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Posttransplant cyclophosphamide (PTCy) graft-versus-host disease (GVHD) prophylaxis has enabled haploidentical (Haplo) transplantation to be performed with results similar to those after matched unrelated donor (MUD) transplantation with traditional prophylaxis. The relative value of transplantation with MUD vs Haplo donors when both groups receive PTCy/calcineurin inhibitor/mycophenolate GVHD prophylaxis is not known. We compared outcomes after 2036 Haplo and 284 MUD transplantations with PTCy GVHD prophylaxis for acute leukemia or myelodysplastic syndrome in adults from 2011 through 2018. Cox regression models were built to compare outcomes between donor types. Recipients of myeloablative and reduced-intensity regimens were analyzed separately. Among recipients of reduced-intensity regimens, 2-year graft failure (3% vs 11%), acute grades 2 to 4 GVHD (hazards ratio [HR], 0.70; P = .022), acute grades 3 and 4 GVHD (HR, 0.41; P = .016), and nonrelapse mortality (HR, 0.43; P = .0008) were lower after MUD than with Haplo donor transplantation. Consequently, disease-free (HR, 0.74; P = .008; 55% vs 41%) and overall (HR, 0.65; P = .001; 67% vs 54%) survival were higher with MUD than with Haplo transplants. Among recipients of myeloablative regimens, day-100 platelet recovery (95% vs 88%) was higher and grades 3 and 4 acute (HR, 0.39; P = .07) and chronic GVHD (HR, 0.66; P = .05) were lower after MUD than with Haplo donor transplantation. There were no differences in graft failure, relapse, nonrelapse mortality, and disease-free and overall survival between donor types with myeloablative conditioning regimens. These data extend and confirm the importance of donor-recipient HLA matching for allogeneic transplantation. A MUD is the preferred donor, especially for transplantations with reduced-intensity conditioning regimens.
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Ciclofosfamida/uso terapéutico , Enfermedad Injerto contra Huésped/prevención & control , Trasplante de Células Madre Hematopoyéticas/métodos , Inmunosupresores/uso terapéutico , Leucemia Mieloide Aguda/terapia , Síndromes Mielodisplásicos/terapia , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Acondicionamiento Pretrasplante , Trasplante Haploidéntico/métodos , Trasplante Homólogo/métodos , Resultado del Tratamiento , Donante no EmparentadoRESUMEN
Cell and gene therapy is a promising and disruptive new field of medicine for diseases lacking effective treatments. Collaboration among stakeholders has become critically important as investigators, health care providers, manufacturers, couriers, data registries, regulators and payers all become more invested in the success of this field. Many organizations have collaborated with each other to increase clarity, advocate for improvements and share lessons learned. These efforts appear to be making an impact, although the potential for duplicative efforts could slow progress. The second Regenerative Medicine InterCHANGE, hosted by the Foundation for the Accreditation of Cellular Therapy, took place at the Phacilitate Leaders World/World Stem Cell Summit conference in Miami, Florida, on January 24, 2020. Participants from several organizations outlined needs to advance cell and gene therapies. Efforts to address these include standardization, workforce development and advocacy. This article summarizes the major challenges and opportunities discussed during the InterCHANGE.
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Tratamiento Basado en Trasplante de Células y Tejidos , Medicina Regenerativa , Terapia Genética , Humanos , Estándares de Referencia , Recursos HumanosRESUMEN
Adoptive therapy with regulatory T cells (Tregs) to prevent graft-versus-host disease (GVHD) would benefit from a strategy to improve homing to the sites of inflammation. We hypothesized that adding fucose to human Tregs, forming the Sialyl Lewis X moiety on P-selectin glycoprotein ligand-1, would improve their trafficking pattern. The selectin pathway recruiter, α-1,3-fucosyltransferase-VI enzyme, significantly increased Treg surface fucosylation (66% vs 8%). In a xenogenic GVHD mouse model, fucosylated Tregs showed prolonged periods of in vivo persistence. When given at a lower dose compared with the untreated Tregs, the murine recipients of fucosylated Tregs maintained weight, had ameliorated clinical GVHD, and improved survival (70% vs 30%; P < .0001). These preclinical data indicate that fucosylated human Tregs is an effective strategy for prevention of GVHD and, as such, warrants consideration for future clinical trials.
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Modelos Animales de Enfermedad , Fucosa/metabolismo , Enfermedad Injerto contra Huésped/inmunología , Enfermedad Injerto contra Huésped/prevención & control , Linfocitos T Reguladores/inmunología , Animales , Western Blotting , Células Cultivadas , Selectina E/metabolismo , Femenino , Sangre Fetal/citología , Sangre Fetal/inmunología , Sangre Fetal/metabolismo , Citometría de Flujo , Fucosiltransferasas/metabolismo , Enfermedad Injerto contra Huésped/mortalidad , Humanos , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones SCID , Oligosacáridos/metabolismo , Antígeno Sialil Lewis XRESUMEN
Delayed engraftment is a major limitation of cord blood transplantation (CBT), due in part to a defect in the cord blood (CB) cells' ability to home to the bone marrow. Because this defect appears related to low levels of fucosylation of cell surface molecules that are responsible for binding to P- and E-selectins constitutively expressed by the marrow microvasculature, and thus for marrow homing, we conducted a first-in-humans clinical trial to correct this deficiency. Patients with high-risk hematologic malignancies received myeloablative therapy followed by transplantation with 2 CB units, one of which was treated ex vivo for 30 minutes with the enzyme fucosyltransferase-VI and guanosine diphosphate fucose to enhance the interaction of CD34(+) stem and early progenitor cells with microvessels. The results of enforced fucosylation for 22 patients enrolled in the trial were then compared with those for 31 historical controls who had undergone double unmanipulated CBT. The median time to neutrophil engraftment was 17 days (range, 12-34 days) compared with 26 days (range, 11-48 days) for controls (P = .0023). Platelet engraftment was also improved: median was 35 days (range, 18-100 days) compared with 45 days (range, 27-120 days) for controls (P = .0520). These findings support ex vivo fucosylation of multipotent CD34(+) CB cells as a clinically feasible means to improve engraftment efficiency in the double CBT setting. The trial is registered to www.clinicaltrials.gov as #NCT01471067.
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Plaquetas/citología , Sangre Fetal/citología , Fucosa/metabolismo , Neoplasias Hematológicas/terapia , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Neutrófilos/trasplante , Adolescente , Adulto , Anciano , Plaquetas/inmunología , Estudios de Cohortes , Selectina E/metabolismo , Estudios de Factibilidad , Femenino , Sangre Fetal/inmunología , Fucosiltransferasas/metabolismo , Enfermedad Injerto contra Huésped , Neoplasias Hematológicas/inmunología , Neoplasias Hematológicas/mortalidad , Células Madre Hematopoyéticas/inmunología , Humanos , Masculino , Persona de Mediana Edad , Neutrófilos/citología , Neutrófilos/inmunología , Selectina-P/metabolismo , Transfusión de Plaquetas , Pronóstico , Tasa de Supervivencia , Adulto JovenRESUMEN
A subset of regulatory B cells (Bregs) in mice negatively regulate T-cell immune responses through the secretion of regulatory cytokines such as IL-10 and direct cell-cell contact and have been linked to experimental models of autoimmunity, inflammation, and cancer. However, the regulatory function of Bregs in human disease is much less clear. Here we demonstrate that B cells with immunoregulatory properties are enriched within both the CD19(+)IgM(+)CD27(+) memory and CD19(+)CD24(hi)CD38(hi) transitional B-cell subsets in healthy human donors. Both subsets suppressed the proliferation and interferon-γ production of CD3/CD28-stimulated autologous CD4(+) T cells in a dose-dependent manner, and both relied on IL-10 secretion as well as cell-cell contact, likely mediated through CD80 and CD86, to support their full suppressive function. Moreover, after allogeneic stem cell transplantation, Bregs from patients with chronic graft-versus-host disease (cGVHD) were less frequent and less likely to produce IL-10 than were Bregs from healthy donors and patients without cGVHD. These findings suggest that Bregs may be involved in the pathogenesis of cGVHD and support future investigation of regulatory B cell-based therapy in the treatment of this disease.
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Subgrupos de Linfocitos B/inmunología , Linfocitos B Reguladores/inmunología , Enfermedad Injerto contra Huésped/inmunología , Inmunoglobulina M/inmunología , Memoria Inmunológica , ADP-Ribosil Ciclasa 1/metabolismo , Antígenos CD19/metabolismo , Subgrupos de Linfocitos B/metabolismo , Linfocitos B Reguladores/metabolismo , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Antígeno CD24/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Estudios de Casos y Controles , Comunicación Celular , Células Cultivadas , Enfermedad Crónica , Citocinas/biosíntesis , Enfermedad Injerto contra Huésped/metabolismo , Humanos , Inmunomodulación , Inmunofenotipificación , Mediadores de Inflamación/metabolismo , Interleucina-10/biosíntesis , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis TumoralRESUMEN
Continued growth in the cell therapy industry and commercialization of cell therapies that successfully advance through clinical trials has led to increased awareness around the need for specialized and complex materials utilized in their manufacture. Ancillary materials (AMs) are components or reagents used during the manufacture of cell therapy products but are not intended to be part of the final products. Commonly, there are limitations in the availability of clinical-grade reagents used as AMs. Furthermore, AMs may affect the efficacy of the cell product and subsequent safety of the cell therapy for the patient. As such, AMs must be carefully selected and appropriately qualified during the cell therapy development process. However, the ongoing evolution of cell therapy research, limited number of clinical trials and registered cell therapy products results in the current absence of specific regulations governing the composition, compliance, and qualification of AMs often leads to confusion by suppliers and users in this field. Here we provide an overview and interpretation of the existing global framework surrounding AM use and investigate some common misunderstandings within the industry, with the aim of facilitating the appropriate selection and qualification of AMs. The key message we wish to emphasize is that in order to most effectively mitigate risk around cell therapy development and patient safety, users must work with their suppliers and regulators to qualify each AM to assess source, purity, identity, safety, and suitability in a given application.
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Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Humanos , Internacionalidad , Control Social Formal , Terminología como AsuntoRESUMEN
Mesenchymal stromal cells (MSCs) as a pharmaceutical for ailments characterized by pathogenic autoimmune, alloimmune and inflammatory processes now cover the spectrum of early- to late-phase clinical trials in both industry and academic sponsored studies. There is a broad consensus that despite different tissue sourcing and varied culture expansion protocols, human MSC-like cell products likely share fundamental mechanisms of action mediating their anti-inflammatory and tissue repair functionalities. Identification of functional markers of potency and reduction to practice of standardized, easily deployable methods of measurements of such would benefit the field. This would satisfy both mechanistic research as well as development of release potency assays to meet Regulatory Authority requirements for conduct of advanced clinical studies and their eventual registration. In response to this unmet need, the International Society for Cellular Therapy (ISCT) addressed the issue at an international workshop in May 2015 as part of the 21st ISCT annual meeting in Las Vegas. The scope of the workshop was focused on discussing potency assays germane to immunomodulation by MSC-like products in clinical indications targeting immune disorders. We here provide consensus perspective arising from this forum. We propose that focused analysis of selected MSC markers robustly deployed by in vitro licensing and metricized with a matrix of assays should be responsive to requirements from Regulatory Authorities. Workshop participants identified three preferred analytic methods that could inform a matrix assay approach: quantitative RNA analysis of selected gene products; flow cytometry analysis of functionally relevant surface markers and protein-based assay of secretome. We also advocate that potency assays acceptable to the Regulatory Authorities be rendered publicly accessible in an "open-access" manner, such as through publication or database collection.
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Bioensayo/métodos , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/citología , Biomarcadores/metabolismo , Citometría de Flujo/métodos , HumanosRESUMEN
RATIONALE: Transendocardial stem cell injection (TESI) with mesenchymal stem cells improves remodeling in chronic ischemic cardiomyopathy, but the effect of the injection site remains unknown. OBJECTIVE: To address whether TESI exerts its effects at the site of injection only or also in remote areas, we hypothesized that segmental myocardial scar and segmental ejection fraction improve to a greater extent in injected than in noninjected segments. METHODS AND RESULTS: Biplane ventriculographic and endocardial tracings were recorded. TESI was guided to 10 sites in infarct-border zones. Sites were mapped according to the 17-myocardial segment model. As a result, 510 segments were analyzed in 30 patients before and 13 months after TESI. Segmental early enhancement defect (a measure of scar size) was reduced by TESI in both injected (-43.7 ± 4.4%; n=95; P<0.01) and noninjected segments (-25.1 ± 7.8%; n=148; P<0.001; between-group comparison P<0.05). Conversely, segmental ejection fraction (a measure of contractile performance) improved in injected scar segments (19.9 ± 3.3-26.3 ± 3.5%; P=0.003) but not in noninjected scar segments (21.3 ± 2.6-23.5 ± 3.2%; P=0.20; between-group comparison P<0.05). Furthermore, segmental ejection fraction in injected scar segments improved to a greater degree in patients with baseline segmental ejection fraction <20% (12.1 ± 1.2-19.9 ± 2.7%; n=18; P=0.003), versus <20% (31.7 ± 3.4-35.5 ± 3.3%; n=12; P=0.33, between-group comparison P<0.0001). CONCLUSIONS: These findings illustrate a dichotomy in regional responses to TESI. Although scar size reduction was evident in all scar segments, scar size reduction and ventricular functional responses preferentially occurred at the sites of TESI versus non-TESI sites. Furthermore, improvement was greatest when segmental left ventricular dysfunction was severe.
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Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Cicatriz/patología , Cicatriz/terapia , Trasplante de Células Madre Mesenquimatosas/métodos , Infarto del Miocardio/patología , Infarto del Miocardio/terapia , Anciano , Cicatriz/diagnóstico por imagen , Femenino , Humanos , Inyecciones , Masculino , Persona de Mediana Edad , Desarrollo de Músculos/fisiología , Infarto del Miocardio/diagnóstico por imagen , Volumen Sistólico/fisiología , Tomografía Computarizada Espiral , Resultado del Tratamiento , Disfunción Ventricular Izquierda/fisiopatología , Disfunción Ventricular Izquierda/terapiaRESUMEN
RATIONALE: Although accumulating data support the efficacy of intramyocardial cell-based therapy to improve left ventricular (LV) function in patients with chronic ischemic cardiomyopathy undergoing CABG, the underlying mechanism and impact of cell injection site remain controversial. Mesenchymal stem cells (MSCs) improve LV structure and function through several effects including reducing fibrosis, neoangiogenesis, and neomyogenesis. OBJECTIVE: To test the hypothesis that the impact on cardiac structure and function after intramyocardial injections of autologous MSCs results from a concordance of prorecovery phenotypic effects. METHODS AND RESULTS: Six patients were injected with autologous MSCs into akinetic/hypokinetic myocardial territories not receiving bypass graft for clinical reasons. MRI was used to measure scar, perfusion, wall thickness, and contractility at baseline, at 3, 6, and 18 months and to compare structural and functional recovery in regions that received MSC injections alone, revascularization alone, or neither. A composite score of MRI variables was used to assess concordance of antifibrotic effects, perfusion, and contraction at different regions. After 18 months, subjects receiving MSCs exhibited increased LV ejection fraction (+9.4 ± 1.7%, P=0.0002) and decreased scar mass (-47.5 ± 8.1%; P<0.0001) compared with baseline. MSC-injected segments had concordant reduction in scar size, perfusion, and contractile improvement (concordant score: 2.93 ± 0.07), whereas revascularized (0.5 ± 0.21) and nontreated segments (-0.07 ± 0.34) demonstrated nonconcordant changes (P<0.0001 versus injected segments). CONCLUSIONS: Intramyocardial injection of autologous MSCs into akinetic yet nonrevascularized segments produces comprehensive regional functional restitution, which in turn drives improvement in global LV function. These findings, although inconclusive because of lack of placebo group, have important therapeutic and mechanistic hypothesis-generating implications. CLINICAL TRIAL REGISTRATION URL: http://clinicaltrials.gov/show/NCT00587990. Unique identifier: NCT00587990.
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Cardiomiopatías/terapia , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Puente de Arteria Coronaria , Trasplante de Células Madre Mesenquimatosas/métodos , Isquemia Miocárdica/terapia , Miocardio/patología , Disfunción Ventricular Izquierda/terapia , Cicatriz/patología , Cicatriz/terapia , Fibrosis/patología , Fibrosis/terapia , Estudios de Seguimiento , Humanos , Inyecciones , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Fenotipo , Estudios Prospectivos , Factores de Tiempo , Resultado del TratamientoRESUMEN
BACKGROUND: Poor engraftment due to low cell doses restricts the usefulness of umbilical-cord-blood transplantation. We hypothesized that engraftment would be improved by transplanting cord blood that was expanded ex vivo with mesenchymal stromal cells. METHODS: We studied engraftment results in 31 adults with hematologic cancers who received transplants of 2 cord-blood units, 1 of which contained cord blood that was expanded ex vivo in cocultures with allogeneic mesenchymal stromal cells. The results in these patients were compared with those in 80 historical controls who received 2 units of unmanipulated cord blood. RESULTS: Coculture with mesenchymal stromal cells led to an expansion of total nucleated cells by a median factor of 12.2 and of CD34+ cells by a median factor of 30.1. With transplantation of 1 unit each of expanded and unmanipulated cord blood, patients received a median of 8.34×10(7) total nucleated cells per kilogram of body weight and 1.81×10(6) CD34+ cells per kilogram--doses higher than in our previous transplantations of 2 units of unmanipulated cord blood. In patients in whom engraftment occurred, the median time to neutrophil engraftment was 15 days in the recipients of expanded cord blood, as compared with 24 days in controls who received unmanipulated cord blood only (P<0.001); the median time to platelet engraftment was 42 days and 49 days, respectively (P=0.03). On day 26, the cumulative incidence of neutrophil engraftment was 88% with expansion versus 53% without expansion (P<0.001); on day 60, the cumulative incidence of platelet engraftment was 71% and 31%, respectively (P<0.001). CONCLUSIONS: Transplantation of cord-blood cells expanded with mesenchymal stromal cells appeared to be safe and effective. Expanded cord blood in combination with unmanipulated cord blood significantly improved engraftment, as compared with unmanipulated cord blood only. (Funded by the National Cancer Institute and others; ClinicalTrials.gov number, NCT00498316.).
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Trasplante de Células Madre de Sangre del Cordón Umbilical , Neoplasias Hematológicas/terapia , Trasplante de Células Madre Mesenquimatosas , Adolescente , Adulto , Recuento de Células Sanguíneas , Plaquetas , Causas de Muerte , Técnicas de Cultivo de Célula , Refuerzo Inmunológico de Injertos , Enfermedad Injerto contra Huésped , Neoplasias Hematológicas/mortalidad , Humanos , Células Madre Mesenquimatosas , Persona de Mediana Edad , Neutrófilos , Quimera por Trasplante , Trasplante Homólogo , Adulto JovenRESUMEN
BACKGROUND AIMS: Mesenchymal stromal cells (MSCs), derived from several tissues including bone marrow and adipose tissue, are being evaluated in clinical trials for a range of diseases. Virtually all tissues of the body contain stromal cells, yet it is unknown whether these sources are similar in phenotype and function. METHODS: We have isolated stromal cells from several human tissues including bone marrow (BM-MSCs), heart (heart stroma, HS), adipose (adipose stroma, AS) and liver (liver stroma, LS) and compared the morphology, phenotype and functional properties of these stromal cell populations. RESULTS: The cellular phenotype of each population was identical, namely, CD105+, CD73+, CD90+, CD34- and CD45-. In addition, morphology and differentiation potential were comparable. Co-culture studies revealed similar supportive potential of BM-MSCs, AS and LS with hematopoietic cells or tumor cells. In contrast, significant inhibition of proliferation of both cells types was obtained with HS, with significant loss of viability with tumor cells, demonstrating a unique functional property of HS with regard to tumor cell proliferation. CONCLUSIONS: Although stromal cells from different tissues have similar morphology and phenotype, their functional properties vary, requiring critical evaluation of stromal cells before use in non-homologous settings. HS may play a key role in inhibiting proliferation of tumor cells in the heart, providing the reason for the low occurrence of tumor development. Given the tumor-supportive property of BM-MSCs and AS, the use of these cells in cardiac tissue may result in replacement of a tumor-inhibitory stroma with a tumor-supportive microenvironment.
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Tejido Adiposo/citología , Células de la Médula Ósea/citología , Hígado/citología , Miocardio/citología , Células del Estroma/citología , Antígenos CD/metabolismo , Antígenos CD34/metabolismo , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Técnicas de Cocultivo , Humanos , Células K562 , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , FenotipoRESUMEN
Regenerative medicine offers new hope for many debilitating diseases that result in damage to tissues and organs. The concept is straightforward with replacement of damaged cells with new functional cells. However, most tissues and organs are complex structures involving multiple cell types, supportive structures, a microenvironment producing cytokines and growth factors and a vascular system to supply oxygen and other nutrients. Therefore repair, particularly in the setting of ischemic damage, may require delivery of multiple cell types providing new vessel formation, a new microenvironment and functional cells. The field of stem cell biology has identified a number of stem cell sources including embryonic stem cells and adult stem cells that offer the potential to replace virtually all functional cells of the body. The focus of this article is a discussion of the potential of mesenchymal stromal cells (MSCs) from cord blood (CB) for regenerative medicine approaches.
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Sangre Fetal/citología , Células Madre Mesenquimatosas/citología , Ensayos Clínicos como Asunto , Humanos , Trasplante de Células Madre Mesenquimatosas , Nicho de Células MadreRESUMEN
BACKGROUND AIMS: Hematopoietic stem cell transplantation of mobilized peripheral blood progenitor cell (PBPC) products results in rapid platelet engraftment, whereas the use of cord blood (CB) shows significant delays. The difference in the quality and number of megakaryocyte (MK) progenitors that may be responsible for the delay in platelet engraftment has not been fully defined. The objective of this study was to quantify the cells of the MK lineage in PBPC and CB products to determine whether potential differences exist. METHODS: We examined PBPC or CB for differences in surface markers and subpopulations as well as polyploidization status within the MK lineage. Colony-forming assays were used to determine whether differences exist in the clonogenic MK progenitor cell. Finally, we transplanted PBPC and CB mononuclear cells into NOD/SCID/IL2Rγ-/- (NSG) mice to study platelet engraftment rates. RESULTS: Equivalent MK populations and polyploidization was observed in PBPCs and CB. MK progenitors were present only in CD34+ cells and had little difference in colony growth between PBPC and CB. Additionally, MK subpopulations were similar in either product with a slightly more progenitor-enriched phenotype in CB. Finally, when PBPC or CB was transplanted at similar doses, equivalent platelet engraftment rates were observed. CONCLUSIONS: PBPC and CB contain similar frequencies of MK populations, and, when transplanted in comparable doses, CB is as effective as PBPCs in producing platelet engraftment in vivo. Understanding the differences in MK populations between PBPC and CB could help generate protocols to improve platelet engraftment after CB transplantation.
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Plaquetas/citología , Leucocitos Mononucleares/trasplante , Células Progenitoras de Megacariocitos/citología , Animales , Antígenos CD34/inmunología , Células Cultivadas , Sangre Fetal/citología , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Leucocitos Mononucleares/citología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Transfusión de PlaquetasRESUMEN
BACKGROUND AIMS: Naturally occurring regulatory T cells (Treg) are emerging as a promising approach for prevention of graft-versus-host disease (GvHD), which remains an obstacle to the successful outcome of allogeneic hematopoietic stem cell transplantation. However, Treg only constitute 1-5% of total nucleated cells in cord blood (CB) (<3 × 106 cells), and therefore novel methods of Treg expansion to generate clinically relevant numbers are needed. METHODS: Several methodologies are currently being used for ex vivo Treg expansion. We report a new approach to expand Treg from CB and demonstrate their efficacy in vitro by blunting allogeneic mixed lymphocyte reactions and in vivo by preventing GvHD through the use of a xenogenic GvHD mouse model. RESULTS: With the use of magnetic cell sorting, naturally occurring Treg were isolated from CB by the positive selection of CD25⺠cells. These were expanded to clinically relevant numbers by use of CD3/28 co-expressing Dynabeads and interleukin (IL)-2. Ex vivo-expanded Treg were CD4âº25⺠FOXP3âº127(lo) and expressed a polyclonal T-cell receptor, Vß repertoire. When compared with conventional T-lymphocytes (CD4âº25â» cells), Treg consistently showed demethylation of the FOXP3 TSDR promoter region and suppression of allogeneic proliferation responses in vitro. CONCLUSIONS: In our NOD-SCID IL-2Rγ(null) xenogeneic model of GvHD, prophylactic injection of third-party, CB-derived, ex vivo-expanded Treg led to the prevention of GvHD that translated into improved GvHD score, decreased circulating inflammatory cytokines and significantly superior overall survival. This model of xenogenic GvHD can be used to study the mechanism of action of CB Treg as well as other therapeutic interventions.
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Sangre Fetal/trasplante , Enfermedad Injerto contra Huésped/terapia , Linfocitos T Reguladores/citología , Trasplante Homólogo/efectos adversos , Animales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/trasplante , Proliferación Celular , Tratamiento Basado en Trasplante de Células y Tejidos , Sangre Fetal/citología , Enfermedad Injerto contra Huésped/patología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Humanos , Ratones , Linfocitos T Reguladores/trasplanteRESUMEN
BACKGROUND AIMS: Advantages associated with the use of cord blood (CB) transplantation include the availability of cryopreserved units, ethnic diversity and lower incidence of graft-versus-host disease compared with bone marrow or mobilized peripheral blood. However, poor engraftment remains a major obstacle. We and others have found that ex vivo fucosylation can enhance engraftment in murine models, and now ex vivo treatment of CB with fucosyltransferase (FT) VI before transplantation is under clinical evaluation (NCT01471067). However, FTVII appears to be more relevant to hematopoietic cells and may alter acceptor substrate diversity. The present study compared the ability of FTVI and FTVII to improve the rapidity, magnitude, multi-lineage and multi-tissue engraftment of human CB hematopoietic stem and progenitor cells (HSPCs) in vivo. METHODS: CD34-selected CB HSPCs were treated with recombinant FTVI, FTVII or mock control and then injected into immunodeficient mice and monitored for multi-lineage and multi-tissue engraftment. RESULTS: Both FTVI and FTVII fucosylated CB CD34⺠cells in vitro, and both led to enhanced rates and magnitudes of engraftment compared with untreated CB CD34⺠cells in vivo. Engraftment after treatment with either FT was robust at multiple time points and in multiple tissues with similar multi-lineage potential. In contrast, only FTVII was able to fucosylate T and B lymphocytes. CONCLUSIONS: Although FTVI and FTVII were found to be similarly able to fucosylate and enhance the engraftment of CB CD34⺠cells, differences in their ability to fucosylate lymphocytes may modulate graft-versus-tumor or graft-versus-host effects and may allow further optimization of CB transplantation.
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Sangre Fetal/efectos de los fármacos , Fucosiltransferasas/administración & dosificación , Enfermedad Injerto contra Huésped/terapia , Animales , Modelos Animales de Enfermedad , Sangre Fetal/citología , Sangre Fetal/trasplante , Enfermedad Injerto contra Huésped/patología , Trasplante de Células Madre Hematopoyéticas/métodos , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , RatonesRESUMEN
IMPORTANCE: Whether culture-expanded mesenchymal stem cells or whole bone marrow mononuclear cells are safe and effective in chronic ischemic cardiomyopathy is controversial. OBJECTIVE: To demonstrate the safety of transendocardial stem cell injection with autologous mesenchymal stem cells (MSCs) and bone marrow mononuclear cells (BMCs) in patients with ischemic cardiomyopathy. DESIGN, SETTING, AND PATIENTS: A phase 1 and 2 randomized, blinded, placebo-controlled study involving 65 patients with ischemic cardiomyopathy and left ventricular (LV) ejection fraction less than 50% (September 1, 2009-July 12, 2013). The study compared injection of MSCs (n=19) with placebo (n = 11) and BMCs (n = 19) with placebo (n = 10), with 1 year of follow-up. INTERVENTIONS: Injections in 10 LV sites with an infusion catheter. MAIN OUTCOMES AND MEASURES: Treatment-emergent 30-day serious adverse event rate defined as a composite of death, myocardial infarction, stroke, hospitalization for worsening heart failure, perforation, tamponade, or sustained ventricular arrhythmias. RESULTS: No patient had a treatment-emergent serious adverse events at day 30. The 1-year incidence of serious adverse events was 31.6% (95% CI, 12.6% to 56.6%) for MSCs, 31.6% (95% CI, 12.6%-56.6%) for BMCs, and 38.1% (95% CI, 18.1%-61.6%) for placebo. Over 1 year, the Minnesota Living With Heart Failure score improved with MSCs (-6.3; 95% CI, -15.0 to 2.4; repeated measures of variance, P=.02) and with BMCs (-8.2; 95% CI, -17.4 to 0.97; P=.005) but not with placebo (0.4; 95% CI, -9.45 to 10.25; P=.38). The 6-minute walk distance increased with MSCs only (repeated measures model, P = .03). Infarct size as a percentage of LV mass was reduced by MSCs (-18.9%; 95% CI, -30.4 to -7.4; within-group, P = .004) but not by BMCs (-7.0%; 95% CI, -15.7% to 1.7%; within-group, P = .11) or placebo (-5.2%; 95% CI, -16.8% to 6.5%; within-group, P = .36). Regional myocardial function as peak Eulerian circumferential strain at the site of injection improved with MSCs (-4.9; 95% CI, -13.3 to 3.5; within-group repeated measures, P = .03) but not BMCs (-2.1; 95% CI, -5.5 to 1.3; P = .21) or placebo (-0.03; 95% CI, -1.9 to 1.9; P = .14). Left ventricular chamber volume and ejection fraction did not change. CONCLUSIONS AND RELEVANCE: Transendocardial stem cell injection with MSCs or BMCs appeared to be safe for patients with chronic ischemic cardiomyopathy and LV dysfunction. Although the sample size and multiple comparisons preclude a definitive statement about safety and clinical effect, these results provide the basis for larger studies to provide definitive evidence about safety and to assess efficacy of this new therapeutic approach. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00768066.
Asunto(s)
Trasplante de Médula Ósea/métodos , Trasplante de Células Madre Mesenquimatosas/métodos , Isquemia Miocárdica/terapia , Anciano , Trasplante de Médula Ósea/efectos adversos , Cardiomiopatías , Progresión de la Enfermedad , Método Doble Ciego , Femenino , Hospitalización , Humanos , Masculino , Trasplante de Células Madre Mesenquimatosas/efectos adversos , Persona de Mediana Edad , Infarto del Miocardio , Accidente Cerebrovascular , Análisis de Supervivencia , Trasplante Autólogo , Resultado del Tratamiento , Disfunción Ventricular Izquierda/terapiaRESUMEN
BACKGROUND: Mesenchymal stem cell (MSC) therapy may prevent neonatal hyperoxia-induced lung injury (HILI). There are, however, no clear data on the therapeutic efficacy of MSC therapy in established HILI, the duration of the reparative effects, and the exact mechanisms of repair. The main objective of this study was to evaluate whether the long-term reparative effects of a single intratracheal (IT) dose of MSCs or MSC-conditioned medium (CM) are comparable in established HILI. METHODS: Newborn rats exposed to normoxia or hyperoxia from postnatal day (P)2)-P16 were randomized to receive IT MSCs, IT CM, or IT placebo (PL) on P9. Alveolarization and angiogenesis were evaluated at P16, P30, and P100. RESULTS: At all time periods, there were marked improvements in alveolar and vascular development in hyperoxic pups treated with MSCs or CM as compared with PL. This was associated with decreased expression of inflammatory mediators and an upregulation of angiogenic factors. Of note, at P100, the improvements were more substantial with MSCs as compared with CM. CONCLUSION: These data suggest that acute effects of MSC therapy in HILI are mainly paracrine mediated; however, optimum long-term improvement following HILI requires treatment with the MSCs themselves or potentially repetitive administration of CM.
Asunto(s)
Medios de Cultivo Condicionados/farmacología , Hiperoxia/complicaciones , Lesión Pulmonar/tratamiento farmacológico , Lesión Pulmonar/terapia , Trasplante de Células Madre Mesenquimatosas/métodos , Análisis de Varianza , Animales , Animales Recién Nacidos , Femenino , Hipertrofia Ventricular Derecha/patología , Lesión Pulmonar/etiología , Masculino , Neovascularización Fisiológica/efectos de los fármacos , Alveolos Pulmonares/efectos de los fármacos , Alveolos Pulmonares/crecimiento & desarrollo , Ratas , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: Stem cell factor (SCF) and its receptor, c-kit, are modulators of angiogenesis. Neonatal hyperoxia-induced lung injury (HILI) is characterized by disordered angiogenesis. The objective of this study was to determine whether exogenous SCF improves recovery from neonatal HILI by improving angiogenesis. METHODS: Newborn rats assigned to normoxia (RA: 20.9% O2) or hyperoxia (90% O2) from postnatal day (P) 2 to 15, received daily injections of SCF 100 µg/kg or placebo (PL) from P15 to P21. Lung morphometry was performed at P28. Capillary tube formation in SCF-treated hyperoxia-exposed pulmonary microvascular endothelial cells (HPMECs) was determined by Matrigel assay. RESULTS: As compared with RA, hyperoxic-PL pups had decrease in alveolarization and in lung vascular density, and this was associated with increased right ventricular systolic pressure (RVSP), right ventricular hypertrophy, and vascular remodeling. In contrast, SCF-treated hyperoxic pups had increased angiogenesis, improved alveolarization, and attenuation of pulmonary hypertension as evidenced by decreased RVSP, right ventricular hypertrophy, and vascular remodeling. Moreover, in an in vitro model, SCF increased capillary tube formation in hyperoxia-exposed HPMECs. CONCLUSION: Exogenous SCF restores alveolar and vascular structure in neonatal rats with HILI by promoting neoangiogenesis. These findings suggest a new strategy to treat lung diseases characterized by dysangiogenesis.
Asunto(s)
Hiperoxia/tratamiento farmacológico , Lesión Pulmonar/tratamiento farmacológico , Factor de Células Madre/uso terapéutico , Animales , Animales Recién Nacidos , Hiperoxia/fisiopatología , Lesión Pulmonar/fisiopatología , Neovascularización Fisiológica/efectos de los fármacos , Placebos , Alveolos Pulmonares/efectos de los fármacos , Alveolos Pulmonares/crecimiento & desarrollo , Ratas , Ratas Sprague-Dawley , Factor de Células Madre/farmacologíaRESUMEN
RATIONALE: Transcatheter, intramyocardial injections of bone marrow-derived cell therapy produces reverse remodeling in large animal models of ischemic cardiomyopathy. OBJECTIVE: We used cardiac MRI (CMR) in patients with left ventricular (LV) dysfunction related to remote myocardial infarction (MI) to test the hypothesis that bone marrow progenitor cell injection causes functional recovery of scarred myocardium and reverse remodeling. METHODS AND RESULTS: Eight patients (aged 57.2±13.3 years) received transendocardial, intramyocardial injection of autologous bone marrow progenitor cells (mononuclear or mesenchymal stem cells) in LV scar and border zone. All patients tolerated the procedure with no serious adverse events. CMR at 1 year demonstrated a decrease in end diastolic volume (208.7±20.4 versus 167.4±7.32 mL; P=0.03), a trend toward decreased end systolic volume (142.4±16.5 versus 107.6±7.4 mL; P=0.06), decreased infarct size (P<0.05), and improved regional LV function by peak Eulerian circumferential strain in the treated infarct zone (-8.1±1.0 versus -11.4±1.3; P=0.04). Improvements in regional function were evident at 3 months, whereas the changes in chamber dimensions were not significant until 6 months. Improved regional function in the infarct zone strongly correlated with reduction of end diastolic volume (r(2)=0.69, P=0.04) and end systolic volume (r(2)=0.83, P=0.01). CONCLUSIONS: These data suggest that transcatheter, intramyocardial injections of autologous bone marrow progenitor cells improve regional contractility of a chronic myocardial scar, and these changes predict subsequent reverse remodeling. The findings support the potential clinical benefits of this new treatment strategy and ongoing randomized clinical trials.
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Trasplante de Células Madre Mesenquimatosas/métodos , Recuperación de la Función/fisiología , Disfunción Ventricular Izquierda/fisiopatología , Disfunción Ventricular Izquierda/cirugía , Remodelación Ventricular/fisiología , Adulto , Anciano , Humanos , Inyecciones , Imagen por Resonancia Magnética , Masculino , Trasplante de Células Madre Mesenquimatosas/efectos adversos , Persona de Mediana Edad , Contracción Miocárdica/fisiología , Infarto del Miocardio/complicaciones , Proyectos Piloto , Resultado del Tratamiento , Disfunción Ventricular Izquierda/etiologíaRESUMEN
RATIONALE: The regenerative potential of the heart is insufficient to fully restore functioning myocardium after injury, motivating the quest for a cell-based replacement strategy. Bone marrow-derived mesenchymal stem cells (MSCs) have the capacity for cardiac repair that appears to exceed their capacity for differentiation into cardiac myocytes. OBJECTIVE: Here, we test the hypothesis that bone marrow derived MSCs stimulate the proliferation and differentiation of endogenous cardiac stem cells (CSCs) as part of their regenerative repertoire. METHODS AND RESULTS: Female Yorkshire pigs (n=31) underwent experimental myocardial infarction (MI), and 3 days later, received transendocardial injections of allogeneic male bone marrow-derived MSCs, MSC concentrated conditioned medium (CCM), or placebo (Plasmalyte). A no-injection control group was also studied. MSCs engrafted and differentiated into cardiomyocytes and vascular structures. In addition, endogenous c-kit(+) CSCs increased 20-fold in MSC-treated animals versus controls (P<0.001), there was a 6-fold increase in GATA-4(+) CSCs in MSC versus control (P<0.001), and mitotic myocytes increased 4-fold (P=0.005). Porcine endomyocardial biopsies were harvested and plated as organotypic cultures in the presence or absence of MSC feeder layers. In vitro, MSCs stimulated c-kit(+) CSCs proliferation into enriched populations of adult cardioblasts that expressed Nkx2-5 and troponin I. CONCLUSIONS: MSCs stimulate host CSCs, a new mechanism of action underlying successful cell-based therapeutics.