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Joel F Habener

Publications and source records attributed to Joel F Habener.

At least 19 recordsLinked to original sources
PubMed2006

Snail2, a mediator of epithelial-mesenchymal transitions, expressed in progenitor cells of the developing endocrine pancreas.

The mammalian pancreas develops by the expansion and morphogenesis of the epithelial cells of the foregut endoderm via the sequential activation of transcription factors that direct differentiation into the various pancreatic lineages. Implicit in this growth and differentiation are the temporal and spatial processes of cell migration and three-dimensional organization, which cooperate to form a properly functioning organ. In many organ systems, such as the kidney, heart, and neural crest derivatives, migration and tissue morphogenesis is accomplished by the transient conversion of stationary epithelial cells to migratory mesenchymal-like cells in a process known as epithelial-mesenchymal transition (EMT). We report the identification of the expression of the transcription factor Snail2/Slug, a known inducer of EMT and cell movement, in both the endocrine and exocrine cells of the developing mouse pancreas. Snail2 is expressed in Neurogenin3-positive endocrine progenitor cells, and expression is maintained during endocrine cell differentiation where it becomes increasingly restricted to the insulin-producing beta cells and somatostatin-producing delta cells. In the adult pancreas, the expression of Snail2 is maintained at low but detectable levels in all beta cells, indicating a latent role for Snail2 in the mature islet. These findings of Snail2 expression during endocrine pancreas development are relevant to the recent evidence demonstrating the involvement of EMT in the expansion of human islet tissue in vitro. EMT-like events appear to be involved in the development of the mammalian pancreas in vivo.

Animals↗
PubMed2006

Inhibition of cyclin-dependent kinase 5 activity protects pancreatic beta cells from glucotoxicity.

Type 2 diabetes (T2D) and Alzheimer disease are degenerative diseases that may share common pathophysiologic mechanisms. Neuronal dysfunction in Alzheimer patients has been linked to overactivity of the cyclin-dependent kinase 5 (CDK5) and its activator p35. Both of these proteins are expressed in the insulin-producing beta cells of the pancreas. Further, glucose enhances p35 gene expression, promoting the formation of active p35/CDK5 complexes that regulate the expression of the insulin gene. In T2D, chronic elevations of glucose, glucotoxicity, impair beta cell function. We therefore postulated that CDK5 and p35 may be responsible for this beta cell impairment and that inhibition of CDK5 might have a beneficial effect. To test this hypothesis, the pancreatic cell line INS-1 was selected as a known in vitro model of glucotoxicity, and roscovitine (10 mum) was used as a CDK5 inhibitor. Chronic exposure of INS-1 cells to high glucose (20-30 mm) reduced both insulin mRNA levels and the activity of an insulin promoter reporter gene. Inhibition of CDK5 prevented this decrease of insulin gene expression. We used DNA binding (gel shift) assays and Western immunoblots to demonstrate that cellular levels of the transcription factor PDX-1, normally decreased by glucotoxicity, were preserved with CDK5 inhibition, as was the binding of PDX-1 to the insulin promoter. Analyses of nuclear and cytoplasmic PDX-1 protein levels revealed that CDK5 inhibition restores nuclear PDX-1, without affecting its cytoplasmic concentration, suggesting that CDK5 regulates the nuclear/cytoplasm partitioning of PDX-1. Using a Myc-tagged PDX-1 construct, we showed that the translocation of PDX-1 from the nucleus to the cytoplasm during glucotoxic conditions was prevented when CDK5 was inhibited. These studies indicate that CDK5 plays a role in the loss of beta cell function under glucotoxic conditions and that CDK5 inhibitors could have therapeutic value for T2D.

Animals↗
PubMed2006

Transcription factor snail modulates hormone expression in established endocrine pancreatic cell lines.

The development of differentiated cells from undifferentiated progenitor cells is one of the central tenets of developmental biology. However, under conditions of tissue morphogenesis, regeneration, and cancer, this process of development is reversed and fully differentiated cells transition to an undifferentiated phenotype. Here we present evidence that the zinc-finger transcription factor Snail modulates this transition in differentiated pancreatic endocrine cell lines. During passage and growth of these cell lines, Snail expression is induced in a subset of cells within the culture, concomitant with a decrease in insulin and/or glucagon expression. As the cells cluster and exit the cell division cycle, nuclear levels of Snail are reduced and hormone expression is resumed. Snail represses proinsulin and proglucagon gene transcription, and reduction of Snail levels by small interfering RNA treatment increases proinsulin gene expression. We propose that Snail modulates the dynamic balance between differentiated and dedifferentiated cells allowing their migration and proliferation. These findings may be relevant to providing approaches for the enhancement of beta-cell growth in individuals with diabetes mellitus.

Cell Differentiation↗
PubMed2006

Glucagon-like peptide-1 receptor and proglucagon expression in mouse skin.

Glucagon-like peptide-1 (GLP-1) is an insulinotropic hormone expressed by alternative post-translational processing of proglucagon in the intestines, endocrine pancreas, and brain. The multiple antidiabetogenic actions of GLP-1 include stimulation of the proliferation and differentiation of the insulin-producing beta cells in the pancreas. The GLP-1 receptor is widely distributed and has been identified in the endocrine pancreas, intestinal tract, brain, lung, kidney, and heart. Here we report the expression of the GLP-1 receptor and proglucagon in the skin of newborn mice located predominantly in the hair follicles, as well as in cultures of skin-derived cells that also express nestin, a marker of cultured cells that have dedifferentiated by epithelial to mesenchymal transition. In cultured skin cells, GLP-1 activates the MAPK/ERK signal transduction pathway, associated with cellular proliferation, differentiation, and cytoprotection. No evidence was found for the activation of cAMP or Ca2+ signaling pathways. Further, redifferentiation of cultured skin-derived cells by incubation in differentiation medium containing GLP-1 induced expression of the proinsulin-derived peptide, C-peptide. These findings suggest a possible paracrine/autocrine role for GLP-1 and its receptor in skin development and possibly also in folliculogenesis.

Animals↗
PubMed2005

Redifferentiation of insulin-secreting cells after in vitro expansion of adult human pancreatic islet tissue.

Cellular replacement therapy holds promise for the treatment of diabetes mellitus but donor tissue is severely limited. Therefore, we investigated whether insulin-secreting cells could be differentiated in vitro from a monolayer of cells expanded from human donor pancreatic islets. We describe a three-step culture protocol that allows for the efficient generation of insulin-producing cell clusters from in vitro expanded, hormone-negative cells. These clusters express insulin at levels of up to 34% that of average freshly isolated human islets and secrete C-peptide upon membrane depolarization. They also contain cells expressing the other major islet hormones (glucagon, somatostatin, and pancreatic polypeptide). The source of the newly differentiated endocrine cells could either be indigenous stem/progenitor cells or the proliferation-associated dedifferentiation and subsequent redifferentiation of mature endocrine cells. The in vitro generated cell clusters may be efficacious in providing islet-like tissue for transplantation into diabetic recipients.

Adult↗
PubMed2005

IPF-1/MODY4 gene missense mutation in an Italian family with type 2 and gestational diabetes.

Maturity-onset diabetes of the young (MODY) is a monogenic autosomal-dominant form of diabetes mellitus with onset before 25 years of age. Genetic variation in insulin promoter factor-1 (IPF1) (MODY4) is uncommon but may contribute to early- or late-onset diabetes as part of a polygenic background. IPF1 is a homeodomain transcription factor required for pancreas development. Our aim was to identify whether IPF1 gene mutations play a role in Italian early-onset type 2 diabetic (T2D) patients and what functional impact mutations may have in the beta cell. We screened 40 Italian early-onset type 2 diabetic probands for IPF1 mutations, performed oral glucose tolerance tests in the unaffected family members, and performed in vitro functional studies of the mutant variant. In an extended family (Italy-6) of 46 members with clinical phenotypes of gestational diabetes, MODY, and T2D, a single nucleotide change of CCT to ACT was identified at codon 33 resulting in a Pro to Thr substitution (P33T) in the IPF1 transactivation domain that also contributes to an altered metabolic status in the unaffected NM subjects. Of the 22 genotyped Italy-6 members, 9 carried the P33T allele (NM), of whom 5 have either T2D or elevated fasting glucose levels. Oral glucose tolerance tests showed higher glucose levels at 90 minutes in unaffected NM compared with unaffected NN subjects. Of the 5 female pregnant carriers of the IPF1 mutation, 4 had pregnancies complicated by reduced birth weights, miscarriages, or early postnatal deaths. In studies in vitro, the IPF1 mutant protein (P33T) showed a reduction in DNA-binding and transcriptional activation functions as compared to the wild-type IPF1 protein. Our findings suggest that the P33T IPF1 mutation may provide an increased susceptibility to the development of gestational diabetes and MODY4 in the Italy-6 pedigree.

Abortion, Spontaneous↗
PubMed2004

Minireview: transcriptional regulation in pancreatic development.

Considerable progress has been made in the understanding of the sequential activation of signal transduction pathways and the expression of transcription factors during pancreas development. Much of this understanding has been obtained by analyses of the phenotypes of mice in which the expression of key genes has been disrupted (knockout mice). Knockout of the genes for Pdx1, Hlxb9, Isl1, or Hex results in an arrest of pancreas development at a very early stage (embryonic d 8-9). Disruption of genes encoding components of the Notch signaling pathway, e.g. Hes1 or neurogenin-3, abrogates development of the endocrine pancreas (islets of Langerhans). Disruption of transcription factor genes expressed more downstream in the developmental cascade (Beta2/NeuroD, Pax4, NKx2.2, and Nkx6.1) curtails the formation of insulin-producing beta-cells. An understanding of the importance of transcription factor genes during pancreas development has provided insights into the pathogenesis of diabetes, in which the mass of insulin-producing beta-cells is reduced.

Animals↗
PubMed2004

Cell-density-dependent regulation of actin gene expression due to changes in actin treadmilling.

The actins are essential cytoskeletal proteins required for the survival and growth of cells. The transitions between soluble (G-actin) and filamentous (F-actin) forms of actin (actin treadmilling) are complexly regulated. Here we show that the expression of the cytoplasmic beta-actin and gamma-actin genes is down-regulated in mouse fibroblasts when the cell density of the culture increases. Conversely, a dense culture replated at lower density results in increases in actin mRNA levels within a few hours. Concomitant with these changes in mRNA levels, we observe increased depolymerization of actin microfilaments at higher densities resulting in an elevated G-actin to F-actin ratio. By using actin polymerization inhibitors, we show that the density-dependent change in actin gene expression is dependent on changes in the ratio of G-actin vs. F-actin levels. Therefore, actin treadmilling and actin gene regulation are not coregulated by cell density, but represent a linear signal transduction pathway in which actin treadmilling regulates actin gene transcription. The physiological transition represented by the growth of a sparse fibroblast population into a confluent and growth-arrested population represents a useful model for the study of how the actin treadmill exerts its action on the gene expression program of cells.

3T3 Cells↗
PubMed2004

Pancreas duodenum homeobox-1 transcriptional activation requires interactions with p300.

The homeodomain transcription factor, pancreas duodenum homeobox (PDX)-1, is essential for pancreas development, insulin production, and glucose homeostasis. Mutations in pdx-1(ipf-1) are associated both with maturity-onset diabetes of the young and type 2 diabetes. PDX-1 interacts with multiple transcription factors and coregulators, including the coactivator p300, to activate the transcription of the insulin gene and other target genes within pancreatic beta-cells. In characterizing the protein-protein interactions of PDX-1 and p300, we identified mutations in PDX-1 that disrupt its function and are associated with increased or decreased interactions with p300. Several mutant PDX-1 proteins that are associated with heritable forms of diabetes in humans, in particular the mutant P63fsdelC, exhibited increased binding to a carboxy-terminal segment of p300 in the setting of decreased DNA-binding activities, suggesting that sequestration of p300 by mutant PDX-1 proteins may be an additional mechanism by which insulin gene expression is reduced in heterozygous carriers of pdx-1(ipf-1) mutations. The introduction of the point mutations S66A/Y68A in the highly conserved amino-terminal PDX-1 transactivation domain reduced the ability of PDX-1 to interact with p300, substantially diminished the transcriptional activation of PDX-1, and reduced the synergistic activation of glucose-responsive insulin promoter enhancer sequences by PDX-1, E12, and E47. We propose that interactions of PDX-1 with p300 are required for the transcriptional activation of PDX-1 target genes. Impairment of interactions between PDX-1 and p300 in pancreatic beta-cells may limit insulin production and lead to the development of diabetes.

Animals↗
PubMed2004

Glucose-induced expression of the cyclin-dependent protein kinase 5 activator p35 involved in Alzheimer's disease regulates insulin gene transcription in pancreatic beta-cells.

The deposition of amyloid within the insulin-producing islets of Langerhans in the pancreas is a common pathological finding in patients with type 2 diabetes. Its relationship with age and the progression of the disease resembles the pathological deposition of beta-amyloid in the brains of Alzheimer's patients. Endocrine cells of pancreatic islets and cells of neuronal lineages express a shared subset of specialized genes. The hyperactivity of the cyclin-dependent protein kinase CDK5, involved in the development and differentiation of the nervous system, is associated with Alzheimer's disease. Overactivity of CDK5 occurs by proteolytic cleavage and cellular mislocalization of its activator, p35. These alterations in p35/CDK5 signaling pathway may mediate, at least in part, the functional abnormalities characteristic of Alzheimer's disease. In this study we report that both the p35 and CDK5 genes are expressed in insulin-producing beta-cells of the pancreas. We detect in beta-cells the formation of an active p35/CDK5 complex with specific kinase activity. Notably, elevations of the extracellular concentration of glucose result in increases in p35 mRNA and protein levels that parallel elevations of p35/CDK5 activity. Functional studies show that p35 stimulates the activity of the insulin promoter and that the stimulation requires CDK5 because stimulation is blocked by roscovitine, an inhibitor of CDK5 activity, a dominant negative form of CDK5, and small interfering RNAs against p35. Our findings indicate that the expression of p35 and CDK5 in insulin-producing beta-cells ensembles a new signaling pathway, the activity of which is controlled by glucose, and its functional role may comprise the regulation of various biological processes in beta-cells, such as is the case for expression of the insulin gene.

Alzheimer Disease↗
PubMed2004

Triple genetic variation in the HNF-4alpha gene is associated with early-onset type 2 diabetes mellitus in a philippino family.

Maturity-onset diabetes of the young-type 1 (MODY1) is a form of monogenic type 2 diabetes mellitus (T2DM) with long-term complications due to mutations in the HNF-4alpha gene. The HNF-4alpha gene is involved in hepatic differentiation and expression of genes regulating glucose transport, glycolysis, and lipid metabolism. The abnormal glucose-stimulated insulin secretion in MODY1 subjects may be due to reduced glucose transport and glycolysis. To date, 14 mutations in the HNF-4alpha gene have been identified as a cause of either MODY1 or late-onset type 2 diabetes. So far, no screening has been performed in subjects from the Philippines. We recruited a Philippino family with autosomal dominant early-onset type 2 diabetes and screened the proband for mutations in the genes for HNF-1alpha, GCK, HNF-4alpha, IPF-1, HNF-6, and NGN3. We identified a new missense mutation in exon 5 (V199I) of the HNF-4alpha gene and 2 new single-nucleotide substitutions in intron 4, IVS4-nt4 (G --> A) and IVS4-nt20 (C --> T), all cosegregating with diabetes in the 3 affected available siblings. These variations were not present in 100 normal healthy subjects. Bioinformatic analysis suggests that these variations in the whole, and overall the IVS4-nt4 variation located at splicing site, may affect the splicing potential of intron 4. We have biochemically and clinically characterized the Philippine-1 family. We suggest that the V199I missense mutation located in the ligand binding/dimerization domain of HNF-4alpha contributes to type 2 diabetes in the Philippine-1 family. The intron variations may contribute susceptibility to diabetes.

Adult↗
PubMed2004

Human pancreatic islet-derived progenitor cell engraftment in immunocompetent mice.

The potential for the use of stem/progenitor cells for the restoration of injured or diseased tissues has garnered much interest recently, establishing a new field of research called regenerative medicine. Attention has been focused on embryonic stem cells derived from human fetal tissues. However, the use of human fetal tissue for research and transplantation is controversial. An alternative is the isolation and utilization of multipotent stem/progenitor cells derived from adult donor tissues. We have previously reported on the isolation, propagation, and partial characterization of a population of stem/progenitor cells isolated from the pancreatic islets of Langerhans of adult human donor pancreata. Here we show that these human adult tissue-derived cells, nestin-positive islet-derived stem/progenitor cells, prepared from human adult pancreata survive engraftment and produce tissue chimerism when transplanted into immunocompetent mice either under the kidney capsule or by systemic injection. These xenografts seem to induce immune tolerance by establishing a mixed chimerism in the mice. We propose that a population of stem/progenitor cells isolated from the islets of the pancreas can cross xenogeneic transplantation immune barriers, induce tissue tolerance, and grow.

Animals↗
PubMed2004

A perspective on pancreatic stem/progenitor cells.

The prevalence of both type 1 and type 2 diabetes mellitus is increasing throughout the world along with the ensuant morbidity and early mortality because of premature microvascular and macrovascular disease. Current insulin and drug therapies control diabetes, but do not cure it. Cell-based therapies offer the possibilities of a permanent cure for diabetes. Recently, success in the transplantation of pancreatic islets in the livers of type 1 diabetics has afforded the opportunity for a potential cure. However, the severe shortage of donor islets for transplantation limits the usefulness of this therapy. One approach is to exploit the use of stem cells, either embryo-derived or adult tissue-derived, as substrates to create islet tissue suitable for transplantation. Cells isolated from embryo blastocysts and from adult pancreas, liver, and bone marrow can be expanded extensively in vitro and differentiated into islet-like clusters that produce insulin, and, in some instances, can achieve glycemic control when transplanted into streptozotocin-induced diabetic mice. It is, now, also possible to envision the direct systemic administration of stem cells that would home in on and regenerate injured islets, or to administer stem cell stimulators that would enhance endogenous pancreatic stem cells to expand and differentiate into functional, insulin-producing beta-cells. This perspective discusses the potential applications of cellular medicines, in the new discipline of regenerative medicine, to achieve a cure for diabetes.

Adult↗
PubMed2004

Glucagon-like peptide 1 agonists and the development and growth of pancreatic beta-cells.

Glucagon-like peptide 1 (GLP-1) is an intestine-derived insulinotropic hormone that stimulates glucose-dependent insulin production and secretion from pancreatic beta-cells. Other recognized actions of GLP-1 are to suppress glucagon secretion and hepatic glucose output, delay gastric emptying, reduce food intake, and promote glucose disposal in peripheral tissues. All of these actions are potentially beneficial for the treatment of type 2 diabetes mellitus. Several GLP-1 agonists are in clinical trials for the treatment of diabetes. More recently, GLP-1 agonists have been shown to stimulate the growth and differentiation of pancreatic beta-cells, as well as to exert cytoprotective, antiapoptotic effects on beta-cells. Recent evidence indicates that GLP-1 agonists act on receptors on pancreas-derived stem/progenitor cells to prompt their differentiation into beta-cells. These new findings suggest an approach to create beta-cells in vitro by expanding stem/progenitor cells and then to convert them into beta-cells by treatment with GLP-1. Thus GLP-1 may be a means by which to create beta-cells ex vivo for transplantation into patients with insulinopenic type 1 diabetes and severe forms of type 2 diabetes.

Animals↗
PubMed2004

No evidence for significant transdifferentiation of bone marrow into pancreatic beta-cells in vivo.

Several recent studies have suggested that the adult bone marrow harbors cells that can differentiate into tissues from all three germ layers. Other reports have contradicted these findings or attributed them to cell fusion. In this study, we investigated whether bone marrow-derived cells contribute to the renewal of adult pancreatic endocrine cells, in particular insulin-producing beta-cells, in vivo. To address this issue, we studied mice transplanted with green fluorescent protein (GFP)-positive, sex-mismatched bone marrow. We also extended our studies to pancreatic injury models (partial pancreatectomy and streptozotocin administration). All animals showed stable full donor chimerism in the peripheral blood and microscopic analysis at 4-6 weeks and 3 months after transplantation, indicating that the GFP(+) and Y chromosome-positive donor bone marrow contributed substantially to blood, lymphatic, and interstitial cells in the pancreas. However, after examining >100,000 beta-cells, we found only 2 beta-cells positive for GFP, both of which were in control animals without pancreatic injury. Thus our study results did not support the concept that bone marrow contributes significantly to adult pancreatic beta-cell renewal.

Animals↗
PubMed2004

Cure of overt diabetes in NOD mice by transient treatment with anti-lymphocyte serum and exendin-4.

Treatment of overtly diabetic NOD mice with anti-lymphocyte serum (ALS), a polyclonal anti-T-cell antibody, abrogates autoimmunity and achieves partial clinical remission. Here we investigated whether the addition of exendin-4, a hormone that stimulates insulin secretion and beta-cell replication and differentiation, improves induction of remission by ALS. Transient treatment of overtly diabetic NOD mice with ALS and exendin-4 achieved complete remission in 23 of 26 mice (88%) within 75 days, accompanied by progressive normalization of glucose tolerance, improved islet histology, increased insulin content in the pancreas, and insulin release in response to a glucose challenge. Syngeneic islets transplanted into mice cured by treatment with ALS plus exendin-4 remained intact, and cotransfer of lymphocytes from cured mice delayed diabetes induction by adoptive transfer, suggesting the long-lasting presence of autoimmune regulatory cells. Although ALS alone also achieved reversal of diabetes, the frequency of remission was low (40%). No treatment or exendin-4 alone failed to produce remission. These results show that exendin-4 synergistically augments the remission-inducing effect of ALS. The addition of beta-cell growth factors, such as exendin-4, to immunotherapy protocols with anti-T-cell antibodies presents a potential novel approach to the cure of patients with new-onset type 1 diabetes.

Adoptive Transfer↗
PubMed2003

Regulation of glucagon-like peptide-1 receptor and calcium-sensing receptor signaling by L-histidine.

Receptor-specific agonists of the extracellular calcium-sensing receptor (CaSR) potentiate glucose-induced insulin secretion, an effect similar to that of glucagon-like peptide-1 (GLP-1). We have sequenced the full open reading frame of the CaSR from rat insulinoma (INS-1) cells and find that the predicted amino acid sequence of the receptor is identical with that of the receptor from the parathyroid gland. This receptor couples to both Gq/11 and Gi/o, and this dual coupling may partly explain the varying effects of nonspecific agonists on secretion reported previously. L-Histidine (L-His) increases the sensitivity of the CaSR to extracellular Ca2+ and potentiates glucose-dependent insulin secretion from INS-1 cells. This potentiation is partially inhibited at low extracellular [Ca2+] where the CaSR is ineffective. Coexpression of the CaSR and GLP-1 receptor (GLP-1R) produces a pertussis toxin-sensitive inhibition of GLP-1-induced cAMP production in response to elevated extracellular [Ca2+]. However, l-His potentiates cAMP response element reporter activity in INS-1 cells and in human embryonic kidney-293 cells expressing either the GLP-1R alone or the CaSR and GLP-1R. INS-1 cells express the RNA for the CaSR at a lower level than that for the GLP-1R. This difference in expression level of the receptors may explain the potentiation of insulin secretion by L-His despite coupling of the CaSR to Gi/o. In conclusion, L-His can potentiate both GLP-1R- and CaSR-activated signaling pathways, and these effects may play a role in the potentiation of glucose-induced insulin secretion in response to meals containing protein in addition to carbohydrates and fat.

Animals↗