Comparison between radiometry and spectrophotometry for the determination of angiotensin-converting enzyme activity in cerebrospinal fluid.

Determination of angiotensin-converting enzyme (ACE) activity in cerebrospinal fluid (CSF) can help for establishing the diagnosis of neurosarcoidosis. We investigated the performance characteristics of two assays for ACE determination in 57 CSF, radiometry with [glycine-1-14C] benzoyl-L-histidyl-L-leucine and spectrophotometry with furylacryloyl-phenylalanyl-L-glycyl-L-glycine (FAPGG) as substrates. We compared both kinetic assays to an ELISA specific for human ACE. Within run and between run imprecisions were 14-17% for radiometry, 6-19% for spectrophotometry and 5-8% for ELISA. The limit of detection was 0.04 U/L for radiometry, 1.0 U/L for spectrophotometry and 0.156 µg/L for ELISA. The limit of quantification was 0.06 U/L for radiometry, 1.5 U/L for spectrophotometry, but not known for ELISA. The domain for quantification was 0.06-4.0 U/L for radiometry, 1.5-24 U/L for spectrophotometry and 0.156-10 µg/L for ELISA. Deming regression and Bland-Altman plots show good correlations between the three assays, but with high slopes, because both kinetic assays use different substrates and ELISA measures ACE molecule but not activity. Radiometry was more sensitive than spectrophotometry, which has a limit of detection above most pathological levels. ELISA could be an alternative to radiometry but only after complete evaluation, determination of normal values and assessment of its clinical value. We claim for standardization of ACE determination as well as in serum as in other biological fluids, in particular CSF.

Human Tissue Angiotensin Converting Enzyme (ACE) Activity Is Regulated by Genetic Polymorphisms, Posttranslational Modifications, Endogenous Inhibitors and Secretion in the Serum, Lungs and Heart.

Objective: Inhibitors of the angiotensin converting enzyme (ACE) are the primarily chosen drugs to treat heart failure and hypertension. Moreover, an imbalance in tissue ACE/ACE2 activity is implicated in COVID-19. In the present study, we tested the relationships between circulating and tissue (lung and heart) ACE levels in men. Methods: Serum, lung (n = 91) and heart (n = 72) tissue samples were collected from Caucasian patients undergoing lung surgery or heart transplantation. ACE I/D genotype, ACE concentration and ACE activity were determined from serum and tissue samples. Clinical parameters were also recorded. Results: A protocol for ACE extraction was developed for tissue ACE measurements. Extraction of tissue-localized ACE was optimal in a 0.3% Triton-X-100 containing buffer, resulting in 260 ± 12% higher ACE activity over detergent-free conditions. SDS or higher Triton-X-100 concentrations inhibited the ACE activity. Serum ACE concentration correlated with ACE I/D genotype (II: 166 ± 143 ng/mL, n = 19, ID: 198 ± 113 ng/mL, n = 44 and DD: 258 ± 109 ng/mL, n = 28, p < 0.05) as expected. In contrast, ACE expression levels in the lung tissue were approximately the same irrespective of the ACE I/D genotype (II: 1423 ± 1276 ng/mg, ID: 1040 ± 712 ng/mg and DD: 930 ± 1273 ng/mg, p > 0.05) in the same patients (values are in median ± IQR). Moreover, no correlations were found between circulating and lung tissue ACE concentrations and activities (Spearman's p > 0.05). In contrast, a significant correlation was identified between ACE activities in serum and heart tissues (Spearman's Rho = 0.32, p < 0.01). Finally, ACE activities in lung and the serum were endogenously inhibited to similar degrees (i.e., to 69 ± 1% and 53 ± 2%, respectively). Conclusion: Our data suggest that circulating ACE activity correlates with left ventricular ACE, but not with lung ACE in human. More specifically, ACE activity is tightly coordinated by genotype-dependent expression, endogenous inhibition and secretion mechanisms.

Investigation of Chlorella pyrenoidosa Protein as a Source of Novel Angiotensin I-Converting Enzyme (ACE) and Dipeptidyl Peptidase-IV (DPP-IV) Inhibitory Peptides.

Chlorella pyrenoidosa (C. pyrenoidosa) is a microalgae species with a remarkably high protein content that may potentially become a source of hypotensive and hypoglycemic peptides. In this study, C. pyrenoidosa proteins were extracted and hydrolyzed overnight with pepsin and trypsin with final degrees of hydrolysis of 18.7% and 35.5%, respectively. By LC-MS/MS, 47 valid peptides were identified in the peptic hydrolysate (CP) and 66 in the tryptic one (CT). At the concentration of 1.0 mg/mL, CP and CT hydrolysates inhibit in vitro the angiotensin-converting enzyme (ACE) activity by 84.2 ± 0.37% and 78.6 ± 1.7%, respectively, whereas, tested at cellular level at the concentration of 5.0 mg/mL, they reduce the ACE activity by 61.5 ± 7.7% and 69.9 ± 0.8%, respectively. At the concentration of 5.0 mg/mL, they decrease in vitro the DPP-IV activity by 63.7% and 69.6% and in Caco-2 cells by 38.4% and 42.5%, respectively. Short peptides (≤10 amino acids) were selected for investigating the potential interaction with ACE and DPP-IV by using molecular modeling approaches and four peptides were predicted to block both enzymes. Finally, the stability of these peptides was investigated against gastrointestinal digestion.

Multiple firm nodules in a young woman: an unusual presentation of sarcoidosis.

A 27-year-old woman presented to the dermatology department with a 1-year history of multiple asymptomatic violaceous lesions on her upper and lower extremities, trunk and abdomen. The lesions were firm on palpation. She had no other associated symptoms and the rest of the examination was unremarkable. An incisional biopsy showed multiple confluent granulomas composed of histiocytes devoid of necrosis surrounded by a rim of lymphocytes extending to the subcutaneous fat consistent with the diagnosis of subcutaneous sarcoidosis. The serum ACE assay was elevated at 134 IU/L. Other blood tests including complete blood count, renal and liver function tests, serum calcium and phosphate were within normal ranges and chest X-ray was unremarkable. Complete remission was achieved with an intralesional triamcinolone injection (10 mg/mL) for a few sessions. Subcutaneous sarcoidosis is a rare variation and its diagnosis requires a high index of suspicion.

Thyroid disorders and SARS-CoV-2 infection: From pathophysiological mechanism to patient management.

The World Health Organization (WHO) declared the COVID-19 epidemic to be a global pandemic in March 2020. COVID-19 is an infection caused by SARS-CoV-2, a coronavirus that utilizes the angiotensin-2 converting enzyme to penetrate thyroid and pituitary cells, and may result in a "cytokine storm". Based on the pathophysiological involvement of the pituitary-thyroid axis, the current review discusses the diagnosis of abnormal thyroid function test, and the management of patients presenting with thyrotoxicosis, thyroid-associated orbitopathy and hypothyroidism in the context of SARS-CoV-2 infection.

Involvement of the digestive system in covid-19. A review. / Afectación del aparato digestivo en la covid-19. Una revisión sobre el tema.

The SARS-CoV-2 pandemic is leading to high mortality and a global health crisis. The primary involvement is respiratory; however, the virus can also affect other organs, such as the gastrointestinal tract and liver. The most common symptoms are anorexia and diarrhea. In about half of the cases, viral RNA could be detected in the stool, which is another line of transmission and diagnosis. covid19 has a worse prognosis in patients with comorbidities, although there is not enough evidence in case of previous digestive diseases. Digestive endoscopies may give rise to aerosols, which make them techniques with a high risk of infection. Experts and scientific organizations worldwide have developed guidelines for preventive measures. The available evidence on gastrointestinal and hepatic involvement, the impact on patients with previous digestive diseases and operating guidelines for Endoscopy Units during the pandemic are reviewed.

Placental barrier against COVID-19.

Vertical transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and possible induction of pregnancy complications, including miscarriage, fetal malformations, fetal growth restriction and/or stillbirth, are serious concerns for pregnant individuals with COVID-19. According to clinical information, the incidence of vertical transmission of SARS-CoV-2 is limited to date. However, even if a neonate tests negative for SARS-CoV-2, frequent abnormal findings, including fetal and maternal vascular malperfusion, have been reported in cases of COVID-19-positive mothers. Primary receptor of SARS-CoV-2 is estimated as angiotensin-converting enzyme 2 (ACE2). It is highly expressed in maternal-fetal interface cells, such as syncytiotrophoblasts, cytotrophoblasts, endothelial cells, and the vascular smooth muscle cells of primary and secondary villi. However other route of transplacental infection cannot be ruled out. Pathological examinations have demonstrated that syncytiotrophoblasts are often infected with SARS-CoV-2, but fetuses are not always infected. These findings suggest the presence of a placental barrier, even if it is not completely effective. As the frequency and molecular mechanisms of intrauterine vertical transmission of SARS-CoV-2 have not been determined to date, intensive clinical examinations by repeated ultrasound and fetal heart rate monitoring are strongly recommended for pregnant women infected with COVID-19. In addition, careful investigation of placental samples after delivery by both morphological and molecular methods is also strongly recommended.

Why do children seem to be more protected against COVID-19? A hypothesis.

Today it remains unclear why children seem to be less likely to get infected by COVID-19 or why they appear to be less symptomatic after infections. All individuals, especially children, are exposed to various viruses including human coronavirus (CoVs) that can generally lead to respiratory infections. We hypothesize that recurrent CoVs exposure may induce an effective antiviral B and T-cell-mediated adaptive immune response, which could also be protective against COVID-19. Based on the high-homology between the Spike protein epitopes of taxonomically-related coronaviruses, we theorize that past/recurrent contact with CoVs might shield children also against the circulating COVID-19 through a possible neutralizing antibody response previously CoVs-induced. This would open up possible lines of research for the development of live-attenuated virus vaccines from CoVs. Future research is desirable to confirm or disprove such hypothesis.

Consistent localization of SARS-CoV-2 spike glycoprotein and ACE2 over TMPRSS2 predominance in placental villi of 15 COVID-19 positive maternal-fetal dyads.

INTRODUCTION: While the COVID-19 pandemic continues to have a significant global health impact, rates of maternal to infant vertical transmission remain low (<5%). Parenchymal changes of placentas from COVID-19 infected mothers have been reported by several groups, but the localization and relative abundance of SARS-CoV-2 viral proteins and cellular entry machinery has not been fully characterized within larger placental tissue cohorts. METHODS: An extended placental tissue cohort including samples from 15 COVID-19 positive maternal-fetal dyads (with n = 5 cases with evidence of fetal transmission) in comparison with 10 contemporary COVID-19 negative controls. Using comparative immunofluorescence, we examined the localization and relative tissue abundance of SARS-CoV2 spike glycoprotein (CoV2 SP) along with the co-localization of two SARS-CoV2 viral entry proteins angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). RESULTS/CONCLUSIONS: CoV2 SP was present within the villous placenta in COVID-19 positive pregnancies with and without evidence of fetal transmission. We further identified the predominance of ACE2 expression in comparison with TMPRSS2. Importantly, both CoV2 SP and ACE2 expression consistently localized primarily within the outer syncytiotrophoblast layer placental villi, a key physiologic interface between mother and fetus. Overall this study provides an important basis for the ongoing evaluation of SARS-CoV-2 physiology in pregnancy and highlights the importance of the placenta as a key source of primary human tissue for ongoing diagnostic and therapeutic research efforts to reduce the global burden of COVID-19.

[Measurement of angiotensin-I-converting enzyme in the blood: help for method validation]. / Dosage de l'enzyme de conversion de l'angiotensine-I sanguine : aide à la validation de méthode.

Blood angiotensin-converting enzyme (ACE) assay is now realized by the determination of enzyme activity on synthetic substrate, mostly furylacryloyl-phenylalanyl-L-glycyl-L-glycine (FAPGG). The matrix can be serum or heparin-plasma, with or without a separator; the assay developed on serum or plasma is not adapted to other matrix such as cerebrospinal fluid where the ACE activity is much lower. This assay has been adapted on a number of automated biochemistry analyzers with the specifications of the supplier of reagents, sometimes with modification of volumes or times for analysis. Samples can be stored at +4̊C for at least for one week, freezing at -20̊C is possible but refreezing is not advised. The assay is linear from 10 to 200 UI/L. Fidelity is excellent after calibration of the assay. Accuracy can be calculated from IQA and EQA results, and the analytical uncertainty is between 2% and 5% in function of the serum ACE value. Usual values will be soon available from studies on age brackets and sex, because ACE activity seems to be more elevated in boys during adolescence. At signature, it is interesting to have medical information on the diagnosis of sarcoidosis or its treatment including ACE inhibitors as a proof of intake; we can give a commentary on elevation of serum ACE activity from other causes than sarcoidosis and the causes for low activities.

ACE2 Expression in Pancreas May Cause Pancreatic Damage After SARS-CoV-2 Infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel coronavirus that causes coronavirus disease 2019 (COVID-19) in human beings, has caused a serious public health issue.1 Attention to pancreatic injury is lacking, which may impact patients' prognosis. In this study, we explored the expression and distribution of angiotensin-converting enzyme 2 (ACE2), the receptor of SARS-CoV-2, in the pancreas. Combined with clinical data, we showed that pancreatic injury can occur in some COVID-19 patients.

In vitro antioxidant and angiotensin-converting enzyme inhibitory activity of fermented milk with different culture combinations.

This study investigated the effects of Lactobacillus plantarum (Lp) and Bifidobacterium animalis ssp. lactis (Ba) in co-cultures with Streptococcus thermophilus (St) on changes in the acidification profile, proteolytic activity, peptide production, in vitro antioxidant activity, and angiotensin-converting enzyme (ACE) inhibitory properties of fermented milks during 21 d of storage at 4°C. The pH values and proteolysis in all batches showed a gradual decrease and increase during storage, respectively. The ACE-inhibitory activity and total antioxidant capacity of all co-fermented milk samples followed a similar pattern, with maximum values on d 6 of storage. The St starter, in conjunction with Ba or Lp or both, enhanced proteolysis, peptide generation, and ACE-inhibitory and antioxidant activity, but decreased pH values compared with St alone. The St-Ba-Lp samples showed higher DPPH• (1,1-diphenyl-2-picrylhydrazyl) radical scavenging activity, hydroxyl radical scavenging activity, and total antioxidant capacity, but similar superoxide anion scavenging activity compared to St-Ba or St-Lp samples. The St-Ba samples showed higher DPPH• radical scavenging activity but lower hydroxyl radical scavenging activity than St-Lp samples. In the ACE-inhibitory assays, the St-Lp samples exhibited relatively low activity among the co-fermented milks, digested or not. The presence of Ba and Lp in fermentation together did not affect ACE-inhibitory activity in undigested fermented milks compared with the presence of Ba alone, and St-Ba-Lp fermented milks demonstrated an increase in ACE-inhibitory activity after simulated gastrointestinal digestion in storage. Pepsin digestion largely improved ACE-inhibitory activity, except in St-Lp samples, in which the activity was reduced. Further hydrolysis by trypsin reduced final activity in digestion. This study suggests that co-cultured fermentation with probiotics improves in vitro antioxidant and ACE inhibition activity in fermented milks, and this effect is partly due to the higher proteolytic activity of probiotics.

Genetic Association of Angiotensin-Converting Enzyme (ACE) Gene I/D Polymorphism with Preterm Birth in Korean Women: Case-Control Study and Meta-Analysis.

BACKGROUND AND OBJECTIVES: The ACE gene encodes the angiotensin-converting enzyme (ACE), a component of the renin-angiotensin system. Increased ACE activity may cause abnormal regulation of placental circulation and angiogenesis, resulting in adverse pregnancy outcomes. Previous studies have reported that the insertion/deletion (I/D) polymorphism of the ACE gene is associated with the development of preterm birth (PTB). However, results of the association between ACE gene I/D and PTB are inconsistent in various populations. Therefore, we performed a case-control study and a meta-analysis to evaluate the association between ACE I/D polymorphism and PTB. Materials and Methods: We analyzed a total of 254 subjects (111 patients with PTB and 143 women at ≥38 weeks gestation) for the case-control study. For the meta-analysis, we searched Google Scholar, PubMed, and NCBI databases with the terms "ACE," "angiotensin-converting enzyme," "preterm birth," "preterm delivery," and their combinations. Results: Our results of the case-control study indicated that ACE I/D polymorphism is significantly associated with PTBs in the overdominant genetic model (odds ratio (OR) 0.57, 95% confidence interval (CI) 0.347-0.949, p = 0.029) and that the ID genotype of ACE I/D polymorphism has a protective effect for PTB (OR 0.57, 95% CI 0.333-0.986, p = 0.043). Similarly, the meta-analysis showed that the OR for the ACE gene ID genotype was 0.66 (95% CI 0.490-0.900, p < 0.01). Conclusion: The ACE gene ID genotype has a significant association with PTB and is a protective factor for PTB. A larger sample set and functional studies are required to further elucidate of our findings.

Plasma and tissue angiotensin-converting enzyme 2 activity and plasma equilibrium concentrations of angiotensin peptides in dogs with heart disease.

BACKGROUND: Angiotensin-converting enzyme 2 (ACE2) is a homologue of angiotensin-converting enzyme (ACE) and produces angiotensin peptides (APs), such as angiotensin 1-9 and 1-7 that are vasodilatory and natriuretic, and act to counterbalance angiotensin II. HYPOTHESIS: Evidence of ACE2 can be found in tissues and plasma of dogs. Equilibrium concentrations of renin angiotensin aldosterone system (RAAS) APs differ in dogs with heart disease compared to healthy dogs and recombinant human ACE2 (rhACE2) alters relative concentrations of APs. ANIMALS: Forty-nine dogs with and 34 dogs without heart disease. METHODS: Immunohistochemistry and assays for tissue and plasma ACE2 activity and equilibrium concentrations of plasma RAAS APs were performed. RESULTS: Immunolabeling for ACE2 was present in kidney and myocardial tissue. Median plasma ACE2 activity was significantly increased in dogs with congestive heart failure (CHF; 6.9 mU/mg; interquartile range [IQR], 5.1-12.1) as compared to control (2.2 mU/mg; IQR, 1.8-3.0; P = .0003). Plasma equilibrium analysis of RAAS APs identified significant increases in the median concentrations of beneficial APs, such as angiotensin 1-7, in dogs with CHF (486.7 pg/mL; IQR, 214.2-1168) as compared to those with preclinical disease (41.0 pg/mL; IQR, 27.4-45.1; P < .0001) or control (11.4 pg/mL; IQR, 7.1-25.3; P = .01). Incubation of plasma samples from dogs with CHF with rhACE2 increased beneficial APs, such as angiotensin 1-9 (preincubation, 10.3 pg/mL; IQR, 4.4-37.2; postincubation, 2431 pg/mL; IQR, 1355-3037; P = .02), while simultaneously decreasing maladaptive APs, such as angiotensin II (preincubation, 53.4 pg/mL; IQR, 28.6-226.4; postincubation, 2.4 pg/mL; IQR, 0.50-5.8; P = .02). CONCLUSIONS AND CLINICAL IMPORTANCE: Recognition of the ACE2 system expands the conventional view of the RAAS in the dog and represents an important potential therapeutic target.

Effects of Chronic Exposure to Mercury on Angiotensin-Converting Enzyme Activity and Oxidative Stress in Normotensive and Hypertensive Rats / Efeitos da Exposição Crônica ao Mercúrio sobre a Atividade da Enzima Conversora de Angiotensina e Estresse Oxidativo em Ratos Normotensos e Hipertensos

Abstract Background: Mercury's deleterious effects are associated with increased cardiovascular risk. Objective: To determine whether chronic exposure to inorganic mercury increases the activity of angiotensin-converting enzyme and its relationship with oxidative stress in several organs and tissues. Methods: We studied male Wistar and spontaneously hypertensive rats (SHR) (3-month-old) exposed or not to HgCl2 for 30 days. At the end of treatment, we investigated the following: changes in body weight, hemodynamic parameters, angiotensin-converting enzyme (ACE) activity and oxidative stress in the heart, aorta, lung, brain and kidney in hypertensive compared to normotensive animals. A value of p < 0.05 was considered significant. Results: Chronic exposure to HgCl2 did not affect weight gain in either group. Systolic blood pressure, measured weekly, did not increase in Wistar rats but showed a small increase in SHR rats. We also observed increases in left ventricular end-diastolic pressure and ACE activity in the plasma and hearts of normotensive rats. In the SHR+Hg group, ACE activity increased in plasma but decreased in kidney, lung, heart, brain and aorta. Oxidative stress was assessed indirectly by malondialdehyde (MDA) production, which increased in Hg-treated rats in both plasma and heart. In the SHR+Hg group, MDA increased in heart and aorta and decreased in lungs and brain. Conclusion: These results suggest that chronic exposure to inorganic mercury aggravates hypertension and produces more expressive changes in ACE activity and oxidative stress in SHRs. Such exposure affects the cardiovascular system, representing a risk factor for the development of cardiovascular disorders in normotensive rats and worsening of pre-existing risks for hypertension.

Resumo Fundamento: Os efeitos deletérios do mercúrio estão associados ao risco cardiovascular aumentado. Objetivo: Determinar se a exposição crônica ao mercúrio inorgânico aumenta a atividade da enzima conversora de angiotensina e sua relação com o estresse oxidativo em vários órgãos e tecidos. Métodos: Estudamos ratos Wistar e ratos espontaneamente hipertensos (SHR) (3 meses de idade) expostos ou não a HgCl2 por 30 dias. Ao final do tratamento, investigamos: alterações de peso, parâmetros hemodinâmicos, atividade da enzima conversora de angiotensina (ECA) e estresse oxidativo no coração, aorta, pulmão, cérebro e rim de animais hipertensos comparados a animais normotensos. Um valor de p < 0,05 foi considerado significativo. Resultados: A exposição crônica ao HgCl2 não afetou o ganho de peso em nenhum dos grupos. A pressão arterial sistólica, medida semanalmente, não aumentou em ratos Wistar, mas mostrou um pequeno aumento nos ratos SHR. Também observamos aumentos na pressão diastólica final do ventrículo esquerdo e na atividade da ECA no plasma e no coração de ratos normotensos. No grupo SHR + Hg, a atividade da ECA aumentou no plasma, mas diminuiu no rim, pulmão, coração, cérebro e aorta. O estresse oxidativo foi avaliado indiretamente pela produção de MDA, que aumentou nos ratos tratados com Hg tanto no plasma quanto no coração. No grupo SHR + Hg, o MDA aumentou no coração e na aorta e diminuiu nos pulmões e no cérebro. Conclusão: Estes resultados sugerem que a exposição crônica ao mercúrio inorgânico agrava a hipertensão e produz mudanças mais expressivas na atividade da ECA e no estresse oxidativo em SHRs. Essa exposição afeta o sistema cardiovascular, representando um fator de risco para o desenvolvimento de distúrbios cardiovasculares em ratos normotensos e para piorar riscos pré-existentes para hipertensão.

Effects of Chronic Exposure to Mercury on Angiotensin-Converting Enzyme Activity and Oxidative Stress in Normotensive and Hypertensive Rats.

BACKGROUND: Mercury's deleterious effects are associated with increased cardiovascular risk. OBJECTIVE: To determine whether chronic exposure to inorganic mercury increases the activity of angiotensin-converting enzyme and its relationship with oxidative stress in several organs and tissues. METHODS: We studied male Wistar and spontaneously hypertensive rats (SHR) (3-month-old) exposed or not to HgCl2 for 30 days. At the end of treatment, we investigated the following: changes in body weight, hemodynamic parameters, angiotensin-converting enzyme (ACE) activity and oxidative stress in the heart, aorta, lung, brain and kidney in hypertensive compared to normotensive animals. A value of p < 0.05 was considered significant. RESULTS: Chronic exposure to HgCl2 did not affect weight gain in either group. Systolic blood pressure, measured weekly, did not increase in Wistar rats but showed a small increase in SHR rats. We also observed increases in left ventricular end-diastolic pressure and ACE activity in the plasma and hearts of normotensive rats. In the SHR+Hg group, ACE activity increased in plasma but decreased in kidney, lung, heart, brain and aorta. Oxidative stress was assessed indirectly by malondialdehyde (MDA) production, which increased in Hg-treated rats in both plasma and heart. In the SHR+Hg group, MDA increased in heart and aorta and decreased in lungs and brain. CONCLUSION: These results suggest that chronic exposure to inorganic mercury aggravates hypertension and produces more expressive changes in ACE activity and oxidative stress in SHRs. Such exposure affects the cardiovascular system, representing a risk factor for the development of cardiovascular disorders in normotensive rats and worsening of pre-existing risks for hypertension.