The purpose of the present study was to determine the effect on breathing in the awake state of carotid body denervation (CBD) over 1-2 wk after denervation. Studies were completed on adult goats repeatedly before and 1) for 15 days after bilateral CBD (n = 8), 2) for 7 days after unilateral CBD (n = 5), and 3) for 15 days after sham CBD (n = 3). Absence of ventilatory stimulation when NaCN was injected directly into a common carotid artery confirmed CBD. There was a significant (P < 0.01) hypoventilation during the breathing of room air after unilateral and bilateral CBD. The maximum PaCO2 increase (8 Torr for unilateral and 11 Torr for bilateral) occurred approximately 4 days after CBD. This maximum was transient because by 7 (unilateral) to 15 (bilateral) days after CBD, PaCO2 was only 3-4 Torr above control. CO2 sensitivity was attenuated from control by 60% on day 4 after bilateral CBD and by 35% on day 4 after unilateral CBD. This attenuation was transient, because CO2 sensitivity returned to control temporally similar to the return of PaCO2 during the breathing of room air. During mild and moderate treadmill exercise 1-8 days after bilateral CBD, PaCO2 was unchanged from its elevated level at rest, but, 10-15 days after CBD, PaCO2 decreased slightly from rest during exercise. These data indicate that 1) carotid afferents are an important determinant of rest and exercise breathing and ventilatory CO2 sensitivity, and 2) apparent plasticity within the ventilatory control system eventually provides compensation for chronic loss of these afferents.

Serum accumulation of the gut microbial metabolite trimethylamine N-oxide (TMAO) is associated with high caloric intake and type 2 diabetes (T2D). Impaired pancreatic β-cell function is a hallmark of diet-induced T2D, which is linked to hyperglycemia and hyperlipidemia. While TMAO production via the gut microbiome-liver axis is well defined, its molecular effects on metabolic tissues are unclear, since studies in various tissues show deleterious and beneficial TMAO effects. We investigated the molecular effects of TMAO on functional β-cell mass. We hypothesized that TMAO may damage functional β-cell mass by inhibiting β-cell viability, survival, proliferation, or function to promote T2D pathogenesis. We treated INS-1 832/13 β-cells and primary rat islets with physiological TMAO concentrations and compared functional β-cell mass under healthy standard cell culture (SCC) and T2D-like glucolipotoxic (GLT) conditions. GLT significantly impeded β-cell mass and function by inducing oxidative and endoplasmic reticulum (ER) stress. TMAO normalized GLT-mediated damage in β-cells and primary islet function. Acute 40µM TMAO recovered insulin production, insulin granule formation, and insulin secretion by upregulating the IRE1α unfolded protein response to GLT-induced ER and oxidative stress. These novel results demonstrate that TMAO protects β-cell function and suggest that TMAO may play a beneficial molecular role in diet-induced T2D conditions.

Objectives Elevated circulating levels of the dietary metabolite trimethylamine N-oxide (TMAO) is associated with chronic diseases including cardiovascular disease (CVD) and obesity. While TMAO production via the gut microbiome-liver axis and distribution through the circulation is clear, its molecular effects on metabolic tissues are still unclear. Some clinical studies suggest that elevated TMAO levels increase the risk of type 2 diabetes (T2D) where pancreatic β cell insulin secretion is insufficient for blood glucose management. T2D promoting mechanisms limit functional β cell mass by reducing β cell viability and survival, inhibiting proliferation or decreasing insulin secretory function. We hypothesized that TMAO decreases functional β cell mass by one of these mechanisms to aggravate the T2D phenotype. Methods Using the INS-1 832/13 β cell line and primary murine islets, we screened the effect of various TMAO concentrations on cell viability, proliferation, and function. These parameters were measured under standard and glucolipotoxic (GLT) culture conditions to mimic T2D. We investigated TMAO effects, GLT effects and combined effects. Results TMAO minimally affected viability, proliferation or function under standard culture conditions across 96-hours of treatment. Culturing with GLT impaired viability, proliferation and function after 24 hours of treatment, mimicking T2D onset. Interestingly, adding 40–80 μM TMAO protected against GLT mediated functional impairments in cells and islets. Further, GLT increased oxidative stress by 2.5-fold and adding TMAO was significantly protective. Electron microscopy reveals that GLT alters insulin granule density whereas TMAO maintains proper granule structure. Conclusions These results reject our hypothesis. While TMAO has minor effects on β cells in standard culture conditions, TMAO is sufficient to improve GLT mediated β cell damage by decreasing oxidative stress and maintaining insulin granule formation. These results suggest an early compensatory role for TMAO in countering oxidative damage caused by glucolipotoxicity in β cell function during T2D onset. Funding Sources Funding for this study was provided by the Beatson Foundation and the US Department of Agriculture.

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Diabetes is one of the fastest growing non‐infectious diseases in the world. Current treatments are composed of pharmaceutical agents that enhance insulin sensitivity and eventual insulin monotherapy. Type 2 diabetes is characterized by insulin insensitivity of peripheral tissue, glucose intolerance, and β‐cell dysfunction. Dietary interventions may benefit patients with diabetes, and various plant derived flavonoids have been shown to exert anti‐diabetic effects. While these flavonoids are large, difficult to absorb, and rarely found in circulation, gut bacteria metabolize these into smaller metabolites which can be observed in circulation. We hypothesize that these gut bacteria derived flavanoid metabolites are absorbed and have direct effects on β‐cell function. Male outbred wistar rats were fed one of three diets in the presence or absence of antibiotic treatment: standard diet, standard diet supplemented with catechin hydrate and epicatechin, or standard diet supplemented with grape seed extract. Total urine was collected from the animals (representing the total amount of absorbed metabolites), then metabolites were extracted and reconstituted in water. Here we present data regarding the in vitro effects of these absorbed gut bacteria derived flavanoids on INS‐1 832/13 β‐cell insulin secretion and proliferation. This study sheds further light on the potential ability of flavanoids and their gut bacteria derived metabolites to enhance functional β‐cell mass.

Dietary flavanols are known for disease preventative properties but are often poorly absorbed. Gut microbiome flavanol metabolites are more bioavailable and may exert protective activities. Using metabolite mixtures extracted from the urine of rats supplemented with flavanols and treated with or without antibiotics, we investigated their effects on INS-1 832/13 β-cell glucose stimulated insulin secretion (GSIS) capacity. We measured insulin secretion under non-stimulatory (low) and stimulatory (high) glucose levels, insulin secretion fold induction, and total insulin content. We conducted treatment-level comparisons, individual-level dose responses, and a responder vs. non-responder predictive analysis of metabolite composition. While the first two analyses did not elucidate treatment effects, metabolites from 9 of the 28 animals demonstrated significant dose responses, regardless of treatment. Differentiation of responders vs. non-responder revealed that levels of native flavanols and valerolactones approached significance for predicting enhanced GSIS, regardless of treatment. Although treatment-level patterns were not discernable, we conclude that the high inter-individual variability shows that metabolite bioactivity on GSIS capacity is less related to flavanol supplementation or antibiotic treatment and may be more associated with the unique microbiome or metabolome of each animal. These findings suggest flavanol metabolite activities are individualized and point to the need for personalized nutrition practices.

One in 10 people suffer from diabetes mellitus, where Type 2 (T2D) makes up 90% of cases worldwide. T2D is characterized by hyperglycemia and insulin resistance stemming from dysregulation of insulin secreting pancreatic ß‐cells. The increased demand for insulin posed by insulin resistance leads to further exhaustion and apoptosis of the ß‐cells. Therefore, increasing functional ß‐cell mass by enhancing ß‐cell survival, proliferation and function, may be leveraged to help treat individuals with T2D diabetes. Despite spending 12% of the global health expenditure on diabetes, treatment limitations call for more research and superior therapeutics to protect and recover ß‐cells. Phytochemical flavanols, including naturally occurring polymers of epicatechin, are known to exert antioxidant bioactivity and improve cell health. We have previously demonstrated monomeric epicatechin enhances ß‐cell glucose‐stimulated insulin secretion. We hypothesize that other dietary flavanols may similarly be sufficient to enhance ß‐cell survival, proliferation and function. The objective of this study was to screen 13 dietary flavanols including epicatechin polymers, cinnamtannins, and procynidins, to define their in vitro ability to modulate ß‐cell survival, proliferation and function. Here we present our findings for these compounds, emphasizing their potential beneficial effects as alternative and complementary treatments for T2D.