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High dose immunosuppression followed by autologous hematopoietic stem cell transplantation (AHSCT) induces prolonged clinical remission in multiple sclerosis (MS) patients. However, how patient immune profiles are associated with clinical outcomes has not yet been completely elucidated. In this study, 37 MS patients were assessed for neurological outcomes, thymic function and long-term immune reconstitution after AHSCT. Patients were followed for a mean (SD) of 68.5 (13.9) months post-transplantation and were retrospectively clustered into progression- and non-progression groups, based on Expanded Disease Status Scale (EDSS) outcomes at last visit. After AHSCT, both patient groups presented increased regulatory T-cell subset counts, early expansion of central- and effector-memory CD8(+)T-cells and late thymic reactivation. However, the non-progression group presented early expansion of PD-1(+)CD8(+)T-cells and of PD-1-expressing CD19(+) B-cells. Here, we suggest that along with increased numbers of regulatory T-cell subsets, PD-1 inhibitory signaling is one possible immunoregulatory mechanism by which AHSCT restores immune tolerance in MS patients., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Autologous haematopoietic stem-cell transplantation (AHSCT) has been experimented as a treatment in patients affected by severe forms of multiple sclerosis (MS) who failed to respond to standard immunotherapy. The rationale of AHSCT is to 'reboot' the immune system and reconstitute a new adaptive immunity. The aim of our study was to identify, through a robust and unbiased transcriptomic analysis, any changes of gene expression in T-cells potentially underlying the treatment effect in patients who underwent non-myeloablative AHSCT for treatment of MS. We evaluated by microarray DNA-chip technology the gene expression of peripheral CD4+ and CD8+ T-cell subsets sorted from patients with MS patients before AHSCT, at 6 months, 1 year and 2 years after AHSCT and from healthy control subjects. Hierarchical clustering analysis revealed that reconstituted CD8+ T-cells of MS patients at 2 years post-transplantation, aggregated together with healthy controls, suggesting a normalization of gene expression in CD8+ cells post-therapy. When we compared the gene expression in MS patients before and after therapy, we detected a large number of differentially expressed genes (DEG) in both CD8+ and CD4+ T-cell subsets at all time points after transplantation. We catalogued the biological function of DEG and we selected 27 genes known to be involved in immune function for accurate quantification of gene expression by real-time PCR. The analysis confirmed and extended with quantitative data, a number of significant changes in both the CD4+ and CD8+ T-cells subsets from MS post-transplant. Notably, CD8+ T-cells revealed more extensive changes in the expression of genes involved in effector immune responses.

Context: In 2007, the effects of the autologous nonmyeloablative hematopoietic stem cell transplantation (HSCT) in 15 patients with type 1 diabetes mellitus (DM) were reported. Most patients became insulin free with normal levels of glycated hemoglobin A(1c) (HbA(1c)) during a mean 18.8-month follow-up. To investigate if this effect was due to preservation of beta-cell mass, continued monitoring was performed of C-peptide levels after stem cell transplantation in the 15 original and 8 additional patients., Objective: To determine C-peptide levels after autologous nonmyeloablative HSCT in patients with newly diagnosed type 1 DM during a longer follow-up., Design, Setting, and Participants: A prospective phase 1/2 study of 23 patients with type 1 DM (aged 13-31 years) diagnosed in the previous 6 weeks by clinical findings with hyperglycemia and confirmed by measurement of serum levels of anti-glutamic acid decarboxylase antibodies. Enrollment was November 2003-April 2008, with follow-up until December 2008 at the Bone Marrow Transplantation Unit of the School of Medicine of Ribeirão Preto, Ribeirão Preto, Brazil. Hematopoietic stem cells were mobilized via the 2007 protocol., Main Outcome Measures: C-peptide levels measured during the mixed-meal tolerance test, before, and at different times following HSCT. Secondary end points included morbidity and mortality from transplantation, temporal changes in exogenous insulin requirements, and serum levels of HbA(1c)., Results: During a 7- to 58-month follow-up (mean, 29.8 months; median, 30 months), 20 patients without previous ketoacidosis and not receiving corticosteroids during the preparative regimen became insulin free. Twelve patients maintained this status for a mean 31 months (range, 14-52 months) and 8 patients relapsed and resumed insulin use at low dose (0.1-0.3 IU/kg). In the continuous insulin-independent group, HbA(1c) levels were less than 7.0% and mean (SE) area under the curve (AUC) of C-peptide levels increased significantly from 225.0 (75.2) ng/mL per 2 hours pretransplantation to 785.4 (90.3) ng/mL per 2 hours at 24 months posttransplantation (P < .001) and to 728.1 (144.4) ng/mL per 2 hours at 36 months (P = .001). In the transient insulin-independent group, mean (SE) AUC of C-peptide levels also increased from 148.9 (75.2) ng/mL per 2 hours pretransplantation to 546.8 (96.9) ng/mL per 2 hours at 36 months (P = .001), which was sustained at 48 months. In this group, 2 patients regained insulin independence after treatment with sitagliptin, which was associated with increase in C-peptide levels. Two patients developed bilateral nosocomial pneumonia, 3 patients developed late endocrine dysfunction, and 9 patients developed oligospermia. There was no mortality., Conclusion: After a mean follow-up of 29.8 months following autologous nonmyeloablative HSCT in patients with newly diagnosed type 1 DM, C-peptide levels increased significantly and the majority of patients achieved insulin independence with good glycemic control., Trial Registration: clinicaltrials.gov Identifier: NCT00315133.

In this review, we present (1) the scientific basis for the use of high-dose immunosuppression followed by autologous peripheral blood hematopoietic stem cell transplantation for newly diagnosed type 1 diabetes (T1D); (2) an update of the clinical and laboratory outcome of 20 patients transplanted at the University Hospital of the Ribeirão Preto Medical School, University of São Paulo, Brazil, and followed up to January/2008, including 4 relapses among 19 patients without previous ketoacidosis; (3) a commentary on criticisms to our article that appeared in four articles from the scientific literature; and (4) a discussion of the prospectives for cellular therapy for T1D.

Context: Type 1 diabetes mellitus (DM) results from a cell-mediated autoimmune attack against pancreatic beta cells. Previous animal and clinical studies suggest that moderate immunosuppression in newly diagnosed type 1 DM can prevent further loss of insulin production and can reduce insulin needs., Objective: To determine the safety and metabolic effects of high-dose immunosuppression followed by autologous nonmyeloablative hematopoietic stem cell transplantation (AHST) in newly diagnosed type 1 DM., Design, Setting, and Participants: A prospective phase 1/2 study of 15 patients with type 1 DM (aged 14-31 years) diagnosed within the previous 6 weeks by clinical findings and hyperglycemia and confirmed with positive antibodies against glutamic acid decarboxylase. Enrollment was November 2003-July 2006 with observation until February 2007 at the Bone Marrow Transplantation Unit of the School of Medicine of Ribeirão Preto, Ribeirão Preto, Brazil. Patients with previous diabetic ketoacidosis were excluded after the first patient with diabetic ketoacidosis failed to benefit from AHST. Hematopoietic stem cells were mobilized with cyclophosphamide (2.0 g/m2) and granulocyte colony-stimulating factor (10 microg/kg per day) and then collected from peripheral blood by leukapheresis and cryopreserved. The cells were injected intravenously after conditioning with cyclophosphamide (200 mg/kg) and rabbit antithymocyte globulin (4.5 mg/kg)., Main Outcome Measures: Morbidity and mortality from transplantation and temporal changes in exogenous insulin requirements (daily dose and duration of usage). Secondary end points: serum levels of hemoglobin A1c, C-peptide levels during the mixed-meal tolerance test, and anti-glutamic acid decarboxylase antibody titers measured before and at different times following AHST., Results: During a 7- to 36-month follow-up (mean 18.8), 14 patients became insulin-free (1 for 35 months, 4 for at least 21 months, 7 for at least 6 months; and 2 with late response were insulin-free for 1 and 5 months, respectively). Among those, 1 patient resumed insulin use 1 year after AHST. At 6 months after AHST, mean total area under the C-peptide response curve was significantly greater than the pretreatment values, and at 12 and 24 months it did not change. Anti-glutamic acid decarboxylase antibody levels decreased after 6 months and stabilized at 12 and 24 months. Serum levels of hemoglobin A(1c) were maintained at less than 7% in 13 of 14 patients. The only acute severe adverse effect was culture-negative bilateral pneumonia in 1 patient and late endocrine dysfunction (hypothyroidism or hypogonadism) in 2 others. There was no mortality., Conclusions: High-dose immunosuppression and AHST were performed with acceptable toxicity in a small number of patients with newly diagnosed type 1 DM. With AHST, beta cell function was increased in all but 1 patient and induced prolonged insulin independence in the majority of the patients.

Autologous hematopoietic stem cell transplantation (AHSCT) increases C-peptide levels and induces insulin independence in patients with type 1 diabetes. This study aimed to investigate how clinical outcomes may associate with the immunological status, especially concerning the balance between immunoregulation and autoreactivity. Twenty-one type 1 diabetes patients were monitored after AHSCT and assessed every 6 months for duration of insulin independence, C-peptide levels, frequencies of islet-specifc autore-active CD8+ T cells (CTL), regulatory lymphocyte subsets, thymic function, and T-cell repertoire diversity. In median follow-up of 78 (range 15-106) months, all patients became insulin-independent, resuming insulin after median of 43 (range 6-100) months. Patients were retrospectively divided into short- or prolonged-remission groups, according to duration of insulin independence. For the entire follow-up, CD3+CD4+ T-cell numbers remained lower than baseline in both groups, whereas CD3+CD8+ T-cell levels did not change, resulting in a CD4/CD8 ratio inversion. Memory CTL comprehended most of T cells detected on long-term follow-up of patients after AHSCT. B cells reconstituted to baseline levels at 2-3 months post-AHSCT in both patient groups. In the prolonged-remission-group, baseline islet-specifc T-cell autoreactivity persisted after transplantation, but regulatory T cell counts increased. Patients with lower frequencies of autoreactive islet-specifc T cells remained insulin-free longer and presented greater C-peptide levels than those with lower frequencies of these cells. Therefore, immune monitoring identifed a subgroup of patients with superior clinical outcome of AHSCT. Our study shows that improved immunoregulation may balance autoreactivity endorsing better metabolic outcomes in patients with lower frequencies of islet-specific T cells. Development of new strategies of AHSCT is necessary to increase frequency and function of T and B regulatory cells and decrease efficiently autoreactive islet-specific T and B memory cells in type 1 diabetes patients undergoing transplantation. [ABSTRACT FROM AUTHOR]

Aims and objectives. To identify differences in cyclosporine levels between blood samples collected from a peripheral venous access, catheter line used for drug infusion and catheter line not used for drug infusion in adult patients receiving allogeneic haematopoietic stem cell transplantation. Background. Cyclosporine is an immunosuppressant that prevents graft-versus-host disease, has a narrow therapeutic window and causes nephrotoxicity. For cyclosporine infusion, a tunnelled central venous access device is used; however, because of the lipophilic properties of the drug, it can adsorb to the catheter surface and falsely raise cyclosporine concentrations in blood specimens. Design. Prospective observational study. Methods. The study collected 135 blood samples from 16 patients. In 13 subjects, samples were obtained from the three lines at three time points (1, 7 and 14 days after the start of cyclosporine infusion), and for three subjects, samples were only obtained at 1 and 7 days after the start of infusion. The 5-ml blood discard method was used for samples collected from the catheter. Using this procedure, the catheter line was washed with saline solution, 5 ml of blood and saline solution were aspirated from the catheter line and discarded, and then sample blood used for the test was collected. The paired t-test with the Bonferroni correction was used to analyse the differences in cyclosporine serum levels. Results. Significant differences were observed when the drug serum levels obtained in the line used for drug infusion were compared with the levels obtained in the line not used for infusion or the peripheral venous line. No differences in drug levels were identified in blood collected from the peripheral venous line and the line not used for drug infusion. Conclusion. Drug adsorption occurs in the line used for infusion. Therefore, the blood sample collected from the line not used for cyclosporine infusion can be considered reliable for drug concentration determination. Relevance to clinical practice. Nurses should standardise one line of the tunnelled central venous access device for cyclosporine infusion, which avoids the need for evasive procedures and provides patients with more comfort. [ABSTRACT FROM AUTHOR]

In this review, we present (1) a brief discussion of hematopoietic stem cell transplantation (HSCT) for severe and refractory autoimmune diseases (AIDs) from its beginning in 1996 through recently initiated prospective randomized clinical trials; (2) an update (up to July 2009) of clinical and laboratory outcomes of 23 patients with newly diagnosed type 1 diabetes mellitus (T1DM), who underwent autologous HSCT at the Bone Marrow Transplantation Unit of the Ribeirão Preto Medical School, University of São Paulo, Brazil; (3) a discussion of possible mechanisms of action of HSCT in AIDs, including preliminary laboratory data obtained from our patients; and (4) a discussion of future perspectives of stem cell therapy for T1DM and type 2 DM, including the use of stem cell sources other than adult bone marrow and the combination of cell therapy with regenerative compounds. [ABSTRACT FROM AUTHOR]