Insulin regulation of glutathione and contractile phenotype in diabetic rat ventricular myocytes

2007 ◽  
Vol 292 (3) ◽  
pp. H1619-H1629 ◽  
Author(s):  
Shumin Li ◽  
Xun Li ◽  
Yu-Long Li ◽  
Chun-Hong Shao ◽  
Keshore R. Bidasee ◽  
...  
Keyword(s):  
De Novo ◽  
Ventricular Myocytes ◽  
Cardiovascular Complications ◽  
Diabetic Rat ◽  
Sprague Dawley ◽  
Cell Functions ◽  
Glutamylcysteine Synthetase ◽  
Rat Myocytes ◽  
Glucose 6 Phosphate Dehydrogenase

Cardiovascular complications of diabetes mellitus involve oxidative stress and profound changes in reduced glutathione (GSH), an essential tripeptide that controls many redox-sensitive cell functions. This study examined regulation of GSH by insulin to identify mechanisms controlling cardiac redox state and to define the functional impact of GSH depletion. GSH was measured by fluorescence microscopy in ventricular myocytes isolated from Sprague-Dawley rats made diabetic by streptozotocin, and video and confocal microscopy were used to measure mechanical properties and Ca2+ transients, respectively. Spectrophotometric assays of tissue extracts were also done to measure the activities of enzymes that control GSH levels. Four weeks after injection of streptozotocin, mean GSH concentration ([GSH]) in isolated diabetic rat myocytes was ∼36% less than in control, correlating with decreased activities of two major enzymes regulating GSH levels: glutathione reductase and γ-glutamylcysteine synthetase. Treatment of diabetic rat myocytes with insulin normalized [GSH] after a delay of 3–4 h. A more rapid but transient upregulation of [GSH] occurred in myocytes treated with dichloroacetate, an activator of pyruvate dehydrogenase. Inhibitor experiments indicated that insulin normalized [GSH] via the pentose pathway and γ-glutamylcysteine synthetase, although the basal activity of glucose-6-phosphate dehydrogenase was not different between diabetic and control hearts. Diabetic rat myocytes were characterized by significant mechanical dysfunction that correlated with diminished and prolonged Ca2+ transients. This phenotype was reversed by in vitro treatment with insulin and also by exogenous GSH or N-acetylcysteine, a precursor of GSH. Our data suggest that insulin regulates GSH through pathways involving de novo GSH synthesis and reduction of its oxidized form. It is proposed that a key function of glucose metabolism in heart is to supply reducing equivalents required to maintain adequate GSH levels for the redox control of Ca2+ handling proteins and contraction.

Redox regulation of Ito remodeling in diabetic rat heart

2005 ◽  
Vol 288 (3) ◽  
pp. H1417-H1424 ◽  
Author(s):  
Xun Li ◽  
Zhi Xu ◽  
Shumin Li ◽  
George J. Rozanski
Keyword(s):  
Ion Channels ◽  
Rat Heart ◽  
Redox Regulation ◽  
Control Level ◽  
Diabetic Rat ◽  
Cell Functions ◽  
Control Heart ◽  
Rat Hearts ◽  
Glucose 6 Phosphate Dehydrogenase

Oxidative stress and the resulting change in cell redox state are proposed to contribute to pathogenic alterations in ion channels that underlie electrical remodeling of the diseased heart. The present study examined whether K+ channel remodeling is controlled by endogenous oxidoreductase systems that regulate redox-sensitive cell functions. Diabetes was induced in rats by streptozotocin, and experiments were conducted after 3–5 wk of hyperglycemia. Spectrophotometric assays of ventricular tissue extracts from diabetic rat hearts revealed divergent changes in two major oxidoreductase systems. The thioredoxin (TRX) system in diabetic rat heart was characterized by a 52% decrease in TRX reductase (TRXR) activity from control heart ( P < 0.05), whereas TRX activity was 1.7-fold greater than control heart ( P < 0.05). Diabetes elicited similar changes in the glutaredoxin (GRX) system: glutathione reductase was decreased 35% from control level ( P < 0.05), and GRX activity was 2.5-fold greater than in control heart ( P < 0.05). The basal activity of glucose-6-phosphate dehydrogenase, which generates NADPH required by the TRX and GRX systems, was not altered by diabetes. Voltage-clamp studies showed that the characteristically decreased density of the transient outward K+ current ( Ito) in isolated diabetic rat myocytes was normalized by in vitro treatment with insulin (0.1 μM) or the metabolic activator dichloroacetate (1.5 mM). The effect of these agonists on Ito was blocked by inhibitors of glucose-6-phosphate dehydrogenase. Moreover, inhibitors of TRXR, which controls the reducing activity of TRX, also blocked upregulation of Ito by insulin and dichloroacetate. These data suggest that K+ channels underlying Ito are regulated in a redox-sensitive manner by the TRX system and the remodeling of Ito that occurs in diabetes may be due to decreased TRXR activity. We propose that oxidoreductase systems are an important repair mechanism that protects ion channels and associated regulatory proteins from irreversible oxidative damage.

Abstract 14044: Engineering Bioactive Nanoparticles to Rejuvenate Vascular Progenitor Cells

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Loan T Bui ◽  
Shanique Edwards ◽  
Laura Alderfer ◽  
Laura Haneline ◽  
Donny Hanjaya-putra
Keyword(s):  
Cell Migration ◽  
Progenitor Cells ◽  
De Novo ◽  
Cardiovascular Complications ◽  
Release Kinetic ◽  
Future Health ◽  
Immunodeficient Mice ◽  
Vascular Progenitor Cells

Introduction: Fetal exposure to gestational diabetes mellitus (GDM) predisposes children to future health complications including hypertension and cardiovascular disease. A key mechanism by which these complications occur is through stress-induced dysfunction of vascular progenitor cells, including endothelial colony-forming cells (ECFCs). In particular, overexpression of transgelin (TAGLN), also known as SM22α, in GDM-ECFCs is associated with actin cytoskeletal rearrangement, which results in reduced cell migration and impaired vasculogenesis. We hypothesized that bioactive nanoparticles (NPs) conjugated on the surface of GDM-ECFCs can provide a sustained pseudo-autocrine stimulation to improve in vitro and in vivo vasculogenesis. Methods & Results: We designed multilamellar lipid NPs with an average size of 147±63 nm in diameter to deliver small molecules SB-431542 (TGF-β inhibitor) directly to the surface of GDM-ECFCs. Bioactive NPs can be robustly conjugated to the surface of ECFCs using thiol-maleimide coupling without altering cell viability and key progenitor phenotypes. By controlling the release kinetic of TGF-β inhibitor from the NPs, we can normalize TAGLN expression and improve cell migration, a critical key step in establishing functional vascular networks. Moreover, bioactive NPs can restore the vasculogenic potential of GDM-ECFCs in both 2D Matrigel and 3D collagen assays. Finally, when transplanted into immunodeficient mice, GDM-ECFCs conjugated with bioactive NPs exhibit robust de novo blood vessel formation with high engraftment rate, comparable to normal ECFCs. Conclusions: Collectively, these findings highlight a simple, yet promising strategy to rejuvenate GDM-ECFCs and improve their therapeutic potentials, which can be clinically-translated to address various cardiovascular complications, as well as toward a range of approaches in tissue repair and regenerative medicine.

Glutathione and K+ channel remodeling in postinfarction rat heart

2002 ◽  
Vol 282 (6) ◽  
pp. H2346-H2355 ◽  
Author(s):  
George J. Rozanski ◽  
Zhi Xu
Keyword(s):  
Oxidative Stress ◽  
Glutathione Reductase ◽  
Ventricular Myocytes ◽  
K Channel ◽  
Glycolytic Flux ◽  
Cell Functions ◽  
Tissue Extracts ◽  
Rat Hearts ◽  
Glutamylcysteine Synthetase ◽  
Pentose Pathway

Electrical remodeling of the diseased ventricle is characterized by downregulation of K+ channels that control action potential repolarization. Recent studies suggest that this shift in electrophysiological phenotype involves oxidative stress and changes in intracellular glutathione (GSH), a key regulator of redox-sensitive cell functions. This study examined the role of GSH in regulating K+ currents in ventricular myocytes from rat hearts 8 wk after myocardial infarction (MI). Colorimetric analysis of tissue extracts showed that endogenous GSH levels were significantly less in post-MI hearts compared with controls, which is indicative of oxidative stress. This change in GSH status correlated with significant decreases in activities of glutathione reductase and γ-glutamylcysteine synthetase. Voltage-clamp studies of isolated myocytes from post-MI hearts demonstrated that downregulation of the transient outward K+ current ( I to) could be reversed by pretreatment with exogenous GSH or N-acetylcysteine, a precursor of GSH. Upregulation of I to was also elicited by dichloroacetate, which increases glycolytic flux through the GSH-related pentose pathway. This metabolic effect was blocked by inhibitors of glutathione reductase and the pentose pathway. These data indicate that oxidative stress-induced alteration in the GSH redox state plays an important role in I to channel remodeling and that GSH homeostasis is influenced by pathways of glucose metabolism.

TRI-IODOTHYRONINE INCREASES INTRA-CELLULAR CALCIUM LEVELS IN SINGLE RAT MYOCYTES

1991 ◽  
Vol 7 (1) ◽  
pp. 77-79 ◽  
Author(s):  
R.B. Lomax ◽  
P.H. Cobbold ◽  
A.P. Allshire ◽  
K.S.R. Cuthbertson ◽  
W.R. Robertson
Keyword(s):  
Ventricular Myocytes ◽  
Single Cells ◽  
Mean Amplitude ◽  
Free Calcium ◽  
Acute Administration ◽  
Cytosolic Free Calcium ◽  
Rat Hearts ◽  
Rat Myocytes ◽  
Stimulation Of

ABSTRACT We have studied the effects of acute administration of tri-iodothyronine (T3) on cytosolic free calcium levels [Ca2+]i in single rat myocytes microinjected with aequorin. Ventricular myocytes were isolated by perfusing rat hearts with collagenase, and healthy, rod-shaped cells were injected to <1% of their volume with aequorin. The photons emitted from single cells were measured and a conversion to [Ca2+]i made on the basis of an in vitro calibration after the remaining aequorin had been discharged by cell lysis. Only cells that depolarized reversibly (showing elevated [Ca2+]i levels) when superfused with 80mM KC1, and which gave a substantial signal on lysis with distilled water were used. The [Ca2+]i rose from a resting value of 150±56nM (mean ± SD, n=14) by 127±47nM on depolarization with 80mM KC1. Application of T3 (1-100nM) led to an increase (P<0.05) in [Ca2+]i (mean amplitude of 152±35nM) before returning to baseline. The median duration of these events was 10 min (range = 1.4-34.4 min). The time to response was shorter when lOOnM T3 was applied (median and range; 6.8, 0-14 min) than when 1nM T3 was used (16, 7.0-56.1 min) (P<0.05). To conclude, physiological concentrations of thyroid hormones caused rapid but transient stimulation of [Ca2+]i in single rat myocytes.

scholarly journals Anti-VEGF therapy prevents Müller intracellular edema by decreasing VEGF-A in diabetic retinopathy

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Tianqin Wang ◽  
Chaoyang Zhang ◽  
Hai Xie ◽  
Mengmeng Jiang ◽  
Haibin Tian ◽  
...  
Keyword(s):  
Intracellular Sodium ◽  
Diabetic Rat ◽  
Vascular Endothelial ◽  
Sprague Dawley ◽  
Sodium Potassium ◽  
Intracellular Potassium ◽  
Sprague Dawley Rats ◽  
Endothelial Growth Factor ◽  
Inwardly Rectifying

Abstract Background Although vascular endothelial growth factor A (VEGF-A) is known to play a key role in causing retinal edema, whether and how VEGF-A induces intracellular edema in the retina still remains unclear. Methods Sprague-Dawley rats were rendered diabetic with intraperitoneal injection of streptozotocin. Intravitreal injection of ranibizumab was performed 8 weeks after diabetes onset. rMC-1 cells (rat Müller cell line) were treated with glyoxal for 24 h with or without ranibizumab. The expression levels of inwardly rectifying K+ channel 4.1 (Kir4.1), aquaporin 4 (AQP4), Dystrophin 71 (Dp71), VEGF-A, glutamine synthetase (GS) and sodium-potassium-ATPase (Na+-K+-ATPase) were examined using Western blot. VEGF-A in the supernatant of the cell culture was detected with ELISA. The intracellular potassium and sodium levels were detected with specific indicators. Results Compared with normal control, protein expressions of Kir4.1 and AQP4 were down-regulated significantly in diabetic rat retinas, which were prevented by ranibizumab. The above changes were recapitulated in vitro. Similarly, the intracellular potassium level in glyoxal-treated rMC-1 cells was increased, while the intracellular sodium level and Na+-K+-ATPase protein level remained unchanged, compared with control. However, ranibizumab treatment decreased intracellular sodium, but not potassium. Conclusion Ranibizumab protected Müller cells from diabetic intracellular edema through the up-regulation of Kir4.1 and AQP4 by directly binding VEGF-A. It also caused a reduction in intracellular osmotic pressure.

Modulatory effect of leptin on nitric oxide production and lipid metabolism in term placental tissues from control and streptozotocin-induced diabetic rats

2004 ◽  
Vol 16 (3) ◽  
pp. 363 ◽  
Author(s):  
Verónica White ◽  
Elida González ◽  
Evangelina Capobianco ◽  
Carolina Pustovrh ◽  
Carlos Soñez ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Lipid Metabolism ◽  
De Novo ◽  
Lipid Synthesis ◽  
Placental Tissue ◽  
Diabetic Rats ◽  
Late Gestation ◽  
Diabetic Rat ◽  
No Production

Leptin production by placental tissues contributes to its circulating levels and functions. The diabetic pathology induces alterations in leptin levels. In the present study, leptin levels were evaluated in placental tissue from control and neonatal streptozotocin-induced (n-STZ) diabetic rats during late gestation. The effects of leptin levels on the generation of nitric oxide (NO), prostaglandin (PG) E2 production and lipid metabolism were examined. Leptin levels were diminished in placentas from n-STZ diabetic rats compared with controls (P < 0.01). These differences were also evident when leptin was evaluated immunohistochemically. Addition of leptin (1 nm) in vitro enhanced NO production in control (66%) and diabetic placentas (134%) by stimulating NO synthase activity (by 38% and 54%, respectively). The addition of leptin increased PGE2 production in placentas from control (173%) and diabetic rats (83%) and produced a 50% decrease in placental lipid levels (phospholipids, triacylglycerides, cholesterol and cholesteryl ester) without involving a reduction in de novo lipid synthesis. These data indicate that leptin enhances the production of placental NO and PGE2, vasoactive agents that modify placental blood flow, and that leptin stimulates placental lipid metabolism, probably generating more lipids for transfer to the fetus. In the diabetic rat, placental leptin was reduced, probably as a response to the maternal environment to locally regulate the transfer of nutrients to the developing fetus.

Metabolic basis of decreased transient outward K+ current in ventricular myocytes from diabetic rats

1996 ◽  
Vol 271 (5) ◽  
pp. H2190-H2196 ◽  
Author(s):  
Z. Xu ◽  
K. P. Patel ◽  
G. J. Rozanski
Keyword(s):  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Diabetic Rats ◽  
K Channel ◽  
Diabetic Rat ◽  
Channel Function ◽  
Early Stages ◽  
Metabolic Basis ◽  
External Glucose

The purpose of this study was to examine the mechanisms of alterations in cardiac K+ channel function in early stages of experimental diabetes mellitus induced by streptozotocin. Transient outward (Ito) and inward rectifier (IK1) K+ currents were recorded by the whole cell voltage-clamp technique in ventricular myocytes isolated from hearts of 2- to 4-wk diabetic and age-matched control rats. Ito density in myocytes from diabetic rats was approximately 30% less than control (at +60 mV; P < 0.01) under basal recording conditions in the presence of 18 mM external glucose, whereas IK1 density was not different between groups. When external glucose concentration was decreased to 5 mM for 4-6 h, basal Ito density was not changed in either group of myocytes. To further examine the possible metabolic basis of reduced Ito density in myocytes from diabetic rats, we separately tested three structurally different compounds that affect substrate utilization in cardiac myocytes: insulin (0.1 microM), dichloroacetate (1.5 mM), and L-carnitine (10 mM). Each compound completely normalized Ito density in myocytes from diabetic rats treated in vitro for 4-6 h. The same agents had no effect on Ito density in control myocytes, nor was IK1 altered in either group of cells. These data provide the first evidence to support the hypothesis that there is a metabolic basis for decreased Ito density in diabetic rat ventricular myocytes in early stages of this model. Furthermore, our data suggest that depressed glucose metabolism in the diabetic heart may be a key factor underlying changes in Ito channel function, because agents that increase glucose utilization normalize Ito density within a short time period.

scholarly journals CHD8 Regulates Cellular Homeostasis and Neuronal Function Genes Across Multiple Models of CHD8 Haploinsufficiency

2018 ◽  
Author(s):  
A. Ayanna Wade ◽  
Kenneth Lim ◽  
Rinaldo Catta-Preta ◽  
Alex S. Nord
Keyword(s):  
Gene Expression ◽  
Gene Regulation ◽  
Chromatin Remodeling ◽  
De Novo ◽  
Autism Spectrum ◽  
Loss Of Function ◽  
Cell Functions ◽  
High Affinity

ABSTRACTThe packaging of DNA into chromatin determines the transcriptional potential of cells and is central to eukaryotic gene regulation. Recent sequencing of patient mutations has linked de novo loss-of-function mutations to chromatin remodeling factors with specific, causal roles in neurodevelopmental disorders. Characterizing cellular and molecular phenotypes arising from haploinsufficiency of chromatin remodeling factors could reveal convergent mechanisms of pathology. Chromodomain helicase DNA binding protein 8 (CHD8) encodes a chromatin remodeling factor gene and has among the highest de novo loss-of-function mutations rates in patients with autism spectrum disorder (ASD). Mutations to CHD8 are expected to drive neurodevelopmental pathology through global disruptions to gene expression and chromatin state, however, mechanisms associated with CHD8 function have yet to be fully elucidated. We analyzed published transcriptomic and epigenomic data across CHD8 in vitro and in vivo knockdown and knockout models to identify convergent mechanisms of gene regulation by CHD8. We found reproducible high-affinity interactions of CHD8 near promoters of genes necessary for basic cell functions and gene regulation, especially chromatin organization and RNA processing genes. Overlap between CHD8 interaction and differential expression suggests that reduced dosage of CHD8 directly relates to decreased expression of these genes. In addition, genes important for neuronal development and function showed consistent dysregulation, though there was a reduced rate and decreased affinity for CHD8 interactions near these genes. This meta-analysis verifies CHD8 as a critical regulator of gene expression and reveals a consistent set of high affinity CHD8 interaction targets observed across human and mouse in vivo and in vitro studies. Our findings highlight novel core functions of CHD8 and indicate direct and downstream gene regulatory impacts that are likely to be associated with neuropathology underlying CHD8-associated neurodevelopmental disorder.

scholarly journals Glucose 6-Phosphate Dehydrogenase from Trypanosomes: Selectivity for Steroids and Chemical Validation in Bloodstream Trypanosoma brucei

Molecules ◽  
2021 ◽  
Vol 26 (2) ◽  
pp. 358
Author(s):  
Cecilia Ortíz ◽  
Francesca Moraca ◽  
Marc Laverriere ◽  
Allan Jordan ◽  
Niall Hamilton ◽  
...  
Keyword(s):  
De Novo ◽  
Pentose Phosphate ◽  
Small Subset ◽  
Cell Physiology ◽  
Single Digit ◽  
In Vitro Proliferation ◽  
African Trypanosomes ◽  
Thiol Redox ◽  
Glucose 6 Phosphate Dehydrogenase

Glucose 6-phosphate dehydrogenase (G6PDH) fulfills an essential role in cell physiology by catalyzing the production of NADPH+ and of a precursor for the de novo synthesis of ribose 5-phosphate. In trypanosomatids, G6PDH is essential for in vitro proliferation, antioxidant defense and, thereby, drug resistance mechanisms. So far, 16α-brominated epiandrosterone represents the most potent hit targeting trypanosomal G6PDH. Here, we extended the investigations on this important drug target and its inhibition by using a small subset of androstane derivatives. In Trypanosoma cruzi, immunofluorescence revealed a cytoplasmic distribution of G6PDH and the absence of signal in major organelles. Cytochemical assays confirmed parasitic G6PDH as the molecular target of epiandrosterone. Structure-activity analysis for a set of new (dehydro)epiandrosterone derivatives revealed that bromination at position 16α of the cyclopentane moiety yielded more potent T. cruzi G6PDH inhibitors than the corresponding β-substituted analogues. For the 16α brominated compounds, the inclusion of an acetoxy group at position 3 either proved detrimental or enhanced the activity of the epiandrosterone or the dehydroepiandrosterone derivatives, respectively. Most derivatives presented single digit μM EC50 against infective T. brucei and the killing mechanism involved an early thiol-redox unbalance. This data suggests that infective African trypanosomes lack efficient NADPH+-synthesizing pathways, beyond the Pentose Phosphate, to maintain thiol-redox homeostasis.

Potassium currents in ventricular myocytes from genetically diabetic rats

1997 ◽  
Vol 273 (4) ◽  
pp. E695-E700 ◽  
Author(s):  
Katsuharu Tsuchida ◽  
Hiroshi Watajima
Keyword(s):  
Action Potential ◽  
Matched Control ◽  
Ventricular Myocytes ◽  
Outward Current ◽  
Potassium Currents ◽  
Diabetic Rat ◽  
Transient Outward Current ◽  
Duration Of Action ◽  
Insulin Dependent ◽  
Rat Myocytes

Our previous study demonstrated the longer duration of action potential in ventricular myocytes from genetically diabetic WBN/Kob rats without change in calcium channel density compared with age-matched controls [Tsuchida, K., H. Watajima, and S. Otamo. Am. J. Physiol. 267 ( Heart Circ. Physiol. 36): H2280–H2289, 1994]. In the present study we examined the alteration of potassium currents, especially transient outward current, in ventricular myocytes of genetically diabetic WBN/Kob rats. WBN/Kob rats gradually develop hyperglycemia with aging and show some similarity to non-insulin-dependent diabetes mellitus models, which differ from the insulin-dependent streptozotocin-treated rat model. The density of the intracellular calcium ion-independent transient outward current ( I to) from 17- to 19-mo diabetic rat myocytes was significantly smaller than that from age-matched control rat myocytes. In addition, the density of I to from 17- to 19-mo rat myocytes was significantly less than that from 2-mo rat myocytes, suggesting that aging-induced alteration of I to was accelerated by the diabetic state. The steady-state inactivation curves of I to, the recovery from I toinactivation, and the other outward currents were not significantly altered between diabetic myocytes and age-matched control myocytes. In conclusion, the prolonged duration of action potential from genetically diabetic rat myocytes is mainly due to the depressed I to.

Sign in / Sign up

Export Citation Format

Share Document