An ongoing debate surrounds the existence of stem cells in the adult endowed with capacity to differentiate into multiple lineages. We examined the possibility that adult bone marrow cells participate in recovery from chemical diabetes through neogenesis of insulin-producing cells. Small-sized cells negative for lineage markers derived by counterflow centrifugal elutriation from the bone marrow were transplanted into mice made diabetic with streptozotocin and sublethal irradiation. These cells homed efficiently to the injured islets and contributed to tissue revascularization. Islet-homed CD45-negative donor cells identified by sex chromosomes downregulated GFP, expressed PDX-1 and proinsulin, and converted the hormone precursor to insulin. An estimated 7.6% contribution of newly formed insulin-producing cells to islet cellularity increased serum insulin and stabilized glycemic control starting at 5 weeks post-transplant and persisting for 20 weeks. Newly differentiated cells displayed normal diploid genotype and there was no evidence of fusion between the grafted stem cells or their myeloid progeny and injured β-cells. Considering the extensive functional incorporation of insulin-producing donor cells in the injured islets, we conclude that the adult bone marrow contains a subset of small cells endowed with plastic developmental capacity.

The contribution of stem cells derived from adult tissues to the recovery of pancreatic islets from chemical injury is controversial. Analysis of nonhematopoietic differentiation of bone marrow-derived cells has yielded positive and negative results under different experimental conditions. Using the smallest subset of bone marrow cells lacking immuno-hematopoietic lineage markers, we have detected incorporation and conversion into insulin-producing cells. Donor cells identified by genomic markers silence green fluorescent protein (GFP) expression as a feature of differentiation, in parallel to expressing PDX-1 and proinsulin. Here we elaborate potential experimental difficulties that might result in false-negative results. The use of GFP as a reporter protein is suboptimal for differentiation experiments: (a) the bone marrow of GFP donors partially expresses the reporter protein, (b) differentiating bone marrow cells silence GFP expression, and (c) the endocrine pancreas is constitutively negative for GFP. In addition, design of the experiments, data analysis, and interpretation encounter numerous objective and subjective difficulties. Rigorous evaluation under optimized experimental conditions confirms the capacity of adult bone marrow-derived stem cells to adopt endocrine developmental traits, and demonstrates that GFP downregulation and silencing is a feature of differentiation.

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Regenerative medicine opens new avenues and promises towards more effective therapies for autoimmune disorders. Current therapeutic strategies for type I diabetes focus on three major directions, with distinct advantages and disadvantages: arrest of autoimmunity, islet transplantation and generation of neoislets. There is mounting evidence that candidate stem cells residing in the hematopoietic compartments participate in regeneration of pancreatic islets following chemical and autoimmune injury in vivo. The apparent major mechanisms include immunomodulation, revascularization, support of endogenous beta-cell regeneration and differentiation into insulin-producing cells. Review of the current evidence suggests that some divergent observations depend primarily on the experimental design, which both limits and accentuates developmental events. The flood of publications reporting negative results appears to reflect primarily suboptimal experimental conditions for differentiation of putative stem cells, rather than limited developmental plasticity. Stem cells modulate the course of autoimmune diabetes through multiple mechanisms, including de novo generation of units capable to sense, produce and secrete insulin. Therefore, the charged debate over controversies surrounding developmental plasticity should not impede attempts to design curative therapies for this disease.

We have recently reported that small-sized bone marrow cells (BMCs) isolated by counterflow centrifugal elutriation and depleted of lineage markers (Fr25lin − ) have the capacity to differentiate and contribute to regeneration of injured islets. In this study, we assess some of the characteristics of these cells compared to elutriated hematopoietic progenitors (R/O) and whole BMCs in a murine model of streptozotocin-induced chemical diabetes. The GFP bright CD45+ progeny of whole BMCs and R/O progenitors progressively infiltrate the pancreas with evolution of donor chimerism; are found at islet perimeter, vascular, and ductal walls; and have a modest impact on islet recovery from injury. In contrast, Fr25lin− cells incorporate in the islets, convert to GFP dim CD45 − PDX-1 + phenotypes, produce proinsulin, and secrete insulin with significant contribution to stabilization of glucose homeostasis. The elutriated Fr25lin − cells express low levels of CD45 and are negative for SCA-1 and c-kit, as removal of cells expressing these markers did not impair conversion to produce insulin. BMCs mediate two synergistic mechanisms that contribute to islet recovery from injury: support of islet remodeling by hematopoietic cells and neogenesis of insulin-producing cells from stem cells.