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There is a lack of in vitro models able to plausibly represent the inflammation microenvironment of knee osteoarthritis (OA). We analyzed the molecules released from OA tissues (synovial membrane, cartilage, infrapatellar fat pad) and investigated whether the stimulation of human synovial fibroblasts (SFs), with synthetic cytokines (IL-1β and TNF-α or IFN-γ) or conditioned media (CM) from OA tissues, influence the SFs' response, in the sense of pro-inflammatory cytokines, chemokines, growth factors, and degradative enzymes modulation. Human SFs were obtained from OA synovial membranes. SFs and their CM were analyzed for biomarkers, proliferation rate, protein profile and gene expression, before and after stimulation. Real-time PCR and multiplex assays quantified OA-related gene expression and biomolecule production. Unlike other activators, CM from OA synovial membrane (CM-SM), significantly up-regulated all genes of interest (IL-6, IL-8, MMP-1, MMP-3, RANTES, MCP-1, TSG-6, YKL-40) in SFs. Multiplex immunoassay analysis showed that levels of OA-related cytokines (IL-6, IL-8, MCP 1, IL-1Ra), chemokine (RANTES) and growth factor (VEGF), produced by CM-SM stimulated SFs, increased significantly compared to non-stimulated SFs. Molecules released from the SM from OA patients induces OA-like changes in vitro, in specific OA synovial populations (SFs). These findings promote the use and establish a compelling in vitro model that simulates the versatility and complexity of the OA disease. This model, in the future, will allow us to study new cell therapies or test drugs by reducing or avoiding animal models.

At present, there is no effective way to treat the consequences of spinal cord injury (SCI). SCI leads to the death of neural and glial cells and widespread neuroinflammation with persisting for several weeks after the injury. Mesenchymal stem cells (MSCs) therapy is one of the most promising approaches in the treatment of this injury. The aim of this study was to characterize the expression profile of multiple cytokines, chemokines, growth factors, and so-called neuromarkers in the serum of an SCI patient treated with autologous bone marrow-derived MSCs (BM-MSCs). SCI resulted in a significant increase in the levels of neuromarkers and proteins involved in the inflammatory process. BM-MSCs administration resulted in significant changes in the levels of neuromarkers (S100, GFAP, and pNF-H) as well as changes in the expression of proteins and growth factors involved in the inflammatory response following SCI in the serum of a patient with traumatic SCI. Our preliminary results encouraged that BM-MSCs with their neuroprotective and immunomodulatory effects could affect the repair process after injury.

Mesenchymal stem cells (MSCs) are of great interest to scientists due to their application in cell therapy of many diseases, as well as regenerative medicine and tissue engineering. Recently, there has been growing evidence surrounding the research based on extracellular vesicles (EVs), especially small EVs (sEVs)/exosomes derived from MSCs. EVs/exosomes can be secreted by almost all cell types and various types of EVs show multiple functions. In addition, MSCs-derived exosomes have similar characteristics and biological activities to MSCs and their therapeutic applications are considered as a safe strategy in cell-free therapy. The aim of this study was the characterization of MSCs isolated from the chorion (CHo-MSCs) of human full-term placenta, as well as the isolation and analysis of small EVs obtained from these cells. Accordingly, in this study, the ability of small EVs' uptake is indicated by synovial fibroblasts, osteoblasts and periosteum-derived MSCs. Improvement in the understanding of the structure, characteristics, mechanism of action and potential application of MSCs-derived small EVs can provide new insight into improved therapeutic strategies.

Objective: Osteoarthritis (OA) commonly affects weight-bearing joints and is characterized by articular cartilage breakdown combined with osteophyte formation at the joint margins and chronic nonspecific inflammation of synovium. Understanding the profile of inflammation in a patient population is an essential starting point to predict or prevent OA progression. The aim of this study was to identify the profile of selected biomolecules in synovial fluid (SF) and investigate the correlation according to gender, age, and severity of the disease within patients from among the general knee OA population., Design: In our study SF samples were aspirated from the knees of 65 OA patients (46 patients with early knee OA and 19 patients with end-stage knee OA according to the Kellgren-Lawrence grading scale). The concentration of interleukins (IL-6, IL-8), matrix metalloproteinases (MMP-1, MMP-3, MMP-13), MMPs inhibitors (TIMP-1, TIMP-2), cartilage oligomeric matrix protein (COMP), and adiponectin was analyzed using a multiplex ELISA-based approach., Conclusions: Our results indicate significant linear correlation of MMP-13 and COMP concentration with age ( P < 0.05), but not with OA severity. In fact, 3 of the examined biomolecules, MMP-3 ( P < 0.01), TIMP-1 ( P < 0.01), and COMP ( P < 0.05) significantly correlate with the grade of knee OA and might be associated with OA severity.

Autologous blood products, such as platelet-rich plasma (PRP), are gaining increasing interest in different fields of regenerative medicine. Although growth factors, the main components of PRP, are thought to stimulate reparation processes, the exact mechanism of action and main effectors of PRP are not fully understood. Plasma contains a high amount of extracellular vesicles (EVs) produced by different cells, including anucleated platelets. Platelet-derived EVs (PL-EVs) are the most abundant type of EVs in circulation. Numerous advantages of PL-EVs, including their ability to be released locally, their ease of travel through the body, their low immunogenicity and tumourigenicity, the modulation of signal transduction as well as the ease with which they can be obtained, has attracted increased attention n. This review focuses briefly on the biological characteristics and isolation methods of PL-EVs, including exosomes derived from platelets (PL-EXOs), and their involvement in the pathology of diseases. Evidence that shows how PL-EVs can be used as a novel tool in medicine, particularly in therapeutic and regenerative medicine, is also discussed in this review.

Mesenchymal stem cells (MSCs) have been demonstrated to have a great potential in the treatment of several diseases due to their differentiation and immunomodulatory capabilities and their ability to be easily cultured and manipulated. Recent investigations revealed that their therapeutic effect is largely mediated by the secretion of paracrine factors including exosomes. Exosomes reflect biophysical features of MSCs and are considered more effective than MSCs themselves. Alternative approaches based on MSC-derived exosomes can offer appreciable promise in overcoming the limitations and practical challenges observed in cell-based therapy. Furthermore, MSC-derived exosomes may provide a potent therapeutic strategy for various diseases and are promising candidates for cell-based and cell-free regenerative medicine. This review briefly summarizes the development of MSCs as a treatment for human diseases as well as describes our current knowledge about exosomes: their biogenesis and molecular composition, and how they exert their effects on target cells. Particularly, the therapeutic potential of MSC-derived exosomes in experimental models and recent clinical trials to evaluate their safety and efficacy are summarized in this study. Overall, this paper provides a current overview of exosomes as a new cell-free therapeutic agent.

A combination of biochemical, biophysical and biological techniques was used to study calf thymus DNA interaction with newly synthesized 7-MEOTA-tacrine thiourea 12-17 and urea heterodimers 18-22, and to measure interference with type I and II topoisomerases. Their biological profile was also inspected in vitro on the HL-60 cell line using different flow cytometric techniques (cell cycle distribution, detection of mitochondrial membrane potential dissipation, and analysis of metabolic activity/viability). The compounds exhibited a profound inhibitory effect on topoisomerase activity (e.g. compound 22 inhibited type I topoisomerase at 1 µM concentration). The treatment of HL-60 cells with the studied compounds showed inhibition of cell growth especially with hybrids containing thiourea (14-17) and urea moieties (21 and 22). Moreover, treatment of human dermal fibroblasts with the studied compounds did not indicate significant cytotoxicity. The observed results suggest beneficial selectivity of the heterodimers as potential drugs to target cancer cells.

Objective This study aimed to compare microfracture and application of adipose-derived stem cells (ADSCs) by local adherent technique enhanced by platelet-rich plasma (PRP) to provide a new approach for the repair of cartilage defect. Design Full-thickness cylindrical defects were created in the medial femoral condyle in 9 New Zealand White rabbits (5 months old, 4.65 ± 0.20 kg). Two groups of rabbits ( n = 3) were either treated with ADSCs (Group 1) or the microfracture technique (Group 2) following intraarticular injection of PRP 3 times in weekly intervals. Rabbits in control group ( n = 3) remained untreated. The outcome was assessed macroscopically, histologically, and immunohistochemically. Results At the end of week 12, Group 1 showed better defect filling compared with Group 2. Specimens treated with the combination of ADSCs and PRP exhibited significant differences from the other groups in all criteria of International Cartilage Repair Society macroscopic scoring system. Conclusions Intraarticular injection of autologous PRP in combination with transplantation of autologous ADSCs by local adherent technique enhances the quality of cartilage defect repair with better results in comparison with microfracture surgery in a rabbit model.

Articular cartilage has limited capacity for natural regeneration and repair. In the present study, we evaluated kartogenin (KGN), a bioactive small heterocyclic molecule, for its effect on in vitro proliferation and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells (hBMSCs) in monolayer culture and in co-culture models in vitro. OA osteochondral cylinders and hBMSCs were collected during total knee replacement. The effect of KGN on hBMSCs during 21 days of culture was monitored by real-time proliferation assay, immunofluorescence staining, histological assay, scanning electron microscopy (SEM) (imaging and multiplex enzyme-linked immunosorbent assay) ELISA assay. The rate of proliferation of hBMSCs was significantly increased by treatment with 10 µM KGN during nine days of culture. Histological and SEM analyses showed the ability of hBMSCs in the presence of KGN to colonize the surface of OA cartilage and to produce glycosaminoglycans and proteoglycans after 21 days of co-culture. KGN treated hBMSCs secreted higher concentrations of TIMPs and the secretion of pro-inflammatory molecules (MMP 13, TNF-α) were significantly suppressed in comparison with control without hBMSCs. Our preliminary results support the concept that 10 µM KGN enhances proliferation and chondrogenic differentiation of hBMSCs and suggest that KGN is a potential promoter for cell-based therapeutic application for cartilage regeneration., Competing Interests: The authors declare no conflict of interest.

Background: Polycystic ovary syndrome (PCOS) is commonly associated with endocrine, metabolic, cardiovascular and other morbidities. However its association with autoimmune diseases is still controversial., Aim: The aim of this study was to assess the prevalence of non organ-specific and antithyroid, antibodies in PCOS women compared to healthy controls., Methods: The study included 152 women with PCOS and 76 healthy controls for the evaluation of non organ-specific autoimmunity and 64 PCOS and 68 controls for the study of organ-specific autoimmunity. All sera were tested for autoantibodies.using the ELISA method., Results: There were no significant differences in the prevalence of ANA, SSA, SSB, anti-dsDNA, anti-RNP, ANCA/MPO or ANCA/PR3 between PCOS and controls. The prevalence of ACLA IgG was higher in controls than PCOS (5.4% v.s. 0%, P=0.011). Patients had a higher prevalence of anti-TPO antibodies (18.75% v.s. 7.35%, P=0.045) and slightly but not significantly higher prevalence of autoimmune thyroiditis (18.75% v.s. 10.29%) than controls., Conclusion: The prevalence of non organ-specific autoantibodies in PCOS women is low and similar to controls. On the other hand, we found a slightly higher prevalence of thyroid autoimmunity in PCOS women.

Diabetes mellitus type 1 is a form of diabetes mellitus that results from the autoimmune destruction of insulin-producing beta cells in the pancreas. The current gold standard therapy for pancreas transplantation has limitations because of the long list of waiting patients and the limited supply of donor pancreas. Mesenchymal stem cells (MSCs), a relatively new potential therapy in various fields, have already made their mark in the young field of regenerative medicine. Recent studies have shown that the implantation of MSCs decreases glucose levels through paracrine influences rather than through direct transdifferentiation into insulin-producing cells. Therefore, these cells may use pro-angiogenic and immunomodulatory effects to control diabetes following the cotransplantation with pancreatic islets. In this review, we present and discuss new approaches of using MSCs in the treatment of diabetes mellitus type 1.

In the present paper we develop a new non-cell based (cell-free) therapeutic approach applied to BV2 microglial cells and spinal cord derived primary microglia (PM) using conditioned media from rat bone marrow stromal cells (BMSCs-CM). First we collected conditioned media (CM) from either naive or injured rat spinal cord tissue (SCI-CM, inflammatory stimulation agent) and from rat bone marrow stromal cells (BMSCs-CM, therapeutic immunomodulation agent). They were both subsequently checked for the presence of chemokines and growth, neurotrophic and neural migration factors using proteomics analysis. The data clearly showed that rat BMSCs-CM contain in vitro growth factors, neural migration factors, osteogenic factors, differentiating factors and immunomodulators, whereas SCI-CM contain chemokines, chemoattractant factors and neurotrophic factors. Afterwards we determined whether the BMSCs-CM affect chemotactic activity, NO production, morphological and pro-apoptotic changes of either BV2 or PM cells once activated with SCI-CM. Our results confirm the anti-migratory and NO-inhibitory effects of BMSCs-CM on SCI-CM-activated microglia with higher impact on primary microglia. The cytotoxic effect of BMSCs-CM occurred only on SCI-CM-stimulated BV2 cells and PM, not on naive BV2 cells, nor on PM. Taken together, the molecular cocktail found in BMSCs-CM is favorable for immunomodulatory properties.

A novel series of trisubstituted acridines were synthesized with the aim of mimicking the effects of BRACO19. These compounds were synthesized by modifying the molecular structure of BRACO19 at positions 3 and 6 with heteroacyclic moieties. All of the derivatives presented in the study exhibited stabilizing effects on the human telomeric DNA quadruplex. UV-vis spectroscopy, circular dichroism, linear dichroism and viscosimetry were used in order to study the nature of the DNA binding in more detail. The results show that all of the novel derivatives were able to fold the single-stranded DNA sequences into antiparallel G-quadruplex structures, with derivative 15 exhibiting the highest stabilizing capability. Cell cycle analysis revealed that a primary trend of the "braco"-like derivatives was to arrest the cells in the S- and G2M-phases of the cell cycle within the first 72h, with derivative 13 and BRACO19 proving particularly effective in suppressing cell proliferation. All studies derivatives were less toxic to human fibroblast cell line in comparison with HT 29 cancer cell line., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Mesenchymal stromal cells (MSCs) in the pancreatic microenvironment can improve diabetes mellitus (DM). The aim of the present study was to determine whether different pancreatic microenvironments influence the improvement of hyperglycemia and insulin deficiency., Methods: MSCs isolated from rat bone marrow were transplanted directly into different pancreatic microenvironments in male DM rats. DM was induced in the rats by streptozotocin injection. The rats were divided into 5 groups: normal control rats, DM control rats, and 3 experimental groups (DM rats plus MSCs injected into the head of the pancreas, the tail of the pancreas, or the whole pancreas). The body weight and blood glucose of the rats were monitored during the experiment after transplantation of the MSCs. Histopathologic and immunohistochemical analyses were used to detect the presence and number of islets and insulin production in the pancreatic tissue of the rats after MSC transplantation., Results: At 28 days after MSC transplantation, we observed a statistically significant decrease in the blood glucose level and an increase in weight in DM rats compared with DM control rats (P < 0.0001 and P < 0.03, respectively). A comparison of each of the DM rat groups treated with MSCs showed no significant differences in the blood glucose levels or body weight., Conclusion: Our results suggest that transplantation of MSCs could improve DM in the pancreatic microenvironment in an animal model with streptozotocin-induced DM. The different pancreatic areas into which the MSCs were implanted had no significant influence on the improvement in hyperglycemia and insulin deficiency., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background and Aims: Umbilical cord blood (UCB) has been identified as a good source of hematopoietic and nonhematopoietic stem cells that can be easily isolated. In the present study we investigated the possibility of whether stem cells in mononuclear UCB grown under defined conditions can produce progeny with neural phenotype., Methods: A combination of antigen-driven magnetic cell sorting (MACs) method and defined culture conditions specific for cells of neural lineages were used for isolation, expansion and differentiation of CD133+/- cells from UCB. Both UCB-derived fractions were expanded by exposure to growth factors (EGF, bFGF). Differentiation was induced by replacing them with fetal bovine serum. Using immunocytochemistry, the cell markers for neural (MAP2, GFAP, RIP) and non-neural lineages (S-100, von Willebrand factor) were detected., Results: The analysis revealed occurrence of fully mature neural and non-neural lineages, which showed qualitative and quantitative differences between population of CD133+ and CD133- cells. The expression levels of MAP2 and RIP in CD133+ were significantly higher than in CD133-, more GFAP positive cells were found in the CD133-. At the same time, S-100 was expressed by 32.47 ± 6.24% of CD133- cells and 29.42 ± 1.32% of CD133- cell expressed a von Willebrand factor antigen., Conclusions: Our results indicate that stem cells derived from umbilical cord blood are easy to obtain, proliferate and are able to differentiate towards the cells of neural lineages, which represents a promising way for their utilization in cell-based therapies for CNS injuries and diseases., (Copyright © 2011 IMSS. Published by Elsevier Inc. All rights reserved.)

Transplantation of bone marrow mesenchymal stromal cells (MSCs) has been shown to improve the functional recovery in various models of spinal cord injury (SCI). However, the issues of the optimal dose, timing, and route of MSC application are crucial factors in achieving beneficial therapeutic outcomes. The objective of this study was to standardize the intrathecal (IT) catheter delivery of rat MSCs after SCI in adult rats. MSCs labeled with PKH-67 were administered by IT delivery to rats at 3 or 7 days after SCI as one of the following treatment regimens: (1) a single injection (5×10(5) MSCs/rat), or (2) as three daily injections (5×10(5) MSCs/rat/d for a total of 1.5×10(6) MSCs/rat over 3 days, injected on days 3, 4, and 5, or days 7, 8, and 9 following SCI. The animals were behaviorally tested for 4 weeks using the Basso, Beattie, and Bresnahan (BBB) locomotor rating scale, and histologically assessed for MSC survival, distribution, and engraftment properties after 28 days. Rats treated with a single injection of MSCs at 3 or 7 days post-injury showed a modest, non-significant improvement in function and low survival of grafted MSCs, which were found attached to the pia mater or accumulated around the anterior spinal artery. In contrast, rats treated with three daily injections of MSCs at days 7, 8, and 9, but not on days 3, 4, and 5, showed significantly higher motor function recovery (BBB score 16.8±1.7) at 14-28 days post-injury. Transplanted PKH-67 MSCs were able to migrate and incorporate into the central lesion. However, only a limited number of surviving MSCs, ranging from 24,128±1170 to 116,258±8568 cells per graft, were observed within the damaged white matter. These results suggest that repetitive IT transplantation, which imposes a minimal burden on the animals, may improve behavioral function when the dose, timing, and targeted IT delivery of MSCs towards the lesion cavity are optimized.