Angiotensin-(1-12): Does It Exist? A Critical Evaluation in Humans, Rats, and Mice.

BACKGROUND: Angiotensin-(1-12), measured by a self-developed, polyclonal antibody-based radioimmunoassay, has been suggested to act as an alternative precursor of angiotensin II. A more reliable detection method would be liquid chromatography-tandem mass spectrometry. METHODS: We set up the quantification of human and murine angiotensin-(1-12) by liquid chromatography-tandem mass spectrometry and then used this method to measure angiotensin-(1-12) in human and mouse blood samples, as well as in mouse brain and kidney. We also verified ex vivo angiotensin-(1-12) generation and metabolism in human blood samples incubated at 37 °C. RESULTS: Stabilization of blood in guanidine hydrochloride was chosen for sample collection since this allowed full recovery of spiked angiotensin-(1-12). Angiotensin-(1-12) was undetectable in human blood samples when incubating nonstabilized plasma at 37 °C, while angiotensin-(1-12) added to nonstabilized human plasma disappeared within 10 minutes. Stabilized human blood samples contained angiotensin II, while angiotensin-(1-12) was undetectable. Blood, hearts, and kidneys, but not brains, of wild-type mice and rats contained detectable levels of angiotensin II, while angiotensin-(1-12) was undetectable. In renin knockout mice, all angiotensins, including angiotensin-(1-12), were undetectable at all sites, despite a 50% rise in angiotensinogen. Angiotensin-(1-12) metabolism in human blood plasma was not affected by renin inhibition. Yet, blockade of angiotensin-converting enzyme and aminopeptidase A, but not of chymase, neutral endopeptidase, or prolyl oligopeptidase, prolonged the half-life of angiotensin-(1-12), and angiotensin-converting enzyme inhibition prevented the formation of angiotensin II. CONCLUSIONS: We were unable to detect intact angiotensin-(1-12) in humans or mice, either in blood or tissue, suggesting that this metabolite is an unlikely source of endogenous angiotensins.

Deciphering global patterns of forest canopy rainfall interception (FCRI): A synthesis of geographical, forest species, and methodological influences.

Forest canopy rainfall interception (FRCI) is an essential hydrological process that governs water and biogeochemical cycles in forest ecosystems. Identifying patterns and relationships of FCRI using a systematic review is key to improving our knowledge supporting new experiment research, modeling, and application. In this meta-analysis, we aimed to delineate the canopy interception (CI), throughfall (TF), and stemflow (SF) concerning geographical and forest variables and experimental methodologies. We leveraged peer-reviewed 170 articles across 234 sites globally, extracting TF, CI, SF, geographical, forest, and experimental aspects. We applied multivariate statistical procedures to discern the principal influences on TF, CI, and SF and examined their multicollinearity. In addition, we developed Generalized Linear Models (GLM) for CI and TF. Global TF experiments indicate that the predominant rainfall devices, number of sample trees, number of events, and monitoring length are 10-20 devices (81% fixed), 3-6 trees, 30-50 events, and 10-30 months. Predominant global values of TF, CI, and SF are 70-80% (median = 73%), 20%-30% (median = 23.9%), and <1.0% (median = 1.87%), respectively. Global models of CI and TF were responsive to T, LAI, and D (respectively, R2adj of 0.196** and 0.206**). Temperate forests mirrored the global model (R2adj of 0.274** and 0.31**, respectively). The Subtropical CI model was fitted based on P and DBH (R2adj = 0.245*), and the TF model was based on E, D, and LAI (R2adj = 0.532**); the Mediterranean CI model was based on T, Basal, and LAI (R2adj = 0.45*), while TF was based on P, Basal, and LAI (R2adj = 0.671**). The Tropical CI model was based on T and H (R2adj = 0.396*), and the TF model, LAI, and P (R2adj = 0.35*). This meta-analysis underscores the importance of comprehending the hydrological processes in forested areas as they are pivotal in mitigating climate change impacts.

The contribution of the AT1 receptor to erythropoiesis.

The renin-angiotensin system (RAS) comprises a broad set of functional peptides and receptors that play a role in cardiovascular homeostasis and contribute to cardiovascular pathologies. Angiotensin II (Ang II) is the most potent peptide hormone produced by the RAS due to its high abundance and its strong and pleiotropic impact on the cardiovascular system. Formation of Ang II takes place in the bloodstream and additionally in tissues in the so-called local RAS. Of the two Ang II receptors (AT1 and AT2) that Ang II binds to, AT1 is the most expressed throughout the mammalian body. AT1 expression is not restricted to cells of the cardiovascular system but in fact AT1 protein is found in nearly all organs, hence, Ang II takes part in several modulatory physiological processes one of which is erythropoiesis. In this review, we present multiple evidence supporting that Ang II modulates physiological and pathological erythropoiesis processes trough the AT1 receptor. Cumulative evidence indicates that Ang II by three distinct mechanisms influences erythropoiesis: 1) stimulation of renal erythropoietin synthesis; 2) direct action on bone marrow precursor cells; and 3) modulation of sympathetic nerve activity to the bone marrow. The text highlights clinical and preclinical evidence focusing on mechanistic studies using rodent models.

Quantifying the impact of gut microbiota on inflammation and hypertensive organ damage.

AIMS: Hypertension (HTN) can lead to heart and kidney damage. The gut microbiota has been linked to HTN, although it is difficult to estimate its significance due to the variety of other features known to influence HTN. In the present study, we used germ-free (GF) and colonized (COL) littermate mice to quantify the impact of microbial colonization on organ damage in HTN. METHODS AND RESULTS: 4-week-old male GF C57BL/6J littermates were randomized to remain GF or receive microbial colonization. HTN was induced by subcutaneous infusion with angiotensin (Ang) II (1.44 mg/kg/day) and 1% NaCl in the drinking water; sham-treated mice served as control. Renal damage was exacerbated in GF mice, whereas cardiac damage was more comparable between COL and GF, suggesting that the kidney is more sensitive to microbial influence. Multivariate analysis revealed a larger effect of HTN in GF mice. Serum metabolomics demonstrated that the colonization status influences circulating metabolites relevant to HTN. Importantly, GF mice were deficient in anti-inflammatory faecal short-chain fatty acids (SCFA). Flow cytometry showed that the microbiome has an impact on the induction of anti-hypertensive myeloid-derived suppressor cells and pro-inflammatory Th17 cells in HTN. In vitro inducibility of Th17 cells was significantly higher for cells isolated from GF than conventionally raised mice. CONCLUSION: The microbial colonization status of mice had potent effects on their phenotypic response to a hypertensive stimulus, and the kidney is a highly microbiota-susceptible target organ in HTN. The magnitude of the pathogenic response in GF mice underscores the role of the microbiome in mediating inflammation in HTN.

Maternal Hypermethioninemia Affects Neurons Number, Neurotrophins Levels, Energy Metabolism, and Na+,K+-ATPase Expression/Content in Brain of Rat Offspring.

In the current study, we verified the effects of maternal hypermethioninemia on the number of neurons, apoptosis, nerve growth factor, and brain-derived neurotrophic factor levels, energy metabolism parameters (succinate dehydrogenase, complex II, and cytochrome c oxidase), expression and immunocontent of Na+,K+-ATPase, edema formation, inflammatory markers (tumor necrosis factor-alpha and interleukin-6), and mitochondrial hydrogen peroxide levels in the encephalon from the offspring. Pregnant Wistar rats were divided into two groups: the first one received saline (control) and the second group received 2.68 µmol methionine/g body weight by subcutaneous injections twice a day during gestation (approximately 21 days). After parturition, pups were killed at the 21st day of life for removal of encephalon. Neuronal staining (anti-NeuN) revealed a reduction in number of neurons, which was associated to decreased nerve growth factor and brain-derived neurotrophic factor levels. Maternal hypermethioninemia also reduced succinate dehydrogenase and complex II activities and increased expression and immunocontent of Na+,K+-ATPase alpha subunits. These results indicate that maternal hypermethioninemia may be a predisposing factor for damage to the brain during the intrauterine life.

Methylphenidate Causes Behavioral Impairments and Neuron and Astrocyte Loss in the Hippocampus of Juvenile Rats.

Although the use, and misuse, of methylphenidate is increasing in childhood and adolescence, there is little information about the consequences of this psychostimulant chronic use on brain and behavior during development. The aim of the present study was to investigate hippocampus biochemical, histochemical, and behavioral effects of chronic methylphenidate treatment to juvenile rats. Wistar rats received intraperitoneal injections of methylphenidate (2.0 mg/kg) or an equivalent volume of 0.9 % saline solution (controls), once a day, from the 15th to the 45th day of age. Results showed that chronic methylphenidate administration caused loss of astrocytes and neurons in the hippocampus of juvenile rats. BDNF and pTrkB immunocontents and NGF levels were decreased, while TNF-α and IL-6 levels, Iba-1 and caspase 3 cleaved immunocontents (microglia marker and active apoptosis marker, respectively) were increased. ERK and PKCaMII signaling pathways, but not Akt and GSK-3ß, were decreased. SNAP-25 was decreased after methylphenidate treatment, while GAP-43 and synaptophysin were not altered. Both exploratory activity and object recognition memory were impaired by methylphenidate. These findings provide additional evidence that early-life exposure to methylphenidate can have complex effects, as well as provide new basis for understanding of the biochemical and behavioral consequences associated with chronic use of methylphenidate during central nervous system development.

Methylphenidate Decreases ATP Levels and Impairs Glutamate Uptake and Na+,K+-ATPase Activity in Juvenile Rat Hippocampus.

The study of the long-term neurological consequences of early exposure with methylphenidate (MPH) is very important since this psychostimulant has been widely misused by children and adolescents who do not meet full diagnostic criteria for ADHD. The aim of this study was to examine the effect of early chronic exposure with MPH on amino acids profile, glutamatergic and Na+,K+-ATPase homeostasis, as well as redox and energy status in the hippocampus of juvenile rats. Wistar male rats received intraperitoneal injections of MPH (2.0 mg/kg) or saline solution (controls), once a day, from the 15th to the 45th day of age. Results showed that MPH altered amino acid profile in the hippocampus, decreasing glutamine levels. Glutamate uptake and Na+,K+-ATPase activity were decreased after chronic MPH exposure in the hippocampus of rats. No changes were observed in the immunocontents of glutamate transporters (GLAST and GLT-1), and catalytic subunits of Na+,K+-ATPase (α1, α2, and α3), as well as redox status. Moreover, MPH provoked a decrease in ATP levels in the hippocampus of chronically exposed rats, while citrate synthase, succinate dehydrogenase, respiratory chain complexes activities (II, II-III, and IV), as well as mitochondrial mass and mitochondrial membrane potential were not altered. Taken together, our results suggest that chronic MPH exposure at early age impairs glutamate uptake and Na+,K+-ATPase activity probably by decreasing in ATP levels observed in rat hippocampus.

Protective effect of green tea extract against proline-induced oxidative damage in the rat kidney.

We investigated, in vivo (acute and chronic), the effects of proline on thiobarbituric acid-reactive substances (TBA-RS) and on the activities of antioxidant enzymes such as catalase (CAT), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) in renal tissues (cortex and medulla) of rats. For acute administration, 29-day-old rats received a single subcutaneous injection of proline (18.2µmol/g body weight) or an equivalent volume of 0.9% saline solution and were sacrificed 1h later. For chronic treatment, proline was injected subcutaneously in the rats twice a day from the 6th to the 28th day of age, and the animals were killed 12h after the last injection. The results showed that acute administration of proline enhanced CAT, SOD and GSH-Px activities, as well as, TBARS in the cortex and decreased CAT activity in the medulla, while chronic treatment increased the activities of SOD in the cortex and increased CAT, SOD and GSH-Px in the medulla of rats. Furthermore, the green tea extract treatment for one week or from the 6th to the 28th day of age prevented the alterations caused by acute and chronic, respectively, proline administration. Herein, we demonstrated that proline alters antioxidant defenses and induces lipid peroxidation in the kidney of rats and the green tea extract was capable to counteract the proline-induced alterations.

Antioxidant effect of simvastatin throught oxidative imbalance caused by lisdexamfetamine dimesylate.

The present study aims to directly investigate the behavioral and antioxidant effects of simvastatin in a model of bipolar mania induced by lisdexamfetamine dimesylate. Wistar rats were treated for 30 days with simvastatin. On the 24th day after the start of treatment, each rat was administered lisdexamfetamine dimesylate for 7 days. The results suggest that simvastatin combined with lisdexamfetamine dimesylate induced a significant increased locomotion and lisdexamfetamine dimesylate administration causes an oxidative imbalance determined by an increment in lipid peroxidation, protein oxidation and alterations in the activities of antioxidant enzymes in brain areas; moreover, in the presence of simvastatin, most of these effects were prevented. These findings contribute to a better understanding of the critical roles of lisdexamfetamine dimesylate in the treatment of neuropsychiatric disorders, associated with increased oxidative stress and changes in antioxidant enzymatic defense. In view of the central role played by lisdexamfetamine dimesylate, the established antioxidant effect of simvastatin therapy is of major interest.

Chronic Treatment with a Clinically Relevant Dose of Methylphenidate Increases Glutamate Levels in Cerebrospinal Fluid and Impairs Glutamatergic Homeostasis in Prefrontal Cortex of Juvenile Rats.

The understanding of the consequences of chronic treatment with methylphenidate is very important since this psychostimulant is extensively prescribed to preschool age children, and little is known about the mechanisms underlying the persistent changes in behavior and neuronal function related with the use of methylphenidate. In this study, we initially investigate the effect of early chronic treatment with methylphenidate on amino acids profile in cerebrospinal fluid and prefrontal cortex of juvenile rats, as well as on glutamatergic homeostasis, Na(+),K(+)-ATPase function, and balance redox in prefrontal cortex of rats. Wistar rats at early age received intraperitoneal injections of methylphenidate (2.0 mg/kg) or an equivalent volume of 0.9% saline solution (controls), once a day, from the 15th to the 45th day of age. Twenty-four hours after the last injection, the animals were decapitated and the cerebrospinal fluid and prefrontal cortex were obtained. Results showed that methylphenidate altered amino acid profile in cerebrospinal fluid, increasing the levels of glutamate. Glutamate uptake was decreased by methylphenidate administration, but GLAST and GLT-1 were not altered by this treatment. In addition, the astrocyte marker GFAP was not altered by MPH. The activity and immunocontent of catalytic subunits (α1, α2, and α3) of Na(+),K(+)-ATPase were decreased in prefrontal cortex of rats subjected to methylphenidate treatment, as well as changes in α1 and α2 gene expression of catalytic α subunits of Na(+),K(+)-ATPase were also observed. CAT activity was increased and SOD/CAT ratio and sulfhydryl content were decreased in rat prefrontal cortex. Taken together, our results suggest that chronic treatment with methylphenidate at early age induces excitotoxicity, at least in part, due to inhibition of glutamate uptake probably caused by disturbances in the Na(+),K(+)-ATPase function and/or in protein damage observed in the prefrontal cortex.

Antioxidant effect of simvastatin throught oxidative imbalancecaused by lisdexamfetamine dimesylate

The present study aims to directly investigate the behavioral and antioxidant effects of simvastatin in a model of bipolar mania induced by lisdexamfetamine dimesylate. Wistar rats were treated for 30 days with simvastatin. On the 24th day after the start of treatment, each rat was administered lisdexamfetamine dimesylate for 7 days. The results suggest that simvastatin combined with lisdexamfetamine dimesylate induced a significant increased locomotion and lisdexamfetamine dimesylate administration causes an oxidative imbalance determined by an increment in lipid peroxidation, protein oxidation and alterations in the activities of antioxidant enzymes in brain areas; moreover, in the presence of simvastatin, most of these effects were prevented. These findings contribute to a better understanding of the critical roles of lisdexamfetamine dimesylate in the treatment of neuropsychiatric disorders, associated with increased oxidative stress and changes in antioxidant enzymatic defense. In view of the central role played by lisdexamfetamine dimesylate, the established antioxidant effect of simvastatin therapy is of major interest...

Gestational hypermethioninaemia alters oxidative/nitrative status in skeletal muscle and biomarkers of muscular injury and inflammation in serum of rat offspring.

In this study we evaluated oxidative/nitrative stress parameters (reactive oxygen species production, lipid peroxidation, sulfhydryl content, superoxide dismutase, catalase and nitrite levels), as well as total protein content in the gastrocnemius skeletal muscle of the offspring of rats that had been subjected to gestational hypermethioninaemia. The occurrence of muscular injury and inflammation was also measured by creatine kinase activity, levels of creatinine, urea and C-reactive protein and the presence of cardiac troponin I in serum. Wistar female rats (70-90 days of age) received methionine (2.68 µmol/g body weight) or saline (control) twice a day by subcutaneous injections during the gestational period (21 days). After the rats gave birth, pups were killed at the twenty-first day of life for removal of muscle and serum. Methionine treatment increased reactive oxygen species production and lipid peroxidation and decreased sulfhydryl content, antioxidant enzymes activities and nitrite levels, as well as total protein content in skeletal muscle of the offspring. Creatine kinase activity was reduced and urea and C-reactive protein levels were increased in serum of pups. These results were accompanied by reduced muscle mass. Our findings showed that maternal gestational hypermethioninaemia induced changes in oxidative/nitrative status in gastrocnemius skeletal muscle of the offspring. This may represent a mechanism which can contribute to the myopathies and loss of muscular mass that is found in some hypermethioninaemic patients. In addition, we believe that these results may be relevant as gestational hypermethioninaemia could cause damage to the skeletal muscle during intrauterine life.

Hypoxanthine induces oxidative stress in kidney of rats: protective effect of vitamins E plus C and allopurinol.

In the present study, we investigated the in vitro effect of hypoxanthine on the activities of antioxidant enzymes such as catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase, as well as on thiobarbituric-acid-reactive substances (TBA-RS), in the renal cortex and medulla of rats. Results showed that hypoxanthine, at a concentration of 10.0 µM, enhanced the activities of CAT and SOD in the renal cortex of 15-, 30- and 60-day-old rats, enhanced SOD activity in the renal medulla of 60-day-old rats and enhanced TBA-RS levels in the renal medulla of 30-day-old rats, as compared with controls. Furthermore, we also verified the influence of allopurinol (an inhibitor of xanthine oxidase), as well as of the antioxidants, trolox and ascorbic acid on the effects elicited by hypoxanthine on the parameters tested. Allopurinol and/or administration of antioxidants prevented most alterations caused by hypoxanthine in the oxidative stress parameters evaluated. Data suggest that hypoxanthine alters antioxidant defences and induces lipid peroxidation in the kidney of rats; however, in the presence of allopurinol and antioxidants, some of these alterations in oxidative stress were prevented. Our findings lend support to a potential therapeutic strategy for this condition, which may include the use of appropriate antioxidants for ameliorating the damage caused by hypoxanthine.

[Evaluation of ectoparasites and hemoparasites in dogs kept in apartments and houses with yards in the city of Juiz de Fora, Minas Gerais, Brazil]. / Avaliação ectoparasitológica e hemoparasitológica em cães criados em apartamentos e casas com quintal na cidade de Juiz de Fora, MG.

Fleas and ticks transmit various pathogens while feeding on the blood of dogs. This study sought to verify the occurrence of ectoparasitism and hemoparasitism in dogs from two urban areas in the city of Juiz de Fora, Minas Gerais, Brazil. Between February and August 2003, 101 dogs were studied: 50 came from apartments in the downtown region and 51 from houses with grassy yards. The ectoparasites were collected and conserved in etanol 70%. The occurrence of hemoparasites was verified by examining blood smears from sample taken from the dogs'ears. The blood smears were stained with Giemsa and 100 fields per slide were examined, studying the erythrocytes to determine parasitism. From among the dogs living in apartments, we found (with respective prevalence and mean intensity): Ctenocephalides felis (12%), (3.3+/-2.0); Rhipicephalus sanguineus (2%); and ixodid nymphs (2%). In this environment in the dogs were not found hemoparasites. From the houses with grassy yards, we observed the following prevalence levels and mean intensities: C. felis (14%), (2.28+/-1.9); R. sanguineus (35%), (7.8+/-9.8); ixodid nymph (18%), (1.4+/-0.7); and ixodid larvae (4%), (12+/-14.4). The hemoparasites found were: Ehrlichia canis (16%) and Babesia canis (2%).