From Angiotensin to Renin to Prorenin and from the Adrenal to the Kidney to the Placenta and the Lungs: An Historical Journey

In 1966 I carried out a study on the role of calcium on angiotensin’s stimulant effects on the adrenal medulla. Since then I have been studying the renin-angiotensin system (RAS) for over a half-century in a wide variety of biological preparations, while awareness of its complexity has exploded. My journey has involved studies on genes, proteins, organelles, cells, tissues, glands, organs and whole animals. This chapter reviews what my colleagues and I have learned from these different levels of organization and is not meant to be an update on all features of the RAS. My studies have included experiments on: perfused cat adrenal glands; genetic and second messenger control of catecholamine synthesis and secretion from cultured bovine chromaffin cells and from rats in vivo; renin storage and release in the rat kidney and secretory granules; properties of isolated renin, prorenin and renin-like proteins; hormonal and second messenger control of prorenin secretion from human utero-placental tissues; renin/prorenin in a variety of tumors; and the effect of RAS drugs in a rodent model of pulmonary fat embolism. This most recent study has direct clinical application. I conclude with what I have learned about biomedical research and lessons for the future

Fat Embolism: What We Have Learned from Animal Models

Pulmonary fat embolism may not be diagnosed before unrelated autopsy and have little clinical impact or lead to acute lung injury with fulminant fat embolism syndrome (FES). The fat may come from various anatomic locations, bone marrow being the most common. There is no specific treatment. This review discusses animal models that can lead to a better understanding of pathophysiological mechanisms underlying this condition and indicates the importance of specific cellular constituents. A hypothesis is postulated that there is a vicious cycle involving oleic acid and angiotensin II (both of which are pulmonary toxicants): oleic acid is derived from lipid embolism by pulmonary lipases that are stimulated by angiotensin; oleic acid also promotes local generation of angiotensin. The potential role of fatty acid receptors and the resolution of this cycle are discussed. Studies show there is potential for long-term effects that might not be revealed in the immediate post-recovery period. Evidence is reviewed that animals are vulnerable to “second hit” effects at a time remote from the initial event. Some beneficial pharmacological treatments are described. These include different drugs acting on the renin-angiotensin system (RAS) that could eventually serve alone or in combination for treatment or prevention. Future therapeutic developments are discussed

Chronic Fibrotic Changes in Experimental Pulmonary Embolization in the Rat Model

Comparative Medicine - OneHealth and Comparative Medicine Poster SessionIntroduction: Fat embolism, a subclinical event, occurs in many clinical settings, such as long bones fractures, liposuction and during cardiopulmonary bypass. Some cases, especially with trauma, result in fat embolism syndrome (FES), a serious manifestation of fat embolism. FES is reported to occur in 5-10% of major trauma cases and can produce profound respiratory problems that may culminate in adult respiratory distress syndrome (ARDS). Embolized fat is hydrolyzed by lipase into free fatty acids which have been shown by previous histological studies to be toxic to the lung. An animal model of fat embolism has been developed utilizing triolein given intravenously (i.v.) to rats. We hypothesized that i.v. triolein will produce histological changes in the lung that are similar to the changes seen in human FES. Methods: Following University animal care approval, unanesthetized Sprague Dawley rats (study n=13, control n=12) were injected with either triolein, 0.2 mL (study) or saline, 0.2 mL (control). Weights were recorded until necropsy at 3 weeks (n=13) and 6 weeks (n=12). Morphometric measurements were made on both H&E and fat-stained tissues from the lungs, heart, kidneys and spleen. All vessels were examined using high magnification fields. Arterial wall thickness (lumen patency) was calculated by vessel luminal and external diameters. The medial-adventitial ratio was calculated from the outer medial diameter divided by the outer adventitial diameter. These values were keyed into statistical software and analysis as a function of time and treatment was calculated using t-tests with significance noted at a p<0.05. Results: Gross pathological changes were seen in lung, heart, kidneys, liver and spleen of the triolein group. Pulmonary histological examination revealed diffuse intra-alveolar hemorrhages and edema with peri-bronchial inflammation. Vasculitis was more prominent in the peri-bronchial areas as well. Pulmonary arteries revealed significant medial thickening as compared with the control groups with lumen patency p=0.004. Adventitia/media ratio, with large variability in the triolein group, was not statistically significant. Conclusions: Our data showed that injected triolein remains in the rat lung after 3 and 6 weeks with associated vascular and septal damage in the lung tissue compared to controls. Discussion: This study is a continuation of our previous study showing an increase of severe pulmonary damage within 3-6 hours following triolein induced fat embolism in the rat, reaching a peak at 96 hrs post injection. Despite unmedicated recovery of general condition and body weight and reopening of the pulmonary arteries and arterioles, collagen and vasculitis persisted up to 6 weeks. Further studies are needed to verify the eventual recovery or the organ evolution toward chronic fibrosis

The placental renin-angiotensin system and oxidative stress in pre-eclampsia

There is an inverse correlation between human birthweight and umbilical venous angiotensin II (AngII) concentrations. Oxidative stress and increased pro-renin receptor (PRR) both enhance the cleavage of angiotensin I from angiotensinogen (AGT). Pre-eclampsia, a hypertensive disorder of pregnancy, manifests as high blood pressure and proteinuria, and is a state of increased oxidative stress. Objectives, study design and main outcome measures Hypothesis: Pre-eclampsia will be associated with increased placental expression of components of the renin–angiotensin system, which could result in reduced infant birthweight. Biopsies were taken 1 cm from the placental edge from 27 normotensive controls and 23 pre-eclamptic White European women. Immunohistochemistry was performed for AGT, PRR, glutathione peroxidase 3 (GPx3) and the AT1R and AT2R AngII receptors. Protein expression was semi-quantitatively assessed (H-score). Results: AT1R expression was significantly increased in pre-eclamptic placentae, and negatively correlated with birthweight (r = −0.529, P = 0.009). AT1R expression was also negatively correlated with GPx3 expression overall (r = −0.647; P = 0.005). AT2R expression positively correlated with AGT (r = 0.615, P = 0.002) in the pre-eclamptic placentae only. Conclusions: The raised AT1R expression in pre-eclampsia, together with inadequate antioxidant protection, possibly through lower GPx activity, might enhance the vasoconstrictor effect of locally-generated AngII, contributing to the restricted fetal growth characteristic of pre-eclampsia. Conversely, the AT2R:AGT association within the pre-eclamptic placenta may provide a compensatory mechanism

Effect of oxygen on the expression of renin-angiotensin system components in a human trophoblast cell line

During the first trimester, normal placental development occurs in a low oxygen environment that is known to stimulate angiogenesis via upregulation of vascular endothelial growth factor (VEGF). Expression of the placental renin-angiotensin system (RAS) is highest in early pregnancy. While the RAS and oxygen both stimulate angiogenesis, how they interact within the placenta is unknown. We postulated that low oxygen increases expression of the proangiogenic RAS pathway and that this is associated with increased VEGF in a first trimester human trophoblast cell line (HTR-8/SVneo). HTR-8/SVneo cells were cultured in one of three oxygen tensions (1%, 5% and 20%). RAS and VEGF mRNA expression were determined by qPCR. Prorenin, angiotensin converting enzyme (ACE) and VEGF protein levels in the supernatant, as well as prorenin and ACE in cell lysates, were measured using ELISAs. Low oxygen significantly increased the expression of both angiotensin II type 1 receptor (AGTR1) and VEGF (both P < 0.05). There was a positive correlation between AGTR1 and VEGF expression at low oxygen (r = 0.64, P < 0.005). Corresponding increases in VEGF protein were observed with low oxygen (P < 0.05). Despite no change in ACE1 mRNA expression, ACE levels in the supernatant increased with low oxygen (1% and 5%, P < 0.05). Expression of other RAS components did not change. Low oxygen increased AGTR1 and VEGF expression, as well as ACE and VEGF protein levels, suggesting that the proangiogenic RAS pathway is activated. This highlights a potential role for the placental RAS in mediating the proangiogenic effects of low oxygen in placental development

13C-phenylalanine breath test detects altered phenylalanine kinetics in schizophrenia patients

Phenylalanine is an essential amino acid required for the synthesis of catecholamines including dopamine. Altered levels of phenylalanine and its metabolites in blood and cerebrospinal fluid have been reported in schizophrenia patients. This study attempted to examine for the first time whether phenylalanine kinetics is altered in schizophrenia using L-[1-13C]phenylalanine breath test (13C-PBT). The subjects were 20 chronically medicated schizophrenia patients (DSM-IV) and the same number of age- and sex-matched controls. 13C-phenylalanine (99 atom% 13C; 100 mg) was administered orally and the breath 13CO2 /12CO2 ratio was monitored for 120 min. The possible effect of antipsychotic medication (risperidone (RPD) or haloperidol (HPD) treatment for 21 days) on 13C-PBT was examined in rats. Body weight (BW), age and diagnostic status were significant predictors of the area under the curve of the time course of Δ13CO2 (‰) and the cumulative recovery rate (CRR) at 120 min. A repeated measures analysis of covariance controlled for age and BW revealed that the patterns of CRR change over time differed between the patients and controls and that Δ13CO2 was lower in the patients than in the controls at all sampling time points during the 120 min test, with an overall significant difference between the two groups. Chronic administration of RPD or HPD had no significant effect on 13C-PBT indices in rats. Our results suggest that 13C-PBT is a novel laboratory test that can detect altered phenylalanine kinetics in chronic schizophrenia patients. Animal experiments suggest that the observed changes are unlikely to be attributable to antipsychotic medication

Genetic analyses of diverse populations improves discovery for complex traits

Genome-wide association studies (GWAS) have laid the foundation for investigations into the biology of complex traits, drug development and clinical guidelines. However, the majority of discovery efforts are based on data from populations of European ancestry1–3. In light of the differential genetic architecture that is known to exist between populations, bias in representation can exacerbate existing disease and healthcare disparities. Critical variants may be missed if they have a low frequency or are completely absent in European populations, especially as the field shifts its attention towards rare variants, which are more likely to be population-specific4–10. Additionally, effect sizes and their derived risk prediction scores derived in one population may not accurately extrapolate to other populations11,12. Here we demonstrate the value of diverse, multi-ethnic participants in large-scale genomic studies. The Population Architecture using Genomics and Epidemiology (PAGE) study conducted a GWAS of 26 clinical and behavioural phenotypes in 49,839 non-European individuals. Using strategies tailored for analysis of multi-ethnic and admixed populations, we describe a framework for analysing diverse populations, identify 27 novel loci and 38 secondary signals at known loci, as well as replicate 1,444 GWAS catalogue associations across these traits. Our data show evidence of effect-size heterogeneity across ancestries for published GWAS associations, substantial benefits for fine-mapping using diverse cohorts and insights into clinical implications. In the United States—where minority populations have a disproportionately higher burden of chronic conditions13—the lack of representation of diverse populations in genetic research will result in inequitable access to precision medicine for those with the highest burden of disease. We strongly advocate for continued, large genome-wide efforts in diverse populations to maximize genetic discovery and reduce health disparities. © 2019, The Author(s), under exclusive licence to Springer Nature Limited

Sociotechnical agendas: reviewing future directions for energy and climate research

The field of science and technology studies (STS) has introduced and developed a “sociotechnical” perspective that has been taken up by many disciplines and areas of inquiry. The aims and objectives of this study are threefold: to interrogate which sociotechnical concepts or tools from STS are useful at better understanding energy-related social science, to reflect on prominent themes and topics within those approaches, and to identify current research gaps and directions for the future. To do so, the study builds on a companion project, a systematic analysis of 262 articles published from 2009 to mid-2019 that categorized and reviewed sociotechnical perspectives in energy social science. It identifies future research directions by employing the method of “co-creation” based on the reflections of sixteen prominent researchers in the field in late 2019 and early 2020. Drawing from this co-created synthesis, this study first identifies three main areas of sociotechnical perspectives in energy research (sociotechnical systems, policy, and expertise and publics) with 15 topics and 39 subareas. The study then identifies five main themes for the future development of sociotechnical perspectives in energy research: conditions of systematic change; embedded agency; justice, power, identity and politics; imaginaries and discourses; and public engagement and governance. It also points to the recognized need for pluralism and parallax: for research to show greater attention to demographic and geographical diversity; to stronger research designs; to greater theoretical triangulation; and to more transdisciplinary approaches