Inhibition of inflammation-mediated DPP-4 expression by linagliptin increases M2 macrophages in atherosclerotic lesions.

BACKGROUND AND AIMS: Dipeptidyl peptidase-4 (DPP-4) inhibitors have been reported to suppress atherosclerosis progression in atherosclerotic mouse models through unclear mechanisms. In this study, we investigated the effect of the DPP-4 inhibitor, linagliptin, on macrophage polarization in vitro and in vivo. METHODS: Mouse bone marrow macrophages (BMMs) were used in in vitro assays. High fat diet (HFD)-fed Apoe-/- mice were treated orally with linagliptin (10 mg/kg-1•day-1) or a vehicle (water) control. RESULTS: In in vitro assays using BMMs, treatment with LPS and IFNγ decreased the mRNA-expression levels of alternatively activated macrophage (M2) markers, and linagliptin treatment prevented these reductions. The mRNA levels of M2 markers and the number of M2 macrophages in the aorta were higher in linagliptin groups than in control groups. Linagliptin decreased the size of atherosclerotic lesions in HFD-fed Apoe-/- mice. Interestingly, inflammatory stimulation increased DPP-4 expression, and linagliptin suppressed these effects in BMMs. Treatment with DPP-4 small-interfering RNA (siRNA) reproduced linagliptin-mediated alteration of M2 polarization. CONCLUSIONS: Linagliptin increased M2 macrophage polarization by inhibiting DPP-4 expression and activity. These findings may indicate the beneficial effects of DPP-4 inhibitors on the progression of diabetic macrovascular complications.

Impact of hepatic HSP72 on insulin signaling.

Heat shock protein 72 (HSP72) is a major inducible molecule in the heat shock response that enhances intracellular stress tolerance. Decreased expression of HSP72 is observed in type 2 diabetes, which may contribute to the development of insulin resistance and chronic inflammation. We used HSP72 knockout (HSP72-KO) mice to investigate the impact of HSP72 on glucose metabolism and endoplasmic reticulum (ER) stress, particularly in the liver. Under a high-fat diet (HFD) condition, HSP72-KO mice showed glucose intolerance, insulin resistance, impaired insulin secretion, and enhanced hepatic gluconeogenic activity. Furthermore, activity of the c-Jun NH2-terminal kinase (JNK) was increased and insulin signaling suppressed in the liver. Liver-specific expression of HSP72 by lentivirus (lenti) in HFD-fed HSP72-KO mice ameliorated insulin resistance and hepatic gluconeogenic activity. Furthermore, increased adipocyte size and hepatic steatosis induced by the HFD were suppressed in HSP72-KO lenti-HSP72 mice. Increased JNK activity and ER stress upon HFD were suppressed in the liver as well as the white adipose tissue of HSP72-KO lenti-HSP72 mice. Thus, HSP72 KO caused a deterioration in glucose metabolism, hepatic gluconeogenic activity, and ß-cell function. Moreover, liver-specific recovery of HSP72 restored glucose homeostasis. Therefore, hepatic HSP72 may play a critical role in the pathogenesis of type 2 diabetes.

Inhibition of Local Macrophage Growth Ameliorates Focal Inflammation and Suppresses Atherosclerosis.

OBJECTIVE: Macrophages play a central role in various stages of atherosclerotic plaque formation and progression. The local macrophages reportedly proliferate during atherosclerosis, but the pathophysiological significance of macrophage proliferation in this context remains unclear. Here, we investigated the involvement of local macrophage proliferation during atherosclerosis formation and progression using transgenic mice, in which macrophage proliferation was specifically suppressed. APPROACH AND RESULTS: Inhibition of macrophage proliferation was achieved by inducing the expression of cyclin-dependent kinase inhibitor 1B, also known as p27kip, under the regulation of a scavenger receptor promoter/enhancer. The macrophage-specific human p27kip Tg mice were subsequently crossed with apolipoprotein E-deficient mice for the atherosclerotic plaque study. Results showed that a reduced number of local macrophages resulted in marked suppression of atherosclerotic plaque formation and inflammatory response in the plaque. Moreover, fewer local macrophages in macrophage-specific human p27kip Tg mice helped stabilize the plaque, as evidenced by a reduced necrotic core area, increased collagenous extracellular matrix, and thickened fibrous cap. CONCLUSIONS: These results provide direct evidence of the involvement of local macrophage proliferation in formation and progression of atherosclerotic plaques and plaque stability. Thus, control of macrophage proliferation might represent a therapeutic target for treating atherosclerotic diseases.

Acetate alters expression of genes involved in beige adipogenesis in 3T3-L1 cells and obese KK-Ay mice.

The induction of beige adipogenesis within white adipose tissue, known as "browning", has received attention as a novel potential anti-obesity strategy. The expression of some characteristic genes including PR domain containing 16 is induced during the browning process. Although acetate has been reported to suppress weight gain in both rodents and humans, its potential effects on beige adipogenesis in white adipose tissue have not been fully characterized. We examined the effects of acetate treatment on 3T3-L1 cells and in obese diabetic KK-Ay mice. The mRNA expression levels of genes involved in beige adipocyte differentiation and genes selectively expressed in beige adipocytes were significantly elevated in both 3T3-L1 cells incubated with 1.0 mM acetate and the visceral white adipose tissue from mice treated with 0.6% acetate for 16 weeks. In KK-Ay mice, acetate reduced the food efficiency ratio and increased the whole-body oxygen consumption rate. Additionally, reduction of adipocyte size and uncoupling protein 1-positive adipocytes and interstitial areas with multilocular adipocytes appeared in the visceral white adipose tissue of acetate-treated mice, suggesting that acetate induced initial changes of "browning". In conclusion, acetate alters the expression of genes involved in beige adipogenesis and might represent a potential therapeutic agent to combat obesity.

Statins meditate anti-atherosclerotic action in smooth muscle cells by peroxisome proliferator-activated receptor-γ activation.

The peroxisome proliferator-activated receptor-γ (PPARγ) is an important regulator of lipid and glucose metabolism, and its activation is reported to suppress the progression of atherosclerosis. We have reported that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) activate PPARγ in macrophages. However, it is not yet known whether statins activate PPARγ in other vascular cells. In the present study, we investigated whether statins activate PPARγ in smooth muscle cells (SMCs) and endothelial cells (ECs) and thus mediate anti-atherosclerotic effects. Human aortic SMCs (HASMCs) and human umbilical vein ECs (HUVECs) were used in this study. Fluvastatin and pitavastatin activated PPARγ in HASMCs, but not in HUVECs. Statins induced cyclooxygenase-2 (COX-2) expression in HASMCs, but not in HUVECs. Moreover, treatment with COX-2-siRNA abrogated statin-mediated PPARγ activation in HASMCs. Statins suppressed migration and proliferation of HASMCs, and inhibited lipopolysaccharide-induced expression of monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) in HASMCs. These effects of statins were abrogated by treatment with PPARγ-siRNA. Treatment with statins suppressed atherosclerotic lesion formation in Apoe(-/-) mice. In addition, transcriptional activity of PPARγ and CD36 expression were increased, and the expression of MCP-1 and TNF-α was decreased, in the aorta of statin-treated Apoe(-/-) mice. In conclusion, statins mediate anti-atherogenic effects through PPARγ activation in SMCs. These effects of statins on SMCs may be beneficial for the prevention of atherosclerosis.

Insulin requirement profiles in Japanese hospitalized subjects with type 2 diabetes treated with basal-bolus insulin therapy.

To assess the total daily inulin dose (TDD) and contribution of basal insulin to TDD and to identify the predictive factors for insulin requirement profiles in subjects with type 2 diabetes, we retrospectively examined insulin requirement profiles of 275 hospitalized subjects treated with basal-bolus insulin therapy (BBT) (mean age, 60.1 ± 12.9 years; HbA1c, 10.2 ± 4.5%). Target plasma glucose level was set between 80 and 129 mg/dL before breakfast and between 80 and 179 mg/dL at 2-hour after each meal without causing hypoglycemia. We also analyzed the relationship between the insulin requirement profiles (TDD and basal/total daily insulin ratio [B/TD ratio]) and insulin-associated clinical parameters. The mean TDD was 0.463 ± 0.190 unit/kg/day (range, 0.16-1.13 unit/kg/day). The mean B/TD ratio was 0.300 ± 0.099 (range, 0.091-0.667). A positive correlation of TDD with B/TD ratio was revealed by linear regression analysis (r=0.129, p=0.03). Stepwise multiple regression analysis identified post-breakfast glucose levels before titrating insulin as an independent determinant of the insulin requirement profile [Std ß (standard regression coefficient) = 0.228, p<0.01 for TDD, Std ß = -0.189, p<0.01 for B/TD ratio]. The TDD was <0.6 unit/kg/day and the B/TD ratio was <0.4 in the majority (70.2%) of subjects in the present study. These findings may have relevance in improving glycemic control and decreasing the risk of hypoglycemia and weight gain in subjects with type 2 diabetes treated with BBT.

Expression, purification, and characterization of cold-adapted inorganic pyrophosphatase from psychrophilic Shewanella sp. AS-11.

In the presence of divalent cations, inorganic pyrophosphatase is activated to hydrolyze inorganic pyrophosphate to inorganic phosphate. Here, we clone, express, purify, and characterize inorganic pyrophosphatase from the psychrophilic Shewanella sp. AS-11 (Sh-PPase). The recombinant Sh-PPase was expressed in Escherichia coli BL21 (DE3) at 20°C using pET16b as an expression vector and purified from the cell extracts by a combination of ammonium sulfate fractionation and anion-exchange chromatography. Sh-PPase was found to be a family II PPase with a subunit molecular mass of 34 kD that preferentially utilizes Mn²âº over Mg²âº ions for activity. The functional characteristics of Sh-PPase, such as activity, temperature dependency, and thermal inactivation, were greatly influenced by manganese ions. Manganese ion activation increased the enzyme's activity at low temperatures; therefore, it was required to gain the cold-adapted characteristics of Sh-PPase.

Autoimmune Polyglandular Syndrome Type 3 Complicated with IgG4-related Disease.

A 52-year-old Japanese woman developed type 1 diabetes mellitus (type 1 DM) at 41 years old. She became complicated with Hashimoto's disease and showed swelling of both submandibular glands, which was diagnosed as IgG4-related disease (IgG4-RD). This is a rare case of a Japanese patient with autoimmune polyglandular syndrome type 3A (APS-3A) coexisting with autoimmune thyroid disease (AITD) and type 1 DM complicated by IgG4-RD. Bilateral submandibular gland resection was successfully performed without steroid therapy. We discuss the possibility that the immunological pathogenic mechanisms of APS-3A and IgG4-RD are related.

Gly or Ala substitutions for Pro(210)Thr(211)Asn(212) at the ß8-ß9 turn of subtilisin Carlsberg increase the catalytic rate and decrease thermostability.

A comparison of the primary structures among psychrophilic, mesophilic, and thermophilic subtilases revealed that the turn between the ß8 and ß9 strands (ß8-ß9 turn, BPN' numbering) of psychrophilic subtilases are more flexible than those of their mesophilic and thermophilic counterparts. To investigate the relationship between structure of this turn and enzyme activity as well as thermostability of mesophilic subtilisin Carlsberg (sC), we analyzed 6 mutants of sC with a single, double, or triple Gly or Ala substitutions for Pro(210)Thr(211)Asn(212) at the ß8-ß9 turn. Among the single Gly substitutions, the P210G substitution most significantly (1.5-fold) increased the specific activity on N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide (AAPF) substrate and 12-fold decreased the thermostability. All mutants tested showed the increased k(cat) for the AAPF substrate and reduced thermostability compared with the wild-type sC. The k(cat) values of the P210G, P210G/T211G, and P210G/T211G/N212G mutants were 1.5-, 1.7-, and 1.8-fold higher than that of the wild-type sC. There were significant positive correlations between k(cat) and thermal inactivation rates as well as k(cat) and K(m) of the wild-type and mutants. These results demonstrate that the structure of ß8-ß9 turn, despite its distance from the active site, has significant effects on the catalytic rate and thermostability of sC through a global network of intramolecular interactions and suggest that the lack of flexibility of this turn stabilizes the wild-type sC against thermal inactivation in compensation for some loss of catalytic activity.

Apocynin suppresses the progression of atherosclerosis in apoE-deficient mice by inactivation of macrophages.

Production of reactive oxygen species (ROS) and other proinflammatory substances by macrophages plays an important role in atherogenesis. Apocynin (4-hydroxy-3-methoxy-acetophenone), which is well known as a NADPH oxidase inhibitor, has anti-inflammatory effects including suppression of the generation of ROS. However, the suppressive effects of apocynin on the progression of atherosclerosis are not clearly understood. Thus, we investigated anti-atherosclerotic effects of apocynin using apolipoprotein E-deficient (apoE(-/-)) mice in vivo and in mouse peritoneal macrophages in vitro. In atherosclerosis-prone apoE(-/-) mice, apocynin suppressed the progression of atherosclerosis, decreased 4-hydroxynonenal-positive area in atherosclerotic lesions, and mRNA expression of monocyte chemoattractant protein-1 (MCP-1) and interleukin-6 (IL-6) in aorta. In mouse peritoneal macrophages, apocynin suppressed the Ox-LDL-induced ROS generation, mRNA expression of MCP-1, IL-6 and granulocyte/macrophage colony-stimulating factor, and cell proliferation. Moreover, immunohistochemical studies revealed that apocynin decreased the number of proliferating cell nuclear antigen-positive macrophages in atherosclerotic lesions of apoE(-/-) mice. These results suggested that apocynin suppressed the formation of atherosclerotic lesions, at least in part, by inactivation of macrophages. Therefore, apocynin may be a potential therapeutic material to prevent the progression of atherosclerosis.

Regulation of TNFα converting enzyme activity in visceral adipose tissue of obese mice.

Tumor necrosis factor α (TNFα) is a pro-inflammatory cytokine and one of the major mediators of obesity-induced insulin resistance. TNFα is generated through TNFα converting enzyme (TACE)-mediated cleavage of the transmembrane precursor pro-TNFα. Inhibition of TACE resulted in the improvement in glucose and insulin levels in diabetic animals, suggesting a crucial role of TACE activity in glucose metabolism. However, the regulation of TACE activity in insulin-sensitive tissues has not been fully determined. This study aimed to investigate the impact of TACE in insulin-sensitive tissues in the early stage of the development of obesity. C57BL6 mice were fed standard chow (B6-SC) or high-fat/high-sucrose diet (B6-HF/HS). KK-Ay mice were fed SC ad libitum (Ay-AL) or fed reduced amounts of SC (caloric restriction (CR); Ay-CR). As control for Ay-AL, KK mice fed SC ad libitum (KK-AL) were used. TACE activity in visceral adipose tissue (VAT), but not in liver or skeletal muscle, was significantly elevated in B6-HF/HS and Ay-AL compared with B6-SC and KK-AL, respectively. Phosphorylation of JNK and p38MAPK, but not ERK, in VATs from B6-HF/HS and Ay-AL was also significantly elevated. Ay-CR showed significantly lower TACE, JNK and p38MAPK activities in VAT and serum TNFα level compared with those of Ay-AL. In contrast, intraperitoneal injection of TNFα activated TACE, JNK and p38MAPK activities in VAT in KK mice. In conclusion, during the development of obesity, TACE activity is elevated only in VAT, and CR effectively reduced TACE activity and TACE-mediated pro-TNFα shedding in VAT.

Effects of combination therapy with vildagliptin and valsartan in a mouse model of type 2 diabetes.

BACKGROUND: Dipeptidyl peptidase-4 (DPP-4) inhibitors modulate incretin hormones and exert anti-diabetic effects in type 2 diabetes mellitus. Treatment with angiotensin II type 1 receptor blockers (ARB) is a proven successful intervention for hypertension with type 2 diabetes. The present study investigated the combined effects of the DPP-4 inhibitor vildagliptin and the ARB valsartan in a mouse model of type 2 diabetes. METHODS: C57BL/6 J mice fed with high-fat diet (HFD) or db/db mice were treated with placebo, phloridzin (PHZ), vildagliptin alone (ViL), valsartan alone (VaL) or ViL with VaL (ViLVaL) for 8 weeks. RESULTS: Glucose metabolism was improved in response to PHZ, ViL and ViLVaL in both HFD and db/db mice. Upon glucose challenge, ViLVaL showed the greatest suppression of blood glucose excursions, with increased insulin secretion, in db/db mice. ViLVaL treatment also showed an improvement of insulin sensitivity in db/db mice. Serum inflammatory cytokines were significantly decreased, and adiponectin was highest, in the ViLVaL group. ViLVaL improved insulin signaling and attenuated stress signaling in liver with amelioration of hepatic steatosis due to activated fatty acid oxidation in db/db mice. Furthermore, immunohistochemical analysis of the pancreas revealed that the combination treatment resulted in an increased expression of insulin and PDX-1, and increased insulin content. CONCLUSIONS: The combination therapy of ViL and VaL improves both pancreatic beta-cell function and insulin sensitivity, with a reduction of the inflammatory and cell stress milieu in mouse models of T2DM. Our results suggest that this combination therapy exerts additive or even synergistic benefits to treat T2DM.

Association between circulating leukocyte subtype counts and carotid intima-media thickness in Japanese subjects with type 2 diabetes.

BACKGROUND: An increased leukocyte count is an independent risk factor for cardiovascular events, but the association between leukocyte subtype counts and carotid atherosclerosis in patients with diabetes has not been determined. We therefore investigated the correlation between leukocyte subtype counts and intima-media thickness of the common carotid artery (CCA-IMT) in subjects with type 2 diabetes. METHODS: This cross-sectional study involved 484 in-patients with type 2 diabetes (282 males and 202 females), who were hospitalized for glycemic control and underwent carotid ultrasonography at Kumamoto University Hospital between 2005 and 2011. Mean and maximum CCA-IMT was measured by high-resolution B-mode ultrasonography. RESULTS: Univariate analyses revealed that mean CCA-IMT was positively correlated with age, systolic blood pressure, brachial-ankle pulse wave velocity (PWV), urinary albumin excretion and duration of diabetes, but was negatively correlated with diastolic blood pressure and fasting plasma glucose. Maximum CCA-IMT was positively and negatively correlated with the same factors as mean CCA-IMT except for fasting plasma glucose. Mean CCA-IMT was positively correlated with total leukocyte (r = 0.124, p = 0.007), monocyte (r = 0.373, p < 0.001), neutrophil (r = 0.139, p = 0.002) and eosinophil (r = 0.107, p = 0.019) counts. Maximum CCA-IMT was positively correlated with total leukocyte (r = 0.154, p < 0.001), monocyte (r = 0.398, p < 0.001), neutrophil (r = 0.152, p < 0.001) and basophil counts (r = 0.102, p = 0.027). Multiple regression analyses showed that monocyte count, age and PWV were significant and independent factors associated with mean CCA-IMT (adjusted R2 = 0.239, p < 0.001), and that monocyte count, age and urinary albumin excretion were significant and independent factors associated with maximum CCA-IMT (adjusted R2 = 0.277, p < 0.001). CONCLUSIONS: Monocyte counts were positively correlated with both mean CCA-IMT and maximum CCA-IMT in patients with type 2 diabetes. Monocyte count may be a useful predictor of macrovascular complications in patients with type 2 diabetes. TRIAL REGISTRATION: Trial registry no: UMIN000003526.

Efficacy and safety of sitagliptin as add-on therapy on glycemic control and blood glucose fluctuation in Japanese type 2 diabetes subjects ongoing with multiple daily insulin injections therapy.

To assess the efficacy and safety of adding sitagliptin, an oral dipeptidyl peptidase-4 inhibitor, in subjects with type 2 diabetes inadequately controlled with multiple daily insulin injections therapy (MDI). HbA1c, 1,5-anhydroglucitol (1,5-AG), body mass index (BMI), insulin doses, six-point self-measured plasma glucose (SMPG) profiles were assessed before, after 12 weeks, and after 24 weeks of MDI with 50 mg/day of sitagliptin in 40 subjects with type 2 diabetes. Safety endpoints included hypoglycemia and any adverse events. HbA1c significantly decreased during the first 12 weeks ( -0.64±0.60%), and was sustained over 24 weeks ( -0.69±0.85%). 1,5-AG increased significantly from 7.5±4.5 µg/mL at baseline to 9.6±5.5 µg/mL after 24 weeks. The bolus insulin dose at 12 weeks was decreased, and the mean plasma glucose, the SD of daily glucose, M-value, and the mean amplitude of glycemic excursions (MAGE) also decreased significantly as compared with baseline values. BMI and frequency of hypoglycemia were not changed significantly. Univariate linear regression analyses revealed that % change in HbA1c was significantly associated with BMI, and % changes in the indexes of glycemic instability (SD of daily glucose and MAGE) were significantly associated with age. In conclusion, adding sitagliptin to MDI significantly improved glycemic control and decreased the daily glucose fluctuation in subjects with type 2 diabetes inadequately controlled with MDI, without weight gain or an increase in the incidence of hypoglycemia. This trial was registered with UMIN (no. UMIN000010157).

Telmisartan exerts antiatherosclerotic effects by activating peroxisome proliferator-activated receptor-γ in macrophages.

OBJECTIVE: Telmisartan, an angiotensin type I receptor blocker (ARB), protects against the progression of atherosclerosis. Here, we investigated the molecular basis of the antiatherosclerotic effects of telmisartan in macrophages and apolipoprotein E-deficient mice. METHODS AND RESULTS: In macrophages, telmisartan increased peroxisome proliferator-activated receptor-γ (PPARγ) activity and PPAR ligand-binding activity. In contrast, 3 other ARBs, losartan, valsartan, and olmesartan, did not affect PPARγ activity. Interestingly, high doses of telmisartan activated PPARα in macrophages. Telmisartan induced the mRNA expression of CD36 and ATP-binding cassette transporters A1 and G1 (ABCA1/G1), and these effects were abrogated by PPARγ small interfering RNA. Telmisartan, but not other ARBs, inhibited lipopolysaccharide-induced mRNA expression of monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α, and these effects were abrogated by PPARγ small interfering RNA. Moreover, telmisartan suppressed oxidized low-density lipoprotein-induced macrophage proliferation through PPARγ activation. In apolipoprotein E(-/-) mice, telmisartan increased the mRNA expression of ABCA1 and ABCG1, decreased atherosclerotic lesion size, decreased the number of proliferative macrophages in the lesion, and suppressed MCP-1 and tumor necrosis factor-α mRNA expression in the aorta. CONCLUSION: Telmisartan induced ABCA1/ABCG1 expression and suppressed MCP-1 expression and macrophage proliferation by activating PPARγ. These effects may induce antiatherogenic effects in hypertensive patients.

[RAAS and insulin resistance].

The role of the renin-angiotensin-aldosterone system (RAAS) on the development of insulin resistance and type 2 diabetes (T2DM) is an area of growing interest. Most of the deleterious actions of the RAAS on insulin signals appear to be mediated through activation of the serine/threonine kinase, oxidative stress and tissue-inflammation in insulin-sensitive organs. Both experimental and clinical studies demonstrated that angiotensin II (Ang II) and aldosterone could play a role in the development of insulin resistance, diabetes and cardiovascular diseases. Large randomized clinical trials revealed that blockade of the RAAS with either angiotensin I converting enzyme inhibitors or AT1 receptor blockers results in decreased T2DM incidence, with a minor attenuation of markers for insulin resistance. This review focuses on the role of RAAS in the pathogenesis of insulin resistance, as well as on clinical relevance of RAAS blockade in the prevention and treatment of the metabolic syndrome and pre-diabetes.

Caloric restriction decreases ER stress in liver and adipose tissue in ob/ob mice.

Endoplasmic reticulum (ER) stress plays a crucial role in the development of insulin resistance and diabetes. Although caloric restriction (CR) improves obesity-related disorders, the effects of CR on ER stress in obesity remain unknown. To investigate how CR affects ER stress in obesity, ob/ob mice were assigned to either ad libitum (AL) (ob-AL) or CR (ob-CR) feeding (2 g food/day) for 1-4 weeks. The body weight (BW) of ob-CR mice decreased to the level of lean AL-fed littermates (lean-AL) within 2 weeks. BW of lean-AL and ob-CR mice was less than that of ob-AL mice. The ob-CR mice showed improved glucose tolerance and hepatic insulin action compared with ob-AL mice. Levels of ER stress markers such as phosphorylated PKR-like ER kinase (PERK) and eukaryotic translation initiation factor 2α and the mRNA expression of activating transcription factor 4 were significantly higher in the liver and epididymal fat from ob-AL mice compared with lean-AL mice. CR for 2 and 4 weeks significantly reduced all of these markers to less than 35% and 50%, respectively, of the levels in ob-AL mice. CR also significantly reduced the phosphorylation of insulin receptor substrate (IRS)-1 and c-Jun NH(2)-terminal kinase (JNK) in ob/ob mice. The CR-mediated decrease in PERK phosphorylation was similar to that induced by 4-phenyl butyric acid, which reduces ER stress in vivo. In conclusion, CR reduced ER stress and improved hepatic insulin action by suppressing JNK-mediated IRS-1 serine-phosphorylation in ob/ob mice.

Nifedipine induces peroxisome proliferator-activated receptor-gamma activation in macrophages and suppresses the progression of atherosclerosis in apolipoprotein E-deficient mice.

OBJECTIVE: Nifedipine, an L-type calcium channel blocker, protects against the progression of atherosclerosis. We investigated the molecular basis of the antiatherosclerotic effect of nifedipine in macrophages and apolipoprotein E-deficient mice. METHODS AND RESULTS: In macrophages, nifedipine increased peroxisome proliferator-activated receptor-gamma (PPARgamma) activity without increasing PPARgamma-binding activity. Amlodipine, another L-type calcium channel blocker, and 1,2-bis-(o-aminophenoxy)-ethane-N,N,-N',N'-tetraacetic acid tetraacetoxy-methyl ester (BAPTA-AM), a calcium chelator, decreased PPARgamma activity, suggesting that nifedipine does not activate PPARgamma via calcium channel blocker activity. Inactivation of extracellular signal-regulated kinase 1/2 suppressed PPARgamma2-Ser112 phosphorylation and induced PPARgamma activation. Nifedipine suppressed extracellular signal-regulated kinase 1/2 activation and PPARgamma2-Ser112 phosphorylation, and mutating PPARgamma2-Ser112 to Ala abrogated nifedipine-mediated PPARgamma activation. These results suggested that nifedipine inhibited extracellular signal-regulated kinase 1/2 activity and PPARgamma2-Ser112 phosphorylation, leading to PPARgamma activation. Nifedipine inhibited lipopolysaccharide-induced monocyte chemoattractant protein-1 expression and induced ATP-binding cassette transporter A1 mRNA expression, and these effects were abrogated by small interfering RNA for PPARgamma. Furthermore, in apolipoprotein E-deficient mice, nifedipine treatment decreased atherosclerotic lesion size, phosphorylation of PPARgamma2-Ser112 and extracellular signal-regulated kinase 1/2, and monocyte chemoattractant protein-1 mRNA expression and increased ATP-binding cassette transporter A1 expression in the aorta. CONCLUSIONS: Nifedipine unlike amlodipine inhibits PPARgamma-Ser phosphorylation and activates PPARgamma to suppress monocyte chemoattractant protein-1 expression and induce ATP-binding cassette transporter A1 expression in macrophages. These effects may induce antiatherogenic effects in hypertensive patients.

Hypoglycemia Induces Mitochondrial Reactive Oxygen Species Production Through Increased Fatty Acid Oxidation and Promotes Retinal Vascular Permeability in Diabetic Mice.

Aims: Hypoglycemia is associated with increased reactive oxygen species (ROS) production and vascular events. We have previously reported that low-glucose (LG) conditions induce mitochondrial ROS (mtROS) production in aortic endothelial cells (ECs). However, the mechanism by which hypoglycemia promotes diabetic retinopathy (DR) is unclear. Blood-retinal barrier (BRB) disruption occurs in the early stages of DR. We hypothesized that the mechanisms underlying hypoglycemia-induced DR are associated with BRB breakdown due to mtROS generation during hypoglycemia. Here, we aimed to determine whether hypoglycemia exacerbated mtROS production and induced BRB disruption. Results: We observed that hypoglycemia induced mtROS production by increasing fatty acid oxidation (FAO), which was suppressed by overexpression of mitochondrial-specific manganese superoxide dismutase (MnSOD) in retinal ECs. Furthermore, FAO blockade decreased the hypoglycemia-induced mtROS production. Recurrent hypoglycemia increased albumin leak in diabetic mice retina, which was suppressed in diabetic vascular endothelial cell-specific MnSOD transgenic (eMnSOD-Tg) mice. Pharmacological FAO blockade also reduced mtROS production, reduced vascular endothelial growth factor (VEGF) production during hypoglycemia, and prevented retinal vascular permeability in diabetic mice. MnSOD overexpression or carnitine palmitoyltransferase I (CPT1) blockade suppressed vascular endothelial-cadherin phosphorylation under LG in retinal ECs. Innovation and Conclusion: Reduction of mtROS and VEGF production via pharmacological FAO and/or CPT1 blockade may prevent hypoglycemia-induced worsening of DR.

Rapid and dramatic glucose-lowering effect of bromocriptine in an inadequately controlled type 2 diabetes patient with prolactinoma.

Dopamine receptor agonists are typically used to treat Parkinson's disease and certain pituitary tumors, such as prolactinoma or a growth hormone-producing tumor. A 53-year-old woman with a history of prolactinoma was referred to Kumamoto University Hospital (Kumamoto, Japan) with poorly controlled type 2 diabetes. Her glycated hemoglobin and serum prolactin levels were increased (8.8% and 160.3 ng/mL, respectively). Bromocriptine, a dopamine D2 receptor agonist, was administered to reduce her serum prolactin level. Because bromocriptine-QR (quick release) has been approved for the treatment of type 2 diabetes mellitus in the USA, a continuous glucose monitoring system, FreeStyle Libre Pro, was utilized to examine the effect of bromocriptine on glycemic control. After the initial administration of bromocriptine, glucose levels were rapidly and dramatically ameliorated, and the time in range (70-180 mg/dL) improved from <50% to >90% between 1 week before and after the initial administration of bromocriptine.