Browsing by Author "Chicco, Adam J., committee member"
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Item Open Access Elucidating the mechanisms of vascular dysfunction in obesity and type 2 diabetes: the role of the gut microbiota(Colorado State University. Libraries, 2019) Lee, Dustin Michael, author; Gentile, Christopher L., advisor; Weir, Tiffany L., committee member; Johnson, Sarah A., committee member; Chicco, Adam J., committee memberOne of the key processes that links both obesity and type 2 diabetes (T2D) to cardiovascular disease (CVD) is the development of vascular dysfunction, characterized by arterial stiffness and endothelial dysfunction. Vascular dysfunction occurs prior to overt CVD, and the development of vascular dysfunction in obesity and T2D strongly predicts future cardiovascular events and mortality. While the mechanisms of vascular dysfunction continue to be fully elucidated, an abundant body of research suggests that the gut microbiota mediate many cardiometabolic diseases. Disturbances to microbial equilibrium, broadly termed gut dysbiosis, have been implicated in numerous metabolic disorders. In a proof of concept study, our lab has previously demonstrated that suppression of gut dysbiosis reverses vascular dysfunction. Thus, further identifying useful and cost effective treatments that beneficially target the gut microbiota in obesity or T2D to prevent or reverse vascular dysfunction remains an important area of research. The goals of this dissertation research were to 1) examine the underlying causes of vascular dysfunction in models of obesity and T2D and 2) identify novel strategies to prevent or attenuate the development of vascular dysfunction in both obesity and T2D. To investigate the aforementioned, we conducted three separate preclinical studies utilizing a mouse model of T2D, diet-induced obesity, and gut microbiota transplantation. In these studies, we measured aortic pulse wave velocity and endothelium-dependent dilation to examine arterial stiffness and endothelial dysfunction, respectively. Both of these techniques are clinically relevant. We also employed several biochemical techniques to examine the mechanisms by which obesity and T2D lead to vascular dysfunction in our models. In the first study (Chapter 2), we explored epidemiological data suggesting that the antidiabetic drug class, sodium glucose cotransporter 2 inhibitors (SGLT2i), have beneficial effects on cardiovascular outcomes. Utilizing a genetic model of T2D to examine the vascular effects of SGLT2i, we found that treatment with dapagliflozin significantly improved both arterial stiffness and endothelial function. These changes were accompanied by decreased circulating inflammation and subtle alterations to the gut microbiota. In the second study (Chapter 3), we examined the effect of a gut microbiota-derived tryptophan metabolite on cardiometabolic outcomes. Mice fed a western diet displayed increased body weight, arterial stiffness and elevated markers of liver inflammation. Supplementation with the tryptophan metabolite, indole-3-propionic acid, had no effect on these outcomes. Finally, in the third study (Chapter 4) we examined whether human gut dysbiosis represents a causal factor in obesity-related vascular dysfunction. Utilizing human fecal samples from lean and obese subjects, we found that mice colonized with an obese gut microbiota displayed endothelial dysfunction independent of body weight changes. Collectively, these studies provide evidence that 1) SGLT2i-related cardiovascular protection is in part mediated by improvements in vascular dysfunction, 2) gut microbial metabolites have differing effects on host physiology, and 3) the obese human microbiota promotes endothelial dysfunction independent of body weight. Future studies should examine more mechanistic contributions of the gut microbiota that mediate vascular dysfunction in obesity and type 2 diabetes.Item Open Access Engineering in vitro models of non-alcoholic fatty liver disease(Colorado State University. Libraries, 2017) Davidson, Matthew David, author; Khetani, Salman R., advisor; Chicco, Adam J., committee member; Donhue, Seth W., committee member; Kota, Arun K., committee memberDecreased resources and a scarcity of affordable, healthy food is contributing to rising obesity rates throughout the world. Consequentially, non-alcoholic fatty liver disease (NAFLD), which is highly correlated with obesity, rates are also increasing with greater than 30% of the US population currently diagnosed. NAFLD starts as a benign state of fat accumulation within liver hepatocytes but often progresses to more detrimental conditions such as non-alcoholic steatohepatitis (NASH), fibrosis/cirrhosis and hepatocellular carcinoma (HCC). There is no cure for NAFLD or its downstream complications and questions still remain about what factors contribute to disease progression. Specifically, the cause(s) of insulin resistance, lipid accumulation, inflammation, and fibrosis are not completely understood. Many of these questions cannot be elucidated in animal models due to confounding contributions from other organs, differences in animal disease pathology (relative to humans) and dietary restrictions. Additionally, if therapies are to be identified for NALFD, human-relevant systems will need to be used due to species differences in drug metabolism enzymes. Primary human hepatocytes (PHHs) are the gold standard for assessing drug metabolism in vitro, but these cells rapidly lose their liver phenotype in vitro. Here we show that micropatterned co-cultures (MPCCs) of PHHs and stromal cells maintain glucose and lipid metabolism in hepatocytes, which suggests their utility for in vitro disease models of NAFLD. Major advances in culturing methods were developed to increase the insulin sensitivity and overall health of hepatocytes in MPCCs prior to carrying out studies regarding NAFLD-related insulin resistance. The highly insulin sensitive MPCC model was then used to develop models of fatty acid-induced NAFLD and hepatic stellate cell induced NASH phenotypes. Potential disease mechanisms and treatments for fatty acid-induced insulin resistance and NASH disease progression were identified using these models.Item Embargo Exploring microbiome-targeted interventions in the mitigation of endothelial dysfunction(Colorado State University. Libraries, 2024) Risk, Briana D., author; Weir, Tiffany L., advisor; Gentile, Christopher L., advisor; Chicco, Adam J., committee member; Foster, Michelle T., committee memberCardiovascular disease (CVD) has been the leading cause of mortality in the United States for more than seventy years eclipsing cancer and respiratory disease by more than 13%. Despite sincere efforts to decrease the incidence of CVD, various environmental and intrinsic factors contribute to CVD progression, making it challenging to mitigate this complex condition. However, the past decade has shown tremendous growth in understanding the connection between the gut microbiome in its protective, or pathogenic progression of CVD. The gut microbiome, or the consortia of microbial species, genes, and metabolites, that shapes the gastrointestinal environment has a profound impact on the vascular endothelium through mechanisms not yet entirely understood. Therefore, the purpose of the research in this dissertation was to: 1) Utilize next-generation sequencing and metabolomics to characterize microbial contribution to varied endothelial response after a dietary blueberry intervention in post-menopausal females; 2) Determine if a hypertensive microbiome after blueberry treatment confers gastrointestinal and endothelial phenotype in a humanized mouse model; 3) Evaluate the efficacy of a probiotic in reversing endothelial dysfunction in dietarily obese mice, while exploring the contribution to gut barrier integrity and vasoactive metabolite proliferation in a novel cell co-culture.Item Open Access Multi-omic approaches to investigate meat quality variation(Colorado State University. Libraries, 2022) Zhai, Chaoyu, author; Nair, Mahesh N., advisor; Prenni, Jessica E., committee member; Chicco, Adam J., committee member; Belk, Keith E., committee memberVariation in the proteome profile of longissimus lumborum (LL) and psoas major (PM) post-rigor influences meat quality attributes such as tenderness and color stability during retail display. Tandem mass tag (TMT) labeling is a chemical labeling approach using isobaric mass tags for accurate mass spectrometry-based quantification and identification of biological macromolecules. The objective of first study was to use TMT labeling to examine proteome profile variation between beef LL and PM during the early postmortem period (45 min, 12 h, and 36 h). We identified a total of 629 proteins, of which 71 were differentially abundant (fold change > 1.5, P < .05) from three comparisons between the muscles (PM vs. LL at 45 min, 12 h and 36 h). These proteins were mainly involved in oxidative phosphorylation and ATP-related transport, tricarboxylic acid cycle, NADPH regeneration, fatty acid degradation, muscle contraction, calcium signaling, chaperone activity, oxygen transport, as well as degradation of the extracellular matrix. At early postmortem, more abundant antiapoptotic proteins in LL could cause high metabolic stability, enhanced autophagy, and delayed apoptosis, while overabundant metabolic enzymes and pro-apoptotic proteins in PM could accelerate the generation of reactive oxygen species and initiation of cell death. Pulmonary hypertension is a noninfectious disease of cattle at altitudes > 1524 m (5,000 ft). Mean pulmonary arterial pressures (PAP) are used as an indicator for pulmonary hypertension in cattle. High PAP cattle (≥50 mmHg) entering the feedlot at moderate elevations have lower feed efficiency as compared to low PAP cattle (< 50 mmHg). In second study, the impact of pulmonary arterial pressure on mitochondrial function, oxidative phosphorylation (OXPHOS) protein abundance, and meat color was examined using LL from high (98 ± 13 mmHg; n = 5) and low (41 ± 3 mmHg; n = 6) PAP fattened Angus steers (live weight of 588 ± 38 kg) during early postmortem period (2 and 48 h) and retail display (days 1 to 9), respectively. High PAP muscle had greater (P = 0.013) OXPHOS-linked respiration and proton leak-associated respiration than low PAP muscles at 2 h postmortem but rapidly declined to be similar (P = 0.145) to low PAP muscle by 48 h postmortem. OXPHOS protein expression was higher (P = 0.045) in low PAP than high PAP muscle. During retail display, redness, chroma, hue, ratio of reflectance at 630 and 580 nm, and metmyoglobin reducing activity decreased faster (P < 0.05) in high PAP steaks than low PAP. Lipid oxidation significantly increased (P < 0.05) in high PAP steaks but not (P > 0.05) in low PAP. The results indicated that high PAP caused a lower OXPHOS efficiency and greater fuel oxidation rates under conditions of low ATP demand in premortem beef LL muscle; this could explain the lower feed efficiency in high PAP feedlot cattle compared to low PAP counterparts. Mitochondrial integral function (membrane integrity or/and protein function) declined faster in high PAP than low PAP muscle at early postmortem. LL steaks from high PAP animals had lower color stability than those from the low PAP animals during simulated retail display, which could be partially attributed to the loss of muscle mitochondrial function at early postmortem by ROS damage in high PAP muscle. Rapid Evaporative Ionization Mass Spectrometry (REIMS) is a type of ambient ionization mass spectrometry, which enables real-time evaluation of several complex traits from a single measurement. The objectives of third study were (1) to investigate the capability of REIMS to accurately identify and predict cooked sheep meat flavor and carcass characteristics based on consumer response utilizing metabolomic data acquired from different types of raw sample by I-Knife and (2) to compare the data generated by these two electrodes (Meat Probe vs. I-Knife) in their ability to differentiate carcass background and sheep meat flavor. Current study demonstrated that REIMS analysis of raw meat samples can be used to accurately predict and classify cooked sheep meat flavor and carcass characteristics (based on consumer response). Specifically, the lean and fat tissue collected at 45 min postmortem can be used to predict carcass characteristics and post rigor meat flavor. Models for diet, flavor intensity acceptance, off flavors presence, overall acceptance, age, and flavor acceptance achieved prediction accuracies higher than 80%. In addition, data generated using the Meat Probe resulted in models with better or similar prediction accuracies of carcass background (age, diet, and gender) and consumer preference (intensity acceptance, flavor acceptance, off flavors presence, and overall acceptance) as compared to models based on data generated using the I-Knife. The Meat Probe was more user-friendly, faster, and cleaner than I-Knife for REIMS analysis. Further investigations are necessary to evaluate the use of the Meat Probe for REIMS analysis in other applications.Item Open Access Novel modulators of blood pressure with age: a physiological and bioinformatics-based approach(Colorado State University. Libraries, 2021) Bachman, Nate P., author; Braun, Barry, advisor; LaRocca, Thomas J., advisor; Chicco, Adam J., committee member; Gentile, Christopher L., committee memberSystolic blood pressure (SBP) increases with age and is a significant risk factor cardio- and cerebrovascular diseases. While the causes of high blood pressure (hypertension) have been extensively studied, the causes of the age-related rise in blood pressure independent of chronic disease remain unclear. Thus, the identification of novel mechanisms underlying age-related high blood pressure may lead to new strategies to reduce chronic disease risk in older adults. Therefore, the goal of this dissertation was to use both physiological and bioinformatics-based approaches to better elucidate contributors to elevated blood pressure in healthy older adults. The main findings are that 1) inhibition of Rho-kinase (an enzyme that participates in numerous cellular/regulatory pathways) lowers systemic blood pressure in healthy older adults concomitant with reduced vascular resistance but not improved endothelial function, 2) genes expression patterns in peripheral white blood cells differ in healthy older adults with elevated SBP compared to those with normal SBP and transcriptomic (RNA) changes relate to vascular and immune function, and 3) circulating chemokines and whole blood immune-related transcripts track with elevated SBP in healthy older adults. Taken together, this work shows that Rho-kinase, circulating RNA transcripts, and circulating chemokines may be novel therapeutic targets and/or biomarkers of elevated blood pressure in healthy older adults with untreated hypertension.Item Open Access Novel role of acetylcholine in vascular control in humans(Colorado State University. Libraries, 2020) Terwoord, Janée D., author; Dinenno, Frank A., advisor; Amberg, Gregory C., committee member; Chicco, Adam J., committee member; Gentile, Christopher L., committee memberThe vascular endothelium is remarkably sensitive to the molecule acetylcholine (ACh), which binds to muscarinic receptors to initiate endothelium-dependent vasodilation. Although vasodilatory responsiveness to ACh is considered the gold standard index of endothelial function, an obligatory role for ACh in peripheral blood flow control has been challenging to elucidate. Thus, muscarinic ACh receptors on endothelial cells are widely considered to be evolutionary remnants with no real physiological function in humans. Administration of exogenous ACh amplifies endothelial sensitivity to other vasodilatory stimuli and blunts sympathetic vasoconstrictor signaling; therefore, we sought to determine whether endogenous ACh contributes to these processes in vivo. Accordingly, the overall goal of this dissertation research was to evaluate the role of ACh in modulating sympathetic α adrenergic vasoconstriction and eliciting vasodilation in healthy, young adults. The primary findings are that 1) ACh interacts with the endothelium-dependent vasodilator adenosine triphosphate (ATP) to augment vasodilation and limit α1 adrenergic vasoconstriction in the skeletal muscle resistance vasculature, 2) endogenous ACh blunts sympathetic vasoconstriction within active skeletal muscle and is an obligatory mechanism of functional sympatholysis during exercise at high intensities, and 3) ACh mediates flow-induced vasodilation of conduit arteries in response to sustained and transient increases in shear rate induced by handgrip exercise and reactive hyperemia, respectively. Collectively, these studies reveal a novel, physiological role of ACh in peripheral blood flow regulation in humans.Item Open Access On the role of circulating ATP in vascular control at rest and during exercise of aging humans(Colorado State University. Libraries, 2010) Kirby, Brett Sean, author; Dinenno, Frank A., advisor; Earley, Scott, 1963-, committee member; Chicco, Adam J., committee member; Gotshall, Robert William, 1945-, committee memberThe following investigation composes a series of experiments with the overall aim of determining the role for the circulating nucleotide adenosine triphosphate (ATP) in vascular control at rest and during exercise in humans of advanced age. We tested the general hypothesis that ATP has definite vasodilator and sympatholytic vasomotor properties in young adults during exercise, but that these actions are impaired in older adults; and that endogenously circulating levels of ATP are diminished during exercise in the aged population. Specifically, the experiments are outlined as such: 1) to determine whether exogenous ATP can modulate α-adrenergic vasoconstriction in the human forearm of young adults, 2) to determine whether vasodilator responsiveness to exogenous ATP is impaired in aging humans, and the contribution of adenosine to ATP-mediated vasodilation in aging humans, 3) to determine whether the ability of exogenous ATP to modulate α-adrenergic vasoconstriction in the human forearm of older adults is impaired similar to the typical response observed during exercise in aged humans, 4) to determine whether endogenous venous plasma [ATP] and ATP release is diminished during mild-to-moderate exercise in aging humans. Our collective findings indicate that alterations in the contribution of ATP to vascular tone in aging humans exist and may in part be a potential mechanism by which aged adults have reductions in oxygen delivery to active skeletal muscle. In particular, circulating exogenous ATP has the ability to significantly blunt α-adrenergic vasoconstriction in young adults similar to that observed during exercise. In contrast to our hypothesis, the vasodilatory responsiveness and sympatholytic properties of exogenous ATP remain intact in aging humans. However, older adults demonstrate reduced venous plasma [ATP] and impaired ATP release during graded mild-to-moderate handgrip exercise which is associated with attenuations in skeletal muscle vasodilation and blood flow. Taken together, it is our belief that the typically observed impairments in skeletal muscle vasodilation and the inability to offset sympathetic vasoconstrictor tone during exercise is in part due to diminished endogenous levels of circulating ATP in aging humans. On the whole, ATP appears to be a significant regulator of vascular control in humans, and may act as a potential mechanism which in part explains the typically observed reductions in skeletal muscle blood flow and oxygen delivery to active tissue in aged humans thereby predisposing this population to an elevated risk for cardiovascular diseases, age-related declines in exercise capacity/tolerance, and an overall decline in quality of life in this population.Item Open Access Oocyte metabolism – a potential link between mare conditions and impaired fertility(Colorado State University. Libraries, 2023) Di Donato Catandi, Giovana, author; Carnevale, Elaine M., advisor; Krisher, Rebecca L., advisor; Chicco, Adam J., committee member; Chen, Thomas W., committee memberMaternal advanced aging and obesity are known for negatively affecting reproductive outcomes by directly impacting the oocyte and the follicular environment, where the oocyte develops and matures. Success of early embryonic development relies on appropriate ability of the oocyte to produce energy. Whether maternal conditions of the mare impact oocyte metabolic function had not been previously determined. In the studies described throughout this dissertation, novel microsensors were utilized to quantify aerobic and anaerobic metabolism of single equine oocytes. Additional and complementing end points were obtained through high- resolution respirometry of granulosa cells and metabolomic profiling of oocytes and cumulus cells. The overarching hypothesis of this dissertation is that mare conditions known to impair fertility, namely advanced age and obesity, affect oocyte metabolism, ultimately impairing oocyte developmental potential. It was additionally hypothesized that dietary supplementation to old or obese mares would reach and affect the ovarian follicular environment and the oocyte, improving its metabolic function and quality. To test these hypotheses, a series of three projects were conducted to: 1) Investigate effects of mare advanced aging on oocyte metabolism; 2) Determine the potential of diet supplementation to old mares to improve oocyte metabolism; 3) Investigate effects of mare obesity on oocyte metabolism and the potential of diet supplementation on normalizing metabolic alterations. Findings from these projects revealed that mare advanced aging impairs oocyte aerobic and anaerobic metabolic function, contributing to limited embryonic metabolism and development after intracytoplasmic sperm injection (ICSI). Short-term dietary supplementation to old mares with feed additives, specifically formulated to improve mitochondrial metabolism and overall equine health, was able to improve mitochondrial metabolism of granulosa cells and oocytes, promoting greater embryonic rates after ICSI in comparison to a control grain supplementation. Additionally, the findings here reported demonstrate that mare obesity promotes several alterations in the ovarian follicle, including excess of reactive oxygen species production by granulosa cells, lipid accumulation in cumulus cells and oocytes, and excessive oocyte aerobic and anaerobic metabolism. Dietary supplementation to obese mares with similar feed components mitigated many of the obesity-associated follicular changes, likely contributing to oocyte quality. Collectively, these novel discoveries contribute to knowledge and understanding of the direct effects of maternal conditions of the mare on the ovarian follicle and oocyte, elucidating cellular mechanisms by which advanced aging and obesity disturb fertility. Furthermore, these findings reveal the benefits of dietary interventions in improving oocyte metabolism and quality. Dietary supplementation represents a non-invasive and feasible approach to tackle female subfertility. Assuredly the results presented throughout this dissertation will contribute to the equine reproduction industry, with potential to have a translational impact on the human fertility industry, by not only elucidating direct effects of maternal conditions on oocyte metabolism, but also by providing a practical method for rescuing it in vivo.Item Open Access Peripheral blood flow regulation in persons with multiple sclerosis(Colorado State University. Libraries, 2021) Ketelhut, Nathaniel B., author; Fling, Brett W., advisor; Hickey, Matthew S., committee member; Chicco, Adam J., committee member; Myers, Brent, committee memberMultiple sclerosis (MS) is an inflammatory, degenerative disease of the central nervous system that is believed to be autoimmune in nature. The disease affects approximately one million people in the United States and results in a wide variety of symptoms including impaired physical function, reduced exercise capacity, and increased fatigability. Although considerable effort has been invested in improving our understanding of the neuromuscular contributions to these symptoms, no studies have investigated whether cardiovascular autonomic dysfunction compromises skeletal muscle blood flow in persons with MS (PwMS). Indeed, approximately 50% of PwMS have an abnormal response to tests of cardiovascular autonomic function, and skeletal muscle blood flow is positively associated with exercise capacity. Thus, the overall goal of this dissertation was to determine whether PwMS have impaired skeletal muscle blood flow responses to exercise relative to age- and sex-matched healthy controls. The primary findings are that 1) the local control of skeletal muscle blood flow during exercise is intact in PwMS, 2) skeletal muscle blood flow is likely reduced during exercise that engages the autonomic nervous system in MS, which may be due to increased α-adrenergic mediated vascular tone, and 3) that PwMS may experience hypersensitivity to α-adrenergic signaling as evidenced by levels of systemic vascular resistance relative to plasma concentrations of norepinephrine. Together, these studies indicate that compromised skeletal muscle blood flow during exercise may contribute to reduced exercise capacity and increased fatigability in PwMS.Item Open Access Protein synthesis rates in response to exercise and β-adrenergic signaling in human skeletal muscle(Colorado State University. Libraries, 2011) Robinson, Matthew McHutcheson, author; Miller, Benjamin F., advisor; Pagliassotti, Michael J., committee member; Hamilton, Karyn L., committee member; Chicco, Adam J., committee member; Hickey, Matthew S., committee memberSkeletal muscle protein turnover is determined by the synthesis and degradation of skeletal muscle proteins and is the mechanism that determines skeletal muscle protein content. A loss of skeletal muscle mass and function occurs during aging (sarcopenia) due to a net imbalance between synthesis and degradation pathways. Mitochondrial protein turnover, a component of skeletal muscle protein turnover, is decreased with aging. A decline in mitochondrial protein turnover and subsequent decline in mitochondrial function is associated with the progression of chronic diseases associated with aging. Aging populations are commonly prescribed medications to combat age-related chronic diseases. Among commonly prescribed medications are β-adrenergic receptor blockers as anti-hypertensive therapy. Decreased β-adrenergic signaling may impair skeletal muscle adaptations to exercise, particular the mitochondrial fraction, and potentially diminish the benefits of exercise training on skeletal muscle protein synthesis. The regulation of post-exercise mitochondrial protein synthesis by β-adrenergic receptor signaling is not well known in humans. Protein consumption following exercise induces net synthesis of skeletal muscle proteins in younger populations, however the effect appears to be blunted with aging and likely lead to sarcopenia. The net positive protein synthesis following exercise with protein feeding occurs for several hours and may be effective therapy for age-related declines in skeletal muscle mass, yet it is not known whether these short increases will persist over longer periods. The overall objective of our three projects was to investigate the regulation of skeletal muscle protein synthesis in response to exercise, protein consumption, and β-adrenergic signaling in humans. We tested the hypothesis that β-adrenergic signals can regulate mitochondrial biogenesis by examining non-selective β-adrenergic stimulation during resting conditions (Experiment #1) and non-selective β-adrenergic blockade during aerobic exercise (Experiment #2). Furthermore, we tested the hypothesis that protein consumption following exercise can promote skeletal muscle protein synthesis over several weeks of aerobic training (Experiment #3). We used stable isotopic methods to determine rates of skeletal muscle protein synthesis including analysis of the mitochondrial fraction as a measure of mitochondrial biogenesis. Additional measures of mitochondrial biogenesis included mitochondrial DNA content and mRNA content of signaling pathways for mitochondrial adaptations. Deuterium labeling over several weeks was used to measure the synthetic rates of skeletal muscle proteins and DNA during aerobic training. Experiment #1 involved examining the short-term response of skeletal muscle protein synthesis and mitochondrial biogenesis following infusion of a non-selective β-adrenergic agonist. We found that non-selective β-adrenergic activation did not increase skeletal muscle synthesis, whole body protein turnover, or measures of mitochondrial biogenesis. Experiment #2 included investigation of the short-term response of skeletal muscle protein synthesis following infusion of a non-selective β-adrenergic antagonist during a one-hour bout of cycling. Mitochondrial protein fractional synthesis rates were decreased following cycling with non-selective β-adrenergic blockade, yet signals for mitochondrial biogenesis were not different compared to a saline control infusion. Experiment #3 included evaluating the ability for post-exercise protein consumption during aerobic training to stimulate long-term measures of multiple skeletal muscle synthetic processes. We determined that consuming protein compared to carbohydrates after exercise did not lead to differences in protein synthesis or mitochondrial DNA content over several weeks. Interestingly, we measured the amount of newly synthesized DNA in skeletal muscle to be ~5%. Skeletal muscle does not undergo regular cell division, therefore the DNA synthesis was higher than expected. It is likely that the DNA synthesis is due to satellite cell activation. We conclude that β-adrenergic signaling during exercise is a signal for mitochondrial protein synthesis in skeletal muscle. Additionally, the ability for protein consumption following exercise to increase protein synthesis over several hours does not lead to long-term increases in protein synthesis. Collectively, these results provide insight into the regulation of skeletal muscle protein turnover with exercise and β-adrenergic signaling. Understanding potential negative drug and exercise interactions can help improve future therapeutic recommendations for healthy aging.Item Open Access Role of vascular hyperpolarization in muscle blood flow regulation in healthy humans(Colorado State University. Libraries, 2013) Crecelius, Anne Renee, author; Dinenno, Frank A., advisor; Chicco, Adam J., committee member; Tjalkens, Ronald B., committee member; Earley, Scott, committee memberThe following investigation composes a series of experiments with the overall aim of determining the role for vascular hyperpolarization via activation of inwardly-rectifying potassium (KIR) channels and Na+/K+-ATPase in the regulation of vascular tone in response to muscle contractions and ischaemia in young, healthy humans. We tested the general hypothesis that activation of KIR channels and Na+/K+-ATPase contributes in large part to the vasodilatory, hyperaemic, and sympatholytic responses observed in these conditions and this contribution is greater than that of other vasodilators, specifically nitric oxide (NO) and prostaglandins (PGs). The specific aims of each experiment were: 1) to determine whether K+-stimulated vascular hyperpolarization via activation of KIR channels and Na+/K+-ATPase mediates contraction-induced rapid vasodilatation in the human forearm; 2) to determine whether vascular hyperpolarization via activation of KIR channels and Na+/K+-ATPase contributes to the hyperaemic response at the onset of repeated muscle contractions, as well as to steady-state forearm blood flow during rhythmic handgrip exercise; 3) to determine whether vascular hyperpolarization via activation of KIR channels and Na+/K+-ATPase contributes to the observed blunting of sympathetically-mediated vasoconstriction that occurs during moderate intensity rhythmic forearm exercise; and 4) to determine whether vascular hyperpolarization via activation of KIR channels and Na+/K+-ATPase contributes to the observed reactive hyperaemia that occurs in the human forearm following release of temporary ischaemia. Our collective findings demonstrate a significant contribution of KIR channels and Na+/K+-ATPase activation to rapid vasodilatation following a single muscle contraction, the onset of exercise hyperaemia in response to repeated muscle contractions, steady-state muscle blood flow during rhythmic handgrip exercise and reactive hyperaemia following temporary ischaemia. In contrast to our hypothesis, we did not observe a significant contribution of KIR channels and Na+/K+-ATPase to the observed blunting of sympathetic α-adrenergic vasoconstriction that occurs during handgrip exercise. In all studies, any role of NO and PGs was modest, if present at all. Taken together, our findings indicate that during a variety of vasodilatory stimuli, there is a large contribution of pathways that are independent of NO and PGs, specifically activation of KIR channels and Na+/K+-ATPase. Hyperpolarization via activation of KIR channels and Na+/K+-ATPase represents a novel mechanistic pathway in the understanding of in vivo regulation of muscle blood flow in response to contractions and ischaemia. These findings may provide insight into understanding impaired vascular function in patient populations and as such, could represent a novel therapeutic target for reversing microvascular dysfunction.