Angiotensin-converting enzyme insertion/deletion polymorphism and polyneuropathy in type 2 diabetes without macroalbuminuria.

Angiotensin-converting enzyme (ACE) gene polymorphism is thought to be a potent risk factor for nephropathy and retinopathy in diabetes. We investigated the association between polyneuropathy and gene polymorphisms of both the ACE insertion/deletion (I/D) and angiotensinogen (AGT) M235T genes in 84 type 2 diabetic patients without macroalbuminuria (21 with polyneuropathy and 63 without). ACE genotype distribution did not differ significantly between patients with and without polyneuropathy, but the frequency of the I allele was significantly higher in those with polyneuropathy than in those without. In contrast, neither the genotype distribution nor the allele frequencies of the AGT gene differed between the two groups. In logistic regression analysis using a D-additive model, the D allele had a protective effect on polyneuropathy (odds ratio [OR], 0.34; 95% confidence interval [CI], 0.13-0.88). A D-dominant model hypothesis also gave a significant OR (0.28; 95% CI, 0.09-0.90). ACE I/D polymorphism, but not AGT M235T polymorphism, may affect polyneuropathy development in type 2 diabetes without macroalbuminuria.

Moderate-intensity regular exercise decreases serum tumor necrosis factor-alpha and HbA1c levels in healthy women.

OBJECTIVE: To evaluate the effects of moderate-intensity regular exercise on serum levels of tumor necrosis factor-alpha (TNF-alpha) and glucose and lipid metabolism parameters. DESIGN: Longitudinal intervention study of a 5 month exercise training program (30-45 min/day, 4-5 days/week). SUBJECTS: Forty-one healthy Japanese women aged 41-69 y at baseline; 27 participants in the exercise program. MEASUREMENTS: Body mass index (BMI), waist-to-hip ratio (WHR), percentage body fat, and fasting levels for serum TNF-alpha, serum soluble TNF receptor p55 (TNF-RI) and TNF receptor p75 (TNF-RII), serum lipids, HbA1c, and serum insulin before and after exercise. RESULTS: In overweight to obese subjects, serum levels of TNF-alpha, TNF-RI and TNF-RII were significantly higher than those in lean subjects. There were significant correlations between log serum TNF-alpha and BMI, percentage body fat, WHR, HbA1c and log insulin. TNF-RI was significantly correlated with BMI, percentage body fat, WHR and HbA1c. TNF-RII was also associated with BMI, percentage body fat and WHR. However, the correlation between TNF-RII and HbA1c did not reach statistical significance. Neither TNF-RI nor TNF-RII was correlated with log insulin. In contrast, TNF-alpha, TNF-RI and TNF-RII were negatively correlated with HDL cholesterol. Regular exercise decreased BMI, percentage body fat, HbA1c, serum TNF-alpha, TNF-RI and TNF-RII and increased HDL cholesterol levels. In addition, exercise-induced change in serum TNF-alpha was independently correlated with changes in HbA1c and serum insulin, after being adjusted for the change in fat-free mass. CONCLUSION: Changes in serum TNF-alpha that occur with exercise may play an important role in improving glucose metabolism parameters.

Reciprocal changes in leptin and tumor necrosis factor-alpha with exercise in insulin resistant rats.

Leptin and tumor necrosis factor--alpha(TNF-alpha) are important mediators of insulin resistance in obese subjects through their over-expression in adipose tissue. Secretion of leptin into the circulation is a signal for the brain in patients with hyperinsulinemia. Regulation of TNF-alpha affects adipocyte insulin sensitivity and lipid accumulation. Exercise training has been suggested for the prevention and treatment of such disorders. However, how exercise modifies secretion of leptin and TNF-alpha is not known. We investigated leptin and TNF-alpha in a rat model of insulin resistance induced by sucrose. After 4 weeks of sucrose feeding, 4 weeks of voluntary wheel running significantly reduced the concentrations of leptin in mesenteric (MS) and subcutaneous fat (SC) when compared to sedentary sucrose-feeding rats. These results suggest that exercise controls cytokine regulation of leptin and TNF-alpha. The increased TNF-alpha in adipose tissue may activate cytolysis for energy consumption.

Positive correlation between circulating leptin levels and lipid lipoproteins in postmenopausal women administered hormone replacement therapy.

Beneficial effects of hormone replacement therapy are reported on plasma concentrations of lipids and lipoproteins. Plasma leptin levels are reported to reflect lipid metabolism. We treated 40 healthy postmenopausal women with continuous combined HRT (0.625 mg conjugated equine estrogen and 2.5 mg medroxyprogesterone acetate orally) daily for 6 months and then investigated total cholesterol, triglyceride, high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol, which are considered to be factors inducing cardiovascular disease (CVD). We measured the plasma levels of lipids and leptin, which are considered to be altered with HRT. Plasma leptin and lipid levels were measured at baseline and after 3 and 6 months of HRT. The plasma levels of leptin in PMW were not significantly reduced by HRT. Although the plasma levels of total cholesterol and LDL-cholesterol did not change by HRT, the administration of HRT significantly increased plasma levels of triglycerides and HDL-cholesterol in PMW. The correlation between leptin and total cholesterol or triglycerides was positive after HRT while these relations were not correlated at baseline. The correlation between lipid levels and leptin may explain the new role of leptin in plasma lipid levels in HRT.

Reduction of leptin precedes fat loss from running exercise in insulin-resistant rats.

Serum concentrations of leptin, a hormone secreted into the circulation by adipocytes, correlate with body mass index. Circulating of leptin is thought to signal the brain in patients with hyperinsulinemia, a condition reported to be preventable and testable by exercise training. In the present experiments, sucrose-fed rats had reduced concentrations of leptin in portal venous blood after 4 weeks of nonforced wheel-running exercise (1.1 +/- 0.1 vs. 6.2 +/- 1.8 ng/mL, in nonexercised sucrose-fed rats, P < 0.05). Mesenteric and subcutaneous fat stores were similar between groups. After 12 weeks of exercise, portal vein levels of leptin concentrations (5.2 +/- 2.1 vs. 9.9 +/- 0.8 ng/mL, P < 0.05) and mesenteric and subcutaneous fat all were reduced in the exercise group. These results suggest that short-term running exercise reduces circulating leptin before any reduction of adipose mass, and this reduction in the concentration of leptin available to its receptors has beneficial effects on the metabolism of fat and carbohydrates.

Running exercise increases tumor necrosis factor-alpha secreting from mesenteric fat in insulin-resistant rats.

Tumor necrosis factor-alpha (TNF-alpha) is an important mediator of insulin resistance in obese subjects, through its overexpression in fat tissue. However, how exercise can modify the expression of TNF-alpha is controversial. We examined TNF-alpha in adipose tissue using an animal model of insulin resistance that was produced by feeding rats a diet high in sucrose. The rats were allocated to one of three groups: those receiving a starch-based diet (control group): those fed a high-sucrose diet (sucrose-fed group): and those fed a high-sucrose diet and given wheel exercise (exercised group). The animals were allowed to eat and drink ad lib for 4 or 12 weeks (4 wk: control n=7, sucrose-fed n=7, exercised n=10; 12 wk: control n=5, sucrose-fed n=5, exercised n=9). The voluntary wheel exercise was initiated with the feeding of the high-sucrose diet. The rats in the exercise groups ran 15 +/- 3 km/week. We showed that 12-week voluntary running exercise significantly (P<0.05) increased both TNF-alpha protein (5-fold) and mRNA (1.4 fold) in the mesenteric fat of insulin-resistant rats compared to non-exercised sucrose-fed mice. Accordingly, in exercised group, plasma glucose (124 +/- 9 mEq/L vs 141 +/- 11 mEq/L). and free fatty acid (0.98 +/- 0.07 mEq/L vs 1.4 +/- 0.05 mEq/L) concentrating in portal vein blood were reduced compared to sucrose-fed group. The amounts of fatty tissue both in mesenteric and subcutaneous tissues were significantly (P<0.05) decreased through running exercise. We consider that up-regulation of TNF-alpha in mesenteric fat may be a compensatory mechanism for the reduction of fatty acid in adipose tissues and this change could control metabolic homeostasis during exercise to modulate a hyperinsulinemic state.

Combination of OK432 and human interferon-alpha for treating viral-induced diabetes mellitus in mice.

We investigated the therapeutic effects of OK432 (picibanil; CAS39325-1-4), an immunomodulator that is derived from the Su strain of Streptococcus pyogenes. This agent was administered alone or combined with human interferon-alpha in a murine model of insulin-dependent diabetes mellitus. Interferon-alpha inhibits viral replication, reducing the incidence of virus-induced IDDM. Groups of DBA/2 mice (N = 25 per group) received an intraperitoneal injection of OK432 and interferon-alpha daily for 16 d beginning 1 d after inoculation with 500 plaque-forming units of encephalomyocarditis virus (EMCV). The dose of OK432 was one clinical unit (corresponding to 0.1 mg dried cells) per mouse, and that of interferon-alpha was 1 x 10(4) u/g. The animals were killed at random at 3 or 7 d after inoculation with EMCV. The survival rate of mice treated with the combination of OK432 and with interferon-alpha was significantly greater than that of the non-treated infected control animals (P < 0.01). Fasting levels of blood glucose were significantly lower in the mice administered the combination, than in the controls, both on day 3 (68 +/- 21 mg/dl vs. 270 +/- 135 mg/dl, P < 0.01) and on day 7 (101 +/- 29 mg/dl vs. 219 +/- 112 mg/dl, P < 0.01). Serum levels of insulin were significantly higher in the treated mice than in the controls (65 +/- 5 vs. 55 +/- 1 microU/ml, P < 0.05). However, in the mice treated with OK432 or interferon-alpha alone, the survival rate and the blood level of glucose and insulin did not differ from those of infected controls. Natural killer (NK) cell activity was significantly higher in the mice treated with the drug combination than in the controls on both days evaluated: day 3, 65 +/- 5 vs. 55 +/- 1%, n = 3, P < 0.05; day 7, 44 +/- 3 vs. 22 +/- 8%, n = 3, P < 0.05). Serum levels of murine interferon in the treated mice exceeded those in controls on both days evaluated (day 3, 671 U/ml vs. 442 U/ml; day 7, 57 U/ml vs. 43 U/ml). There were no significant differences in NK cell activity or in the interferon level in mice treated with either OK432 or interferon-alpha alone as compared with the infected, non-treated controls. Results suggest that the combination of OK432 and interferon-alpha protects against virally induced IDDM by increasing the activity of NK cells as well as the plasma level of interferon.

[Use of casual plasma glucose levels in community screening for diabetes mellitus].

The aim of this study was to assess the utility of casual plasma glucose as a screening test for diabetes mellitus in lieu of hemoglobin A1c (HbA1c) and urinary glucose. Casual plasma glucose and urinary glucose were measured in 1,024 individuals in a rural community. The individuals were then offered participation in an oral glucose tolerance test (OGTT). A 75 g OGTT based on WHO criteria was performed and HbA1c was measured in 290 respondents. Receiver operating characteristic (ROC) curves for casual plasma glucose and HbA1c were constructed and the areas under ROC curves for the tests were calculated. The area under the ROC curve for HbA1c was significantly higher than that for casual plasma glucose (p < 0.01). The sensitivity and specificity for casual plasma glucose > 140 mg/d/were 61% and 62%, respectively, while the specificity of HbA1c was 93% with a sensitivity of 61%. The sensitivity of urinary glucose was low (only 18%) with a comparable specificity (93%). It is concluded that measurement of casual plasma glucose is inferior to HbA1c as a screening test for diabetes.

Hepatic embolization through extrahepatic collateral pathways after hepatic arterial embolization for the hepatocellular carcinoma.

Extrahepatic collateral pathways developing after repeated transcatheter arterial embolization (TAE) for hepatocellular carcinoma (HCC) make therapeutic arterial embolization for recurrent lesions extremely difficult. TAE was performed through the collateral pathways using a sophisticated micro-catheter with good trackability and pushability and a coaxial system. Twenty-three TAEs were undertaken through the collateral pathways in 13 patients with recurrent HCC who had extrahepatic collateral pathways after the previous hepatic arterial TAE. There were 69 extrahepatic collateral pathways, with partially obstructed hepatic arteries. On the average, three feeding arteries were seen in the liver. The main extrahepatic collateral pathways were the inferior phrenic artery and epicholedocal artery, 18 vessels and 29 vessels, respectively, accounting for about 80% of the total collateral pathways. TAEs were successful in all cases and the number of embolized vessels was 2.1 on average. The average time of the first collateral TAE after the initial conventional hepatic arterial TAE was 2.3 years. Excellent prognosis was observed with a one-year survival rate of 77% and 3-year survival rate of 38% after the collateral TAE. We consider that collateral TAE for recurrent HCC with obstruction of the hepatic artery is the procedure of choice, is technically feasible, and provides better prognosis for the patients.

Cloning of a gene from Escherichia coli that confers resistance to fosmidomycin as a consequence of amplification.

A gene conferring resistance to fosmidomycin (Fs) was cloned from the gene pool of a wild-type strain of Escherichia coli. The cloned DNA fragment was sequenced and shown to encode a putative polypeptide of 406 amino acids (aa) with a molecular weight of 43303. The gene mapped at 10.9 min on the E. coli chromosome and was designated fsr (fosmidomycin resistance). Maxicell analysis revealed that the Fsr protein migrated in sodium dodecyl sulfate-polyacrylamide-gel electrophoresis as a broad band of 35 kDa. A comparison between the aa sequence of Fsr and sequences in a protein database revealed 18% homology to the bacterial drug-export proteins that mediate resistance to tetracycline and chloramphenicol. Hydropathy analysis of the Fsr protein revealed twelve putative transmembrane segments. The degree of FsR of transformants depended on the number of copies of the plasmid that contained fsr. The levels of ubiquinone-8 and undecaprenyl phosphate in cells that harbored a high-copy-number plasmid that included fsr were almost the same as those in the cells without the plasmid. These results suggest that Fsr does not have any direct effect on the biosynthesis of isoprenoid in E. coli, and that the mechanism for FsR involves the efflux of the drug by a process that is facilitated by Fsr.

Decreased serum 1,5-anhydroglucitol in nondiabetic subjects with a family history of NIDDM.

OBJECTIVE: To investigate the effect of a family history of NIDDM on HbAlc and serum 1,5-anhydroglucitol (AG) in nondiabetic subjects. RESEARCH DESIGN AND METHODS: A 75-g oral glucose tolerance test was performed; 258 subjects with normal glucose tolerance and 106 subjects with impaired glucose tolerance (IGT) were selected HbAlc and serum AG were compared between subjects with and without a family history of NIDDM. The relationships between age, BMI, HbAlc, serum AG, fasting and 2-h plasma glucose, and urinary glucose were also examined using principal component analysis with a varimax rotation. RESULTS: In the normal group, only serum AG was lower in subjects with a positive family history than in those with no family history. On the other hand, in the IGT group, subjects with a positive family history were younger and had a higher 2-h plasma glucose, a higher urinary glucose, and a lower serum AG than those with no family history, whereas there was no difference in HbAlc. Principal component analysis identified three factors. The first factor, a linear combination of HbAlc and fasting plasma glucose, was labeled an average glycemic factor. The second factor, which included serum AG, 2-h plasma glucose, and urinary glucose, was labeled an oscillatory glycemic factor. The third factor, which contrasted age against BMI, was labeled an environmental factor. CONCLUSIONS: Serum AG is related to glycosuria even among nondiabetic subjects, and its concentrations are decreased in those with a family history of NIDDM. Our results suggest that serum AG rather than HbAlc reflects early metabolic abnormalities in these subjects.

Effects of age and obesity on glycated haemoglobin and 1,5-anhydroglucitol in screening for type 2 diabetes mellitus.

The diagnostic accuracy of glycated haemoglobin (HbA1c) and 1,5-anhydroglucitol (AG) and their association with age and obesity in the community screening for Type 2 diabetes were investigated. HbA1c and AG were measured in 368 subjects at the time of an oral glucose tolerance test (OGTT) based on WHO criteria. The impact of age and obesity on the probability of diabetes was evaluated by multivariate logistic regression analysis. Receiver operating characteristic (ROC) analysis was used to assess diagnostic accuracy of each method. The areas under ROC curves were compared for subjects aged 30-59 and 60-82 years. In the logistic analysis, age was associated with the probability of diabetes, while body mass index did not contribute to the model. The paired ROC analyses indicated that HbA1c was superior to AG only in the younger subjects. When the areas for two age groups were compared, both HbA1c and AG performed better in the younger than older subjects. We conclude that for the younger subjects, HbA1c value is highly sensitive (100.0%) and specific (90.4%) and is helpful for the diagnosis of Type 2 diabetes. On the other hand for the older subjects, both HbA1c and AG are poor screening tests in reference to the OGTT standard.

Blocking effect of sperm immobilizing antibodies on sperm penetration of human zonae pellucidae.

To investigate the mechanism of the blocking effect of sperm immobilizing antibodies on human fertilization, an in vitro zona penetration test was carried out using media containing the IgG fraction extracted from sperm immobilizing antibody-negative or -positive serum. The sperm penetration rate of the test was 100% (6/6) when spermatozoa were treated with the IgG fraction derived from sperm immobilizing antibody-negative serum, whereas it was only 17% (1/6) when spermatozoa were treated with the IgG fraction derived from sperm immobilizing antibody-positive serum. Electron microscopic observation of the sperm immobilizing antibody-negative and -positive serum-treated spermatozoa showed that the number of acrosome-reacted spermatozoa was significantly greater in the sperm immobilizing antibody-negative serum than in the antibody-positive serum. Therefore, it appears that one of the blocking mechanisms of the spermatozoal penetration of the zona pellucida by sperm immobilizing antibodies may be due to inhibition of the acrosome reaction in the spermatozoa.

Influence of water quality on in vitro fertilization and embryo development for the mouse.

Mouse in vitro fertilization and embryo culture were performed in media prepared with five different water preparations. The results of the experiments improved with the frequency of distillation. Each water preparation was analyzed by the measurement of the electrical conductivities and inorganic ion concentrations and by high-performance liquid chromatography to examine the mutual relation between water quality and the method of water purification. The best results were obtained with Milli-Q water, which had the lowest concentration of inorganic ions and organic compounds. On the contrary, unexpected contamination by organic compounds and zinc ions occurred after multiple distillation, possibly leached from the glassware and silicon tube. The hatching rate seemed to be an appropriate indicator to assess the biological qualities of media for the development of embryos cultured in vitro.

Inhibition of sperm penetration through human zona pellucida by antisperm antibodies.

An in vitro penetration test using human spermatozoa, sera, and eggs stored in a highly concentrated salt solution was designed for examination of the effect of antisperm antibodies on the process of fertilization. Spermatozoa from a healthy fertile donor incubated in modified Biggers, Whiiten and Whittingham (BWW) medium containing 7.5% antisperm-antibody-negative serum, could penetrate through the zonae pellucidae of the stored eggs, but not when the spermatozoa from the same donor had been incubated in modified BWW medium containing 7.5% antisperm-antibody-positive serum. After the antisperm-antibody-positive serum was absorbed with washed spermatozoa, the sperm penetration was not blocked. Therefore, antisperm antibodies appear to block human sperm penetration through the human zona pellucida.