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1.
J Clin Oncol ; : JCO1902408, 2020 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-32031876

RESUMO

PURPOSE: To compare the outcomes of patients with Hodgkin or non-Hodgkin lymphoma undergoing nonmyeloablative haploidentical or unrelated cord blood (UCB) hematopoietic cell transplantation. PATIENTS AND METHODS: We retrospectively studied 740 patients with Hodgkin lymphoma (n = 283, 38%) and non-Hodgkin lymphoma (n = 457, 62%) age 18-75 years who received transplantations from 2009 to 2016. Data were reported to the Lymphoma Working Party of the European Society for Blood and Marrow Transplantation, Eurocord, or Center for International Blood and Marrow Transplant Research. Of the 526 patients who received haploidentical transplantation, 68% received bone marrow and 32% received peripheral blood. All patients received a uniform transplantation conditioning regimen (2 Gy of total-body irradiation, cyclophosphamide, and fludarabine) and graft-versus-host disease prophylaxis (calcineurin inhibitor and mycophenolate). In addition, patients who received a haploidentical transplantation received posttransplantation cyclophosphamide. RESULTS: Compared with haploidentical bone marrow and peripheral-blood transplantations and adjusted for age, lymphoma subtype, and disease status, survival was lower after UCB transplantation (hazard ratio [HR], 1.55; P = .001; and HR, 1.59; P = .005, respectively). Similarly, progression-free survival was lower after UCB transplantations compared with haploidentical bone marrow and peripheral-blood transplantations (HR, 1.44; P = .002; and HR, 1.86; P < .0001), respectively. The 4-year overall and progression-free survival rates after UCB transplantation were 49% and 36%, respectively, compared with 58% and 46% after haploidentical bone marrow transplantation and 59% and 52% after peripheral-blood transplantation, respectively. Lower survival was attributed to higher transplantation-related mortality after UCB transplantation compared with haploidentical bone marrow and peripheral-blood transplantation (HR, 1.91; P = .0001; and HR, 2.27; P = .0002, respectively). CONCLUSION: When considering HLA-mismatched transplantation for Hodgkin or non-Hodgkin lymphoma, the data support haploidentical related donor transplantation over UCB transplantation.

2.
Cytotherapy ; 22(1): 1-5, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31866320

RESUMO

In the current article we summarize the 15-year experience of the Spanish Cell Therapy Network (TerCel), a successful collaborative public initiative funded by the Spanish government for the support of nationwide translational research in this important area. Thirty-two research groups organized in three programs devoted to cardiovascular, neurodegenerative and immune-inflammatory diseases, respectively, currently form the network. Each program has three working packages focused on basic science, pre-clinical studies and clinical application. TerCel has contributed during this period to boost the translational research in cell therapy in Spain, setting up a network of Good Manufacturing Practice-certified cell manufacturing facilities- and increasing the number of translational research projects, publications, patents and clinical trials of the participating groups, especially those in collaboration. TerCel pays particular attention to the public-private collaboration, which, for instance, has led to the development of the first allogeneic cell therapy product approved by the European Medicines Agency, Darvadstrocel. The current collaborative work is focused on the development of multicenter phase 2 and 3 trials that could translate these therapies to clinical practice for the benefit of patients.

3.
Tissue Eng Regen Med ; 16(5): 525-538, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31624707

RESUMO

Background: Mesenchymal stromal cells (MSCs) have potent immunomodulatory and neuroprotective properties, and have been tested in neurodegenerative diseases resulting in meaningful clinical improvements. Regulatory guidelines specify the need to perform preclinical studies prior any clinical trial, including biodistribution assays and tumourigenesis exclusion. We conducted a preclinical study of human bone marrow MSCs (hBM-MSCs) injected by intrathecal route in Non-Obese Diabetic Severe Combined Immunodeficiency mice, to explore cellular biodistribution and toxicity as a privileged administration method for cell therapy in Friedreich's Ataxia. Methods: For this purpose, 3 × 105 cells were injected by intrathecal route in 12 animals (experimental group) and the same volume of culture media in 6 animals (control group). Blood samples were collected at 24 h (n = 9) or 4 months (n = 9) to assess toxicity, and nine organs were harvested for histology and safety studies. Genomic DNA was isolated from all tissues, and mouse GAPDH and human ß2M and ß-actin genes were amplified by qPCR to analyze hBM-MSCs biodistribution. Results: There were no deaths nor acute or chronic toxicity. Hematology, biochemistry and body weight were in the range of normal values in all groups. At 24 h hBM-MSCs were detected in 4/6 spinal cords and 1/6 hearts, and at 4 months in 3/6 hearts and 1/6 brains of transplanted mice. No tumours were found. Conclusion: This study demonstrated that intrathecal injection of hBM-MSCs is safe, non toxic and do not produce tumors. These results provide further evidence that hBM-MSCs might be used in a clinical trial in patients with FRDA.

4.
Proc Natl Acad Sci U S A ; 116(41): 20655-20665, 2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31548426

RESUMO

The contractile perivascular cells, pericytes (PC), are hijacked by glioblastoma (GB) to facilitate tumor progression. PC's protumorigenic function requires direct interaction with tumor cells and contributes to the establishment of immunotolerance to tumor growth. Cancer cells up-regulate their own chaperone-mediated autophagy (CMA), a process that delivers selective cytosolic proteins to lysosomes for degradation, with pro-oncogenic effects. However, the possible impact that cancer cells may have on CMA of surrounding host cells has not been explored. We analyzed the contribution of CMA to the GB-induced changes in PC biology. We have found that CMA is markedly up-regulated in PC in response to the oxidative burst that follows PC-GB cell interaction. Genetic manipulations to block the GB-induced up-regulation of CMA in PC allows them to maintain their proinflammatory function and to support the induction of effective antitumor T cell responses required for GB clearance. GB-induced up-regulation of CMA activity in PC is essential for their effective interaction with GB cells that help tumor growth. We show that CMA inhibition in PC promotes GB cell death and the release of high immunogenic levels of granulocyte-macrophage colony stimulating factor (GM-CSF), through deregulation of the expression of cell-to-cell interaction proteins and protein secretion. A GB mouse model grafted in vivo with CMA-defective PC shows reduced GB proliferation and effective immune response compared to mice grafted with control PC. Our findings identify abnormal up-regulation of CMA as a mechanism by which GB cells elicit the immunosuppressive function of PC and stabilize GB-PC interactions necessary for tumor cell survival.

5.
Blood ; 134(16): 1337-1345, 2019 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-31484647

RESUMO

Achieving and maintaining a high-quality response is the treatment goal for patients with newly diagnosed multiple myeloma (NDMM). The phase 3 PETHEMA/GEM2012 study, in 458 patients aged ≤65 years with NDMM, is evaluating bortezomib (subcutaneous) + lenalidomide + dexamethasone (VRD) for 6 cycles followed by autologous stem cell transplant (ASCT) conditioned with IV busulfan + melphalan vs melphalan and posttransplant consolidation with 2 cycles of VRD. We present grouped response analysis of induction, transplant, and consolidation. Responses deepened over time; in patients who initiated cycle 6 of induction (n = 426), the rates of a very good partial response or better were 55.6% by cycle 3, 63.8% by cycle 4, 68.3% by cycle 5, and 70.4% after induction. The complete response rate of 33.4% after induction in the intent-to-treat (ITT) population, which was similar in the 92 patients with high-risk cytogenetics (34.8%), also deepened with further treatment (44.1% after ASCT and 50.2% after consolidation). Rates of undetectable minimal residual disease (median 3 × 10-6 sensitivity) in the ITT population also increased from induction (28.8%) to transplant (42.1%) and consolidation (45.2%). The most common grade ≥3 treatment-emergent adverse events during induction were neutropenia (12.9%) and infection (9.2%). Grade ≥2 peripheral neuropathy (grouped term) during induction was 17.0%, with a low frequency of grade 3 (3.7%) and grade 4 (0.2%) events. VRD is an effective and well-tolerated regimen for induction in NDMM with deepening response throughout induction and over the course of treatment. This trial was registered at www.clinicaltrials.gov as #NCT01916252 and EudraCT as #2012-005683-10.

6.
J Clin Med ; 8(8)2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31416236

RESUMO

Background: The aim of this study was: to evaluate the biological properties of new hydraulic materials: Bio-C Repair and Bio-C Sealer. Methods: Periodontal ligament stem cells were exposed to several dilutions of Bio-C Repair and Bio-C Sealer. The ion release profile and pH were determined. Metabolic activity, cell migration and cell survival were assessed using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), wound-healing assays and Annexin assays, respectively. Cells were cultured in direct contact with the surface of each material. These were then analyzed via scanning electron microscopy (SEM) and energy dispersive X-ray (EDX). Statistical differences were assessed using a two-way ANOVA (α < 0.05). Results: Similar pH was observed in these cements. Bio-C Sealer released significantly more Ca and Si ions (p < 0.05) in comparison with Bio-C Repair. Undiluted Bio-C Sealer induced a significant reduction on cellular viability, cell survival and cell migration when compared to the control (p < 0.05). Moreover, SEM showed abundant cells adhered on Bio-C Repair and a moderate number of cells attached on Bio-C Sealer. Finally, EDX analysis identified higher percentages of Ca and O in the case of Bio-C repair than with Bio-C sealer, while other elements such as Zr and Si were more abundant in Bio-C sealer. Conclusions: Bio-C Repair displayed higher cell viability, cell adhesion and migration rates than Bio-C Sealer.

7.
Hematol Oncol Stem Cell Ther ; 12(4): 194-203, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31319058

RESUMO

OBJECTIVE/BACKGROUND: Patients with follicular lymphoma (FL) with early therapy failure (ETF) within 2 years of frontline therapy have poor overall survival (OS). We recently reported the results of autologous stem cell transplantation (ASCT) in patients from the Grupo Español de Linfomas y Trasplantes de Médula Ósea (GELTAMO) registry treated with rituximab prior to ASCT and with ETF after first-line immunochemotherapy, leading to 81% 5-year OS since ASCT. We explored whether ASCT is also an effective option in the pre-rituximab era-that is, in patients treated in induction and rescued only with chemotherapy. METHODS: ETF was defined as relapse/progression within 2 years of starting first-line therapy. We identified two groups: the ETF cohort (n = 87) and the non-ETF cohort (n = 47 patients receiving ASCT but not experiencing ETF following first-line therapy). RESULTS: There was a significant difference in 5-year progression-free survival between the ETF and non-ETF cohorts (43% vs. 57%, respectively; p = .048). Nevertheless, in patients with ETF with an interval from first relapse after primary treatment to ASCT of <1 year, no differences were observed in 5-year progression-free survival (48% vs. 66%, respectively; p = .44) or in 5-year OS (69% vs. 77%, p = .4). Patients in the ETF cohort transplanted in complete remission showed a plateau in the OS curves, at 56%, beyond 13.7 years of follow-up. CONCLUSION: ASCT may be a curative option for ETF in patients who respond to rescue chemotherapy, without the need for immunotherapy or other therapies, and should be considered as an early consolidation, especially in patients with difficult access to rituximab.

8.
Front Immunol ; 10: 1151, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31231366

RESUMO

Cell therapy is a progressively growing field that is rapidly moving from preclinical model development to clinical application. Outcomes obtained from clinical trials reveal the therapeutic potential of stem cell-based therapy to deal with unmet medical treatment needs for several disorders with no therapeutic options. Among adult stem cells, mesenchymal stem cells (MSCs) are the leading cell type used in advanced therapies for the treatment of autoimmune, inflammatory and vascular diseases. To date, the safety and feasibility of autologous MSC-based therapy has been established; however, their indiscriminate use has resulted in mixed outcomes in preclinical and clinical studies. While MSCs derived from diverse tissues share common properties depending on the type of clinical application, they markedly differ within clinical trials in terms of efficacy, resulting in many unanswered questions regarding the application of MSCs. Additionally, our experience in clinical trials related to critical limb ischemia pathology (CLI) shows that the therapeutic efficacy of these cells in different animal models has only been partially reproduced in humans through clinical trials. Therefore, it is crucial to develop new research to identify pitfalls, to optimize procedures and to clarify the repair mechanisms used by these cells, as well as to be able to offer a next generation of stem cell that can be routinely used in a cost-effective and safe manner in stem cell-based therapies targeting CLI.

9.
Biol Blood Marrow Transplant ; 25(9): 1786-1791, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31082473

RESUMO

Gonadal impairment is an important late effect with a significant impact on quality of life of transplanted patients. The aim of this study was to compare gonadal function after busulfan (Bu) or treosulfan (Treo) conditioning regimens in pre- and postpubertal children. This retrospective, multicenter study included children transplanted in pediatric European Society for Blood and Marrow Transplantation (EBMT) centers between 1992 and 2012 who did not receive gonadotoxic chemoradiotherapy before the transplant. We evaluated 137 patients transplanted in 25 pediatric EBMT centers. Median age at transplant was 11.04 years (range, 5 to 18); 89 patients were boys and 48 girls. Eighty-nine patients were prepubertal at transplant and 48 postpubertal. One hundred eighteen children received Bu and 19 Treo. A higher proportion of girls treated with Treo in the prepubertal stage reached spontaneous puberty compared with those treated with Bu (P = .02). Spontaneous menarche was more frequent after Treo than after Bu (P < .001). Postpubertal boys and girls treated with Treo had significantly lower luteinizing hormone levels (P = .03 and P = .04, respectively) compared with the Bu group. Frequency of gonadal damage associated with Treo was significantly lower than that observed after Bu. These results need to be confirmed in a larger population.

10.
Stem Cell Res Ther ; 10(1): 126, 2019 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-31029166

RESUMO

BACKGROUND: The treatment of extensive and/or chronic skin wounds is a widespread and costly public health problem. Mesenchymal stem cells (MSCs) have been proposed as a potential cell therapy for inducing wound healing in different clinical settings, alone or in combination with biosynthetic scaffolds. Among them, silk fibroin (SF) seeded with MSCs has been shown to have increased efficacy in skin wound healing experimental models. METHODS: In this report, we investigated the wound healing effects of electrospun SF scaffolds cellularized with human Wharton's jelly MSCs (Wj-MSCs-SF) using a murine excisional wound splinting model. RESULTS: Immunohistopathological examination after transplant confirmed the presence of infiltrated human fibroblast-like CD90-positive cells in the dermis of the Wj-MSCs-SF-treated group, yielding neoangiogenesis, decreased inflammatory infiltrate and myofibroblast proliferation, less collagen matrix production, and complete epidermal regeneration. CONCLUSIONS: These findings indicate that Wj-MSCs transplanted in the wound bed on a silk fibroin scaffold contribute to the generation of a well-organized and vascularized granulation tissue, enhance reepithelization of the wound, and reduce the formation of fibrotic scar tissue, highlighting the potential therapeutic effects of Wj-MSC-based tissue engineering approaches to non-healing wound treatment.

11.
Br J Haematol ; 186(2): 263-268, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30916384

RESUMO

The prognostic significance of low-hypodiploidy has not been extensively evaluated in minimal residual disease (MRD)-oriented protocols for adult acute lymphoblastic leukaemia (ALL). We analysed the outcome of hypodiploid adult ALL patients treated within Programa Español de Tratamientos en Hematología (PETHEMA) protocols. The 5-year cumulative incidence of relapse (CIR) of low-hypodiploid B-cell precursor (BCP)-ALL was significantly higher than that of high-hypodiploids (52% vs. 12%, P = 0.013). Low-hypodiploid BCP-ALL patients aged ≤35 years showed superior survival (71% vs. 21%, P = 0.026) and lower 5-year CIR (17% vs. 66%, P = 0.090) than low-hypodiploids aged >35 years. Older adults and elderly low-hypodiploid BCP-ALL patients show dismal prognosis although achieving an end-induction good MRD response.

12.
Clin Oral Investig ; 23(10): 3915-3924, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30684060

RESUMO

OBJECTIVE: The aim of this study was to analyze the biological effects of MTA Repair HP and ProRoot MTA on human periodontal ligament stem cells (hPDLSCs) after exposure to acidic and neutral environments. MATERIALS AND METHODS: Discs of each material (n = 30) were exposed to phosphate buffered saline (pH = 7.4) or butyric acid (pH = 5.2) for 7 days, and biological testing was carried out in vitro on hPDLSCs. Cell viability and apoptosis assays were performed using eluates of each root-end filling material. To evaluate cell attachment to the different materials, hPDLSCs were directly seeded onto the material surfaces and analyzed by scanning electron microscopy. The chemical composition of the root-end filling materials was determined by energy-dispersive x-ray and eluates were analyzed by inductively coupled plasma-mass spectrometry. Statistical differences were assessed by ANOVA and Tukey test (p < 0.05). RESULTS: Under an acidic environment, both materials displayed similar ion release abilities, with the increased release of Si and Ca ions. Substantial changes in microstructure were observed for both materials after exposure to acidic pH. In addition, material exposure to an acidic environment showed a similar degree of cell adherence, and, surprisingly, MTA Repair HP exhibited higher cell viability rates at pH 5.2 than ProRoot MTA. CONCLUSIONS: Exposure to an acidic environment promoted Si and Ca ion release from ProRoot MTA and MTA Repair HP. Moreover, we observed optimal biological properties of ProRoot MTA and MTA Repair HP in terms of cell viability, cell death, and cell attachment in both environments. CLINICAL RELEVANCE: These results may suggest that MTA Repair HP and ProRoot exhibited optimal biological properties in terms of cell viability, cell death and cell attachment in acidic environment, being considered as materials for root-end filling and perforations.


Assuntos
Pemetrexede , Ligamento Periodontal/citologia , Materiais Restauradores do Canal Radicular , Células-Tronco/efeitos dos fármacos , Compostos de Cálcio , Células Cultivadas , Combinação de Medicamentos , Humanos , Teste de Materiais , Óxidos , Silicatos
13.
Biol Blood Marrow Transplant ; 25(2): 335-342, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30243581

RESUMO

Allogeneic hematopoietic cell transplantation (allo-HCT) using siblings and matched donors has the potential for long-term disease control in a subset of high-risk patients with multiple myeloma (MM); however, the data on using haploidentical donors in this disease are limited. We conducted a retrospective analysis to examine the outcomes of patients with MM who underwent haploidentical allo-HCT within European Society for Blood and Marrow Transplantation/Center for International Blood and Marrow Transplant Research centers. A total of 96 patients underwent haploidentical allo-HCT between 2008 and 2016. With a median follow-up of 24.0 months (range, 13.2 to 24.9 months), 97% (95% confidence interval [CI], 93% to 100%) of patients had neutrophil engraftment by day 28, and 75% (95% CI, 66% to 84%) achieved platelet recovery by day 60. Two-year progression-free survival (PFS) was 17% (95% CI, 8% to 26%), and overall survival (OS) was 48% (95% CI, 36% to 59%). At 2 years, the cumulative risk of relapse/progression was 56% (95% CI, 45% to 67%), and 1-year nonrelapse mortality (NRM) was 21% (95% CI, 13% to 29%). The incidences of acute graft-versus-host-disease (GVHD) grades II-IV by 100 days and chronic GVHD at 2 years were 39% (95% CI, 28% to 49%) and 46% (95% CI, 34% to 59%), respectively. On univariate analysis, use of post-transplantation cyclophosphamide (PT-Cy) (54% [95% CI, 41% to 68%] versus 25% [95% CI, 1% to 48%]; P =.009) and use of bone marrow as source of stem cells (72% [95% CI, 55% to 89%] versus 31% [95% CI, 17% to 46%]; P = .001) were associated with improved OS at 2 years. Disease status, patient sex, intensity of conditioning regimen, recipient/donor sex mismatch, and cytomegalovirus serostatus had no impact on OS, PFS, or NRM. Haploidentical transplantation is feasible for patients with multiply relapsed or high-risk MM, with an encouraging 2-year OS of 48% and an NRM of 21% at 1 year, supporting further investigation of haploidentical allo-HCT in suitable candidates with MM.


Assuntos
Doença Enxerto-Hospedeiro/mortalidade , Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo/mortalidade , Mieloma Múltiplo/veterinária , Adulto , Idoso , Aloenxertos , Intervalo Livre de Doença , Feminino , Seguimentos , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Recidiva , Estudos Retrospectivos , Fatores de Risco , Taxa de Sobrevida
14.
J Wound Care ; 27(12): 806-815, 2018 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-30557111

RESUMO

OBJECTIVE: The amniotic membrane (AM) is a tissue with low immunogenity and high therapeutic potential due to its anti-inflammatory, anti-fibrotic and antimicrobial effects. This paper describes the use of cryopreserved amniotic membrane allografts to treat diabetic foot ulcers (DFUs) in patients with diabetes. METHOD: In this case series, AM was processed to obtain a final medicinal product: cryopreserved amniotic membrane. cryopreserved AM was applied every 7-10 days until total epithelialisation of the DFUs. RESULTS: A total of 14 patients with DFUs (median size: 12.30cm, (range: 0.52-42.5cm2) were treated and followed up until complete closure (median time: 20 weeks, range: 7-56 weeks). Patients received 4-40 AM applications. All patients in this study achieved complete epithelialisation of the wound. No adverse events were observed. CONCLUSION: AM is a feasible and safe treatment in complex DFUs. Furthermore, the treatment is successful in achieving epithelialisation of long-evolution, unhealed wounds resistant to conventional therapies.


Assuntos
Aloenxertos/transplante , Âmnio/transplante , Criopreservação/métodos , Pé Diabético/cirurgia , Cicatrização/fisiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Espanha , Resultado do Tratamento , Adulto Jovem
15.
Sci Rep ; 8(1): 16299, 2018 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-30389962

RESUMO

Mesenchymal stem cell (MSC) transplantation is emerging as an ideal tool to restore the wounded central nervous system (CNS). MSCs isolated from extra-embryonic tissues have some advantages compared to MSCs derived from adult ones, such as an improved proliferative capacity, life span, differentiation potential and immunomodulatory properties. In addition, they are more immunoprivileged, reducing the probability of being rejected by the recipient. Umbilical cords (UCs) are a good source of MSCs because they are abundant, safe, non-invasively harvested after birth and, importantly, they are not encumbered with ethical problems. Here we show that the intravitreal transplant of Wharton´s jelly mesenchymal stem cells isolated from three different human UCs (hWJMSCs) delays axotomy-induced retinal ganglion cell (RGC) loss. In vivo, hWJMSCs secrete anti-inflammatory molecules and trophic factors, the latter alone may account for the elicited neuroprotection. Interestingly, this expression profile differs between naive and injured retinas, suggesting that the environment in which the hWJMSCs are modulates their secretome. Finally, even though the transplant itself is not toxic for RGCs, it is not innocuous as it triggers a transient but massive infiltration of Iba1+cells from the choroid to the retina that alters the retinal structure.


Assuntos
Transplante de Células-Tronco Mesenquimais/métodos , Degeneração Retiniana/terapia , Células Ganglionares da Retina/patologia , Geleia de Wharton/citologia , Animais , Anti-Inflamatórios/metabolismo , Axotomia/efeitos adversos , Modelos Animais de Doenças , Feminino , Humanos , Injeções Intravítreas , Células-Tronco Mesenquimais/metabolismo , Fatores de Crescimento Neural/metabolismo , Ratos , Ratos Sprague-Dawley , Degeneração Retiniana/etiologia , Degeneração Retiniana/patologia , Resultado do Tratamento , Cordão Umbilical/citologia
16.
Cytotherapy ; 20(9): 1110-1123, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30170815

RESUMO

BACKGROUND: The regenerative and immunomodulatory properties of human mesenchymal stromal cells (hMSCs) have raised great hope for their use in cell therapy. However, when intravenously infused, hMSCs fail to reach sites of tissue injury. Fucose addition in α(1,3)-linkage to terminal sialyllactosamines on CD44 creates the molecule known as hematopoietic cell E-/L-selectin ligand (HCELL), programming hMSC binding to E-selectin that is expressed on microvascular endothelial cells of bone marrow (BM), skin and at all sites of inflammation. Here we describe how this modification on BM-derived hMSCs (BM-hMSCs) can be adapted to good manufacturing practice (GMP) standards. METHODS: BM-hMSCs were expanded using xenogenic-free media and exofucosylated using α(1,3)-fucosyltransferases VI (FTVI) or VII (FTVII). Enforced fucosylation converted CD44 into HCELL, and HCELL formation was assessed using Western blot, flow cytometry and cell-binding assays. Untreated (unfucosylated), buffer-treated and exofucosylated BM-hMSCs were each analyzed for cell viability, immunophenotype and differentiation potential, and E-selectin binding stability was assessed at room temperature, at 4°C, and after cryopreservation. Cell product safety was evaluated using microbiological testing, karyotype analysis, and c-Myc messenger RNA (mRNA) expression, and potential effects on genetic reprogramming and in cell signaling were analyzed using gene expression microarrays and receptor tyrosine kinase (RTK) phosphorylation arrays. RESULTS: Our protocol efficiently generates HCELL on clinical-scale batches of BM-hMSCs. Exofucosylation yields stable HCELL expression for 48 h at 4°C, with retained expression after cell cryopreservation. Cell viability and identity are unaffected by exofucosylation, without changes in gene expression or RTK phosphorylation. DISCUSSION: The described exofucosylation protocol using xenogenic-free reagents enforces HCELL expression on hMSCs endowing potent E-selectin binding without affecting cell viability or native phenotype. This described protocol is readily scalable for GMP-compliant clinical production.


Assuntos
Biotecnologia/métodos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Biotecnologia/normas , Diferenciação Celular , Sobrevivência Celular , Células Cultivadas , Criopreservação , Selectina E/metabolismo , Células Endoteliais/metabolismo , Fucose/metabolismo , Fucosiltransferases/metabolismo , Glicosilação , Humanos , Receptores de Hialuronatos/metabolismo , Imunofenotipagem , Transcriptoma
17.
Materials (Basel) ; 11(8)2018 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-30081464

RESUMO

Background: Cell-Based Therapies (CBT) constitute a valid procedure for increasing the quantity and quality of bone in areas with an inadequate bone volume. However, safety and efficacy should be investigated prior to clinical application. The objective of this study was to evaluate the biodistribution, safety and osteogenic capacity of bone marrow-derived human mesenchymal stem cells (hBMMSCs) pre-seeded into ß-tricalcium phosphate (TCP) and implanted into NOD/SCID mice at subcutaneous and intramuscular sites. Methods: hBMMSCs were isolated, characterized and then cultured in vitro on a porous ß-TCP scaffold. Cell viability and attachment were analyzed and then hBMMSCs seeded constructs were surgically placed at subcutaneous and intramuscular dorsal sites into NOD/SCID mice. Acute and subchronic toxicity, cell biodistribution and efficacy were investigated. Results: There were no deaths or adverse events in treated mice during the 48-hour observation period, and no toxic response was observed in mice. In the 12-week subchronic toxicity study, no mortalities, abnormal behavioral symptoms or clinical signs were observed in the saline control mice or the hBMMSCs/ß-TCP groups. Finally, our results showed the bone-forming capacity of hBMMSCs/ß-TCP since immunohistochemical expression of human osteocalcin was detected from week 7. Conclusions: These results show that transplantation of hBMMSCs/ß-TCP in NOD/SCID mice are safe and effective, and might be applied to human bone diseases in future clinical trials.

18.
Am J Hematol ; 93(7): 867-873, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29658143

RESUMO

The means of optimally managing very elderly patients with diffuse large B-cell lymphoma (DLBCL) has not been established. We retrospectively analyzed 252 patients aged 80-100 years, diagnosed with DLBCL or grade 3B follicular lymphoma, treated in 19 hospitals from the GELTAMO group. Primary objective was to analyze the influence of the type of treatment and comorbidity scales on progression-free survival (PFS) and overall survival (OS). One hundred sixty-three patients (63%) were treated with chemotherapy that included anthracyclines and/or rituximab, whereas 15% received no chemotherapeutic treatment. With a median follow-up of 44 months, median PFS and OS were 9.5 and 12.5 months, respectively. In an analysis restricted to the 205 patients treated with any kind of chemotherapy, comorbidity scales did not influence the choice of treatment type significantly. Independent factors associated with better PFS and OS were: age < 86 years, cumulative illness rating scale (CIRS) score < 6, intermediate risk (1-2) R-IPI, and treatment with R-CHOP at full or reduced doses. We developed a prognostic model based on the multivariate analysis of the 108 patients treated with R-CHOP-like: median OS was 45 vs. 12 months (P = .001), respectively, for patients with 0-1 vs. 2-3 risk factors (age > 85 years, R-IPI 3-5 or CIRS > 5). In conclusion, treatment with R-CHOP-like is associated with good survival in a significant proportion of patients. We have developed a simple prognostic model that may aid the selection patients who could benefit from a curative treatment, although it needs to be validated in larger series.


Assuntos
Linfoma Difuso de Grandes Células B/diagnóstico , Idoso de 80 Anos ou mais , Anticorpos Monoclonais Murinos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Ciclofosfamida/uso terapêutico , Gerenciamento Clínico , Doxorrubicina/uso terapêutico , Humanos , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/mortalidade , Prednisona/uso terapêutico , Prognóstico , Estudos Retrospectivos , Rituximab , Espanha , Análise de Sobrevida , Vincristina/uso terapêutico
19.
J Endod ; 44(1): 126-132, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29079052

RESUMO

INTRODUCTION: The aim of the present study was to evaluate the in vitro cytotoxicity of MTA Repair HP, NeoMTA Plus, and Biodentine, new bioactive materials used for dental pulp capping, on human dental pulp stem cells (hDPSCs). METHODS: Biological testing was carried out in vitro on hDPSCs. Cell viability and cell migration assays were performed using eluates of each capping material. To evaluate cell morphology and cell attachment to the different materials, hDPSCs were directly seeded onto the material surfaces and analyzed by scanning electron microscopy. The chemical composition of the pulp-capping materials was determined by energy-dispersive X-ray and eluates were analyzed by inductively coupled plasma-mass spectrometry. Statistical differences were assessed by analysis of variance and Tukey test (P < .05). RESULTS: Cell viability was moderate after 24 and 48 hours in the presence of MTA Repair HP and NeoMTA Plus, whereas at 48 and 72 hours, Biodentine showed higher rates of cell viability than MTA Repair HP and NeoMTA Plus (P < .001). A cell migration assay revealed adequate cell migration rates for MTA Repair HP and NeoMTA Plus, both similar to the control group rates, meanwhile the highest cell migration rate was observed in the presence of Biodentine (P < .001). Scanning electron microscope studies showed a high degree of cell proliferation and adhesion on Biodentine disks but moderate rates on MTA Repair HP and NeoMTA Plus disks. Energy-dispersive X-ray pointed to similar weight percentages of C, O, and Ca in all 3 materials, whereas other elements such as Al, Si, and S were also found. CONCLUSIONS: The new pulp-capping materials MTA Repair HP, NeoMTA Plus, and Biodentine showed a suitable degree of cytocompatibility with hDPSCs, and good cell migration rates, although Biodentine showed higher rates of proliferation time-dependent.


Assuntos
Materiais Biocompatíveis , Compostos de Cálcio , Polpa Dentária/citologia , Agentes de Capeamento da Polpa Dentária e Pulpectomia , Silicatos , Células-Tronco , Sobrevivência Celular , Humanos , Teste de Materiais
20.
J Tissue Eng Regen Med ; 12(3): 808-820, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-28621502

RESUMO

Keratinocyte migration is a mandatory aspect of wound healing. We have previously shown that amniotic membrane (AM) applied to chronic wounds assists healing through a process resulting in the overexpression of c-Jun at the wound's leading edge. We have also demonstrated that AM modifies the genetic programme induced by transforming growth factor-ß (TGF-ß) in chronic wounds. Here we used a scratch assay of mink lung epithelial cells (Mv1Lu) and a spontaneously immortalized human keratinocyte cell line (HaCaT) cells to examine the influence of AM application on the underlying signalling during scratch closure. AM application induced c-Jun phosphorylation at the leading edge of scratch wounds in a process dependent on MAPK and JNK signalling. Strikingly, when the TGF-ß-dependent Smad-activation inhibitor SB431542 was used together with AM, migration improvement was partially restrained, whereas the addition of TGF-ß had a synergistic effect on the AM-induced cell migration. Moreover, antagonizing TGF-ß with specific antibodies in both cell lines or knocking out TGF-ß receptors in Mv1Lu cells had similar effects on cell migration as using SB431542. Furthermore, we found that AM was able to attenuate TGF-ß-Smad signalling specifically at the migrating edge; AM treatment abated Smad2 and Smad3 nuclear localization in response to TGF-ß in a process dependent on mitogen-activated protein kinase kinase 1 (MEK1) activation but independent of EGF receptor or JNK activation. The involvement of Smad signalling on AM effects on HaCaT keratinocytes was further corroborated by overexpression of either Smad2 or Smad3 and the use of Smad phosphorylation-specific inhibitors, revealing a differential influence on AM-induced migration for each Smad. Thus, AM TGF-ß-Smad signalling abating is essential for optimal cell migration and wound closure.


Assuntos
Âmnio/metabolismo , Movimento Celular , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Humanos , Queratinócitos/citologia , Queratinócitos/metabolismo , Vison , Fosforilação , Proteínas Proto-Oncogênicas c-jun/metabolismo , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Cicatrização
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