Introduction

In the United States, cardiovascular disease (CVD) is recognized as the number one cause of death for the general population (He et al., 2021). In 2017, CVD comprised 23% of all U.S. deaths, 24.2% of male deaths, and 21.8% of female deaths, accounting for a total of 647,457 deaths that year (Heron, 2019). In 2017, CVD was most prevalent in older populations, as it was the number one cause of death for individuals aged 65-81 years at 25-28.6% of total deaths, and the second leading cause of death for individuals aged 45-64 years at 20.8% of total deaths (Heron, 2019). In 2023, this trend remains true, with the majority of deaths in individuals aged 65 years and older resulting from CVD (Virani et al., 2021). Since 2017, CVD related deaths have increased to approximately 659,000 deaths per year, causing approximately one in every four deaths (Virani et al., 2021). CVD is not only the number one cause of death in the U.S., but it is also very costly. From 1996 to 2016, CVD related spending increased by $108 billion, from $212 billion in 1996 to $320 billion in 2016 (Birger et al., 2021). Currently, the U.S. spends approximately $363 billion per year on CVD, including medical care and medications (Virani et al., 2021).

Cardiovascular disease is linked to a number of risk factors that are continuously increasing in prevalence due to changing American lifestyle habits. These risk factors include but are not limited to: hypertension, hypercholesterolemia, sedentary lifestyles, poor nutrition, smoking, and substance and alcohol abuse. High blood pressure is attributed to an increase in death rates, with the high blood pressure attributed deaths increasing by 34.2% from 2009-2019 (Tsao et al., 2022). High cholesterol is another CVD risk factor and currently, 10.5% of adults 20 years of age and older have high total cholesterol (Tsao et al., 2022). The U.S. is seeing a decrease in active lifestyles in

individuals post high school, as well as an increase in sedentary behavior by adults (Tsao et al., 2022). U.S. citizens are also consuming more saturated fats, low quality carbohydrates, and added sugars, due to dietary changes and the increased prevalence and accessibility of fast food (Tsao et al., 2022). In the U.S., overall tobacco use is decreasing, however 14% of adults aged 18 and over still use tobacco products and smoke daily (Tsao et al., 2022). Due to the development of e-cigarettes and their gaining popularity, e-cigarette use by adolescents has increased by 18.1% from 2011 to 2020 (Tsao et al., 2022). Alcohol, cannabis, opioid, and stimulant abuse also contribute to CVD risk, as abusers of these products have been found to have a higher risk of developing CVD (Gan et al. 2021). The combined CVD risk factors of high blood pressure, high cholesterol, and smoking are prevalent in approximately 46.5% of the American population, making them highly impactful on CVD development (Fryar et al., 2012).

There is also a link between CVD, type 2 diabetes (T2D), and obesity. From 1999-2018 in the U.S., obesity prevalence increased nationally, with the frequency of obesity reaching 42.4% among adults in 2018 (Tsao et al., 2022; Hales et al., 2020.

Currently, 2.4 million deaths can be attributed to obesity worldwide (Tsao et al., 2022). Diagnosed diabetes affects approximately 28.2 million individuals, pre-diabetes affects approximately 113.6 million individuals, and undiagnosed diabetes affects approximately

9.8 million individuals in the U.S. (Tsao et al., 2022). In 2019, diabetes was determined to be an underlying cause of death in the deaths of 87,647 individuals (Tsao et al., 2022).

Obesity and T2D are connected, as insulin resistance is a key factor in the development of obesity-related cardiometabolic disease (Tsao et al., 2022). Insulin

resistance and diabetes are also linked to hyperglycemia, hyperlipidemia, and endothelial dysfunction, resulting in an increased risk of developing CVD. Hyperglycemia and hyperlipidemia, specifically, have been identified as potential factors relating to CVD development via oxidative stress, which is delineated as an overproduction in reactive oxygen species (ROS). An overproduction of ROS not only creates a vicious cycle of increasing levels of hyperglycemia and hyperlipidemia, as ROS concentrations stimulate these two states, but ROS overproduction also contributes to endothelial dysfunction and altered insulin signaling pathways. Therefore, it is beneficial to investigate the relationship between insulin resistance and oxidative stress, and how the relationship between the two states is affected by obesity.

As a result of increasing levels of risk factors and the prevalence of CVD in the United States, researchers, physicians, and medical staff are in agreement that the key to reducing CVD related mortalities is to increase an individuals' social determinants of health (SODH). That involves reducing poverty, increasing access to healthcare, increasing access to holistic nutrition, making education accessible, and creating safe environments that foster increased physical activity, growth, and development. Making positive dietary changes, including following a Mediterranean diet, reduces CVD risk by 26% for at risk individuals (Delgado-Lista et al., 2022). Furthermore, frequent exercise helps to reduce CVD risk factors (Lin et al., 2020). Pharmacy-based interventions, where pharmacists guide patients on pharmaceutical intervention and provide CVD education also helps to reduce CVD (Tsuyuki et al., 2016). However, the most beneficial way of reducing CVD and CVD associated risk factors is by providing culturally-competent and holistic preventative care (Brewer et al., 2017). By tailoring preventative primary care,

physicians and teams are able to reduce heart healthcare costs and cardiovascular related hospitalizations (Mundt et al., 2015). Also, by utilizing preventative care measures, health care providers could prevent between 50,000-100,000 CVD related deaths annually (Farley et al., 2010).

Despite researchers and medical staff having ways to reduce CVD risk factors and treat CVD once it appears, there are still gaps in knowledge of how CVD presents itself at the biological level. However, researchers have identified endothelial cells as playing a key role in CVD development, as a decline in endothelial cell function can lead to cardiovascular and cardiometabolic disease (Incalza et al., 2018). Therefore, to better understand the relationship between endothelial dysfunction and CVD development, as well as the relationship between endothelial dysfunction and other cardiometabolic diseases, a combined approach to studying how various tissue types affect endothelial cell function must be carried out. More specifically, the impact of ROS on endothelial dysfunction and the factors culminating in the overproduction of ROS need to be identified. Therefore, the purpose of this study is to pinpoint factors that lead to the overproduction of ROS within endothelial cells located in adipose tissue. To examine this, quantitative data will be collected from populations of obese and lean individuals participating in Cesar Meza and Dr. Robert Hickner's study aimed to link vascular and metabolic dysfunction via NADPH oxidase (Nox). The overall research question this larger study explores is: does increased Nox-derived ROS production in obesity impair blood glucose profiles by reducing insulin-mediated suppression of lipolysis? As the present study is a sub study of the overall research project, this study addresses the

question: how does glycocalyx quality correlate with adipose tissue Nox activity and adipose tissue microvascular blood flow?