Increased basal levels of plasma nitric oxide in Type 2 diabetic subjects. Relationship to microvascular complications.

To assess the underlying mechanisms of decreased endothelial function and advanced vascular complications in patients with Type 2 diabetes, we determined basal levels of plasma nitric oxide (NO(x): NO(2)(-) and NO(3)(-)) using a newly developed high-performance liquid chromatography (HPLC)-Griess method in hospitalized 129 diabetic and 76 nondiabetic subjects, and examined their clinical characteristics. Serum lipid peroxide and advanced glycation end products (AGEs) as markers of oxidative stress were also measured, and intima-media thickness (IMT) of the carotid artery was evaluated as a marker of atherosclerosis. In diabetic subjects, microvascular complications were newly evaluated during their admission. There were no differences in age or sex between the diabetic and nondiabetic subjects. Although there was no difference in basal plasma NO(2)(-) levels between the two groups, the basal levels of plasma NO(3)(-) in diabetic subjects were significantly higher than those in nondiabetic subjects. Plasma NO(x) levels in neither diabetic nor nondiabetic subjects correlated with serum lipids, HbA1c, or IMT. In diabetic subjects, plasma NO(3)(-) levels were related not only to the presence of hypertension but also to advanced microvascular complications. Moreover, plasma NO(3)(-) levels were positively correlated with both serum lipid peroxide and AGEs. Multiple regression analysis revealed that serum AGEs level was strongly associated with plasma NO(3)(-) level. Thus, the findings are consistent with the hypothesis that decreased endothelium-dependent vasodilation in diabetic subjects is associated with the impaired action of NO secondary to its inactivation resulting from increased oxidative stress, rather than decreased NO production from vascular endothelium, and that abnormal NO metabolism is related to advanced diabetic microvascular complications.

Involvement of tyrosine kinase in citrate-stimulated aldosterone production in bovine glomerulosa cells.

The present study was designed to assess whether citrate stimulates aldosterone production by isolated bovine adrenal glomerulosa cells in vitro. When the cells were incubated with graded concentrations of citrate up to 4.0 mM, basal aldosterone production was significantly elevated, with a gradual reduction of extracellular ionized calcium concentration. Without citrate, however, adding increasing amounts of calcium chloride to a calcium-free medium did not reproduce the citrate's effect on basal aldosterone production. Genistein, an inhibitor of tyrosine kinases, inhibited the citrate (4 mM)-induced aldosterone production in a dose-dependent manner, with 89.8% of inhibition at a concentration of 10 microM. When the cells were exposed to citrate (4 mM) for 5, 10, and 30 min, tyrosine in Mr 105,000 endogenous protein was dominantly phosphorylated. This study demonstrates for the first time that citrate stimulates aldosterone production in bovine adrenal glomerulosa cells in vitro and also suggests a crucial involvement of protein tyrosine kinase in the steroidogenic action of citrate in the cells.

Treatment of renovascular hypertension using stent implantation in an elderly patient with NIDDM.

A 70-year-old man with NIDDM was diagnosed as having renovascular hypertension (RVH), based on a stenosis of the ostial portion of the left renal artery with markedly elevated plasma renin activity (PRA) in both the left renal vein and the peripheral blood, and positive captopril tests. After percutaneous transluminal renal angioplasty (PTRA), his blood pressure (BP) and PRA normalized. However, since restenosis occurred three months later, stent therapy was applied, and consequently BP and PRA normalized immediately after this procedure. During the one-year follow-up, side effects have not been noted. We propose that stent therapy may be feasible for ostial renal artery stenosis in elderly diabetic patients.

Reversed circadian blood pressure rhythm independently predicts endstage renal failure in non-insulin-dependent diabetes mellitus subjects.

To investigate the significance of reversed circadian blood pressure (BP) rhythm as a predictor for diabetic endstage renal failure, introduction of hemodialysis (HD) was determined as an end point in 325 noninsulin-dependent diabetes mellitus (NIDDM) subjects, in whom 24-h BPs had been monitored during their first admissions between 1988 and 1996. Circadian BP rhythm was analyzed by the COSINOR method, as previously reported. After exclusion of 68 dropout subjects, 257 were recruited for further analyses, in which 194 had normal circadian BP rhythms (N), and the remaining 63 had reversed rhythms (R). During this follow-up period, the numbers of HD-introduced subjects in N and R were 6 and 16, respectively, showing a higher prevalence in the latter (p < 0.001, chi2 test). Follow-up periods were significantly shorter in HD-introduced diabetic subjects of N and R than those in HD-free subjects of each group. In baseline characteristics, there were no differences in age, gender, or serum creatinine between HD-free and HD-introduced subjects of N or R. With regard to microvascular complications, the degree of retinopathy and nephropathy in N and R tended to be more pronounced in HD-introduced subjects than in HD-free subjects. Further, mean levels of circadian mean BP rhythms in HD-introduced subjects of N or R were similarly high, compared with those in HD-free subjects of each group, irrespective of circadian BP pattern. Unadjusted HD-free times were estimated by the Kaplan-Meier method, with a significant difference noted between N and R (p < 0.001; log-rank test). The Cox proportional-hazards model adjusted for circadian BP pattern, age, gender, blood pressure level, glycemic control, duration of diabetes, serum total protein, and serum creatinine demonstrated that circadian BP pattern, age, gender (female), blood pressure level (hypertension), and serum creatinine exhibited significant high relative risks. Thus, our data suggest that reversed circadian BP rhythm is an independent predictor of endstage renal failure in NIDDM subjects.

Mitochondrial DNA mutations are associated with both decreased insulin secretion and advanced microvascular complications in Japanese diabetic subjects.

To assess the roles of various mitochondrial (Mt) DNA mutations in diabetic and nondiabetic subjects, we screened Mt DNAs at the 3243 base pair (bp) and its adjacent portion in unrelated Japanese diabetic and nondiabetic subjects. Furthermore, to clarify the clinical features of diabetic subjects harboring a Mt DNA mutation, we evaluated the ability of insulin secretion and microvascular complications in diabetic subjects. Five hundred thirty-seven diabetic patients and 612 unrelated nondiabetic subjects were recruited into this study. In Mt DNA analyses, Mt DNA was isolated from peripheral leukocytes of the subjects, and then an Mt DNA fragment surrounding the tRNA(Leu(UUR)) site was amplified by the polymerase chain reaction (PCR) using two sets of primers. These fragments were further digested with three kinds of restriction endonucleases and were subjected to agarose gel electrophoresis. When a mutation was present, Mt DNA fragments were directly sequenced with an autosequencer. Baseline characteristics in all subjects were examined, and microvascular complications and insulin secretory capacity in diabetic subjects were newly evaluated. Eight kinds of Mt DNA mutations, which were point mutations, were found in 74 subjects. Each affected subject had only one mutation in the Mt DNA examined. Among them, the mutations at np 3316, 3394, 3593, and 3391 were accompanied by amino acid replacement. Thirty-eight diabetic patients were affected (7.1%), including two subjects with a point mutation at np 3243, and 26 nondiabetic subjects were affected (4.2%). Thus, there was a higher prevalence in diabetic subjects than in nondiabetic subjects. There was no significant difference in the prevalence of maternally inherited diabetes between these two groups. The mean level of urinary C-peptide excretion was lower in diabetic subjects with an Mt DNA mutation (DM+) than in those without it (DM-). Although the prevalence of hypertension in DM+ was higher than that in DM-, diabetic retinopathy and nephropathy in DM+ were problematic, in comparison with those in DM-, when statistical corrections were performed for the effect of hypertension. Furthermore, a strategy based on logistic regression analysis revealed that advanced retinopathy and decreased urinary C-peptide excretion in all diabetic subjects studied were strongly related to the presence of Mt DNA mutation. Our results suggest that Mt DNA mutations in Japanese diabetic subjects are related to the development of diabetes, and also that these mutations are associated with not only a decrease in insulin secretion but also advanced diabetic microvascular complications.

Hypertension due to coexisting pheochromocytoma and aldosterone-producing adrenal cortical adenoma.

A 49-year-old male was diagnosed as having primary aldosteronism at age 39, and he was treated with antihypertensive drugs. In 1995, a computed tomogram revealed a mass in the right adrenal gland. Radiological examinations and endocrinological data revealed the presence of a pheochromocytoma in the right and an adrenocortical tumor in the left adrenal gland. Right adrenalectomy and left partial adrenalectomy were performed. Histologically, the right adrenal mass was compatible with pheochromocytoma, and the left adrenal mass was an adrenocortical adenoma. Endocrinological data as well as blood pressure returned to normal after operation.

Reversed circadian blood pressure rhythm is associated with occurrences of both fatal and nonfatal vascular events in NIDDM subjects.

To assess the significance of reversed circadian blood pressure (BP) rhythms as a predictive factor of vascular events in NIDDM, vital status after an average 4-year follow-up was determined in 325 NIDDM subjects in whom the circadian BP profile had been monitored between 1988 and 1996. Circadian BP rhythm was analyzed by the COSINOR (a compound word for cosine and vector) method, as previously described. After exclusion of 37 dropped-out subjects, 288 were recruited to the further analysis, of which 201 had a normal circadian BP rhythm (group N) and the remaining 87 had a reversed one (group R). There was no difference in sex, HbA1c, the prevalence of smokers, serum lipids, or serum electrolytes between groups N and R at baseline, whereas age, the prevalence of hypertension, serum creatinine, and diabetic complications were more pronounced in group R than in group N. During the follow-up period (which averaged 52 months in group N and 36 months in group R), fatal and nonfatal vascular (cerebrovascular, cardiovascular, peripheral vascular arteries, and retinal artery) events occurred in 20 subjects in group N and 56 in group R. Unadjusted survival times and event-free times were estimated by the Kaplan-Meier product-limit method, and there was a significant difference in both unadjusted survival and event-free survival rates between groups N and R (P < 0.001 each; log-rank test). The Cox proportional-hazards model adjusted for age, sex, circadian BP pattern, duration of diabetes, therapy for diabetes, various diabetic complications, and hypertension demonstrated that circadian BP pattern and age exhibited significant, high adjusted relative risks for fatal events, and that diabetic nephropathy, postural hypotension, and hypertension as well as circadian BP pattern exhibited significant, high adjusted relative risks with respect to the occurrence of various nonfatal vascular events. These results suggest that reversed circadian BP rhythm is associated with occurrences of both fatal and nonfatal vascular events in NIDDM subjects.

Mitochondrial DNA point mutation at nucleotide pair 3316 in a Japanese family with heterogeneous phenotypes of diabetes.

A mitochondrial DNA (mtDNA) point mutation at nucleotide pair (np) 3316 has been reported in relation to diabetes. We recently encountered a non-obese family with this type of mutation. The proband in the affected family, a 49-year-old woman who had been previously diagnosed as having an insulin-requiring non-insulin-dependent diabetes mellitus (NIDDM), was referred to our hospital for treatment of diabetic gangrene in her left foot. Her insulin secretory capacity was markedly reduced, but the insulin sensitivity evaluated by the euglycemic hyperinsulinemic clamp technique was normal. In addition, her serum lactate level was markedly increased after a 5 min ambulation, although her serum pyruvate and ketones remained within the normal range. Twenty-year-old twin sons had been treated with insulin since the age of 7, when both were diagnosed with insulin-dependent diabetes mellitus (IDDM). The proband's mother, a 68-year-old, was nondiabetic at this time. MtDNA analysis revealed a point mutation at np 3316 in all family members, which was homoplasmic for the mutation on a photograph of agarose gel electrophoresis containing ethidium bromide under ultraviolet light. This mutation seemed to be maternally transmitted in the family, and the onset of diabetes was occurring earlier and the insulin secretory capacity was declining from generation to generation, so that these findings suggest that the point mutation at np 3316 is associated with various phenotypes of diabetes.

Potentiating effects of calcium chelators on basal and stimulated aldosterone production by bovine adrenal glomerulosa cells in vitro.

We previously reported the potentiation of basal aldosterone production by the addition of the calcium chelator ethyleneglycol-bis (beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) to an extracellular solution of bovine adrenal glomerulosa cells in vitro. To assess whether the addition of the calcium chelators ethylenediamine-N,N,N',N'-tetraacetic acid (EDTA) and EGTA can potentiate basal and stimulated aldosterone production, we compared the effect of EDTA with that of EGTA on basal and the agonist (potassium; 8 mM, angiotensin II; 10 nM, ACTH; 10 nM)-stimulated aldosterone production by the cells in vitro. These two chelators lowered the extracellular ionized calcium ([Ca2+]o) concentration in a similar manner. The levels of basal and the agonist-stimulated aldosterone production in the presence of EDTA (1 mM) and EGTA (1 mM) were significantly (P < 0.01 or less) increased when compared with those in the absence of EDTA and EGTA, respectively. These results show that the addition of EDTA and EGTA to an extracellular solution potentiates basal and the agonist-stimulated aldosterone production in vitro. Although an increase in basal aldosterone production in the presence of EDTA (1 mM) and EGTA (1 mM) was completely inhibited by the voltage-dependent calcium channel antagonist nifedipine (1 microM) or the calmodulin antagonist pimozide (1 microM), the potentiation of the agonist-stimulated aldosterone production does not seem to be induced by CA2+/calmodulin-dependent nor cAMP-dependent systems. These findings suggest that calcium chelators such as EDTA and EGTA may possess activating effect on basal and stimulated aldosterone production in bovine adrenal glomerulosa cells.

Enhancement of potassium-, and angiotensin II-stimulated aldosterone production by the calcium chelator EGTA in bovine adrenal glomerulosa cells in vitro.

The present study was designed to assess the effect of the calcium chelator EGTA (ethylenglycolbis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid; 1.0 mM) on potassium (8 mM)- and angiotensin II (AII; 10 nM)-stimulated aldosterone production in bovine adrenal glomerulosa cells in vitro. The combined administration of EGTA and potassium, or of EGTA and AII, yielded a significant increase in the levels of aldosterone production. The net increment in aldosterone production after the combined administration of EGTA and potassium, or that after the combined administration of EGTA and AII was also significantly higher than the sum of that evoked by EGTA and potassium alone, or the sum of that evoked by EGTA and AII alone, respectively. At similar concentrations of extracellular ionized calcium ([Ca2+]out) or magnesium, the levels of agonist-stimulated aldosterone production were markedly elevated by the administration of EGTA. These results indicate that lowering [Ca2+]out concentrations with EGTA enhances potassium- and AII-stimulated aldosterone production in bovine adrenal glomerulosa cells in vitro. This enhancement may be predominantly due to another effect of EGTA, in addition to the stimulation of calcium entry into the cells.

Rapidly progressive glomerulonephritis concomitant with diabetic nephropathy.

We describe a rare case of a rapidly progressive glomerulonephritis (RPGN) superimposed on diabetic nephropathy. A 68-year-old woman with non-insulin-dependent diabetes mellitus (NIDDM) complicated with diabetic triopathy demonstrated a rapid deterioration of renal function. Her urinary sediment contained many red blood cell (RBC) cells and casts, suggesting an additional renal disease accompanying diabetic nephropathy. Renal biopsy revealed crescent formation in many glomeruli characteristic of the pauci-immune type of RPGN. Steroid pulse therapy transiently halted the deterioration in renal function, but the patient died of pneumonia complicated with methicillin-resistant staphylococcus aureus (MRSA) infection. The unusual findings in diabetic nephropathy indicated the coexistence of primary glomerulonephritis and diabetic glomerulosclerosis in this case.

Effects of posture on the plasma hormonal and renal water-electrolyte excretory responses to acute water loading in diabetic subjects with hypoadrenergic orthostatic hypotension.

The effects of posture on the plasma hormonal and renal water-electrolyte excretory responses to acute water loading (20 mL/kg BW, orally) were studied in six non-insulin-dependent diabetes mellitus (NIDDM) subjects with hypoadrenergic orthostatic hypotension (HOH), eight NIDDM subjects without HOH, and seven nondiabetic subjects. The three groups were similar with respect to basal levels of mean blood pressure (MBP), serum sodium and osmolality, plasma renin activity (PRA), the plasma volume regulatory hormones alpha-atrial natriuretic peptide (ANP), arginine vasopressin (AVP) and aldosterone, and urinary water and sodium excretion. In the supine state, while allowing the subjects to stand only to void, water loading resulted in no changes in MBP and similar responses of these plasma and urinary parameters in the three groups. In the standing state, water loading produced responses of MBP, and plasma and urinary parameters comparable to those in the supine state in the diabetic group without HOH and the nondiabetic group. In the diabetic group with HOH, however, MBP and hourly urinary water and sodium excretion rates were low compared to those in the other two groups. During water loading, plasma ANP decreased, and, despite the fall of MBP, plasma AVP remained unchanged, and PRA and plasma aldosterone increased normally in the diabetic group with HOH. These results demonstrate that, in NIDDM subjects with HOH, changing from lying to standing induces deranged renal water and sodium handling after water loading, accompained by a decrease in plasma ANP, and inadequate responses of plasma AVP, PRA, and plasma aldosterone to hypotension.

Altered circadian blood pressure rhythm and progression of diabetic nephropathy in non-insulin dependent diabetes mellitus subjects: an average three year follow-up study.

BACKGROUND: In addition to autonomic dysfunction, diabetic nephropathy has been identified as another factor inducing a reversed circadian blood pressure (BP) rhythm in diabetic subjects. This study was carried out to assess the relationship between alterations in circadian BP rhythm and progression of diabetic nephropathy in subjects with non-insulin dependent diabetes mellitus (NIDDM). METHODS: Ambulatory 24-hour BP, 24-hour urinary albumin excretion rate (UAE), and plasma hormonal factors were measured during an average 3-year follow-up in 16 hospitalized subjects with NIDDM. Twelve age-matched control subjects were also studied. RESULTS: During an average 3-year follow-up, diabetic subjects had no significant progression of severe nephropathy and/or somatic neuropathy and showed no transition from a normal to a reversed mean blood pressure (mBP) pattern. However, mBP during whole day or nighttime, but not daytime, at baseline in diabetic subjects was high as compared with control subjects exhibiting an increased night/day mBP ratio and a decreased night/day mBP difference. The mBPs during various time periods (whole day, daytime, and nighttime) at follow-up in diabetic subjects were more elevated than those at baseline, showing a more increased night/day mBP ratio and a more decreased night/day mBP difference. In diabetic subjects, UAE during follow-up was increased, and UAE increments were well correlated with changes in mBP during whole day and nighttime. Plasma renin activity (PRA) and plasma aldosterone (PA) were decreased, while plasma alpha-atrial natriuretic peptide (ANP) was increased at follow-up, compared with at baseline. The mBP increments during various time periods were well correlated with changes in these hormonal factors, and UAE increments were well correlated with changes in PA and plasma ANP. CONCLUSIONS: The altered circadian BP rhythm observed in diabetic subjects may occur at the early stage of diabetic nephropathy with opposite changes in plasma renin-aldosterone and plasma ANP.

Painless thyroiditis occurring during long-term treatment with interferon alfa in a patient with chronic active hepatitis C.

We describe here painless thyroiditis during interferon (IFN) therapy in a 65-year-old man with chronic active hepatitis C. The patient had hypothyroidism in the late stage of a 24-week course of treatment with IFN-alpha. After cessation of the treatment a small, firm goiter was noticed, and chronic focal thyroiditis was diagnosed histologically. Analyses of the stock serum samples drawn before, during, and after IFN-alpha therapy revealed transient hyperthyroidism followed by transient hypothyroid states with aggravation of antithyroid hormone antibody titers. These findings suggest that long-term IFN-alpha therapy caused painless thyroiditis with aggravation of autoimmunity in our patient with preexisting chronic thyroiditis.

Hypertension and unilateral renal ischemia (Page kidney) due to compression of a retroperitoneal paraganglioma.

We describe a 17-year-old hypertensive man with a left abdominal mass on the computed tomographic scan. Radiologic studies revealed a large tumor near the left renal hilus, resulting in renal compression and displacement toward the posterolateral abdominal cavity. Aortography excluded stenosis of the renal arterial vessels. Plasma and urinary catecholamines were normal and plasma renin activity (PRA) was high. The left renal vein renin was 2.1-fold higher than the right one. After resecting the tumor including the left kidney and adrenal, high blood pressure and elevated PRA returned to normal. Histologically, the tumor was a paraganglioma, and the affected kidney showed hyperplasia of the juxtaglomerular apparatus. Thus, the hypertension in our patient was probably due to renal ischemia extrinsically compressed by a non-functioning retroperitoneal paraganglioma (Page kidney).

Non-obese Cushing's syndrome in an aged woman with non-insulin-dependent diabetes mellitus.

A 76-year-old diabetic woman with non-obese Cushing's syndrome developed poor glycemic control with glibenclamide. She presented with a slight weight loss while bedridden due to a fall. Cushing's syndrome in this patient was suspected because of hypercortisolemia with eosinopenia, and adrenal Cushing's syndrome was diagnosed by endocrine and radiological examinations. A right adrenal adenoma was confirmed by autopsy. In this patient, progressive obesity and other common features of Cushing's syndrome may have been concealed by aging itself and coexisting diabetes mellitus.

Effects of long-term cilazapril treatment on glucose and lipid metabolism in hypertensive patients with non-insulin-dependent diabetes mellitus.

The effects of long-term cilazapril treatment on glucose and lipid metabolism were assessed in 25 hypertensive patients with non-insulin-dependent diabetes mellitus (NIDDM). Patients were treated with 0.5 to 1 mg of cilazapril once daily or a combination of cilazapril and other antihypertensive drugs once daily for 48 weeks. Both systolic and diastolic blood pressures were significantly reduced (P < 0.001) throughout the study with no significant changes in heart rate and no adverse effects such as cough. There were no significant changes in body mass index or serum levels of glycated hemoglobin A1c, fructosamine, total cholesterol, triglycerides, lipoproteins (high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and very-low-density lipoprotein cholesterol), or apolipoproteins (apo A-I, apo C-II, apo C-III, apo B, and apo E). Cilazapril caused a significant increase (P < 0.05) in levels of apo A-II and a significant decrease (P < 0.05) in the apo B:apo A-I ratio, an index of arteriosclerosis. These results suggest that cilazapril has favorable effects on glucose and lipid metabolism and that it may be useful as the first or second choice of antihypertensive drugs in hypertensive patients with NIDDM.

Effect of 24 weeks of treatment with epalrestat, an aldose reductase inhibitor, on peripheral neuropathy in patients with non-insulin-dependent diabetes mellitus.

The effects of treatment with epalrestat, an aldose reductase inhibitor, on peripheral neuropathy were studied in 45 patients with non-insulin-dependent diabetes mellitus (NIDDM). Epalrestat 150 mg three times daily was given for 24 weeks. Subjective symptoms, such as spontaneous pain in the lower extremities and numbness and hypoesthesia of the extremities or trunk, were significantly (P < 0.001) relieved after 12 and 24 weeks of epalrestat treatment. Vibratory perception thresholds, as measured by using a tuning fork (C-128) and a vibrometer, were improved after 24 weeks of treatment. Furthermore, there were no adverse effects on glucose or lipid metabolism during treatment. These results suggest that long-term (24-week) epalrestat therapy can be used effectively to treat peripheral neuropathy in NIDDM patients without affecting glucose or lipid metabolism.