Hitting the (bio)mark part 2: analyzing, interpreting, and reporting biomarker data in cardiovascular research.

Incorporating biomarkers into cardiovascular studies, including nursing research, is a common approach when identifying underlying mechanisms and providing targets for intervention. However, effective utilization of biomarker data demands careful consideration. In the analysis, interpretation, and reporting phase, there are many facets to consider, including non-normality of the data, normalization procedures, and potential confounding influences of other clinical data. Furthermore, as many studies focus on patient-reported outcomes, it is important that the analysis and interpretation of biomarkers in relation to patient-reported outcomes is rigorous and reproducible. In this article, part 2 of 2, we provide an overview of considerations for the analysis, interpretation, and reporting phases of biomarker studies. We also provide an example of these steps.

Hitting the (bio)mark part 1: selecting and measuring biomarkers in cardiovascular research.

Cardiovascular studies, including nursing research, frequently integrate biomarkers for diagnostic, prognostic, monitoring and therapeutic insights. However, effective utilization of biomarker data demands careful consideration. In the study design phase, researchers must select biomarkers that align with study objectives while considering resources and logistical factors. Additionally, a nuanced understanding of disease pathophysiology and biomarker characteristics is needed. During data collection, suitable experimental conditions and assays need to be defined. Whether researchers opt to manage these steps internally or outsource some, a comprehensive understanding of biomarker selection and experiments remains crucial. In this article, part 1 of 2, we provide an overview of considerations for the design to measurement phases of biomarker studies.

sPDGFRß and neuroinflammation are associated with AD biomarkers and differ by race: The ASCEND Study.

INTRODUCTION: There remains an urgent need to identify preclinical pathophysiological mechanisms of Alzheimer's disease (AD) development in high-risk, racially diverse populations. We explored the relationship between cerebrospinal fluid (CSF) markers of vascular injury and neuroinflammation with AD biomarkers in middle-aged Black/African American (B/AA) and non-Hispanic White (NHW) participants. METHODS: Adults (45-65 years) with a parental history of AD were enrolled (n = 82). CSF and blood biomarkers were collected at baseline and year 2. RESULTS: CSF total tau (t-tau), phosphorylated tau (p-tau), and amyloid beta (Aß)40 were elevated at year 2 compared to baseline. CSF soluble platelet-derived growth factor receptor ß (sPDGFRß) levels, a marker of pericyte injury, correlated positively with t-tau, p-tau, Aß40 markers of vascular injury, and cytokines at baseline and year 2. CSF sPDGFRß and tau were significantly lower in B/AA than NHW. DISCUSSION: Vascular dysfunction and neuroinflammation may precede cognitive decline and disease pathology in the very early preclinical stages of AD, and there are race-related differences in these relationships. HIGHLIGHTS: Cerebrospinal fluid (CSF) Alzheimer's disease (AD) biomarkers changed over 2 years in high-risk middle-aged adults. Markers of vascular dysfunction were associated with the CSF biomarkers amyloid beta and tau. AD biomarkers were lower in Black compared to non-Hispanic White individuals. Markers of vascular dysfunction were lower among Black individuals.

The Effects of Exercise on Telomere Length in Persons With Heart Failure.

BACKGROUND: Telomere length is reduced in persons with heart failure (HF). Inflammation is a putative mechanism contributing to telomere shortening. Although physical activity is known to increase telomere length, its effects in HF are unknown. OBJECTIVE: The aim of this study was to examine the effects of exercise on telomere length and its relationship with interleukin (IL)-1ß in persons with HF. METHODS: This secondary analysis of a 3-month home-based aerobic exercise intervention measured total telomere length and IL-1ß levels in persons with HF (69% with reduced ejection fraction). RESULTS: Total telomere length increased and plasma IL-1ß levels decreased in the exercise group from baseline to 3 months. Total telomere length was negatively associated with IL-1ß at baseline (r = -0.441 P = .001). CONCLUSIONS: The association between telomere length and IL-1ß suggests a relationship between inflammation and cellular aging. Moderate-intensity exercise may help maintain cellular functions. Further research is needed to examine the effects on outcomes in persons with HF.

Heart failure symptom burden, dietary intake, and inflammation: An integrative review of the literature.

Heart failure (HF) is characterized by high symptom burden including, but not limited to fatigue, dyspnea, and edema. Up to 21.5% of HF patients experience significant depressive symptoms, much higher than 7.1% in adults without HF. Diet, metabolites, and other inflammatory mechanisms have gained notable attention in recent studies for contributions to symptoms in HF. Symptoms for black adults (B/As) with HF are often influenced by lifestyle factors, which may influence their higher mortality rates; few studies address these factors. Distinguishing the links between key elements with diet, inflammation, and symptoms may bring clarity for new dietary strategies in HF clinical care. The purpose of this integrative review is to examine the existing literature regarding relationships among physiologic pathways in HF along with physical and emotional symptoms in the context of inflammation, dietary intake, tumor necrosis factor-alpha (TNF-α), a biomarker of inflammation, and trimethylamine-N-Oxide (TMAO). Based on available evidence, inflammation may be a key link between physical symptoms, diet, depression, TMAO, and TNF-α in persons with HF and warrants further examination to clarify pathological links to solidify evidence for better guidance with dietary modifications. The literature reviewed in this study demonstrates that more work is needed to examine dietary planning, social support, and differences between men and women in the B/A community. Results of this literature review call attention to the essential, personalized care needs related to symptom monitoring and dietary planning which is expected to decrease symptom burden in the HF population.

Intersectionality in Alzheimer's Disease: The Role of Female Sex and Black American Race in the Development and Prevalence of Alzheimer's Disease.

It is well known that vascular factors and specific social determinants of health contribute to dementia risk and that the prevalence of these risk factors differs according to race and sex. In this review, we discuss the intersection of sex and race, particularly female sex and Black American race. Women, particularly Black women, have been underrepresented in Alzheimer's disease clinical trials and research. However, in recent years, the number of women participating in clinical research has steadily increased. A greater prevalence of vascular risk factors such as hypertension and type 2 diabetes, coupled with unique social and environmental pressures, puts Black American women particularly at risk for the development of Alzheimer's disease and related dementias. Female sex hormones and the use of hormonal birth control may offer some protective benefits, but results are mixed, and studies do not consistently report the demographics of their samples. We argue that as a research community, greater efforts should be made to not only recruit this vulnerable population, but also report the demographic makeup of samples in research to better target those at greatest risk for the disease.

Impact of early pericardial fluid chymase activation after cardiac surgery.

Introduction: Chymase is a highly destructive serine protease rapidly neutralized in the circulation by protease inhibitors. Here we test whether pericardial fluid (PCF) chymase activation and other inflammatory biomarkers determine intensive care unit length of stay, and explore mechanisms of chymase delivery by extracellular vesicles to the heart. Methods: PCF was collected from adult patients (17 on-pump; 13 off-pump) 4 h after cardiac surgery. Extracellular vesicles (EVs) containing chymase were injected into Sprague-Dawley rats to test for their ability to deliver chymase to the heart. Results: The mean intensive care unit (ICU) stay and mean total length of stay was 2.17 ± 3.8 days and 6.41 ± 1.3 days respectively. Chymase activity and 32 inflammatory markers did not differ in on-pump vs. off-pump cardiac surgery. Society of Thoracic Surgeons Predicted Risk of Morbidity and Mortality Score (STS-PROM), 4-hour post-surgery PCF chymase activity and C-X-C motif chemokine ligand 6 (CXCL6) were all independent predictors of ICU and total hospital length of stay by univariate analysis. Mass spectrometry of baseline PCF shows the presence of serine protease inhibitors that neutralize chymase activity. The compartmentalization of chymase within and on the surface of PCF EVs was visualized by immunogold labeling and transmission electron microscopy. A chymase inhibitor prevented EV chymase activity (0.28 fmol/mg/min vs. 14.14 fmol/mg/min). Intravenous injection of PCF EVs obtained 24 h after surgery into Sprague Dawley rats shows diffuse human chymase uptake in the heart with extensive cardiomyocyte damage 4 h after injection. Discussion: Early postoperative PCF chymase activation underscores its potential role in cardiac damage soon after on- or off-pump cardiac surgery. In addition, chymase in extracellular vesicles provides a protected delivery mechanism from neutralization by circulating serine protease inhibitors.

Potential Protective Mechanisms of S-equol, a Metabolite of Soy Isoflavone by the Gut Microbiome, on Cognitive Decline and Dementia.

S-equol, a metabolite of soy isoflavone daidzein transformed by the gut microbiome, is the most biologically potent among all soy isoflavones and their metabolites. Soy isoflavones are phytoestrogens and exert their actions through estrogen receptor-ß. Epidemiological studies in East Asia, where soy isoflavones are regularly consumed, show that dietary isoflavone intake is inversely associated with cognitive decline and dementia; however, randomized controlled trials of soy isoflavones in Western countries did not generally show their cognitive benefit. The discrepant results may be attributed to S-equol production capability; after consuming soy isoflavones, 40-70% of East Asians produce S-equol, whereas 20-30% of Westerners do. Recent observational and clinical studies in Japan show that S-equol but not soy isoflavones is inversely associated with multiple vascular pathologies, contributing to cognitive impairment and dementia, including arterial stiffness and white matter lesion volume. S-equol has better permeability to the blood-brain barrier than soy isoflavones, although their affinity to estrogen receptor-ß is similar. S-equol is also the most potent antioxidant among all known soy isoflavones. Although S-equol is available as a dietary supplement, no long-term trials in humans have examined the effect of S-equol supplementation on arterial stiffness, cerebrovascular disease, cognitive decline, or dementia.

Racial Differences in XO (Xanthine Oxidase) and Mitochondrial DNA Damage-Associated Molecular Patterns in Resistant Hypertension.

BACKGROUND: We previously reported increased plasma XO (xanthine oxidase) activity in patients with resistant hypertension. Increased XO can cause mitochondrial DNA damage and promote release of fragments called mitochondrial DNA damage-associated molecular patterns (mtDNA DAMPs). Here, we report racial differences in XO activity and mtDNA DAMPs in Black and White adults with resistant hypertension. METHODS: This retrospective study includes 91 resistant hypertension patients (44% Black, 47% female) with blood pressure >140/90 mm Hg on ≥4 medications and 37 normotensive controls (30% Black, 54% female) with plasma XO activity, mtDNA DAMPs, and magnetic resonance imaging of left ventricular morphology and function. RESULTS: Black-resistant hypertension patients were younger (mean age 52±10 versus 59±10 years; P=0.001), with higher XO activity and left ventricular wall thickness, and worse diastolic dysfunction than White resistant hypertension patients. Urinary sodium excretion (mg/24 hour per kg) was positively related to left ventricular end-diastolic volume (r=0.527, P=0.001) and left ventricular mass (r=0.394, P=0.02) among Black but not White resistant hypertension patients. Patients with resistant hypertension had increased mtDNA DAMPs versus controls (P<0.001), with Black mtDNA DAMPS greater than Whites (P<0.001). Transmission electron microscopy of skeletal muscle biopsies in resistant hypertension patients demonstrates mitochondria cristae lysis, myofibrillar loss, large lipid droplets, and glycogen accumulation. CONCLUSIONS: These data warrant a large study to examine the role of XO and mitochondrial mtDNA DAMPs in cardiac remodeling and heart failure in Black adults with resistant hypertension.

Exercise and Cognitive Training Intervention Improves Self-Care, Quality of Life and Functional Capacity in Persons With Heart Failure.

This study evaluated a 12-week, home-based combined aerobic exercise (walking) and computerized cognitive training (EX/CCT) program on heart failure (HF) self-care behaviors (Self-care of HF Index [SCHFI]), disease specific quality of life (Kansas City Cardiomyopathy Questionnaire [KCCQ]), and functional capacity (6-minute walk distance) compared to exercise only (EX) or a usual care attention control (AC) stretching and flexibility program. Participants (N = 69) were older, predominately female (54%) and African American (55%). There was significant improvement in self-care management, F(2, 13) = 5.7, p < .016; KCCQ physical limitation subscale, F(2, 52) = 3.4, p < .039; and functional capacity (336 ± 18 vs 388 ± 20 m, p < .05) among the EX/CCT participants. The underlying mechanisms that EX and CCT targets and the optimal dose that leads to improved outcomes are needed to design effective interventions for this rapidly growing population.

Adaptation of Metabolomics and Microbiomic Research Protocols During the COVID-19 Pandemic.

BACKGROUND: When the COVID-19 pandemic hit in 2020, researchers in the P30 Center for the Study of Symptom Science, Metabolomics, and Multiple Chronic Conditions at Emory University's Nell Hodgson Woodruff School of Nursing faced major challenges in recruitment and data collection because of limited access to the clinic and community facilities and the risk of COVID-19 exposure associated with in-person study contact. OBJECTIVES: The purpose of this article is to (a) describe how a cadre of pilot/supplement principal investigators adapted their studies to allow for safe and trustworthy data collection during the COVID-19 pandemic (March 2020 through date of publication) and (b) discuss steps that facilitated the technical aspects of remote data collection, especially involving biological specimens. RESULTS: Four pilot studies and two administrative supplements within the center-all at different stages of execution-adopted various alternative remote recruitment, enrollment, and data and specimen collection approaches to continue their research endeavors in a way that maximized the safety of both the research participants and the research teams. DISCUSSION: The article concludes with a discussion on the importance of a participant-centered approach when using remote methods, actions, or steps initiated to facilitate the technical aspects of remote data collection and reflections on the continued use of remote research strategies beyond the COVID-19 pandemic.

Caregiver subjective and physiological markers of stress and patient heart failure severity in family care dyads.

Greater family caregiver exposure to uncontrolled patient symptoms is predictive of greater caregiver psychological and physiological stress in dementia and other chronic illnesses, but these phenomena have not been well-studied in heart failure (HF) - a disease with high symptom burden. The purpose of this study was to test the hypothesis that worse patient functional status (as reflected by increasing HF symptoms) would be associated with elevated psychological and physiological stress for the caregiver. This was a secondary analysis of data from 125 HF caregivers in the Caregiver Opportunities for Optimizing Lifestyle (COOL) study. Psychological stress was measured on four dimensions: care-related strain/burden (Oberst Caregiving Burden Scale), depression (Center for Epidemiological Studies Depression Scale), anxiety (State-Trait Anxiety Index), and general stress (Perceived Stress Scale). Physiological stress was measured by markers of HPA axis function (elevated cortisol awakening response [CAR]), endothelial dysfunction (increased PAI-1), and inflammation (increased IL-6, hsCRP). HF patient functional status was quantified by caregiver assessment of New York Heart Association (NYHA) Class. Generalized linear models were used to test associations between patient NYHA Class and stress (one model per indicator). NYHA Class (ordinal) was backwards difference coded in each model to examine caregiver stress in relation to increasing levels of HF severity. Caregivers were mostly female and in their mid-fifties, with a slight majority of the sample being African American and the patient's spouse. Overall, patient functional status was associated with greater caregiver psychological and physiological stress. In terms of psychological stress, higher NYHA Class was significantly associated with greater caregiver anxiety and general stress, but not with caregiver burden or depression. In terms of physiological stress, higher NYHA Class was associated with elevated markers in all models (elevated CAR and higher IL-6, hsCRP, and PAI-1). Across models, most associations between NYHA Class and stress were present at relatively early stages of functional limitation (i.e. Class II), while others emerged when functional limitations became more severe. To inform timing and mechanisms for much-needed caregiver interventions, research is needed to determine which aspects of HF symptomatology are most stressful for caregivers across the HF trajectory.

Daytime sleepiness predicts inflammation and ambulatory blood pressure in sleep apnoea.

INTRODUCTION: Sleepiness in obstructive sleep apnoea is associated with cardiovascular risk; however, the biological mechanisms are not known. This study explored whether those with subjective sleepiness have increased plasma tumour necrosis factor-related protein 1 (C1qTNF1), a novel adipose-derived hormone (adipokine), and 24-h ambulatory blood pressure (ABP) compared to those without sleepiness in newly diagnosed, treatment-naïve participants with obstructive sleep apnoea. METHODS: Overall, 94 participants were included in the analysis. Participants completed the Epworth Sleepiness Scale (ESS), 24-h ABP was monitored, and plasma C1qTNF1 was measured. Sleepy participants were defined as ESS≥10 and nonsleepy as ESS<10. Multiple linear regression was used to explore differences in C1qTNF1, and 24-h mean arterial pressure (MAP) between sleepy and nonsleepy participants, adjusting for age, sex, body mass index, apnoea-hypopnoea index, and smoking status. RESULTS: C1qTNF1 was significantly higher in sleepy participants (n=57) compared to nonsleepy participants (n=37) (ß=0.41 NPX, 95% CI 0.02, 0.80; p=0.04). The 24-h MAP was significantly higher in sleepy participants compared to nonsleepy participants (ß=4.06 mmHg, 95% CI 0.36, 7.77; p=0.03). CONCLUSIONS: Our findings show that sleepiness is associated with inflammation and higher 24-h MAP in sleep apnoea.

Reduced Left Atrial Emptying Fraction and Chymase Activation in Pathophysiology of Primary Mitral Regurgitation.

Increasing left atrial (LA) size predicts outcomes in patients with isolated mitral regurgitation (MR). Chymase is plentiful in the human heart and affects extracellular matrix remodeling. Chymase activation correlates to LA fibrosis, LA enlargement, and a decreased total LA emptying fraction in addition to having a potential intracellular role in mediating myofibrillar breakdown in LA myocytes. Because of the unreliability of the left ventricular ejection fraction in predicting outcomes in MR, LA size and the total LA emptying fraction may be more suitable indicators for timing of surgical intervention.

An Intervention to Improve Physical Function and Caregiver Perceptions in Family Caregivers of Persons With Heart Failure.

Objective: This randomized controlled trial was conducted to determine whether a 12-week home-based aerobic and resistance exercise program would improve physical function and caregiving perceptions among family caregivers (FCGs) of persons with heart failure. Method: Overall, 127 FCGs were randomized to one of three groups: usual care attention control (UCAC), psychoeducation only (PE), and psychoeducation plus exercise (PE + EX). Physical function measures (6-min walk test, handgrip, and upper and lower strength) and caregiving perceptions (Bakas Caregiving Outcomes Scale) were obtained at baseline and at 6 months. Results: FCGs in the PE + EX showed significant improvement in 6-min walk distance (p = .012), handgrip, and lower extremity strength compared with the PE and UCAC groups. The combined group had the greatest improvement in caregiver perceptions (p < .001). Conclusion: FCGs in the PE + EX group improved the most in physical function and caregiver perception outcomes. Directions for future research are provided.

Exercise and Cognitive Training as a Strategy to Improve Neurocognitive Outcomes in Heart Failure: A Pilot Study.

OBJECTIVE: Mild cognitive impairment, especially memory loss, is prevalent in patients with heart failure (HF) and contributes to poor clinical outcomes and higher mortality. METHODS: This study evaluated a combined aerobic exercise and cognitive training (EX/CT) program on memory, executive function, attention, processing speed and reaction time compared to exercise only or a usual care attention control (UCAC) stretching and flexibility program. Participants completed a standardized neurocognitive battery at baseline, 3 months, and 6 months along with demographic, clinical, and functional capacity (6-minute walk test). A linear mixed model analysis was used with comorbidity as a covariate. RESULTS: Sixty-nine participants were enrolled, the mean age was 61 ± 10 years, 54% were women, 55% were African American, and the mean left ventricular ejection fraction percentage was 35 ± 15. A significant group by time interaction for verbal memory was found at 3 months (F [2, 53] = 4.3, p = 0.018) but was not sustained at 6 months in the EX/CT group. Processing speed/attention differed across treatment groups between baseline and 6 months, but improvement occurred among UCAC participants. There were also significant group differences in the 6MWT distance occurring at 3 months (F [2, 52] = 3.5, p = 0.036); however, significant improvement was observed within the EX/CT group only. There were no significant differences in 6MWT in the other groups at 3 or 6 months. CONCLUSION: An EX/CT intervention was associated with improved memory in persons with HF and warrants further investigation in a larger trial. The relationship between functional capacity and cognitive function also needs further study.

Plasma xanthine oxidase activity is related to increased sodium and left ventricular hypertrophy in resistant hypertension.

BACKGROUND: The extra-renal effects of aldosterone on left ventricular (LV) structure and function are exacerbated by increased dietary sodium in persons with hypertension. Previous studies demonstrated endothelial dysfunction and increased oxidative stress with high salt diet in normotensive salt-resistant subjects. We hypothesized that increased xanthine oxidase (XO), a product of endothelial cells, is related to 24-h urinary sodium and to LV hypertrophy and function in patients with resistant hypertension (RHTN). METHODS: The study group included persons with RHTN (n = 91), defined as a blood pressure > 140/90 mmHg on ≥ 3 medications at pharmacologically effective doses. Plasma XO activity and 24-h urine were collected, and cardiac magnetic resonance imaging (MRI) was performed to assess LV function and morphology. Sixty-seven normotensive persons on no cardiovascular medications served as controls. A subset of RHTN (n = 19) received spironolactone without salt restriction for six months with follow-up XO activity measurements and MRI analyses. RESULTS: XO activity was increased two-fold in RHTN vs. normal and was positively correlated with LV mass, LV diastolic function, and 24-h urinary sodium. In RHTN patients receiving spironolactone without salt restriction, LV mass decreased, but LV diastolic function and XO activity did not improve. Baseline urinary sodium was positively associated with rate of change of LV mass to volume ratio and the LV E/A ratio. CONCLUSIONS: These results demonstrate a potential role of endothelium-derived oxidative stress and excess dietary salt in the pathophysiology of LV hypertrophy and diastolic dysfunction in persons with RHTN unaffected by the addition of spironolactone.

Effects of Exercise on ASC Methylation and IL-1 Cytokines in Heart Failure.

INTRODUCTION/PURPOSE: Inflammation contributes to heart failure (HF) progression and the interleukin (IL)-1 cytokine IL-1ß is implicated in this process. The adaptor protein apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is necessary for inflammasome activation of IL-1ß. Lower ASC methylation is associated with worse outcomes in HF. The purpose of this study was to examine the effects of exercise on changes in ASC methylation and activation of the IL-1 family cytokine IL-1ß in persons with HF. METHODS: Participants (N = 54) were randomized to receive exercise intervention (n = 38) or attention control (n = 16) for 3 months. Percent methylation of the ASC gene, plasma IL-1ß, and ASC mRNA and were obtained at baseline, 3 months, and 6 months. RESULTS: ASC methylation was higher in the exercise group as compared to control at 3 months (6.10% ± 0.5% vs 5.80% ± 0.4%; P = 0.04) and 6 months (6.07 ± 0.4 vs 5.82 ± 0.4; P = 0.04). Plasma IL-1ß was lower in the exercise group at 3 months (1.43 ± 0.5 pg·mL vs 2.09 ± 1.3 pg·mL; P = 0.02) and 6 months (1.49 ± 0.5 pg·mL vs 2.13 ± 1.4 pg·mL; P = 0.004). ASC mRNA expression was negatively associated with ASC methylation at baseline (r = -0.97, P = 0.001), 3 months (r = -0.90, P = 0.001), and 6 months (r = -0.81, P = 0.001). ASC mRNA was lower than baseline at 3 months (P = 0.004) and 6 months (P = 0.002) among those in the exercise group. ASC methylation was positively associated with 6-min walk test at baseline (r = 0.517, P < 0.001), 3 months (r = 0.464, P = 0.004), and 6 months (r = 497, P = 0.05). CONCLUSIONS: Exercise was related to increased mean percent ASC methylation and decreased IL-1ß and ASC mRNA gene expression in HF. Epigenetic regulation of ASC can be a biological mechanism by which exercise can promote better outcomes in HF.

The Third Time's a Charm: Psychometric Testing and Update of the Atlanta Heart Failure Knowledge Test.

BACKGROUND AND OBJECTIVE: Since first being published in 2009, the Atlanta Heart Failure Knowledge Test (AHFKT) has proven a reliable and valid instrument and has been used in multiple studies. Given advances in heart failure (HF) self-care, we proposed to reevaluate the psychometric properties of the AHFKTv2 across these recent studies and update the instrument. METHODS: Demographic, clinical, and baseline AHFKTv2 data from 4 intervention studies in persons with HF were combined for this analysis (N = 284). The 30 questions of the AHFKT are focused on 5 HF self-care knowledge domains: pathophysiology, nutrition, behavior, medications, and symptoms. Characteristics of the sample were analyzed using descriptive statistics; validity testing with t tests and Mann-Whitney 2-group tests and Pearson r and Spearman ρ correlations; and reliability calculations and factor analysis were performed based on tetrachoric correlations. RESULTS: Participants were 22 to 84 years of age, 66% were African American, 63% were male, and 94% had New York Heart Association class II to III HF. Mean AHFKT score was 80.6% (±11%). Hypotheses that higher levels of knowledge would be associated with higher education level (t = -2.7, P < .01) and less sodium consumption (ρ = -0.22, P = .03) were validated. Factor analysis revealed 1 general knowledge factor with good reliability, Cronbach's α was .87. Item response analysis identified individual questions requiring review and revision. CONCLUSION: Comprehensive psychometric evaluation of the AHFKTv2 confirmed its internal consistency reliability and validity and provided direction for production of the AHFKTv3 available for use in research and clinical practice.