RESUMEN
BACKGROUND AIMS: The biodistribution of human MSCs after systemic delivery is incompletely understood. We investigated the changes in cell size and cell surface markers of human MSCs after intravenous (IV) injection in immune competent mice. METHODS: Male human MSCs were labeled with fluorescent vital dye PKH67 and tracked after IV administration in C57/BL6 mice. MSCs were tracked in blood and different murine tissues by human SRY gene quantitative polymerase chain reaction (qPCR) analysis, flow cytometry and fluorescence microscopy. Calibrated microbeads were used to track the size of transplanted MSCs. RESULTS: The majority of injected MSCs were detected by qPCR in the lungs 5 min after transplantation, whereas <0.1% were detected in other tissues over 24 h. Flow cytometric and fluorescence microscopic analysis indicated that MSCs continuously decreased in size after transplantation and underwent fragmentation. The majority of PKH+ MSCs and their fragments were found in lungs and liver. PKH+ MSCs rapidly became positive for annexin V, propidium iodide and calreticulin, indicating loss of cell integrity. In addition, PKH+ fragments co-stained with antibodies against C3b, F4/80 and/or GR-1 indicating opsonization. Preincubation of MSCs in hyperosmolaric hydroxyethyl starch (HyperHAES) decreased MSCs size before transplantation, delayed the loss of viability markers and increased the frequency of traceable MSCs up to 24 h after transplantation. CONCLUSIONS: PKH67 labeled MSCs are fragmented after IV injection in mice, acquire apoptotic and phagocytic cell markers and accumulate in the lungs and liver.
Asunto(s)
Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/citología , Animales , Apoptosis , Biomarcadores/análisis , Tamaño de la Célula , Supervivencia Celular , Citometría de Flujo/métodos , Xenoinjertos , Humanos , Inyecciones Intravenosas , Ratones Endogámicos C57BL , Compuestos Orgánicos/farmacocinética , Distribución TisularRESUMEN
Multipotent mesenchymal stromal cells derived from human placenta (pMSCs), and unrestricted somatic stem cells (USSCs) derived from cord blood share many properties with human bone marrow-derived mesenchymal stromal cells (bmMSCs) and are currently in clinical trials for a wide range of clinical settings. Here we present gene expression profiles of human cord blood-derived unrestricted somatic stem cells (USSCs), human placental-derived mesenchymal stem cells (hpMSCs), and human bone marrow-derived mesenchymal stromal cells (bmMSCs), all derived from four different donors. The microarray data are available on the ArrayExpress database (www.ebi.ac.uk/arrayexpress) under accession number E-TABM-880. Additionally, the data has been integrated into a public portal, www.stemformatics.org. Our data provide a resource for understanding the differences in MSCs derived from different tissues.
RESUMEN
Mobilization of hematopoietic stem and progenitor cells (HPCs) is induced by treatment with granulocyte-colony stimulating factor, chemotherapy, or irradiation. We observed that these treatments are accompanied by a release of chemotactic activity into the blood. This plasma activity is derived from the bone marrow, liver, and spleen and acts on HPCs via the chemokine receptor CXCR4. A human blood peptide library was used to characterize CXCR4-activating compounds. We identified CXCL12[22-88] and N-terminally truncated variants CXCL12[24-88], CXCL12[25-88], CXCL12[27-88], and CXCL12[29-88]. Only CXCL12[22-88] could effectively bind to CXCR4 and induce intracellular calcium flux and chemotactic migration of HPCs. CXCL12[25-88] and CXCL12[27-88] revealed neither agonistic nor antagonistic activities in vitro, whereas CXCL12[29-88] inhibited CXCL12[22-88]-induced chemotactic migration. Since binding to glycosaminoglycans (GAG) modulates the function of CXCL12, binding to heparin was analyzed. Surface plasmon resonance kinetic analysis showed that N-terminal truncation of Arg22-Pro23 increased the dissociation constant KD by one log10 stage ([22-88]: KD: 5.4 ± 2.6 µM; [24-88]: KD: 54 ± 22.4 µM). Further truncation of the N-terminus decreased the KD ([25-88] KD: 30 ± 4.8 µM; [27-88] KD: 23 ± 1.6 µM; [29-88] KD: 19 ± 5.4 µM), indicating increasing competition for heparin binding. Systemic in vivo application of CXCL12[22-88] as well as CXCL12[27-88] or CXCL12[29-88] induced a significant mobilization of HPCs in mice. Our findings indicate that plasma-derived CXCL12 variants may contribute to the regulation of HPC mobilization by modulating the binding of CXCL12[22-88] to GAGs rather than blocking the CXCR4 receptor and, therefore, may have a contributing role in HPC mobilization.
Asunto(s)
Quimiocina CXCL12/sangre , Quimiotaxis , Células Madre Hematopoyéticas/fisiología , Animales , Plaquetas/metabolismo , Señalización del Calcio , Células Cultivadas , Trasplante de Células Madre Hematopoyéticas , Humanos , Leucocitos/metabolismo , Ratones Endogámicos DBA , Unión Proteica , Procesamiento Proteico-Postraduccional , ProteolisisRESUMEN
Haematopoietic stem cell (HSC) transplantation is an established cell-based therapy for a number of haematological diseases. To enhance this therapy, there is considerable interest in expanding HSCs in artificial niches prior to transplantation. This study compared murine HSC expansion supported through co-culture on monolayers of either undifferentiated mesenchymal stromal cells (MSCs) or osteoblasts. Sorted Lineage(-) Sca-1(+) c-kit(+) (LSK) haematopoietic stem/progenitor cells (HPC) demonstrated proliferative capacity on both stromal monolayers with the greatest expansion of LSK shown in cultures supported by osteoblast monolayers. After transplantation, both types of bulk-expanded cultures were capable of engrafting and repopulating lethally irradiated primary and secondary murine recipients. LSKs co-cultured on MSCs showed comparable, but not superior, reconstitution ability to that of freshly isolated LSKs. Surprisingly, however, osteoblast co-cultured LSKs showed significantly poorer haematopoietic reconstitution compared to LSKs co-cultured on MSCs, likely due to a delay in short-term reconstitution. We demonstrated that stromal monolayers can be used to maintain, but not expand, functional HSCs without a need for additional haematopoietic growth factors. We also demonstrated that despite apparently superior in vitro performance, co-injection of bulk cultures of osteoblasts and LSKs in vivo was detrimental to recipient survival and should be avoided in translation to clinical practice.
RESUMEN
BACKGROUND: Multipotent mesenchymal stromal cells suppress T-cell function in vitro, a property that has underpinned their use in treating clinical steroid-refractory graft-versus-host disease after allogeneic hematopoietic stem cell transplantation. However the potential of mesenchymal stromal cells to resolve graft-versus-host disease is confounded by a paucity of pre-clinical data delineating their immunomodulatory effects in vivo. DESIGN AND METHODS: We examined the influence of timing and dose of donor-derived mesenchymal stromal cells on the kinetics of graft-versus-host disease in two murine models of graft-versus-host disease (major histocompatibility complex-mismatched: UBI-GFP/BL6 [H-2(b)]âBALB/c [H-2(d)] and the sibling transplant mimic, UBI-GFP/BL6 [H-2(b)]âBALB.B [H-2(b)]) using clinically relevant conditioning regimens. We also examined the effect of mesenchymal stromal cell infusion on bone marrow and spleen cellular composition and cytokine secretion in transplant recipients. RESULTS: Despite T-cell suppression in vitro, mesenchymal stromal cells delayed but did not prevent graft-versus-host disease in the major histocompatibility complex-mismatched model. In the sibling transplant model, however, 30% of mesenchymal stromal cell-treated mice did not develop graft-versus-host disease. The timing of administration and dose of the mesenchymal stromal cells influenced their effectiveness in attenuating graft-versus-host disease, such that a low dose of mesenchymal stromal cells administered early was more effective than a high dose of mesenchymal stromal cells given late. Compared to control-treated mice, mesenchymal stromal cell-treated mice had significant reductions in serum and splenic interferon-γ, an important mediator of graft-versus-host disease. CONCLUSIONS: Mesenchymal stromal cells appear to delay death from graft-versus-host disease by transiently altering the inflammatory milieu and reducing levels of interferon-γ. Our data suggest that both the timing of infusion and the dose of mesenchymal stromal cells likely influence these cells' effectiveness in attenuating graft-versus-host disease.
Asunto(s)
Enfermedad Injerto contra Huésped/inmunología , Trasplante de Células Madre Hematopoyéticas/métodos , Células Madre Mesenquimatosas/inmunología , Células del Estroma/inmunología , Animales , Células Cultivadas , Técnicas de Cocultivo , Citocinas/inmunología , Citocinas/metabolismo , Femenino , Enfermedad Injerto contra Huésped/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Mediadores de Inflamación/inmunología , Mediadores de Inflamación/metabolismo , Interferón gamma/inmunología , Interferón gamma/metabolismo , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Células del Estroma/citología , Células del Estroma/metabolismo , Análisis de Supervivencia , Linfocitos T/inmunología , Linfocitos T/metabolismo , Factores de TiempoRESUMEN
Cardiovascular diseases, particularly acute myocardial infarction, are the leading causes of death worldwide. Important advances have been made in the secondary treatment for cardiovascular diseases such as heart transplantation and medical and surgical therapies. Although these therapies alleviate symptoms, and may even improve survival, none can reverse the disease process and directly repair the lasting damage. Thus, the cure of cardiovascular diseases remains a major unmet medical need. Recently, cellular therapy has been proposed as a candidate treatment for this. Many stem and progenitor cell populations have each been suggested as a potential basis for such therapy. This review assesses some of the more notable exogenous adult cell candidates and provides insights into the mechanisms by which they may mediate improvement in cardiac function following acute myocardial infarction. Research into the cellular therapy field is of great importance for the further planning of clinical trials for cardiac cellular myoplasty.
RESUMEN
Mesenchymal stem cells or multipotent mesenchymal stromal cells (both referred to as MSC) have been shown in some studies to have a beneficial effect on myocardial recovery after infarct. Current strategies for MSC delivery to heart involve intravenous, intraarterial, and intramuscular delivery. Different routes of MSC delivery and a lack of knowledge of the mechanisms that MSC utilise to migrate in vivo has most likely led to the marked variations in results that have been found. This review aims to summarise the current knowledge of MSC migratory mechanisms and looks to future methods of MSC manipulation prior to delivery in order to enhance MSC migration and engraftment.