Aldose reductase activation is a key component of myocardial response to ischemia.
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Aldose reductase, a member of the aldo-keto reductase family, has been implicated in the development of vascular and neurological complications in diabetes. Despite recent studies from our laboratory demonstrating protection of ischemic hearts by an aldose reductase inhibitor, the presence and influence of aldose reductase in cardiac tissue remain unknown. Our goal in this study was to isolate and characterize the kinetic properties of cardiac aldose reductase, as well as to study the impact of flux via this enzyme on glucose metabolism and contractile function in hearts subjected to ischemia-reperfusion. Results demonstrate that ischemia increases myocardial aldose reductase activity and that these increases are, in part, due to activation by nitric oxide. The kinetic parameter of cardiac aldose reductase (Kcat) was significantly higher in ischemic tissues. Aldose reductase inhibition increased glycolysis and glucose oxidation. Aldose reductase inhibited hearts, when subjected to ischemia/reperfusion, exhibited less ischemic injury and was associated with lower lactate/pyruvate ratios (a measure of cytosolic NADH/NAD+), greater tissue content of adenosine triphosphate, and improved cardiac function. These findings indicate that aldose reductase is a component of ischemic injury and that pharmacological inhibitors of aldose reductase present a novel adjunctive approach for protecting ischemic hearts. (+info)
YUA001, a novel aldose reductase inhibitor isolated from alkalophilic Corynebacterium sp. YUA25. II. Chemical modification and biological activity.
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A series of novel N-substituted tyramine (2-p-hydroxyphenylethylamine) derivatives (1 to approximately 11) were synthesized and evaluated for their inhibitory activity against pig kidney aldose reductase (EC 1, 1, 1, 21). Of these compounds, N-2-p-hydroxyphenylethyl maleamic acid (10) exhibits the strongest aldose reductase inhibitory activity, which is 22 times more potent than that of YUA001. (+info)
Osmoregulatory alterations in taurine uptake by cultured human and bovine lens epithelial cells.
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PURPOSE: Comparative assessment of cultured human lens epithelial cells (HLECs) and bovine lens epithelial cells (BLECs) established the nature of the relationship between taurine-concentrating capability and intracellular polyol accumulation or extracellular hypertonicity. METHODS: The kinetic characteristics of active taurine accumulation based on the measurement of in vitro [3H]-taurine uptake were resolved by side-to-side review of cultured HLECs and BLECs pre-exposed to either galactose-supplemented medium or extracellular hypertonicity. Competitive RT-PCR was used to appraise variation in taurine transporter (TauT) mRNA abundance from cells maintained in hyperosmotic medium over a 72-hour exposure period. RESULTS: The capacity to accumulate [3H]-taurine was significantly lowered after prolonged (20-hour) incubation of cultured BLECs in 40 mM galactose in contrast to HLECs, the latter cells' velocity curve being indistinguishable from control cells in physiological medium. Inhibition of the intracellular taurine transport site appeared to be noncompetitive, in that there was a marked reduction in the V(max) without significant alteration in the K(m) to a high-affinity transport site. Galactitol content in BLECs exceeded five times that found in HLECs. The coadministration of the aldose reductase inhibitor, sorbinil, with 40 mM galactose completely prevented the inhibitory effect of galactose on [3H]-taurine uptake. Acute exposure (3 hours) of HLECs and BLECs to a range of 10 to 40 mM galactitol or 10 to 40 mM galactose plus sorbinil-supplemented medium suggested by Dixon plot that neither galactitol nor galactose interacted with the extracellular taurine transport site. In contrast, [3H]-taurine accumulation was markedly elevated in both HLECs and BLECs after prolonged exposure to galactose-free medium made hyperosmotic by supplementation with sodium chloride. The enhanced taurine uptake capacity involved increase in peak velocity (V(max)) without significant change in Michaelis-Menten constant (K(m)). Cultured HLECs and BLECs responded to hypertonicity with an inducible but transitory upregulation of TauT mRNA. CONCLUSIONS: These results demonstrate that lens epithelial cells express a high-affinity TauT protein capable of active uptake, but predisposed to inhibition by intracellular galactitol when the sugar alcohol is present in sufficiently high concentration to interfere with cell metabolism. Furthermore, lens epithelial cells respond to hypertonic stress by raising taurine transport activity. The increase in taurine uptake is due to an increase in the number of high-affinity TauTs expressed as a result of an increase in the manifestation of taurine mRNA stemming from exposure to hypertonic medium. (+info)
Nitric oxide (NO) plays an important role in the control of numerous vascular functions including basal Na+-K+-ATPase activity in arterial tissue. Hyperglycemia inhibits Na+-K+-ATPase activity in rabbit aorta, in part, through diminished bioactivity of NO. The precise mechanism(s) for such observations, however, are not yet clear. The purpose of this study was to examine the role of superoxide in modulating NO-mediated control of Na+-K+-ATPase in response to hyperglycemia. Rabbit aorta incubated with hyperglycemic glucose concentrations (44 mM) demonstrated a 50% reduction in Na+-K+-ATPase activity that was abrogated by superoxide dismutase. Hyperglycemia also produced a 50% increase in steady-state vascular superoxide measured by lucigenin-enhanced chemiluminescence that was closely associated with reduced Na+-K+-ATPase activity. Specifically, the hyperglycemia-induced increase in vascular superoxide was endothelium dependent, inhibited by L-arginine, and stimulated by N(omega)-nitro-L-arginine. Aldose reductase inhibition with zopolrestat also inhibited the hyperglycemia-induced increase in vascular superoxide. In each manipulation of vascular superoxide, a reciprocal change in Na+-K+-ATPase activity was observed. Finally, a commercially available preparation of Na+-K+-ATPase was inhibited by pyrogallol, a superoxide generator. These data suggest that hyperglycemia induces an increase in endothelial superoxide that inhibits the stimulatory effect of NO on vascular Na+-K+-ATPase activity. (+info)
The crystal structure of rat liver AKR7A1. A dimeric member of the aldo-keto reductase superfamily.
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The structure of the rat liver aflatoxin dialdehyde reductase (AKR7A1) has been solved to 1.38-A resolution. Although it shares a similar alpha/beta-barrel structure with other members of the aldo-keto reductase superfamily, AKR7A1 is the first dimeric member to be crystallized. The crystal structure also reveals details of the ternary complex as one subunit of the dimer contains NADP(+) and the inhibitor citrate. Although the underlying catalytic mechanism appears similar to other aldo-keto reductases, the substrate-binding pocket contains several charged amino acids (Arg-231 and Arg-327) that distinguish it from previously characterized aldo-keto reductases with respect to size and charge. These differences account for the substrate specificity for 4-carbon acid-aldehydes such as succinic semialdehyde and 2-carboxybenzaldehyde as well as for the idiosyncratic substrate aflatoxin B(1) dialdehyde of this subfamily of enzymes. Structural differences between the AKR7A1 ternary complex and apoenzyme reveal a significant hinged movement of the enzyme involving not only the loops of the structure but also parts of the alpha/beta-barrel most intimately involved in cofactor binding. (+info)
Time- and dose-dependent differential upregulation of three genes by 17 beta-estradiol in endothelial cells.
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The purpose of this study was to identify genetic targets in the vasculature for estrogen by profiling genes expressed in female human aortic endothelial cells exposed to various doses of 17 beta-estradiol at differing concentrations and for differing periods of time. Our approach employed a RT-PCR-based cloning strategy of DNA differential display analysis, with differential expression verified by semiquantitative PCR performed with gene-specific primers. A significant increase in mRNA expression in response to 17 beta-estradiol was observed for the following three genes: aldose reductase (3.4-fold), caspase homologue-alpha protein (4.2-fold), and plasminogen activator inhibitor-1 intron e (2.3-fold). For all three upregulated genes, estradiol-induced upregulation occurred with a similar time course and temporally clustered to the first 24 h after hormone treatment. In addition, the effect of estradiol dose on gene expression was consistent and occurred at physiological concentrations. Our results describe previously uncharacterized estradiol-sensitive time- and dose-dependent regulation of genes with potential importance to vascular function in human endothelial cells. (+info)
Gene expression of enzymes comprising the polyol pathway in various rat tissues determined by the competitive RT-PCR method.
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The quantitative measurements of aldose reductase (AR) and sorbitol dehydrogenase (SDH) gene expression in various rat tissues were performed by the competitive reverse transcription-polymerase chain reaction (RT-PCR). AR mRNA was detectable in all tissues analyzed with pronounced differences in the amounts. SDH mRNA was most abundant in testes and liver, but was absent from lens. The estimation of the AR cDNA/SDH cDNA ratio showed that the relative abundance of SDH to AR differs among tissues. These results indicate that different tissues contain varying amounts of AR mRNA and SDH mRNA; that is, each tissue has its own polyol pathway activity. (+info)
Rottlerin inhibits tonicity-dependent expression and action of TonEBP in a PKCdelta-independent fashion.
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Novel protein kinase C (PKC) isoforms PKCdelta and PKCepsilon have recently been implicated in signaling by hypertonic stress. We investigated the role of the putative PKCdelta inhibitor rottlerin on tonicity-dependent gene regulation. In the renal medullary mIMCD3 cell line, rottlerin blocked tonicity-dependent transcription of a tonicity enhancer (TonE)-driven luciferase reporter gene, as well as tonicity-dependent transcription of the physiological tonicity effector gene aldose reductase, but not urea-dependent transcription. Consistent with these data, rottlerin inhibited tonicity-dependent expression of TonE binding protein (TonEBP) at the mRNA and protein levels. Another inhibitor of both novel and conventional PKC isoforms, GF-109203X, suppressed TonEBP-dependent transcription but failed to influence tonicity-inducible TonEBP expression. Global PKC downregulation with protracted phorbol ester treatment, however, failed to influence tonicity-dependent signaling, arguing against a PKCdelta-dependent mechanism of rottlerin action in this model. In addition, hypertonic stress failed to induce phosphorylation of PKCdelta. Furthermore, in a PC-12 cell model with a comparable degree of tonicity-dependent transcription, constitutive overexpression of dominant negative-acting PKCdelta or PKCepsilon effectively decreased tonicity signaling to extracellular signal-regulated kinase activation, as expected, but failed to influence TonE-dependent transcription. TonE-dependent transcription, however, remained rottlerin sensitive in this PC-12 cell model. In the aggregate, these data indicate that rottlerin dramatically inhibits tonicity-dependent TonEBP expression and TonE-dependent transcription but, despite its reputed mode of action, does so through a PKCdelta-independent pathway. (+info)