scholarly journals Glucocorticoids increase insulin binding and the amount of insulin-receptor mRNA in human cultured lymphocytes

1988 ◽  
Vol 249 (3) ◽  
pp. 715-719 ◽  
Author(s):  
Y Shibasaki ◽  
H Sakura ◽  
M Odawara ◽  
M Shibuya ◽  
Y Kanazawa ◽  
...  
Keyword(s):  
Cell Surface ◽  
Insulin Receptor ◽  
De Novo ◽  
Insulin Receptors ◽  
Fold Increase ◽  
Insulin Binding ◽  
Dependent Manner ◽  
Receptor Mrna ◽  
Dose Dependent ◽  
Receptor Mrnas

The effect of steroid hormones on insulin binding and the amount of insulin-receptor mRNA was examined in IM-9 lymphocytes. Cortisol and cortexolone, but not oestrogen, increased both the binding of insulin and the amount of insulin-receptor mRNA in a time- and dose-dependent manner. Cortisol was most potent, and induced a 2-fold increase in insulin binding and a 4-fold increase in mRNA. The elevation in binding was due to an increased number of insulin receptors at the cell surface. The increase in mRNA involved all four of the insulin-receptor mRNAs and could not be inhibited by cycloheximide. The cortisol-induced increase in mRNA was associated with a 3-4-fold increase in the synthesis of pro-receptor. The relative potency of the three steroids indicated that these effects were mediated by an interaction with the glucocorticoid receptor. The results of this study suggest that cortisol can increase the number of insulin receptors at the cell surface by increasing the amounts of insulin-receptor mRNA and the synthesis de novo of insulin receptors.

In vitro regulation of human monocyte insulin receptors by insulin and 1-methyl-3-isobutylxanthine

1983 ◽  
Vol 245 (5) ◽  
pp. E494-E501
Author(s):  
R. H. Whitson ◽  
S. A. Kaplan
Keyword(s):  
Insulin Receptor ◽  
Cultured Cells ◽  
Phosphodiesterase Inhibitor ◽  
Insulin Receptors ◽  
Human Monocyte ◽  
Insulin Binding ◽  
Receptor Downregulation ◽  
Independent Process ◽  
Dose Dependent

Monocytes separated from human blood by Ficoll-Hypaque and adherence to polystyrene flasks were maintained successfully in culture for 7 days. The cultured cells showed normal morphology and good viability. The insulin binding properties of the cultured monocytes were also identical to those of fresh monocytes. In vitro pretreatment of the monocytes with insulin decreased both the number and affinity of insulin receptors, resulting in a 72% reduction in the binding of tracer quantities of 125I-insulin. Insulin-induced receptor down regulation was dose-dependent and specific to the insulin receptor. Monocytes pretreated with insulin in the presence of the cyclic nucleotide phosphodiesterase inhibitor 1-methyl-3-isobutylxanthine (MIX) lost significantly fewer insulin receptors than monocytes treated with insulin alone. Tracer binding to these cells was 62% of control levels. MIX had no effect on basal insulin binding. The cAMP analogues N6,O2'-dibutyryl cAMP and 8-bromo-cAMP did not counteract insulin-induced receptor downregulation by themselves and did not significantly enhance the effects of MIX. These results indicate that MIX may counteract insulin receptor downregulation by a cAMP-independent process.

Increased hepatic insulin proreceptor-to-receptor ratio in diabetes: a possible processing defect

1991 ◽  
Vol 261 (5) ◽  
pp. E562-E571
Author(s):  
D. Dardevet ◽  
K. Komori ◽  
C. Grunfeld ◽  
S. A. Rosenzweig ◽  
M. G. Buse
Keyword(s):  
Insulin Receptor ◽  
Insulin Dose ◽  
Total Yield ◽  
Insulin Receptors ◽  
Fold Increase ◽  
Diabetic Rats ◽  
Beta Subunits ◽  
Uncontrolled Diabetes ◽  
Lentil Lectin ◽  
Dose Dependent

Hepatic insulin proreceptors and receptors were studied in control and in ketotic diabetic rats 2-4 wk after streptozotocin treatment. Solubilized preparations were partially purified by wheat germ agglutinin-agarose (WGA) and lentil lectin agarose (LLA) chromatography to enrich eluates in insulin receptors and proreceptors, respectively. After phosphorylation with [gamma-32P]ATP, an approximately 190-kDa glycoprotein was identified in LLA eluates as the insulin proreceptor, based on insulin dose-dependent tyrosine autophosphorylation, immunoprecipitation with insulin receptor-specific antibodies, and high-mannose glycosylation. Mature approximately 95 kDa phosphorylated beta-subunits were present in both LLA and WGA eluates. LLA also showed phosphorylated partially processed beta-subunits (approximately 85 kDa) and proreceptors (approximately 190 kDa). Proreceptors comprised less than 1% of the total yield of hepatic insulin receptors. The incorporation of 32P into proreceptors (per gram liver or DNA) was 4.7- or 4.5-fold greater in diabetic vs. control rats, whereas receptor labeling increased only 1.8- or 1.5-fold in diabetic rats. beta-Subunit autophosphorylation per receptor was identical in control and diabetic rats. The phosphorylation data suggested a diabetes-associated 2.6-fold increase in proreceptor-to-receptor ratios. When assessed by cross-linking with 125I-labeled insulin or by immunoblotting, proreceptor-to-receptor ratios were increased 1.5- and 3.1-fold, respectively, in diabetic rats. The data suggest that uncontrolled diabetes may alter insulin receptor processing.

scholarly journals Lectin-induced alterations of the interaction of insulin and insulin-like growth factor 1 receptors with their ligands

2008 ◽  
Vol 73 (8-9) ◽  
pp. 793-804 ◽  
Author(s):  
Romana Masnikosa ◽  
Anna Nikolic ◽  
Olgica Nedic
Keyword(s):  
Growth Factor ◽  
Cell Membrane ◽  
Wheat Germ ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Dependent Manner ◽  
Con A ◽  
High Affinity ◽  
Igf I ◽  
Dose Dependent

In order to study whether the carbohydrate moieties of the human placental IGF-I receptor (IGF1R), IGF-II receptor (IGF2R) and insulin receptors (IRs) play a role in ligand binding, solubilised cell membrane preparations were incubated with 125I-labelled IGF-I, IGF-II and insulin in the presence of lectins with different sugar specificities. Three incubation procedures were tested: ligand-first, co-incubation and lectin-first incubation. Wheat germ agglutinin (WGA), concanavalin A (Con A) and phytohaemagglutinin (PHA) altered the binding of IGF-I and insulin to their high-affinity receptors in a lectin specific and dose-dependent manner, whereas these lectins did not affect the interaction of IGF-II with its receptor(s). Moreover, the same lectins either inhibited or enhanced IGF-I and insulin binding, depending on the incubation scheme. These results also suggest that IR-A and IR-B from human placenta might be differently glycosylated.

scholarly journals Absence of down-regulation of the insulin receptor by insulin. A possible mechanism of insulin resistance in the rat

1983 ◽  
Vol 210 (2) ◽  
pp. 373-378 ◽  
Author(s):  
A P Walker ◽  
D J Flint
Keyword(s):  
Insulin Resistance ◽  
Insulin Receptor ◽  
Culture Medium ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Recycling Process ◽  
Down Regulation ◽  
Pregnant Rats ◽  
Pregnancy And Lactation ◽  
Dose Dependent

Insulin resistance occurs in rat adipocytes during pregnancy and lactation despite increased or normal insulin binding respectively; this suggests that a post-receptor defect exists. The possibility has been examined that, although insulin binding occurs normally, internalization of insulin or its receptor may be impaired in these states. Insulin produced a dose-dependent reduction in the number of insulin receptors on adipocytes from virgin rats maintained in culture medium, probably due to internalization of the hormone-receptor complex. In contrast, adipocytes from pregnant and lactating rats did not exhibit this ‘down-regulation’ phenomenon. Down regulation was, however, apparent in all groups when the experiments were performed in Tris buffer (where receptor recycling is inhibited), suggesting that in pregnant and lactating rats insulin receptors are rapidly recycled back to the plasma membrane, whereas in virgin rats this recycling process is less effective. Internalization of insulin was also determined by using 125I-labelled insulin. Adipocytes from pregnant and lactating rats appeared to internalize similar amounts of insulin to virgin rats. In the presence of the lysosomal inhibitor chloroquine, adipocytes from pregnant rats internalized more insulin than virgin or lactating rats. These results suggest that adipocytes from pregnant and lactating rats internalize insulin and its receptor normally, whereas intracellular processing of the insulin receptor may differ from that in virgin rats. In addition the rate of lysosomal degradation of insulin may be altered in adipocytes from pregnant rats.

Intracellular (Golgi) receptors for insulinlike peptides in rat liver

1980 ◽  
Vol 58 (10) ◽  
pp. 1075-1081 ◽  
Author(s):  
Barry I. Posner ◽  
Roberto M. Gonzalez ◽  
Harvey J. Guyda
Keyword(s):  
Insulin Receptor ◽  
Rat Liver ◽  
Insulin Binding ◽  
Inhibitory Effect ◽  
Mass Action ◽  
Dependent Manner ◽  
Temperature Dependent ◽  
Biochemical Basis ◽  
High Level ◽  
Dose Dependent

An insulinlike peptide (ILAs) has been isolated in our laboratory from human serum. The binding of 125I-labeled ILAs was studied in subcellular fractions from rat liver and found to be much greater in microsomes than purified plasmalemma. The high level of microsomal binding was due to a particular enrichment of binding sites in Golgi elements. Binding to Golgi was time and temperature dependent and was augmented by an increase of either subcellular fraction or 125I-labeled ILAs in accordance with a mass action process. Degradation of 125I-labeled ILAs was greatest in the Golgi vesicle fraction and was reduced by incubation at 4 °C. Bound 125I-labeled ILAs could be eluted and was found to retain integrity. Binding was pH dependent with a broad optimum at pH 7.7–8.5. Dissociation of bound 125I-labeled ILAs was time and temperature dependent. It was greater at 37 than 4 °C, and was uninfluenced by unlabeled ILAs. The ILAs receptor was stable at 4 °C but was markedly decreased by preincubation at 37 °C. The binding of 125I-labeled ILAs was inhibited by unlabeled ILAs and related insulinlike peptides (the insulinlike growth factors, IGF-1 and IGF-2) in a dose-dependent manner. Insulin and its analogues had only a partially inhibitory effect, and structurally unrelated peptides were without inhibitory efficacy. In contrast ILAs and IGF-1 and IGF-2 inhibited 125I-labeled insulin binding to its receptors in a dose-dependent fashion.These observations identify a receptor for insulinlike peptides in the Golgi elements of rat liver. It is distinct from the insulin receptor previously observed in these elements. The dual interaction of ILAs and other insulinlike peptides with both the insulin receptor and their own unique receptor constitutes the presumed biochemical basis for the two types of action effected by this family of peptides, namely, an effect on metabolism comparable to insulin and an effect on cellular anabolism and growth.

scholarly journals Cell-surface insulin receptor cycling and its implication in the glycogenic response in cultured foetal hepatocytes

1986 ◽  
Vol 239 (3) ◽  
pp. 609-615 ◽  
Author(s):  
P Soubigou ◽  
E Pringault ◽  
C Plas
Keyword(s):  
Cell Surface ◽  
Insulin Receptor ◽  
Polyacrylamide Gel Electrophoresis ◽  
Insulin Receptors ◽  
Rat Hepatocytes ◽  
Insulin Binding ◽  
Surface Proteins ◽  
Short Exposure ◽  
Intermittent Exposure ◽  
Initial Binding

The insulin-receptor cycle was investigated in cultured foetal rat hepatocytes by determining the variations in insulin-binding sites at the cell surface after short exposure to the hormone. Binding of 125I-insulin was measured at 4 degrees C after dissociation of prebound native insulin. Two protocols were used: exchange binding assay and binding after acid treatment; both gave the same results. Cell-surface 125I-insulin-receptor binding decreased sharply (by 40%) during the first 5 min of 10 nM-insulin exposure (t1/2 = 2 min) and remained practically constant thereafter; subsequent removal of the hormone restored the initial binding within 10 min. This fall-rise sequence corresponded to variations in the number of insulin receptors at the cell surface, with no detectable change in receptor affinity. The reversible translocation of insulin receptors from the cell surface to a compartment not accessible to insulin at 4 degrees C was hormone-concentration- and temperature-dependent. SDS/polyacrylamide-gel electrophoresis after cross-linking of bound 125I-insulin to cell-surface proteins with disuccinimidyl suberate showed that these variations were not associated with changes in Mr of binding components, in particular for the major labelled band of Mr 130,000. The insulin-receptor cycle could be repeated after intermittent exposure to insulin. Continuous or intermittent exposure to the hormone gave a similar glycogenic response, contrary to the partial effect of a unique short (5-20 min) exposure. A relationship could be established between the repetitive character of the rapid insulin-receptor cycle and the maximal expression of the biological effect in cultured foetal hepatocytes.

scholarly journals New Insights on NETosis Induced by Entamoeba histolytica: Dependence on ROS from Amoebas and Extracellular MPO Activity

Antioxidants ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 974
Author(s):  
César Díaz-Godínez ◽  
Joshue Fabián Jorge-Rosas ◽  
Mario Néquiz ◽  
Santiago Martínez-Calvillo ◽  
Juan P. Laclette ◽  
...  
Keyword(s):  
Cell Surface ◽  
Nadph Oxidase ◽  
Entamoeba Histolytica ◽  
Pathogen Detection ◽  
Dependent Manner ◽  
Antimicrobial Proteins ◽  
Nadph Oxidase Activity ◽  
Mitochondrial Ros ◽  
Pre Treatment ◽  
Dose Dependent

NETosis is a neutrophil process involving sequential steps from pathogen detection to the release of DNA harboring antimicrobial proteins, including the central generation of NADPH oxidase dependent or independent ROS. Previously, we reported that NETosis triggered by Entamoeba histolytica trophozoites is independent of NADPH oxidase activity in neutrophils, but dependent on the viability of the parasites and no ROS source was identified. Here, we explored the possibility that E. histolytica trophozoites serve as the ROS source for NETosis. NET quantitation was performed using SYTOX® Green assay in the presence of selective inhibitors and scavengers. We observed that respiratory burst in neutrophils was inhibited by trophozoites in a dose dependent manner. Mitochondrial ROS was not also necessary, as the mitochondrial scavenger mitoTEMPO did not affect the process. Surprisingly, ROS-deficient amoebas obtained by pre-treatment with pyrocatechol were less likely to induce NETs. Additionally, we detected the presence of MPO on the cell surface of trophozoites after the interaction with neutrophils and found that luminol and isoluminol, intracellular and extracellular scavengers for MPO derived ROS reduced the amount of NET triggered by amoebas. These data suggest that ROS generated by trophozoites and processed by the extracellular MPO during the contact with neutrophils are required for E. histolytica induced NETosis.

scholarly journals Dectin-1-Mediated Production of Pro-Inflammatory Cytokines Induced by Yeast β-Glucans in Bovine Monocytes

2021 ◽  
Vol 12 ◽  
Author(s):  
Ana R. V. Pedro ◽  
Tânia Lima ◽  
Ricardo Fróis-Martins ◽  
Bárbara Leal ◽  
Isabel C. Ramos ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Surface ◽  
Inflammatory Cytokines ◽  
Surface Expression ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
Feed Supplements ◽  
Surface Receptor ◽  
Dose Dependent ◽  
Pro Inflammatory Cytokines

Yeast-derived products containing β-glucans have long been used as feed supplements in domesticated animals in an attempt to increase immunity. β-glucans are mainly recognized by the cell surface receptor CLEC7A, also designated Dectin-1. Although the immune mechanisms elicited through Dectin-1 activation have been studied in detail in mice and humans, they are poorly understood in other species. Here, we evaluated the response of bovine monocytes to soluble and particulate purified β-glucans, and also to Zymosan. Our results show that particulate, but not soluble β-glucans, can upregulate the surface expression of costimulatory molecules CD80 and CD86 on bovine monocytes. In addition, stimulated cells increased production of IL-8 and of TNF, IL1B, and IL6 mRNA expression, in a dose-dependent manner, which correlated positively with CLEC7A gene expression. Production of IL-8 and TNF expression decreased significantly after CLEC7A knockdown using two different pairs of siRNAs. Overall, we demonstrated here that bovine monocytes respond to particulate β-glucans, through Dectin-1, by increasing the expression of pro-inflammatory cytokines. Our data support further studies in cattle on the induction of trained immunity using dietary β-glucans.

Physiological concentrations of insulin and T3 stimulate 3T3-L1 adipocyte acyl-CoA synthetase gene transcription

1996 ◽  
Vol 270 (5) ◽  
pp. E873-E881 ◽  
Author(s):  
M. S. Kansara ◽  
A. K. Mehra ◽  
J. Von Hagen ◽  
E. Kabotyansky ◽  
P. J. Smith
Keyword(s):  
Gene Transcription ◽  
Fold Increase ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Long Chain Fatty Acids ◽  
Reciprocal Regulation ◽  
Stearoyl Coa Desaturase ◽  
Dose Dependent ◽  
Long Chain ◽  
Stimulation Of

Acyl-CoAsynthetase (ACS) is a key gene for cellular utilization of long-chain fatty acids. We characterized its regulation by physiological concentrations of insulin that acutely regulate metabolism. Our results demonstrate that subnanomolar insulin rapidly and maximally stimulates ACS gene transcription in the absence of protein synthesis; 0.5 nM insulin produced a 2.3 +/- 0.1-fold increase in ACS mRNA levels and induced ACS gene transcription 2.4 +/- 0.3-fold. The insulin sensitivity of ACS was compared with lipoprotein lipase (LPL) and stearoyl-CoA desaturase-1 (SCD-1), which were both less sensitive to insulin. Physiological triiodothyronine (10 nm) also induced ACS mRNA 2.4 +/- 0.1-fold and gene transcription 2.8 +/- 0.3-fold and coordinately induced LPL and SCD-1 mRNA and gene transcription. Because insulin and adenosine 3',5'-cyclic monophosphate often regulate genes involved in lipid and carbohydrate metabolism in a reciprocal manner, we evaluated effects of 1-methyl-3-isobutylxanthine (MIX).ACS mRNA levels were strongly downregulated by MIX in a dose-dependent manner, and ACS gene transcription inhibited in a coordinate manner with LPL and SCD-1. These data demonstrate a uniquely sensitive pattern of stimulation of ACS gene transcription by insulin with reciprocal regulation by MIX, and they suggest a significant role for ACS as a tightly regulated “gatekeeper” gene participating in the control of adipocyte metabolism.

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