Increased Urinary Angiotensin-Converting Enzyme 2 in Renal Transplant Patients with Diabetes

Angiotensin-converting enzyme 2 (ACE2) is expressed in the kidney and may be a renoprotective enzyme, since it converts angiotensin (Ang) II to Ang-(1-7). ACE2 has been detected in urine from patients with chronic kidney disease. We measured urinary ACE2 activity and protein levels in renal transplant patients (age 54 yrs, 65% male, 38% diabetes, n = 100) and healthy controls (age 45 yrs, 26% male, n = 50), and determined factors associated with elevated urinary ACE2 in the patients. Urine from transplant subjects was also assayed for ACE mRNA and protein. No subjects were taking inhibitors of the renin-angiotensin system. Urinary ACE2 levels were significantly higher in transplant patients compared to controls (p = 0.003 for ACE2 activity, and p≤0.001 for ACE2 protein by ELISA or western analysis). Transplant patients with diabetes mellitus had significantly increased urinary ACE2 activity and protein levels compared to non-diabetics (p<0.001), while ACE2 mRNA levels did not differ. Urinary ACE activity and protein were significantly increased in diabetic transplant subjects, while ACE mRNA levels did not differ from non-diabetic subjects. After adjusting for confounding variables, diabetes was significantly associated with urinary ACE2 activity (p = 0.003) and protein levels (p<0.001), while female gender was associated with urinary mRNA levels for both ACE2 and ACE. These data indicate that urinary ACE2 is increased in renal transplant recipients with diabetes, possibly due to increased shedding from tubular cells. Urinary ACE2 could be a marker of renal renin-angiotensin system activation in these patients

Urinary ACE2 activity and protein in control subjects and renal transplant (Tx) recipients.

<p>(A) Graph depicts box plots of urinary ACE2 activity in control subjects (n = 50) and Tx recipients (n = 100). For each box plot, median values are indicated by the line within the box, with value shown beside or above the line. The box represents 50% of the values (25^th and 75^th percentiles), with the upper bar representing the 90^th percentile and the lower bar representing the 10^th percentile. Open circles indicate outliers. * p = 0.003, Control vs Tx recipients. (B) Graph depicts box plots of urinary ACE2 protein by ELISA in control subjects and Tx recipients. ** p<0.001, Control vs Tx recipients. (C) Graph depicts box plots of urinary ACE2 protein by western analyses in control subjects and Tx recipients. Densitometry analysis was performed on both urinary ACE2 bands (120 kDa and 90 kDa), and the sum of the two bands was used for quantitative comparisons. **p = 0.001, Control vs Tx recipients.</p

Subject demographic data.

<p>Values are medians with interquartile range in parentheses. Abbreviations: eGFR: estimated glomerular filtration rate, ACR: urine albumin to Cr ratio, CNI: calcineurin inhibitor. ^a calcineurin inhibitor, mycophenolate mofetil or azathioprine, and corticosteroid. N/A: not applicable.</p

Urinary Ang II and Ang-(1-7).

<p>(A) Graph shows box plots of RIA for Ang II in urine specimens from transplant recipients without diabetes (no Diabetes) or with diabetes (Diabetes). For each box plot, median values are indicated by the line within the box, with value shown above the line. The box represents 50% of the values (25^th and 75^th percentiles), with the upper bar representing the 90^th percentile and the lower bar representing the 10^th percentile. Open circles indicate outliers. * p = 0.027, Diabetes vs. No Diabetes. (B) Graph shows box plots of EIA for Ang-(1-7) in urine specimens from transplant recipients without or with diabetes. There was no significant difference between the two groups (p = 0.126).</p

Urinary ACE2 activity and protein in renal transplant recipients: Effect of diabetes.

<p>(A) Graph depicts box plots of urinary ACE2 activity in transplant recipients without diabetes (No Diabetes), or with diabetes (Diabetes). For each box plot, median values are indicated by the line within the box, with value shown above the line. The box represents 50% of the values (25^th and 75^th percentiles), with the upper bar representing the 90^th percentile and the lower bar representing the 10^th percentile. Open circles indicate outliers. * p<0.001, Diabetes vs. No Diabetes; n = 62 (No Diabetes), n = 38 (Diabetes). (B) Graph depicts box plots of urinary ACE2 protein by ELISA in transplant recipients without diabetes (No Diabetes), or with diabetes (Diabetes). *p<0.001, Diabetes vs. No Diabetes. (C) Graph depicts box plots of urinary ACE2 protein by western analysis in transplant patients without diabetes (No Diabetes), or with diabetes (Diabetes). *p<0.001, Diabetes vs. No Diabetes. Above graph is representative immunoblot for ACE2 in urine, showing bands at 120 kDa and 90 kDa. Densitometry analysis was performed on both bands, and the sum of the two bands was used for quantitative comparisons. The protein bands for ACE2 in urine specimens were not observed when membranes were incubated with the secondary antibody alone, bypassing the primary antibody step. Lanes 1–3, No Diabetes. Lanes 4–6, Diabetes. Lane 7: recombinant human ACE2 protein (hACE2), used as a positive control. (D) Representative immunoblot for urinary ACE2 treated without (−) or with (+) the deglycosylase enzyme PNGase F. Lanes 1+, 2+, 3+, and 4+ show a reduction in the sizes of urinary ACE2 fragments to ∼85 kDa and ∼65 kDa in urine samples treated with PNGase F. Lanes 1 and 2, No Diabetes. Lanes 3 and 4, Diabetes. Lane 5: recombinant human ACE2 protein (hACE2).</p

Multiple linear regression adjusting for common variables in transplant recipients.

<p>All analyses adjusted for eGFR, age, gender, albuminuria, diabetes, hypertension, and use of calcineurin inhibitors. Abbreviations: eGFR: estimated glomerular filtration rate, ACR: urine albumin to Cr ratio.</p

Integration of Women’s Cardiovascular Health Content Into Healthcare Provider Education: Results of a Rapid Review and National Survey

Despite its importance, formal education in healthcare training programs on sex- and gender-specific cardiovascular disease (CVD) risk factors, symptoms, treatment, and outcomes is lacking. We completed rapid reviews of the academic and grey literature to describe the current state of women-specific CVD education in medical, nursing, and other healthcare education programs. Second, we analyzed results from a Canada-wide survey of healthcare professional education programs to identify gaps in curricula related to sex- and gender-specific training in CVD. Our academic review yielded only 15 peer-reviewed publications, and our online search only 20 healthcare education programs, that note that they specifically address women, or sex and gender, and CVD in their curricula. Across both searches, the majority of training and education programs were from the USA, varied greatly in length, delivery mode, and content covered, and lacked consistency in evaluation. Of surveys sent to 213 Canadian universities and other entry-to-practice programs, 80 complete responses (37.6%) were received. A total of 47 respondents (59%) reported that their programs included women-specific CVD content. Among those programs without content specific to CVD in women, 69.0% stated that its inclusion would add “quite a bit” or “a great deal” of value to the program. This study highlights the emerging focus on and substantial gaps in women-specific CVD training and education across healthcare education programs. All medical, nursing, and healthcare training programs are implored to incorporate sex- and gender-based CVD content into their regular curricula as part of a consolidated effort to minimize gaps in cardiovascular care. Résumé: Malgré la prévalence des maladies cardiovasculaires (CV), les programmes d’enseignement en santé accordent peu d’attention aux facteurs de risque, aux symptômes, aux traitements et aux issues selon le sexe ou le genre. Premièrement, nous avons fait une revue rapide de la littérature universitaire et la littérature grise pour faire état de la formation sur les maladies CV spécifiques aux femmes dans les programmes d’enseignement en médecine, en soins infirmiers et autres domaines de la santé. Deuxièmement, nous avons analysé les résultats d’une enquête menée à l’échelle du Canada sur des programmes de formation professionnelle pour cerner les lacunes dans les programmes au chapitre de la formation sur les maladies CV en fonction du sexe et du genre. Notre analyse de la littérature universitaire a permis de relever seulement 15 publications révisées par des pairs à ce sujet, et notre recherche en ligne a mis au jour seulement 20 programmes d’enseignement qui comportent un volet portant spécifiquement sur les femmes, ou bien le sexe et le genre, et les maladies CV. Ces deux enquêtes ont révélé que la majorité des programmes de formation et d’enseignement étaient aux États-Unis et qu’ils présentaient une grande diversité sur le plan de la durée, du mode d’enseignement et du contenu abordé. De plus, les méthodes d’évaluation n’étaient pas uniformes. Parmi les sondages envoyés à 213 universités et programmes d’admission à la pratique au Canada, 80 réponses complètes (37,6 %) ont été reçues. Quarante-sept des établissements qui ont répondu (59 %) ont signalé que leurs programmes comprenaient du contenu portant sur les maladies CV spécifiques aux femmes. Parmi les établissements dont les programmes ne comportaient aucun contenu spécifique aux femmes, 69,0 % ont indiqué qu’une telle inclusion ajouterait « beaucoup » ou « énormément » de valeur au programme. Cette étude met en lumière l’attention nouvelle accordée à la formation et à l’enseignement sur les maladies CV spécifiques aux femmes ainsi que les lacunes substantielles observées à cet égard dans les programmes d’enseignement en santé. Les programmes de formation en médecine, en soins infirmiers et en santé sont vivement invités à intégrer du contenu spécifique au sexe et au genre pour ce qui est des maladies CV dans un effort concerté visant à réduire les lacunes dans les soins cardiovasculaires

State of the science in women's cardiovascular disease : a Canadian perspective on the influence of sex and gender

Cardiovascular disease (CVD) is the leading cause of premature death for women in Canada.1 Although it has long been recognized that estrogen impacts vascular responses in women, there is emerging evidence that physiologic and pathophysiologic cardiovascular responses are uniquely affected across the spectrum of a woman's life. Despite a global understanding that manifestations and outcomes of CVD are known to differ between men and women, uptake of the recognition of sex and gender influences on the clinical care of women has been slow or absent.2 To highlight the need for better research, diagnosis, treatment, awareness, and support of women with CVD in Canada, the Canadian Women's Heart Health Alliance (CWHHA), supported by the University of Ottawa Heart Institute, and in collaboration with the Heart and Stroke Foundation of Canada (HSFC), undertook a comprehensive review of the evidence on sex‐ and gender‐specific differences in comorbidities, risk factors, disease awareness, presentation, diagnosis, and treatment across the entire spectrum of CVD. The intent of this review was not to directly compare women and men on epidemiological and outcome measures of CVD, but to synthesize the state of the evidence for CVD in women and identify significant knowledge gaps that hinder the transformation to clinical practice and care that is truly tailored for women, a significant health challenge that has only been recognized in Canada relatively recently. This review highlights the scarcity of Canadian data on CVD in women as part of the ongoing struggle to increase awareness of and improve outcomes for women with CVD. Because of a paucity of published Canada‐specific evidence, the purpose of this review is to provide an infrastructure to summarize world‐wide published evidence, including knowledge gaps that must be understood to then make effective recommendations to alleviate the glaring “unders” of CVD for women in Canada: under‐aware, under‐diagnosed and under‐treated, under‐researched, and under‐support