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We sought to assess the potential of human cord blood-derived mesenchymal stem cells (CB-MSCs) to derive insulin-producing, glucose-responsive cells. We show here that differentiation protocols based on stepwise culture conditions initially described for human embryonic stem cells (hESCs) lead to differentiation of cord blood-derived precursors towards a pancreatic endocrine phenotype, as assessed by marker expression and in vitro glucose-regulated insulin secretion. Transplantation of these cells in immune-deficient animals shows human C-peptide production in response to a glucose challenge. These data suggest that human cord blood may be a promising source for regenerative medicine approaches for the treatment of diabetes mellitus.

The objective of this study was to determine whether the combination therapy of intrapancreatic autologous stem cell infusion (ASC) and hyperbaric oxygen treatment (HBO) before and after ASC can improve islet function and metabolic control in patients with type 2 diabetes mellitus (T2DM). This prospective phase 1 study enrolled 25 patients with T2DM who received a combination therapy of intrapancreatic ASC and periinfusion HBO between March 2004 and October 2006 at Stem Cells Argentina Medical Center Buenos Aires, Argentina. Clinical variables (body mass index, oral hypoglycemic drugs, insulin requirement) and metabolic variables (fasting plasma glucose, C-peptide, HbA1c, and calculation of C-peptide/glucose ratio) were assessed over quartile periods starting at baseline and up to 1 year follow-up after intervention. Means were calculated in each quartile period and compared to baseline. Seventeen male and eight female patients were enrolled. Baseline variables expressed as means ± SEs were: age 55 ± 2.14 years, diabetes duration 13.2 ± 1.62 years, insulin dose 34.8 ± 2.96 U/day, and BMI 27.11 ± 0.51. All metabolic variables showed significant improvement when comparing baseline to 12 months follow-up, respectively: fasting glucose 205.6 ± 5.9 versus 105.2 ± 14.2 mg/dl, HbA1c 8.8 ± 0.2 versus 6.0 ± 0.4%, fasting C-peptide 1.5 ± 0.2 versus 3.3 ± 0.3 ng/ml, C-peptide/glucose ratio 0.7 ± 0.2 versus 3.5 ± 0.3, and insulin requirements 34.8 ± 2.9 versus 2.5 ± 6.7 U/day. BMI remained constant over the 1-year follow-up. Combined therapy of intrapancreatic ASC infusion and HBO can improve metabolic control and reduce insulin requirements in patients with T2DM. Further randomized controlled clinical trials will be required to confirm these findings.

The measurement of cellular oxygen consumption rate (OCR) is a potential tool for the assessment of metabolic potency of isolated islets of Langerhans prior to clinical transplantation. We used a commercially available 96-well plate fluoroprobe, the BD Oxygen Biosensor System (OBS), to estimate OCR in 27 human islet preparations, and compared these results to those of concurrent mouse transplantations. OCR was estimated both from the dO 2 at steady state and from the transient rate of change of dO 2 during the initial culture period immediately after seeding (“dO 2 slope”). To demonstrate the validity of the OBS-derived values, it was shown that they scaled linearly with islet equivalent number/DNA concentration and with each other. These measurements were obtained for each preparation of islets incubated in media supplemented with either low (2.2 mM) or high (22 mM) glucose. Concurrently, one to three athymic nude mice were transplanted with 2,000 IEQs under the kidney capsule. The OCR Index, defined as the ratio of the DNA-normalized “dO 2 slope” in high glucose to that in low glucose, proved highly predictive of mouse transplant results. Of the 69 mice transplanted, those receiving islets where the OCR Index exceeded 1.27 were 90% likely to reverse within 3 days, whereas those receiving islets with an OCR Index below 1.27 took significantly longer, often failing to reverse at all over a 35-day time period. These results suggest that the OBS could be a useful tool for the pretransplant assessment of islet cell potency.

Variables such as pH, pCO 2 , and pO 2 have been established in the literature as critical factors that could affect the outcome of the islet cell processing and, therefore, the quality of the cells that could be transplanted. This report describes a highly accurate continuous multiparametric monitoring system and its evaluation for continuous monitoring of physiological variables during critical steps of the islet isolation procedure as well as during in vitro culture of the insulin-producing cells. Close monitoring of these variables could be of assistance to improve the outcome of islet cell processing, allowing to identify as soon as possible problems that could be corrected during the procedure, as well as during in vitro preservation, or shipment to remote sites.

As islet cell transplantation gains increasing interest following results published by the Edmonton group, results that have been successfully reproduced by several centers nationwide and abroad, the need of guidelines to standardize the procedure becomes highly important. We detail the key steps of the infusion procedure utilizing a closed gravity fed bag system utilized at our institution since 1990, which consists of a 600-ml transfer bag and a 150-ml rinse bag connected via sterile tubing. The use of gravity allows for a control rate of infusion as well as providing a safety mechanism through natural reduction of flow that parallels any increase in portal pressure, therefore allowing the operator to prevent precipitous pressure rises. Reports on significant rise in portal pressures during islet cell infusion as well as portal vein thrombosis have been published. Infusion at these centers was carried out using a syringe method. Using our technique, portal vein thrombosis (partial or complete) was not detected in any of the infusions performed at our institution. This method may be of assistance to minimize some of the observed complications associated with islet transplant procedures and has now been adapted by most centers performing clinical islet transplantation.

We have tested the effect of donor bone marrow cell (DBMC) infusion on the survival of pancreatic islet allografts in the rat, without the use of cytoablative recipient conditioning. Lewis and diabetic Brown Norway rats were used as donors and recipients, respectively. Donor islets were placed beneath the left renal capsule. Infusion of DBMC and temporary immunosuppression followed by delayed islet transplantation resulted in indefinite survival of all islet grafts (MST >180 days). Control animals demonstrated recurrent hyperglycemia (islet allografts rejection). Donor bone marrow derived cells were detected in the spleen and cervical lymph nodes of BN recipients of LEW bone marrow but not in the recipients of islet transplants alone. Second set full thickness skin grafts were performed in normal BN and in recipients of a previously successful ITX. Donor specific skin grafts were accepted in the animals that had received DBMC 40 days before the islet allograft, while animals receiving DBMC at the time of the islet allograft rejected the donor specific skin graft similarly to the controls. However, these animals did not reject a second set donor-specific islet transplant. The results indicate that radiation conditioning of the recipients was not necessary to induce microchimerism and graft acceptance in this rodent model of islet allotransplantation.

Graft-versus-host disease (GVHD) remains a major complication of bone marrow transplantation. This report describes reversal of GVHD by infusion of stored recipient bone marrow following combined liver-bone marrow allotransplantation. Graft-versus-host disease developed at the end of the first postoperative week. The skin involvement progressively spread to approximately 80% of the body surface and was not affected by modification of the immunosuppressive treatment. On the 42nd and 43rd postoperative day 1.23 × 108 and 1.6 × 108 autologous bone marrow cells per kg of recipient body weight were infused. The skin rush began to dramatically improve and resolved within 2 wk from the autologous marrow infusion. Autologous bone marrow storage previous to allogeneic bone marrow transplantation for tolerance induction could constitute a safety net in case of occurrence of GVHD.

The future implementation of stem cell therapies to treat conditions thus far considered incurable has been envisioned as logical consequence of the fast-paced progress in stem cell research over the last few years. Still, many practical obstacles stand in the way to the routine application of these novel technologies in medicine. The conference “Stem Cell Therapies in Reparative Medicine,” held aboard the cruise vessel Majesty of the Seas (Miami, USA- Nassau, Bahamas, April 19–22, 2002), focused on the analysis of these problems from different perspectives, including developmental biology (cell proliferation, fate determination, and enrichment), immunology (allorejection and prevention of autoimmunity recurrence), and clinical therapy, emphasizing the impact of stem cell technologies on the emerging field of tissue engineering and the treatment of alpha-1 antitrypsin deficiency.