Bone marrow stromal cell-derived hepcidin has antimicrobial and immunomodulatory activities.

Bone marrow stromal cells (BMSCs) have immunomodulatory activities in numerous species and have been used in clinical trials. BMSCs also make antibacterial agents. Since hepcidin is known to have antimicrobial effects in fish, we wondered if it might also be used as an antimicrobial agent by mammalian BMSCs. In the present study, we show hepcidin expression in both mouse (mBMSC) and human BMSCs (hBMSC). We observed a hBMSC hepcidin-dependent degradation of ferroportin in HEK-293 reporter cells in vitro. In human and mouse bone marrows (BM) we detected hepcidin-positive BMSCs in close proximity to hematopoietic progenitors. The conditioned culture medium of hBMSCs significantly reduced bacterial proliferation that was partially blocked by a hepcidin-neutralizing antibody. Similarly, medium in which hepcidin-deficient (Hamp-/-) mouse BMSCs had been grown was significantly less effective in reducing bacterial counts than the medium of wild-type cells. In a zymosan-induced peritonitis mouse model we found that mBMSC-derived hepcidin reduced the number of invading polymorphonuclear (PMN) cells in the peritoneal cavity. Our results show that BMSC-derived hepcidin has antimicrobial properties in vitro and also reduces inflammation in vivo. We conclude that hepcidin should be added to the expanding arsenal of agents available to BMSCs to fight infections and inflammation.

Bone Marrow-Derived Mesenchymal Stromal Cells (MSCs) Modulate the Inflammatory Character of Alveolar Macrophages from Sarcoidosis Patients.

Sarcoidosis is a devastating inflammatory disease affecting many organs, especially the lungs and lymph nodes. Bone marrow-derived mesenchymal stromal cells (MSCs) can "reprogram" various types of macrophages towards an anti-inflammatory phenotype. We wanted to determine whether alveolar macrophages from sarcoidosis subjects behave similarly by mounting an anti-inflammatory response when co-cultured with MSCs. Fifteen sarcoidosis and eight control subjects underwent bronchoscopy and bronchoalveolar lavage (BAL). Unselected BAL cells (70-94% macrophages) were isolated and cultured with and without MSCs from healthy adults. Following stimulation of the cultured cells with lipopolysaccharide, the medium was removed to measure interleukin 10 and tumor necrosis factor alpha (IL-10 and TNF-α). In two additional sarcoidosis subjects, flow cytometry was used to study intracellular cytokines and surface markers associated with alveolar macrophages to confirm the results. Unselected BAL cells from sarcoidosis subjects co-cultured with MSCs showed a reduction in TNF-α (pro-inflammatory M1) and an increase in IL-10 (anti-inflammatory M2) in 9 of 11 samples studied. Control subject samples showed few, if any, differences in cytokine production. Unselected BAL cells from two additional patients analyzed by flow cytometry confirmed a switch towards an anti-inflammatory state (i.e., M1 to M2) after co-culture with MSCs. These results suggest that, similarly to other macrophages, alveolar macrophages also respond to MSC contacts by changing towards an anti-inflammatory phenotype. Based on our results, we hypothesize that mesenchymal stromal cells applied to the airways might alleviate lung inflammation and decrease steroid need in patients with sarcoidosis.

Mesenchymal stromal cells from infants with simple polydactyly modulate immune responses more efficiently than adult mesenchymal stromal cells.

Bone marrow-derived stromal cells or mesenchymal stromal cells (BMSCs or MSCs, as we will call them in this work) are multipotent progenitor cells that can differentiate into osteoblasts, adipocytes and chondrocytes. In addition, MSCs have been shown to modulate the function of a variety of immune cells. Donor age has been shown to affect the regenerative potential, differentiation, proliferation and anti-inflammatory potency of MSCs; however, the impact of donor age on their immunosuppressive activity is unknown. In this study, we evaluated the ability of MSCs derived from very young children and adults on T-cell suppression and cytokine secretion by monocytes/macrophages. MSCs were obtained from extra digits of children between 10 and 21 months and adults between 28 and 64 years of age. We studied cell surface marker expression, doubling time, lineage differentiation potential and immunosuppressive function of the MSCs. Young MSCs double more quickly and differentiate into bone and fat cells more efficiently than those from older donors. They also form more and dense colonies of fibroblasts (colony forming unit-fibroblast [CFU-F]). MSCs from both young and adult subjects suppressed T-cell proliferation in a mitogen-induced assay at 1:3 and 1:30 ratios. At a 1:30 ratio, however, MSCs from adults did not, but MSCs from infants did suppress T-cell proliferation. In the mixed lymphocyte reaction assay, MSCs from infants produced similar levels of suppression at all three MSC/T-cell ratios, but adult MSCs only inhibited T-cell proliferation at a 1:3 ratio. Cytokine analyses of co-cultures of MSCs and macrophages showed that both adult and young MSCs suppress tumor necrosis factor alpha (TNF-α) and induce interleukin-10 (IL-10) production in macrophage co-culture assay in a similar manner. Overall, this work shows that developing MSCs display a higher level of immunosuppression than mature MSCs.

Immunogenic potential of human bone marrow mesenchymal stromal cells is enhanced by hyperthermia.

BACKGROUND AIMS: Bone marrow-derived mesenchymal stromal cells (MSCs) have been reported to suppress T-cell proliferation and used to alleviate the symptoms of graft-versus-host disease (GVHD). MSCs are a mixed cell population and at this time there are no tools to isolate the cells responsible for the T-cell suppression. We wanted to find a way to enhance the immune-modulatory actions of MSCs and tried varying the temperature at which they were cultured. METHODS: We cultured human MSCs derived from healthy volunteers at different temperatures and tested their ability to switch macrophage character from pro-inflammatory to anti-inflammatory (M1 type to M2 type). Using an enzyme-linked immunosorbent assay (ELISA), we showed that when MSCs are cultured at higher temperatures their ability to induce co-cultured macrophages to produce more interleukin-10, (IL-10) (an anti-inflammatory cytokine) and less tumor necrosis factor alpha, (TNFα) (a pro-inflammatory cytokine) is increased. We performed Western blots and immunocytochemistry to screen for changes that might underlie this effect. RESULTS: We found that in hyperthermia the heat shock protein, HSF1, translocated into the nucleus of MSCs. It appears to induce the COX2/PGE2 (Cyclooxygenase2/Prostaglandin E2) pathway described earlier as a major mechanism of MSC-directed immune-suppression. CONCLUSION: Hyperthermia increases the efficacy of MSC-driven immune-suppression. We propose that changing the time of MSC administration to patients to mid-to-late afternoon when the body temperature is naturally highest might be beneficial. Warming the patient could also be considered.

Vasopressin stimulates the proliferation and differentiation of red blood cell precursors and improves recovery from anemia.

Arginine vasopressin (AVP) made by hypothalamic neurons is released into the circulation to stimulate water resorption by the kidneys and restore water balance after blood loss. Patients who lack this antidiuretic hormone suffer from central diabetes insipidus. We observed that many of these patients were anemic and asked whether AVP might play a role in red blood cell (RBC) production. We found that all three AVP receptors are expressed in human and mouse hematopoietic stem and progenitor cells. The AVPR1B appears to play the most important role in regulating erythropoiesis in both human and mouse cells. AVP increases phosphorylation of signal transducer and activator of transcription 5, as erythropoietin (EPO) does. After sublethal irradiation, AVP-deficient Brattleboro rats showed delayed recovery of RBC numbers compared to control rats. In mouse models of anemia (induced by bleeding, irradiation, or increased destruction of circulating RBCs), AVP increased the number of circulating RBCs independently of EPO. In these models, AVP appears to jump-start peripheral blood cell replenishment until EPO can take over. We suggest that specific AVPR1B agonists might be used to induce fast RBC production after bleeding, drug toxicity, or chemotherapy.