Intravenous administration of bone marrow-derived multipotent mesenchymal stromal cells enhances the recruitment of CD11b(+) myeloid cells to the lungs and facilitates B16-F10 melanoma colonization.

The discovery that the regenerative properties of bone marrow multipotent mesenchymal stromal cells (BM-MSCs) could collaterally favor neoplastic progression has led to a great interest in the function of these cells in tumors. However, the effect of BM-MSCs on colonization, a rate-limiting step of the metastatic cascade, is unknown. In this study, we investigated the effect of BM-MSCs on metastatic outgrowth of B16-F10 melanoma cells. In in vitro experiments, direct co-culture assays demonstrated that BM-MSCs stimulated the proliferation of B16-F10 cells in a dose-dependent manner. For in vivo experiments, luciferase-expressing B16-F10 cells were injected through tail vein and mice were subsequently treated with four systemic injections of BM-MSCs. In vivo bioluminescent imaging during 16 days demonstrated that BM-MSCs enhanced the colonization of lungs by B16-F10 cells, which correlated with a 2-fold increase in the number of metastatic foci. Flow cytometry analysis of lungs demonstrated that although mice harboring B16-F10 metastases displayed more endothelial cells, CD4 T and CD8 T lymphocytes in the lungs in comparison to metastases-free mice, BM-MSCs did not alter the number of these cells. Interestingly, BM-MSCs inoculation resulted in a 2-fold increase in the number of CD11b(+) myeloid cells in the lungs of melanoma-bearing animals, a cell population previously described to organize "premetastatic niches" in experimental models. These findings indicate that BM-MSCs provide support to B16-F10 cells to overcome the constraints that limit metastatic outgrowth and that these effects might involve the interplay between BM-MSCs, CD11b(+) myeloid cells and tumor cells.

Mast cells control insulitis and increase Treg cells to confer protection against STZ-induced type 1 diabetes in mice.

Quantitative alterations in mast cell numbers in pancreatic lymph nodes (PLNs) have been reported to be associated with type 1 diabetes (T1D) progression, but their potential role during T1D remains unclear. In this study, we evaluated the role of mast cells in T1D induced by multiple low-dose streptozotocin (MLD-STZ) treatments, using two strains of mast cell-deficient mice (W/W(v) or Wsh/Wsh) and the adoptive transfer of mast cells. Mast cell deficient mice developed severe insulitis and accelerated hyperglycemia, with 100% of mice becoming diabetic compared to their littermates. In parallel, these diabetic mice had decreased numbers of T regulatory (Treg) cells in the PLNs. Additionally, mast cell deficiency caused a significant reduction in IL-10, TGF-ß, and IL-6 expression in the pancreatic tissue. Interestingly, IL-6-deficient mice are more susceptible to T1D associated with reduced Treg-cell numbers in the PLNs, but mast cell transfer from wild-type mice induced protection to T1D in these mice. Finally, mast cell adoptive transfer prior to MLD-STZ administration conferred resistance to T1D, promoted increased Treg cells, and decreased IL-17-producing T cells in the PLNs. Taken together, our results indicate that mast cells are implicated in resistance to STZ-induced T1D via an immunological tolerance mechanism mediated by Treg cells.

Mitochondrial DNA Activates the NLRP3 Inflammasome and Predisposes to Type 1 Diabetes in Murine Model.

Although a correlation between polymorphisms of NOD-like receptor family-pyrin domain containing 3 (NLRP3) and predisposition to type 1 diabetes (T1D) has been identified, the potential function and activation of the NLRP3 inflammasome in T1D have not been clarified. The present study shows that non-obese diabetic mice exhibited increased NLRP3, and pro-IL-1ß gene expression in pancreatic lymph nodes (PLNs). Similar increases in gene expression of NLRP3, apoptosis associated speck like protein (ASC) and pro-IL-1ß were induced by multiple low doses of streptozotocin (STZ) in C57BL/6 mice. In addition, diabetic C57BL/6 mice also exhibited increased IL-1ß protein expression in the pancreatic tissue at day 7, which remained elevated until day 15. Diabetic mice also showed increased positive caspase-1 macrophages in the PLNs, which were decreased in NLRP3-/- mice, but not in ASC-/- mice, after STZ treatment. NLRP3- and IL-1R-deficient mice, but not ASC-deficient mice, showed reduced incidence of diabetes, less insulitis, lower hyperglycemia, and normal insulin levels compared to wild-type (WT) diabetic mice. Notably, these mice also displayed a decrease in IL-17-producing CD4 and CD8 T cells (Th17 and Tc17) and IFN-γ-producing CD4 and CD8 T cells (Th1 and Tc1) in the PLNs. Following STZ treatment to induce T1D, NLRP3-deficient mice also exhibited an increase in myeloid-derived suppressor cell and mast cell numbers in the PLNs along with a significant increase in IL-6, IL-10, and IL-4 expression in the pancreatic tissue. Interestingly, diabetic mice revealed increased circulating expression of genes related to mitochondrial DNA, such as cytochrome b and cytochrome c, but not NADH dehydrogenase subunit 6 (NADH). Mitochondrial DNA (mDNA) from diabetic mice, but not from non-diabetic mice, induced significant IL-1ß production and caspase-1 activation by WT macrophages, which was reduced in NLRP3-/- macrophages. Finally, mDNA administration in vivo increased Th17/Tc17/Th1/Tc1 cells in the PLNs and precipitated T1D onset, which was abolished in NLRP3-/- mice. Overall, our results demonstrate that mDNA-mediated NLRP3 activation triggers caspase-1-dependent IL-1ß production and contributes to pathogenic cellular responses during the development of STZ-induced T1D.

Transcriptional profiling reveals intrinsic mRNA alterations in multipotent mesenchymal stromal cells isolated from bone marrow of newly-diagnosed type 1 diabetes patients.

BACKGROUND: Bone marrow multipotent mesenchymal stromal cells (MSCs) are a diverse subset of precursors that contribute to the homeostasis of the hematopoietic niche. MSCs can be isolated and expanded in vitro and have unique immunomodulatory and regenerative properties that make them attractive for the treatment of autoimmune diseases, including type 1 diabetes (T1D). Whether autologous or allogeneic MSCs are more suitable for therapeutic purposes has not yet been established. While autologous MSCs may present abnormal function, allogeneic cells may be recognized and rejected by the host immune system. Thus, studies that investigate biological characteristics of MSCs isolated from T1D patients are essential to guide future clinical applications. METHODS: Bone marrow-derived MSCs from recently diagnosed type 1 diabetes patients (T1D-MSCs) were compared with those from healthy individuals (C-MSCs) for morphological and immunophenotypic characteristics and for differentiation potential. Bioinformatics approaches allowed us to match absolute and differential gene expression of several adhesion molecules, immune mediators, growth factors, and their receptors involved with hematopoietic support and immunomodulatory properties of MSCs. Finally, the differentially expressed genes were collated for functional pathway enrichment analysis. RESULTS: T1D-MSCs and C-MSCs were similar for morphology, immunophenotype, and differentiation potential. Our absolute gene expression results supported previous literature reports, while also detecting new potential molecules related to bone marrow-derived MSC functions. T1D-MSCs showed intrinsic abnormalities in mRNA expression, including the immunomodulatory molecules VCAM-1, CXCL12, HGF, and CCL2. Pathway analyses revealed activation of sympathetic nervous system and JAK STAT signaling in T1D-MSCs. CONCLUSIONS: Collectively, our results indicate that MSCs isolated from T1D patients present intrinsic transcriptional alterations that may affect their therapeutic potential. However, the implications of these abnormalities in T1D development as well as in the therapeutic efficacy of autologous MSCs require further investigation.