Abstract: Type 1 diabetes (T1D) is an autoimmune disease in which the patient's immune system recognizes their pancreatic islet insulin-producing cells and destroys them. To cure T1D in a comprehensive manner, not only must the islet cells be replaced, the patient's immune system must also be properly regulated mostly in the form of suppression. Blood-derived new stem cells have shown promise in both aspects of this treatment.

Millions of type 1 diabetes (T1D) patients worldwide must have daily insulin injections to survive. However, insulin injection is not a cure; it does not halt the persistent autoimmune responses. This compelling need brings a sense of urgency to find a cure for T1D that can not only overcome the shortage of insulin-producing beta-cells, but also halt the progression of autoimmunity. Recently, in vitro co-culture of lymphocytes with stem cells, including human cord blood stem cells (CB-SC) (Zhao et al., 2009; Zhao et al., 2007) or mesenchymal stem cells (Uccelli et al., 2008), has been shown to modulate lymphocytes. Application of these stem cells holds promise for the treatment of T1D. This review will focus on the recent progress of cord blood stem cells.

Functional Defects of Regulatory T cells in T1D

T1D is a complex, chronic autoimmune disease with a serious clinical outcome. Despite the possible involvement of viral infection, environmental factors, and genetic predisposition that can increase the risk of T1D, the specific factor or factors that trigger autoimmunity remain elusive. Due to this complexity and the uncertain mechanism of disease, it may be difficult to treat or prevent the disease by blocking only one cytokine or signaling pathway (von Herrath and Nepom, 2009; Faustman, 2008; Fife and Bluestone, 2008; You et al., 2008). Regulatory T cells (Tregs) play a crucial role in maintaining homeostasis and self-tolerance through their inhibitory impacts on autoreactive effector T cells, such as releasing immunosuppressive cytokines interleukin-10 (IL-10) and/or transforming growth factor-β 1 (TGF-β1). Compelling evidence demonstrates that abnormalities of Tregs, either in cell number or in function, are associated with the initiation and progression of T1D, both in diabetic patients and animal models (Bayry et al., 2008; Brusko et al., 2005; Lindley et al., 2005; Pop et al., 2005; You et al., 2005).

Currently, administration of molecularly engineered CD3 (a pan-T cell biomarker) antibody against T cells within a few weeks of the initial onset of glycosuria has led to marked preservation of pancreatic islet cells’ ability to produce C-peptide (an insulin by-product) and a decrease in the insulin dose requirement compared to controls who did not receive this monoclonal antibody (Keymeulen et al., 2005; McDevitt and Unanue, 2008). Insulin is still needed after CD3 antibody therapy in new-onset T1D (Keymeulen et al., 2005). However, the safety and adverse effects in many patients are a concern in CD3 antibody therapy (Keymeulen et al., 2005; McDevitt and Unanue, 2008; Herold et al. 2005). With another immunotherapy using the recombinant human glutamic acid decarboxylase (GAD 65) as a therapeutic vaccine, patients who participated in the initial study and showed preservation of C-peptide levels and a decrease in insulin dose requirement were the patients with the shortest time lapse between onset of glycosuria and administration of GAD 65. However, patients with recent-onset diabetes that occurred more than 6 months prior to the treatment with GAD 65 showed no therapeutic effect (Uibo and Lernmark, 2008; Ludvigsson et al., 2008). Importantly, studies on biological mechanisms in both clinical trials revealed that induction of Tregs is the common turning point attributable to the observed therapeutic effects (Uibo and Lernmark, 2008; Belghith et al., 2003; You et al., 2007).

Cord Blood Stem Cells Correct the Functional Defects of Regulatory T cells (Tregs) in T1D

T1D is a T cell-mediated autoimmune disease that leads to a major loss of insulin-producing beta cells. Manipulation of Tregs has become an attractive approach for the prevention and treatment of T1D. Nevertheless, only a limited number of studies have focused on restoration of impaired Treg function to confer protection against autoimmune diabetes. We identified a novel type of stem cells from human umbilical cord blood, designated cord blood stem cells (CB-SC) (Zhao et al., 2006). For the first time, we used CB-SC to correct functional defects of CD4⁺CD62L⁺ Tregs through modulating global gene expression profiles, leading to reversal of overt diabetes in an autoimmune diabetic NOD mouse model (Zhao et al., 2009; Zhao et al., 2007). Notably, treatment with CB-SC-modulated CD4⁺CD62L⁺ Tregs (mCD4⁺CD62L⁺ Tregs) in diabetic NOD mice can simultaneously overcome the autoimmunity via systemic and local immune modulations and the shortage of insulin-producing cells via stimulating the β-cell regeneration (Zhao et al., 2009). However, the freshly isolated CD4⁺CD62L⁺ Tregs failed to show therapeutic potential on T1D (Zhao et al., 2009). Thus, results indicate that CB-SC can correct the functional defects of Tregs. Current mechanistic studies in healthy human donors have revealed that CB-SC can modulate lymphocyte proliferation and related phenotypes via surface molecular programmed death-ligand 1 and soluble factor nitric oxide (Zhao et al., 2007). To this end, it will achieve more favorable results by administering autologous Tregs after being educated by CB-SC via in vitro co-culture for the treatment of T1D.

Induction of Immune Tolerance Via “TGF-β1 Ring” in Pancreatic Islets

IL-10 and TGF-β1 are representative cytokines that contribute to the induction of immune tolerance. Treatment with mCD4⁺CD62L⁺ Tregs in diabetic NOD mice markedly increased plasma levels of IL-10 and TGF-β1. These suppressor cytokines can help create a tolerogenic environment following treatment with mCD4⁺CD62L⁺ Tregs. Specifically, TGF-β1 represents one of the best characterized cytokines contributing to the induction of immune suppression and maintaining self-tolerance (Li and Flavell, 2008). A number of studies suggest that paracrine and/or autocrine sources of TGF-β1 are essential for the regulation of T cell tolerance (Longenecker et al., 2002; Du et al., 2006; Kaplan et al., 2007). To elucidate the de novo molecular mechanism underlying the protection of newly generated islet β cells following treatment with mCD4⁺CD62L⁺ Tregs, we have found that TGF-β1-positive cells, along with their released TGF-β1 in the matrix, formed a “TGF-β1 ring” surrounding pancreatic islets. This ring can protect newly generated islets against the autoimmune re-attack by inducing apoptosis of auto-aggressive effector lymphocytes (Zhao et al., 2009).

Shortage of insulin-producing cells is another key issue for T1D. Abrogation of autoimmunity without an adequate residual beta-cell mass will not restore euglycemia. Thus, approaches of reconstituting normal, physiological insulin production must be part of any therapy aimed at treating T1D (von Herrath and Nepom, 2009; Pasquali et al., 2008). Previous studies have demonstrated that this new type of blood stem cell can turn into insulin-producing cells (Zhao et al., 2006; Zhao et al., 2007). Notably, treatment with CB-SC-modulated Tregs not only diminished the autoimmunity, but also eliminated hyperglycemia (Zhao et al., 2009). Mechanistic studies on glycemic control have revealed that increase in total beta-cell mass and reconstitution of islet architecture via a marked increase in residual beta-cell proliferation play a key role in the restoration of euglycemia after treatment with CB-SC-modulated Tregs (Zhao et al., 2009). Treatment with CB-SC-modulated Tregs enhances expression of TGF-β1 and formation of the TGF-β1 ring in pancreatic islets that can contribute to the local protection of newly generated pancreatic islets from the re-destruction by autoreactive immune cells.

These new blood stem cells naturally exist in human blood circulation including cord blood and adult peripheral blood. They can be isolated using relatively straightforward methodologies and can be cultured and expanded in vitro (Zhao et al., 2006). They show promise in managing the many facets of T1D treatment. Therapeutic applications of these stem cells have the potential to exert enormous clinical impact on T1D and towards the development of novel stem cell-modulated Tregs to treat T1D.

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