Parallel development and characterisation of an anti-oxidant stent coating and an in vitro biological model for qualitative assessment

Restenosis is a major cause of coronary artery stent failure and is linked to vascular endothelium damage with resultant oxidative and inflammatory stress. Drug eluting stents (DES) have failed to eliminate this risk; particularly for diabetic patients. Here, we have assessed the potential therapeutic effects of a novel antioxidant stent coating using (i) a common chemical assay, DPPH (2,2-diphenyl-1-picrylhydrazyl) and (ii) superoxide scavenging ability using NBT (nitroblue tetrazolium) reduction and have identified high antioxidant potential that is not dependent on drug incorporation within the coating. To assess the biological effect of this novel antioxidant coating, we have initially used human umbilical vein endothelial cells (HUVECs) treated with pro-inflammatory cytokines to mimic the inflammation stress encountered post-stent placement. Pro-inflammatory signalling was assessed by measuring phospho-P65 (pP65) expression using quantitative western blotting and oxidative stress assessed by measuring reactive oxygen species (ROS) generation using DCFDA. In addition, we have specifically examined expression and activation (via oxidation) of an enzyme called Ca2+/Calmodulin-dependent protein kinase II-delta (CaMKIIδ) that is known to be a central component of vascular pathology during acute and chronic inflammation and oxidative stress. HUVECs were stimulated with pro-inflammatory cytokines, Tissue Necrotic Factor alpha (TNFα) and Interleukin 1-beta (IL-1β). Stimulation with IL-1β (10 ng/ml) for 1h resulted in the highest overall level of P65 phosphorylation (7.80 ± 1.41 (fold increase ± S.E.M. in pP65 expression in IL-1β -stimulated cells c.f. unstimulated controls, n=4, p<0.05). Conversely, stimulation with TNFα (10 ng/ml) for 6h led to the highest overall increase in ROS (5.02 ± 0.54, fold-stimulation ± S.E.M. in ROS in TNFα -stimulated cells c.f. unstimulated controls, n=3, p<0.05). We have shown that CaMKIIδ is highly expressed in HUVECs and that stimulation with IL-1β (10 ng/ml) for 3h significantly induced CaMKII oxidation (1.56 ± 0.06 (fold-increase ± S.E.M. in oxCaMKII in IL-1β stimulated cells c.f. unstimulated controls, n=4, p<0.05). Preventing the oxidation-induced activation of CaMKII may present a therapeutic mechanism by which antioxidant stents can improve endothelial recovery and future work will examine the reduction or reversal of this effect in the presence of our novel stent coating. In conclusion, we have developed a novel anti-oxidant stent coating and established an in vitro system to test biological activity. This approach will now be applied to more physiologically relevant cell types before examining the novel coating’s efficacy

Transition from pre-diabetes to diabetes and predictors of risk in Mexican-Americans

Background: No studies have examined risk factors for the transition from pre-diabetes to diabetes in populations with widespread obesity and diabetes. We determined proximal changes and factors affecting the transition among Mexican-Americans with pre-diabetes. Methods: Participants with pre-diabetes (n=285) were recruited from our randomly sampled population-based Cameron County Hispanic Cohort. These participants were followed for an average of 27 months with repeat examination every 3 to 4 months. Metabolic health was defined as having less than 2 metabolic abnormalities (e.g., hypertension, elevated low-density lipoprotein, etc). Diabetes was identified as fasting blood glucose ≥126 mg/dL, glycated hemoglobin ≥6.5% and/or on hypoglycemic medication. Results: Ninety-six of 285 (33.7%) participants transitioned to overt diabetes. The increased risk of diabetes in the metabolically unhealthy varying with follow-up time was 81% (adjusted odds ratio [OR]: 1.81; 95% CI: 1.09–3.02). The risk of diabetes increased 8% for each kg/m2 of increase in body mass index (BMI, OR: 1.08; 95% CI: 1.05–1.11) independent of covariates. Transition to diabetes was accompanied by a mean increase in BMI of 0.28 kg/m2, and deterioration in metabolic health of 9% (OR: 1.09; 95% CI: 1.003–1.18) compared with those who did not transition. Conclusions: Deteriorating metabolic health and/or increasing BMI significantly raises the risk of transitioning from pre-diabetes to diabetes. Transition itself was accompanied by further increase in BMI and deterioration in metabolic health. These data underline the importance of improving metabolic health and avoiding weight gain in pre-diabetes as simple but clear diabetes prevention targets, and emphasize the importance of lifestyle management

Undiagnosed Diabetes and Pre-Diabetes in Health Disparities

Globally half of all diabetes mellitus is undiagnosed. We sought to determine the extent and characteristics of undiagnosed type 2 diabetes mellitus and pre-diabetes in Mexican Americans residing in the United States. This disadvantaged population with 50% lifetime risk of diabetes is a microcosm of the current pandemic. We accessed baseline data between 2004 and 2014 from 2,838 adults recruited to our Cameron County Hispanic Cohort (CCHC); a two-stage randomly selected \u27Framingham-like\u27 cohort of Mexican Americans on the US Mexico border with severe health disparities. We examined prevalence, risk factors and metabolic health in diagnosed and undiagnosed diabetes and pre-diabetes. Two thirds of this Mexican American population has diabetes or pre-diabetes. Diabetes prevalence was 28.0%, nearly half undiagnosed, and pre-diabetes 31.6%. Mean BMI among those with diabetes was 33.5 kg/m2 compared with 29.0 kg/m2 for those without diabetes. Significant risk factors were low income and educational levels. Most with diabetes had increased waist/hip ratio. Lack of insurance and access to health services played a decisive role in failure to have diabetes diagnosed. Participants with undiagnosed diabetes and pre-diabetes had similar measures of poor metabolic health similar but generally not as severe as those with diagnosed diabetes. More than 50% of a minority Mexican American population in South Texas has diabetes or pre-diabetes and is metabolically unhealthy. Only a third of diabetes cases were diagnosed. Sustained efforts are imperative to identify, diagnose and treat individuals in underserved communities

Integrating and evaluating interdisciplinary sustainability and STEM curriculum in geographical education: A case of three teaching modalities

The effectiveness of interdisciplinary sustainability curriculum remains understudied in geography education. Accordingly, we deployed and evaluated an interdisciplinary sustainability and STEM module for in-person and online sections of a fall 2018 Human Geography course. Results indicate that sustainability knowledge improved after the interdisciplinary curricular intervention irrespective of course modality. Another focus is to explore student reactions to teaching modality due to COVID-19 disruptions. Results indicate that online student sustainability knowledge also improved during COVID-19 (fall 2020). For students in a section converted from in-person to blended, sustainability knowledge did not improve. Implications are provided

High prevalence of subclinical atherosclerosis by carotid ultrasound among Mexican Americans: discordance with 10-year risk assessment using the Framingham risk score

ackground: Framingham risk scores (FRS) were validated in a mostly Caucasian population. Evaluation of subclinical atherosclerosis by carotid ultrasound may improve ascertainment of risk in nonwhite populations. This study aimed to evaluate carotid intima-media thickness (cIMT) and carotid plaquing among Mexican Americans, and to correlate these markers with coronary risk factors and the FRS. Methods/results: Participants (n = 141) were drawn from the Cameron County Hispanic Cohort. Carotid artery ultrasound was performed and cIMT measured. Carotid plaque was defined as areas of thickening \u3e50% of the thickness of the surrounding walls. Mean age was 53.1 ± 11.7 years (73.8% female). Most were overweight or obese (88.7%) and more than half (53.2%) had the metabolic syndrome. One third (34.8%) had abnormal carotid ultrasound findings (either cIMT ≥75th percentile for gender and age or presence of plaque). Among those with abnormal carotid ultrasound, the majority were classified as being at low 10-year risk for cardiovascular events. Carotid ultrasound reclassified nearly a third of the cohort as being at high risk. This discordance between 10-year FRS and carotid ultrasound was noted whether risk was assessed for hard coronary events or global risk. Concordance between FRS and carotid ultrasound findings was best when long-term (30-year) risk was assessed and no subject with an abnormal carotid ultrasound was categorized as low risk by the 30-year FRS algorithm. Conclusions: Integration of carotid ultrasound findings to coronary risk assessments and use of longer term prediction models may provide better risk assessment in this minority population, with earlier initiation of appropriate therapies

Characterising endothelial CaMKII oxidation in a cellular model of inflammation to investigate the therapeutic potential of a novel anti-oxidant stent coating

Drug eluting stent use is widespread but restenosis due to endothelial dysfunction remains a challenge. Ca2+/Calmodulin-dependent protein kinase II-delta (CaMKIIδ) is implicated as a key modulator of cardiovascular pathology and hyper-activation of CaMKIIδ promotes endothelial inflammation and oxidative stress [1]. This study has characterised different endothelial cell responses for the purpose of testing a novel antioxidant stent coating. Interleukin 1-beta (IL-1β) or tissue necrosis factor-alpha (TNFα) have been used to mimic pro-inflammatory signalling and CaMKII activation observed following endothelial stress. Human umbilical vein and coronary artery endothelial cells (HUVECs and HCAECs) were treated with IL-1β or TNFα (both 10ng/ml) up to 6 h. Quantitative immunoblotting was used to assess pro-inflammatory signalling, phospho-P65 (pP65; Cell Signalling) and CaMKII activation, phospho-CaMKII (pCaMKII; Thermo Fisher) and oxidised-CaMKII (oxCaMKII; GeneTex) expression. Increased pP65 expression was observed following 30 min stimulation with either IL-1β or TNFα in HUVECs[2]. Similarly, significant increases in pP65 expression were observed in HCAECs following stimulation with both cytokines (IL-1β: 1.93±0.29 and TNFα: 1.54±0.15 mean fold-change c.f. control ±S.E.M; n=4, p<0.05). This indicates both cell types produce similar pro-inflammatory responses following cytokine stimulation. In contrast, CaMKII activation in response to cytokine stimulation was different across cell types. Significant activation (via oxidation) of CaMKII was observed in HUVECs only after IL-1β stimulation[2]. In HCAECs however, activation of CaMKII was observed only after TNFα stimulation, via both oxidation (TNFα (6h): 1.50±0.12, mean fold-change c.f. control ±S.E.M; n=4, p<0.05), and phosphorylation (TNFα (30min): 1.22±0.05, mean fold-change c.f. control ±S.E.M; n=4, p<0.05).This work highlights the differences in responses of HUVECs and HCAECs to stimulation by inflammatory cytokines. Results demonstrate the importance of considering the endothelial model used for research into therapeutic intervention

Metabolic Health Has Greater Impact on Diabetes than Simple Overweight/Obesity in Mexican Americans

To compare the risk for diabetes in each of 4 categories of metabolic health and BMI. Methods. Participants were drawn from the Cameron County Hispanic Cohort, a randomly selected Mexican American cohort in Texas on the US-Mexico border. Subjects were divided into 4 phenotypes according to metabolic health and BMI: metabolically healthy normal weight, metabolically healthy overweight/obese, metabolically unhealthy normal weight, and metabolically unhealthy overweight/obese. Metabolic health was defined as having less than 2 metabolic abnormalities. Overweight/obese status was assessed by BMI higher than 25 kg/m2. Diabetes was defined by the 2010 ADA definition or by being on a diabetic medication. Results. The odds ratio for diabetes risk was 2.25 in the metabolically healthy overweight/obese phenotype (95% CI 1.34, 3.79), 3.78 (1.57, 9.09) in the metabolically unhealthy normal weight phenotype, and 5.39 (3.16, 9.20) in metabolically unhealthy overweight/obese phenotype after adjusting for confounding factors compared with the metabolically healthy normal weight phenotype. Conclusions. Metabolic health had a greater effect on the increased risk for diabetes than overweight/obesity. Greater focus on metabolic health might be a more effective target for prevention and control of diabetes than emphasis on weight loss alone

The Definition of Insulin Resistance Using HOMA-IR for Americans of Mexican Descent Using Machine Learning

Objective The lack of standardized reference range for the homeostasis model assessment-estimated insulin resistance (HOMA-IR) index has limited its clinical application. This study defines the reference range of HOMA-IR index in an adult Hispanic population based with machine learning methods. Methods This study investigated a Hispanic population of 1854 adults, randomly selected on the basis of 2000 Census tract data in the city of Brownsville, Cameron County. Machine learning methods, support vector machine (SVM) and Bayesian Logistic Regression (BLR), were used to automatically identify measurable variables using standardized values that correlate with HOMA-IR; K-means clustering was then used to classify the individuals by insulin resistance. Results Our study showed that the best cutoff of HOMA-IR for identifying those with insulin resistance is 3.80. There are 39.1% individuals in this Hispanic population with HOMA-IR\u3e3.80. Conclusions Our results are dramatically different using the popular clinical cutoff of 2.60. The high sensitivity and specificity of HOMA-IR\u3e3.80 for insulin resistance provide a critical fundamental for our further efforts to improve the public health of this Hispanic population