Collagen-containing scaffolds enhance attachment and proliferation of non-cultured bone marrow multipotential stromal cells

Large bone defects are ideally treated with autografts, which have many limitations. Therefore, osteoconductive scaffolds loaded with autologous bone marrow (BM) aspirate are increasingly used as alternatives. The purpose of this study was to compare the growth of multipotential stromal cells (MSCs) from unprocessed BM on a collagen-containing bovine bone scaffold (Orthoss® Collagen) with a non-collagen-containing bovine bone scaffold, Orthoss®. Another collagen-containing synthetic scaffold, Vitoss® was included in the comparison. Colonization of scaffolds by BM MSCs (n = 23 donors) was evaluated using microscopy, colony forming unit-fibroblast assay and flow-cytometry. The number of BM MSCs initially attached to Orthoss® Collagen and Vitoss® was similar but greater than Orthoss® (p = 0.001 and p = 0.041, respectively). Furthermore, the number of MSCs released from Orthoss® Collagen and Vitoss® after 2-week culture was also higher compared to Orthoss® (p = 0.010 and p = 0.023, respectively). Interestingly, collagen-containing scaffolds accommodated larger numbers of lymphocytic and myelomonocytic cells. Additionally, the proliferation of culture-expanded MSCs on Orthoss® collagen and Vitoss® was greater compared to Orthoss® (p = 0.047 and p = 0.004, respectively). Collectively, collagen-containing scaffolds were superior in supporting the attachment and proliferation of MSCs when they were loaded with unprocessed BM aspirates. This highlights the benefit of collagen incorporation into bone scaffolds for use with autologous bone marrow aspirates as autograft substitutes

Mesenchymal Stem Cell functionalization for enhanced therapeutic applications

To date, the therapeutic efficacy of human mesenchymal stem cells (hMSCs) has been investigated in various clinical trials with moderate or in some cases inconsistent results. The still elusive reproducibility relates in part with constitutive differences in the cell preparation, translated into variable “cell potencies”. Other factors include poor cell homing and survival, and age/disease-associated host tissue impairment. It is well accepted that within in vivo niches MSCs exist as heterogeneous cell populations with different stemness propensities and supportive functions. Phenotype-based MSC purification of homogeneous subsets can result in cell populations with distinct biological functions. In addition, preclinical studies have shown that MSC functionalization in vitro, via cell priming, can boost their immunomodulatory, trophic and reparative capacities in vivo. Therefore, in the present review we discuss how phenotype-based MSC purification and MSC priming technologies can contribute to an improved MSC-based product for safer and more effective therapeutic applications

Gene expression and functional comparison between multipotential stromal cells from lateral and medial condyles of knee osteoarthritis patients

Osteoarthritis (OA) is the most common degenerative joint disorder. Multipotential stromal cells (MSCs) have a crucial role in joint repair, but how OA severity affects their characteristics remains unknown. Knee OA provides a good model to study this, as osteochondral damage is commonly more severe in the medial weight-bearing compartment compared to lateral side of the joint. This study utilised in vitro functional assays, cell sorting, gene expression and immunohistochemistry to compare MSCs from medial and lateral OA femoral condyles. Despite greater cartilage loss and bone sclerosis in medial condyles, there was no significant differences in MSC numbers, growth rates or surface phenotype. Culture-expanded and freshly-purified medial-condyle MSCs expressed higher levels of several ossification-related genes. Using CD271-staining to identify MSCs, their presence and co-localisation with TRAP-positive chondroclasts was noted in the vascular channels breaching the osteochondral junction in lateral condyles. In medial condyles, MSCs were additionally found in small cavities within the sclerotic plate. These data indicate subchondral MSCs may be involved in OA progression by participating in cartilage destruction, calcification and sclerotic plate formation and that they remain abundant in severe disease. Biological or biomechanical modulation of these MSCs may be a new strategy towards cartilage and bone restoration in knee OA

Functional and Molecular Analysis of Human Osteoarthritic Chondrocytes Treated with Bone Marrow-Derived MSC-EVs

\ua9 2024 by the authors.Osteoarthritis (OA) is a degenerative joint disease, causing impaired mobility. There are currently no effective therapies other than palliative treatment. Mesenchymal stromal cells (MSCs) and their secreted extracellular vesicles (MSC-EVs) have shown promise in attenuating OA progression, promoting chondral regeneration, and modulating joint inflammation. However, the precise molecular mechanism of action driving their beneficial effects has not been fully elucidated. In this study, we analyzed MSC-EV-treated human OA chondrocytes (OACs) to assess viability, proliferation, migration, cytokine and catabolic protein expression, and microRNA and mRNA profiles. We observed that MSC-EV-treated OACs displayed increased metabolic activity, proliferation, and migration compared to the controls. They produced decreased proinflammatory (Il-8 and IFN-γ) and increased anti-inflammatory (IL-13) cytokines, and lower levels of MMP13 protein coupled with reduced expression of MMP13 mRNA, as well as negative microRNA regulators of chondrogenesis (miR-145-5p and miR-21-5p). In 3D models, MSC-EV-treated OACs exhibited enhanced chondrogenesis-promoting features (elevated sGAG, ACAN, and aggrecan). MSC-EV treatment also reversed the pathological impact of IL-1β on chondrogenic gene expression and extracellular matrix component (ECM) production. Finally, MSC-EV-treated OACs demonstrated the enhanced expression of genes associated with cartilage function, collagen biosynthesis, and ECM organization and exhibited a signature of 24 differentially expressed microRNAs, associated with chondrogenesis-associated pathways and ECM interactions. In conclusion, our data provide new insights on the potential mechanism of action of MSC-EVs as a treatment option for early-stage OA, including transcriptomic analysis of MSC-EV-treated OA, which may pave the way for more targeted novel therapeutics

Gene expression signatures of synovial fluid multipotential stromal cells (MSCs) in advanced knee osteoarthritis and following knee joint distraction

Osteoarthritis (OA) is the most common musculoskeletal disorder. Although joint replacement remains the standard of care for knee OA patients, knee joint distraction (KJD), which works by temporarily off-loading the joint for 6–8 weeks, is becoming a novel joint-sparing alternative for younger OA sufferers. The biological mechanisms behind KJD structural improvements remain poorly understood but likely involve joint-resident regenerative cells including multipotent stromal cells (MSCs). In this study, we hypothesized that KJD leads to beneficial cartilage-anabolic and anti-catabolic changes in joint-resident MSCs and investigated gene expression profiles of synovial fluid (SF) MSCs following KJD as compared with baseline. To obtain further insights into the effects of local biomechanics on MSCs present in late OA joints, SF MSC gene expression was studied in a separate OA arthroplasty cohort and compared with subchondral bone (SB) MSCs from medial (more loaded) and lateral (less loaded) femoral condyles from the same joints. In OA arthroplasty cohort (n = 12 patients), SF MSCs expressed lower levels of ossification- and hypotrophy-related genes [bone sialoprotein (IBSP), parathyroid hormone 1 receptor (PTH1R), and runt-related transcription factor 2 (RUNX2)] than did SB MSCs. Interestingly, SF MSCs expressed 5- to 50-fold higher levels of transcripts for classical extracellular matrix turnover molecules matrix metalloproteinase 1 (MMP1), a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5), and tissue inhibitor of metalloproteinase-3 (TIMP3), all (p < 0.05) potentially indicating greater cartilage remodeling ability of OA SF MSCs, compared with SB MSCs. In KJD cohort (n = 9 patients), joint off-loading resulted in sustained, significant increase in SF MSC colonies’ sizes and densities and a notable transcript upregulation of key cartilage core protein aggrecan (ACAN) (weeks 3 and 6), as well as reduction in pro-inflammatory C–C motif chemokine ligand 2 (CCL2) expression (weeks 3 and 6). Additionally, early KJD changes (week 3) were marked by significant increases in MSC chondrogenic commitment markers gremlin 1 (GREM1) and growth differentiation factor 5 (GDF5). In combination, our results reveal distinct transcriptomes on joint-resident MSCs from different biomechanical environments and show that 6-week joint off-loading leads to transcriptional changes in SF MSCs that may be beneficial for cartilage regeneration. Biomechanical factors should be certainly considered in the development of novel MSC-based therapies for OA

Theoretical Study on Superconductivity in Boron-Doped Diamond

We consider superconductivity in boron (B) doped diamond using a simplified model for the valence band of diamond. We treat the effects of substitutional disorder of B ions by the coherent potential approximation (CPA) and those of the attractive force between holes by the ladder approximation under the assumption of instantaneous interaction with the Debye cutoff. We thereby calculate the quasiparticle life time, the evolution of the single-particle spectra due to doping, and the effect of disorder on the superconducting critical temperature $T_c$. We in particular compare our results with those for supercell calculations to see the role of disorder, which turns out to be of crucial importance to $T_c$.Comment: 9 pages, 13 figures, submitted to J. Phys. Soc. Jpn., Errors in embedded eps figure files have been correcte

A new class of glycomimetic drugs to prevent free fatty acid-induced endothelial dysfunction

Background: Carbohydrates play a major role in cell signaling in many biological processes. We have developed a set of glycomimetic drugs that mimic the structure of carbohydrates and represent a novel source of therapeutics for endothelial dysfunction, a key initiating factor in cardiovascular complications. Purpose: Our objective was to determine the protective effects of small molecule glycomimetics against free fatty acid­induced endothelial dysfunction, focusing on nitric oxide (NO) and oxidative stress pathways. Methods: Four glycomimetics were synthesized by the stepwise transformation of 2,5­dihydroxybenzoic acid to a range of 2,5­substituted benzoic acid derivatives, incorporating the key sulfate groups to mimic the interactions of heparan sulfate. Endothelial function was assessed using acetylcholine­induced, endotheliumdependent relaxation in mouse thoracic aortic rings using wire myography. Human umbilical vein endothelial cell (HUVEC) behavior was evaluated in the presence or absence of the free fatty acid, palmitate, with or without glycomimetics (1µM). DAF­2 and H2DCF­DA assays were used to determine nitric oxide (NO) and reactive oxygen species (ROS) production, respectively. Lipid peroxidation colorimetric and antioxidant enzyme activity assays were also carried out. RT­PCR and western blotting were utilized to measure Akt, eNOS, Nrf­2, NQO­1 and HO­1 expression. Results: Ex vivo endothelium­dependent relaxation was significantly improved by the glycomimetics under palmitate­induced oxidative stress. In vitro studies showed that the glycomimetics protected HUVECs against the palmitate­induced oxidative stress and enhanced NO production. We demonstrate that the protective effects of pre­incubation with glycomimetics occurred via upregulation of Akt/eNOS signaling, activation of the Nrf2/ARE pathway, and suppression of ROS­induced lipid peroxidation. Conclusion: We have developed a novel set of small molecule glycomimetics that protect against free fatty acidinduced endothelial dysfunction and thus, represent a new category of therapeutic drugs to target endothelial damage, the first line of defense against cardiovascular disease

Runoff Changes in the Šumava/Bohemian Forest Region

RUNOFF CHANGES IN THE ŠUMAVA / BOHEMIAN FOREST REGION Abstract: The goal of this thesis is to the evaluation of possible causes of changes in runoff regime in the Šumava region from time and spacial point of view. The thesis includes research and applied part. The research part is dedicated to methods of evaluation of runoff changes and their possible causes in teh Šumava region such as anthropogenic factors (changes in the river network, dams, drainage, land - use changes), natural factors (climate changes, peatbogs influence) and disturbances (wind calamities and bark beetle outbreaks, floods). In the applied part there is an analysis of precipitaion - runoff regime for long-term time series of average annual, monthly and minimal daily discharges and monthly precipitations for selected gauging stations in upper Otava, Ostružná, Volyňka, upper Blanice and Teplá Vltava basins using simple and double mass curves and Mann - Kendall test. In conclusion the achieved results were evaluated, discussed and compared with subject publications. Key words: disturbance, Mann - Kendall test, runoff, discharge, mass curves, precipitation, Šumava, change

Usefulness of Mesenchymal Cell Lines for Bone and Cartilage Regeneration Research

The unavailability of sufficient numbers of human primary cells is a major roadblock for in vitro repair of bone and/or cartilage, and for performing disease modelling experiments. Immortalized mesenchymal stromal cells (iMSCs) may be employed as a research tool for avoiding these problems. The purpose of this review was to revise the available literature on the characteristics of the iMSC lines, paying special attention to the maintenance of the phenotype of the primary cells from which they were derived, and whether they are effectively useful for in vitro disease modeling and cell therapy purposes. This review was performed by searching on Web of Science, Scopus, and PubMed databases from 1 January 2015 to 30 September 2019. The keywords used were ALL = (mesenchymal AND ("cell line" OR immortal*) AND (cartilage OR chondrogenesis OR bone OR osteogenesis) AND human). Only original research studies in which a human iMSC line was employed for osteogenesis or chondrogenesis experiments were included. After describing the success of the immortalization protocol, we focused on the iMSCs maintenance of the parental phenotype and multipotency. According to the literature revised, it seems that the maintenance of these characteristics is not guaranteed by immortalization, and that careful selection and validation of clones with particular characteristics is necessary for taking advantage of the full potential of iMSC to be employed in bone and cartilage-related research.status: publishe

Repeated antifungal use audits are essential for selecting the targets for intervention in antifungal stewardship

A previous audit to assess the quality of antifungal use was performed in our hospital in 2011. After 5&nbsp;years of antifungal stewardship program (AFS), we performed a follow-up audit in order to describe the long-term effect of such program. Using a predefined score, we evaluated the antifungal use in 100 consecutive adult inpatients receiving systemic antifungals. Results of the present audit were compared with those of a previous one, performed in 2011, before the implementation of our AFS. After 5&nbsp;years, AFS program has induced a change in the population who received antifungal drugs in our hospital with a reduction in medical patients and a relative higher prescription among hematological ones. As for indications, empirical use decreased very significantly (from 62 to 30%, p &lt; 0.001), while tailored treatment (from 20 to 41%, p = 0.001) and prophylaxis (from 15 to 27%, p = 0.03) increased. Compared to 2011, we also observed an improvement in the optimal choice of antifungal drug, route of administration, and microbiological adjustment. However, no significant improvement was observed regarding adequacy of length of therapy or optimal dosage or administration route. Although we observed an increase in the number of optimal DOTs used, the potential estimated savings continued to be high (~ €44,199 for every 100 patients receiving antifungals). Our study is the first to show the impact on the use of antifungal drugs exerted by a prolonged non-coercive AFS program. We also demonstrate the utility of a periodic audit of antifungal use in order to point out new goals for future interventions