In African American type 2 diabetic patients, is vitamin D deficiency associated with lower blood levels of hydrogen sulfide and cyclic adenosine monophosphate, and elevated oxidative stress?

African Americans (AA) have a higher incidence of cardiovascular disease and vitamin D (VD) deficiency compared with Caucasians. Hydrogen sulfide (H(2)S) is an important signaling molecule. This study examined the hypothesis that blood levels of H(2)S are lower in AA type 2 diabetic patients (T2D). Fasting blood was obtained from T2D and healthy controls. Results showed a significant decrease in plasma levels of cyclic adenosine monophosphate (cAMP) and H(2)S in AA T2D but not in Caucasian T2D when compared with those of respective age- and race-matched healthy controls. Plasma VD levels were significantly lower in AA T2D compared with Caucasian T2D. Cell culture studies demonstrate that 1,25(OH)(2)-VD supplementation significantly increased expression of cystathionine-γ-lyase (CSE), H(2)S formation, and cAMP secretion, but decreased reactive oxygen species in high glucose-treated U937 monocytes. This suggests that VD supplementation upregulates CSE and H(2)S formation and decreases oxidative stress, and that VD deficiency may contribute to the malfunctioning of H(2)S signaling and thus a higher incidence of vascular inflammation in AA. These results lead to the hypothesis that VD supplementation can replenish blood concentrations of H(2)S and cAMP and lower oxidative stress and cardiovascular disease in AA T2D.

The effect of sleep apnea and insomnia on blood levels of leptin, insulin resistance, IP-10, and hydrogen sulfide in type 2 diabetic patients.

INTRODUCTION: Sleep deficits associated with sleep apnea and insomnia increase the risk of vascular inflammation and insulin resistance. This study examined the hypothesis that inflammation markers are higher in those diabetic patients who experience sleep deficits compared with those without any history of a sleep disorder. METHODS: Fasting blood was obtained after written informed consent, and sleep disorder histories were obtained from type 2 diabetic patients (n=81) attending clinics of the Louisiana State University Health Sciences Center. RESULTS: There was a significant correlation between body weight and leptin, and leptin in turn was significantly correlated with 10-kDa interferon-γ-induced protein (IP-10) levels and insulin resistance in type 2 diabetic patients. Fasting blood levels of leptin, IP-10, and insulin resistance were significantly elevated in patients with sleep deficits compared with diabetics with normal sleep patterns. There were no differences in glycosylated hemoglobin (HbA1c) or fasting glucose in patients with sleep deficits compared with those with normal sleep patterns. Sleep deficits increase circulating levels of leptin, IP-10, and insulin resistance compared to levels seen in patients with diabetes who reported no difficulty with sleep. Patients with sleep apnea had significantly lower hydrogen sulfide (H(2)S) levels compared with patients with normal sleep patterns or patients with insomnia. Low levels of circulating H(2)S could contribute to higher vascular inflammation in patients with sleep apnea. CONCLUSIONS: These results suggest that sleep apnea is associated with a decrease in circulating H(2)S and sleep disorders increase the risk of inflammation and insulin resistance, which can contribute to the increased risk of vascular disease in subjects with type 2 diabetes.

Effect of chromium dinicocysteinate supplementation on circulating levels of insulin, TNF-α, oxidative stress, and insulin resistance in type 2 diabetic subjects: randomized, double-blind, placebo-controlled study.

SCOPE: Chromium and cysteine supplementation have been shown to improve glucose metabolism in animal studies. This study examined the hypothesis that chromium dinicocysteinate (CDNC), a complex of chromium and l-cysteine, is beneficial in lowering oxidative stress, vascular inflammation, and glycemia in type 2 diabetic subjects. METHODS AND RESULTS: Type 2 diabetic subjects enrolled in this study were given placebo for 1 month for stabilization and then randomized into one of three groups: placebo (P), chromium picolinate (CP), or CDNC, after which they received daily oral supplementation for 3 months. Of the 100 patients enrolled in the study, 74 patients completed it. There were 25 patients in the P supplemented group, 25 in the CP supplemented and 24 in the CDNC supplemented group who completed the study. Blood markers of glycemia, vascular inflammation, HOMA insulin resistance, and oxidative stress were determined at randomization and after 3 months of supplementation with P, CP, or CDNC. There was a significant decrease at 3 months in insulin resistance (p = 0.02) and in the levels of protein oxidation (p = 0.02) and TNF-α (p = 0.01) in the CDNC supplemented cohort compared to baseline. However, there was no statistically significant change in these markers in the CP supplemented group compared to baseline. Insulin levels significantly decreased (p = 0.01) for subjects receiving CDNC but not CP. There was no significant impact of supplementation on HbA(1c) or glucose levels in either of the groups. CONCLUSION: CDNC supplementation lowers insulin resistance by reducing blood levels of TNF-α, insulin, and oxidative stress in type 2 diabetic subjects. Therefore, CDNC supplementation has potential as an adjunct therapy for individuals with type 2 diabetes.

Oxidative stress and autonomic nerve function in early type 1 diabetes.

INTRODUCTION: The biochemical mechanisms by which hyperglycemia causes microvascular disease and neuropathy are poorly understood. Experimental studies have established that oxidative stress is present in diabetic rodents with neuropathy, and that antioxidant therapy is protective. Oxidative stress is also present in human diabetes, but its clinical importance is uncertain. MATERIAL AND METHODS: We examined several biochemical measures of oxidative stress in 37 patients with recent-onset (less than 2 years) type 1 diabetes annually in a 3-year longitudinal study. We also performed a comprehensive annual evaluation of somatosensory and autonomic nerve function. A total of 41 control subjects were studied. RESULTS: Malondialdehyde excretion, a measure of lipid peroxidation, was 1.5l ± .1 µmol/g creatinine in the control subjects, but 2.43 ± . 3 in the diabetic patients in year one, 2.39 ± .2 in year two and 1.92 ± .15 in year three, which was different from controls across all years; p < .005. Serum NOx (nitrate and nitric) was 34.0 ± 4.9 µmol/L in the controls, but 52.4 ± 5 in the diabetics in year one, 50.0 ± 5.1 in year two, and 49.0 ± 5.2 in year three, which was different from controls; p < .01. We measured sudomotor function and observed that the poorly controlled diabetic patients had relatively increased sweating above the waist and relatively decreased sweating below the waist, a typical pattern for sympathetic nerve injury. The ratio of sweating above to sweating below the waist was .385 ± .04 in controls, 0.70 ± .14 in diabetic patients in year one, .51 ± .14 in year two and .496 ± .12 in year three (different from controls; p < .01 across all years). Urinary MDA correlated negatively with total sweat (r = -39, p < .01); NOx also correlated negatively with total sweat (r = -.34, p < .025). Abnormalities in the processing of renin (the renin/prorenin ratio), a test of renal sympathetic neurons, was also documented in early type 1 diabetes. CONCLUSIONS: Oxidative stress and excessive serum NOx are associated with sympathetic dysfunction in early type 1 diabetes.

Can diabetes I and early blindness be prevented using a tylenol combination which inhibits oxidative and nitrosative stress?

Since oxidative/nitrosative stress cause diabetes, can we prevent this chemistry generating the disease? Streptozotocin causes diabetes by entering the pancreatic beta cell generating excessive nitric oxide which reacts with oxygen creating a toxin possibly peroxynitrite, dinitrogen trioxide, dinitrogen tetraoxide and so forth. The toxic compounds damage the DNA causing beta cell death. This prevents insulin synthesis, storage and release. By using antioxidant substances that destroy the nitric-oxide-based toxins (e.g., carboxy-PTIO (oxidizes nitric oxide), polyphenolic-quercetin and monophenolic acetaminophen (Tylenol)) which are oxidation and nitration targets can the diabetes I causing toxins in animals be destroyed? Will this tri-drug combination completely prevent the deleterious effects of diabetes namely poor blood glucose control and blindness from cataracts for the entire length of the experiment (one year). These disease reversal experiments were accomplished in rats where the streptozotocin-diabetic effects were completely thwarted. In vitro experiments were accomplished to provide the scientific basis for the experimental results in animals.

Oxidative/nitrosative stresses trigger type I diabetes: preventable in streptozotocin rats and detectable in human disease.

Recently we demonstrated that streptozotocin (STZ) diabetes (type I) in rats is preventable using a simultaneous equimolar injection of carboxy-PTIO (c-PTIO). Both changes in blood sugar and cataracts are prevented. This apparently occurs because the nitric oxide (NO) (from STZ) generated in the beta cells is oxidized to nitrite by c-PTIO preventing diabetes. STZ generates NO producing a NO-based toxin. The toxin damages DNA by nicking and activates poly-ADP-ribose causing necrosis and triggering inflammation. Is there evidence that O/N stress occurs in early human type I diabetes? We studied 40 children with or without early type I diabetes and observed that urate is decreased 25% in all these diabetic children each over the age of 3 years. Urate is a major portion of blood-antioxidant load. Surely this decrease in urate indicates ongoing O/N stress. Does O/N stress initiate disease? STZ studies in rats indicates that this is correct.

Lipid peroxidation in early type 1 diabetes mellitus is unassociated with oxidative damage to DNA.

Oxidative stress damages DNA in experimental diabetes, and in vitro studies have suggested that it is linked to lipid peroxidation. The objective of the study was to determine whether lipid peroxidation, as assessed with malondialdehyde excretion in recent-onset type 1 diabetes mellitus, is associated with oxidative damage to DNA, as assessed from 8-hydroxydeoxyguanosine excretion. A 3-year longitudinal study of recent-onset type 1 diabetes mellitus was performed. Age- and sex-matched control subjects were studied once. Patients were studied as inpatients at West Virginia University Hospitals. Thirty-seven patients with recent-onset (2-22 months) type 1 diabetes mellitus (male, 10; female, 27) were enrolled in a longitudinal study of oxidative stress. The mean age of the patients was 20 years. None of the patients had hyperlipidemia or were treated with lipid-lowering drugs. Only 1 patient had hypertension and was being treated with beta-adrenergic blocking therapy. Thirty-six patients completed the study; one withdrew after the second evaluation. Lipid peroxidation was assessed by measuring malondialdehyde excretion. Oxidative damage to DNA was assessed from 8-hydroxydeoxyguanosine excretion. Malondialdehyde excretion was increased in the diabetic patients at the first evaluation (2.43 +/- 0.31 micromol/g creatinine), second evaluation (2.34 +/- 0.24), and third evaluation (1.93 +/- 0.15) compared with control subjects (1.51 +/- 0.11) (P < .005). 8-Hydroxydeoxyguanosine excretion, however, was not increased in the diabetic patients. There was no correlation between malondialdehyde and 8-hydroxydeoxyguanosine excretion. We confirmed the presence of oxidative stress in early diabetes as assessed from malondialdehyde excretion. We were unable, however, to confirm oxidative damage to DNA in this cohort of patients; and there was no evidence of a correlation between lipid peroxidation and DNA damage.

Effect of octreotide on postmenopausal hot flushes.

We have previously documented that octreotide therapy suppresses sweating and palpitations in patients with the postural tachycardia syndrome. We now report that octreotide also suppresses these and related symptoms in patients with postmenopausal hot flushes.

Luminescence experiments involved in the mechanism of streptozotocin diabetes and cataract formation.

Streptozotocin (STZ)-induced diabetes is linked to excessive nitric oxide (NO), and possibly peroxynitrite (OONO(-)) and/or other nitrogen oxides, e.g. nitrogen trioxide (N(2)O(3)), which damages DNA of pancreatic beta cells, causing death and loss of insulin. Simultaneous injection of carboxy-PTIO (CPTIO) and STZ prevents diabetes and cataract formation in rats, whereas 4-hydroxy-Tempo (4HT) does not. CPTIO oxidizes nitric oxide to nitrite, which prevents production of the diabetogenic toxin. Peroxynitrite may not be involved, since 4HT (converts O(2)(-) to H(2)O(2)) injected with STZ produces diabetes. All six of the control rats injected with STZ became diabetic and developed cataracts after 3 months. Eight rats injected with STZ and CPTIO were non-diabetic with no cataracts up to a year. This work establishes the idea that excessive nitric oxide is a primary initiator in STZ diabetes. Luminescence experiments using OONO(-) generation from SIN-1 with L-012 indicates that 4HT is an effective inhibitor, while CPTIO is ineffective. Experiments with dilute solutions of nitrogen trioxide added to ladder or plasmid DNA reveal extensive nicking of DNA, thereby raising the possibility that other oxides of nitrogen could be involved with the damage to DNA. It can be concluded that diabetes can be prevented by oxidizing excessive NO from STZ.

Treatment of autonomic neuropathy, postural tachycardia and orthostatic syncope with octreotide LAR.

The purpose of this study was to determine whether autonomic neuropathy and the postural tachycardia syndrome can be treated with octreotide LAR (Long Acting Release). This was an open-label pilot project. Protocol 1 Patients with autonomic neuropathy (n = 4) were given increasing doses of octreotide LAR once a month for three months. Blood pressure was measured in the sitting posture every two weeks. Pretreatment mean blood pressure averaged 83.8 +/- 7.1 mm Hg. After four, six and eight weeks of therapy the blood pressures averaged 96.3 +/- 6.4, 98.2 +/- 6.1 (p < .025), and 104.1 +/- 3.1 (p < .025) respectively. Therapy led to a dramatic improvement in symptoms in one patient but another had an unacceptable elevation in supine blood pressure. Protocol 2 Patients with POTS or orthostatic intolerance were given 10, 20, or 30 mg of octreotide LAR over three months. Seven patients entered and five completed the study. After two months treatment, standing time increased from 36.0 +/- 9.2 to 59.2 +/- .8 minutes (p < .01). Heart rate in the standing position was suppressed from 106 +/- .83 to 93.2 +/- .8 beats per minute (p < .05). Orthostatic dizziness and chronic fatigue improved. We conclude that octreotide LAR can be used to treat autonomic neuropathy but there is a risk of an excessive pressor response. Octreotide LAR improved standing time and suppressed tachycardia in patients with orthostatic intolerance.

Antibodies to GAD65 and peripheral nerve function in the DCCT.

Antibodies to the smaller isoform of glutamic acid decarboxylase (GAD65Ab) have been linked to the presence of neuropathy in Type 1 diabetes in several small studies. We attempted to confirm this association by measuring GAD65Ab, GAD65Ab epitopes and IA-2Ab in 511 patients who participated in the Diabetes Control and Complications Trial (DCCT). We also tested for correlations between these autoantibodies and C-peptide and glycemic control. We only included patients for whom serum was available from the first 4 years of their illness. The presence or absence of neuropathy was determined by electrophysiological studies, autonomic testing and clinical evaluation at baseline and 5 years into the trial or at close out. Samples from controls (patients without neuropathy at 5 years) were selected for patients who had similar C-peptide responses to a standardized meal at baseline. The GAD65Ab index correlated with HgbA(1c) only in the adult participants and only at baseline. The adults initially in poor control (upper tertile for glycemia) had higher GAD65Ab and lower C-peptides. The GAD65Ab index was not significantly different in patients with confirmed clinical neuropathy at 5 years versus controls matched for C-peptide (.248+/-.03 versus .278+/-.03). Epitope analysis, based on the blocking of conformational epitopes by recombinant Fab, revealed that the binding to multiple epitopes was decreased in the patients with neuropathy.

Treatment of postural tachycardia syndrome: a comparison of octreotide and midodrine.

We assessed the potency of octreotide and midodrine, and their combination, in the treatment of the postural tachycardia syndrome (POTS) and orthostatic intolerance (OI). Nine patients with POTS and six patients with OI stood for up to 1 hour while their HR and BP were monitored. Patients received on separate days, midodrine 10 mg 1 hour before testing, octreotide 0.9 micro g/kg 8 minutes before testing or combination therapy. Standing time in the patients with POTS was 41.2 +/- 8.4 minutes and not improved by midodrine or octreotide, but increased to 56.3 +/- 2.7 (P < 0.01) minutes following combination therapy. The standing heart rate in POTS, 114 +/- 0.7 bpm, was suppressed by midodrine 92.8 +/- 0.7 (P < 0.001), octreotide 90.6 +/- 0.78 (P < 0.001), and combination therapy 84.7 +/- 0.7 (P < 0.001). Combination therapy was better than monotherapy (P < 0.001) but only for the first 10 minutes of standing. Standing time of 36.3 +/- 3.5 minutes in the patients with OI improved with midodrine, octreotide and combination therapy (55.5 +/- 3.1, 56.5 +/- 3.5, and 56.6 +/- 3.3, respectively, P < 0.05 for each). Standing heart rate in OI was 100 +/- .76 bpm; following midodrine it was 80.3 +/- .69 (P < 0.05), following octreotide it was 84.8 +/- .86, and following combination therapy it was 71.2 +/- .9 (P < 0.01). The RR interval versus time area under the curve (The Orthostatic Index) was 21.1 +/- 4 in patients with OI. After midodrine it was 41.4 +/- 3.5 (P < 0.01), after octreotide 40.3 +/- 3.8 (P < 0.01) and after the combination it was 47.3 +/- 4.6 (P < 0.001). Midodrine and octreotide suppressed tachycardia in POTS and improved standing times in OI. The two drugs had similar potencies; combination therapy was not significantly better than monotherapy.

Sympathetic sudomotor disturbance in early type 1 diabetes mellitus is linked to lipid peroxidation.

The present study was performed to determine whether increased lipid peroxidation, as assessed from malondialdehyde (MDA) excretion, is associated with deterioration in peripheral nerve function in early type 1 diabetes mellitus. These parameters were measured annually for 3 years in 36 patients who entered the study less than 2 years after the diagnosis of diabetes. Malondialdehyde excretion was 1.51 +/- 0.20 micromol/g creatinine in the controls, and 2.43 +/- 0.21, 2.39 +/- 0.22, and 1.93 +/- 0.21 micromol/g creatinine at the first, second, and third evaluations, respectively (P < .005). The increased MDA was seen only in the female participants. Malondialdehyde excretion was increased in those with high vs low hemoglobin Alc across all years (P < .05). Malondialdehyde excretion correlated negatively with sudomotor function below the waist. The mean sweat production from the 3 evaluations correlated with mean MDA excretion across all years in the proximal leg (r = -0.42, P < .005) and distal leg (r = -0.40, P < .01). Below the waist, sweating correlated with MDA (r = -0.40, P < .01) as did total sweat (r = -0.38, P < .01). The response amplitudes of the peroneal nerves correlated negatively with MDA excretion (for the mean values at the second 2 evaluations, P < .005, r = -0.45). Tests of sensory function correlated inconsistently with MDA excretion. In summary, lipid peroxidation, as assessed from malondialdehyde excretion, is associated with sudomotor dysfunction in early diabetes.

Nitrosative stress in early Type 1 diabetes. David H. P. Streeten Memorial Lecture.

Although hyperglycemia has been shown to cause peripheral nerve dysfunction in patients with diabetes, the biochemical mechanisms for this effect are poorly understood. The excessive production of reactive oxygen species and reactive nitrogen species has been proven to be detrimental in experimental diabetes, but there is little evidence that these metabolic events take place clinically and are physiologically important in man. To assess this we measured nitrite and nitrate (indices of nitric oxide production), nitrotyrosine (an index of peroxynitrite), 8-isoprostaglandin F-2 alpha, an isoprostane reflective of oxidative stress and lipid peroxidation, and uric acid, an index of antioxidant defense in patients with recently diagnosed Type 1 diabetes and aged-matched controls. The diabetic patients were followed for three years. We documented the overproduction of nitric oxide and increased lipid peroxidation in early diabetes and showed these changes had detectable adverse effects on peripheral nerve function especially sympathetic sudomotor nerves. We documented the suppression of uric acid and showed this was associated with multiple abnormalities in autonomic function. In addition, we present indirect evidence that overproduction of reactive oxygen species and reactive nitrogen species have adverse effects on beta cell function and blood pressure.

Peroxynitrite versus nitric oxide in early diabetes.

BACKGROUND: Peroxynitrite is a toxic compound formed during the inactivation of nitric oxide (NO) by the superoxide anion. The physiologic significance of this pathway of NO metabolism has never been documented in vivo. Because peroxynitrite provides a pathway for the inactivation of NO we postulated that peroxynitrite's correlation with physiologic parameters would be the opposite of those associated with NO, which is a vasodilator and suppresses sudomotor function. We assessed the significance of peroxynitrite by comparing its associations with blood pressure (BP) and sudomotor responses with those of NO. METHODS: Thirty-seven patients with type 1 diabetes enrolled in a longitudinal study of oxidative stress. Nitric oxide was assessed from nitrite and nitrate (collectively NOx) and peroxynitrite was assessed from the nitrotyrosine (nTy) content of protein. METHODS: nTy was 13.3 +/- 2.0 micromol/L in the control subjects and 26.8 +/- 4.4 micromol/L, 26.1 +/- 4.3 micromol/L, and 32.7 +/- 4.3 micromol/L in the diabetic patients (P <.01) at the time of the first, second, and third evaluations, respectively. Patients with increased nitrotyrosine/tyrosine (nTy/Ty) had higher mean BP than those with low nTy/Ty (81.1 +/- l.9 mm Hg v 75.5 +/- 1.7 at the third evaluation, P <.025). The ratio of nTy/NOx correlated with BP at the first (P <.05), second (P <.05), and third (P <.01) evaluations. Patients with high nTy/Ty had increased sudomotor responses (5.85 +/- 0.75 microL of total sweat) at the third evaluation compared to those with low nTy/Ty (3.32 +/- 0.43 microL, P <.005) and normal controls (3.90 +/- 0.41 microL, P <.05). The associations of nTy with BP and sudomotor responses were the opposite of those with NOx. CONCLUSIONS: The conversion of NO oxide to peroxynitrite is physiologically significant in humans.

Oxidative stress and insulin requirements in patients with recent-onset type 1 diabetes.

The purpose of this study was to analyze biochemical measures of oxidative stress and assess their relationship to insulin requirements early in type 1 diabetes. Thirty-seven patients enrolled in a 3-yr longitudinal study of the effects of oxidative stress on the early natural history of this disorder. We measured plasma nitrite and nitrate (collectively NOx), nitrotyrosine, and 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2alpha)). Plasma NOx was 34.0 +/- 4.9 micro mol/liter in the control subjects and 52.4 +/- 5.1, 50.0 +/- 5.1, and 49.0 +/- 5.2 micro mol/liter in the diabetic patients at the first, second, and third evaluations, respectively (P < 0.01). Nitrotyrosine was 13.3 +/- 2.0 micro mol/liter in controls and 26.8 +/- 4.4, 26.1 +/- 4.3, and 32.7 +/- 4.3 micro mol/liter in the diabetic patients (P < 0.01). 8-Iso-PGF(2alpha) was higher in the poorly controlled than in the well controlled patients. NOx correlated with insulin dose at the first (P < 0.05), second (P < 0.025), and third (P < 0.05) evaluations. 8-Iso-PGF(2alpha) correlated with insulin dose at the first (P < 0.01) and third (P < 0.0025) evaluations. Systemic measures of oxidative stress correlate with insulin requirements in early type 1 diabetes. These results suggest that oxidative stress is taking place in the pancreas and damaging the beta-cell.

Nitrosative stress, uric Acid, and peripheral nerve function in early type 1 diabetes.

The present study was performed to determine whether nitric oxide overproduction is associated with deterioration in peripheral nerve function in type 1 diabetes. We measured peripheral nerve function and biochemical indicators of nitrosative stress annually for 3 years in 37 patients with type 1 diabetes. Plasma nitrite and nitrate (collectively NO(x)) were 34.0 +/- 4.9 micro mol/l in the control subjects and 52.4 +/- 5.1, 50.0 +/- 5.1, and 49.0 +/- 5.2 in the diabetic patients at the first, second, and third evaluations, respectively (P < 0.01). Nitrotyrosine (NTY) was 13.3 +/- 2.0 micro mol/l in the control subjects and 26.8 +/- 4.4, 26.1 +/- 4.3, and 32.7 +/- 4.3 in the diabetic patients (P < 0.01). Uric acid was suppressed by 20% in the diabetic patients (P < 0.001). Composite motor nerve conduction velocity for the median, ulnar, and peroneal nerves was decreased in patients with high versus low NTY (mean Z score -0.522 +/- 0.25 versus 0.273 +/- 0.22; P < 0.025). Patients with high NO(x) had decreased sweating, and those with suppressed uric acid had decreased autonomic function. In conclusion, nitrosative stress in early diabetes is associated with suppressed uric acid and deterioration in peripheral nerve function.