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T Inoguchi

Publications and source records attributed to T Inoguchi.

At least 37 records · Page 2Linked to original sources
PubMed1994

Characterization of the mechanism for the chronic activation of diacylglycerol-protein kinase C pathway in diabetes and hypergalactosemia.

Similar vascular pathological conditions are observed in diabetic animals and those with diet-induced hypergalactosemia. Both diabetes and hypergalactosemia are believed to cause vascular dysfunction via a common biochemical mechanism. In this study, we have found that both diabetes and hypergalactosemia in the short term (2-4 months) can increase total diacylglycerol (DAG) levels by 52 +/- 9 and 74 +/- 13% in the retina and aorta, respectively, of diabetic dogs, and by 94 +/- 9 and 78 +/- 11% in the retina and aorta, respectively, in dogs with hypergalactosemia as compared with normal control animals (P < 0.01). The elevation of DAG levels was maintained for 5 years in the aortas of diabetic and hypergalactosemic dogs. To characterize the mechanism of the DAG increases, we have determined that total DAG levels were significantly increased in cultured macro- and microvascular cells exposed to elevated glucose (22 mM) and galactose (16.5 mM) levels. These increased levels were not prevented by sorbinil, an aldose reductase inhibitor. One of the sources of the increased DAG levels was probably derived from de novo synthesis from both hexoses as determined by radiolabeling studies. Intracellularly, the DAG elevation activated protein kinase C (PKC) activity with increases of 58 +/- 12% (P < 0.05) and 66 +/- 8% (P < 0.01) in the membrane fraction of cultured aortic smooth muscle cells exposed to elevated glucose and galactose levels, respectively. These findings have clearly demonstrated a possible common biochemical mechanism by which hyperglycemia and hypergalactosemia can chronically activate the DAG-PKC pathway in the vasculature and could be a possible explanation for the development of diabetic vascular complications.

Animals↗
PubMed1993

Correlation of diacylglycerol level and protein kinase C activity in rat retina to retinal circulation.

The increases in diacylglycerol (DAG) level and protein kinase C (PKC) activity have been characterized biochemically and functionally in the retina and the brain of diabetic rats as well as in cultured vascular cells. PKC specific activities were increased in the membraneous fraction of retina from streptozotocin (STZ)-induced diabetic rats and the genetically determined diabetic BB rats, respectively, after 1 or 2 wk of diabetes, compared with control. The ratio of total PKC activities from membraneous and cytosol fractions was also increased in the retina of diabetic rats. With diabetes, all the isoenzymes and the total DAG level were increased in the rat retina, whereas no changes were found in the rat brain. Insulin treatment normalized plasma glucose levels and partially prevented the increases in the membraneous PKC activity and all the isoenzymes in the retina. In the retinal endothelial cells, the total DAG level and PKC specific activities are increased by 36 and 22%, respectively, in the membraneous pool when the glucose levels are changed from 5.5 to 22 mM. Activation of PKC activity and isoform beta II by the vitreal injection of phorbol dibutyrate mimicked the abnormal retinal blood circulation observed in diabetic rats (2.22 +/- 0.24 vs. 1.83 +/- 0.40 s). Thus diabetes and elevated glucose levels will increase DAG level and PKC activities and its isoenzyme specifically in vascular cells and may affect retinal hemodynamics.

Amino Acid Sequence↗
PubMed1992

Vitamin E binds to specific binding sites and enhances prostacyclin production by cultured aortic endothelial cells.

We evaluated the effect of d-alpha-tocopherol (vitamin E) on the production of prostacyclin (PGI2) by cultured bovine aortic endothelial cells. Vitamin E at physiological doses significantly enhanced the production of PGI2 by aortic endothelial cells when added to the culture simultaneously with histamine, the Ca2+ ionophore A23187 (A23187), plasma-derived serum (PDS), or arachidonic acid. This effect was found to occur in a time- and dose-dependent manner, and the maximal enhancement was produced by 9.28 microM of vitamin E for 1 h incubations. Significantly lower amounts of lipid peroxides were measured in endothelial cells stimulated by 10% PDS with 9.28 microM of vitamin E than in those stimulated without vitamin E for over 24 h, although the stimulation during the initial 1 to 12 h period did not have a significant effect on lipid peroxide formation in cultured aortic endothelial cells. We also demonstrated that bovine aortic endothelial cells have specific binding sites for [3H]vitamin E that exhibited time- and temperature-dependent saturability. At 4 degrees C, the nonspecific binding was 8-12% of the total binding, and the specific binding reached equilibrium by 2 h. Specific binding increased with the concentration of [3H]vitamin E and became saturated at concentrations between 1.5 microM and 2.0 microM per 2.0 x 10(5) cells. Raising the unlabeled vitamin E concentration from 97.7 nM to 1,000 microM reduced the specific binding of 2.0 microM [3H]vitamin E.(ABSTRACT TRUNCATED AT 250 WORDS)

6-Ketoprostaglandin F1 alpha↗
PubMed1992

Preferential elevation of protein kinase C isoform beta II and diacylglycerol levels in the aorta and heart of diabetic rats: differential reversibility to glycemic control by islet cell transplantation.

In the present study, we have measured protein kinase C (PKC) specific activities and total diacylglycerol (DAG) level in the aorta and heart of rats, which showed that after 2 weeks of streptozotocin (STZ)-induced diabetes, membranous PKC specific activity and total DAG content were increased significantly by 88% and 40% in the aorta and by 21% and 72% in the heart, respectively. Hyperglycemia was identified as being a causal factor since elevated glucose levels increased DAG levels in cultured aortic endothelial and smooth muscle cells. Analysis by immunoblotting revealed that only alpha and beta II PKC isoenzymes are detected in these two tissues and vascular cells among those studied. In STZ-induced diabetic rats, beta II isoenzyme is preferentially increased in both aorta and heart, whereas PKC alpha did not change significantly. The increases in membranous PKC specific activity and DAG level are observed in both spontaneous diabetes-prone diabetic BB rats as well as in STZ-induced diabetic BB and Sprague-Dawley rats, which persisted for up to 5 weeks. After 2 weeks of diabetes without treatment, the normalization of blood glucose levels for up to 3 weeks with islet cell transplants in STZ-induced diabetic BB rats reversed the biochemical changes only in the heart, but not in the aorta. These results suggest that PKC activity and DAG level may be persistently activated in the macrovascular tissues from diabetic animals and indicate a possible role for these biochemical parameters in the development of diabetic chronic vascular complications.

Animals↗
PubMed1992

Vitamin E restores reduced prostacyclin synthesis in aortic endothelial cells cultured with a high concentration of glucose.

Reduced prostacyclin (PGI2) production by the vascular wall may play an important role in the pathogenesis of vascular lesions such as atherosclerosis. The present study was undertaken to evaluate the effect of vitamin E on the production of PGI2 and other prostaglandins (prostaglandin E2 [PGE2], thromboxane A2 [TXA2], and 15-hydroxyeicosatetraenoic acid [15-HETE]) by bovine aortic endothelial cells cultured in a high concentration of glucose (300 mg/dL). Compared with endothelial cells cultured in 100 mg/dL glucose, the production of PGI2 and other prostaglandins, except 15-HETE, was significantly reduced in cultures containing 300 mg/dL glucose when stimulated by histamine, the Ca2+ ionophore, A23187, or human plasma-derived serum (PDS). The addition of vitamin E to each stimulant significantly restored the production of PGI2, PGE2, and TXA2, products of the cyclo-oxygenase pathway, in aortic endothelial cells cultured in 300 mg/dL glucose. This effect of vitamin E on the stimulation of prostaglandin production was generally specific for D-alpha-tocopherol, but not for the other vitamin E analogs tested. However, vitamin E and the stimulants had no effect on the production of 15-HETE, a product of the lipoxygenase pathway. Moreover, vitamin E alone, without stimulants, did not affect prostaglandin production in cultured bovine aortic endothelial cells. These results suggest that vitamin E may restore reduced PGI2, PGE2, or TXA2 production by bovine aortic endothelial cells cultured in a high concentration of glucose. It seems likely that vitamin E may restore depressed PGI2 production by the vascular wall in hyperglycemic conditions such as those seen in patients with diabetes mellitus.

Animals↗
PubMed1992

Partial purification of serum prostacyclin stimulatory activity by heparin-agarose column; abnormality detected in diabetics.

Human plasma-derived serum (PDS) stimulated the production of 6-keto-PGF1 alpha (a stable metabolite of PGI2) by cultured bovine aortic endothelial cells. The stimulation was both time- and dose-dependent. The main prostacyclin stimulatory activity (PSA) in human PDS remained biologically active after dialysis and was inhibited by the simultaneous addition of heparin. The maximum inhibition of PSA was obtained with 10 micrograms/ml heparin. PDS obtained from patients with non-insulin-dependent diabetes mellitus (NIDDM, n = 24) showed significantly less PSA than that from the control subjects (n = 11). A decrease in PSA was also found in diabetic patients using dialyzed PDS. The PSA in human PDS had a specific binding affinity to heparin-agarose and the bound PSA was eluted by a linear gradient of NaCl, which showed two major PSA peaks at 1.0 and 1.5 M NaCl. The dialyzed, mixed PDS from patients with NIDDM and the control subjects was independently applied to a heparin-agarose column and eluted by a linear gradient of NaCl. Comparing the PSA in each peak between the diabetic and the control dialyzed PDS, the PSA at 1.5 M NaCl was markedly decreased in the diabetic patients, but the PSA at 1.0 M NaCl did not change significantly. These observations suggest that the decreased PSA in human diabetic PDS may result mainly from the decrease in the activity of a specific non-dialyzed factor(s) which can bind to heparin. The decreased PSA in serum seems to be responsible in part for decreased PGI2 synthesis by the vascular wall of diabetics.

6-Ketoprostaglandin F1 alpha↗
PubMed1992

Effect of glucose on Na, K-ATPase activity in cultured bovine aortic endothelial cells.

The effect of high concentrations of glucose on Na, K-ATPase activity and the polyol pathway was studied using cultured bovine aortic endothelial cells. Na, K-ATPase activity was expressed as ouabain-sensitive K+ uptake. A significant decrease in Na, K-ATPase activity with an intracellular accumulation of sorbitol was found in confluent endothelial cells incubated with 400 mg/dl glucose for 96 h. However, there was no significant change in the Na, K-ATPase activity or sorbitol content of the cells incubated with 100 mg/dl glucose plus 300 mg/dl mannitol. The decrease in Na, K-ATPase induced by the high glucose concentration was restored by the simultaneous addition of 10(-4) M ponalrestat (ICI 128,436; Statil), an aldose reductase inhibitor. The addition of this agent also significantly reduced the increase in sorbitol induced by high glucose levels. These results suggest that the decrease in Na, K-ATPase activity induced in cultured aortic endothelial cells by high concentrations of glucose may be caused in part by the accumulation of sorbitol.

Aldehyde Reductase↗
PubMed1992

Induction of resistance to endothelin-1's biochemical actions by elevated glucose levels in retinal pericytes.

Because retinal pericytes have contractile properties and are affected by diabetes, we have studied the responsiveness of pericytes to ET-1, a potent vasoconstrictor, in the presence of various concentrations of glucose. Cultured calf retinal pericytes were exposed to glucose levels of 5.5 or 25 mM for up to 8 days. Radioreceptor studies that used [125I]ET-1 showed that pericytes contained high-affinity binding sites with Kd of 3 x 10(-10) M, and these binding affinities were unaffected by glucose concentration. Receptor number appears to be elevated, but this increase was NS. Responsiveness of pericytes to ET-1 was studied with respect to stimulation of DAG and IP3 levels and PKC activities. In contrast to receptor binding, exposure to 25 mM glucose for > 6 days blunted pericyte responsiveness to ET-1. The time course of ET-1 stimulation as measured by [3H]glycerol labeling, and IP3 level showed a 98% increase in [3H]DAG at 10 min and a fourfold increase for IP3, respectively. Cells exposed to 25 mM glucose only had a 32% increase for DAG, and no increase for IP3 was observed. Dose-response studies on the stimulation of [3H]DAG increase showed the range of ET-1's effect to be between 10(-9) and 10(-7) M. At maximum, cells exposed to 5.5 mM glucose had a 70% increase versus only a 30% increase in those exposed to 25 mM glucose. Similarly, ET-1 only increased the total DAG levels in pericytes exposed to 5.5 mM glucose by 41%. PKC activity also was measured because DAG is one of its cellular activators.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Platelet stimulation for prostacyclin production in aortic endothelial cell cultures: alteration in diabetes mellitus.

We evaluated the effect of platelets on prostacyclin (PGI2) production in bovine aortic endothelial cell cultures. Human platelet extract significantly stimulated PGI2 production by cultured aortic endothelial cells in a time- and dose-dependent manner, suggesting that platelets contain PGI2-stimulatory activity (PSA). Supernatant fluid separated from platelets activated by collagen also exhibited PSA. The factor(s) causing the PSA of platelets was non-dialysable and heat-stable (56 degrees C for 30 min or 100 degrees C for 3 min), was completely inhibited by trypsin pretreatment, and exhibited an affinity to heparin agarose. Furthermore, gel filtration chromatography showed that the factor(s) responsible for the platelet PSA was eluted at three different peaks with approximate molecular weights of 50,000, 25,000 and 11,000. The PSA of platelet extract from patients with non-insulin-dependent diabetes mellitus (NIDDM) (n = 10) was compared to that from age-matched control subjects (n = 10). Platelet extract from patients with NIDDM stimulated cultured aortic endothelial cells to produce greater amounts of PGI2 than did that from control subjects. These data suggest that the increased PSA of platelets isolated from diabetic patients may contribute to the abnormal interaction between platelets and the vascular wall in diabetic patients.

Animals↗
PubMed1991

Haemostatic factors associated with vascular thrombosis in patients with systemic lupus erythematosus and the lupus anticoagulant.

To elucidate the mechanism of vascular thrombosis in patients with systemic lupus erythematosus and the lupus anticoagulant changes in factors associated with haemostasis were investigated. The lupus anticoagulant was associated with an increased incidence of thrombosis, particularly cerebral thrombosis. Concentrations of fibrinopeptide A and fibrinopeptide B beta 15-42 were significantly raised in the plasma of patients with systemic lupus erythematosus and the anticoagulant compared with concentrations in patients without the lupus anticoagulant. The tendency towards formation of thrombosis was not found in all lupus patients with the anticoagulant, however. Concentrations of thromboxane B2 were remarkably raised in the plasma of the two patients with the lupus anticoagulant who had recently had thrombosis. Concentrations of 6-keto-prostaglandin F1 alpha, protein C, antithrombin III, and plasminogen were similar in both groups. No significant decrease in serum stimulatory activity on prostacyclin production by cultured aortic endothelial cells was noted in lupus patients with the anticoagulant, but inhibition was present in the two patients with recent thrombosis. These results indicate that although patients with the lupus anticoagulant are not always in a hypercoagulable state, haemostatic abnormalities found in some patients with the anticoagulant may be predictive of thrombotic events.

6-Ketoprostaglandin F1 alpha↗
PubMed1990

Modification of prostacyclin-stimulatory activity in sera by glucose, insulin, low density lipoprotein, linoleic acid and linoleic acid hydroperoxide.

Reduced prostacyclin (PGI2) production by the vascular wall has been proposed as one of the possible causes of diabetic vascular complications. We found an activity which stimulated PGI2 production by cultured endothelial cells (PGI2-stimulatory activity, PSA) in human plasma-derived serum (PDS). The PSA was less in patients with diabetes mellitus. The present study was undertaken to evaluate how metabolic factors relevant to diabetic angiopathy modify the PSA. Pooled PDS was prepared from 10 healthy volunteers. The 6-keto-PGF1 alpha (6KF, a stable metabolite of PGI2) production by cultured bovine aortic endothelial cells was maximally stimulated by Dulbecco's modified Eagle's medium (DMEM) containing 10% pooled PDS after incubation for 60 min. The production of 6KF was reduced in a dose-dependent manner by the addition of 10% pooled PDS with glucose and linoleic acid hydroperoxide (lipid peroxide). In contrast, human low density lipoprotein (LDL) and linoleic acid (unsaturated fatty acid) enhanced the production of 6KF by 10% pooled PDS in a dose-dependent manner. Insulin, however, showed no effect on the production of 6KF by 10% pooled PDS. These results suggest that the reduced PSA in diabetics may be the result, in part, of a modification of the PSA by diabetic metabolic factors such as glucose and lipid peroxide.

6-Ketoprostaglandin F1 alpha↗
PubMed1990

Effects of captopril on urinary excretion of albumin and prostaglandins in patients with diabetic nephropathy.

Recently the beneficial effects of captopril (angiotensin-converting-enzyme inhibitor) on diabetic nephropathy, especially proteinuria, have been reported by some investigators. The mechanism whereby proteinuria is reduced, however, have not been clarified yet. The present study was undertaken to evaluate the effects of captopril on urinary albumin excretion in relation to urinary prostaglandins (PGs) excretion in patients with non-insulin-dependent diabetes mellitus (NIDDM). Captopril (37.5 mg/day) was orally administered to 13 diabetic patients for an eight-week period. A single administration of captopril (12.5 mg) was performed in the same patients. The spot urine samples were collected in the early morning and 2 hr after the single administration of captopril. The albumin index (mg/gram-creatinine) was markedly decreased in eight patients (Group A) within four weeks, but no decrease was found in five patients (Group B). Furthermore, in Group A, by the single administration of captopril urinary excretions of 6-keto-PCF1 alpha (a stable metabolite of PGI2) and PGE2 were significantly (p less than 0.05) increased while urinary TXB2 (a stable metabolite of TXA2) excretion did not change significantly. The urinary 6-keto-PGF1 alpha/TXB2 ratio, which is significantly (p less than 0.05) low in diabetic patients was significantly (p less than 0.01) increased up to the normal value in Group A. In Group B, however, there were no significant changes in urinary PGs excretion. These results suggest that captopril enhances the production of intrarenal vasodilatory PGs such as PGI2 and PGE2, which may be deeply involved in the reduction of intraglomerular capillary pressure and urinary protein excretion in diabetic patients.

6-Ketoprostaglandin F1 alpha↗
PubMed1990

Effects of vitamin E administration on platelet function in diabetes mellitus.

A decrease in the vitamin E content of human diabetic platelets is closely associated with the accelerated platelet aggregation and platelet prostaglandin metabolism seen in patients with diabetes mellitus. We investigated the effect of vitamin E supplementation on these abnormalities of physiological function and prostaglandin metabolism in 14 non-insulin dependent diabetics with proliferative retinopathy. ADP-induced platelet aggregation was inhibited in vitro by the addition of vitamin E in a dose-dependent manner. However, in lower concentrations considered to be physiological doses in vivo, significantly greater inhibition was observed in diabetic platelets than in the control platelets. Next, alpha-tocopheryl nicotinate was administered to diabetics at a daily dose of 600 mg. The platelet vitamin E content was restored to control levels in 13 of the 14 patients after 2-4 weeks of daily administration. The ADP-induced platelet aggregation rate, platelet thromboxane B2 (TXB2, a stable metabolite of TXA2, a vasoconstrictor production, and plasma TXB2 level were low in all 14 diabetics. In contrast, plasma 6-keto-PGF 1 alpha (a stable metabolite of PGI2, a vasodilator) was significantly increased and therefore the 6-keto-PGF 1 alpha/TXB2 ratio in plasma was restored to within normal limits. These results indicate that vitamin E may improve platelet function and prostaglandin metabolism in diabetes mellitus and may be able to provide further beneficial effects in relation to the development of diabetic vascular complications.

6-Ketoprostaglandin F1 alpha↗
PubMed1989

Antithrombin III stimulates prostacyclin production by cultured aortic endothelial cells.

Prostacyclin (PGI2) is a potent vasodilator and an inhibitor of platelet aggregation. We found that antithrombin III (AT III), an anticoagulant present in circulating blood, stimulated PGI2 production by cultured bovine aortic endothelial cells in a dose- and time-dependent manner. The stimulation of PGI2 production by AT III was observed at physiological concentrations and was inhibited by the addition of anti-AT III antiserum and heparin. These results suggest that AT III may stimulate PGI2 production by binding to heparin-like molecules on the endothelial cell membrane.

6-Ketoprostaglandin F1 alpha↗
PubMed1989

Decrease in insulin binding to aortic endothelial cells cultured with high glucose concentration.

We studied the effects of glucose on specific insulin binding to cultured endothelial cells from the bovine aorta. We cultured the cells in Dulbecco's modified Eagle's medium, containing 100 or 300 mg/dl glucose or 100 mg/dl glucose plus 200 mg/dl mannitol. We added 125I-insulin to monolayers of these cells and counted the radioactivity resulting. Specific insulin binding increased with time and dosage. Maximal binding resulted from 0.17 nM 125I-insulin being incubated for 30 min at 37 degrees C, and amounted to 0.35% per 10(5) cells being bound in cultures of 100 mg/dl glucose. The higher concentration of glucose led to significantly less binding (P less than 0.05) (0.24 +/- 0.03% vs. 0.38 +/- 0.02%, n = 6). The addition of mannitol, on the other hand, did not affect binding. All three incubation conditions produced curvilinear competition curves. Scatchard analysis showed that insulin bound significantly less (P less than 0.05) to endothelial cells in 300 mg/dl glucose than to those in 100 mg/dl glucose (4.0 +/- 1.2 nmol/l vs. 12.8 +/- 3.1, n = 6, mean +/- SEM). Insulin binding capacity, however, did not change. We conclude that glucose can reduce insulin binding to endothelial cells and that it may reduce receptor-related insulin transport into and out of the cells.

Animals↗
PubMed1989

Reduced stimulatory activity on prostacyclin production by cultured endothelial cells in serum from aged and diabetic patients.

Reduced prostacyclin (PGI2) generation by the vascular wall shows a close relationship with the development of atherosclerosis. The present study found plasma-derived serum (PDS) to contain an activity which stimulated PGI2 production by cultured bovine aortic endothelial cells. Diabetic and aged patients with atherosclerotic disease were examined for abnormalities in that stimulatory activity in PDS. PDS obtained from both diabetics (NIDDM) and aged patients showed a significant reduction in the stimulation of PGI2 production by cultured bovine aortic endothelial cells compared with age-matched controls and young healthy volunteers, respectively. It was suggested that the reduced PGI2 stimulatory activity in PDS may be one of the pathogenic mechanisms of vascular lesions such as atherosclerosis in diabetics and aged humans.

Adult↗
PubMed1989

Abnormality in prostacyclin-stimulatory activity in sera from diabetics.

A reduction in prostacyclin (PGI2) production by vascular wall may cause platelet hyperaggregability in diabetics, which is considered to be a possible pathogenesis of diabetic vascular complications. In the present study, the presence of PGI2-stimulatory activity (PSA) in rat and human plasma-derived serum (PDS) was confirmed by cultured bovine aortic endothelial cells. PSA in PDS was significantly decreased in streptozotocin-induced diabetic rats and in patients with non-insulin-dependent diabetes mellitus (NIDDM). PDS from patients with NIDDM showed less PSA prior to the clinical onset of diabetic vascular complications, such as retinopathy and proteinuria. The reduction in PSA was still observed in dialyzed PDS from the patients with NIDDM. The nondialyzable PSA was heat-stable at 56 degrees C for 30 minutes and partially stable at 100 degrees C for five minutes. This activity was not extractable with diethylether and was precipitable with trichloroacetic acid. The study of Sephadex G-50 column chromatography showed that a major part of PSA in dialyzed PDS was found in the area of the molecular weight of 12,000 to 17,000 daltons. In conclusion, the reduction in PSA from diabetics may cause a reduction of PGI2 production by vascular wall, subsequently contributing to the development of diabetic vascular complications.

6-Ketoprostaglandin F1 alpha↗