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T Otonkoski

Publications and source records attributed to T Otonkoski.

At least 37 records · Page 2Linked to original sources
PubMed2000

Neonatal Type I diabetes associated with maternal echovirus 6 infection: a case report.

AIMS/HYPOTHESIS: Neonatal diabetes mellitus is rare, and it has not been associated with beta-cell autoimmunity. Enteroviral infections during pregnancy have been implicated as a risk factor for the later development of Type I (insulin-dependent) diabetes mellitus. We now report of a baby girl who was born severely growth-retarded with neonatal insulin-deficient diabetes, and look for evidence of intrauterine enteroviral infections and beta-cell targeted autoimmunity. METHODS: Diabetes-associated autoimmunity was studied by measurement of several types of islet cell reactive autoantibodies. The infant's T-cell responses to insulin and enterovirus antigens were recorded and enterovirus antibodies were measured both from the mother and the child. RESULTS: Several types of diabetes-associated autoantibodies were detected postnatally, including insulin autoantibodies, conventional islet cell autoantibodies and glutamic acid decarboxylase antibodies, whereas no autoantibodies were observed in the mother. The infant's T-cells showed reactivity to insulin and purified enterovirus particles. Based on serological studies, the pathogenetic process could have been triggered by an echovirus 6 infection during pregnancy. The patient's diabetes has been permanent, although there were signs of endogenous insulin production for several months. Exocrine pancreatic insufficiency was diagnosed at the age of 1 year. CONCLUSION/INTERPRETATION: These observations suggests that enteroviral infections may induce beta-cell autoimmunity even in utero.

Autoimmunity↗
PubMed2000

Genetics of neonatal hyperinsulinism.

Congenital hyperinsulinism (HI) is a clinically and genetically heterogeneous entity. The clinical heterogeneity is manifested by severity ranging from extremely severe, life threatening disease to very mild clinical symptoms, which may even be difficult to identify. Furthermore, clinical responsiveness to medical and surgical management is extremely variable. Recent discoveries have begun to clarify the molecular aetiology of this disease and thus the mechanisms responsible for this clinical heterogeneity are becoming more clear. Mutations in 4 different genes have been identified in patients with this clinical syndrome. Most cases are caused by mutations in either of the 2 subunits of the beta cell ATP sensitive K(+) channel (K(ATP)), whereas others are caused by mutations in the beta cell enzymes glucokinase and glutamate dehydrogenase. However, for as many as 50% of the cases, no genetic aetiology has yet been determined. The study of the genetics of this disease has provided important new information about beta cell physiology. Although the clinical ramifications of these findings are still limited, in some situations genetic studies might greatly aid in patient management.

B-Lymphocytes↗
PubMed2000

Practical management of hyperinsulinism in infancy.

Hyperinsulinism in infancy is one of the most difficult problems to manage in contemporary paediatric endocrinology. Although the diagnosis can usually be achieved without difficulty, it presents the paediatrician with formidable day to day management problems. Despite recent advances in understanding the pathophysiology of hyperinsulinism, the neurological outcome remains poor, and there is often a choice of unsatisfactory treatments, with life long sequelae for the child and his or her family. This paper presents a state of the art overview on management derived from a consensus workshop held by the European network for research into hyperinsulinism (ENRHI). The consensus is presented as an educational aid for paediatricians and children's nurses. It offers a practical guide to management based on the most up to date knowledge. It presents a proposed management cascade and focuses on the clinical recognition of the disease, the immediate steps that should be taken to stabilise the infant during diagnostic investigations, and the principles of definitive treatment.

Genetic Therapy↗
PubMed2000

Dominantly inherited hyperinsulinism caused by a mutation in the sulfonylurea receptor type 1.

ATP-sensitive potassium channels play a major role in linking metabolic signals to the exocytosis of insulin in the pancreatic beta cell. These channels consist of two types of protein subunit: the sulfonylurea receptor SUR1 and the inward rectifying potassium channel Kir6.2. Mutations in the genes encoding these proteins are the most common cause of congenital hyperinsulinism (CHI). Since 1973, we have followed up 38 pediatric CHI patients in Finland. We reported previously that a loss-of-function mutation in SUR1 (V187D) is responsible for CHI of the most severe cases. We have now identified a missense mutation, E1506K, within the second nucleotide binding fold of SUR1, found heterozygous in seven related patients with CHI and in their mothers. All patients have a mild form of CHI that usually can be managed by long-term diazoxide treatment. This clinical finding is in agreement with the results of heterologous coexpression studies of recombinant Kir6.2 and SUR1 carrying the E1506K mutation. Mutant K(ATP) channels were insensitive to metabolic inhibition, but a partial response to diazoxide was retained. Five of the six mothers, two of whom suffered from hypoglycemia in infancy, have developed gestational or permanent diabetes. Linkage and haplotype analysis supported a dominant pattern of inheritance in a large pedigree. In conclusion, we describe the first dominantly inherited SUR1 mutation that causes CHI in early life and predisposes to later insulin deficiency.

ATP-Binding Cassette Transporters↗
PubMed2000

Mechanisms of coxsackievirus-induced damage to human pancreatic beta-cells.

Enteroviruses may be involved in the pathogenesis of insulin-dependent diabetes mellitus, either through direct beta-cell infection or as triggers of autoimmunity. In the present study we investigated the patterns of infection in adult human islet cell preparations (consisting of 56+/-14% beta-cells) by several coxsackieviruses. The cells were infected with prototype strains of coxsackievirus B (CBV) 3, 4, and 5 as well as coxsackievirus A9 (CAV-9). The previously characterized diabetogenic strain of coxsackievirus B4 (CBV-4-E2) was used as a reference. All viruses replicated well in beta-cells, but only CBVs caused cell death. One week after infection, the insulin response of the beta-cells to glucose or glucose plus theophylline was most severely impaired by CBV-3 and CBV-5 infections. CBV-4 also caused significant functional impairment, whereas CAV-9-infected cells responded like uninfected controls. After 2 days of infection, about 40% of CBV-5-infected cells had undergone morphological changes characteristic of pyknosis, i.e. highly distorted nuclei with condensed but intact chromatin. Both mitochondria and plasma membrane were intact in these cells. DNA fragmentation was found in 5.9+/-1.1% of CBV-5-infected beta-cell nuclei (2.1+/-0.3% in controls; P<0.01). CAV-9 infection did not induce DNA fragmentation. One week after infection the majority of infected cells showed characteristics of secondary necrosis. Medium nitrite and inducible nitric oxide synthase messenger ribonucleic acid levels were not significantly up-regulated by CBV infection. These results suggest that several enteroviruses may infect human beta-cells. The infection may result in functional impairment or death of the beta-cell or may have no apparent immediate adverse effects, as shown here for CAV-9. Coxsackie B viruses cause functional impairment and beta-cell death characterized by nuclear pyknosis. Apoptosis appears to play a minor role during a productive CBV infection in beta-cells.

Adult↗
PubMed2000

Impaired migration and delayed differentiation of pancreatic islet cells in mice lacking EGF-receptors.

Pancreatic acini and islets are believed to differentiate from common ductal precursors through a process requiring various growth factors. Epidermal growth factor receptor (EGF-R) is expressed throughout the developing pancreas. We have analyzed here the pancreatic phenotype of EGF-R deficient (-/-) mice, which generally die from epithelial immaturity within the first postnatal week. The pancreata appeared macroscopically normal. The most striking feature of the EGF-R (-/-) islets was that instead of forming circular clusters, the islet cells were mainly located in streak-like structures directly associated with pancreatic ducts. Based on BrdU-labelling, proliferation of the neonatal EGF-R (-/-) beta-cells was significantly reduced (2.6+/-0.4 versus 5.8+/-0.9%, P<0.01) and the difference persisted even at 7-11 days of age. Analysis of embryonic pancreata revealed impaired branching morphogenesis and delayed islet cell differentiation in the EGF-R (-/-) mice. Islet development was analyzed further in organ cultures of E12.5 pancreata. The proportion of insulin-positive cells was significantly lower in the EGF-R (-/-) explants (27+/-6 versus 48+/-8%, P<0.01), indicating delayed differentiation of the beta cells. Branching of the epithelium into ducts was also impaired. Matrix metalloproteinase (MMP-2 and MMP-9) activity was reduced 20% in EGF-R (-/-) late-gestation pancreata, as measured by gelatinase assays. Furthermore, the levels of secreted plasminogen activator inhibitor-1 (PAI-1) were markedly higher, while no apparent differences were seen in the levels of active uPA and tPa between EGF-R (-/-) and wild-type pancreata. Our findings suggest that the perturbation of EGF-R-mediated signalling can lead to a generalized proliferation defect of the pancreatic epithelia associated with a delay in beta cell development and disturbed migration of the developing islet cells as they differentiate from their precursors. Upregulated PAI-1 production and decreased gelatinolytic activity correlated to this migration defect. An intact EGF-R pathway appears to be a prerequisite for normal pancreatic development.

Animals↗
PubMed1999

Differentiation and maturation of porcine fetal islet cells in vitro and after transplantation.

BACKGROUND: Porcine fetal pancreas is a potential source of beta cells for transplantation. The immaturity of the cells is a problem. We have defined the optimal conditions for in vitro propagation of this tissue before transplantation. METHODS: Porcine fetal pancreas tissue was obtained for tissue culture at various stages of development. Serum-containing and serum-free media and a variety of potential differentiation factors were tested. In vitro, the numbers of endocrine islet cells and their proliferation were quantified and functional maturity of the beta cells was assessed by perifusion. Growth and maturation of the cells was assessed 3 months after transplantation into nude mice. RESULTS: Highest beta cell mass was obtained from end-gestational, as compared with early fetal or neonatal, pancreas. Nicotinamide and sodium butyrate effectively increased the insulin content and the number of endocrine cells in culture. In combination, these factors led up to a 90-fold increase in the insulin content of islet-like cell clusters (ICC) as compared with untreated controls. However, a high level of cell death through apoptosis was observed in these maximally stimulated endocrine cells, and they did not survive as grafts when transplanted into nude mice. Instead, a serum-free culture medium containing 10 mM nicotinamide and 0.1 mM isobutylmethylxanthine was found to support both differentiation and proliferation of endocrine cells as loose ICCs. Insulin release from these ICCs was sensitive to glucose. When transplanted under the kidney capsule of normoglycemic nude mice, a high level of beta cell differentiation and function was evident only in the ICCs cultured in the serum-free medium, and in freshly isolated ICCs. When transplanted to hyperglycemic nude recipients, the cells cultured in serum-free medium for 3 weeks reversed hyperglycemia more consistently and rapidly than freshly isolated ICCs. CONCLUSIONS: Optimal maturation of porcine fetal pancreatic cells is obtained in serum-free medium supplemented with nicotinamide. Butyrate is a potent stimulus for beta cell differentiation but leads to increased apoptotic cell death.

Animals↗
PubMed1999

Transplantation of human fetal pancreas: fresh vs. cultured fetal islets or ICCS.

The paucity of human adult islets available for transplantation in IDDM makes the use of human fetal pancreas a potential alternative. Fetal pancreatic endocrine cells grow and differentiate over time when fresh explants or cultured islet-like cell clusters (ICCs) are transplanted under the kidney capsule in athymic nude mice. We have recently developed a procedure to isolate fetal islets, which differ from ICCs in their beta-cell content. This study was undertaken to compare the maturation and growth of grafts from purified fetal islets, containing mostly beta-cells, to grafts of mostly undifferentiated endocrine cell precursors, cultured as ICCs, and fresh, uncultured tissue. Total insulin content was highest in the fresh tissue pre-transplant while insulin levels fell precipitously during culture as either fetal islets or ICCs. Although 500 fetal islets contained more insulin than 500 ICCS before transplantation, the insulin content of the resulting grafts was the same 3 months post-transplantation. The degree of stimulation following glucose challenge was comparable, as was the histological appearance. However 70 mg of fresh tissue was needed to generate the fetal islets while only 30 mg was needed for the ICCs. Grafts of 30 mg fresh tissue also had similar total insulin contents and stimulation following glucose challenge, but, when normalized to DNA there was a significantly higher concentration of insulin in the grafts from ICCs or fetal islets. Moreover there were distinct morphological differences, with fibrous and ductal elements prominent in the grafts from fresh tissue, which were also much larger and more diffuse, with cystic elements evident macroscopically. Quantitative immunohistochemical analysis showed that grafts from cultured tissue were 48.3+/-5% positive for immunoreactive insulin compared with grafts from fresh tissue which were only 13.3+/-1.4% positive for insulin. In conclusion cultured ICCs, a heterogeneous mixture of hormone-containing and undifferentiated endocrine cells, are a preferable source for transplantation than either purified fetal islets or uncultured tissue.

Animals↗
PubMed1999

A point mutation inactivating the sulfonylurea receptor causes the severe form of persistent hyperinsulinemic hypoglycemia of infancy in Finland.

Mutations in genes encoding the ATP-regulated potassium (K(ATP)) channels of the pancreatic beta-cell (SUR1 and Kir6.2) are the major known cause of persistent hyperinsulinemic hypoglycemia of infancy (PHHI). We collected all cases of PHHI diagnosed in Finland between 1983 and 1997 (n = 24). The overall incidence was 1:40,400, but in one area of Central Finland it was as high as 1:3,200. Haplotype analysis using polymorphic markers spanning the SUR1/Kir6.2 gene cluster confirmed linkage to the 11p region. Sequence analysis revealed a novel point mutation in exon 4 of SUR1, predicting a valine to aspartic acid change at amino acid 187 (V187D). Of the total cases, 15 affected individuals harbored this mutation in heterozygous or homozygous form, and all of these had severe hyperinsulinemia that responded poorly to medical treatment and required subtotal pancreatectomy. No K(ATP) channel activity was observed in beta-cells isolated from a homozygous patient or after coexpression of recombinant Kir6.2 and SUR1 carrying the V187D mutation. Thus, the mutation produces a nonfunctional channel and, thereby, continuous insulin secretion. This unique SUR1 mutation explains the majority of PHHI cases in Finland and is strongly associated with a severe form of the disease. These findings provide diagnostic and prognostic utility for suspected PHHI patients.

ATP-Binding Cassette Transporters↗
PubMed1998

Cow milk feeding induces antibodies to insulin in children--a link between cow milk and insulin-dependent diabetes mellitus?

Exposure to cow milk (CM)-based formulas in early infancy has been associated with an increased risk of insulin-dependent diabetes mellitus (IDDM), but studies on the possible pathogenic mechanism(s) linking CM and IDDM are contradicting. We hypothesized that if CM formulas contained bovine insulin (BI), exposure to them could lead to immunization against insulin, which is the only known beta-cell-specific autoantigen in IDDM. We measured immunoglobulin G (IgG) antibodies by enzyme immunoassay (EIA) to BI and human insulin (HI) in children who received, during the first 9 months of life, either a formula containing whole CM proteins or a formula containing hydrolyzed casein (HC) peptides. BI was detectable by radioimmunoassay (RIA) and immunoblotting in the CM-based formula. At 6 months of age the children who received CM formula had higher levels of IgG antibodies to BI than children who received either HC formula or children who were exclusively breast-fed (median levels 0.480 versus 0.185, P = 0.04; and 0.480 versus 0.160, P = 0.04; respectively). Also, at 9 months of age, children in the CM group differed from the HC group (0.403 versus 0.230; P = 0.02). Antibodies to BI and HI showed a positive correlation and cross-reacted in inhibition studies. The high incidence of insulin-binding antibodies in young children with IDDM may be explained by oral immunization to BI present in CM. Exposure to BI, which differs from HI only by three amino acids, may break the tolerance to insulin.

Animals↗
PubMed1998

Growth factor-mediated proliferation and differentiation of insulin-producing INS-1 and RINm5F cells: identification of betacellulin as a novel beta-cell mitogen.

It is not clear which growth factors are crucial for the survival, proliferation, and differentiation of pancreatic beta-cells. We used the relatively differentiated rat insulinoma cell line INS-1 to elucidate this issue. Responsiveness of the DNA synthesis of serum-starved cells was studied to a wide variety of growth factors. The most potent stimulators were PRL, GH, and betacellulin, a member of the epidermal growth factor (EGF) family that has not previously been shown to be mitogenic for beta-cells. In addition to these, only vascular endothelial growth factor, insulin-like growth factor-1 and -2, had significant mitogenic activity, whereas hepatocyte growth factor, nerve growth factor-beta, platelet-derived growth factors, basic fibroblast growth factor, EGF, transforming growth factor-alpha (TGF-alpha), neu differentiation factor, and TGF-beta were inactive. None of these factors affected the insulin content of INS-1 cells. In contrast, certain differentiation factors, including nicotinamide, sodium butyrate, activin A, and 1,25-dihydroxyvitamin D3 inhibited the DNA synthesis and increased the insulin content. Also all-trans-retinoic acid had an inhibitory effect on cell DNA synthesis but no effect on insulin content. From these findings betacellulin emerges as a novel growth factor for the beta-cell. Half-maximal stimulation of INS-1 DNA synthesis was obtained with 25 pM betacellulin. Interestingly, betacellulin had no effect on RINm5F cells, whereas both EGF and TGF-alpha were slightly mitogenic. These effects may possibly be explained by differential expression of the erbB receptor tyrosine kinases. In RINm5F cells a spectrum of erbB gene expression was detected (EGF receptor/erbB-1, erbB-2/neu, and erbB-3), whereas INS-1 cells showed only expression of EGF receptor. Expression of the erbB-4 gene was undetectable in these cell lines. In summary, our results suggest that the INS-1 cell line is a suitable model for the study of beta-cell growth and differentiation because the responses to previously identified beta-cell mitogens were essentially similar to those reported in primary cells. In addition, we have identified betacellulin as a possible modulator of beta-cell growth.

Animals↗
PubMed1998

Culture of adult human islet preparations with hepatocyte growth factor and 804G matrix is mitogenic for duct cells but not for beta-cells.

It has recently been reported that human adult beta-cells proliferate during culture on an extracellular matrix prepared from rat 804G cells and in the presence of hepatocyte growth factor (HGF). The present study compares the mitogenic effect of this condition on human beta-cells and on neighboring non-endocrine duct cells. Islet cell-enriched fractions were prepared from adult human organ donors and cultured in suspension or on 804G matrix, with or without HGF. The combination of 804G matrix and HGF increased the number of 5-bromo-2'-deoxyuridine-positive (BrdU+) cells within 48 h reaching a maximum after 4 days. In sections, virtually all BrdU+ cells were negative for insulin or glucagon and for preproinsulin mRNA but expressed the ductal cell markers cytokeratin 19 and 7, carbonic anhydrase-II, and carbohydrate antigen 19-9. After 4 days of culture, the cytokeratin 19+ ductal cells exhibited a BrdU-labeling index of 30% (P < 0.01 vs. 2% without HGF and matrix), whereas <0.1% of insulin-positive and <1% of glucagon-positive cells were labeled. Formation of bilayers with ductal cells covering the endocrine cells may cause erroneous interpretation on double positivity in unsectioned tissue. It is concluded that culture of human islet cell preparations with HGF and 804G matrix stimulates the proliferation of the duct cells but not of the underlying beta-cells.

Adult↗
PubMed1997

Functional beta-cell mass after transplantation of human fetal pancreatic cells: differentiation or proliferation?

The scarcity of human adult islets available for transplantation in IDDM makes the use of human fetal pancreatic cells desirable. Human fetal pancreatic cells grow and differentiate after transplantation in nude mice. It is unclear whether proliferation of preexisting endocrine cells or differentiation of precursor cells is mainly responsible for the increased islet mass and if beta-cell enrichment before transplantation enhances the functional outcome of the graft. To answer these questions, we transplanted purified human fetal islets, islet-like cell clusters (ICCs), and fresh tissue under the kidney capsule of nude mice. Insulin content was highest in the fresh tissue but fell rapidly during culture as either fetal islets or ICCs. Although fetal islets contained fourfold more insulin than ICCs before transplantation, the insulin content of the resulting grafts was the same after 3 months in vivo. The degree of stimulation after glucose challenge was comparable; however, more tissue was needed to generate the fetal islets. Grafts of fresh tissue also had similar total insulin contents, but when normalized to DNA, insulin concentration was significantly higher in the grafts from cultured tissue. Moreover, there were distinct morphological differences; the grafts from fresh tissue were more fibrous, with prominent ductal and cystic elements. Grafts from cultured tissue were two- to threefold enriched in endocrine tissue when compared with grafts originating from fresh tissue. These results suggest that islet cells identified in the grafted ICCs are mainly derived through differentiation of endocrine precursors and that cultured ICCs are more preferable than either fetal islets or uncultured tissue for transplantation.

Age Factors↗
PubMed1996

ICA69 is expressed equally in the human endocrine and exocrine pancreas.

Islet cell autoantigen 69 kDa (ICA69) has been reported as a polypeptide antigen expressed in pancreatic beta cells, and autoimmunity against this antigen has been associated with insulin-dependent diabetes mellitus. We have studied the cell type specificity and ontogeny of ICA69 gene expression in man. The ICA69 gene was expressed in all adult human tissues. The level of expression was three-to five-times higher in the pancreas than in the brain, liver, intestine, kidney, spleen, lung or adrenal glands. Pancreatic ICA69 expression increased with age, adult levels being five times higher than the levels present at 13 weeks of gestation. Total RNA from four separate preparations of isolated human islets revealed levels of ICA69 mRNA similar to those found in the pancreas as a whole, although another islet antigen, glutamic acid decarboxylase 65, was highly enriched in the islets. In situ hybridization and immunohistochemical staining of sections of the fetal and adult pancreas revealed expression of the ICA69 gene and protein throughout the acinar, ductal, and islet tissue, but not in the mesenchyme. Analysis of ICA69 mRNA levels in human cell lines indicated expression in neural, endothelial and epithelial cells, but not in fibroblasts. In conclusion, ICA69, although highest in the pancreas, is widely distributed in other human tissues, excluding connective tissue. Within the human pancreas, ICA69 is not enriched in the islets or in the beta cells.

Adult↗
PubMed1996

A role for hepatocyte growth factor/scatter factor in fetal mesenchyme-induced pancreatic beta-cell growth.

We have investigated the role of hepatocyte growth factor/scatter factor (HGF/SF) in the growth and/or differentiation of pancreatic islet beta-cells. We found that in the human fetal pancreas immunoreactive HGF/SF receptor (c-met proto-oncogene product) is preferentially associated with the developing beta-cells. In the adult pancreas, c-met messenger RNA is highly enriched in the islets and the immunoreactive protein is also restricted to the islet beta-cells. HGF/SF messenger RNA content of fetal pancreas-derived fibroblasts is more than 10-fold higher than that of adult fibroblasts. Culture of human fetal pancreatic epithelial cells in conditioned medium from the fetal pancreatic fibroblasts caused a 2.4-fold stimulation of the formation of islet-like cell clusters that was due to both mitogenic and morphogenic effects. Beta-cell proliferation in the cell clusters was stimulated 3.5-fold by the conditioned medium, and this was associated with a marked decrease in insulin content. All of the effects of the conditioned medium were blocked by anti-HGF/SF antibody. Specificity was confirmed by overriding the blocking effect of the antibody with excess recombinant HGF/SF. Conditioned medium from adult pancreatic fibroblasts stimulated islet-like cell cluster formation only slightly, and did not affect beta-cell replication. These results suggest that HGF/SF secreted by fetal fibroblasts is mitogenic to beta-cells. Taken together, our findings indicate an important role for HGF/SF in fetal mesenchyme-induced pancreatic beta-cell growth.

Adult↗