Background and aims: Retinal arteriolar narrowing and venular widening are associated with increased cardiovascular risk, even at young ages. Whether diet contributes to early microvascular changes in children is not widely explored. We explored the associations of frequency of healthy and unhealthy food group intake with retinal vessel calibers in black and white children.Methods and results: This study included school-aged (5-9 years) black (N = 433, 7.46 f 0.98 years), and white (N = 403, 7.43 f 0.82 years) children. Anthropometric and blood pressure measurements were taken, along with retinal vessel calibers (central retinal arteriolar (CRAE) and venular (CRVE) equivalents). Frequencies of food group intake were assessed using a foodfrequency questionnaire. A factor analysis was performed to describe food group patters. Independent associations between retinal vessel calibers and frequencies of food group intake and food group patters were explored. In black children, cookies, cakes, and biscuits were associated with narrower arterioles (p < 0.05). In white children, cold sweetened beverages were associated with narrower arterioles (p = 0.02), whereas salty snacks were associated with narrower arterioles (p = 0.01) and wider venules (p < 0.05). Fruits were positively associated with CRAE (p = 0.03) in white children only.Conclusion: A higher frequency of unhealthy food group consumption was associated with retinal arteriolar narrowing and venular widening in both black and white children. However, fruit intake was shown beneficial for retinal microvascular health in white children only. Our findings may highlight the importance of promoting healthy eating patterns from early childhood which may reduce the risk of premature cardiovascular disease development.(c) 2023 The Authors. Published by Elsevier B.V. on behalf of The Italian Diabetes Society, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition and the Department of Clinical Medicine and Surgery, Federico II University.

Background and aim: Adverse changes in large arteries and retinal microvascular calibres are predictors for hypertension and stroke. These adverse changes were reported in response to the suppressed night-time dipping as well as in hypertensive individuals with exaggerated morning surge. However, it is unknown whether young healthy individuals with normal morning surge, but with non-dipping night-time blood pressure profile, will already present with early onset of micro and macrovascular changes. We investigated the relationship of morning surge with retinal vessel calibres and large artery stiffness in healthy young dippers and non-dippers. Methods: We included black and white participants (N = 323), aged 20–30 years, determined sleep-trough surge. We determined central retinal artery and vein equivalents from fundus images, as well as carotid-femoral pulse wave velocity. Results: Dippers had a greater morning surge (within normal range) (p < 0.001) and a greater proportion of individuals with exaggerated morning surge (p < 0.001). In dippers only, we found a positive relationship between central retinal artery equivalent and morning surge (p = 0.040). In sensitivity analysis, the association between central retinal artery equivalent and morning surge in dippers lost significance (p = 0.09) when 24-hour diastolic blood pressure was replaced with night-time diastolic blood pressure. However, an independent negative association between central retinal artery equivalent and night-time diastolic blood pressure (p < 0.001) emerged. Normal morning surge of dippers was positively associated with central retinal artery equivalent (dependent on night-time dipping). Conclusion: Normal night-time dipping may therefore be involved in preserving retinal arteriolar diameter of dippers, which may reflect a lower vascular resistance.

Objective Identifying individuals at increased risk of early vascular ageing (EVA) is paramount to inform intervention and prevention strategies and curb the increasing burden of cardiovascular disease. Methods We stratified and phenotyped pre-screened young apparently healthy South African adults (20-30 yrs) (n=1,041) into vascular ageing profile groups based on carotid femoral pulse wave velocity (cfPWV) percentiles (healthy vascular ageing [HVA]; average vascular ageing [AVA] and EVA). We further compared various anthropometric, cardiovascular (CV), oxidative stress and lifestyle risk factors and determined factor scores to explore associations between CV measures and factor clusters to explore associations in those at risk of EVA. Results Young adults in the EVA group displayed marked phenotypic characteristics in terms of anthropometry, CV, and lifestyle risk factors, even though cfPWV were within healthy ranges. Blood pressure (brachial and central) and cfPWV were all incrementally higher across all three vascular ageing groups (p-trend <= 0.011). Hypertension, lifestyle risk factors such as self-reported smoking and alcohol consumption were all highest in the EVA group (p-trend <= 0.046). Additionally, in the EVA group only, cfPWV (adj. R-2=0.028; b=0.171; p=0.042) associated positively with Factor 2 (oxidative stress and antioxidant capacity). No associations existed between Factor 1 (basic lipids) and any anthropometric or CV measures (p.0.050). Conclusion Young adults with higher cfPWV presented with a less favourable vascular profile and more unhealthy lifestyle behaviours compared to groups with lower cfPWV. In the EVA group, cfPWV positively associated with a cluster of oxidative stress and antioxidant capacity. Early lifestyle behaviours may have the ability to modify the balance between oxidants and antioxidants, potentially contributing to early onset arterial stiffness.

Background: Carotid-femoral pulse wave velocity (PWV) is the gold-standard noninvasive measure of arterial stiffness. Data comparing tonometry-based devices such as the SphygmoCor XCEL to simpler brachial-cuff-based estimates of PWV, such as from the Mobil-O-Graph in African populations are sparse. We therefore aimed to compare PWV measured by the Mobil-O-Graph and the SphygmoCor XCEL device in a sample of South African women and children. Methods: Women (n = 85) 29 years [interquartile range (IQR): 29-69] and their children/grandchildren (n = 27) 7 years (IQR: 4-11) were recruited for PWV measurement with Mobil-O-Graph and SphygmoCor XCEL on the same day. Wilcoxon signed-rank test, regression analysis, spearman correlation and Bland-Altman plots were used for PWV comparison between devices. Results: For adults, the SphygmoCor XCEL device had a significantly higher PWV (7.3 m/s, IQR: 6.4-8.5) compared with the Mobil-O-Graph (5.9 m/s, IQR: 5.0-8.1, P = 0.001) with a correlation coefficient of 0.809 (P <= 0.001). Bland--Altman analysis indicated an acceptable level of agreement but significant bias (mean difference PWV: 0.90 +/- 1.02 m/s; limits of agreement: -1.10 to 2.90). The odds of having a PWV difference more than 1 m/s decreased with a higher age [odds ratio (OR): 0.95, 95% confidence interval (95% CI) = 0.92-0.98] and increased with greater height (OR: 1.10, 95% CI = 1.01-1.21, P = 0.03) in multivariable analysis. In children, the Bland-Altman indicated an excellent level of agreement (-0.03 +/- 0.63 m/s; limits of agreement: -1.26 to 1.21), but no correlation was found (r(s) = 0.08, P = 0.71). Conclusion: Particularly in younger and taller women, the Mobil-O-Graph significantly underestimated PWV compared with the SphygmoCor. Although no correlation was found between the two devices for children, further research is required due to the small sample size. Furthermore, the clinical value of both methods in young African populations requires further investigation.

Background and aims: Childhood obesity and high blood pressure (BP) are main determinants for cardiovascular disease development with regional and ethnic differences. Narrower arteriolar (CRAE) and wider venular (CRVE) retinal vessel diameters are sensitive markers of early vascular compromise in children. We aimed to compare retinal vessel diameters and investigate associations and odds ratios with body mass index (BMI) and BP in a multi-national/ethnic childhood study. Methods: BMI, systolic (SBP) and diastolic BP (DBP) were screened in 929 black and white South African (SA) and 1171 Swiss children (aged 5-9 years). Retinal assessments were performed using a retinal vessel analyzer to determine CRAE and CRVE. Results: Black SA children had wider CRVE compared to white SA and Swiss children (all p < 0.001). However, BMI or BP was not associated with CRVE in black SA children. Higher BMI and BP associated with narrower CRAE in all children, except for BMI in black SA children, in whom narrower CRAE was found for every unit increase in SBP (beta = -0.199 mu m, p = 0.001) and DBP (beta = -0.312 mu m, p < 0.001). Obesity (OR:1.38[1.01; 1.89]), hypertension (OR:1.90[1.53; 2.36]) and black ethnicity (OR:1.50[1.18; 1.92]) increased the likelihood for arteriolar narrowing. Conclusions: Black SA children presented with wider retinal venules compared to their white SA and Swiss peers, which was unexplained by conventional risk factors. The overall risk of arteriolar narrowing was driven by obesity, hypertension and ethnicity. Our findings indicate the importance to differentiate cardiovascular risk by microvascular phenotype in different populations and ethnicity early in life.

Objectives and methods: Hypertension is a growing health concern in childhood populations and individuals of African descent. As the kidneys play a significant role in blood pressure regulation, we compared alpha-1 microglobulin (A1M) as a marker of proximal tubular function between young healthy black and white children (n = 957; aged: 5-9 years) and explored its association with blood pressure. Results: The black children had higher levels of A1M (P < 0.001) and higher DBP (P < 0.001) when compared with their white counterparts. In multiple regression analysis, SBP (adj. R-2 = 0.173, beta = 0.151; P < 0.001) and DBP (adj. R-2 = 0.110, beta = 0.179; P < 0.001) associated positively with A1M in the black children. In binary logistic regression, each standard deviation increase in A1M increased the odds of having elevated blood pressure by 28% (P = 0.002) in the black group, independent of age, sex, BMI z-score and body height. No significance was reached in the white children. Conclusion: Our findings highlight the importance of a marker of proximal tubular function, especially in children of black ethnicity, in the setting of elevated blood pressure. Early childhood screening for elevated blood pressure remains essential in order to promote primary prevention of hypertension and early onset kidney damage in children.

Objectives: Low or high sympatho-adrenal-medullary axis (SAM) and hypothalamic-pituitary-adrenal axis (HPA) dysregulation reflect chronic stress. Retinal vessel dynamics may relate to SAM, HPA activity and stroke risk. Our objectives were therefore to assess the relationships between retinal vessel, SAM and HPA responses, and to determine stroke risk. Methods: A prospective bi-ethnic gender cohort (n = 275, 45 +/- 9 years) was included. Urine/serum/saliva samples for SAM [norepinephrine:creatinine ratio (u-NE)] and HPA [adrenocorticotrophic hormone (ACTH), cortisol] were obtained at baseline, three-year follow up and upon flicker light-induced provocation. Diastolic ocular perfusion pressure was measured as a marker of hypo-perfusion. Retinal arterial narrowing and venous widening calibres were quantified from digital images in the mydriatic eye. A validated stress and stroke risk score was applied. Results: An interaction term was fitted for venous dilation in u-NE tertiles (p = 0.05) and not in u-NE median/quartiles/ quintiles. Independent of race or gender, tertile 1 (low u-NE) had a 112% increase in u-NE, decreases in cortisol, and no changes in ACTH over three years (positive feedback). Tertile 3 (high u-NE) contradictorily had decreases in u-NE and cortisol, and increases in ACTH (negative feedback). In tertile 1, reduced arterial dilation, and faster arterial vasoconstriction and narrowing were related to higher SAM activity and hypo-perfusion (p = 0.05), whereas delayed venous dilation, recovery and widening were related to cortisol hypo-secretion (p = 0.05). In tertile 1, delayed venous recovery responses predicted stress and stroke risk [odds ratio 4.8 (1.2-19.6); p = 0.03]. These associations were not found in u-NE tertiles 2 and 3. Conclusions: In response to low norepinephrine, a reflex increase in SAM activity occurred, enhancing arterial vasoconstriction and hypo-perfusion. Concomitant HPA dysregulation attenuated retinal vein vasoactivity and tone, reflecting delayed vein recovery responses and non-adaptation to stress. These constrained vein recovery responses are indicative of increased chronic stress and stroke risk.

Aims: Plasminogen activator inhibitor-1 (PAI-1), traditionally associated with fibrinolysis, is increasingly implicated in impaired vascular function. However, studies on its association with microvascular function are limited to the cutaneous and coronary microvascular beds in older and diseased individuals. To better understand its potential involvement in the early stages of disease development, we investigated the associations of retinal vasodilatory responses to flicker light with PAI-1 activity (PAI-1act) in young and healthy individuals. Methods: We included healthy Black and White women and men (n = 518; aged 20-30 years), and measured plasma PAI-1act and retinal vasodilatory responses to flicker light provocation. We also collected demographic and lifestyle data, measured blood pressure, anthropometry, blood lipids, inflammatory and other biomarkers. Results: In multivariate regression analyses, maximal retinal venular dilation associated independently and inversely with PAI-1act (adj. R2 = 0.11; 13 = -0.15; p = 0.001) in the total group. In exploratory subgroup analyses, this association remained in White women (adj. R2 = 0.07; 13 = -0.23; p = 0.005), and was more robust with younger age and lower blood pressure and in non-smokers, but also with greater central adiposity, higher low-density lipoprotein cholesterol and inflammation (all p < 0.05). Conclusions: Our data suggest that in young individuals, PAI-1 may already be associated with subclinical microvascular dysfunction.

Platelet extravasation during inflammation is under-appreciated. In wild-type (WT) mice, a central corneal epithelial abrasion initiates neutrophil (PMN) and platelet extravasation from peripheral limbal venules. The same injury in mice expressing low levels of the β2-integrin, CD18 (CD18hypo mice) shows reduced platelet extravasation with PMN extravasation apparently unaffected. To better define the role of CD18 on platelet extravasation, we focused on two relevant cell types expressing CD18: PMNs and mast cells. Following corneal abrasion in WT mice, we observed not only extravasated PMNs and platelets but also extravasated erythrocytes (RBCs). Ultrastructural observations of engorged limbal venules showed platelets and RBCs passing through endothelial pores. In contrast, injured CD18hypo mice showed significantly less venule engorgement and markedly reduced platelet and RBC extravasation; mast cell degranulation was also reduced compared to WT mice. Corneal abrasion in mast cell-deficient (KitW-sh/W-sh) mice showed less venule engorgement, delayed PMN extravasation, reduced platelet and RBC extravasation and delayed wound healing compared to WT mice. Finally, antibody-induced depletion of circulating PMNs prior to corneal abrasion reduced mast cell degranulation, venule engorgement, and extravasation of PMNs, platelets, and RBCs. In summary, in the injured cornea, platelet and RBC extravasation depends on CD18, PMNs, and mast cell degranulation.

Background: Obesity and hypertension prevalence among children are a concern, with limited evidence available on sex and ethnic differences in childhood blood pressure. We aimed to determine the number of children with hypertension and obesity to identify unique adiposity and blood pressure characteristics by sex and ethnicity, and to estimate the odds of having elevated blood pressure with increasing adiposity. Methods: We included 1062 healthy children (5-9 years of age) in an observational school-based study in South Africa. Pediatric validated automated devices were used to measure brachial blood pressure and performed pulse wave analysis to assess central hemodynamics. Standard anthropometry was carried out to determine body composition and demographic questionnaires were completed. Results: Almost 20% of children were overweight/obese and 14.1% had elevated blood pressure or hypertension (22.8%). Ethnic differences included greater adiposity in white compared with black children (all P < 0.0001), but higher DBP and total vascular resistance in black compared with white children (both P < 0.05). DBP and total vascular resistance were also higher in girls than boys (both P < 0.01). A 51-60% increased risk of developing elevated blood pressure was observed for 1SD (standard deviation) increase of sex-specific BMI [1.60 (1.4-1.8); P < 0.0001] and waist/height ratio [1.51 (1.3-1.7); P Conclusion: Unique sex and ethnic differences in body composition and blood pressure exist in prepubescent children, with overweight/obesity increasing the risk of elevated blood pressure. Our findings support primary prevention strategies to combat the growing burden of hypertension and obesity-related diseases in youth.

Background & Aims Nitric oxide, a major inhibitory nonadrenergic, noncholinergic neurotransmitter that relaxes smooth muscle, may be implicated in the pathophysiology of visceral hypersensitivity in irritable bowel syndrome (IBS). Impaired bioavailability of the nitric oxide precursor molecule L-arginine and higher concentrations of methylarginines (endogenous inhibitors of nitric oxide synthesis) are known to impair nitric oxide synthesis in numerous gastrointestinal cell types. We therefore examined serum concentrations of L-arginine and the methylarginines in a nested case-control study, to assess whether these factors are associated with adult IBS. Methods Data on clinical characteristics, methylarginines, and L-arginine (measured using LC-MS/MS) were collected from a random population-based cohort of Australian adults (median age = 64 years; IQR = 60-70). Cases of IBS, defined according to Rome III criteria (N = 156), and controls (N = 332) were identified from within the cohort at the 5-year follow-up. Results In adjusted logistic regression analyses, L-arginine, asymmetric dimethylarginine, symmetric dimethylarginine, L-arginine/asymmetric dimethylarginine ratio, and Kessler-10 psychological distress scores were significantly associated with IBS (p > 0.05). Similar results were found for IBS subtypes. Higher serum L-arginine concentration had the strongest association with IBS diagnosis, with an odds ratio of 9.03 for those with serum L-arginine at the 75th (84 mu mol/L) versus 25th (46 mu mol/L) percentile (95% CI: 5.99-13.62). L-arginine had the best discriminative ability with a bias-adjusted area under the receiver operator characteristic curve of 0.859. Conclusions Higher serum concentrations of L-arginine and endogenous methylarginines are strongly associated with IBS in adults.

PURPOSE:The purpose of this study was to use a mouse model of diet-induced obesity to determine if corneal dysfunction begins prior to the onset of sustained hyperglycemia and if the dysfunction is ameliorated by diet reversal. METHODS:Six-week-old male C57BL/6 mice were fed a high fat diet (HFD) or a normal diet (ND) for 5-15 weeks. Diet reversal (DiR) mice were fed a HFD for 5 weeks, followed by a ND for 5 or 10 weeks. Corneal sensitivity was determined using aesthesiometry. Corneal cytokine expression was analyzed using a 32-plex Luminex assay. Excised corneas were prepared for immunofluorescence microscopy to evaluate diet-induced changes and wound healing. For wounding studies, mice were fed a HFD or a ND for 10 days prior to receiving a central 2mm corneal abrasion. RESULTS:After 10 days of HFD consumption, corneal sensitivity declined. By 10 weeks, expression of corneal inflammatory mediators increased and nerve density declined. While diet reversal restored nerve density and sensitivity, the corneas remained in a heightened inflammatory state. After 10 days on the HFD, corneal circadian rhythms (limbal neutrophil accumulation, epithelial cell division and Rev-erbα expression) were blunted. Similarly, leukocyte recruitment after wounding was dysregulated and accompanied by delays in wound closure and nerve recovery. CONCLUSION:In the mouse, obesogenic diet consumption results in corneal dysfunction that precedes the onset of sustained hyperglycemia. Diet reversal only partially ameliorated this dysfunction, suggesting a HFD diet may have a lasting negative impact on corneal health that is resistant to dietary therapeutic intervention.

Background People living with the Human Immunodeficiency Virus (PLHIV) have an increased susceptibility to develop non-communicable diseases such as cardiovascular disease (CVD). Infection with HIV contributes to the development of CVD independent of traditional risk factors, with endothelial dysfunction being the central physiological mechanism. While HIV-related mortality is declining due to antiretroviral treatment (ART), the number of deaths due to CVD is rising in South Africa - the country with the highest number of PLHIV and the world's largest ART programme. The EndoAfrica study was developed to determine whether HIV infection and ART are associated with cardiovascular risk markers and changes in vascular structure and function over 18 months in adults from different provinces of South Africa. This paper describes the rationale, methodology and baseline cohort profile of the EndoAfrica study conducted in the North West Province, South Africa. Methods In this case-control study, conducted between August 2017 and June 2018, 382 volunteers of African descent (276 women; 106 men), comprising of 278 HIV infected and 104 HIV free individuals were included. We measured health behaviours, a detailed cardiovascular profile, and performed biomarker analyses. We compared baseline characteristics, blood pressure, vascular function and biochemical markers between those infected and HIV free. Results At baseline, the HIV infected participants were older (43 vs 39 years), less were employed (21% vs 40%), less had a tertiary education (7% vs 16%) and their body mass index was lower (26 vs 29 kg/m(2)) than that of the HIV free participants. While the cardiovascular profile, flow-mediated dilation and pulse wave velocity did not differ, glycated haemoglobin was lower (p = 0.017) and total cholesterol, high density lipoprotein cholesterol, triglycerides, gamma-glutamyltransferase and tobacco use were higher (allp < 0.047) in PLHIV. Conclusion Despite PLHIV being older, preliminary cross-sectional analysis suggests that PLHIV being treated with ART do not have poorer endothelial or vascular function compared to the HIV free participants. More detailed analyses on the baseline and follow-up data will provide further clarity regarding the cardiovascular profile of South Africans living with HIV.

Background and aims: Heart rate variability (HRV) is a main determinant of autonomic function and related to the development of hypertension and cardiovascular (CV) disease. Hyper-tension develops in black populations at an earlier age, which could be due to differences in the autonomic nervous system activity and sodium/potassium handling in black and white popula-tions. We investigated whether HRV is associated with 24 h urinary sodium and potassium excretion and blood pressure (BP) in a young bi-ethnic cohort. Methods and results: We examined 423 black and 483 white healthy adults (aged 24.5 +/- 3.1 years) for 24 h HRV, including standard deviation of normal RR intervals (SDNN) reflecting auto-nomic variations over time, and root mean square of successive differences (RMSSD) reflecting parasympathetic activity. We measured 24 h urinary sodium and potassium concentration and BP. The black group had lower SDNN and potassium excretion as well as higher RMSSD, sodium and Na/k ratio compared to the white group (all p < 0.05). Only in black individuals, urinary po-tassium excretion was independently and negatively associated with SDNN (0[95% CI];-0.26 [-0.50;-0.02]ms) and RMSSD (-0.14[-0.27;-0.01]ms, p < 0.05). One unit increase in sodium/po-tassium (Na/K) ratio was associated with higher SDNN (0[95% CI]; 3.04[0.89; 5.19]ms) and RMSSD (1.60[0.41; 2.78]ms) in the black cohort only (both p < 0.001). In both groups elevated 24 h diastolic BP was associated with lower RMSSD (p < 0.05). Conclusion: Lower potassium excretion and higher Na/K ratio related independently to higher HRV in young and healthy black adults. A better ethnic-specific understanding of sodium and po-tassium handling is required as part of preventive cardiology, especially in black individuals. Clinical trial registration: ClinicalTrials.gov Identifier: NCT03292094; URL: https://clinicaltrials. gov/ct2/show/NCT03292094. (c) 2020 The Italian Diabetes Society, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

Counseling graduate students may begin practicum with low self-efficacy regarding their counseling abilities and skills. In the current study, we implemented a small-series ( N = 11) single-case research design to assess the effectiveness of the practicum experience to increase counseling students’ self-efficacy. Analysis of participants’ scores on the Counselor Activity Self-Efficacy Scale yielded treatment effects indicating that the practicum experience encompassing direct services, group supervision, and triadic supervision may be effective for increasing counselor self-efficacy. Given that the practicum experience with triadic supervision was a promising approach for improving counseling graduate students’ self-efficacy, we provide implications for counselor educators to integrate triadic supervision and self-efficacy to the forefront of discussions.

Background Leptin is a vasoactive peptide that has been linked to diseases associated with macrovascular deterioration. What is still uncertain is its involvement in the microvasculature. Since microvascular changes are presumed to precede macrovascular deterioration, we examined whether measures of the retinal microvasculature are associated with leptin in healthy, young Black and White individuals. Method We included 283 Black and 289 White men and women (aged 20-30 years). We determined serum leptin, calculated central retinal artery and vein equivalents and arterio-venous ratio. We also measured retinal vessel responses to light flicker provocation. Results Black men were leaner and had lower leptin than White men, whereas Black women had increased adiposity and leptin compared to White women (all p<0.001). Black groups had narrower retinal arteries, and greater maximum arteriolar and venular dilations in response to light flicker than the White groups (p<0.001). Arterio-venous ratio associated negatively with leptin (all p <= 0.044) in all groups (except Black women), but was lost upon adjustment for body mass index and other covariates. We found an inverse association between maximal venular dilation and leptin only in Black men in single and multiple regression analyses (Std beta = -0.22; R-2=0.05; p=0.035). No associations were found between other retinal measures with leptin in the other groups. Conclusion We found an independent, negative association between retinal vein dilation with leptin in healthy, young Black men, suggesting a potential detrimental role for leptin in regulating microvascular responses in a population group known to be at greater risk of cardiovascular disease development.

Platelets and neutrophils are often recruited at sites of microvascular inflammation. While neutrophil extravasation in inflammation is well characterized, platelet extravasation in inflammation is poorly understood and has only been documented in a small number of studies. We showed previously that mice expressing low levels of the leukocyte integrin CD18 (CD18hypo) had reduced total recruitment of platelets (intravascular and extravascular) in the corneal limbus following corneal epithelial wound injury. In this study we sought to determine the role of CD18 in platelet extravasation. Following corneal wound injury, wild‐type (WT) mice had evidence of platelet extravasation across limbal venules; however, CD18hypo mice had markedly reduced platelet extravasation. In view of isolated reports that platelets express CD18, we compared blood platelet counts and ex vivo platelet function under shear stress between CD18hypo and WT mice. Blood platelet counts were measured with a Scil Vet ABC hematology analyzer (Scil animal care company). Flow chamber assays were conducted using a microfluidic BioFlux System (Fluxion Biosciences) with plates coated with type I collagen (25 μg/ml; Helena) and blocked with 5% Bovine serum albumin. The labeled blood was perfused over the collagen‐coated surfaces at two values of fixed shear stress (16 dyne/cm2 and 30 dyne/cm2). Platelet counts and platelet adhesion to collagen at both levels of shear stress were comparable between CD18hypo and WT mice. Since a variety of leukocytes express CD18, including mast cells and neutrophils, we performed experiments to assess the role of these leukocytes in platelet extravasation. We found that CD18hypo mice had a small but statistically significant decrease in mast cell number and a marked decrease in degranulated mast cells as compared to WT mice following wound injury. Further, corneal epithelial wound injury in mice deficient in mast cells (KitW‐sh/W‐sh) resulted in a marked decrease in platelet extravasation as compared to WT mice. Finally, antibody‐induced depletion of circulating neutrophils resulted in a marked decrease in platelet extravasation relative to control mice. Overall, these studies demonstrate that platelet extravasation in a corneal wound injury model of inflammation depends on CD18, mast cells, and neutrophils.Support or Funding InformationSupported by: NIH EY018239, EY17120, HL079368 and VA BLR&D Merit Review Award I01 BX002551. M.A.B. was supported by NHLBI T32 HL139425Extravascular platelets (green) are evident surrounding venules 24 hours postwounding in wild type mice.Figure 1Platelets from CD18 hypomorphic mice had similar adhesion under shear as those from Wild‐type mice; blood platelet counts were similar in both strainsFigure 2