IL-17A expressed on neutrophil extracellular traps promotes mesenchymal stem cell differentiation toward bone-forming cells in ankylosing spondylitis.

Ankylosing spondylitis (AS) is an inflammatory disease characterized by excessive bone formation. We investigated the presence of neutrophil extracellular traps (NETs) in AS and how they are involved in the osteogenic capacity of bone marrow mesenchymal stem cells (MSCs) through interleukin-17A (IL-17A). Peripheral neutrophils and sera were obtained from patients with active AS and healthy controls. NET formation and neutrophil/NET-associated proteins were studied using immunofluorescence, immunoblotting, qPCR, and ELISA. In vitro co-culture systems of AS NET structures and MSCs isolated from controls were deployed to examine the role of NETs in the differentiation of MSCs toward osteogenic cells. Analysis was performed using specific staining and qPCR. Neutrophils from patients with AS were characterized by enhanced formation of NETs carrying bioactive IL-17A and IL-1ß. IL-17A-enriched AS NETs mediated the differentiation of MSCs toward bone-forming cells. The neutrophil expression of IL-17A was positively regulated by IL-1ß. Blocking IL-1ß signaling on neutrophils with anakinra or dismantling NETs using DNase-I disrupted osteogenesis driven by IL-17A-bearing NETs. These findings propose a novel role of neutrophils in AS-related inflammation, linking IL-17A-decorated NETs with the differentiation of MSCs toward bone-forming cells. Moreover, IL-1ß triggers the expression of IL-17A on NETs offering an additional therapeutic target in AS.

The Role of Bone Marrow Mesenchymal Stem Cell Derived Extracellular Vesicles (MSC-EVs) in Normal and Abnormal Hematopoiesis and Their Therapeutic Potential.

Mesenchymal stem cells (MSCs) represent a heterogeneous cellular population responsible for the support, maintenance, and regulation of normal hematopoietic stem cells (HSCs). In many hematological malignancies, however, MSCs are deregulated and may create an inhibitory microenvironment able to induce the disease initiation and/or progression. MSCs secrete soluble factors including extracellular vesicles (EVs), which may influence the bone marrow (BM) microenvironment via paracrine mechanisms. MSC-derived EVs (MSC-EVs) may even mimic the effects of MSCs from which they originate. Therefore, MSC-EVs contribute to the BM homeostasis but may also display multiple roles in the induction and maintenance of abnormal hematopoiesis. Compared to MSCs, MSC-EVs have been considered a more promising tool for therapeutic purposes including the prevention and treatment of Graft Versus Host Disease (GVHD) following allogenic HSC transplantation (HSCT). There are, however, still unanswered questions such as the molecular and cellular mechanisms associated with the supportive effect of MSC-EVs, the impact of the isolation, purification, large-scale production, storage conditions, MSC source, and donor characteristics on MSC-EV biological effects as well as the optimal dose and safety for clinical usage. This review summarizes the role of MSC-EVs in normal and malignant hematopoiesis and their potential contribution in treating GVHD.

Differential expression of cell cycle and WNT pathway-related genes accounts for differences in the growth and differentiation potential of Wharton's jelly and bone marrow-derived mesenchymal stem cells.

BACKGROUND: In view of the current interest in exploring the clinical use of mesenchymal stem cells (MSCs) from different sources, we performed a side-by-side comparison of the biological properties of MSCs isolated from the Wharton's jelly (WJ), the most abundant MSC source in umbilical cord, with bone marrow (BM)-MSCs, the most extensively studied MSC population. METHODS: MSCs were isolated and expanded from BM aspirates of hematologically healthy donors (n = 18) and from the WJ of full-term neonates (n = 18). We evaluated, in parallel experiments, the MSC immunophenotypic, survival and senescence characteristics as well as their proliferative potential and cell cycle distribution. We also assessed the expression of genes associated with the WNT- and cell cycle-signaling pathway and we performed karyotypic analysis through passages to evaluate the MSC genomic stability. The hematopoiesis-supporting capacity of MSCs from both sources was investigated by evaluating the clonogenic cells in the non-adherent fraction of MSC co-cultures with BM or umbilical cord blood-derived CD34+ cells and by measuring the hematopoietic cytokines levels in MSC culture supernatants. Finally, we evaluated the ability of MSCs to differentiate into adipocytes and osteocytes and the effect of the WNT-associated molecules WISP-1 and sFRP4 on the differentiation potential of WJ-MSCs. RESULTS: Both ex vivo-expanded MSC populations showed similar morphologic, immunophenotypic, survival and senescence characteristics and acquired genomic alterations at low frequency during passages. WJ-MSCs exhibited higher proliferative potential, possibly due to upregulation of genes that stimulate cell proliferation along with downregulation of genes related to cell cycle inhibition. WJ-MSCs displayed inferior lineage priming and differentiation capacity toward osteocytes and adipocytes, compared to BM-MSCs. This finding was associated with differential expression of molecules related to WNT signaling, including WISP1 and sFRP4, the respective role of which in the differentiation potential of WJ-MSCs was specifically investigated. Interestingly, treatment of WJ-MSCs with recombinant human WISP1 or sFRP4 resulted in induction of osteogenesis and adipogenesis, respectively. WJ-MSCs exhibited inferior hematopoiesis-supporting potential probably due to reduced production of stromal cell-Derived Factor-1α, compared to BM-MSCs. CONCLUSIONS: Overall, these data are anticipated to contribute to the better characterization of WJ-MSCs and BM-MSCs for potential clinical applications.

Impaired proliferative potential of bone marrow mesenchymal stromal cells in patients with myelodysplastic syndromes is associated with abnormal WNT signaling pathway.

It has been shown that bone marrow mesenchymal stromal cells (MSCs) from patients with myelodysplastic syndromes (MDSs) display defective proliferative potential. We have probed the impaired replicative capacity of culture-expanded MSCs in MDS patients (n=30) compared with healthy subjects (n=32) by studying senescence characteristics and gene expression associated with WNT/transforming growth factor-ß1 (TGFB1) signaling pathways. We have also explored the consequences of the impaired patient MSC proliferative potential by investigating their differentiation potential and the capacity to support normal CD34(+) cell growth under coculture conditions. Patient MSCs displayed decreased gene expression of the senescence-associated cyclin-dependent kinase inhibitors CDKN1A, CDKN2A, and CDKN2B, along with PARG1, whereas the mean telomere length was upregulated in patient MSCs. MDS-derived MSCs exhibited impaired capacity to support normal CD34(+) myeloid and erythroid colony formation. No significant changes were observed between patients and controls in gene expression related to TGFB1 pathway. Patient MSCs displayed upregulated non-canonical WNT expression, combined with downregulated canonical WNT expression and upregulated canonical WNT inhibitors. MDS-derived MSCs displayed defective osteogenic and adipogenic lineage priming under non-differentiating culture conditions. Pharmacological activation of canonical WNT signaling in patient MDSs led to an increase in cell proliferation and upregulation in the expression of early osteogenesis-related genes. This study indicates abnormal WNT signaling in MSCs of MDS patients and supports the concept of a primary MSC defect that might have a contributory effect in MDS natural history.

Mesenchymal stem cells derived from Wharton's Jelly of the umbilical cord: biological properties and emerging clinical applications.

In recent years there seems to be an unbounded interest concerning mesenchymal stem cells (MSCs). This is mainly attributed to their exciting characteristics including long-term ex vivo proliferation, multilineage potential and immunomodulatory properties. In this regard MSCs emerge as attractive candidates for various therapeutic applications. MSCs were originally isolated from the bone marrow (BM) and this population is still considered as the gold standard for MSC applications. Nevertheless the BM has several limitations as source of MSCs, including MSC low frequency in this compartment, the painful isolation procedure and the decline in MSC characteristics with donor's age. Thus, there is accumulating interest in identifying alternative sources for MSCs. To this end MSCs obtained from the Wharton's Jelly (WJ) of umbilical cords (UC) have gained much attention over the last years since they can be easily isolated, without any ethical concerns, from a tissue which is discarded after birth. Furthermore WJ-derived MSCs represent a more primitive population than their adult counterparts, opening new perspectives for cell-based therapies. In this review we will at first give an overview of the biology of WJ-derived UC-MSCs. Then their potential application for the treatment of cancer and immune mediated disorders, such graft versus host disease (GVHD) and systemic lupus erythematosus (SLE) will be discussed, and finally their putative role as feeder layer for ex vivo hematopoietic stem cell (HSC) expansion will be pointed out.

Mesenchymal stem cells in immune-mediated bone marrow failure syndromes.

Immune-mediated bone marrow failure syndromes (BMFS) are characterized by ineffective marrow haemopoiesis and subsequent peripheral cytopenias. Ineffective haemopoiesis is the result of a complex marrow deregulation including genetic, epigenetic, and immune-mediated alterations in haemopoietic stem/progenitor cells, as well as abnormal haemopoietic-to-stromal cell interactions, with abnormal release of haemopoietic growth factors, chemokines, and inhibitors. Mesenchymal stem/stromal cells (MSCs) and their progeny (i.e., osteoblasts, adipocytes, and reticular cells) are considered as key cellular components of the bone marrow haemopoietic niche. MSCs may interfere with haemopoietic as well as immune regulation. Evidence suggests that bone marrow MSCs may be involved in immune-mediated BMFS underlying pathophysiology, harboring either native abnormalities and/or secondary defects, caused by exposure to activated marrow components. This review summarizes previous as well as more recent information related to the biologic/functional characteristics of bone marrow MSCs in myelodysplastic syndromes, acquired aplastic anemia, and chronic idiopathic neutropenia.