By the end of 2003, cell therapy for T1DM started being performed in humans, and the world's first study was performed by the Divisions of Immunology and Endocrinology of the Hospital das Clínicas of the Faculty of Medicine of Ribeirão Preto - University of São Paulo - Brazil [12, 13]. The basic inclusion criteria are age between 12 and 35 years and a diagnosis of T1DM less than 6 weeks prior to inclusion confirmed by positive serum levels of anti-GAD antibodies.

In the first stage of the protocol, called mobilization, a small dose of cyclophosphamide is administered intravenously to mobilize hematopoietic stem cells from the bone marrow to the peripheral blood. Afterwards, we apply daily subcutaneous injections of the granulocyte colony stimulating factor to proliferate circulating stem cells; these cells are then collected and frozen.

Ten to fifteen days later, we start the second phase, called the conditioning regimen, in which we use high dose immunosuppression with intravenous cyclophosphamide 200 mg/kg plus intravenous rabbit anti-thymocyte globulin 4.5 mg/kg with aim of 'turning off' the immune system, mainly the peripheral T cells that retain immunologic memory. Later, we intravenously re-infuse the previously collected hematopoietic stem cells, which do not have immunologic memory, to regenerate a new immune system that will not attack pancreatic beta cells. This procedure is called 'immunologic resetting' and although no beta cells are regenerated, those not yet destroyed are preserved. It is important to emphasize that evidence in the medical literature indicates that hematopoietic stem cells are unable to be differentiated into beta cells. This is why we include only newly diagnosed patients - that is, patients who still have residual beta cell mass to be preserved.

Using this procedure, we may maintain or increase the secretion of endogenous insulin, improving the metabolic control and obviously the risk of chronic complications. We emphasize that if we obtain increased C-peptide levels for long periods and keep good glycemic control, we may offer patients a lower risk of chronic complications from diabetes, even without suspending insulin therapy.

Up to December 2008, we have performed this procedure in 23 patients: 17 men and 6 women aged 13 to 31 with an average body mass index of 19.7 kg/m2, an average glycemia level of 395.6 mg/dl and an average glycosylated hemoglobin level of 8.4% at the time of diagnosis, and undergoing anti-GAD treatment ranging from 1.1 to 102 U/ml and using an average dose of 0.4 UI/kg/day soon before starting the treatment.

All patients are recommended to be on a diet and to exercise regularly within a regimen of intensive insulin therapy and carbohydrate counting to try and achieve the following goals: pre-prandial glycemia <120 mg/dL, postprandial glycemia <140 mg/dL and glycosylated hemoglobin (HbA1c) <7%.

Patients receive psychological guidance to understand that they still have T1DM and that they must maintain a routine of automonitoring of capillary glycemia and carbohydrate counting to refrain from food abuse. Compliance with these guidelines is a challenge for the multidisciplinary team.

Of the 23 patients included, 20 remained free from insulin for some period. Of these, 12 have been continuously insulin-free since treatment, 8 became transiently insulin-free and 3 maintained daily insulin doses. In the group of continuously insulin-free patients, most stopped daily insulin injections soon after stem cell infusion and the mean period free from insulin is 31 months (ranging from 14 to 52 months). There was a significant reduction in HbA1c compared with pretreatment values, with all measures below 7% during follow-up. In parallel, this group of patients had important increases in mean peak-stimulated C-peptide levels (0.8 nmol/L pre-treatment versus 2.9 nmol/L after 3 years; P < 0.05). Of note, all patients reported important improvement in their quality of life; complete data will be published soon. Moreover, this long period without exogenous insulin use associated with progressive increases in C-peptide levels in multiple patients practically rules out the hypothesis of a prolonged honeymoon phase in this group.

Eight patients became transiently insulin-free for periods of 6 to 47 months (mean 17.7 months) However, they have excellent glucose control using small doses of insulin. Interestingly, in two patients who relapsed after being insulin free for 43 and 47 months, respectively, we opted to add sitagliptin (a dipeptidyl peptidase-4 inhibitor) 100 mg per day orally 4 and 2 months after resumption of insulin treatment. Both became insulin-free again 1 and 2 months later, and have remained free from exogenous insulin for more than 5 and 6 months, respectively. Peak-stimulated C-peptide levels declined in both patients before insulin treatment was resumed; however, C-peptide levels increased again after sitagliptin prescription, corresponding with the period of insulin-independence. Aside from this, we believe that suppression of glucagon levels can also explain the rapid insulin suspension after sitagliptin use. Additionally, immunomodulating effects of sitagliptin are speculated. So, this is the first report of complete insulin suspension in individuals with T1DM using sitagliptin [14].

In the whole group of eight patients who were transiently insulin-free, there was a significant increase in C-peptide levels from 0.6 nmol/L pre-treatment to 1.7 nmol/L 4 years after treatment (P < 0.05). To the best of our knowledge, this is the most prolonged period of C-peptide increase (up to 4 years) in interventional trials aiming at beta cell preservation.

Only three patients did not experience any period free from insulin. One presented with diabetic ketoacidosis at diagnosis and received glucocorticoids to prevent rabbit anti-thymocyte globulin reactions, one developed diabetic ketoacidosis before enrollment, and one had inadvertently received steroids (300 mg hydrocortisone) along with stem cell infusion. None of them achieved HbA1c levels less than 7% in spite of progressive increase in daily insulin doses (>0.8 IU/kg/day).

The majority of the adverse effects related to treatment were mild, including nausea, vomiting, fever, hyporexia and alopecia. With regard to severe adverse effects, two patients presented bilateral nosocomial pneumonia that completely responded to intravenous broad spectrum antibiotics; there was no mortality. During long-term follow-up, there was one case each of Graves' disease, transient hypergonadotropic hypogonadism, and autoimmune hypothyroidism. Nine patients had post-transplant hypospermia. Two patients fathered children 2 years after transplant. There was no mortality.