A 1-year, prospective, observational study of Japanese outpatients with type 1 and type 2 diabetes switching from insulin glargine or detemir to insulin degludec in basal-bolus insulin therapy (Kumamoto Insulin Degludec Observational study).

AIMS/INTRODUCTION: The aim of the present prospective observational study was to assess long-term efficacy and safety of insulin degludec as a part of a basal-bolus therapy for Japanese patients with type 1 or type 2 diabetes in routine clinical practice. MATERIALS AND METHODS: In the present study, 93 type 1 diabetes patients and 135 type 2 diabetes patients treated with insulin glargine or detemir were switched from their basal insulin to insulin degludec. The primary end-points were the changes in glycated hemoglobin (HbA1c) from baseline at 3, 6 and 12 months. The secondary end-points were changes in body mass index, insulin dose, frequency of hypoglycemia and adverse events. RESULTS: HbA1c levels from baseline were significantly reduced at 3, 6, and 12 months by 0.4, 0.4 and 0.3% in type 1 diabetes patients, respectively, and by 0.5, 0.5 and 0.3% in type 2 diabetes patients, respectively. Body mass index in type 1 diabetes patients increased significantly (P < 0.05), whereas that in type 2 diabetes patients did not change. Basal insulin dose decreased significantly at 3 months after switching (P < 0.05), and returned baseline dose at 12 months in type 1 diabetes and type 2 diabetes patients. The frequency of both total and nocturnal hypoglycemia decreased significantly in type 1 diabetes and type 2 diabetes patients (P < 0.05). The result of multiple regression analysis showed that baseline HbA1c was a significant independent variable of the percentage change in HbA1c with switching. CONCLUSION: In both type 1 diabetes and type 2 diabetes patients, switching from insulin glargine or insulin detemir to insulin degludec led to improvement of glycemic control with a significant reduction of hypoglycemia.

TZDs reduce mitochondrial ROS production and enhance mitochondrial biogenesis.

Although it has been reported that thiazolidinediones (TZDs) may reduce cardiovascular events in type 2 diabetic patients, its precise mechanism is unclear. We previously demonstrated that hyperglycemia-induced production of reactive oxygen species from mitochondria (mtROS) contributed to the development of diabetic complications, and metformin normalized mt ROS production by induction of MnSOD and promotion of mitochondrial biogenesis by activating the PGC-1alpha pathway. In this study, we examined whether TZDs could inhibit hyperglycemia-induced mtROS production by activating the PGC-1alpha pathway. We revealed that pioglitazone and ciglitazone attenuated hyperglycemia-induced ROS production in human umbilical vein endothelial cells (HUVECs). Both TZDs increased the expression of NRF-1, TFAM and MnSOD mRNA. Moreover, pioglitazone increased mtDNA and mitochondrial density. These results suggest that TZDs normalize hyperglycemia-induced mtROS production by induction of MnSOD and promotion of mitochondrial biogenesis by activating PGC-1alpha. This phenomenon could contribute to the prevention of diabetic vascular complications.

Endothelial MnSOD overexpression prevents retinal VEGF expression in diabetic mice.

We previously proposed that hyperglycemia-induced mitochondrial ROS overproduction is a key event in the development of diabetic complications. In this study, we established a novel transgenic mouse (eMnSOD-Tg), which specifically expressed MnSOD in endothelial cells, by employing a Tie2 promoter/enhancer, and investigated the impact of mitochondrial ROS production on diabetic retinopathy in vivo. Using immunohistochemistry, overexpression of MnSOD in endothelial cells was confirmed in eMnSOD-Tg mice. By introduction of diabetes by streptozotocin, levels of urinary 8-hydroxydeoxyguanosine, a marker of mitochondrial oxidative stress, and expression of VEGF mRNA and protein and fibronectin mRNA in retinas were increased in wild-type littermates. However, these observations were ameliorated in eMnSOD-Tg mice, although control and eMnSOD-Tg mice showed a comparable level of hyperglycemia. In the present study, we newly developed a line of transgenic mice, which specifically express MnSOD in endothelium. In addition, overexpression of mitochondrial-specific SOD in endothelium could prevent diabetic retinopathy in vivo.

Impact of mitochondrial ROS production in the pathogenesis of insulin resistance.

Tumor necrosis factor-alpha (TNF-alpha) inhibits insulin action, in part, by activating c-jun NH(2)-terminal kinases (JNK). However, the precise mechanisms by which TNF-alpha activates JNK are unknown. Recently, we confirmed that hyperglycemia increased mitochondrial reactive oxygen species (ROS) production, and which can associate with the pathogenesis of diabetic vascular complications. In addition, apoptosis signal-regulating kinase 1 (ASK1) was reported to activate the JNK and p38 signaling pathways and is required for TNF-alpha-induced apoptosis. Here we demonstrate that TNF-alpha increases mitochondrial ROS production and ASK1 activity, and that these TNF-alpha-induced phenomena associate with JNK activation, increase in Ser(307) phosphorylation of IRS-1 and decrease in insulin-stimulated tyrosine phosphorylation of IRS-1, all of which are believed to be the molecular basis of TNF-alpha-induced insulin resistance. We claim that mitochondrial ROS production may be a key factor not only in diabetic vascular complications, but also in the development of type 2 diabetes. This integrating paradigm could provide a new conceptual framework for further research and therapies for the treatment of type 2 diabetes.

Impact of mitochondrial ROS production on diabetic vascular complications.

Vascular complications are the leading cause of morbidity and mortality in patients with diabetes. Four main molecular mechanisms have been implicated in glucose-mediated vascular disease. There are: glucose-induced activation of protein kinase C (PKC) isoforms; increased formation of glucose-derived advanced glycation end-products (AGE); increased glucose flux through the aldose reductase pathway; and increased production of reactive oxygen species (ROS). Here we demonstrate that hyperglycemia-induced production of ROS is abrogated by inhibitors of mitochondrial metabolism, or by overexpression of uncoupling protein-1 or manganese superoxide dismutase. Normalization of mitochondrial ROS production by each of these agents prevents glucose-induced activation of PKC, formation of AGE, and accumulation of sorbitol in bovine vascular endothelial cells. We also claim that 8-hydroxydeoxyguanosine, which represents mitochondrial oxidative damage was elevated in patients with either retinopathy, albuminuria or increased intima-media thickness of carotid arteries. These results suggest that hyperglycemia induces mitochondrial ROS production, and which can associate to the pathogenesis of diabetic vascular complications.

Impact of mitochondrial reactive oxygen species and apoptosis signal-regulating kinase 1 on insulin signaling.

Tumor necrosis factor (TNF)-alpha inhibits insulin action; however, the precise mechanisms are unknown. It was reported that TNF-alpha could increase mitochondrial reactive oxygen species (ROS) production, and apoptosis signal-regulating kinase 1 (ASK1) was reported to be required for TNF-alpha-induced apoptosis. Here, we examined roles of mitochondrial ROS and ASK1 in TNF-alpha-induced impaired insulin signaling in cultured human hepatoma (Huh7) cells. Using reduced MitoTracker Red probe, we confirmed that TNF-alpha increased mitochondrial ROS production, which was suppressed by overexpression of either uncoupling protein-1 (UCP)-1 or manganese superoxide dismutase (MnSOD). TNF-alpha significantly activated ASK1, increased serine phosphorylation of insulin receptor substrate (IRS)-1, and decreased insulin-stimulated tyrosine phosphorylation of IRS-1 and serine phosphorylation of Akt, and all of these effects were inhibited by overexpression of either UCP-1 or MnSOD. Similar to TNF-alpha, overexpression of wild-type ASK1 increased serine phosphorylation of IRS-1 and decreased insulin-stimulated tyrosine phosphorylation of IRS-1, whereas overexpression of dominant-negative ASK1 ameliorated these TNF-alpha-induced events. In addition, TNF-alpha activated c-jun NH(2)-terminal kinases (JNKs), and this observation was partially inhibited by overexpression of UCP-1, MnSOD, or dominant-negative ASK1. These results suggest that TNF-alpha increases mitochondrial ROS and activates ASK1 in Huh7 cells and that these TNF-alpha-induced phenomena contribute, at least in part, to impaired insulin signaling.

Activation of AMP-activated protein kinase reduces hyperglycemia-induced mitochondrial reactive oxygen species production and promotes mitochondrial biogenesis in human umbilical vein endothelial cells.

We previously proposed that the production of hyperglycemia-induced mitochondrial reactive oxygen species (mtROS) is a key event in the development of diabetes complications. The association between the pathogenesis of diabetes and its complications and mitochondrial biogenesis has been recently reported. Because metformin has been reported to exert a possible additional benefit in preventing diabetes complications, we investigated the effect of metformin and 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) on mtROS production and mitochondrial biogenesis in cultured human umbilical vein endothelial cells. Treatment with metformin and AICAR inhibited hyperglycemia-induced intracellular and mtROS production, stimulated AMP-activated protein kinase (AMPK) activity, and increased the expression of peroxisome proliferator-activated response-gamma coactivator-1alpha (PGC-1alpha) and manganese superoxide dismutase (MnSOD) mRNAs. The dominant negative form of AMPKalpha1 diminished the effects of metformin and AICAR on these events, and an overexpression of PGC-1alpha completely blocked the hyperglycemia-induced mtROS production. In addition, metformin and AICAR increased the mRNA expression of nuclear respiratory factor-1 and mitochondrial DNA transcription factor A (mtTFA) and stimulated the mitochondrial proliferation. Dominant negative-AMPK also reduced the effects of metformin and AICAR on these observations. These results suggest that metformin normalizes hyperglycemia-induced mtROS production by induction of MnSOD and promotion of mitochondrial biogenesis through the activation of AMPK-PGC-1alpha pathway.

A case of female pseudohermaphroditism caused by aromatase deficiency.

Female pseudohermaphroditism is caused by several etiologies. Here we report a case of aromatase deficiency who showed ambiguous genitalia and maternal virilization during pregnancy. The mother had noticed her own virilization from 16 wk of gestation without androgen exposure and had low urinary estriol levels (5~10 µg/ml at 35 wk of gestation). At birth, the patient presented severe virilization (Prader V), and was assigned as a male with a micropenis and unpalpable testes but the patient had a normal female karyotype and a uterus and cystic ovaries found by magnetic resonance imaging. The patient had a increase in serum 17α-hydroxy progesterone levels (basal 4.9 → 37 ng/ml after a single 0.25 mg/m(2) infusion of ACTH), but the increase in adrenal androgen was not sufficient to virilize the external genitalia. Dehydroepiandrosterone, 17α-hydroxy pregnenolone and deoxycorticosterone were within the normal ranges. These findings suggested a diagnosis of nonadrenal female pseudohermaphroditism. From the clinical features and biochemical data, we endocrinologically diagnosed her as having an aromatase deficiency. The aromatase gene is now under investigation for definite diagnosis. We finally agreed that aromatase deficiency should be suspected when both the mother and the newborn have been virilized.

Dysregulation of IL-13 production by cord blood CD4+ T cells is associated with the subsequent development of atopic disease in infants.

Early intervention strategies in allergic diseases will be dependent on identification of newborns at high risk for later development of atopic disease. In this cohort study of 106 neonates, we investigated whether cytokine production property and responsiveness to IL-12 of neonatal CD4(+) T cells were associated with the subsequent development of atopic disease and whether a skewed cytokine production property was intrinsic to helper T cells. To exclude the effects of contaminating cells, highly purified cord blood CD4(+) T cells were stimulated with anti-CD3 MAb and recombinant B7-2 molecule in the presence or absence of IL-12. Production of IL-13 and interferon-gamma was determined by ELISA. The infants were assessed at 12 mo for the development of atopic diseases. CD4(+) T cells of neonates who manifested allergic symptoms (atopic group) produced higher levels of IL-13 compared with those of the nonatopic group in both the presence and absence of IL-12. No significant difference was noted between the two groups with respect to interferon-gamma production. Moreover, higher IL-13 production was also observed in neonates with chronic eczema than those with short-term eczema. Our data suggest that increased production of IL-13 by neonatal CD4(+) T cells is a useful marker of newborns at high risk for subsequent development of atopic diseases and that an intrinsic abnormality of CD4(+) T cell is associated with the pathogeneses of atopic disease, especially atopic dermatitis in infants.