactive protein content, composite activity indices), and peripheral blood lymphocyte subpopulation analysis. In RA patients with knee synovitis, ultrasound-guided fine-needle aspiration biopsy of the synovial membrane was performed. Results and discussion . A positive association between the number of CD4⁺ T cells in the synovium and clinical disease activity in RA is revealed, whereas an inverse relationship was observed in peripheral blood. These findings provide additional insights into local immune responses in RA and highlight the potential of synovial biopsy analysis for therapy personalization. Conclusions . In patients with RA of varying disease activity, divergent associations were identified between the number of CD4⁺ T helper cells in the synovial membrane of affected joints and in peripheral blood.

Regulation of inflammation events in inflammatory/autoimmune rheumatic diseases, particularly axial spondyloarthritis, is critically important for alleviating clinical symptoms. Recent studies have shown the key role of PD-1 and Tim-3 inhibitory receptors in regulating the functional phenotype of monocytes in inflammatory/autoimmune diseases. The aim of this study was to investigate the expression of the PD-1 and Tim-3 inhibitory receptors in different subsets of monocytes in patients with axial spondyloarthritis (axSpA), and to evaluate their association with clinical features, as well as with tolerogenic marker Mer tyrosine kinase (MerTK). The expression of PD-1 and Tim-3 in classical, intermediate, and non-classical monocytes was determined by means of multicolor flow cytometry in peripheral blood mononuclear cells in 60 patients with axial spondyloarthritis and 40 healthy donors. The analysis of circulating monocyte subpopulations revealed an increased proportion of classical monocytes in patients, which positively correlated with systemic inflammation markers: erythrocyte sedimentation rate and C-reactive protein. Like as in donor group, the highest expression of PD-1 and Tim-3 in axSpA patients was observed in intermediate and non-classical monocyte subsets. However, the relative proportions of PD-1 ⁺ and PD-1 ⁺ Tim-3 ⁺ cells in intermediate and non-classical monocytes were significantly decreased in axSpA patients, whereas Tim-3 expression levels did not differ from the donor values. Notably, the proportion of PD-1 ⁺ cells directly correlated with expression of MerTK, a factor mediating the anti-inflammatory activity of monocytes/macrophages in all monocyte subsets. A statistically significant decrease in PD-1 expression and co-expression of PD-1 and Tim-3 was detected in HLA-B27 antigen carriers, patients with advanced and late-stage disease, and those with peripheral axSpA, including hip joint involvement. On the contrary, in HLA-B27-negative patients and those with early/non-radiographic stage disease, PD-1 expression was comparable to donor levels, while Tim-3 ⁺ cell contents were increased among classical monocytes. In patients receiving first-line therapy, the decreased PD-1 expression and PD-1 ⁺ Tim-3 ⁺ co-expression directly correlated with disease activity. The obtained data that include a reduced PD-1 expression and PD-1 ⁺ Tim-3 ⁺ co-expression in intermediate and non-classical monocytes in patients with genetic predisposition, advanced disease stages, and peripheral joint involvement, as well as the association between PD-1 ⁺ cell number and MerTK expression in all monocyte subsets, suggest a dysregulation of inhibitory checkpoint receptor expression under inflammatory conditions. These findings could presume a diminished anti-inflammatory capacity of monocytes.

Osteoporosis is becoming a “silent pandemic” because of its ever-increasing prevalence and the absence of clinical manifestations until a bone fracture happens. The purpose of this review is to summarize the actual data on the pathogenesis of osteoporosis and its treatment options. The disease develops through a multifactorial process involving an imbalance between bone remodeling and different factors like genetics, non-coding RNA regulation, osteoimmune dysregulation, oxidative stress, cellular senescence, and fat–bone interactions. Existing medications have beneficial effects by preserving and increasing bone density and reducing the risk of fractures. Among them, there are bisphosphonates, strontium ranelate, calcitonin, estrogen-progestin therapy, selective estrogen receptor modulators, and parathyroid hormone analogues. Otherwise, they suffer from certain disadvantages, such as adverse effects, including serious ones, and limitations associated with comorbidity. Targeting pathways underlying bone metabolism could significantly improve the therapeutic options and provide new tools in the fight against osteoporosis. We consider here targeted therapeutics that are already in clinical practice, as well as the most promising novel agents that are now under development: antibodies, siRNAs, aptamers, and small molecules.

Innate lymphoid cells (ILCs) are the innate analogues of lymphocytes that do not express antigen-specific receptors and are primarily found in tissues and mucosa. ILCs are divided into three groups based on the transcription factors and cytokines they secrete. Group 1 ILCs produce interferon (IFN)-γ in response to IL-12 and are dependent on the transcription factor T-bet; group 2 ILCs (ILC2s) predominantly produce type 2 cytokines (IL-5, IL-4, IL-9, and IL-13) in response to IL-33, IL-25, and thymic stromal lymphopoietin (TSLP) and are dependent on GATA3. Group 3 ILCs include ILC3s and lymphoid tissue inducer cells (LTi). The latter group secretes IL-17 and IL-22 in response to IL-1β and IL-23 and functionally depends on RORγt. Recently, early ILC precursors were found in peripheral blood, which were defined by the CD5 marker and are likely to be of thymic origin. These cells can, on demand, enter the bloodstream (like monocytes), move with the blood flow into tissues for subsequent differentiation into a mature phenotype. In this work, we assessed the content of CD5+ILC2 in peripheral blood of patients with rheumatoid arthritis (RA), which is characterized by chronic inflammation in the joints and uncontrolled cell proliferation, thus maintaining the inflammatory events. In this work, we used peripheral blood from patients with RA (n = 7) and conditionally healthy donors (n = 13). The obtained peripheral blood mononuclear cells (PBMPs) were stained with the following panel of antibodies: anti-lineage (CD2/3/14/16/19/20/56/235a), antiCD11c and anti-FceR1 alpha-FITC, anti-CD294-PE, anti-CD127-PerCP/Cy5.5, antiCD117-APC, anti-CD5-BV-450. Innate lymphoid cells were defined as Lin-CD127+, CD294+ILCs were identified as ILC2. The proportion of CD5+ cells among ILC2 was also evaluated. The cell phenotype was analyzed by flow cytometry. We showed that the proportion of ILC2 among all PBMPs was significantly lower in patients with RA compared to the donor group, and the number of CD5+ILC2 among ILC2 was significantly higher than in the control group. The obtained results are unique and provide us with new data on the changing percentage of ILC2 among PC MNCs and CD5+ILC2 among ILC2.

Complex mechanisms of thymus functioning affect its structural matrix thus promoting gradual accumulation of genetic mutations, changes in gene expression and premature immunosenescence. The study of early thymic cell migrants (ETM) is necessary to identify possible diagnostic biomarkers and potential checkpoints of autoimmune diseases (AID). We have studied peripheral blood samples of patients with rheumatoid arthritis (RA), psoriatic arthritis (PsA), psoriasis (PS) and healthy donors. The aim of the study was to assess the ratio of recent cell migrants from the thymus and circulating naïve T regulatory cells in healthy persons and autoimmune diorders. The relative numbers of recent thymic migrants CD4+CD45RA+CD31+ (ETM), and the contents of FoxP3+T regulatory cells (Treg) in this population were determined by flow cytometry. The number of proliferating cells was determined by intracellular Ki-67 marker. The results show that the number of ETM was reduced in the group of patients with RA and patients with PsA compared with the healthy donor group (17.3%, 18.6% vs. 23.6%), and did not change significantly in patients with psoriasis. There were no significant differences between donors and patients with AIDs on the contents of proliferating ETM and Tregs. The proliferation rates of T regulatory cells in patients with psoriatic pathology were comparable to donor values, with a trend for its increase in RA patients. The numbers of proliferating naive FoxP3+Tregs in RA patients was significantly higher than the numbers of proliferating ETM (15.8% vs 3.1%, p 0.05, respectively). A decrease in relative number of ETM in AID may be associated with activation of T lymphocytes and higher proportion of central memory T cells. The number of Tregs among ETM, and their proliferation rates in patients with psoriatic disorder suggest the absence of disturbed Treg generation in thymus. The ambiguous results obtained in patients with RA are of potential significance for diagnostics and therapy. However, they require additional confirmation in a larger sample of patients.

Currently, there is insufficient data in the literature on the efficacy and safety of platelet-rich plasma (PRP) therapy in patients with osteoarthritis (OA) of the knee. The aim of this study was to evaluate the efficacy and safety of local therapy with PRP and hyaluronic acid (HA) by intra-articular injection in patients with stage II-III knee OA.Material and methods. Patients (n = 32, age 60.9 ± 7.7 years) were divided into 2 groups: the first (18 patients) received a combination of PRP and HA intra-articularly. PRP was injected according to the schedule (1, 7, and 14 days) and HA (1 day). In the second group (14 patients), HA was injected intra-articularly in isolation. Manipulations were performed on one knee joint. Prior to treatment, all patients underwent radiography of the knee joint and ultrasound in three zones (upper pole of the patella, lateral and medial condyles of the tibia) to evaluate changes in cartilage thickness in the dynamic range. Diagnostic arthroscopy of the affected joint was performed to objectify the severity of cartilage lesions one day prior to the first intra-articular drug injection. Control ultrasound was performed at 12 and 24 weeks of followup. The clinical efficacy of the therapy was assessed using WOMAC, Lequesne, and visual analog pain scale.Results. Analysis of the dynamics of pain syndrome showed an analgesic effect of therapy and improvement of the functional status of patients in the first and second groups after 3 months. In both groups, after 3 months of observation, according to ultrasound data, a statistically significant increase in cartilaginous tissue in the area of the upper pole of the patella was revealed, as well as in the area of the lateral condyle of the tibia in the second group after 3 months with a further increase in cartilage thickness by the 6th month of observation. In the third localization, the medial condyle of the tibia, both groups showed no statistically significant increase in cartilage thickness.Conclusions. PRP and HA are effective and safe in the treatment of stage II–III knee OA with chronic pain syndrome. Regression of clinical effects occurred 6 months after initiation of therapy, while the combination of therapy methods did not contribute to prolongation of positive results. Thus, the combination of the indicated treatment methods (PRP + HA) does not seem appropriate.

Introduction. Bioluminescent labeling of tumor cells is becoming the standard in preclinical studies of novel anti-cancer drugs. Despite the variety of luciferases, many are not suitable for in vivo imaging.Aim: to compare different Nanolux substrates in vitro and demonstrate the feasibility of using the NLuc/h-coelenterazine pair for in vivo imaging of disseminated tumor cells using the IVIS Spectrum system.Materials and methods. The target cell line and T-cells expressing chimeric antigen receptors (CAR) were obtained via lentiviral cell transduction. Luminescence was measured with a Luminoskan™ Microplate Luminometer, while in vivo imaging in an NSG mouse model of CAR T-cell therapy was performed using the IVIS Spectrum system.Results. Various substrates were compared in vitro for luminescence brightness and stability.Additionally, the use of h-coelenterazine in combination with Nluc luciferase was demonstrated for in vivo cell imaging in mice. A genetically modified Nalm6-NLuc-CopGFP cell line was successfully obtained.Conclusion. The comparison of various Nluc substrates in vitro and in vivo identified an optimal enzyme/substrate pair, which can serve as a valuable tool for assessing efficacy, safety and toxicity of compounds and cellular products being developed for antitumor therapy. The plasmid encoding Nluc can also be used to modify other cell lines necessary for the development and characterization of new gene therapy approaches.

Osteoporosis is a metabolic skeletal disease characterized by a high risk of fractures with minimal trauma due to decreased bone mass, microarchitecture, bone mineralization, and bone strength. The range of drugs for the treatment of osteoporosis is limited, and patients often have certain restrictions on their use. It is important to search for new therapeutic options for the treatment of osteoporosis. The leading role of the Wnt pathway in the process of bone remodeling has been proven. Oligonucleotide aptamers represent promising alternative to antibodies for targeted regulation of Wnt pathway, particularly through an inhibition of functional activity of Wnt antagonist sclerostin. In this study, we produced a series of chemically modified DNA aptamers targeting sclerostin and assessed their properties by in vitro experiments with osteoblasts and osteoclasts. We conducted that aptamers do not have cytotoxicity against osteoblasts and osteoclasts. Based on these results, aptamers are promising for further investigation of their targeting interactions with Wnt-pathway inhibitors.

Vitaly Omelchenko, M.D., Elena Letyagina, M.D., Maxim Korolev, M.D.. J Rheum Dis -0001;0:. https://doi.org/10.4078/jrd.2024.0040