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1.
Sci Rep ; 9(1): 8589, 2019 06 13.
Article in English | MEDLINE | ID: mdl-31197210

ABSTRACT

Insulin secretion from pancreatic beta-cells is dependent on zinc ions as essential components of insulin crystals, zinc transporters are thus involved in the insulin secretory process. Zip14 (SLC39a14) is a zinc importing protein that has an important role in glucose homeostasis. Zip14 knockout mice display hyperinsulinemia and impaired insulin secretion in high glucose conditions. Endocrine roles for Zip14 have been established in adipocytes and hepatocytes, but not yet confirmed in beta-cells. In this study, we investigated the role of Zip14 in the INS-1E beta-cell line. Zip14 mRNA was upregulated during high glucose stimulation and Zip14 silencing led to increased intracellular insulin content. Large-scale proteomics showed that Zip14 silencing down-regulated ribosomal mitochondrial proteins, many metal-binding proteins, and others involved in oxidative phosphorylation and insulin secretion. Furthermore, proliferation marker Mki67 was down-regulated in Zip14 siRNA-treated cells. In conclusion, Zip14 gene expression is glucose sensitive and silencing of Zip14 directly affects insulin processing in INS-1E beta-cells. A link between Zip14 and ribosomal mitochondrial proteins suggests altered mitochondrial RNA translation, which could disturb mitochondrial function and thereby insulin secretion. This highlights a role for Zip14 in beta-cell functioning and suggests Zip14 as a future pharmacological target in the treatment of beta-cell dysfunction.


Subject(s)
Cation Transport Proteins/metabolism , Gene Expression Regulation , Insulin Secretion/genetics , Insulin-Secreting Cells/metabolism , Proteomics , Animals , Apoptosis/drug effects , Apoptosis/genetics , Cation Transport Proteins/genetics , Cell Line , Cell Survival/drug effects , Cell Survival/genetics , Down-Regulation/drug effects , Down-Regulation/genetics , Gene Expression Regulation/drug effects , Gene Silencing , Glucose/pharmacology , Insulin/genetics , Insulin/metabolism , Insulin Secretion/drug effects , Metallothionein/genetics , Metallothionein/metabolism , Mitochondria/drug effects , Mitochondria/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Small Interfering/metabolism , Rats , Reproducibility of Results , Up-Regulation/drug effects , Up-Regulation/genetics , Zinc Transporter 8/genetics , Zinc Transporter 8/metabolism
2.
Article in English | MEDLINE | ID: mdl-28303117

ABSTRACT

Polycystic ovary syndrome (PCOS) is associated with infertility, increased androgen levels, and insulin resistance. In adipose tissue, zinc facilitates insulin signaling. Circulating zinc levels are altered in obesity, diabetes, and PCOS; and zinc supplementation can ameliorate metabolic disturbances in PCOS. In adipose tissue, expression of zinc influx transporter ZIP14 varies with body mass index (BMI), clinical markers of metabolic syndrome, and peroxisome proliferator-activated receptor gamma (PPARG). In this study, we investigated expression levels of ZIP14 and PPARG in subcutaneous adipose tissue of 36 PCOS women (17 lean and 19 obese women) compared with 23 healthy controls (7 lean and 16 obese women). Further, expression levels of zinc transporter ZIP9, a recently identified androgen receptor, and zinc efflux transporter ZNT1 were investigated, alongside lipid profile and markers of glucose metabolism [insulin degrading enzyme, retinol-binding protein 4 (RBP4), and glucose transporter 4 (GLUT4)]. We find that ZIP14 expression is reduced in obesity and positively correlates with PPARG expression, which is downregulated with increasing BMI. ZNT1 is upregulated in obesity, and both ZIP14 and ZNT1 expression significantly correlates with clinical markers of altered glucose metabolism. In addition, RBP4 and GLUT4 associate with obesity, but an association with PCOS as such was present only for PPARG and RBP4. ZIP14 and ZNT1 does not relate to clinical androgen status and ZIP9 is unaffected by all parameters investigated. In conclusion, our findings support the existence of a zinc dyshomeostasis in adipose tissue in metabolic disturbances including PCOS-related obesity.

3.
Biometals ; 29(2): 287-98, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26867900

ABSTRACT

Zinc trafficking in pancreatic beta cells is tightly regulated by zinc transporting (ZNTs) proteins. The role of different ZNTs in the beta cells is currently being clarified. ZNT8 transports zinc into insulin granules and is critical for a correct insulin crystallization and storage in the granules whereas ZNT3 knockout negatively affects beta cell function and survival. Here, we describe for the first time the sub-cellular localization of ZNT3 by immuno-gold electron microscopy and supplement previous data from knockout experiments with investigations of the effect of ZNT3 in a pancreatic beta cell line, INS-1E overexpressing ZNT3. In INS-1E cells, we found that ZNT3 was abundant in insulin containing granules located close to the plasma membrane. The level of ZNT8 mRNA was significantly decreased upon over-expression of ZNT3 at different glucose concentrations (5, 11 and 21 mM glucose). ZNT3 over-expression decreased insulin content and insulin secretion whereas ZNT3 over-expression improved the cell survival after 24 h at varying glucose concentrations (5, 11 and 21 mM). Our data suggest that ZNT3 and ZNT8 (known to regulate insulin secretion) have opposite effects on insulin synthesis and secretion possibly by a transcriptional co-regulation since mRNA expression of ZNT3 was inversely correlated to ZNT8 and ZNT3 over-expression reduced insulin synthesis and secretion in INS-1E cells. ZNT3 over-expression improved cell survival.


Subject(s)
Cation Transport Proteins/metabolism , Insulin-Secreting Cells/metabolism , Insulin/metabolism , Animals , Cation Transport Proteins/genetics , Cell Line , Cell Survival , Gene Expression , Humans , Insulin Secretion , Protective Factors , Protein Transport , Rats , Zinc Transporter 8
4.
BMC Obes ; 2: 46, 2015.
Article in English | MEDLINE | ID: mdl-26623077

ABSTRACT

BACKGROUND: The expansion and function of adipose tissue are important during the development of insulin resistance and inflammation in obesity. Zinc dyshomeostasis is common in obese individuals. In the liver, zinc influx transporter ZIP14, affects proliferation and glucose metabolism but the role of ZIP14 in adipose tissue is still unknown. This study investigates ZIP14 gene expression in human adipose tissue before and after weight loss as well as the regulation of ZIP14 during early adipogenesis. METHODS: Fourteen obese individuals were investigated before and after a 10 week weight loss intervention and compared to 14 non-obese controls. Gene expressions of ZIP14 and peroxisome proliferator-activated receptor γ (PPARγ) were measured in subcutaneous adipose tissue and correlated with metabolic and inflammatory markers. Further, we investigated gene expression of ZIP14 and PPARγ during early adipogenesis of 3T3-L1 pre-adipocytes, together with an in silico analysis of PPARγ binding motifs in the promoter sequence of ZIP14. RESULTS: ZIP14 was down-regulated in obese individuals compared to non-obese controls (p = 0.0007) and was up-regulated after weight loss (p = 0.0005). Several metabolic markers of clinical importance, including body mass index, triglyceride, and insulin resistance, were inversely correlated with ZIP14. During early adipogensis an up-regulation of ZIP14 gene expression was found. PPARγ gene expression was positively correlated with the ZIP14 gene expression in both adipose tissue and during adipogenesis. However, in silico analysis revealed that the ZIP14 promoter does not contain PPARγ-binding motifs. CONCLUSIONS: We hypothesize that ZIP14-mediated zinc influx might directly influence PPARγ activity and that ZIP14 may regulate expansion and function of adipose tissue and serve as a potential biomarker for metabolic stress.

6.
BMC Res Notes ; 7: 84, 2014 Feb 07.
Article in English | MEDLINE | ID: mdl-24502363

ABSTRACT

BACKGROUND: Zinc is essential for the activities of pancreatic ß-cells, especially insulin storage and secretion. Insulin secretion leads to co-release of zinc which contributes to the paracrine communication in the pancreatic islets. Zinc-transporting proteins (zinc-regulated transporter, iron-regulated transporter-like proteins [ZIPs] and zinc transporters [ZnTs]) and metal-buffering proteins (metallothioneins, MTs) tightly regulate intracellular zinc homeostasis. The present study investigated how modulation of cellular zinc availability affects ß-cell function using INS-1E cells. RESULTS: Using INS-1E cells, we found that zinc supplementation and zinc chelation had significant effects on insulin content and insulin secretion. Supplemental zinc within the physiological concentration range induced insulin secretion. Insulin content was reduced by zinc chelation with N,N,N',N-tektrakis(2-pyridylmethyl)-ethylenediamine. The changes in intracellular insulin content following exposure to various concentrations of zinc were reflected by changes in the expression patterns of MT-1A, ZnT-8, ZnT-5, and ZnT-3. Furthermore, high zinc concentrations induced cell necrosis while zinc chelation induced apoptosis. Finally, cell proliferation was sensitive to changes in zinc the concentration. CONCLUSION: These results indicate that the ß-cell-like function and survival of INS-1E cells are dependent on the surrounding zinc concentrations. Our results suggest that regulation of zinc homeostasis could represent a pharmacological target.


Subject(s)
Chelating Agents/pharmacology , Insulin/metabolism , Islets of Langerhans/drug effects , Zinc/pharmacology , Animals , Apoptosis/drug effects , Cation Transport Proteins/biosynthesis , Cation Transport Proteins/genetics , Cell Division/drug effects , Cell Line , DNA, Complementary/genetics , Dose-Response Relationship, Drug , Ethylenediamines/pharmacology , Gene Expression Profiling , Gene Expression Regulation/drug effects , In Vitro Techniques , Insulin Secretion , Islets of Langerhans/cytology , Islets of Langerhans/metabolism , Metallothionein/biosynthesis , Metallothionein/genetics , Necrosis , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Rats , Real-Time Polymerase Chain Reaction , Transcription, Genetic/drug effects , Zinc Transporter 8
7.
PLoS One ; 9(1): e83758, 2014.
Article in English | MEDLINE | ID: mdl-24400077

ABSTRACT

BACKGROUND AND AIMS: Glucagon-like peptide-1 (GLP-1) may provide beneficial cardiovascular effects, possibly due to enhanced myocardial energetic efficiency by increasing myocardial glucose uptake (MGU). We assessed the effects of GLP-1 on MGU in healthy subjects during normo- and hypoglycemia. MATERIALS AND METHODS: We included eighteen healthy men in two randomized, double-blinded, placebo-controlled cross-over studies. MGU was assessed with GLP-1 or saline infusion during pituitary-pancreatic normo- (plasma glucose (PG): 4.5 mM, n = 10) and hypoglycemic clamps (PG: 3.0 mM, n = 8) by positron emission tomography with (18)fluoro-deoxy-glucose ((18)F-FDG) as tracer. RESULTS: In the normoglycemia study mean (± SD) age was 25±3 years, and BMI was 22.6±0.6 kg/m(2) and in the hypoglycemia study the mean age was 23±2 years with a mean body mass index of 23±2 kg/m(2). GLP-1 did not change MGU during normoglycemia (mean (+/- SD) 0.15+/-0.04 and 0.16+/-0.03 µmol/g/min, P = 0.46) or during hypoglycemia (0.16+/-0.03 and 0.13+/-0.04 µmol/g/min, P = 0.14). However, the effect of GLP-1 on MGU was negatively correlated to baseline MGU both during normo- and hypoglycemia, (P = 0.006, r(2) = 0.64 and P = 0.018, r(2) = 0.64, respectively) and changes in MGU correlated positively with the level of insulin resistance (HOMA 2IR) during hypoglycemia, P = 0.04, r(2) = 0.54. GLP-1 mediated an increase in circulating glucagon levels at PG levels below 3.5 mM and increased glucose infusion rates during the hypoglycemia study. No differences in other circulating hormones or metabolites were found. CONCLUSIONS: While GLP-1 does not affect overall MGU, GLP-1 induces changes in MGU dependent on baseline MGU such that GLP-1 increases MGU in subjects with low baseline MGU and decreases MGU in subjects with high baseline MGU. GLP-1 preserves MGU during hypoglycemia in insulin resistant subjects. ClinicalTrials.gov registration numbers: NCT00418288: (hypoglycemia) and NCT00256256: (normoglycemia).


Subject(s)
Glucagon-Like Peptide 1/pharmacology , Glucose/metabolism , Heart/drug effects , Hypoglycemia/metabolism , Myocardium/metabolism , Adult , Blood Glucose , Cross-Over Studies , Fluorodeoxyglucose F18/metabolism , Hormones/metabolism , Humans , Hypoglycemia/diagnosis , Male , Metabolome , Positron-Emission Tomography , Young Adult
8.
Biometals ; 26(3): 369-85, 2013 Jun.
Article in English | MEDLINE | ID: mdl-23653168

ABSTRACT

Multiple sclerosis (MS) is a neurodegenerative disease caused by recurring attacks of neuroinflammation leading to neuronal death. Immune-suppressing gold salts are used for treating connective tissue diseases; however, side effects occur from systemic spread of gold ions. This is limited by exploiting macrophage-induced liberation of gold ions (dissolucytosis) from gold surfaces. Injecting gold beads in hyaluronic acid (HA) as a vehicle into the cavities of the brain can delay clinical signs of disease progression in the MS model, experimental autoimmune encephalitis (EAE). This study investigates the anti-inflammatory properties of metallic gold/HA on the gene expression of tumor necrosis factor (Tnf-α), Interleukin (Il)-1ß, Il-6, Il-10, Colony-stimulating factor (Csf)-v2, Metallothionein (Mt)-1/2, Bcl-2 associated X protein (Bax) and B cell lymphoma (Bcl)-2 in cultured J774 macrophages and in rodents with early stages of EAE. Cells grew for 5 days on gold/HA or HA, then receiving 1,000 ng/mL lipopolysaccharide (LPS) as inflammatory challenge. In the EAE experiment, 12 Lewis rats received gold injections and control groups included 11 untreated and 12 HA-treated EAE rats and five healthy animals. The experiment terminated day 9 when the first ten animals showed signs of EAE, only one of which were gold-treated (1p = 0.0367). Gene expression in the macrophages showed a statistically significant decrease in Il-6, Il-1ß and Il-10-response to LPS; interestingly HA induced a statistically significant increase of Il-10. In the EAE model gene expression of inflammatory cytokines increased markedly. Compared to EAE controls levels of Tnf-α, Il-1ß, Il-10, Il-6, IL-2, Ifn-γ, Il-17, transforming growth factor (Tgf)-ß, superoxide dismutase (Sod)-2, Mt-2 and fibroblast growth factor (Fgf)-2 were lower in the gold-treated group. HA-treated animals expressed similar or intermediate levels. Omnibus testing for reduced inflammatory response following gold-treatment was not significant, but tendencies towards a decrease in the Sod-2, Fgf-2, Il-1ß response and a higher Bdnf and IL-23 gene expression were seen. In conclusion, our findings support that bio-liberation of gold from metallic gold surfaces have anti-inflammatory properties similar to classic gold compounds, warranting further studies into the pharmacological potential of this novel gold-treatment and the possible synergistic effects of hyaluronic acid.


Subject(s)
Encephalomyelitis, Autoimmune, Experimental/drug therapy , Gene Expression Regulation/drug effects , Hyaluronic Acid/pharmacology , Immunosuppression Therapy , Macrophages/drug effects , Neurons/drug effects , Organogold Compounds/pharmacology , Animals , Cell Survival/drug effects , Cells, Cultured , Encephalomyelitis, Autoimmune, Experimental/genetics , Encephalomyelitis, Autoimmune, Experimental/immunology , Female , Gene Expression Profiling , Hyaluronic Acid/chemistry , Macrophages/immunology , Mice , Neurons/metabolism , Organogold Compounds/chemistry , Rats , Rats, Inbred Lew , Real-Time Polymerase Chain Reaction
9.
Nephrol Dial Transplant ; 27(11): 4211-8, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22910041

ABSTRACT

BACKGROUND: End-stage renal failure (ESRF) patients demonstrate augmented growth hormone (GH) secretion, but normal insulin-like growth factor-I (IGF-I) concentrations, indicating a state of GH resistance. To test this hypothesis, we compared the IGF-I response with exogenous GH in haemodialysis patients and healthy controls, with special focus on free GH and bioactive IGF-I. METHODS: Ultrafiltered free GH and total GH were measured in serum collected hourly for 24 h at baseline and after 7 days of recombinant human (rh) GH (50 µg/kg/day) treatment in 11 non-diabetic haemodialysis patients and 10 matched controls. Serum levels of bioactive IGF-I (determined by cell-based IGF-I receptor activation assay), total IGF-I and the GH-binding protein (GHBP) were assayed twice daily. RESULTS: At baseline, patients showed elevated total GH (24 ± 5 versus 9 ± 1 µg/L × h, P < 0.02), free GH (21 ± 5 versus 7 ± 1 µg/L × h, P < 0.02), reduced GHBP (1.5 ± 0.3 versus 2.5 ± 0.2 nmol/L, P < 0.01), high-normal total IGF-I (173 ± 18 versus 135 ± 14 µg/L, P = 0.12) and subnormal bioactive IGF-I (2.1 ± 0.3 versus 2.8 ± 0.2 µg/L, P < 0.05) when compared with controls. After 7 days of rhGH treatment, there was a greater GH increase in the non-diabetic haemodialysis patients than in controls (total GH: 293 ± 33 versus 166 ± 13 µg/L × h, P < 0.001; free GH: 284 ± 40 versus 126 ± 15 µg/L × h, P < 0.001). GHB remained unaffected and total IGF-I increased to the same extent in patients and controls (701 ± 87 versus 572 ± 33 µg/L, P = 0.17), whereas bioactive IGF-I tended to be lower in patients (5.37 ± 0.55 versus 6.63 ± 0.25 µg/L, P < 0.10). When adjusting for the actual increments in plasma GH, the ability of exogenous GH to stimulate bioactive IGF-I levels was reduced by ~50% in ESRF (P < 0.02), whereas the response of total IGF-I remained normal (74%; P= 0.18) CONCLUSIONS: The study demonstrates that ESRF is associated with markedly elevated serum levels of free GH. Furthermore changes in bioactive, but not immunoreactive, IGF-I indicated that the hepatic sensitivity to GH was reduced by 50% in ESRF patients. Clearly, the physiological importance of our observations awaits further studies, but they suggest that changes in total IGF-I may not necessarily reflect changes in the endogenous activity of IGF-I in ESRF patients on GH treatment.


Subject(s)
Growth Hormone/blood , Growth Hormone/therapeutic use , Insulin-Like Growth Factor I/analysis , Kidney Failure, Chronic/blood , Kidney Failure, Chronic/drug therapy , Renal Dialysis/statistics & numerical data , Adult , Female , Humans , Male , Middle Aged , Ultrafiltration
10.
Biometals ; 25(1): 1-8, 2012 Feb.
Article in English | MEDLINE | ID: mdl-21866305

ABSTRACT

The review collects the emerging information about zinc transporter 3 (ZnT3). ZnT3 has been associated with Alzheimer's disease, airway diseases and diabetes. ZnT3 was discovered and cloned in 1996. Since then, the major interest in the protein has been in its ability to transport zinc into pre-synaptic vesicles of glutamatergic neurones and its role during the development of amyloid ß plaques in Alzheimer's disease. Increasing evidence suggests that ZnT3 is present in various cell types like different cell types in the brain, cells from adipose tissue, beta-cells from pancreatic islets, epithelial cells, cells from testis, prostate cancer cells and cells from retina. The expression of ZnT3 is regulated by age, hormones, fatty acids, zinc chelation, and glucose.


Subject(s)
Cation Transport Proteins/metabolism , Zinc/metabolism , Aging/physiology , Alzheimer Disease/physiopathology , Animals , Brain/metabolism , Cation Transport Proteins/genetics , Cell Death/physiology , Diabetes Mellitus/physiopathology , Gonadal Hormones/metabolism , Humans
11.
Expert Opin Ther Targets ; 15(10): 1153-62, 2011 Oct.
Article in English | MEDLINE | ID: mdl-21749267

ABSTRACT

INTRODUCTION: Epidemiological evidence suggests an association between type 2 diabetes (T2DM) and Alzheimer's disease (AD), in that one disease increases the risk of the other. T2DM and AD share several molecular processes which underlie the tissue degeneration in either disease. Disturbances in insulin signaling may be the link between the two conditions. Drugs originally developed for T2DM are currently being considered as possible novel agents in the treatment of AD. AREAS COVERED: This review discusses the potential role of glucagon-like peptide -1 (GLP-1) treatment in AD. GLP-1 receptors are expressed in areas of the brain important to memory and learning, and GLP-1 has growth-factor-like properties similar to insulin. A key neuropathological feature of AD is the accumulation of amyloid-beta (Aß). In preclinical studies, GLP-1 and longer lasting analogues have been shown to have both neuroprotective and neurotrophic effects, and to protect synaptic activity in the brain from Aß toxicity. EXPERT OPINION: A convincing amount of evidence has shown a beneficial effect of GLP-1 agonist treatment on cognitive function, memory and learning in experimental models of AD. GLP-1 analogues may therefore be the new therapeutic agent of choice for intervention in AD.


Subject(s)
Alzheimer Disease/drug therapy , Amyloid beta-Peptides/drug effects , Diabetes Mellitus, Type 2/drug therapy , Glucagon-Like Peptide 1/therapeutic use , Alzheimer Disease/epidemiology , Alzheimer Disease/metabolism , Alzheimer Disease/physiopathology , Amyloid beta-Peptides/metabolism , Animals , Cognition , Diabetes Mellitus, Type 2/epidemiology , Glucagon-Like Peptide 1/pharmacology , Humans , Learning , Memory , Neuroprotective Agents/pharmacology , Neuroprotective Agents/therapeutic use , Rats
12.
Histol Histopathol ; 26(6): 689-97, 2011 06.
Article in English | MEDLINE | ID: mdl-21472684

ABSTRACT

In clinical medicine metallic silver is used as anti-bacterial coating on various catheters, bandages and prostheses. By means of dissolucytosis, i.e. extracellular macrophage-mediated bio-liberation of metal ions, silver ions are continuously liberated from silver surfaces starting within minutes of exposure. The present study investigates how bio-liberation and subsequent cellular uptake of silver ions affects cell viability and cell signalling within the first 3-24 hours of exposure when J774 macrophages are grown directly on a silver surface. Autometallography (AMG) was applied to demonstrate cytoplasmatic silver uptake and localisation after 1, 3, 12 and 24 hours of exposure to metallic silver. From 12 hours onwards the cells were completely filled with silver enhanced silver-sulphur nanocrystals (AMG-silver grains). At the ultrastructural level, the silver accumulations were located to lysosome-like structures. An immunoassay cell death kit found silver-induced apoptosis after 12 and 24 hours of exposure. Necrosis was seen at the same times. Judged by mRNA analysis silver exposure statistically significantly induces TNF-α and m-CSF gene expression, especially at 3 hours. Furthermore, anti-inflammatory IL-10 transcription is reduced by silver uptake and 24 hours of silver exposure induces massive iNOS-2 gene expression. At the same time silver exposure increases the gene expression of metallothionein (MT-I/MT-II), a cystein-rich protein known for its role in detoxifying heavy metals. Our data suggest that silver ions liberated from metallic silver surfaces accumulate in lysosomes, reduce macrophage viability by apoptosis and necrosis and induce a pro-inflammatory response.


Subject(s)
Apoptosis/drug effects , Inflammation/chemically induced , Macrophages/drug effects , Macrophages/ultrastructure , Silver/toxicity , Animals , Cell Line , Macrophages/metabolism , Mice , Microscopy, Electron, Transmission , Reverse Transcriptase Polymerase Chain Reaction , Silver/metabolism
13.
APMIS ; 119(2): 93-102, 2011 Feb.
Article in English | MEDLINE | ID: mdl-21208276

ABSTRACT

Zinc is essential for the crystallization of insulin in pancreatic ß-cells and is thought to induce apoptosis in a dose-dependent manner, thereby regulating ß-cell mass. Therefore, a tight intracellular regulation of Zn²(+) is required. The zinc-transporter family SLC30A is an important factor in the regulation of zinc homeostasis. The aim of this study was to examine the effect of the zinc transporters ZnT3 and ZnT8 on insulin metabolism and apoptosis. Both these proteins are present in pancreatic ß-cells and have been linked to diabetes. The objective of our study was to perform a considerable siRNA-mediated knock-down of ZnT3 and ZnT8 in INS-1E cells, a pancreatic ß-cell model, and afterwards examine the impact on cell viability and insulin metabolism. Increased levels of apoptosis were observed after knock-down of both ZnT3 and ZnT8. Insulin secretion was significantly reduced by ZnT3 knock-down, whereas knock-down of ZnT8 resulted in increased intracellular content of insulin accompanied by a relatively lowered secretion. Both zinc transporters in this way seem to play a role in ß-cell survival and the ability of these cells to react appropriately to surrounding glucose concentrations.


Subject(s)
Apoptosis , Cation Transport Proteins/physiology , Insulin-Secreting Cells/metabolism , Insulin/biosynthesis , RNA, Small Interfering/genetics , Animals , Cation Transport Proteins/antagonists & inhibitors , Cation Transport Proteins/genetics , Cells, Cultured , Insulin/metabolism , Insulin Secretion , Insulin-Secreting Cells/pathology , Rats , Zinc Transporter 8
14.
PLoS One ; 4(5): e5684, 2009 May 25.
Article in English | MEDLINE | ID: mdl-19492079

ABSTRACT

BACKGROUND: Ion transporters of the Slc30A- (ZnT-) family regulate zinc fluxes into sub-cellular compartments. beta-cells depend on zinc for both insulin crystallization and regulation of cell mass. METHODOLOGY/PRINCIPAL FINDINGS: This study examined: the effect of glucose and zinc chelation on ZnT gene and protein levels and apoptosis in beta-cells and pancreatic islets, the effects of ZnT-3 knock-down on insulin secretion in a beta-cell line and ZnT-3 knock-out on glucose metabolism in mice during streptozotocin-induced beta-cell stress. In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression. Zinc chelation by DEDTC lowered INS-1E insulin content and insulin expression. Furthermore, zinc depletion increased ZnT-3- and decreased ZnT-8 gene expression whereas the amount of ZnT-3 protein in the cells was decreased. Zinc depletion and high glucose induced apoptosis and necrosis in INS-1E cells. The most responsive zinc transporter, ZnT-3, was investigated further; by immunohistochemistry and western blotting ZnT-3 was demonstrated in INS-1E cells. 44% knock-down of ZnT-3 by siRNA transfection in INS-1E cells decreased insulin expression and secretion. Streptozotocin-treated mice had higher glucose levels after ZnT-3 knock-out, particularly in overt diabetic animals. CONCLUSION/SIGNIFICANCE: Zinc transporting proteins in beta-cells respond to variations in glucose and zinc levels. ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels. Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment.


Subject(s)
Carrier Proteins/metabolism , Glucose/pharmacology , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Insulin/metabolism , Membrane Proteins/metabolism , Stress, Physiological/drug effects , Zinc/pharmacology , Animals , Blood Glucose/drug effects , Carrier Proteins/genetics , Cation Transport Proteins/genetics , Cation Transport Proteins/metabolism , Cell Death/drug effects , Cell Line , Chelating Agents/pharmacology , Dose-Response Relationship, Drug , Fasting/blood , Gene Expression Regulation/drug effects , Gene Knockdown Techniques , Glucose/metabolism , Hyperglycemia/metabolism , Insulin/genetics , Insulin Secretion , Insulin-Secreting Cells/cytology , Membrane Proteins/genetics , Membrane Transport Proteins , Mice , Mice, Knockout , Rats , Streptozocin , Zinc Transporter 8
15.
BMC Endocr Disord ; 9: 7, 2009 Feb 25.
Article in English | MEDLINE | ID: mdl-19243577

ABSTRACT

BACKGROUND: Beta-cells are extremely rich in zinc and zinc homeostasis is regulated by zinc transporter proteins. beta-cells are sensitive to cytokines, interleukin-1beta (IL-1beta) has been associated with beta-cell dysfunction and -death in both type 1 and type 2 diabetes. This study explores the regulation of zinc transporters following cytokine exposure. METHODS: The effects of cytokines IL-1beta, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) on zinc transporter gene expression were measured in INS-1-cells and rat pancreatic islets. Being the more sensitive transporter, we further explored ZnT8 (Slc30A8): the effect of ZnT8 over expression on cytokine induced apoptosis was investigated as well as expression of the insulin gene and two apoptosis associated genes, BAX and BCL2. RESULTS: Our results showed a dynamic response of genes responsible for beta-cell zinc homeostasis to cytokines: IL-1beta down regulated a number of zinc-transporters, most strikingly ZnT8 in both islets and INS-1 cells. The effect was even more pronounced when mixing the cytokines. TNF-alpha had little effect on zinc transporter expression. IFN-gamma down regulated a number of zinc transporters. Insulin expression was down regulated by all cytokines. ZnT8 over expressing cells were more sensitive to IL-1beta induced apoptosis whereas no differences were observed with IFN-gamma, TNF-alpha, or a mixture of cytokines. CONCLUSION: The zinc transporting system in beta-cells is influenced by the exposure to cytokines. Particularly ZnT8, which has been associated with the development of diabetes, seems to be cytokine sensitive.

16.
Mol Cell Endocrinol ; 264(1-2): 68-73, 2007 Jan 29.
Article in English | MEDLINE | ID: mdl-17118530

ABSTRACT

Zinc ions influence adipose tissue metabolism by regulating leptin secretion and by promoting free fatty acid release and glucose uptake. The mechanisms controlling zinc metabolism in adipose tissue are unknown. We therefore examined the gene-expression levels of a number of zinc-transporting proteins in adipose tissue, comparing subcutaneous fat with visceral fat from lean and obese humans. Both ZnT-proteins responsible for zinc transport from cytosol to extracellular compartments and intracellular vesicles and Zip-proteins responsible for zinc transport to the cytoplasm were expressed in all samples. This suggests that zinc metabolism in adipocytes is actively controlled by zinc-transporters. The expression levels were different in lean and obese subjects suggesting a role for these proteins in obesity. Furthermore, the expression levels were different from subcutaneous fat to intra-abdominal fat suggesting that the metabolic activity in adipocytes is to some extent dependent upon zinc and the activity of zinc-transporting proteins or vice versa.


Subject(s)
Adipocytes/metabolism , Carrier Proteins/biosynthesis , Gene Expression Regulation , Obesity/metabolism , Subcutaneous Fat/metabolism , Zinc/metabolism , Adult , Fatty Acids/metabolism , Female , Glucose/metabolism , Humans , Leptin/metabolism , Middle Aged
17.
Diabetes ; 55(3): 574-81, 2006 Mar.
Article in English | MEDLINE | ID: mdl-16505218

ABSTRACT

Tomosyn, a syntaxin-binding protein, is capable of dissociating mammalian homolog of the Caenorhabditis elegans unc-18 gene from syntaxin and is involved in the regulation of exocytosis. We have investigated the expression, cellular localization, and functional role of tomosyn in pancreatic beta-cells. Western blotting revealed a 130-kDa protein corresponding to tomosyn in insulin-secreting beta-cell lines. RT-PCR amplification showed that b-, m-, and s-tomosyn isoform mRNAs are expressed in beta-cell lines and rat pancreatic islets. Immunohistochemistry revealed punctate tomosyn immunoreactivity in the cytoplasm of insulin-, glucagon-, pancreatic polypeptide-, and somatostatin-containing islet cells. Syntaxin 1 coimmunoprecipitated with tomosyn in extracts of insulin-secreting cells. Overexpression of m-tomosyn in mouse beta-cells significantly decreased exocytosis, whereas inhibition of tomosyn expression by small interfering RNA increased exocytosis. Hence, in the pancreatic beta-cell, tomosyn negatively regulates insulin exocytosis.


Subject(s)
Exocytosis , Insulin-Secreting Cells/metabolism , Insulin/metabolism , Nerve Tissue Proteins/physiology , R-SNARE Proteins/physiology , Animals , Calcium/metabolism , Cells, Cultured , Colforsin/pharmacology , Glucose/pharmacology , Male , Mice , Mice, Obese , Nerve Tissue Proteins/analysis , Nerve Tissue Proteins/genetics , R-SNARE Proteins/analysis , R-SNARE Proteins/genetics , RNA, Messenger/analysis , Rats , Rats, Sprague-Dawley , Syntaxin 1/analysis
18.
Proc Natl Acad Sci U S A ; 102(29): 10303-8, 2005 Jul 19.
Article in English | MEDLINE | ID: mdl-16014415

ABSTRACT

Cytosolic free Ca2+ plays an important role in the molecular mechanisms leading to regulated insulin secretion by the pancreatic beta cell. A number of Ca2+-binding proteins have been implicated in this process. Here, we define the role of the Ca2+-binding protein neuronal Ca2+ sensor-1 (NCS-1) in insulin secretion. In pancreatic beta cells, NCS-1 increases exocytosis by promoting the priming of secretory granules for release and increasing the number of granules residing in the readily releasable pool. The effect of NCS-1 on exocytosis is mediated through an increase in phosphatidylinositol (PI) 4-kinase beta activity and the generation of phosphoinositides, specifically PI 4-phosphate and PI 4,5-bisphosphate. In turn, PI 4,5-bisphosphate controls exocytosis through the Ca2+-dependent activator protein for secretion present in beta cells. Our results provide evidence for an essential role of phosphoinositide synthesis in the regulation of glucose-induced insulin secretion by the pancreatic beta cell. We also demonstrate that NCS-1 and its downstream target, PI 4-kinase beta, are critical players in this process by virtue of their capacity to regulate the release competence of the secretory granules.


Subject(s)
1-Phosphatidylinositol 4-Kinase/metabolism , Calcium-Binding Proteins/metabolism , Exocytosis/physiology , Insulin/metabolism , Islets of Langerhans/enzymology , Nerve Tissue Proteins/metabolism , 1-Phosphatidylinositol 4-Kinase/physiology , Animals , Cell Fractionation , Cells, Cultured , Electric Capacitance , Enzyme Activation/physiology , Enzyme-Linked Immunosorbent Assay , Female , Glucose/metabolism , Green Fluorescent Proteins , Immunoblotting , Immunohistochemistry , Insulin Secretion , Islets of Langerhans/metabolism , Mice , Mice, Inbred Strains , Neuronal Calcium-Sensor Proteins , Neuropeptides , Patch-Clamp Techniques
19.
Basic Clin Pharmacol Toxicol ; 97(1): 22-8, 2005 Jul.
Article in English | MEDLINE | ID: mdl-15943755

ABSTRACT

The study examined static and dynamic parathyroid hormone and calcitonin secretion after radioiodine, and was a retrospective study of patients having received radioiodine for benign conditions 8-12 years earlier. In one group of patients parathyroid hormone and calcium were measured as single blood tests, in a second group of patients parathyroid hormone and calcitonin secretion capacity were measured during a hypocalcaemic citrate-clamp and a hypercalcaemic stimulation test. Baseline calcium and parathyroid hormone were normal within expected ranges 8-12 years after radioiodine. Stimulated parathyroid hormone or calcitonin secretion did not differ from an age- and gender-matched control group. Radioiodine in doses used for benign thyroid diseases appears safe with regard to parathyroid hormone and calcitonin secretion.


Subject(s)
Iodine Radioisotopes/adverse effects , Parathyroid Glands/radiation effects , Thyroid Diseases/radiotherapy , Thyroid Gland/radiation effects , Aged , Calcitonin/blood , Calcium/blood , Female , Humans , Male , Middle Aged , Parathyroid Glands/physiopathology , Parathyroid Hormone/blood , Retrospective Studies , Thyroid Diseases/physiopathology , Thyroid Gland/physiopathology
20.
Basic Clin Pharmacol Toxicol ; 96(6): 410-2, 2005 Jun.
Article in English | MEDLINE | ID: mdl-15910403

ABSTRACT

Polycystic ovary syndrome is characterized among other things by oligo-amenorrhea and may account for more than 75% of cases with anoluvatory infertility. Due to its positive effects on polycystic ovary syndrome-induced infertility metformin has become one of the most common drugs used in this group of patients. The efficacy of the drug as well as the first reports on metformin used in pregnancy has encouraged the continued use of the drug after conception. This MiniReview reviews the current pros and cons of metformin use in pregnancy while awaiting the results of ongoing randomised, controlled clinical trials addressing the subject.


Subject(s)
Metformin/adverse effects , Metformin/therapeutic use , Polycystic Ovary Syndrome/drug therapy , Abnormalities, Drug-Induced , Abortion, Spontaneous , Anencephaly/chemically induced , Animals , Anophthalmos/chemically induced , Blastocyst/drug effects , Female , Fertility/drug effects , Humans , Hypoglycemic Agents/adverse effects , Hypoglycemic Agents/therapeutic use , Ovulation/drug effects , Pregnancy , Pregnancy Outcome
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