Anxiolytic effect of alamandine in male transgenic rats with low brain angiotensinogen is dependent on activation of MrgD receptors.

Alamandine is a peptide hormone belonging to the renin-angiotensin system (RAS). It acts through the Mas-related G-protein coupled receptor type D, MrgD, which is expressed in different tissues, including the brain. In the present study, we hypothesize that a lack of alamandine, through MrgD, could cause the anxiety-like behavior in transgenic rats with low brain angiotensinogen [TGR(ASrAOGEN)680]. Adult male transgenic rats exhibited a significant increase in the latency to feeding time in the novelty suppressed feeding test and a decrease in the percentage of time and entries in the open arms in the elevated plus maze. These effects were reversed by intracerebroventricular infusion of alamandine. Pretreatment with D-Pro7-Ang-(1-7), a Mas and MrgD receptor antagonist, prevented the anxiolytic effects induced by this peptide. However, its effects were not altered by the selective Mas receptor antagonist, A779. In conclusion, our data indicates that alamandine, through MrgD, attenuates anxiety-like behavior in male TGR(ASrAOGEN)680, which reinforces the importance of the counter-regulatory RAS axis as promising target for the treatment of neuropsychiatric disorders.

ISCEV standard for clinical pattern electroretinography (2024 update).

The pattern electroretinogram (PERG) is a localized retinal response evoked by a contrast-reversing pattern, usually a black and white checkerboard, which provides information about macular and retinal ganglion cell function. This document, from the International Society for Clinical Electrophysiology of Vision (ISCEV; www.iscev.org ) presents an updated and revised Standard for clinical PERG testing. This replaces the 2013 and all earlier versions. Minimum protocols for basic PERG stimuli, recording methods and reporting are specified, to promote consistency of methods for diagnosis and monitoring purposes, while responding to evolving clinical practices and technology. The main changes in the updated ISCEV Standard for clinical PERG include expanded guidance about large stimulus fields, stimulus parameters for simultaneous PERG and pattern visual evoked potential recording, baseline drift correction, and use of consistent ambient room lighting. These changes aim to provide a clinically relevant document about current practice which will facilitate good quality recordings and inter-laboratory comparisons.

ISCEV and IPS guideline for the full-field stimulus test (FST).

The full-field stimulus test (FST) is a psychophysical technique designed for the measurement of visual function in low vision. The method involves the use of a ganzfeld stimulator, as used in routine full-field electroretinography, to deliver full-field flashes of light. This guideline was developed jointly by the International Society for Clinical Electrophysiology of Vision (ISCEV) and Imaging and Perimetry Society (IPS) in order to provide technical information, promote consistency of testing and reporting, and encourage convergence of methods for FST. It is intended to aid practitioners and guide the formulation of FST protocols, with a view to future standardisation.

Diabetes and hypertension in elderly women: interactions between severity and failure to control inflammation.

Elderly women are more susceptible to the development of chronic non-communicable diseases. Among these, diabetes mellitus (DM) and systemic arterial hypertension (SAH) stand out. This work aimed to carry out an expanded study on the interactions of anthropometric, biochemical and inflammatory parameters associated with the risk of severity in elderly women with hypertension and diabetes. The study involved the evaluation of 126 elderly women with hypertension and diabetes mellitus. The women were divided according disease severity (low, moderate, high and very high). Anthropometric data were collected by bioimpedance analysis. The inflammatory and biochemical data were obtained from volunteer blood samples. Waist circumference, waist circumference/height ratio, and systolic and diastolic pressures increased with severity. Biochemical marker levels increased with risk of severity, except HDLc. In the very high risk group, there was a higher IL-1ß, IFN-γ and TNF-α production, however, lower IL-10 levels were observed. The very high risk group showed change values for the IL-10/IL-1ß, IL-10/IL-17 and IL-10/TNF-α ratios. The results showed to be extensively altered in the very high risk group, where the inflammatory profile loses its responsiveness. This is the first study that shows an expanded view of the different parameters evaluated in elderly women with hypertension and diabetes.

Impact of genetic deletion of MrgD or Mas receptors in depressive-like behaviour in mice.

OBJECTIVES: To evaluate the impact of genetic deletion of receptors of the counterregulatory arms of the renin-angiotensin system in depressive-like behaviours. METHODS: 8-12 weeks-old male mice wild type (WT, C57BL/6J) and mice with genetic deletion of MrgD (MrgD KO) or Mas receptors (Mas KO) were subjected to the Forced Swim Test (FST) and the Tail Suspension Test (TST). Brain-derived neurotrophic factor (BDNF) levels were measured by enzyme-linked immunosorbent assay (ELISA). Blockade of Mas was performed by acute intracerebroventricular (icv) injection of its selective antagonist, A779. RESULTS: No statistical difference in immobility time was observed between MrgD KO and WT male animals subjected to FST and TST. However, acute icv injection of A779 significantly increased the immobility time of MrgD KO male mice subjected to FST and TST, suggesting the involvement of Mas in preventing depressive-like behaviour. Indeed, Mas KO male animals showed increased immobility time in FST and TST, evidencing a depressive-like behaviour in these animals, in addition to a reduction in BDNF levels in the prefrontal cortex and hippocampus. No changes in BDNF levels were observed in MrgD KO male animals. CONCLUSION: Our data showed that Mas plays an important role in the neurobiology of depression probably by modulating BDNF expression. On the contrary, lack of MrgD did not alter depressive-like behaviour, which was supported by the lack of alterations in BDNF levels.

Pattern ERGs suggest a possible retinal contribution to the visual acuity loss in acute optic neuritis.

PURPOSE: Macular involvement in optic neuritis (ON) is well-recognised but poorly understood and may be of clinical relevance. This study explores macular structure-function correlates in acute ON. METHODS: This cross-sectional cohort study recruited ON patients within 14 days of symptom onset. Subjects underwent pattern electroretinography (PERG), pattern visual evoked potentials (PVEP) and optical coherence tomography (OCT) imaging. PERG P50 and N95 components were correlated with OCT data. RESULTS: Twenty-six individuals with ON were recruited, comprising eleven multiple sclerosis (MS-ON), six myelin oligodendrocyte glycoprotein associated (MOG-ON) and nine with isolated ON. These were compared with 28 healthy controls. PVEPs were undetectable in 11 (42%) of individuals with ON. When detectable, PVEP P100 was delayed (median 136 ms range 110-173 ms) and amplitude reduced (median 6 µV, range 3-14 µV) in ON compared with controls (both p < 0.001). PERG P50 component amplitudes, largely reflecting macular function, were reduced in affected eyes (median 2.3 µV; range 0.8-5.0 µV) compared with controls (3.3 µV; range 2.8-5.7 µV) and compared with fellow eyes (p < 0.001). The N95:P50 ratio was below the reference range in the affected eyes of five patients. Eight cases (32%) had subnormal P50 amplitudes (< 2.0 µV), and these patients had poorer visual acuity (p = 0.020). P50 amplitudes were positively correlated with an increase in inner nuclear layer thickness (rs = 0.36; p = 0.009) and macular ganglion cell and inner plexiform layer (mGCIPL) thickness (rs = 0.44, p = 0.022). CONCLUSION: PERG P50 component reduction reveals dysfunction of inner macular layers in acute ON and correlates with structural alterations on OCT. These early macular pathologic processes are likely to contribute to the visual loss.

Physical activity reduces intradermal bacterial load in a murine model submitted to forced swim training - a pilot study.

Regular exercise is beneficial to health. This study evaluated the effects of moderate and intense physical exercise modalities on intradermal infection by Staphylococcus aureus in a murine model. Mice that practiced moderate exercise had lower bacterial load on lymph nodes and less inflammatory infiltrate in dermis. They presented greater weight, however, less amount of epididymal fat: the weight was increased while they had fat diminished. A positive correlation was observed between lipid content and bacterial load in mice trained at moderate intensity. Animals that were under high intensity exercises presented superior bacterial load on the lymph nodes, increased neutrophil count and circulating lymphocytes, and had leukocyte recruitment to the dermis augmented, when compared to the ones in moderate exercise. These findings suggest that moderate physical activity modulates the immune response in dermal infection caused by S. aureus in a murine model.

Angiotensin-(1-7) prevents T3-induced cardiomyocyte hypertrophy by upregulating FOXO3/SOD1/catalase and downregulating NF-ĸB.

Clinical studies have shown a correlation between thyroid disorders and cardiac diseases. High levels of triiodothyronine (T3) induce cardiac hypertrophy, a risk factor for cardiac complications and heart failure. Previous results have demonstrated that angiotensin-(1-7) is able to block T3-induced cardiac hypertrophy; however, the molecular mechanisms involved in this event have not been fully elucidated. Here, we evidenced the contribution of FOXO3 signaling to angiotensin-(1-7) effects. Angiotensin-(1-7) treatment increased nuclear FOXO3 levels and reduced p-FOXO3 levels (inactive form) in isolated cardiomyocytes. Knockdown of FOXO3 by RNA silencing abrogated the antihypertrophic effect of angiotensin-(1-7). Increased expression of antioxidant enzymes superoxide dismutase 1 (SOD1 and catalase) and lower levels of reactive oxygen species and nuclear factor-κB (NF-κB) were observed after angiotensin-(1-7) treatment in vitro. Consistent with these results, transgenic rats overexpressing angiotensin-(1-7) displayed increased nuclear FOXO3 and SOD1 levels and reduced NF-κB levels in the heart. These results provide a new molecular mechanism responsible for the antihypertrophic effect of angiotensin-(1-7), which may contribute to future therapeutic targets.

Hemodynamic phenotyping of transgenic rats with ubiquitous expression of an angiotensin-(1-7)-producing fusion protein.

Activation of the angiotensin (Ang)-converting enzyme (ACE) 2/Ang-(1-7)/MAS receptor pathway of the renin-angiotensin system (RAS) induces protective mechanisms in different diseases. Herein, we describe the cardiovascular phenotype of a new transgenic rat line (TG7371) that expresses an Ang-(1-7)-producing fusion protein. The transgene-specific mRNA and the corresponding protein were shown to be present in all evaluated tissues of TG7371 with the highest expression in aorta and brain. Plasma Ang-(1-7) levels, measured by radioimmunoassay (RIA) were similar to control Sprague-Dawley (SD) rats, however high Ang-(1-7) levels were found in the hypothalamus. TG7371 showed lower baseline mean arterial pressure (MAP), assessed in conscious or anesthetized rats by telemetry or short-term recordings, associated with increased plasma atrial natriuretic peptide (ANP) and higher urinary sodium concentration. Moreover, evaluation of regional blood flow and hemodynamic parameters with fluorescent microspheres showed a significant increase in blood flow in different tissues (kidneys, mesentery, muscle, spleen, brown fat, heart and skin), with a resulting decrease in total peripheral resistance (TPR). TG7371 rats, on the other hand, also presented increased cardiac and global sympathetic tone, increased plasma vasopressin (AVP) levels and decreased free water clearance. Altogether, our data show that expression of an Ang-(1-7)-producing fusion protein induced a hypotensive phenotype due to widespread vasodilation and consequent fall in peripheral resistance. This phenotype was associated with an increase in ANP together with an increase in AVP and sympathetic drive, which did not fully compensate the lower blood pressure (BP). Here we present the hemodynamic impact of long-term increase in tissue expression of an Ang-(1-7)-fusion protein and provide a new tool to investigate this peptide in different pathophysiological conditions.

Alamandine through MrgD receptor induces antidepressant-like effect in transgenic rats with low brain angiotensinogen.

Alamandine (Ala1-Arg2-Val3-Tyr4-Ile5-His6-Pro7), a heptapeptide hormone of the renin-angiotensin system (RAS), exerts its effects through the Mas-related G-protein coupled receptor of the type D, MrgD, which is expressed in different tissues, including the brain. In the present study, we tested the hypothesis that alamandine could attenuate the depression-like behavior observed in transgenic rats with low brain angiotensinogen, TGR (ASrAOGEN)680. Transgenic rats exhibited a significant increase in the immobility time in forced swim test, a phenotype reversed by intracerebroventricular infusion of alamandine. Pretreatment with D-Pro7-Ang-(1-7), a Mas/MrgD receptor antagonist, prevented the antidepressant-like effect induced by this peptide demonstrating, for the first time, that alamandine through MrgD receptor, can modulate depression-like behavior in TGR (ASrAOGEN)680. This result shows an action of alamandine which strengthens the importance of the counter-regulatory arms of the RAS in fight and treatment of neuropsychiatric diseases.

Relevance of angiotensin-(1-7) and its receptor Mas in pneumonia caused by influenza virus and post-influenza pneumococcal infection.

Resolution failure of exacerbated inflammation triggered by Influenza A virus (IAV) prevents return of pulmonary homeostasis and survival, especially when associated with secondary pneumococcal infection. Therapeutic strategies based on pro-resolving molecules have great potential against acute inflammatory diseases. Angiotensin-(1-7) [Ang-(1-7)] is a pro-resolving mediator that acts on its Mas receptor (MasR) to promote resolution of inflammation. We investigated the effects of Ang-(1-7) and the role of MasR in the context of primary IAV infection and secondary pneumococcal infection and evaluated pulmonary inflammation, virus titers and bacteria counts, and pulmonary damage. Therapeutic treatment with Ang-(1-7) decreased neutrophil recruitment, lung injury, viral load and morbidity after a primary IAV infection. Ang-(1-7) induced apoptosis of neutrophils and efferocytosis of these cells by alveolar macrophages, but had no direct effect on IAV replication in vitro. MasR-deficient (MasR-/-) mice were highly susceptible to IAV infection, displaying uncontrolled inflammation, increased viral load and greater lethality rate, as compared to WT animals. Ang-(1-7) was not protective in MasR-/- mice. Interestingly, Ang-(1-7) given during a sublethal dose of IAV infection greatly reduced morbidity associated with a subsequent S. pneumoniae infection, as seen by decrease in the magnitude of neutrophil influx, number of bacteria in the blood leading to a lower lethality. Altogether, these results show that Ang-(1-7) is highly protective against severe primary IAV infection and protects against secondary bacterial infection of the lung. These effects are MasR-dependent. Mediators of resolution of inflammation, such as Ang-(1-7), should be considered for the treatment of pulmonary viral infections.

Primary cutaneous peripheral T-cell lymphoma, not otherwise specified: results of a multicentre European Organization for Research and Treatment of Cancer (EORTC) cutaneous lymphoma taskforce study on the clinico-pathological and prognostic features.

BACKGROUND: Cutaneous peripheral T-cell lymphoma, not otherwise specified (PTL NOS) is an aggressive, but poorly characterized neoplasm. OBJECTIVES: The European Organization for Research and Treatment of Cancer cutaneous lymphoma taskforce (EORTC CLTF) investigated 33 biopsies of 30 patients with primary cutaneous PTL NOS to analyse their clinical, histological, immunophenotypic features and outcome. METHODS: Retrospective analysis of clinical data and histopathological features by an expert panel. RESULTS: Cutaneous PTL NOS manifested clinically either with solitary or disseminated rapidly grown ulcerated tumours or disseminated papulo-nodular lesions. Histologically, a mostly diffuse or nodular infiltrate in the dermis and often extending into the subcutis was found. Epidermotropism was rarely present and only mild and focal. Unusual phenotypes were frequent, e.g. CD3+ /CD4- /CD8- and CD3+ /CD4+ /CD8+ . Moreover, 18% of the cases exhibited an aberrant expression of the B-cell marker CD20 by the tumour cells. All solitary tumours were located on the limbs and presented a high expression of GATA-3 but this did not correlate with outcome and therefore could not serve as a prognostic factor. The prognosis was shown to be generally poor with 10 of 30 patients (33%) dying of lymphoma within the follow-up of 36 months (mean value; range 3-144). The survival rates were 61% after 3 years (CI, 43-85%) and 54% after 5 years (CI, 36-81%). Small to medium-sized morphology of tumour cells was associated with a better outcome than medium to large or large tumour cells. Age, gender, clinical stage, CD4/CD8 phenotype and GATA-3 expression were not associated with prognosis. Chemotherapy was the most common treatment modality, but surgical excision and/or radiotherapy may represent an appropriate first-line treatment for solitary lesions. CONCLUSIONS: Cutaneous PTL NOS shows an aggressive course in most patients independent of initial presentation, age and phenotype. Cytomorphology was identified as a prognostic factor. The data indicate a need for more effective treatment modalities in PTL NOS.

Dynamics of Transmission of Urban Arbovirus Dengue, Zika and Chikungunya in Southwestern Region of Bahia, Brazil.

Arboviral diseases are disseminated all over the world. In Brazil, they remain neglected, alerting public authorities to possible outbreaks. Over here, we report the epidemiological indicators of Dengue from 2010 to 2015, Zika between 2015 and 2016, and Chikungunya from 2014 to 2016, within 19 municipalities of Southwestern Region of Bahia, Brazil. The data were collected from Brazilian national public information systems (SISFAD, SINAN, and IBGE) and by Endemic Control Agents. The analysis consisted of a description of vector characteristics, Home Infestation Index and characterization of human reported cases. The years 2011 and 2013 were recorded as having the highest frequencies of positive properties for the presence of the arboviruse vectors. Most municipalities presented high annual values of Home Infestation Index indicating an alert situation (62.28%). In the evaluated period, there were (i) 9,196 cases of Dengue, (ii) 636 cases of Zika and (iii) 224 cases of Chikungunya reported. This is the first report of the epidemiological characteristics of these arboviruses in the 19 municipalities of Bahia. It is believed that the data collected may contribute to public health policies aimed at controlling future epidemics of these arboviruses.

Ureaplasma diversum clearance in lung mice infection is mediated by neutrophils.

Pneumonia in cattle is one of the causes of morbidity rates and economic loss. The host response to lung infections caused by Ureaplasma diversum in bovines is virtually unknown. Here in the immune response was evaluated in a murine model for an experimental pulmonary infection by U. diversum. Therefore, AJ, BALB/C and C57BL/6 mice received intratracheal inoculation of U. diversum and were evaluated after 1, 2, 3, 7 and 14 days and the clinical specimens were collected. In bronchoalveolar lavages (BAL) an increase of inflammatory cells was observed. Neutrophils were the main cells recruited to the site of infection and the infiltration was coincided with the production of pro-inflammatory cytokines. We found a large amount of neutrophil in this initial period, followed by a decrease 7 and 14 days post infection, accompanied by bacterial clearance. Our results evidenced the presence of U. diversum within the neutrophil that suggests a phagocytic role of this cell in the elimination of the infection. The immune response features reported here are the initial evidence that healthy immune systems may control these microorganisms. This may be the first step to design new strategies immune based to control the infections in naturally infected hosts.

Gender-related differences in the modulation of anthropometric, biochemical, and immune markers by physical activity in hypertensive and diabetic individuals.

Systemic arterial hypertension (SAH) and type 2 diabetes mellitus (T2DM) compose the two major noncommunicable chronic inflammatory diseases. Physical activity has been shown as a promising complementary approach to control the systemic inflammation. However, it is still unclear whether this modulation is gender-dependent. The objective of this study was evaluate the gender-related influence of physical activity on the inflammatory response and biochemical profile of individuals with SAH and T2DM. An international physical activity questionnaire was applied to 376 individuals diagnosed with SAH and T2DM in order to access their exercises routine and was evaluated the influence of physical activity in biochemical, anthropometrical, and immunological markers involved in these disorders in men and women. Even though active individuals have exhibited lower serum levels of IL-1ß, IFN-γ, TNF-α, and IL-17A, the ratios between IL-10 and all inflammatory cytokines were higher in men than in women. Physically active individuals also demonstrated increased HDL/LDL and HDL/VLDL ratios. Moreover, multiple correlations revealed that in active women both IL-10 and TNF-α serum levels positively correlate with fasting glucose levels, and were negatively associated with HDL levels. Our findings suggest that gender-related differences dictate a distinct crosstalk between inflammatory and biochemical markers in physically active individuals.

Treatment, reuse, leaching characteristics and genotoxicity evaluation of electroplating sludge.

Electroplating sludge (ES) is a waste that is generated by the galvanization industry and is highly toxic because it contains heavy metals. This study examined the physiochemical properties of ES residue, the recovery of the present metals, and the reuse of these metals to add value to this residue and avoid environmental contamination through its inadequate disposal. The potential for leaching of ES was investigated using various tests, and a decrease in the germination speed of Lactuca sativa seeds and the appearance of chromosomal aberrations in the cytotoxicity tests were observed. The reduction of the pH and dynamic leaching conditions favor the leaching of ES heavy metals. An increase in the ES concentration in soil decreases the speed of germination and increases the number of chromosomal aberrations that are related to aneugenic phenomena that promote tumor development. Metals were recovered through solubilization, followed by selective precipitation. The recovery of heavy metals from ES decreased its toxicity by eliminating toxic components. The reuse of the metals in the electroplating process may reduce the cost of disposal of ES, thereby rendering it an economically and environmentally friendly alternative. The products that were galvanized using the recovered solution showed the best results in the corrosion test, thereby demonstrating the viability of the use of this solution in industrial galvanization.

Characterization of the Renin-Angiotensin System in Aged Cavernosal Tissue and its Role in Penile Fibrosis.

BACKGROUND: The renin-angiotensin system (RAS) plays an important role in erectile function. The RAS contains 2 major axes: one deleterious, composed of ACE-Ang II-AT1 receptor, and another protective, composed of ACE2-Ang-(1-7)-Mas receptor. While aging is a well-known cause for development of male sexual disorders, little is known about local regulation of the RAS in age-related erectile dysfunction (ED). AIM: The present study aimed to assess regulation of the RAS in aging-associated ED rat model and evaluate possible options for disease management through pharmacological modulation of the RAS. METHODS: Penile tissues were harvested from 3-, 12-, and 24-month-old Wistar rats. Local expression of major RAS components and ED markers was measured by RT-PCR. Protein expression of RAS components was assessed by western blot. Collagen deposition was measured by Sirius Red and immunohistochemical staining. Evaluation of collagen content was also performed in penile sections of Mas-knockout mice by Sirius Red and Masson's trichrome stainings. Finally, the effect of Ang-(1-7) pretreatment on TGF-ß-induced myofibroblast activation was studied in primary cavernosal and immortalized fibroblasts. OUTCOMES: Experimental results highlighted the essential role of the RAS in modulation of cavernosal fibrosis. RESULTS: The present study demonstrates local expression of angiotensinogen mRNA alongside with major RAS components, which suggests local autonomous functioning of the RAS within penile tissue. Gene expression analysis revealed strong positive correlation between ACE-Ang II-AT1 axis with markers for inflammation and fibrosis. While corpus cavernosum from 24-month-old rats was characterized by increased collagen deposition, protein expression of ACE, AT1, and Mas was shown to be upregulated in the penile tissue of this group. At the same time, penile sections from Mas-knockout mice (FVB/N background) were also shown to have increased collagen deposition. Finally, it was demonstrated that Ang-(1-7) treatment of primary cavernosal and immortalized fibroblasts was able to alleviate TGF-ß-induced fibroblast-to-myofibroblast transition. CLINICAL TRANSLATION: The present study suggests Ang-(1-7) treatment as a possible strategy for pharmacological management of fibrosis-associated ED in aging. STRENGTHS & LIMITATIONS: The link between the RAS and penile fibrosis, indicated by a holistic screening of different ED markers, was confirmed by in vivo and in vitro data. However, results, presented in the manuscript, need to be further reinforced by human data. Important to note, the main goal of the study was to characterize RAS regulation in aging condition rather than state any causal relationships. CONCLUSION: Present study characterizes RAS regulation in aging-associated ED and indicates its important role in cavernosal fibrosis. Bragina ME, Costa-Fraga F, Sturny M, et al. Characterization of the Renin-Angiotensin System in Aged Cavernosal Tissue and its Role in Penile Fibrosis. J Sex Med 2020;17:2129-2140.

Quantitative 1 H NMR spectroscopy (qNMR) in the early process development of a new quorum sensing inhibitor.

2-methyl-5,6,7,8-tetrahydro-2H-chromen-4(3H)-one (called 6-oxo) is presented as a new AI-1 quorum sensing inhibitor for Vibrio harveyi. The development of a chemical process to afford traceable materials for new biological assays demands the development of analytical methods to ensure their purity and quality. This work describes the use of quantitative 1 H nuclear magnetic resonance (NMR) spectroscopy (qNMR) to assess the purity of a sample of 6-oxo (99.88%) and a sample of its major process impurity (E)-1-(2-hydroxycyclohex-2-en-1-yl)but-2-en-1-one (called HCB; 98.28%). To explore the scope of the use of qNMR to quantify the amount of low-content components in samples related to the chemical process for 6-oxo synthesis, this work also determined the amount of 6-oxo in two HCB samples: (a) the high-purity HCB sample described above and (b) a crude HCB sample collected during the chemical process. Despite the complexity of the crude sample, the amount of 6-oxo was readily assessed and could help to estimate the extent to which 6-oxo was already formed during the HCB synthesis. This information can help the understanding of how the process parameters can be modified to improve the performance of the whole process, by controlling the reaction mechanisms working at each step of this chemical process. In this context, our results reinforce qNMR as a complementary analytical tool for the quantification of the main component found in a sample, contributing to the standardization of reference materials and thus allowing the development of analytical methods for process control and traceability of the samples used for biological assays.

Angiotensin-(1-7) Prevents Lipopolysaccharide-Induced Autophagy via the Mas Receptor in Skeletal Muscle.

Skeletal muscle atrophy, which occurs in lipopolysaccharide (LPS)-induced sepsis, causes a severe muscle function reduction. The increased autophagy contributes to sepsis-induced skeletal muscle atrophy in a model of LPS injection, increasing LC3II/LC3I ratio, autophagy flux, and autophagosomes. Angiotensin-(1-7) (Ang-(1-7)) has anti-atrophic effects via the Mas receptor in skeletal muscle. However, the impact of Ang-(1-7) on LPS-induced autophagy is unknown. In this study, we determined the effect of Ang-(1-7) on sepsis-induced muscle autophagy. C57BL6 wild-type (WT) mice and mice lacking the Mas receptor (KO Mas) were injected with LPS together with the systemic administration of Ang-(1-7) to determine autophagy in skeletal muscle. We also evaluated autophagy and p38 and c-Jun N-terminal kinase (JNK)activation. Our results show that Ang-(1-7) prevents LPS-induced autophagy in the diaphragm, tibialis anterior, and gastrocnemius of WT mice, which is demonstrated by a decrease in the LC3II/LC3I ratio and mRNA levels of lc3b and ctsl. This effect was lost in KO Mas mice, suggesting the role of the Mas receptor. The results in C2C12 cells show that Ang-(1-7) reduces several LPS-dependent effects, such as autophagy (LC3II/LC3I ratio, autophagic flux, and autophagosomes), activation of p38 and JNK, B-cell lymphoma-2 (BCL2) phosphorylation, and disassembly of the Beclin1/BCL2 complex. In conclusion, Ang-(1-7)/Mas receptor reduces LPS-induced autophagy in skeletal muscle. In vitro assays indicate that Ang-(1-7) prevents LPS-induced autophagy and modifies the MAPK signaling and the disassembly of a complex involved at the beginning of autophagy.

Genetic deletion of the alamandine receptor MRGD leads to dilated cardiomyopathy in mice.

We have recently described a new peptide of the renin-angiotensin system, alamandine, a derivative of angiotensin-(1-7). Mas-related G protein-coupled receptor member D (MrgD) was identified as its receptor. Although similar cardioprotective effects of alamandine to those of angiotensin-(1-7) have been described, the significance of this peptide in heart function is still elusive. We aimed to evaluate the functional role of the alamandine receptor MrgD in the heart using MrgD-deficient mice. MrgD was localized in cardiomyocytes by immunofluorescence using confocal microscopy. High-resolution echocardiography was performed in wild-type and MrgD-deficient mice (2 and 12 wk old) under isoflurane anesthesia. Standard B-mode images were obtained in the right and left parasternal long and short axes for morphological and functional assessment and evaluation of cardiac deformation. Additional heart function evaluation was performed using Langendorff isolated heart preparations and inotropic measurements of isolated cardiomyocytes. Immunofluorescence indicated that the MrgD receptor is expressed in cardiomyocytes, mainly in the membrane and perinuclear and nuclear regions. Echocardiography showed left ventricular remodeling and severe dysfunction in MrgD-deficient mice. Strikingly, MrgD-deficient mice presented a pronounced dilated cardiomyopathy with a marked decrease in systolic function. Echocardiographic changes were supported by the data obtained in isolated hearts and inotropic measurements in cardiomyocytes. Our data add new evidence for a major role for alamandine/MrgD in the heart. Furthermore, our results indicate that we have identified a new gene implicated in dilated cardiomyopathy, unveiling a new target for translational approaches aimed to treat heart diseases. NEW & NOTEWORTHY The renin-angiotensin system is a key target for cardiovascular therapy. We have recently identified a new vasodepressor/cardioprotective angiotensin, alamandine. Here, we unmasked a key role for its receptor, Mas-related G protein-coupled receptor member D (MrgD), in heart function. The severe dilated cardiomyopathy observed in MrgD-deficient mice warrants clinical and preclinical studies to unveil its potential use in cardiovascular therapy.