Media matters: culture medium-dependent hypervariable phenotype of mesenchymal stromal cells.

Background: Despite a long history of investigation and sustained efforts in clinical testing, the number of market authorisations for mesenchymal stromal cell (MSC) therapies remains limited, with none approved by the United States Food and Drug Administration. Several barriers are impeding the clinical progression of MSC therapies, to the forefront of these is a lack of standardised manufacturing protocols which is further compounded by an absence of biologically meaningful characterisation and release assays. A look at clinical trial registries demonstrates the diversity of MSC expansion protocols with variabilities in cell source, isolation method and expansion medium, among other culture variables, making it extraordinarily difficult to compare study outcomes.

Xeno-free cultured mesenchymal stromal cells release extracellular vesicles with a "therapeutic" miRNA cargo ameliorating cartilage inflammation .

Mesenchymal stromal cells (MSCs)-derived extracellular vesicles (EVs) emerged as an innovative strategy for the treatment of chronic disorders such as osteoarthritis (OA). Biological activity of EVs is generally driven by their cargo, which might be influenced by microenvironment. Therefore, pre-conditioning strategies, including modifications in culture conditions or oxygen tension could directly impact on MSCs paracrine activity.

Needle to needle robot-assisted manufacture of cell therapy products.

Advanced therapeutic medicinal products (ATMPs) have emerged as novel therapies for untreatable diseases, generating the need for large volumes of high-quality, clinically-compliant GMP cells to replace costly, high-risk and limited scale manual expansion processes. We present the design of a fully automated, robot-assisted platform incorporating the use of multiliter stirred tank bioreactors for scalable production of adherent human stem cells. The design addresses a needle-to-needle closed process incorporating automated bone marrow collection, cell isolation, expansion, and collection into cryovials for patient delivery.

Regenerative and Anti-Inflammatory Potential of Regularly Fed, Starved Cells and Extracellular Vesicles In Vivo.

Background: Mesenchymal stem/stromal cells (MSC) have been employed successfully in immunotherapy and regenerative medicine, but their therapeutic potential is reduced considerably by the ischemic environment that exists after transplantation. The assumption that preconditioning MSC to promote quiescence may result in increased survival and regenerative potential upon transplantation is gaining popularity.

Methods: The purpose of this work was to evaluate the anti-inflammatory and regenerative effects of human bone marrow MSC (hBM-MSC) and their extracellular vesicles (EVs) grown and isolated in a serum-free medium, as compared to starved hBM-MSC (preconditioned) in streptozotocin-induced diabetic fractured male C57BL/6J mice.

Characterizing the secretome of licensed hiPSC-derived MSCs.

Although mesenchymal stromal cells (MSCs) from primary tissues have been successfully applied in the clinic, their expansion capabilities are limited and results are variable. MSCs derived from human-induced pluripotent stem cells (hiMSCs) are expected to overcome these limitations and serve as a reproducible and sustainable cell source. We have explored characteristics and therapeutic potential of hiMSCs in comparison to hBMSCs.

Manufacturing Mesenchymal Stromal Cells for the Treatment of Osteoarthritis in Canine Patients: Challenges and Recommendations.

The recent interest in advanced biologic therapies in veterinary medicine has opened up opportunities for new treatment modalities with considerable clinical potential. Studies with mesenchymal stromal cells (MSCs) from animal species have focused on characterization (mostly following protocols developed for human application), experimental testing in controlled studies and clinical use in veterinary patients. The ability of MSCs to interact with the inflammatory environment through immunomodulatory and paracrine mechanisms makes them a good candidate for treatment of inflammatory musculoskeletal conditions in canine species.

GMP-Compliant Production of Autologous Adipose-Derived Stromal Cells in the NANT 001 Closed Automated Bioreactor.

In recent years mesenchymal stromal cells (MSCs) have received a great deal of interest for the treatment of major diseases, but clinical translation and market authorization have been slow. This has been due in part to a lack of standardization in cell manufacturing protocols, as well as a lack of biologically meaningful cell characterization tools and release assays. Cell production strategies to date have involved complex manual processing in an open environment which is costly, inefficient and poses risks of contamination.

Accessing depth-resolved high spatial frequency content from the optical coherence tomography signal.

Optical coherence tomography (OCT) is a rapidly evolving technology with a broad range of applications, including biomedical imaging and diagnosis. Conventional intensity-based OCT provides depth-resolved imaging with a typical resolution and sensitivity to structural alterations of about 5-10 microns. It would be desirable for functional biological imaging to detect smaller features in tissues due to the nature of pathological processes.

Subconjunctival administration of low-dose murine allogeneic mesenchymal stromal cells promotes corneal allograft survival in mice.

Background: Systemic administration of mesenchymal stromal cells (MSCs) has been efficacious in many inflammatory disease settings; however, little data are available on the potential immunomodulatory effects following local MSC administration in the context of corneal transplantation. The purpose of this study was to assess the potential of subconjunctival injection of MSCs to promote corneal allograft survival.

Methods: MSCs were isolated from female C57BL/6 (H-2) or Balb/c (H-2) mice and extensively characterized.

The Secretome Derived From Mesenchymal Stromal Cells Cultured in a Xeno-Free Medium Promotes Human Cartilage Recovery .

Osteoarthritis (OA) is a disabling joint disorder causing articular cartilage degeneration. Currently, the treatments are mainly aimed to pain and symptoms relief, rather than disease amelioration. Human bone marrow stromal cells (hBMSCs) have emerged as a promising paracrine mechanism-based tool for OA treatment.

Survival/Adaptation of Bone Marrow-Derived Mesenchymal Stem Cells After Long-Term Starvation Through Selective Processes.

After in vivo transplantation, mesenchymal stem cells (MSC) face an ischemic microenvironment, characterized by nutrient deprivation and reduced oxygen tension, which reduces their viability and thus their therapeutic potential. Therefore, MSC response to models of in vitro ischemia is of relevance for improving their survival and therapeutic efficacy. The aim of this study was to understand the survival/adaptive response mechanism that MSC use to respond to extreme culture conditions.

Induced Pluripotent Stem Cell-Derived Mesenchymal Stromal Cells Are Functionally and Genetically Different From Bone Marrow-Derived Mesenchymal Stromal Cells.

There has been considerable interest in the generation of functional mesenchymal stromal cell (MSC) preparations from induced pluripotent stem cells (iPSCs) and this is now regarded as a potential source of unlimited, standardized, high-quality cells for therapeutic applications in regenerative medicine. Although iMSCs meet minimal criteria for defining MSCs in terms of marker expression, there are substantial differences in terms of trilineage potential, specifically a marked reduction in chondrogenic and adipogenic propensity in iMSCs compared with bone marrow-derived (BM) MSCs. To reveal the cellular basis underlying these differences, we conducted phenotypic, functional, and genetic comparisons between iMSCs and BM-MSCs.

Manufacturing Differences Affect Human Bone Marrow Stromal Cell Characteristics and Function: Comparison of Production Methods and Products from Multiple Centers.

Human bone marrow stromal cells (BMSCs, also known as bone marrow-derived mesenchymal stem cells) are manufactured using many different methods, but little is known about the spectrum of manufacturing methods used and their effects on BMSC characteristics and function. Seven centers using, and one developing, Good Manufacturing Practices (GMP) processes were surveyed as to their production methods. Among the seven centers, all used marrow aspirates as the starting material, but no two centers used the same manufacturing methods.

Chondrocytes Derived From Mesenchymal Stromal Cells and Induced Pluripotent Cells of Patients With Familial Osteochondritis Dissecans Exhibit an Endoplasmic Reticulum Stress Response and Defective Matrix Assembly.

Unlabelled: : Familial osteochondritis dissecans (FOCD) is an inherited skeletal defect characterized by the development of large cartilage lesions in multiple joints, short stature, and early onset of severe osteoarthritis. It is associated with a heterozygous mutation in the ACAN gene, resulting in a Val-Met replacement in the C-type lectin domain of aggrecan. To understand the cellular pathogenesis of this condition, we studied the chondrogenic differentiation of patient bone marrow mesenchymal stromal cells (BM-MSCs).

Evaluation of the Early In Vivo Response of a Functionally Graded Macroporous Scaffold in an Osteochondral Defect in a Rabbit Model.

Cartilage tissue engineering is a multifactorial problem requiring a wide range of material property requirements from provision of biological cues to facilitation of mechanical support in load-bearing diarthrodial joints. The study aim was to design, fabricate and characterize a template to promote endogenous cell recruitment for enhanced cartilage repair. A polylactic acid poly-ε-caprolactone (PLCL) support structure was fabricated using laser micromachining technology and thermal crimping to create a functionally-graded open pore network scaffold with a compressive modulus of 9.

Thermoresponsive Substrates Used for the Growth and Controlled Differentiation of Human Mesenchymal Stem Cells.

This communication outlines the advances made in the development of thermoresponsive substrates for human mesenchymal stem cell (hMSC) expansion and subsequent controlled specific and multilineage differentiation from a previous study performed by this group. Previously, the development of an inexpensive and technically accessible method for hMSC expansion and harvesting was reported, using the solvent casting deposition method and thermoresponsive poly(N-isopropylacrylamide). Here, the logical continuation of this work is reported with the multipassage expansion of hMSCs with phenotypic maintenance followed by induced adipogenic, osteogenic, and chondrogenic differentiation.

ROCK activity and the Gβγ complex mediate chemotactic migration of mouse bone marrow-derived stromal cells.

Introduction: Bone marrow-derived stromal cells (BMSCs), also known as mesenchymal stem cells, are the focus of intensive efforts worldwide to elucidate their function and biology. Despite the importance of BMSC migration for their potential therapeutic uses, the mechanisms and signalling governing stem cell migration are still not fully elucidated.

Methods: We investigated and detailed the effects of MCP-1 activation on BMSCs by using inhibitors of G protein-coupled receptor alpha beta (GPCR αβ), ROCK (Rho-associated, coiled-coil containing protein kinase), and PI3 kinase (PI3K).

A novel collagen-nanohydroxyapatite microRNA-activated scaffold for tissue engineering applications capable of efficient delivery of both miR-mimics and antagomiRs to human mesenchymal stem cells.

Manipulation of gene expression through the use of microRNAs (miRNAs) offers tremendous potential for the field of tissue engineering. However, the lack of sufficient site-specific and bioactive delivery systems has severely hampered the clinical translation of miRNA-based therapies. In this study, we developed a novel non-viral bioactive delivery platform for miRNA mimics and antagomiRs to allow for a vast range of therapeutic applications.

TNFα and IL-1β influence the differentiation and migration of murine MSCs independently of the NF-κB pathway.

Introduction: Mesenchymal stem cells (MSCs) have the ability to repair and regenerate tissue, home to sites of inflammation, and evade the host immune system. As such, they represent an attractive therapy for the treatment of autoimmune inflammatory diseases. However, results from in vivo murine studies in inflammatory arthritis have been conflicting, and this may be due to the genetic background of the MSCs used.

Basic fibroblast growth factor modifies the hypoxic response of human bone marrow stromal cells by ERK-mediated enhancement of HIF-1α activity.

Human bone marrow stromal cells (hBMSCs, also known as bone marrow-derived mesenchymal stem cells) are promising tools for the cellular therapy of human pathologies related to various forms of hypoxia. Although the current concepts of their clinical use include the expansion of hBMSC in standard cell culture conditions, the effect of the mitogen-driven ex vivo expansion on the adaptation to the hypoxic environment is unknown. Here, we provide data that the basic fibroblast growth factor (FGF2) enhances the induction of a wide range of hypoxia-related adaptive genes in hypoxic hBMSCs.

Chondrogenic differentiation increases antidonor immune response to allogeneic mesenchymal stem cell transplantation.

Allogeneic mesenchymal stem cells (allo-MSCs) have potent regenerative and immunosuppressive potential and are being investigated as a therapy for osteoarthritis; however, little is known about the immunological changes that occur in allo-MSCs after ex vivo induced or in vivo differentiation. Three-dimensional chondrogenic differentiation was induced in an alginate matrix, which served to immobilize and potentially protect MSCs at the site of implantation. We show that allogeneic differentiated MSCs lost the ability to inhibit T-cell proliferation in vitro, in association with reduced nitric oxide and prostaglandin E2 secretion.

Allogeneic murine mesenchymal stem cells: migration to inflamed joints in vivo and amelioration of collagen induced arthritis when transduced to express CTLA4Ig.

Despite the immunosuppressive, homing, and regenerative capabilities of mesenchymal stem cells (MSCs), their ability to migrate to arthritic joints and influence the course of arthritis in vivo remains poorly understood. The objective of this study was to determine if allogeneic MSCs migrate to inflamed joints in vivo and to determine if MSCs expressing the costimulation blocker cytotoxic T lymphocyte associated antigen-4 coupled to immunoglobulin-G (CTLA4Ig) could be used to ameliorate collagen induced arthritis (CIA). The migration of systemically delivered inbred mouse strain (FVB) MSCs to migrate to inflamed joints in CIA was studied using real-time quantitative polymerase chain reaction.

Topical administration of allogeneic mesenchymal stromal cells seeded in a collagen scaffold augments wound healing and increases angiogenesis in the diabetic rabbit ulcer.

There is a critical clinical need to develop therapies for nonhealing diabetic foot ulcers. Topically applied mesenchymal stromal cells (MSCs) provide a novel treatment to augment diabetic wound healing. A central pathological factor in nonhealing diabetic ulcers is an impaired blood supply.