Patient-Specific iPSC-Derived Models Link Aberrant Endoplasmic Reticulum Stress Sensing and Response to Juvenile Osteochondritis Dissecans Etiology.

Juvenile osteochondritis dissecans (JOCD) is a pediatric disease, which begins with an osteonecrotic lesion in the secondary ossification center which, over time, results in the separation of the necrotic fragment from the parent bone. JOCD predisposes to early-onset osteoarthritis. However, the knowledge gap in JOCD pathomechanisms severely limits current therapeutic strategies. To elucidate its etiology, we conducted a study with induced pluripotent stem cells (iPSCs) from JOCD and control patients. iPSCs from skin biopsies were differentiated to iMSCs (iPSC-derived mesenchymal stromal cells) and subjected to chondrogenic and endochondral ossification, and endoplasmic reticulum (ER)-stress induction assays. Our study, using 3 JOCD donors, showed that JOCD cells have lower chondrogenic capability and their endochondral ossification process differs from control cells; yet, JOCD- and control-cells accomplish osteogenesis of similar quality. Our findings show that endoplasmic reticulum stress sensing and response mechanisms in JOCD cells, which partially regulate chondrocyte and osteoblast differentiation, are related to these differences. We suggest that JOCD cells are more sensitive to ER stress than control cells, and in pathological microenvironments, such as microtrauma and micro-ischemia, JOCD pathogenesis pathways may be initiated. This study is the first, to the best of our knowledge, to realize the important role that resident cells and their differentiating counterparts play in JOCD and to put forth a novel etiological hypothesis that seeks to consolidate and explain previously postulated hypotheses. Furthermore, our results establish well-characterized JOCD-specific iPSC-derived in vitro models and identified potential targets which could be used to improve diagnostic tools and therapeutic strategies in JOCD.

Glucose Metabolism: Optimizing Regenerative Functionalities of Mesenchymal Stromal Cells Postimplantation.

Mesenchymal stromal cells (MSCs) are considered promising candidates for regenerative medicine applications. Their clinical performance postimplantation, however, has been disappointing. This lack of therapeutic efficacy is most likely due to suboptimal formulations of MSC-containing material constructs. Tissue engineers, therefore, have developed strategies addressing/incorporating optimized cell, microenvironmental, biochemical, and biophysical cues/stimuli to enhance MSC-containing construct performance. Such approaches have had limited success because they overlooked that maintenance of MSC viability after implantation for a sufficient time is necessary for MSCs to develop their regenerative functionalities fully. Following a brief overview of glucose metabolism and regulation in MSCs, the present literature review includes recent pertinent findings that challenge old paradigms and notions. We hereby report that glucose is the primary energy substrate for MSCs, provides precursors for biomass generation, and regulates MSC functions, including proliferation and immunosuppressive properties. More importantly, glucose metabolism is central in controlling in vitro MSC expansion, in vivo MSC viability, and MSC-mediated angiogenesis postimplantation when addressing MSC-based therapies. Meanwhile, in silico models are highlighted for predicting the glucose needs of MSCs in specific regenerative medicine settings, which will eventually enable tissue engineers to design viable and potent tissue constructs. This new knowledge should be incorporated into developing novel effective MSC-based therapies. Impact statement The clinical use of mesenchymal stromal cells (MSCs) has been unsatisfactory due to the inability of MSCs to survive and be functional after implantation for sufficient periods to mediate directly or indirectly a successful regenerative tissue response. The present review summarizes the endeavors in the past, but, most importantly, reports the latest findings that elucidate underlying mechanisms and identify glucose metabolism as the crucial parameter in MSC survival and the subsequent functions pertinent to new tissue formation of importance in tissue regeneration applications. These latest findings justify further basic research and the impetus for developing new strategies to improve the modalities and efficacy of MSC-based therapies.

Reduced Size Profile of Amniotic Membrane Particles Decreases Osteoarthritis Therapeutic Efficacy.

Osteoarthritis (OA) is a widespread disease that continues to lack approved and efficacious treatments that modify disease progression. Micronized dehydrated human amnion/chorion membrane (µ-dHACM) has been shown to be effective in reducing OA progression, but many of the engineering design parameters have not been explored. The objectives of this study were to characterize the particle size distributions of two µ-dHACM formulations and to investigate the influence of these distributions on the in vivo therapeutic efficacy of µ-dHACM. Male Lewis rats underwent medial meniscus transection (MMT) or sham surgery, and intra-articular injections of saline, µ-dHACM, or reduced particle size µ-dHACM (RPS µ-dHACM) were administered at 24 hours postsurgery (n = 9 per treatment group). After 3 weeks, the animals were euthanized, and left legs harvested for equilibrium partitioning of an ionic contrast agent microcomputed tomography and histological analysis. µ-dHACM and RPS µ-dHACM particles were fluorescently tagged and particle clearance was tracked in vivo for up to 42 days postsurgery. Protein elution from both formulations was quantified in vitro. Treatment with µ-HACM, but not RPS µ-dHACM, reduced lesion volume in the MMT model 3 weeks postsurgery. In contrast, RPS µ-dHACM increased cartilage surface roughness and osteophyte cartilage thickness and volume compared to saline treatment. There was no difference of in vivo fluorescently tagged particle clearance between the two µ-dHACM sizes. RPS µ-dHACM showed significantly greater protein elution in vitro over 21 days. Overall, delivery of RPS µ-dHACM did result in an increase of in vivo joint degeneration and in vitro protein elution compared to µ-dHACM, but did not result in differences in joint clearance in vivo. These results suggest that particle size and factor elution may be tailorable factors that are important to optimize for particulate amniotic membrane treatment to be an effective therapy for OA. Impact Statement Osteoarthritis (OA) is a widespread disease that continues to lack treatments that modify the progression of the disease. Micronized dehydrated human amnion/chorion membrane (µ-dHACM) has been shown to be effective in reducing OA progression, but many of the engineering design parameters have not been explored. This work investigates the effects of particle size profile of the µ-dHACM particles and lays out the methods used in these studies. The results of this work will guide engineers in designing µ-dHACM treatments specifically and disease-modifying OA therapeutics generally, and it demonstrates the utility of novel therapeutic evaluation methods such as contrast-enhanced microcomputed tomography.

Understanding and leveraging cell metabolism to enhance mesenchymal stem cell transplantation survival in tissue engineering and regenerative medicine applications.

In tissue engineering and regenerative medicine, stem cell-specifically, mesenchymal stromal/stem cells (MSCs)-therapies have fallen short of their initial promise and hype. The observed marginal, to no benefit, success in several applications has been attributed primarily to poor cell survival and engraftment at transplantation sites. MSCs have a metabolism that is flexible enough to enable them to fulfill their various cellular functions and remarkably sensitive to different cellular and environmental cues. At the transplantation sites, MSCs experience hostile environments devoid or, at the very least, severely depleted of oxygen and nutrients. The impact of this particular setting on MSC metabolism ultimately affects their survival and function. In order to develop the next generation of cell-delivery materials and methods, scientists must have a better understanding of the metabolic switches MSCs experience upon transplantation. By designing treatment strategies with cell metabolism in mind, scientists may improve survival and the overall therapeutic potential of MSCs. Here, we provide a comprehensive review of plausible metabolic switches in response to implantation and of the various strategies currently used to leverage MSC metabolism to improve stem cell-based therapeutics.

Application of biomaterials to in vitro pluripotent stem cell disease modeling of the skeletal system.

Disease-specific pluripotent stem cells can be derived through genetic manipulation of embryonic stem cells or by reprogramming somatic cells (induced pluripotent stem cells). These cells are a valuable tool to study human diseases in vitro in order to dissect their pathomechanisms and develop novel therapeutics. Although pluripotent stem cell-derived models have successfully recapitulated the abnormalities of some skeletal diseases in vitro, this field is still at its early stages, and it could greatly benefit from the direct application of biomaterial research. Biomaterial-based systems may be utilized to enhance the differentiation processes of pluripotent stem cells in order to create more homogeneous and physiologically relevant in vitro disease models. Moreover, inducing the disease phenotype may be facilitated by the guidance of biomaterials. This review presents a comprehensive summary of existing biomaterial applications in human disease modeling and their potential on skeletal disease models. By utilizing disease-specific pluripotent stem cells, current biomaterial-based systems for in vitro models could be extrapolated to study skeletal diseases in a petri dish.

EPIC-µCT imaging of articular cartilage.

Characterization of articular cartilage morphology and composition using microcomputed tomography (microCT) techniques requires the use of contrast agents to enhance X-ray attenuation of the tissue. This chapter describes the use of an anionic iodinated contrast agent at equilibrium with articular cartilage. In this technique, negatively charged contrast agent molecules distribute themselves inversely with respect to the negatively charged proteoglycans (PGs) within the cartilage tissue (Palmer et al. Proc Natl Acad Sci U S A 103:19255-19260, 2006). This relationship allows for assessment of cartilage degradation, as areas of high X-ray attenuation have been shown to correspond to areas of depleted PGs (Palmer et al. Proc Natl Acad Sci U S A 103:19255-19260, 2006; Xie et al. Osteoarthritis Cartilage 18:65-72, 2010).