Yoshihisa Nakatani

Possible novel therapy for diabetes with cell-permeable JNK-inhibitory peptide

The JNK pathway is known to be activated in several tissues in the diabetic state, and is possibly involved in the development of insulin resistance and suppression of insulin biosynthesis. Here we show a potential new therapy for diabetes using cell-permeable JNK-inhibitory peptide. Intraperitoneal administration of the peptide led to its transduction into various tissues in vivo, and this treatment markedly improved insulin resistance and ameliorated glucose tolerance in diabetic mice. These data indicate that the JNK pathway is critically involved in diabetes and that the cell-permeable JNK-inhibitory peptide may have promise as a new therapeutic agent for diabetes.

Effect of Troglitazene on Microalbuminuria in Patients With Incipient Diabetic Nephropathy

OBJECTIVE Although some studies have suggested a direct action of troglitazone on vascular cells, its effects on diabetic vascular diseases have not been reported. We therefore investigated the effect of troglitazone on microalbuminuria in patients with incipient diabetic nephropathy. RESEARCH DESIGN AND METHODS A total of 30 patients with type 2 diabetes associated with microalbuminuria (urinary albumin-to-creatinine ratio [ACR] [milligrams per gram creatinine] ranging from 30 to 300 mg/g creatinine) were studied. They were randomly divided into two groups: patients treated with metformin (500 mg/day, n = 13) or with troglitazone (400 mg/day, n = 17) for 12 weeks. ACR, lipid profile, blood pressure, glycated hemoglobin, and plasma glucose during meal-load tests were measured every 4 weeks. RESULTS Anthropometric indices (BMI and percent fat), lipid profile, and blood pressure did not change with either treatment. Fasting and postmeal glucose levels decreased similarly in the two groups. Decrements in glycated hemoglobin were greater in the metformin group at 4 and 8 weeks after the initiation of treatment (P < 0.05). Troglitazone reduced ACR (median [25–75th percentiles]) from 70 (49–195) to 40 (31–90) mg/g creatinine at 4 weeks (P = 0.021) and maintained these reduced levels throughout the treatment period (8 weeks: 35 [26–68], P = 0.007; 12 weeks: 43 [26–103], P = 0.047). Metformin did not change ACR throughout the 12 weeks. CONCLUSIONS Troglitazone ameliorated microalbuminuria in diabetic nephropathy. Furthermore, our findings suggest that troglitazone has some effects on vascular cells other than lowering plasma glucose levels. Troglitazone might be useful for diabetic angiopathy, including nephropathy and coronary artery disease.

The Forkhead Transcription Factor Foxo1 Bridges the JNK Pathway and the Transcription Factor PDX-1 through Its Intracellular Translocation

It has been shown that oxidative stress and activation of the c-Jun N-terminal kinase (JNK) pathway induce the nucleocytoplasmic translocation of the pancreatic transcription factor PDX-1, which leads to pancreatic β-cell dysfunction. In this study, we have shown that the forkhead transcription factor Foxo1/FKHR plays a role as a mediator between the JNK pathway and PDX-1. Under oxidative stress conditions, Foxo1 changed its intracellular localization from the cytoplasm to the nucleus in the pancreatic β-cell line HIT-T15. The overexpression of JNK also induced the nuclear localization of Foxo1, but in contrast, suppression of JNK reduced the oxidative stress-induced nuclear localization of Foxo1, suggesting the involvement of the JNK pathway in Foxo1 translocation. In addition, oxidative stress or activation of the JNK pathway decreased the activity of Akt in HIT cells, leading to the decreased phosphorylation of Foxo1 following nuclear localization. Furthermore, adenovirus-mediated Foxo1 overexpression reduced the nuclear expression of PDX-1, whereas repression of Foxo1 by Foxo1-specific small interfering RNA retained the nuclear expression of PDX-1 under oxidative stress conditions. Taken together, Foxo1 is involved in the nucleocytoplasmic translocation of PDX-1 by oxidative stress and the JNK pathway.

Oxidative stress, ER stress, and the JNK pathway in type 2 diabetes

Pancreatic β-cell dysfunction and insulin resistance are observed in type 2 diabetes. Under diabetic conditions, oxidative stress and ER stress are induced in various tissues, leading to activation of the JNK pathway. This JNK activation suppresses insulin biosynthesis and interferes with insulin action. Indeed, suppression of the JNK pathway in diabetic mice improves insulin resistance and ameliorates glucose tolerance. Thus, the JNK pathway plays a central role in pathogenesis of type 2 diabetes and may be a potential target for diabetes therapy.

A Crucial Role of MafA as a Novel Therapeutic Target for Diabetes

MafA, a recently isolated pancreatic β-cell-specific transcription factor, is a potent activator of insulin gene transcription. In this study, we show that MafA overexpression, together with PDX-1 (pancreatic and duodenal homeobox factor-1) and NeuroD, markedly increases insulin gene expression in the liver. Consequently, substantial amounts of insulin protein were induced by such combination. Furthermore, in streptozotocin-induced diabetic mice, MafA overexpression in the liver, together with PDX-1 and NeuroD, dramatically ameliorated glucose tolerance, while combination of PDX-1 and NeuroD was much less effective. These results suggest a crucial role of MafA as a novel therapeutic target for diabetes.

Oxidative Stress and the JNK Pathway in Diabetes

Under diabetic conditions, oxidative stress is induced and the JNK pathway is activated, which is involved in deterioration of pancreatic beta-cell function found in diabetes. Oxidative stress and/or activation of the JNK pathway suppress insulin gene expression, accompanied by reduction of PDX-1 DNA binding activity. Treatment with antioxidants and/or suppression of the JNK pathway protect beta-cells from some of the toxic effects of hyperglycemia. The JNK pathway is also involved in the progression of insulin resistance; suppression of the JNK pathway in obese diabetic mice markedly improves insulin resistance and ameliorates glucose tolerance. The phosphorylation state of key molecules for insulin signaling is altered upon modification of the JNK pathway. Taken together, the JNK pathway plays a crucial role in progression of insulin resistance as well as beta-cell dysfunction found in diabetes and thus could be a potential therapeutic target for diabetes.

Impaired splanchnic and peripheral glucose uptake in liver cirrhosis

BACKGROUND/AIM Patients with liver cirrhosis are insulin-resistant and frequently glucose-intolerant. Although peripheral glucose uptake has been shown to be impaired in liver cirrhosis, little is known about the significance of splanchnic (hepatic) glucose uptake after oral glucose load. METHODS/RESULTS We performed an oral glucose tolerance test and euglycemic hyperinsulinemic clamp with oral glucose load for eight patients with liver cirrhosis and eight patients with chronic active hepatitis. The patients with liver cirrhosis had higher plasma glucose levels 2 h after glucose load than those with chronic active hepatitis (228+/-22 mg/dl vs. 102+/-9 mg/dl, p<0.01). Using the euglycemic hyperinsulinemic clamp with oral glucose load, we simultaneously measured peripheral and splanchnic glucose uptake. Peripheral glucose uptake in liver cirrhosis was 6.1+/-0.7 mg x kg(-1) x min(-1), which was lower than that in healthy volunteers (10.5+/-0.9 mg x kg(-1) x min(-1), p<0.05) and in chronic active hepatitis (8.4+/-0.3 mg x kg(-1) x min(-1), p<0.05). Furthermore, splanchnic glucose uptake in liver cirrhosis was much lower (20.1+/-3.4%) than in healthy volunteers (36.0+/-4.0%, p<0.05) and in chronic active hepatitis (37.2+/-3.1%, p<0.05). CONCLUSION These results suggest that glucose intolerance in patients with liver cirrhosis is caused by a defect of the glucose uptake of both splanchnic and peripheral tissues.

Germinability of coat-lacking spores of Bacillusmegaterium

Upon treatment with acid, the germinability of both intact and coat-lacking spores of Bacillus megaterium ATCC 19213 exhibited similar features. Namely, when the spores previously germinated by alanine in the presence of phosphate buffer were converted to H-spores by treatment with nitric acid, germination proceeded at a very low speed in a same germination medium. When H-spores converted to Ca-spores by treatment with calcium acetate and subsequently germinated, germination proceeded at a speed higher than that of native spores and occurred even in the absence of buffer. These results suggest that the site of exchangeable cations concerned with germinability must not exist in the coat.

Mutation of the Pax6 gene causes impaired glucose-stimulated insulin secretion

To the Editor: The paired-box gene that encodes Pax6 is a member of the Pax gene family and is expressed in eye, nose, pancreas and the central nervous system. It is well known that aniridia, a panocular human eye malformation, is caused by mutations in Pax6 [1]. Mutant rats with small eyes (rSey) were identified during the course of breeding Sprague–Dawley rats [2], and it was shown that about 600 base pairs of Pax6 mRNA are deleted in these rats. At the genomic level, a single base (G) insertion in the exon produces the truncated mRNA [3]. Heterozygotes (rSey/+) have small eyes, while homozygotes (rSey/rSey) lack eyes and a nose, resulting in perinatal death. We previously evaluated glucose tolerance in six patients with aniridia and found that five of the patients possessed a Pax6 mutation and had impaired insulin secretion and glucose intolerance [4]. Based on this finding, we hypothesised that a reduction in Pax6 expression or activity leads to the deterioration of beta cell function. In the present study we evaluated pancreatic beta cell function in rats with a heterozygous mutation in Pax6 (rSey/+ rats) to investigate whether insufficient insulin secretion is attributable to Pax6 inactivation. It should be noted that the rats used in this study were inbred. To evaluate pancreatic beta cell function we performed an IVGTT. Glucose-stimulated first-phase insulin secretion (at 2 min) was significantly lower in rSey/+ rats than in +/+ rats (Fig. 1a), although the two groups had similar plasma glucose levels (Fig. 1b). In contrast, arginine-stimulated insulin secretion in rSey/+ rats was almost the same as that in +/+ rats (Fig. 1c). To further evaluate glucose-stimulated insulin secretion, we performed a perifusion study using islets isolated from rSey/+ and +/+ rats. As shown in Figure 1d, glucose-stimulated insulin secretion was significantly lower in islets isolated from rSey/+ rats than in those from +/+ rats. It should be noted that a marked difference in glucose-stimulated first-phase insulin secretion was observed at 1 min. As in the in vivo study, arginine-stimulated insulin secretion in islets from rSey/+ rats was almost the same as that in islets from +/+ rats (Fig. 1e). KCl-stimulated insulin secretion in rSey/+ islets was also comparable to that in +/+ islets (data not shown). Taken together, these results indicate that the heterozygous mutation in Pax6 specifically impaired glucose-stimulated insulin secretion without influencing arginineor KCl-stimulated insulin secretion. To examine islet morphology and insulin biosynthesis, we performed immunostaining for insulin and examined insulin content in the pancreas. There were no differences in islet morphology (data not shown) or insulin content between pancreata from rSey/+ and +/+ rats. To elucidate the mechanism by which mutation of Pax6 causes impairment of glucosestimulated insulin secretion, we used RT-PCR to examine the expression of various beta-cell-related factors in the islets of the rats. We assumed that there are some defects in the pathway between glucose entry and membrane depolarisation in the beta cells of the rSey/+ rats. Glucose is the primary physiological stimulus for insulin secretion, and this process requires glucose sensing [5]. GLUT2 facilitates the rapid equilibration of glucose across the plasma membrane, and glucokinase is the rate-limiting step in glycolytic flux for insulin secretion [6]. Both of these proteins are crucial for glucosestimulated insulin secretion. The KATP channel, which contains Kir 6.2 and sulphonylurea receptor 1 (SUR1), is also important for glucose-induced insulin secretion. However, we observed no differences in the levels of expression of glucokinase, GLUT2, Kir 6.2 or SUR-1 between the two groups of rats (data not shown). These results indicate that the heterozygous mutation in Pax6 causes impaired glucose-stimulated insulin secretion without influencing islet morphology, insulin content in the pancreas or the expression of various betacell-related factors. Thus, it remains unknown how mutation of Pax6 causes impairment of glucose-stimulated insulin secretion. Although not examined in this study, it was recently reported that the transcription of the gene encoding the glucose-6-phosphatase catalytic subunit-related protein is regulated by Pax6 [7], and it is possible that this is affected by mutation of Pax6. In conclusion, the results of the present study indicate that in rSey/+ rats, glucose-stimulated insulin secretion is specifically impaired, while arginine-stimulated insulin secretion, islet morphology and expression of various beta-cell-related factors are almost the same as in +/+ rats. Since it is well known that a deficiency in glucose-stimulated early-phase insulin secretion is a strong predictor for the development of type 2 diabetes [8], we postulate that decreased Pax6 expression or activity could lead to type 2 diabetes.