Danling Gu

Pancreatic Expression of Keratinocyte Growth Factor Leads to Differentiation of Islet Hepatocytes and Proliferation of Duct Cells

Keratinocyte growth factor, (KGF), a member of the fibroblast growth factor (FGF) family, is involved in wound healing. It also promotes the differentiation of many epithelial tissues and proliferation of epithelial cells as well as pancreatic duct cells. Additionally, many members of the highly homologous FGF family (including KGF), influence both growth and cellular morphology in the developing embryo. We have previously observed elevated levels of KGF in our interferon-gamma transgenic mouse model of pancreatic regeneration. To understand the role of KGF in pancreatic differentiation, we generated insulin promoter-regulated KGF transgenic mice. Remarkably, we have found that ectopic KGF expression resulted in the emergence of hepatocytes within the islets of Langerhans in the pancreas. Additionally, significant intra-islet duct cell proliferation in the pancreata of transgenic KGF mice was observed. The unexpected appearance of hepatocytes and proliferation of intra-islet duct cells in the pancreata of these mice evidently stemmed directly from local exposure to KGF.

α-Cell Neogenesis in an Animal Model of IDDM

Currently there is debate regarding the capacity of pancreatic islets to regenerate in adult animals. Because pancreatic endocrine cells are thought to arise from duct cells, we examined the pancreatic ductal epithelium of the diabetic NOD mouse for evidence of islet neogenesis. We have evidence of duct proliferation as well as ductal cell differentiation, as suggested by bromodeoxyuridine-labeling and the presence of glucagon-containing cells within these ducts. In addition, the ductal epithelia in diabetic NOD mice expressed the neuroendocrine markers neuropeptide Y and tyrosine hydroxylase. These ducts also expressed the homeobox gene product, insulin promoter factor 1. Ductal cell proliferation and expression of these markers was not observed in transgenic NOD mice (NOD-E), which do not develop clinical or histopathological symptoms of IDDM. This suggests that the observed ductal cell proliferation and differentiation was a direct result of β-cell destruction and insulin insufficiency in these adult diabetic mice, which further suggests that these events are recapitulating islet ontogeny observed during embryogenesis. It is possible that comparable processes occur in the human diabetic pancreas.

Endocrine/Exocrine Intermediate Cells in Streptozotocin-Treated Ins-IFN-γ Transgenic Mice

To trace the ontogeny of β cell regrowth in adult transgenic mice that produce interferon-γ in the islets (ins-IFN-γ), their existing β cells were depleted by treatment with high doses of streptozotocin (STZ). Initially, β cell necrosis and degranulation were apparent in STZ-treated mice of both the BALB/c and the ins-IFN-γ transgenic strains. The newly emerging transitional cells were then characterized by ultrastructural analysis. Interestingly, transitional cells harboring both exocrine and endocrine granules appeared frequently in ins-IFN-γ transgenics after high-dose STZ treatment. New β cells were produced primarily by the formation of new islets from the small pancreatic ducts. β cell regeneration in the ins-IFN-γ transgenic mouse model is thus explained primarily by the budding of new islets from the ducts with acinar cells as possible precursors of islet cells.

alpha-Cell neogenesis in an animal model of IDDM.

Currently there is debate regarding the capacity of pancreatic islets to regenerate in adult animals. Because pancreatic endocrine cells are thought to arise from duct cells, we examined the pancreatic ductal epithelium of the diabetic NOD mouse for evidence of islet neogenesis. We have evidence of duct proliferation as well as ductal cell differentiation, as suggested by bromodeoxyuridine-labeling and the presence of glucagon-containing cells within these ducts. In addition, the ductal epithelia in diabetic NOD mice expressed the neuroendocrine markers neuropeptide Y and tyrosine hydroxylase. These ducts also expressed the homeobox gene product, insulin promoter factor 1. Ductal cell proliferation and expression of these markers was not observed in transgenic NOD mice (NOD-E), which do not develop clinical or histopathological symptoms of IDDM. This suggests that the observed ductal cell proliferation and differentiation was a direct result of beta-cell destruction and insulin insufficiency in these adult diabetic mice, which further suggests that these events are recapitulating islet ontogeny observed during embryogenesis. It is possible that comparable processes occur in the human diabetic pancreas.

Necrotizing myopathy induced by overexpression of interferon-γ in transgenic mice

A transgenic mouse model has been established in which the cytokine interferon‐γ (IFN‐γ) is overexpressed through the action of the acetylcholine receptor epsilon promoter acting at the neuromuscular junction. While originally developed as a model for the study of the pathogenesis of myasthenia gravis, there are important differences from both human myasthenia gravis and its animal model, experimental autoimmune myasthenia gravis. By 4 months of age there was a well‐established inflammatory, predominantly necrotizing myopathy, with marked dystrophic calcification. Dystrophic and degenerative changes in terminal axons and adjacent Schwann cells were also apparent. The acetylcholine receptor was not the primary target of the inflammatory response, since at 10 weeks of age the receptor content was not decreased and antibodies were not detected bound to the receptor. The IFNγ transgenic mouse model may provide a clinically relevant model of necrotizing myopathy for investigation of the pathological changes associated with, and presumably precipitated by, overexpression of the proinflammatory cytokine interferon‐γ on the neuromuscular junction, intramuscular nerves and myofibers.

A transgenic model for studying islet development

The regeneration of islet cells in a transgenic mouse strain harboring the interferon-gamma gene (IFN-gamma) linked to the insulin promoter DNA fragment (ins-IFN-gamma) is described. The regeneration follows the loss of islets by an immune response provoked by IFN-gamma and manifests in the proliferation of duct cells, the presence of progenitor cells, and the formation of buds and isletlike structure. All three types (A, B, and D) of four endocrine cells identified by immunolabeling are present. The progenitor cells express neuronal enzymes, tyrosine hydroxylase (TH) and glutamic acid decarboxylase (GAD), as revealed by specific antibodies. The results indicate that the islet regeneration closely resembles the embryonic islet differentiation and serves as a model for studying islet development. The expression of neuronal enzymes by islet progenitor cells signifies yet unknown relationships to the nervous tissue. GAD, recognized as an autoantigen in insulin-dependent diabetes mellitus (IDDM) and stiff-man syndrome, may provide a clue to the mechanism of autoimmune disease.

Treatment of IFN-γ Transgenic Mice With Anti–IFN-γ Reveals the Remodeling Capacity of the Adult Pancreas

Pancreatic expression of γ-interferon (IFN-γ) initiates a cascade of pathogenic changes that include pancreatic inflammation, islet cell destruction, hyperglycemia, and islet regeneration. In this study, we explore the developmental plasticity of the adult pancreas and particularly its ability to return to normoglycemia and to remodel itself from an advanced pathogenic state. This was approached by treating adult transgenic mice with a pulse of anti–IFN-γ antibody and determining the functional and morphological status of the pancreas. We demonstrated that anti–IFN-γ antibody administration led to the reduction of hyperglycemic blood glucose levels in transgenic mice. We also observed that the pancreas returned from a profoundly perturbed state toward normality. Analysis of the mitotic index indicated that cell proliferation previously associated with islet cell regeneration was greatly reduced after anti–IFN-γ administration. Our results highlight the ability of the adult pancreas to remodel itself and return from a complex pathological state to normalcy once the trophic signal inducing this pathology is removed. These data also suggest that anti–IFN-γ administration may have important clinical implications for treatment of chronic pancreatitis in humans.

The role of infiltrating macrophages in islet destruction and regrowth in a transgenic model

The expression of interferon-gamma (IFN-gamma) in pancreatic beta cells leads to a complex pathology that represents the processes of both islet destruction and islet regeneration. Inflammatory cells and the factors elicited from them participate in the development of pathology in this transgenic model. To dissect the role of infiltrating macrophages in these events, the monoclonal directed against the type 3 complement receptor (5C6) was utilized to inhibit the extravasation of macrophages. This was approached by treating transgenic mice with 5C6 for 3 or 4 months, starting from 5-7 days of age. The data presented in this report demonstrate that infiltrating macrophages are important in the observed induction of diabetes in our transgenic model. We also found that infiltrating macrophages did not play a major role in the observed proliferation and islet regeneration, but some interesting subtleties regarding the regulation of this proliferative process emerged.

Murine Transgenic Models of IDDM

Insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease. To study the immune responses leading to the breakdown of tolerance to islet β-cells, genes of putative causative agents and of major histocompatibility complex (MHC) are linked to insulin promoter and incorporated into the germ line of a mouse. The expression of transgenes are tissue specific (β-cell-specific) and their actions are analyzed during the life span of the “host” animals. These transgenic mice not only serve as models of IDDM but also provide new powerful tools for studying the many aspects of peripheral tolerance.