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Eradicating leukemia requires a deep understanding of the interaction between leukemic cells and their protective microenvironment. The CXCL12/CXCR4 axis has been postulated as a critical pathway dictating leukemia stem cell (LSC) chemoresistance in AML due to its role in controlling cellular egress from the marrow. Nevertheless, the cellular source of CXCL12 in the acute myeloid leukemia (AML) microenvironment and the mechanism by which CXCL12 exerts its protective role in vivo remain unresolved. Here, we show that CXCL12 produced by Prx1+ mesenchymal cells but not by mature osteolineage cells provide the necessary cues for the maintenance of LSCs in the marrow of an MLL::AF9-induced AML model. Prx1+ cells promote survival of LSCs by modulating energy metabolism and the REDOX balance in LSCs. Deletion of Cxcl12 leads to the accumulation of reactive oxygen species and DNA damage in LSCs, impairing their ability to perpetuate leukemia in transplantation experiments, a defect that can be attenuated by antioxidant therapy. Importantly, our data suggest that this phenomenon appears to be conserved in human patients. Hence, we have identified Prx1+ mesenchymal cells as an integral part of the complex niche-AML metabolic intertwining, pointing towards CXCL12/CXCR4 as a target to eradicate parenchymal LSCs in AML., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Despite the bone marrow microenvironment being widely recognised as a key player in cancer research, the current animal models that represent a human haematopoietic system lack the contribution of the humanised marrow microenvironment. Here we describe a murine model that relies on the combination of an orthotopic humanised tissue-engineered bone construct (ohTEBC) with patient-specific bone marrow (BM) cells to create a humanised bone marrow (hBM) niche capable of supporting the engraftment of human haematopoietic cells. Results showed that this model supports the engraftment of human CD34 + cells from a healthy BM with human haematopoietic cells migrating into the mouse BM, human BM compartment, spleen and peripheral blood. We compared these results with the engraftment capacity of human CD34 + cells obtained from patients with multiple myeloma (MM). We demonstrated that CD34 + cells derived from a diseased BM had a reduced engraftment potential compared to healthy patients and that a higher cell dose is required to achieve engraftment of human haematopoietic cells in peripheral blood. Finally, we observed that hematopoietic cells obtained from the mobilised peripheral blood of patients yields a higher number of CD34 + , overcoming this problem. In conclusion, this humanised mouse model has potential as a unique and patient-specific pre-clinical platform for the study of tumour-microenvironment interactions, including human bone and haematopoietic cells, and could, in the future, serve as a drug testing platform.

In this study, we examined the effectiveness of systemic subcutaneous delivery of recombinant Insulin-like growth factor (IGF)-I concurrently with primary cultured bone marrow-derived mesenchymal stem cell (MSC) transplant on fracture repair. We found that the fracture callus volume increased in mice with a stabilized tibia fracture that received IGF-I+MSC when compared with that in either untreated or MSC alone treated mice. In evaluating the callus tissue components, we found that the soft and new bone tissue volumes were significantly increased in IGF-I+MSC recipients. Histological and in-situ hybridization analyses confirmed a characteristic increase of newly forming bone in IGF-I+MSC recipients and that healing progressed mostly through endochondral ossification. The increase in soft and new bone tissue volumes correlated with increased force and toughness as determined by biomechanical testing. In conclusion, MSC transplant concurrent with systemic delivery of IGF-I improves fracture repair suggesting that IGF-I+MSC could be a novel therapeutic approach in patients who have inadequate fracture repair.

Importance of the Field: Mesenchymal stem cells have the ability to differentiate into osteoblasts, chondrocytes and adipocytes. Along with differentiation, MSCs can modulate inflammation, home to damaged tissues and secrete bioactive molecules. These properties can be enhanced through genetic-modification that would combine the best of both cell and gene therapy fields to treat monogenic and multigenic diseases., Areas Covered in This Review: Findings demonstrating the immunomodulation, homing and paracrine activities of MSCs followed by a summary of the current research utilizing MSCs as a vector for gene therapy, focusing on skeletal disorders, but also cardiovascular disease, ischemic damage and cancer., What the Reader Will Gain: MSCs are a possible therapy for many diseases, especially those related to the musculoskeletal system, as a standalone treatment, or in combination with factors that enhance the abilities of these cells to migrate, survive or promote healing through anti-inflammatory and immunomodulatory effects, differentiation, angiogenesis or delivery of cytolytic or anabolic agents., Take Home Message: Genetically-modified MSCs are a promising area of research that would be improved by focusing on the biology of MSCs that could lead to identification of the natural and engrafting MSC-niche and a consensus on how to isolate and expand MSCs for therapeutic purposes.

Murine embryonic limb cultures have invaluable roles in studying skeletogenesis. Substance delivery is an underdeveloped area in developmental biology that has primarily relied on Affi-Gel-Blue-agarose-beads. However, the lack of information about the efficiency of agarose-bead loading and release and difficulties for a single-bead implantation represent significant limitations. We optimized the use of glycol chitosan-5beta-cholanic acid conjugates (HGC) as a novel protein delivery system in mouse embryonic limbs. To this purpose, we loaded HGC either with recombinant Noggin, or bovine serum albumin (BSA). The size, morphology and stability of the protein-loaded-HGC were determined by transmission electron microscopy and dynamic-light-scattering. HGC-BSA and HGC-Noggin loading efficiencies were 80-90%. Time-course study revealed that Noggin and BSA were 80-90% released after 48 h. We developed several techniques to implant protein-loaded-HGC into murine embryonic joints from embryonic age E13.5 to E15.5, including a micro-injection system dispensing nanoliters. HGC did not interfere with skeletogenesis. Using CBR-3BA staining, we detected HGC nanoparticles within implanted tissues. Furthermore, a sustained release of BSA and Noggin was demonstrated in HGC-BSA and HGC-Noggin injected regions. HGC-released Noggin was biologically active in blocking the BMP signaling in in vitro mesenchyme limb micromasses as well as in ex-vivo limb cultures. Results reveal that HGC is a valuable protein-delivery system in developmental biology., (Copyright 2010 Elsevier B.V. All rights reserved.)

Background: Mesenchymal stem cells (MSC) are multipotent cells with the ability to differentiate into mesenchyme-derived cells including osteoblasts and chondrocytes., Objective: To provide an overview and expert opinion on the in vivo ability of MSC to home into tissues, their regenerative properties and potential applications for cell-based therapies to treat bone and cartilage disorders., Methods: Data sources including the PubMed database, abstract booklets and conference proceedings were searched for publications pertinent to MSC and their properties with emphasis on the in vivo studies and clinical use in cartilage and bone regeneration and repair. The search included the most current information possible., Conclusion: MSC can migrate to injured tissues and some of their reparative properties are mediated by paracrine mechanisms including their immunomodulatory actions. MSC possess a critical potential in regenerative medicine for the treatment of skeletal diseases, such as osteoarthritis or fracture healing failure, where treatments are partially effective or palliative.

Despite its clinical significance, joint morphogenesis is still an obscure process. In this study, we determine the role of transforming growth factor beta (TGF-beta) signaling in mice lacking the TGF-beta type II receptor gene (Tgfbr2) in their limbs (Tgfbr2(PRX-1KO)). In Tgfbr2(PRX-1KO) mice, the loss of TGF-beta responsiveness resulted in the absence of interphalangeal joints. The Tgfbr2(Prx1KO) joint phenotype is similar to that in patients with symphalangism (SYM1-OMIM185800). By generating a Tgfbr2-green fluorescent protein-beta-GEO-bacterial artificial chromosome beta-galactosidase reporter transgenic mouse and by in situ hybridization and immunofluorescence, we determined that Tgfbr2 is highly and specifically expressed in developing joints. We demonstrated that in Tgfbr2(PRX-1KO) mice, the failure of joint interzone development resulted from an aberrant persistence of differentiated chondrocytes and failure of Jagged-1 expression. We found that TGF-beta receptor II signaling regulates Noggin, Wnt9a, and growth and differentiation factor-5 joint morphogenic gene expressions. In Tgfbr2(PRX-1KO) growth plates adjacent to interphalangeal joints, Indian hedgehog expression is increased, whereas Collagen 10 expression decreased. We propose a model for joint development in which TGF-beta signaling represents a means of entry to initiate the process.