Cell Sorbitol Research Articles

Bioequivalence Study of Epalrestat for Healthy Chinese Subjects.

Epalrestat is a reversible noncompetitive inhibitor of aldose reductase with selective inhibition of aldose reductase. It can inhibit the accumulation of sorbitol in red blood cells in patients with diabetic peripheral neuropathy and can improve patients' conscious symptoms and neurological dysfunction. This study was designed to evaluate the bioequivalence in healthy Chinese subjects of a new test formulation and reference formulation of oral epalrestat (50mg) in the fasting state. The study was performed with 44 healthy Chinese subjects according to a randomized 2-way crossover design. The main pharmacokinetic parameters of test formulation and reference formulation as follows: 4793 and 4781 ng/mL for maximum plasma concentration, 8556 and 8431 ngh/mL for area under the plasma concentration-time curve extrapolated to infinity. The test formulation of epalrestat was bioequivalent to the reference formulation. The bioequivalence study of epalrestat in healthy Chinese subjects suggests that the test and reference formulations have similar pharmacokinetics and both formulations are well tolerated in the dose range studied in healthy Chinese subjects. All these findings provided valuable pharmacokinetic knowledge for further clinical development.

Predictive ADMET study of rhodanine-3-acetic acid chalcone derivatives

Considered one of the chronic diseases in a global state of emergency, diabetes mellitus (DM) may cover an area of 629 million people affected by 2045. The main route of hyperglycemia in the human body is associated with the action of the enzyme aldose reductase on glucose molecules, resulting in the accumulation of sorbitol in blood and tissue cells. Thus, the present study is a high-throughput virtual screening (HTS) of the various molecular datasets available online, as well as an investigation of the pharmacokinetic properties of absorption, distribution, metabolism, excretion, and toxicity (ADMET) and enzyme inhibition aldose reductase from a series of derivatives of rhodanine-3-acetic acid chalcones (D1-8). Four derivatives (D1-4) showed greater oral bioavailability, as they are anions of low relative polarity, showing better viability to interact with the aldose reductase target with a low toxic response to the host.

Accumulation of sorbitol in the sciatic nerve modulates circadian properties of diabetes-induced neuropathic pain hypersensitivity in a diabetic mouse model

The intensity of pain in diabetic neuropathy varies in a circadian time-dependent manner. It is well known that diabetes has two distinct types, which are differentiated based on the cause of the disease. Previous studies have yet to compare the circadian properties of the pain intensity of diabetic neuropathy between type I and type II diabetes. In this study, we demonstrated that the pain intensity of diabetic peripheral neuropathy in a db/db mouse model of type II diabetes showed a significant diurnal oscillation, but such time-dependent oscillation was not detected in a streptozotocin (STZ)-induced type I diabetic mouse model. The polyol pathway-induced accumulation of sorbitol in peripheral nerve cells suppresses Na+/K+-ATPase activity, which is associated with the intensity of pain in diabetic neuropathy. In db/db mice, this accumulation of sorbitol in peripheral nerve cells showed significant diurnal oscillation. In addition, pain intensity and Na+/K+-ATPase activity were decreased at the peak time of sorbitol accumulation in these mice. Although STZ-induced diabetic mice also showed sorbitol accumulation and Na+/K+-ATPase dysfunction, these measures did not oscillate in a time-dependent manner. These findings reveal differences in the circadian properties of pain hypersensitivity in mouse models of type I and type II diabetes, and also provide ideas for developing novel approaches to the management of diabetic neuropathy.

An optimized protocol for the preparation of oxygen-evolving thylakoid membranes from Cyclotella meneghiniana provides a tool for the investigation of diatom plastidic electron transport

BackgroundThe preparation of functional thylakoid membranes from diatoms with a silica cell wall is still a largely unsolved challenge. Therefore, an optimized protocol for the isolation of oxygen evolving thylakoid membranes of the centric diatom Cyclotella meneghiniana has been developed. The buffer used for the disruption of the cells was supplemented with polyethylene glycol based on its stabilizing effect on plastidic membranes. Disruption of the silica cell walls was performed in a French Pressure cell and subsequent linear sorbitol density gradient centrifugation was used to isolate the thylakoid membrane fraction.ResultsSpectroscopic characterization of the thylakoids by absorption and 77 K fluorescence spectroscopy showed that the photosynthetic pigment protein complexes in the isolated thylakoid membranes were intact. This was supported by oxygen evolution measurements which demonstrated high electron transport rates in the presence of the artificial electron acceptor DCQB. High photosynthetic activity of photosystem II was corroborated by the results of fast fluorescence induction measurements. In addition to PSII and linear electron transport, indications for a chlororespiratory electron transport were observed in the isolated thylakoid membranes. Photosynthetic electron transport also resulted in the establishment of a proton gradient as evidenced by the quenching of 9-amino-acridine fluorescence. Because of their ability to build-up a light-driven proton gradient, de-epoxidation of diadinoxanthin to diatoxanthin and diatoxanthin-dependent non-photochemical quenching of chlorophyll fluorescence could be observed for the first time in isolated thylakoid membranes of diatoms. However, the ∆pH, diadinoxanthin de-epoxidation and diatoxanthin-dependent NPQ were weak compared to intact diatom cells or isolated thylakoids of higher plants.ConclusionsThe present protocol resulted in thylakoids with a high electron transport capacity. These thylakoids can thus be used for experiments addressing various aspects of the photosynthetic electron transport by, e.g., employing artificial electron donors and acceptors which do not penetrate the diatom cell wall. In addition, the present isolation protocol yields diatom thylakoids with the potential for xanthophyll cycle and non-photochemical quenching measurements. However, the preparation has to be further refined before these important topics can be addressed systematically.

Inhibition of Aldose Reductase and Red Blood Cell Sorbitol Accumulation by Extract of Capparis spinosa

Background: Activation of polyol pathway, due to increased aldose reductase (ALR2) activity, is one of the several mechanisms that have been involved in the development of secondary complications of diabetes. Therefore, ALR2 inhibition has been recognized as a potential therapeutic goal in the prevention and treatment of various secondary complications of diabetes, such as cataract. Objectives: In this study, we aimed to evaluate the inhibitory effects of hydro alcoholic Capparis spinosa extract (CSE) on ALR2 and sorbitol accumulation in red blood cells (RBCs). Materials and Methods: The hydroalcoholic extract of Capparis spinosa (C. spinosa) was prepared and its inhibitory effects were evaluated by using the partial purified bovine lens ALR2 and its specificity was checked by comparing with its inhibitory effect on aldehyde reductase (ALR1), the other member of aldo-keto reductases family. Because RBC sorbitol accumulation is strongly correlated with the blood glucose level, the effect of CSE on sorbitol accumulation in human RBCs, preincubated with high serum glucose concentration, was determined. Results: The CSE inhibited ALR2 activity, with an IC 50 of 0.54 mg/mL and it was seven and a half times greater than that of ALR1 inhibition, indicating the specificity of CSE for ALR2, rather than ALR1. Also, CSE, at a concentration of 0.54 mg/mL, could reduce sorbitol accumulation in erythrocytes, by 46.8%. Conclusions: These results demonstrated that CSE, mainly through inhibition of ALR2, could prevent or delay the diabetic complications, like cataract formation, as a result of sorbitol accumulation.

A Clinical and Neuropathological Study of Chinese Patients with Diabetic Peripheral Neuropathy

ObjectiveTo examine whether the neuropathological and metabolic changes of peripheral nerves are correlated to clinical features in diabetes mellitus type 2 patients with peripheral neuropathy.Methods147 type 2 diabetic patients with signs/symptoms of diabetic peripheralneuropathy (DPN) aged 53.4±12.3 years and 134 healthy volunteers aged 55.5±11.7 years were investigated for fasting plasma glucose (FPG), hemoglobin A1C (HbA1c), and red blood cell sorbitol (RBC sorbitol) in addition to nerve conduction velocity (NCV). Among the 147 diabetic patients, 10 patients underwent superficial peroneal nerve biopsy for light and electron microscopy.ResultsIn the experimental group, the levels of HbA1c and RBC sorbitol showed significant increase compared with the controlled group, whereas motor nerve conduction velocity (MNCV) and sensory nerve conduction velocity (SNCV) both showed decline and SNCV decreased at a greater extent. Morphologically, there were various degrees of nerve fiber loss, associated with axon degeneration and capillary luminal narrowing in 10 patients undergone nerve biopsy.ConclusionThe metabolic change of sorbitol, the consequently observed changes in NCV and histopathology of peripheral nerves are positively correlated with the duration of diabetes and overall level of blood glucose.

The ATAC Acetyltransferase Complex Coordinates MAP Kinases to Regulate JNK Target Genes

In response to extracellular cues, signal transduction activates downstream transcription factors like c-Jun to induce expression of target genes. We demonstrate that the ATAC (Ada two A containing) histone acetyltransferase (HAT) complex serves as a transcriptional cofactor for c-Jun at the Jun N-terminal kinase (JNK) target genes Jra and chickadee. ATAC subunits are required for c-Jun occupancy of these genes and for H4K16 acetylation at the Jra enhancer, promoter, and transcribed sequences. Under conditions of osmotic stress, ATAC colocalizes with c-Jun, recruits the upstream kinases Misshapen, MKK4, and JNK, and suppresses further activation of JNK. Relocalization of these MAPKs and suppression of JNK activation by ATAC are dependent on the CG10238 subunit of ATAC. Thus, ATAC governs the transcriptional response to MAP kinase signaling by serving as both a coactivator of transcription and as a suppressor of upstream signaling.

ZAC1 Is Up-regulated by Hypertonicity and Decreases Sorbitol Dehydrogenase Expression, Allowing Accumulation of Sorbitol in Kidney Cells

Affymetrix GeneChip technology was employed to detect differentially expressed genes in inner medullary collecting duct (IMCD3) cells grown under isotonic and hypertonic conditions. A marked up-regulation was found for the zinc-finger protein ZAC1 under hypertonic stress (219-fold, p < 0.001). Changes in expression for ZAC1 were verified by quantitative PCR for message and Western blotting for protein. In mouse and human kidney tissues, ZAC1 expression was substantial in the papilla and was absent in the cortex. Furthermore, ZAC1 expression significantly increased in the papilla of mice following 36 h of fluid restriction and decreased in polyuric mice consuming sucrose in water. Because ZAC1 has been described to be a potential negative regulator of sorbitol dehydrogenase (SDH) in hippocampal cells, we examined whether this relationship also occurs in kidney cells under hypertonic stress. We found that stable IMCD3 clones silenced for ZAC1 to varying levels demonstrated an inverse effect on SDH expression. ZAC1 binds to a consensus repression site within the promoter of SDH, pointing to a mechanism whereby ZAC1 acts by repressing SDH transcriptional activity during hypertonic conditions. Taken together, these data strongly suggest that ZAC1 is up-regulated under hypertonic stress and negatively regulates expression of SDH, allowing for accumulation of sorbitol as a compatible organic osmolyte.

ZAC1 is up‐regulated by hypertonicity and decreases sorbitol dehydrogenase expression allowing accumulation of sorbitol in kidney cells

The zinc‐finger protein ZAC1 was determined to be markedly upregulated under hypertonic stress in inner medullary collecting duct (IMCD3) cells by Affymetrix genechip analysis (219‐fold, p&lt;0.001). This change in message was verified by Quantitative PCR and by Western blot for protein. In mouse and human kidney tissues, ZAC1 expression was substantial in the papilla and absent in the cortex. ZAC1 protein expression was localized to the nucleus in cells under hypertonic stress by immunocytochemical analysis. Since a previous study in the hippocampal cell line HW3‐5 reported an inverse correlation between the expression of ZAC1 and sorbitol dehydrogenase (SDH), ‐a key enzyme involved in sorbitol degradation that is down regulated with hypertonicity. We examined if the decrease in SDH expression under hypertonic stress is mediated by the up‐regulation of ZAC1. Stable IMCD3 clones silenced for ZAC1 to varying levels demonstrated a direct effect on SDH expression. Silencing ZAC1 expression under hypertonic conditions caused maintenance of high SDH protein expression which prevented sorbitol accumulation in IMCD3 cells. Taken together, these data indicates that ZAC1 is up‐regulated under hypertonic stress and negatively regulates expression of SDH allowing for accumulation of sorbitol as a compatible organic osmolyte.

In vitro aldose reductase inhibitory activity of some flavonyl-2,4-thiazolidinediones

Aldose reductase (AR) is implicated to play a critical role in diabetes and cardiovascular complications because of the reaction it catalyzes. AR enzyme appears to be the key factor in the reduction of glucose to sorbitol. Synthesis and accumulation of sorbitol in cells due to AR activity is the main cause of diabetic complications, such as diabetic cataract, retinopathy, neuropathy and nephropathy. Aldose reductase inhibitors have been found to prevent sorbitol accumulation in tissues. Numerous compounds have been prepared in order to improve the pharmacological prophile of inhibition of aldose reductase enzyme. In this study, seventeen flavonyl-2,4-thiazolidinediones (flavonyl-2,4-TZD) (Ia–e, IIa–e and IIIa–g) were tested for their ability to inhibit rat kidney AR. Compound Ib showed the highest inhibitory activity (88.69 ± 1.46%) whereas Ia, IIa, IIIa, IIIb also showed significant inhibitory activity (49.26 ± 2.85, 67.29 ± 1.09, 71.11 ± 1.95, 64.86 ± 1.21%, respectively).

Impact of mitochondrial ROS production on diabetic vascular complications

Vascular complications are the leading cause of morbidity and mortality in patients with diabetes. Four main molecular mechanisms have been implicated in glucose-mediated vascular disease. There are: glucose-induced activation of protein kinase C (PKC) isoforms; increased formation of glucose-derived advanced glycation end-products (AGE); increased glucose flux through the aldose reductase pathway; and increased production of reactive oxygen species (ROS). Here we demonstrate that hyperglycemia-induced production of ROS is abrogated by inhibitors of mitochondrial metabolism, or by overexpression of uncoupling protein-1 or manganese superoxide dismutase. Normalization of mitochondrial ROS production by each of these agents prevents glucose-induced activation of PKC, formation of AGE, and accumulation of sorbitol in bovine vascular endothelial cells. We also claim that 8-hydroxydeoxyguanosine, which represents mitochondrial oxidative damage was elevated in patients with either retinopathy, albuminuria or increased intima-media thickness of carotid arteries. These results suggest that hyperglycemia induces mitochondrial ROS production, and which can associate to the pathogenesis of diabetic vascular complications.

Fibrates inhibit aldose reductase activity in the forward and reverse reactions

Fibrates such as bezafibrate, gemfibrozil, clofibric acid, ciprofibrate and fenofibrate, are ligands for peroxisome proliferator-activated receptor α (PPARα), and are used as therapeutic agents in the treatment of hyperlipidemia. Synthesis and accumulation of sorbitol in cells due to aldose reductase (AR) activity is implicated in secondary diabetic complications. In pursuit of finding a lead compound identification to design an effective AR inhibitor employing fragment-based design-like approach, we found that this class of compounds and their nearest neighbors could inhibit AR. Bezafibrate and gemfibrozil displayed a mixed non-competitive inhibition pattern in the glyceraldehyde reduction activity and pure non-competitive inhibition pattern in the benzyl alcohol oxidation activity of AR. Clofibric acid, ciprofibrate and fenofibrate showed pure non-competitive inhibition patterns in the forward reaction. In the reverse reaction, clofibric acid displayed a non-competitive inhibition pattern while ciprofibrate and fenofibrate displayed competitive inhibition patterns. This finding reveals for the first time a novel attribute of the fibrates in the regulation of AR activity and may be useful as lead compounds to control the function of AR in the progression and treatment of secondary diabetic complications in addition to other clinical conditions. Alternatively, these findings demonstrate that AR plays a significant role in the fibrate metabolism under various scenarios.

Quantitative determination of endogenous sorbitol and fructose in human erythrocytes by atmospheric-pressure chemical ionization LC tandem mass spectrometry

Evaluation of different extraction methods for quantification of endogenous sorbitol and fructose in human red blood cells (RBCs) and matrix effects in ESI and APCI showed that protein-precipitation followed by mixed-mode solid-phase extraction was more effective extraction method and APCI more effective ionization method. Then the LC/APCI-MS/MS method was fully validated and successfully applied to analysis of clinical RBC samples. The concentrations of endogenous sorbitol and fructose were determined using calibration curves employing sorbitiol- 13C 6 and fructose- 13C 6 as surrogate analytes. The method has provided excellent intra- and inter-assay precision and accuracy with a linear range of 50.0–10,000 ng/mL (correlation coefficient >0.999) for sorbitol- 13C 6 and 250–50000 ng/mL (correlation coefficient >0.999) for fructose- 13C 6 in human RBCs.

Black and Green Teas Equally Inhibit Diabetic Cataracts in a Streptozotocin-Induced Rat Model of Diabetes

Green and black teas were given at 1.25% in the drinking water to streptozotocin-induced diabetic rats for 3 months. Normal and diabetic control groups were also studied. As expected, diabetic animals had significantly increased glucose in lens and plasma. Lens and red blood cell sorbitol were significantly increased as a result of the aldose reductase pathway activation. Plasma and lens lipid thiobarbituric acid-reactive substances and protein glycation were also significantly elevated. Both teas significantly inhibited diabetic cataracts and caused significant reductions in the biochemical pathway implicated in the development of the pathology. After corrections for glucose, it was found that the teas retard the development of diabetic cataracts by a hypoglycemic effect that in turn inhibits the biochemical indicators of pathology. There were significant correlations between glucose, cataract score, and these indicators. Green tea but not black tea caused a significant decline in triglycerides in the diabetic animals. Tea may be a simple, inexpensive means of preventing or retarding human diabetes and the ensuing complications. Tea also should be investigated as an adjunct therapy for diabetes treatment.

Sorbitol Transporter Expression in Apple Sink Tissues: Implications for Fruit Sugar Accumulation and Watercore Development

In apple (Malus ×domestica Borkh.), where sorbitol is a primary photosynthetic product that is translocated throughout the plant, accumulation of sorbitol in sink cells appears to require an active carrier-mediated membrane transport step. Recent progress in isolation and characterization of genes for sorbitol transporters in sour cherry (Prunus cerasus L.) and mannitol transporters in celery (Apium graveolens L.) suggested that similar transporters may be present in apple tissues. A defect in these transporters could also explain the occurrence of the fruit disorder watercore, characterized by the accumulation of fluids and sorbitol in the apoplasmic free space. Our objectives therefore included isolation and characterization of genes for sorbitol transporters in apple tissues and comparisons of expression of transporter genes, especially in various sink tissues including watercored and non-watercored fruit tissues. We have isolated and characterized two sorbitol transporter genes, MdSOT1 and MdSOT2. Sequence analyses indicated that these are members of the major facilitator transporter superfamily that gives rise to highly hydrophobic integral membrane proteins. Heterologous expression and measurement of sorbitol uptake in yeast indicated that these are specific and with high affinities for sorbitol, with Kms for sorbitol of 1.0 and 7.8 mm for MdSOT1 and MdSOT2, respectively. Sorbitol transporter expression was evident in all sink tissues tested with the exception of watercore-affected fruit tissues. Sorbitol accumulation in apple sink tissues thus involves an apoplasmic active membrane transport step and watercore results from a defect in that process.

A sensitive assay of red blood cell sorbitol level by high performance liquid chromatography: potential for diagnostic evaluation of diabetes

Background The level of sorbitol within erythrocyte or other tissue is increased during diabetic complications such as neuropathy, retinopathy, and nephropathy and is thus considered as an indicator for the diagnosis and monitoring the diabetic status. A highly sensitive method of estimation of RBC sorbitol may be very useful in determining the state of diabetes. In this study a high performance liquid chromatography method was developed to analyze RBC sorbitol level. Method Benzoylation was used for the precolumn derivatization of RBC polyol prior to analyze by HPLC using C-18 reversed phase column and UV detector. Results The sensitivity was 5 ng/ml using standard sorbitol. The linearity was highly satisfactory up to 2.5 μg/ml which was adequate to measure RBC sorbitol in both diabetic and normal subjects. The values were significantly higher for the diabetic patient compared to normal. Within-run assay CVs were <10% and <14% for between-run. Conclusion This method could be useful in estimating RBC sorbitol content for the clinical evaluation of diabetic patients.

Transcriptome profiling and the pathogenesis of diabetic complications.

Diabetes is an escalating problem worldwide and a major cause of vascular disease, renal failure, and blindness, among other complications. The cellular mediators of high glucose-induced injury include activation of protein kinase C, accumulation of cell sorbitol from increased flux through the aldose reductase pathway, and generation of advanced glycosylation end products and reactive oxygen species, among others. Current strategies for preventing and slowing the progression of the macrovascular and microvascular complications of diabetes include optimization of glycemic control and BP, angiotensin-converting enzyme inhibitors and angiotensin II blockers, and HMG CoA reductase inhibitors. However, there is an urgent need to develop new therapeutic strategies, as these interventions, although they may slow, rarely halt the progression of diabetic complications. Central to this process is the elucidation of the molecular events that drive this complex disease and that are potential therapeutic targets. This review discusses the promise offered in this regard by global monitoring of cellular or tissue mRNA expression (so-called transcriptomics) and illustrates the potential of this approach by focusing on recent studies on the pathogenesis of diabetic nephropathy.

Prevention of retinal capillary basement membrane thickening in diabetic dogs by a non-steroidal anti-inflammatory drug.

To investigate the effect of treatment with the non-steroidal anti-inflammatory drug Sulindac on the early vascular pathology of diabetic retinopathy in the dog, and it's effect on recognised biochemical indices of hyperglycaemia-related pathophysiology. Experimental diabetes (streptozotocin/alloxan) was induced in 22 male beagle dogs and 12 of the animals were assigned at random to receive oral Sulindac (10 mg/kg daily). Age- and sex-matched control animals were maintained as non-diabetic controls. After 4 years, several morphological parameters were quantified in the retinal microvasculature of each animal group using an established stereological method. Also, the following diabetes-associated biochemical parameters were analysed: accumulation of advanced glycation end products (AGEs), red blood cell polyol levels and antioxidant status. Diabetes increased red blood cell sorbitol levels when compared to non-diabetic controls (p< or =0.05), however, there was no difference in sorbitol levels between the untreated and the treated diabetic animals. No significant differences were found in red blood cell myoinositol levels between the three groups of animals. Pentosidine and other AGEs were increased two- to three-fold in the diabetic animals (p< or =0.001) although treatment with Sulindac did not affect their accumulation in diabetic skin collagen or alter diabetes-induced rises in plasma malondialdehyde. Retinal capillary basement membrane volume was significantly increased in the untreated diabetic dogs compared to non-diabetic controls or Sulindac-treated diabetic animals (p< or =0.0001). This study has confirmed the beneficial effect of a non-steroidal anti-inflammatory drug on the early vascular pathology of diabetic retinopathy. However the treatment benefit was not dependent on inhibition of polyol pathway activity, advanced glycation, or oxidative stress.

Stimulation of GLUT-1 glucose transporter expression in response to hyperosmolarity.

Glucose transporter isoform-1 (GLUT-1) expression is stimulated in response to stressful conditions. Here we examined the mechanisms mediating the enhanced expression of GLUT-1 by hyperosmolarity. GLUT-1 mRNA, GLUT-1 protein, and glucose transport increased after exposure of Clone 9 cells to 600 mosmol/l (produced by addition of mannitol). The stimulation of glucose transport was biphasic: in the early phase (0-6 h) a approximately 2.5-fold stimulation of glucose uptake was associated with no change in the content of GLUT-1 mRNA, GLUT-1 protein, or GLUT-1 in the plasma membrane, whereas the approximately 17-fold stimulation of glucose transport during the late phase (12-24 h) was associated with increases in both GLUT-1 mRNA (approximately 7.5-fold) and GLUT-1 protein content. Cell sorbitol increased after 3 h of exposure to hyperosmolarity. The increase in GLUT-1 mRNA content was associated with an increase in the half-life of the mRNA from 2 to 8 h. A 44-bp region in the proximal GLUT-1 promoter was necessary for basal activity and for the two- to threefold increases in expression by hyperosmolarity. It is concluded that the increase in GLUT-1 mRNA content is mediated by both enhanced transcription and stabilization of GLUT-1 mRNA and is associated with increases in GLUT-1 content and glucose transport activity.

The Saccharomyces cerevisiae aldose reductase is implied in the metabolism of methylglyoxal in response to stress conditions.

The enzyme aldose reductase plays an important role in the osmo-protection mechanism of diverse organisms. Here, we show that yeast aldose reductase is encoded by the GRE3 gene. Expression of GRE3 is carbon-source independent and up-regulated by different stress conditions, such as NaCl, H2O2, 39 degrees C and carbon starvation. Measurements of enzyme activity and intracellular sorbitol in wild-type cells also indicate that yeast aldose reductase is stress-regulated. Overexpression of GRE3 increases methylglyoxal tolerance in Saccharomyces cerevisiae. Furthermore, high expression of GRE3 complements the deficiency of the glyoxalase system of a glo1delta mutant strain. Consistent with this, in vitro and in vivo assays of yeast aldose reductase activity indicate that methylglyoxal is an endogenous substrate of aldose reductase. Furthermore, addition of NaCl or H2O2 to exponential-phase cells triggers an initial transient increase in the intracellular level of methylglyoxal, which is dependent on the Gre3p and Glo1p function. These observations indicate that the metabolism of methylglyoxal is stimulated under stress conditions; and they support a methylglyoxal degradative pathway, in which this compound is metabolised by the action of aldose reductase.