Bill Haynes

BACKGROUND: There is an unmet need for a safer and more effective treatment for obesity. This study assessed the effects of licogliflozin, a dual inhibitor of sodium‐glucose co‐transporter (SGLT) 1/2, on body weight, metabolic parameters and incretin hormones in patients with type 2 diabetes mellitus (T2DM) and/or obesity.

METHODS: Patients with obesity (BMI, 35‐50 kg/m2) were enrolled into a 12‐week study (N = 88; licogliflozin 150 mg q.d.). Patients with T2DM were enrolled into a… Show more

BACKGROUND: There is an unmet need for a safer and more effective treatment for obesity. This study assessed the effects of licogliflozin, a dual inhibitor of sodium‐glucose co‐transporter (SGLT) 1/2, on body weight, metabolic parameters and incretin hormones in patients with type 2 diabetes mellitus (T2DM) and/or obesity.

METHODS: Patients with obesity (BMI, 35‐50 kg/m2) were enrolled into a 12‐week study (N = 88; licogliflozin 150 mg q.d.). Patients with T2DM were enrolled into a second, two‐part study, comprising a single‐dose cross‐over study (N = 12; 2.5 − 300 mg) and a 14‐day dosing study (N = 30; 15 mg q.d).

RESULTS: Treatment with licogliflozin 150 mg q.d. for 12 weeks in patients with obesity significantly reduced body weight by 5.7% vs placebo (P < 0.001) and improved metabolic parameters such as significantly reduced postprandial glucose excursion (21%; P < 0.001), reduced insulin levels (80%; P < 0.001) and increased glucagon (59%; P < 0.001). In patients with T2DM, a single dose of licogliflozin 300 mg in the morning prior to an oral glucose tolerance test (OGTT) remarkably reduced glucose excursion by 93% (P < 0.001; incremental AUC0‐4h) and suppressed insulin by 90% (P < 0.01; incremental AUC0‐4h). Treatment with licogliflozin 15 mg q.d. for 14 days reduced 24‐hour average glucose levels by 26% (41 mg/dL; P < 0.001) and increased UGE24 to 100 g (P < 0.001) in patients with T2DM. Treatment with licogliflozin was generally safe and well tolerated. Diarrhea (increased numbers of loose stool) was the most common adverse event in all studies (90% with licogliflozin vs 25% with placebo in the 12‐week study)...

CONCLUSION: Licogliflozin treatment (1‐84 days) leads to significant weight loss and favourable changes in a variety of metabolic parameters and incretin hormones. Dual inhibition of SGLT1/2 with licogliflozin in the gut and kidneys is an attractive strategy for treating obesity and diabetes. Show less

Objective - Nonalcoholic fatty liver disease (NAFLD) is becoming a leading cause of advanced chronic liver disease. The progression of NAFLD, including nonalcoholic steatohepatitis (NASH), has a strong genetic component, and the most robust contributor is the patatin-like phospholipase domain-containing 3 (PNPLA3) rs738409 encoding the 148M protein sequence variant. We hypothesized that suppressing the expression of the PNPLA3 148M mutant protein would exert a beneficial effect on the entire… Show more

Objective - Nonalcoholic fatty liver disease (NAFLD) is becoming a leading cause of advanced chronic liver disease. The progression of NAFLD, including nonalcoholic steatohepatitis (NASH), has a strong genetic component, and the most robust contributor is the patatin-like phospholipase domain-containing 3 (PNPLA3) rs738409 encoding the 148M protein sequence variant. We hypothesized that suppressing the expression of the PNPLA3 148M mutant protein would exert a beneficial effect on the entire spectrum of NAFLD.

Methods - We examined the effects of liver-targeted GalNAc3-conjugated antisense oligonucleotide (ASO)-mediated silencing of Pnpla3 in a knock-in mouse model in which we introduced the human PNPLA3 I148M mutation.

Results - ASO-mediated silencing of Pnpla3 reduced liver steatosis (p = 0.038) in homozygous Pnpla3 148M/M knock-in mutant mice but not in wild-type littermates fed a steatogenic high-sucrose diet. In mice fed a NASH-inducing diet, ASO-mediated silencing of Pnpla3 reduced liver steatosis score and NAFLD activity score independent of the Pnpla3 genotype, while reductions in liver inflammation score (p = 0.018) and fibrosis stage (p = 0.031) were observed only in the Pnpla3 knock-in 148M/M mutant mice. These responses were accompanied by reduced liver levels of Mcp1 (p = 0.026) and Timp2 (p = 0.007) specifically in the mutant knock-in mice. This may reduce levels of chemokine attracting inflammatory cells and increase the collagenolytic activity during tissue regeneration.

Conclusion - This study provides the first evidence that a Pnpla3 ASO therapy can improve all features of NAFLD, including liver fibrosis, and suppress the expression of a strong innate genetic risk factor, Pnpla3 148M, which may open up a precision medicine approach in NASH. Show less

Objective: Dedifferentiation could explain reduced functional pancreatic β-cell mass in type 2 diabetes (T2D).

Methods: Here we model human β-cell dedifferentiation using growth factor stimulation in the human β-cell line, EndoC-βH1, and human pancreatic islets.

Results: Fibroblast growth factor 2 (FGF2) treatment reduced expression of β-cell markers, (INS, MAFB, SLC2A2, SLC30A8, and GCK) and activated ectopic expression of MYC, HES1, SOX9, and NEUROG3. FGF2-induced… Show more

Objective: Dedifferentiation could explain reduced functional pancreatic β-cell mass in type 2 diabetes (T2D).

Methods: Here we model human β-cell dedifferentiation using growth factor stimulation in the human β-cell line, EndoC-βH1, and human pancreatic islets.

Results: Fibroblast growth factor 2 (FGF2) treatment reduced expression of β-cell markers, (INS, MAFB, SLC2A2, SLC30A8, and GCK) and activated ectopic expression of MYC, HES1, SOX9, and NEUROG3. FGF2-induced dedifferentiation was time- and dose-dependent and reversible upon wash-out. Furthermore, FGF2 treatment induced expression of TNFRSF11B, a decoy receptor for RANKL and protected β-cells against RANKL signaling. Finally, analyses of transcriptomic data revealed increased FGF2 expression in ductal, endothelial, and stellate cells in pancreas from T2D patients, whereas FGFR1, SOX,9 and HES1 expression increased in islets from T2D patients.

Conclusions: We developed an FGF2-induced model of human β-cell dedifferentiation, identified new markers of dedifferentiation, and found evidence for increased pancreatic FGF2, FGFR1, and β-cell dedifferentiation in T2D. Show less

Human in vitro physiological models studying disease and drug treatment effects are urgently needed as more relevant tools to identify new drug targets and therapies. We have developed a human microfluidic two-organ-chip model to study pancreatic islet–liver cross-talk based on insulin and glucose regulation. We have established a robust co-culture of human pancreatic islet microtissues and liver spheroids maintaining functional responses up to 15 days in an insulin-free medium. Functional… Show more

Human in vitro physiological models studying disease and drug treatment effects are urgently needed as more relevant tools to identify new drug targets and therapies. We have developed a human microfluidic two-organ-chip model to study pancreatic islet–liver cross-talk based on insulin and glucose regulation. We have established a robust co-culture of human pancreatic islet microtissues and liver spheroids maintaining functional responses up to 15 days in an insulin-free medium. Functional coupling, demonstrated by insulin released from the islet microtissues in response to a glucose load applied in glucose tolerance tests on different days, promoted glucose uptake by the liver spheroids. Co-cultures maintained postprandial glucose concentrations in the circulation whereas glucose levels remained elevated in both single cultures. Thus, insulin secreted into the circulation stimulated glucose uptake by the liver spheroids, while the latter, in the absence of insulin, did not consume glucose as efficiently. As the glucose concentration fell, insulin secretion subsided, demonstrating a functional feedback loop between the liver and the insulin-secreting islet microtissues. Finally, inter-laboratory validation verified robustness and reproducibility. Further development of this model using tools inducing impaired glucose regulation should provide a unique in vitro system emulating human type 2 diabetes mellitus. Show less

Nitrite stores decrease after exercise in patients with peripheral artery disease (PAD) and diabetes represents decreased nitric oxide (NO) bioavailability that may contribute to endothelial dysfunction and limit exercise duration. The primary objective of this placebo-controlled study was the safety and tolerability of multiple doses of oral sodium nitrite in patients with PAD, predominantly with diabetes, over a period of 10 weeks. The primary efficacy endpoint was endothelial flow-mediated… Show more

Nitrite stores decrease after exercise in patients with peripheral artery disease (PAD) and diabetes represents decreased nitric oxide (NO) bioavailability that may contribute to endothelial dysfunction and limit exercise duration. The primary objective of this placebo-controlled study was the safety and tolerability of multiple doses of oral sodium nitrite in patients with PAD, predominantly with diabetes, over a period of 10 weeks. The primary efficacy endpoint was endothelial flow-mediated dilatation (FMD) and secondary efficacy endpoints included a 6-minute walk test and quality of life assessment. Of the 55 subjects, the most common side effects attributed to sodium nitrite were a composite of headache and dizziness occurring in 21% with the 40 mg dose and 44% with the 80 mg dose. There was no clinically significant elevation of methemoglobin. FMD non-significantly worsened in the placebo and 40 mg groups, but was stable in the 80 mg group. Diabetic patients receiving 80 mg had significantly higher FMD compared with the placebo and 40 mg groups. There was no significant change in 6-minute walk test or quality of life parameters over time compared to placebo. In conclusion, sodium nitrite therapy is well tolerated in patients with PAD. The possible clinical benefit of sodium nitrite should be studied in a larger and fully powered trial. Show less

Objective: Anxiety predicts cardiovascular events, though the mechanism remains unclear. We hypothesized that anxious symptoms will correlate with impaired resistance and conduit vessel function in participants aged 55–90 years.

Method: Anxious symptoms were measured with the Symptom Checklist-90-Revised in 89 participants with clinically diagnosed atherosclerotic cardiovascular disease and 54 healthy control participants. Vascular function was measured in conduit arteries using brachial… Show more

Objective: Anxiety predicts cardiovascular events, though the mechanism remains unclear. We hypothesized that anxious symptoms will correlate with impaired resistance and conduit vessel function in participants aged 55–90 years.

Method: Anxious symptoms were measured with the Symptom Checklist-90-Revised in 89 participants with clinically diagnosed atherosclerotic cardiovascular disease and 54 healthy control participants. Vascular function was measured in conduit arteries using brachial flow-mediated dilatation (FMD) and in forearm resistance vessels (FRV) using intra-arterial drug administration and plethysmography.

Results: Anxious symptoms were not associated with FMD in either group. Participants with atherosclerosis exhibited significant inverse associations of anxious symptoms with FRV dilatation (β for acetylcholine =−0.302, p=0.004). Adjustment for medication, risk factors and depressive symptoms did not alter the association between anxiety and FRV dysfunction, except for BMI (anxiety β=−0.175, p=0.060; BMI β=−0.494, p<0.001). While BMI was more strongly associated with FRV function than anxiety, combined BMI and anxiety accounted for more variance in FRV function than either separately. Control participants showed no association of anxiety with FRV function.

Conclusion: Anxiety is uniquely and substantially related to poorer resistance vessel function (both endothelial and vascular smooth muscle function) in individuals with atherosclerosis. These relationships were independent of medication, depression and cardiovascular risk factors, with the exception of BMI. These findings support the concept that anxiety potentially increases vascular events through worsening of vascular function in atherosclerotic disease. Show less

The incidence of myocardial infarctions and influenza follow similar seasonal patterns. To determine if acute myocardial infarctions (AMIs) and ischemic strokes are associated with influenza activity, we built time series models using data from the Nationwide Inpatient Sample. In these models, we used influenza activity to predict the incidence of AMI and ischemic stroke. We fit national models as well as models based on four geographic regions and five age groups. Across all models, we found… Show more

The incidence of myocardial infarctions and influenza follow similar seasonal patterns. To determine if acute myocardial infarctions (AMIs) and ischemic strokes are associated with influenza activity, we built time series models using data from the Nationwide Inpatient Sample. In these models, we used influenza activity to predict the incidence of AMI and ischemic stroke. We fit national models as well as models based on four geographic regions and five age groups. Across all models, we found consistent significant associations between AMIs and influenza activity, but not between ischemic strokes and influenza. Associations between influenza and AMI increased with age, were greatest in those over 80 years old, and were present in all geographic regions. In addition, the natural experiment provided by the second wave of the influenza pandemic in 2009 provided further evidence of the relationship between influenza and AMI, because both series peaked in the same non-winter month. Show less

The role of neurohumoral factors in the sodium retention of nephrotic syndrome is controversial. We report a case with abrupt onset of severe nephrotic-range proteinuria and hypoalbuminemia due to membranous glomerulonephritis that was associated with renal salt wasting and hypovolemia without edema. Further evaluation showed hypoaldosteronism, hyporeninemia, and primary autonomic failure principally affecting the sympathetic nervous system, determined by the Valsalva maneuver. Administration… Show more

The role of neurohumoral factors in the sodium retention of nephrotic syndrome is controversial. We report a case with abrupt onset of severe nephrotic-range proteinuria and hypoalbuminemia due to membranous glomerulonephritis that was associated with renal salt wasting and hypovolemia without edema. Further evaluation showed hypoaldosteronism, hyporeninemia, and primary autonomic failure principally affecting the sympathetic nervous system, determined by the Valsalva maneuver. Administration of exogenous mineralocorticoid and oral salt caused edema and accelerated hypertension. The severe hypoaldosteronism likely was due to use of the angiotensin-converting enzyme inhibitor lisinopril, and it improved after this drug treatment was discontinued. The nephrotic proteinuria resolved after treatment with cyclosporine and prednisone, but the primary autonomic failure with hyporeninemic hypoaldosteronism persisted. The case shows that intratubular factors activated by nephrotic proteinuria are not sufficient to produce sodium retention in the absence of aldosterone and an intact sympathetic nervous system. Show less

OBJECTIVE—Leptin is an adipocyte hormone that plays a major role in energy balance. Leptin receptors in the hypothalamus are known to signal via distinct mechanisms, including signal transducer and activator of transcription-3 (STAT3) and phosphoinositol-3 kinase (PI 3-kinase). Here, we tested the hypothesis that extracellular signal–regulated kinase (ERK) is mediating leptin action in the hypothalamus.

RESEARCH DESIGN AND METHODS—Biochemical, pharmacological, and physiological… Show more

OBJECTIVE—Leptin is an adipocyte hormone that plays a major role in energy balance. Leptin receptors in the hypothalamus are known to signal via distinct mechanisms, including signal transducer and activator of transcription-3 (STAT3) and phosphoinositol-3 kinase (PI 3-kinase). Here, we tested the hypothesis that extracellular signal–regulated kinase (ERK) is mediating leptin action in the hypothalamus.

RESEARCH DESIGN AND METHODS—Biochemical, pharmacological, and physiological approaches were combined to characterize leptin activation of ERK in the hypothalamus in rats.

RESULTS—Leptin activates ERK1/2 in a receptor-mediated manner that involves JAK2. Leptin-induced ERK1/2 activation was restricted to the hypothalamic arcuate nucleus. Pharmacological blockade of hypothalamic ERK1/2 reverses the anorectic and weight-reducing effects of leptin. The pharmacological antagonists of ERK1/2 did not attenuate leptin-induced activation of STAT3 or PI 3-kinase. Blockade of ERK1/2 abolishes leptin-induced increases in sympathetic nerve traffic to thermogenic brown adipose tissue (BAT) but does not alter the stimulatory effects of leptin on sympathetic nerve activity to kidney, hindlimb, or adrenal gland. In contrast, blockade of PI 3-kinase prevents leptin-induced sympathetic activation to kidney but not to BAT, hindlimb, or adrenal gland.

CONCLUSIONS—Our findings indicate that hypothalamic ERK plays a key role in the control of food intake, body weight, and thermogenic sympathetic outflow by leptin but does not participate in the cardiovascular and renal sympathetic actions of leptin. Show less

Background: Bipolar disorder is associated with excess cardiovascular mortality. We hypothesized outpatients with bipolar disorder would exhibit excess cardiovascular risk factors, particularly among prevalent users of the second generation antipsychotics associated with weight gain and valproic acid derivatives.

Methods: This chart review of 217 patients with bipolar disorder examined cardiovascular risk factors of the metabolic syndrome. We also evaluated if certain medications were… Show more

Background: Bipolar disorder is associated with excess cardiovascular mortality. We hypothesized outpatients with bipolar disorder would exhibit excess cardiovascular risk factors, particularly among prevalent users of the second generation antipsychotics associated with weight gain and valproic acid derivatives.

Methods: This chart review of 217 patients with bipolar disorder examined cardiovascular risk factors of the metabolic syndrome. We also evaluated if certain medications were cross-sectionally associated with metabolic syndrome.

Results: Fifty-six patients were not weighed and many did not have available lipid profiles. Over three-quarters of those with available data (N=161) were overweight or obese (body-mass index ≥ 25) and nearly half were obese (body-mass index ≥ 30). A prevalence exceeding general population estimates was also observed for hypertriglyceridemia, elevated blood pressure/hypertension, and elevated fasting glucose/diabetes. Among those with all requisite data (n = 60), over 50% met criteria for National Cholesterol Education Program defined metabolic syndrome, nearly double the expected prevalence. A trend toward greater prevalence of metabolic syndrome among prevalent users of the second generation antipsychotics associated with weight gain was observed.

Conclusions: Obesity and the metabolic syndrome were common in patients with bipolar disorder. These patients may be under-evaluated for cardiovascular risk and warrant screening and early intervention. Show less

Background: Endothelin-1 (ET-1) has been implicated in the pathophysiology of chronic kidney disease (CKD) and ET receptor blockade has shown renoprotective effects in animals. We examined the haemodynamic and renal effects of an ET receptor antagonist, TAK-044, in patients with CKD.

Methods: Seven patients with CKD (MAP 103 mmHg; Creatinine 3.5 mg/dl) received three 15 min IV infusions, each separated by at least 7 days, of either placebo or TAK-044 (100 or 750 mg) in a randomized… Show more

Background: Endothelin-1 (ET-1) has been implicated in the pathophysiology of chronic kidney disease (CKD) and ET receptor blockade has shown renoprotective effects in animals. We examined the haemodynamic and renal effects of an ET receptor antagonist, TAK-044, in patients with CKD.

Methods: Seven patients with CKD (MAP 103 mmHg; Creatinine 3.5 mg/dl) received three 15 min IV infusions, each separated by at least 7 days, of either placebo or TAK-044 (100 or 750 mg) in a randomized, double blind crossover study. Systemic and renal haemodynamics, and plasma immunoreactive ET-1, big ET-1 and C-terminal fragment concentrations, were determined.

Results: Compared with placebo, TAK-044 reduced mean arterial pressure (MAP) (100 mg: 7.4 ± 1.9 mmHg, 750 mg: 8.4 ± 2.3 mmHg, P < 0.01) and systemic vascular resistance index (100 mg: 650 ± 140 dyne.s.cm −5 .m −2 , 750 mg: 829 ± 141 dyne.s.cm −5 .m −2 , P < 0.01) at both doses. TAK-044 increased cardiac index and heart rate to a similar degree at both doses. With regards to renal haemodynamics, TAK-044 had no significant effect on the glomerular filtration rate at either dose but tended to increase renal plasma flow (100 mg: 9.6 ± 5.0 ml/min, 750 mg: 25.3 ± 19.5 ml/min) and decreased the effective filtration fraction (100 mg: 3.6 ± 1.1%, 750 mg: 4.7 ± 1.7%, P < 0.01), in a dose-dependent manner. TAK-044 had no significant effect on sodium or lithium clearance, or on fractional excretion of sodium and lithium. Plasma ET-1 concentrations rose more than two-fold after 750 mg TAK-044 while big ET-1 and C-terminal fragment concentrations were unchanged.

Conclusions: These findings suggest an important role for ET-1 in controlling systemic and renal haemodynamics in patients with CKD. The antihypertensive and potentially renoprotective actions of ET receptor antagonists shown in this study may prove useful in slowing the progression of CKD. Clinical trials are now needed to address these key questions for CKD. Show less

Leptin is an adipocyte-derived hormone that plays a key role in the regulation of body weight through its actions on appetite and metabolism. Leptin also increases sympathetic nerve activity (SNA) and blood pressure. We tested the hypothesis that diet-induced obesity is associated with resistance to the metabolic actions of leptin but preservation of its renal SNA and arterial pressure effects, leading to hypertension. Mice were fed a high-fat diet for 10 weeks to induce moderate obesity. The… Show more

Leptin is an adipocyte-derived hormone that plays a key role in the regulation of body weight through its actions on appetite and metabolism. Leptin also increases sympathetic nerve activity (SNA) and blood pressure. We tested the hypothesis that diet-induced obesity is associated with resistance to the metabolic actions of leptin but preservation of its renal SNA and arterial pressure effects, leading to hypertension. Mice were fed a high-fat diet for 10 weeks to induce moderate obesity. The decrease in food intake and body weight induced by intraperitoneal or intracerebroventricular leptin was significantly attenuated in the obese mice. Regional SNA responses to leptin were differentially altered in diet-induced obese mice. Renal SNA response to leptin was preserved, whereas lumbar and brown adipose tissue SNA responses were attenuated in obese mice. Radiotelemetric arterial pressure was ∼10 mmHg higher in obese mice. Furthermore, the increase in arterial pressure in response to long-term (12 days) leptin treatment was preserved in obese mice. Thus, mice with diet-induced obesity exhibit circulating hyperleptinemia and resistance to the metabolic actions of leptin. However, there is preservation of the renal sympathetic and arterial pressure responses to leptin, which represent a potential mechanism for the adverse cardiovascular consequences of obesity. Show less

Adipose tissue represents an important source of angiotensinogen (AGT). We investigated the effect of obesity induced by a high-fat diet on the expression of mouse (mAGT) and human AGT (hAGT) genes in liver, kidney, and heart and different adipose depots in normal mice (C57BL/6J), and in transgenic mice expressing the hAGT gene under the control of its own promoter. Mice were fed a high-fat diet (45% kcal) or normal chow (10% kcal) for 10 and 20 wk. The expression of mAGT and hAGT mRNA was… Show more

Adipose tissue represents an important source of angiotensinogen (AGT). We investigated the effect of obesity induced by a high-fat diet on the expression of mouse (mAGT) and human AGT (hAGT) genes in liver, kidney, and heart and different adipose depots in normal mice (C57BL/6J), and in transgenic mice expressing the hAGT gene under the control of its own promoter. Mice were fed a high-fat diet (45% kcal) or normal chow (10% kcal) for 10 and 20 wk. The expression of mAGT and hAGT mRNA was quantified using an RNAse protection assay. Mice on the high-fat diet exhibited increased weight, fat mass, and plasma leptin. Expression of mAGT or hAGT genes was not affected by high-fat diet in nonadipose tissues, brown adipose tissue, or subcutaneous white fat. In contrast, high-fat diet increased both mAGT and hAGT gene expression in visceral adipose depots (omental, reproductive, and perirenal fat). Thus obesity-induced by a high-fat diet is associated with a tissue-specific increased expression of both mouse and human AGT genes in intra-abdominal adipose tissue. Our findings also suggest that 1.2 kb of regulatory sequences present in the hAGT transgene are sufficient to transcriptionally respond to a high-fat diet in an adipose-specific and depot-specific manner. Show less

The action of insulin in the central nervous system produces sympathetic nervous system activation (also called sympathoactivation), although the neuronal intracellular mechanisms that mediate this are unclear. We hypothesized that PI3K and MAPK, the major pathways involved in insulin receptor signaling, mediate sympathetic nerve responses to insulin. Intracerebroventricular administration of insulin in rat increased multifiber sympathetic nerve activity to the hindlimb, brown adipose tissue… Show more

The action of insulin in the central nervous system produces sympathetic nervous system activation (also called sympathoactivation), although the neuronal intracellular mechanisms that mediate this are unclear. We hypothesized that PI3K and MAPK, the major pathways involved in insulin receptor signaling, mediate sympathetic nerve responses to insulin. Intracerebroventricular administration of insulin in rat increased multifiber sympathetic nerve activity to the hindlimb, brown adipose tissue (BAT), adrenal gland, and kidney. Ex vivo biochemical studies of mediobasal hypothalamic tissue revealed that insulin stimulated the association of insulin receptor substrate–1 with the p85α subunit of PI3K and also tyrosine phosphorylation of p42 and p44 subunits of MAPK in the hypothalamus. In order to determine whether PI3K and/or MAPK were involved in insulin-mediated sympathoactivation, we tested the effect of specific inhibitors of PI3K (LY294002 and wortmannin) and MAPK (PD98059 and U0126) on regional sympathetic responses to insulin. Interestingly, regional sympathoactivation to insulin was differentially affected by blockade of PI3K and MAPK. Inhibition of PI3K specifically blocked insulin-induced sympathoactivation to the hindlimb, while inhibition of MAPK specifically blocked insulin-induced sympathoactivation to BAT. Sympathoactivation to corticotrophin-releasing factor, however, was not affected by inhibition of PI3K and MAPK. These data demonstrate that PI3K and MAPK are specific and regionally selective mediators of the action of insulin on the sympathetic nervous system. Show less

To better understand the relations among leptin, insulin, and body fat during the metabolic progression to diabetes and during drug monotherapy, metabolic parameters were examined in subjects classified as 1) type 2 diabetes; 2) impaired fasting glucose or mild diabetes mellitus; 3) nondiabetic, matched for body mass index (BMI); and 4) nonobese, nondiabetic. Diabetic subjects were also studied during no pharmacological treatment, after 3 months of randomization to metformin or glyburide, and… Show more

To better understand the relations among leptin, insulin, and body fat during the metabolic progression to diabetes and during drug monotherapy, metabolic parameters were examined in subjects classified as 1) type 2 diabetes; 2) impaired fasting glucose or mild diabetes mellitus; 3) nondiabetic, matched for body mass index (BMI); and 4) nonobese, nondiabetic. Diabetic subjects were also studied during no pharmacological treatment, after 3 months of randomization to metformin or glyburide, and after 3 months of cross-over to the opposite drug. Log leptin correlated more with percent body fat (slope, 0.042; confidence interval, 0.036–0.047; r2 = 0.826; P < 0.0001) than with total fat mass, percent truncal or nontruncal fat, or BMI. When normalized to percent fat, leptin did not differ by gender. Leptin normalized to percent fat was 35% less in untreated diabetes than that in BMI-matched controls (P < 0.001). Leptin normalized to percent fat was increased by 25% (P < 0.01) as a result of glyburide therapy compared with pretreatment values, but was unchanged by therapy with metformin. Across a spectrum of subjects with diabetes, impaired fasting glucose/mild diabetes, or BMI-matched nondiabetic controls, normalized leptin significantly correlated with glucose-induced insulin release, but not with insulin sensitivity. Our data suggest that plasma leptin is reduced in untreated type 2 diabetes probably as a consequence of reduced insulin secretion and that circulating leptin concentrations are differentially affected by monodrug therapy. Show less

Background: Cigarette smoking causes endothelial dysfunction, possibly through increased oxidant stress. The enzyme xanthine oxidase produces oxidative free radicals. We tested the hypothesis that xanthine oxidase contributes to endothelial dysfunction in cigarette smokers by administering the inhibitor allopurinol.

Methods and Results: Fourteen cigarette smokers (31±4 pack years) and 14 age- and sex-matched healthy non-smoking control subjects participated in a single-blinded… Show more

Background: Cigarette smoking causes endothelial dysfunction, possibly through increased oxidant stress. The enzyme xanthine oxidase produces oxidative free radicals. We tested the hypothesis that xanthine oxidase contributes to endothelial dysfunction in cigarette smokers by administering the inhibitor allopurinol.

Methods and Results: Fourteen cigarette smokers (31±4 pack years) and 14 age- and sex-matched healthy non-smoking control subjects participated in a single-blinded, randomized, 2-phase crossover study. All subjects had no other risk factors for atherosclerosis. Inhibition of xanthine oxidase was achieved by a single oral dose of 600 mg of allopurinol on the day of the study. Stimulated nitric oxide endothelial responses were assessed by forearm blood flow responses to intraarterial administration of acetylcholine and bradykinin 4 to 7 hours later; basal nitric oxide was assessed using the nitric oxide synthase inhibitor NG-monomethyl-l-arginine (L-NMMA); and nitroprusside was used to assess sensitivity to nitric oxide. Dilatation produced by acetylcholine was significantly less in smokers (254±57%) than healthy controls (390±55%) (P=0.009). Allopurinol reversed endothelial dysfunction in smokers (acetylcholine, 463±78%, P=0.001) without affecting responses in non-smokers (401±80%). Bradykinin responses were also impaired in smokers (P=0.003), and improved with allopurinol, though not significantly (P=0.06). Responses to nitroprusside and L-NMMA were not significantly different between smokers and controls and were not altered by allopurinol.

Conclusions: Smoking-induced endothelial dysfunction of resistance vessels is rapidly reversed with oral allopurinol. These data suggest that xanthine oxidase contributes importantly to endothelial dysfunction caused by cigarette smoking. Show less

Blood pressure is under close control of the autonomic nervous system, principally the sympathetic nervous system. Sympathetic nervous activity can rapidly increase arterial pressure by augmentation of cardiac output and increases in total peripheral resistance. However, quite large and sustained changes in sympathetic nerve activity are required to maintain higher arterial pressure solely through effects on the heart and resistance arterioles. In contrast, even small increases in sympathetic… Show more

Blood pressure is under close control of the autonomic nervous system, principally the sympathetic nervous system. Sympathetic nervous activity can rapidly increase arterial pressure by augmentation of cardiac output and increases in total peripheral resistance. However, quite large and sustained changes in sympathetic nerve activity are required to maintain higher arterial pressure solely through effects on the heart and resistance arterioles. In contrast, even small increases in sympathetic activity can stimulate renin release through activation of ß1 adrenoceptors, and promote renal sodium and water reabsorption through activation of α1 adrenoceptors. More marked renal sympathoactivation will result in renal vasoconstriction through activation of vascular smooth muscle a1 adrenoceptors, exacerbating renal sodium retention. These renal effects can result in a gradual but profound increase in arterial pressure. Given the important physiological role of the sympathetic nervous system in regulation of arterial pressure, it is plausible that increased sympathetic activity could contribute to increased arterial pressure in hypertension.
However, the role of the sympathetic nervous system in the development, maintenance or complications of human hypertension is still unclear. This partly reflects the technical difficulty in measuring the overall activity of a paracrine system that releases an unstable neurotransmitter which is rapidly cleared. New technologies, such as use of labeled norepinephrine kinetic techniques, positron emission tomography and microneurographic direct measurement of sympathetic nerve activity (SNA) have provided clarified some issues but posed further questions. This chapter reviews evidence that suggests increased sympathetic ac¬tivity in human hypertension, discuss putative mechanisms that may underlie sympathoactivation in hypertension, and highlight the potential significance of elevated sympathetic nerve traffic. Show less

To determine whether leptin has insulin sensitizing effects in normal rodents, we measured plasma glucose and insulin concentrations in male Sprague-Dawley rats treated with leptin or vehicle by continuous sc infusion for 48 h. In additional experiments, we examined the acute effect of iv leptin upon insulin sensitivity under conditions of clamped glycemia.

Subcutaneous leptin was administered at 10.0 and 1.0 μg/h. To avoid confounding effects of differences in food intake, both leptin-… Show more

To determine whether leptin has insulin sensitizing effects in normal rodents, we measured plasma glucose and insulin concentrations in male Sprague-Dawley rats treated with leptin or vehicle by continuous sc infusion for 48 h. In additional experiments, we examined the acute effect of iv leptin upon insulin sensitivity under conditions of clamped glycemia.

Subcutaneous leptin was administered at 10.0 and 1.0 μg/h. To avoid confounding effects of differences in food intake, both leptin- and vehicle-treated rats were fasted during the 48-h period of infusion. Infusion of leptin, 10 μg/h, significantly reduced both plasma glucose and insulin. Leptin, 1.0 μg/h, also decreased plasma glucose and insulin, although the effects on insulin did not achieve statistical significance. Leptin at either dose did not alter body weight or epididymal fat mass compared with vehicle treated controls. Leptin, 10 μg/h, decreased circulating insulin-like growth factor-1 levels. No differences in GLUT-4 content in either in brown or epididymal fat were observed as a result of leptin-treatment. Leptin, 10 μg/h, significantly decreased urine osmolality, increased water intake, and reduced renal potassium excretion compared with vehicle-infused rats. In additional rats, we measured the acute effect of iv leptin on insulin sensitivity determined as whole body glucose utilization during hyperinsulinemic glucose clamps performed at glucose targets of 60 and 90 mg/100 ml. Glucose utilization was increased by 29% during the last 135 min of glycemia clamped at 60 mg/100 ml (P < 0.05) and by 30% during the last 135 min of glycemia clamped at 90 mg/dl (P < 0.01) in rats infused with leptin compared with vehicle.

In summary, leptin increased insulin sensitivity in normal rats both under fasting conditions and in the presence of hyperinsulinemia at clamped glucose. These effects did not appear dependent on altered body weight.... Show less

Leptin is a peptide hormone produced by adipose tissue which acts centrally to decrease appetite and increase energy expenditure. Although leptin increases norepinephrine turnover in thermogenic tissues, the effects of leptin on directly measured sympathetic nerve activity to thermogenic and other tissues are not known. We examined the effects of intravenous leptin and vehicle on sympathetic nerve activity to brown adipose tissue, kidney, hindlimb, and adrenal gland in anesthetized… Show more

Leptin is a peptide hormone produced by adipose tissue which acts centrally to decrease appetite and increase energy expenditure. Although leptin increases norepinephrine turnover in thermogenic tissues, the effects of leptin on directly measured sympathetic nerve activity to thermogenic and other tissues are not known. We examined the effects of intravenous leptin and vehicle on sympathetic nerve activity to brown adipose tissue, kidney, hindlimb, and adrenal gland in anesthetized Sprague-Dawley rats. Intravenous infusion of mouse leptin over 3 h (total dose 10-1,000 microg/kg) increased plasma concentrations of immunoreactive murine leptin up to 50-fold. Leptin slowly increased sympathetic nerve activity to brown adipose tissue (+286+/-64% at 1,000 microg/kg; P = 0.002). Surprisingly, leptin infusion also produced gradual increases in renal sympathetic nerve activity (+228+/-63% at 1,000 microg/kg; P = 0.0008).The effect of leptin on sympathetic nerve activity was dose dependent, with a threshold dose of 100 microg/kg. Leptin also increased sympathetic nerve activity to the hindlimb (+287+/-60%) and adrenal gland (388+/-171%). Despite the increase in overall sympathetic nerve activity, leptin did not increase arterial pressure or heart rate. Leptin did not change plasma glucose and insulin concentrations. Infusion of vehicle did not alter sympathetic nerve activity. Obese Zucker rats, known to possess a mutation in the gene for the leptin receptor, were resistant to the sympathoexcitatory effects of leptin, despite higher achieved plasma leptin concentrations. These data demonstrate that leptin increases thermogenic sympathetic nerve activity and reveal an unexpected stimulatory effect of leptin on overall sympathetic nerve traffic. Show less

Background: The importance of endothelin-1 in chronic heart failure (CHF) is unclear. We therefore investigated the effects of endothelin-converting enzyme (ECE) inhibition and endothelin ETA receptor blockade in CHF patients treated with ACE inhibitors. We also compared the function of ETA and ETB receptors in healthy subjects and patients with CHF.

Methods and Results: Locally active doses of study drugs were infused into the nondominant brachial artery while forearm blood flow (FBF)… Show more

Background: The importance of endothelin-1 in chronic heart failure (CHF) is unclear. We therefore investigated the effects of endothelin-converting enzyme (ECE) inhibition and endothelin ETA receptor blockade in CHF patients treated with ACE inhibitors. We also compared the function of ETA and ETB receptors in healthy subjects and patients with CHF.

Methods and Results: Locally active doses of study drugs were infused into the nondominant brachial artery while forearm blood flow (FBF) was measured by venous occlusion plethysmography. In CHF patients (n=10), phosphoramidon (a combined ECE and neutral endopeptidase inhibitor) and BQ-123 (an ETA receptor antagonist) increased FBF by 52±10% (P=.0006) and 31±6% (P=.002), respectively, and thiorphan (a selective neutral endopeptidase inhibitor) reduced FBF by 15±5% (P=.0007). Forearm vasoconstriction to endothelin-1 (an ETA and ETB receptor agonist) was significantly blunted in CHF patients compared with control subjects (both n=10; CHF versus control subjects, P<.001), whereas vasoconstriction to sarafotoxin S6c (an ETB receptor agonist) was significantly enhanced in CHF patients compared with control subjects (both n=10; CHF versus control subjects, P<.05).

Conclusions: ECE inhibitors and ETA receptor antagonists may be useful as vasodilator agents in CHF patients already receiving treatment with an ACE inhibitor. Both ETA and ETB receptors can mediate agonist-induced vasoconstriction in healthy subjects and patients with CHF, but further studies are required to clarify the contribution of each receptor subtype in mediating the effects of endogenous endothelin-1. Show less

Endothelin-1 is an endothelium-derived vasoconstrictor peptide, possibly involved in the pathophysiology of cardiovascular disease. We examined the contribution of endogenously generated endothelin-1 to maintenance of peripheral vascular tone in healthy subjects by local intra-arterial administration of an inhibitor of endothelin converting enzyme, phosphoramidon, and of a selective endothelin receptor A antagonist, BQ-123.

Brachial artery infusion of local doses of proendothelin-1, the… Show more

Endothelin-1 is an endothelium-derived vasoconstrictor peptide, possibly involved in the pathophysiology of cardiovascular disease. We examined the contribution of endogenously generated endothelin-1 to maintenance of peripheral vascular tone in healthy subjects by local intra-arterial administration of an inhibitor of endothelin converting enzyme, phosphoramidon, and of a selective endothelin receptor A antagonist, BQ-123.

Brachial artery infusion of local doses of proendothelin-1, the precursor to endothelin-1, caused a slow-onset dose-dependent forearm vasoconstriction which was abolished by co-infusion of phosphoramidon. Phosphoramidon did not affect responses to endothelin-1. Phosphoramidon caused slow-onset vasodilatation when infused alone, with blood flow increasing by 37% at 90 min (p=0·03). Vasoconstriction to endothelin-1 was abolished by co-infusion of BQ-123 (p=0·006), with forearm blood flow tending to increase. Infusion of BQ-123 alone caused progressive vasodilatation, with blood flow increasing by 64% after 60 min (p=0·007).

These results show that endogenous production of endothelin-1 contributes to the maintenance of vascular tone. Endothelin converting enzyme inhibitors and receptor antagonists may have therapeutic potential as vasodilators. Show less