Adipose tissue hyaluronan production improves systemic glucose homeostasis and primes adipocytes for CL 316,243-stimulated lipolysis

Plasma hyaluronan (HA) increases systemically in type 2 diabetes (T2D) and the HA synthesis inhibitor, 4-Methylumbelliferone, has been proposed to treat the disease. However, HA is also implicated in normal physiology. Therefore, we generated a Hyaluronan Synthase 2 transgenic mouse line, driven by a tet-response element promoter to understand the role of HA in systemic metabolism. To our surprise, adipocyte-specific overproduction of HA leads to smaller adipocytes and protects mice from high-fat-high-sucrose-diet-induced obesity and glucose intolerance. Adipocytes also have more free glycerol that can be released upon beta3 adrenergic stimulation. Improvements in glucose tolerance were not linked to increased plasma HA. Instead, an HA-driven systemic substrate redistribution and adipose tissue-liver crosstalk contributes to the systemic glucose improvements. In summary, we demonstrate an unexpected improvement in glucose metabolism as a consequence of HA overproduction in adipose tissue, which argues against the use of systemic HA synthesis inhibitors to treat obesity and T2D.

Repurposing beta-3 adrenergic receptor agonists for Alzheimer’s disease: beneficial effects in a mouse model

Background Old age, the most important risk factor for Alzheimer’s disease (AD), is associated with thermoregulatory deficits. Brown adipose tissue (BAT) is the main thermogenic driver in mammals and its stimulation, through β3 adrenergic receptor (β3AR) agonists or cold acclimation, counteracts metabolic deficits in rodents and humans. Studies in animal models show that AD neuropathology leads to thermoregulatory deficits, and cold-induced tau hyperphosphorylation is prevented by BAT stimulation through cold acclimation. Since metabolic disorders and AD share strong pathogenic links, we hypothesized that BAT stimulation through a β3AR agonist could exert benefits in AD as well. Methods CL-316,243, a specific β3AR agonist, was administered to the triple transgenic mouse model of AD (3xTg-AD) and non-transgenic controls from 15 to 16 months of age at a dose of 1 mg/kg/day i.p. Results Here, we show that β3AR agonist administration decreased body weight and improved peripheral glucose metabolism and BAT thermogenesis in both non-transgenic and 3xTg-AD mice. One-month treatment with a β3AR agonist increased recognition index by 19% in 16-month-old 3xTg-AD mice compared to pre-treatment (14-month-old). Locomotion, anxiety, and tau pathology were not modified. Finally, insoluble Aβ42/Aβ40 ratio was decreased by 27% in the hippocampus of CL-316,243-injected 3xTg-AD mice. Conclusions Overall, our results indicate that β3AR stimulation reverses memory deficits and shifts downward the insoluble Aβ42/Aβ40 ratio in 16-month-old 3xTg-AD mice. As β3AR agonists are being clinically developed for metabolic disorders, repurposing them in AD could be a valuable therapeutic strategy.

β3 Adrenergic Receptor Stimulation Promotes Reperfusion in Ischemic Limbs in a Murine Diabetic Model

Aims/Hypothesis: Peripheral arterial disease (PAD) is a major burden, resulting in limb claudication, repeated surgical interventions and amputation. There is an unmet need for improved medical management of PAD that improves quality of life, maintains activities of daily life and reduces complications. Nitric oxide (NO)/redox balance is a key regulator of angiogenesis. We have previously shown beneficial effects of a β3 adrenergic receptor (β3AR) agonist on NO/redox balance. We hypothesized that β3AR stimulation would have therapeutic potential in PAD by promoting limb angiogenesis.Methods: The effect of the β3AR agonist CL 316,243 (1–1,000 nmol/L in vitro, 1 mg/kg/day s. c) was tested in established angiogenesis assays with human endothelial cells and patient-derived endothelial colony forming cells. Post-ischemia reperfusion was determined in streptozotocin and/or high fat diet-induced diabetic and non-diabetic mice in vivo using the hind limb ischemia model.Results: CL 316,243 caused accelerated recovery from hind limb ischemia in non-diabetic and type 1 and 2 diabetic mice. Increased eNOS activity and decreased superoxide generation were detected in hind limb ischemia calf muscle from CL 316, 243 treated mice vs. controls. The protective effect of CL 316,243 in diabetic mice was associated with >50% decreases in eNOS glutathionylation and nitrotyrosine levels. The β3AR agonist directly promoted angiogenesis in endothelial cells in vitro. These pro-angiogenic effects were β3AR and NOS-dependent.Conclusion/Interpretation:β3AR stimulation increased angiogenesis in diabetic ischemic limbs, with demonstrable improvements in NO/redox balance and angiogenesis elicited by a selective agonist. The orally available β3AR agonist, Mirabegron, used for overactive bladder syndrome, makes translation to a clinical trial by repurposing of a β3AR agonist to target PAD immediately feasible.

Developmental exposure to DDT or DDE alters sympathetic innervation of brown adipose in adult female mice

Background Exposure to the bioaccumulative pesticide dichlorodiphenyltrichloroethane (DDT) and its metabolite dichlorodiphenyldichloroethylene (DDE) has been associated with increased risk of insulin resistance and obesity in humans and experimental animals. These effects appear to be mediated by reduced brown adipose tissue (BAT) thermogenesis, which is regulated by the sympathetic nervous system. Although the neurotoxicity of DDT is well-established, whether DDT alters sympathetic innervation of BAT is unknown. We hypothesized that perinatal exposure to DDT or DDE promotes thermogenic dysfunction by interfering with sympathetic regulation of BAT thermogenesis. Methods Pregnant C57BL/6 J mice were administered environmentally relevant concentrations of DDTs (p,p’-DDT and o,p’-DDT) or DDE (p,p’-DDE), 1.7 mg/kg and 1.31 mg/kg, respectively, from gestational day 11.5 to postnatal day 5 by oral gavage, and longitudinal body temperature was recorded in male and female offspring. At 4 months of age, metabolic parameters were measured in female offspring via indirect calorimetry with or without the β3 adrenergic receptor agonist, CL 316,243. Immunohistochemical and neurochemical analyses of sympathetic neurons innervating BAT were evaluated. Results We observed persistent thermogenic impairment in adult female, but not male, mice perinatally exposed to DDTs or p,p’-DDE. Perinatal DDTs exposure significantly impaired metabolism in adult female mice, an effect rescued by treatment with CL 316,243 immediately prior to calorimetry experiments. Neither DDTs nor p,p’-DDE significantly altered BAT morphology or the concentrations of norepinephrine and its metabolite DHPG in the BAT of DDTs-exposed mice. However, quantitative immunohistochemistry revealed a 20% decrease in sympathetic axons innervating BAT in adult female mice perinatally exposed to DDTs, but not p,p’-DDE, and 48 and 43% fewer synapses in stellate ganglia of mice exposed to either DDTs or p,p’-DDE, respectively, compared to control. Conclusions These data demonstrate that perinatal exposure to DDTs or p,p’-DDE impairs thermogenesis by interfering with patterns of connectivity in sympathetic circuits that regulate BAT. Graphical abstract

Developmental Exposure to DDT or DDE Alters Sympathetic Innervation of Brown Adipose in Adult Female Mice

Background: Exposure to the bioaccumulative pesticide dichlorodiphenyltrichloroethane (DDT) and its metabolite dichlorodiphenyldichloroethylene (DDE) has been associated with increased risk of insulin resistance and obesity in humans and experimental animals. These effects appear to be mediated by reduced brown adipose tissue (BAT) thermogenesis, which is regulated by the sympathetic nervous system. Although the neurotoxicity of DDT is well-established, whether DDT alters sympathetic innervation of BAT is unknown. We hypothesized that perinatal exposure to DDT or DDE promotes thermogenic dysfunction by interfering with sympathetic regulation of BAT thermogenesis. Methods: Pregnant C57BL/6J mice were exposed by oral gavage to environmentally relevant concentrations of DDT (1.7 mg/kg) or DDE (1.31 mg/kg) from gestational day 11.5 to postnatal day 5, and longitudinal body temperature was recorded in male and female offspring. At 4 months of age, metabolic parameters were measured in female offspring via indirect calorimetry with or without the β3 adrenergic receptor agonist, CL 316,243. Immunohistochemical and neurochemical analyses of sympathetic neurons innervating BAT were evaluated.Results: We observed persistent thermogenic impairment in adult female, but not male, mice perinatally exposed to DDT or DDE. Perinatal DDT exposure significantly impaired metabolism in adult female mice, an effect rescued by treatment with CL 316,243 immediately prior to calorimetry experiments. Neither DDT nor DDE significantly altered BAT morphology or the concentrations of norepinephrine and its metabolite DHPG in the BAT of DDT-exposed mice. However, quantitative immunohistochemistry revealed a 20% decrease in sympathetic axons innervating BAT in adult female mice perinatally exposed to DDT, but not DDE, and 48% and 43% fewer synapses in stellate ganglia of mice exposed to either DDT or DDE, respectively, compared to control. Conclusions: These data demonstrate that perinatal DDT and DDE exposure impairs thermogenesis by interfering with patterns of connectivity in sympathetic circuits that regulate BAT.

A systems biology analysis of adrenergically stimulated adiponectin exocytosis in white adipocytes

ABSTRACTCirculating levels of the adipocyte hormone adiponectin are typically reduced in obesity and this deficiency has been linked to metabolic diseases. It is thus important to understand the mechanisms controlling adiponectin exocytosis. This understanding is hindered by the high complexity of the data and the underlying signaling network. To handle this complexity, we here analyze the data using systems biology mathematical modelling. Previously, we have developed a mathematical model for how different intracellular concentrations of Ca2+, cAMP and ATP affect adiponectin exocytosis (measured as increase in membrane capacitance). However, recent work has shown that adiponectin exocytosis is physiologically triggered via signaling pathways involving adrenergic β3 receptors (β3ARs). Therefore, we have herein developed a more comprehensive model that also includes adiponectin exocytosis stimulated by extracellularly applied epinephrine or the β3AR agonist CL 316,243. Our model can explain all previous patch-clamp data, as well as new data consisting of a combination of the intracellular mediators and extracellular adrenergic stimuli. Without changing the parameters, the model can accurately predict independent validation data with other combinations of patch-clamp pipette solutions and external stimuli. Finally, we use the model to perform new in silico experiments examining situations where corresponding wet lab experiments are difficult to perform. By this approach, we simulated adiponectin exocytosis in single cells, in response to the reduction of β3ARs that is observed in adipocytes from animals with obesity-induced diabetes. Our work brings us one step closer to understanding the intricate regulation of adiponectin exocytosis.

Repurposing beta3-adrenergic receptor agonists for Alzheimer’s disease: Beneficial effects on recognition memory and amyloid pathology in a mouse model

ABSTRACTOld age, the most important risk factor for Alzheimer’s disease (AD), is associated with thermoregulatory deficits. Brown adipose tissue (BAT) is the main thermogenic driver in mammals and its stimulation, through β3-adrenergic receptor (β3AR) agonists or cold acclimation, counteracts metabolic deficits in rodents and humans. Studies in animal models show that AD neuropathology leads to thermoregulatory deficits and cold-induced tau hyperphosphorylation is prevented by BAT stimulation through cold acclimation. Since metabolic disorders and AD share strong pathogenic links, we hypothesized that BAT stimulation through a β3AR agonist could exert benefits in AD as well.Here, we show that β3AR agonist administration (CL-316,243, 1 mg/kg/day i.p., from 15 to 16 months of age) decreased body weight and improved peripheral glucose metabolism and BAT thermogenesis in both non-transgenic and 3xTg-AD mice. One-month treatment with a β3AR agonist increased recognition index by 19% in 16-month-old 3xTg-AD mice compared to pre-treatment (14-month-old). Locomotion, anxiety and tau pathology were not modified. Finally, insoluble Aβ42/Aβ40 ratio was decreased by 27% in the hippocampus of CL-316,243-injected 3xTg-AD mice.Overall, our results indicate that β3AR stimulation reverses memory deficits and shifts downward the insoluble Aβ42/Aβ40 ratio in 16-month-old 3xTg-AD mice. As β3AR agonists are being clinically developed for metabolic disorders, repurposing them in AD could be a valuable therapeutic strategy.

Beta-3 Receptor Agonists

Beta-3 adrenergic receptors (β3-AR) have a more widespread tissue distribution in the human body as compared to beta1- and beta2 (β1/2)-adrenergic receptors, including in the bladder, brain, adipose tissue and cardiovascular system. Thus, β3- AR are potential drug targets for a wide range of therapeutic areas, both cardiovascular and non-cardiovascular including overactive bladder (OAB), depression, metabolic syndrome, obesity and heart failure (HF). β3-AR agonists that are selective to the β3-AR include CL 316,243, amibegron (SR58611A), mirabegron (YM-178) and vibegron (RVT-901). However, in HF, study results regarding a possible inotropic effect of β3-AR agonists remain equivocal and some authors report a negative inotropic effect in HF and β3-AR antagonists are also under study.

Duration of a ”Brown-Like” Phenotype of White Adipose Tissue Induced by the β3 Agonist CL-316,243

Abstract“Browning” i. e. the transformation of white adipose tissue into brown-like adipose tissue could induce efficient burning of excess fat reserves via induction of non-shivering thermogenesis. For example, activation of ß3 adrenergic receptors has been show to induce such changes, however, it is still not clear, how long after termination of such a treatment, beneficial effects might be maintained. To address this question, we treated rats s.c. for 2 weeks with the ß3 agonist CL-316,243 at 1 mg/kg and assessed interscapular brown fat and inguinal white fat pads weight, UCP-1 (a marker for the brown-like fat phenotype) using immunohistochemistry and H&E staining, at different intervals after treatment termination.One 1 day after the treatment cessation there was a decrease of inguinal white fat pad weight and increase of interscapular fat pad. This change vanished at 7 days for inguinal pad and at 14 days for interscapular pad. Histological analysis of interscapular pads showed increased UCP-1 staining and brown-like morphology in H&E staining slices at 1 day, but not other time points. In case of inguinal pad there were brown-like features in H&E slices at 1 day and less after 7 days, but absent at 14 days. UCP-1 staining was only detected 1 day after the treatment.In conclusion, the present results indicate that browning-like changes of white fat may be short lasting after treatment termination and could require maintenance treatment of inductor to achieve desired therapeutic effect. This might be a serious shortcoming of potential therapeutic use.