Photobiomodulation Therapy on Myocardial Infarction in Rats: Transcriptional and Posttranscriptional Implications to Cardiac Remodeling.

BACKGROUND AND OBJECTIVES: Induction of myocardial infarction (MI) in rats by occlusion of the left anterior descending coronary artery is an experimental model used in research to elucidate functional, structural, and molecular modifications associated with ischemic heart disease. Photobiomodulation therapy (PBMT) has become a therapeutic alternative by modulating various biological processes eliciting several effects, including anti-inflammatory and pro-proliferative actions. The main objective of this work was to evaluate the effect of PBMT in the modulation of transcriptional and post-transcriptional changes that occurred in myocardium signal transduction pathways after MI. STUDY DESIGN/MATERIALS AND METHODS: Continuous wave (CW) non-thermal laser parameters were: 660 nm wavelength, power 15 mW, with a total energy of 0.9 J, fluence of 1.15 J/cm2 , spot size of 0.785 cm2 , and time of 60 seconds. Using in silico analysis, we selected and then, quantified the expression of messenger RNA (mRNA) of 47 genes of 9 signaling pathways associated with MI (angiogenesis, cell survival, hypertrophy, oxidative stress, apoptosis, extracellular matrix, calcium kinetics, cell metabolism, and inflammation). Messenger RNA expression quantification was performed in myocardial samples by polymerase chain reaction real-time array using TaqMan customized plates. RESULTS: Our results evidenced that MI modified mRNA expression of several well-known biomarkers related to detrimental cardiac activity in almost all signaling pathways analyzed. However, PBMT reverted most of these transcriptional changes. More expressively, PBMT provoked a robust decrease in mRNA expression of molecules that participate in post-MI inflammation and ECM composition, such as IL-6, TNF receptor, TGFb1, and collagen I and III. Global microRNA (miRNA) expression analysis revealed that PBMT decreased miR-221, miR-34c, and miR-93 expressions post-MI, which are related to deleterious effects in cardiac remodeling. CONCLUSION: Thus, the identification of transcriptional and post-transcriptional changes induced by PBMT may be used to interfere in the molecular dynamics of cardiac remodeling post-MI.

Exercise benefits the cardiac, autonomic and inflammatory responses to organophosphate toxicity.

The organophosphate, diisopropyl fluorophosphate (DFP), may impair cardiovascular, autonomic and immune function while exercise training is thougt to be restorative. Experiments determined effects of wheel exercise in C57B1 male mice, testing cardiovascular and autonomic function and characterization of the immunological profile. Sedentary (S) and exercise (ET) groups were treated with corticosterone (CORT) followed by injection of DFP. This model was associated with systolic and diastolic dysfunction in the S group, measured using echocardiography (ECHO). Chronic exercise ameliorated the cardiac deficit. Autonomic balance, accessed by heart rate variability (HRV), showed increased sympathetic and decreased parasympathetic modulation in S group. Autonomic balance in ET mice was not affected by DFP. Our DFP model resulted in mild neuroinflammation seen by increased IL5, IL12 and MIP2 in brain and plasma IL6 and IL1a. DFP had a negative impact on cardiac/autonomic function and inflammatory markers, effects reduced by exercise. Data suggest a beneficial effect of exercise training on the cardiovascular and autonomic responses to DFP/CORT.

Corticosterone and pyridostigmine/DEET exposure attenuate peripheral cytokine expression: Supporting a dominant role for neuroinflammation in a mouse model of Gulf War Illness.

Gulf War Illness (GWI) is a chronic multi-symptom disorder experienced by as many as a third of the veterans of the 1991 Gulf War; the constellation of "sickness behavior" symptoms observed in ill veterans is suggestive of a neuroimmune involvement. Various chemical exposures and conditions in theater have been implicated in the etiology of the illness. Previously, we found that GW-related organophosphates (OPs), such as the sarin surrogate, DFP, and chlorpyrifos, cause neuroinflammation. The combination of these exposures with exogenous corticosterone (CORT), mimicking high physiological stress, exacerbates the observed neuroinflammation. The potential relationship between the effects of OPs and CORT on the brain versus inflammation in the periphery has not been explored. Here, using our established GWI mouse model, we investigated the effects of CORT and DFP exposure, with or without a chronic application of pyridostigmine bromide (PB) and N,N-diethyl-meta-toluamide (DEET), on cytokines in the liver and serum. While CORT primed DFP-induced neuroinflammation, this effect was largely absent in the periphery. Moreover, the changes found in the peripheral tissues do not correlate with the previously reported neuroinflammation. These results not only support GWI as a neuroimmune disorder, but also highlight the separation between central and peripheral effects of these exposures.

A Logic Model of Neuronal-Glial Interaction Suggests Altered Homeostatic Regulation in the Perpetuation of Neuroinflammation.

Aberrant inflammatory signaling between neuronal and glial cells can develop into a persistent sickness behavior-related disorders, negatively impacting learning, memory, and neurogenesis. While there is an abundance of literature describing these interactions, there still lacks a comprehensive mathematical model describing the complex feed-forward and feedback mechanisms of neural-glial interaction. Here we compile molecular and cellular signaling information from various studies and reviews in the literature to create a logically-consistent, theoretical model of neural-glial interaction in the brain to explore the role of neuron-glia homeostatic regulation in the perpetuation of neuroinflammation. Logic rules are applied to this connectivity diagram to predict the system's homeostatic behavior. We validate our model predicted homeostatic profiles against RNAseq gene expression profiles in a mouse model of stress primed neuroinflammation. A meta-analysis was used to calculate the significance of similarity between the inflammatory profiles of mice exposed to diisopropyl fluorophostphate (DFP) [with and without prior priming by the glucocorticoid stress hormone corticosterone (CORT)], with the equilibrium states predicted by the model, and to provide estimates of the degree of the neuroinflammatory response. Beyond normal homeostatic regulation, our model predicts an alternate self-perpetuating condition consistent with chronic neuroinflammation. RNAseq gene expression profiles from the cortex of mice exposed to DFP and CORT+DFP align with this predicted state of neuroinflammation, whereas the alignment to CORT alone was negligible. Simulations of putative treatment strategies post-exposure were shown to be theoretically capable of returning the system to a state of typically healthy regulation with broad-acting anti-inflammatory agents showing the highest probability of success. The results support a role for the brain's own homeostatic drive in perpetuating the chronic neuroinflammation associated with exposure to the organophosphate DFP, with and without CORT priming. The deviation of illness profiles from exact model predictions suggests the presence of additional factors or of lasting changes to the brain's regulatory circuitry specific to each exposure.

Protective Role of UCP2 in Oxidative Stress and Apoptosis during the Silent Phase of an Experimental Model of Epilepsy Induced by Pilocarpine.

Neuroprotection is a desirable process in many neurological disorders, yet complex mechanisms involved in this field are not completely understood. The pilocarpine epilepsy model causes potent, seizure-induced excitotoxicity cell death and mitochondria impairment. The present study is aimed at investigating the role of UCP2, a ROS negative regulator, in the neuroprotection after cholinergic insult. Our data demonstrated that UCP2 expression was augmented in the rat hippocampus 3 days after status epilepticus (SE), reaching a peak on the fifth day, then returning to basal levels. Concomitantly, phospho-AKT expression levels were higher in the hippocampus during the early silent phase (5 days after SE). Additionally, it was demonstrated that the blockade of UCP2 by antisense oligonucleotides (ASO) in SE rats successfully diminished both UCP2 mRNA and protein contents. SE ASO rats presented increased mitochondrial proapoptotic factor expression, caspase-3 activity, inflammatory cytokine expression, and ROS formation. Moreover, ASO treatment diminished p-AKT expression and antioxidant enzyme activities after pilocarpine insult. In conclusion, the present results highlight the neuroprotective actions of UCP2, acting in the inhibition of apoptotic factors and oxidative stress, to increase neuron survival after SE onset.

Measurement of Mouse Heart Rate Variability using Echocardiographic System.

AIM: We employed an echocardiographic (ECHO) system as the backbone for the collection of electrocardiogram (ECG) and heart rate variability (HRV) data. The system was tested using an exercise model in which C57 male mice were exposed to sham or forced wheel running. METHODS: Peak/peak (RR) interval was recorded over a 3 min period using the ECG platform of the ECHO system. Isoflurane-anesthetized male mice were divided into two groups (n = 8/group): sedentary (S) and forced wheel trained (T). HRV was analyzed in time and frequency domains (Fast Fourier Transform). Exercise training (T) was performed on a motorized wheel at low intensity 1 h/day, 5 days/week, 8 weeks duration. Cardiac morphometry and function were analyzed using ECHO while ECG was the basis to measure HRV. The sampling rate was 8000 Hz. Results show that the trained mice presented a reduction in heart rate as compared to the sedentary group. This was associated with lower cardiac sympathetic and higher parasympathetic modulation leading to an improved sympathetic/parasympathetic ratio (low-frequency band/high-frequency band). The trained group showed a reduction in isovolumetric relaxation time, reduced myocardial performance index, increased relative wall thickness, and left ventricle mass when compared to the sedentary group. CONCLUSION: Results document the utility of combining the ECHO and the ECG platform, allowing for the dual measurement of autonomic and cardiac function in mice.

Increasing Resilience to Traumatic Stress: Understanding the Protective Role of Well-Being.

The brain maintains homeostasis in part through a network of feedback and feed-forward mechanisms, where neurochemicals and immune markers act as mediators. Using a previously constructed model of biobehavioral feedback, we found that in addition to healthy equilibrium another stable regulatory program supported chronic depression and anxiety. Exploring mechanisms that might underlie the contributions of subjective well-being to improved therapeutic outcomes in depression, we iteratively screened 288 candidate feedback patterns linking well-being to molecular signaling networks for those that maintained the original homeostatic regimes. Simulating stressful trigger events on each candidate network while maintaining high levels of subjective well-being isolated a specific feedback network where well-being was promoted by dopamine and acetylcholine, and itself promoted norepinephrine while inhibiting cortisol expression. This biobehavioral feedback mechanism was especially effective in reproducing well-being's clinically documented ability to promote resilience and protect against onset of depression and anxiety.

Breaking Away: The Role of Homeostatic Drive in Perpetuating Depression.

We propose that the complexity of regulatory interactions modulating brain neurochemistry and behavior is such that multiple stable responses may be supported, and that some of these alternate regulatory programs may play a role in perpetuating persistent psychological dysfunction. To explore this, we constructed a model network representing major neurotransmission and behavioral mechanisms reported in literature as discrete logic circuits. Connectivity and information flow through this biobehavioral circuitry supported two distinct and stable regulatory programs. One such program perpetuated a depressive state with a characteristic neurochemical signature including low serotonin. Further analysis suggested that small irregularities in glutamate levels may render this pathology more directly accessible. Computer simulations mimicking selective serotonin reuptake inhibitor (SSRI) therapy in the presence of everyday stressors predicted recidivism rates similar to those reported clinically and highlighted the potentially significant benefit of concurrent behavioral stress management therapy.

Comparative mRNA and MicroRNA Profiling during Acute Myocardial Infarction Induced by Coronary Occlusion and Ablation Radio-Frequency Currents.

The ligation of the left anterior descending coronary artery is the most commonly used experimental model to induce myocardial infarction (MI) in rodents. A high mortality in the acute phase and the heterogeneity of the size of the MI obtained are drawbacks recognized in this model. In an attempt to solve the problem, our group recently developed a new MI experimental model which is based on application of myocardial ablation radio-frequency currents (AB-RF) that yielded MI with homogeneous sizes and significantly reduce acute mortality. In addition, cardiac structural, and functional changes aroused by AB-RF were similar to those seen in animals with MI induced by coronary artery ligation. Herein, we compared mRNA expression of genes that govern post-MI milieu in occlusion and ablation models. We analyzed 48 mRNAs expressions of nine different signal transduction pathways (cell survival and metabolism signs, matrix extracellular, cell cycle, oxidative stress, apoptosis, calcium signaling, hypertrophy markers, angiogenesis, and inflammation) in rat left ventricle 1 week after MI generated by both coronary occlusion and AB-RF. Furthermore, high-throughput miRNA analysis was also assessed in both MI procedures. Interestingly, mRNA expression levels and miRNA expressions showed strong similarities between both models after MI, with few specificities in each model, activating similar signal transduction pathways. To our knowledge, this is the first comparison of genomic alterations of mRNA and miRNA contents after two different MI procedures and identifies key signaling regulators modulating the pathophysiology of these two models that might culminate in heart failure. Furthermore, these analyses may contribute with the current knowledge concerning transcriptional and post-transcriptional changes of AB-RF protocol, arising as an alternative and effective MI method that reproduces most changes seem in coronary occlusion.

Exercise Training Prevents Cardiovascular Derangements Induced by Fructose Overload in Developing Rats.

The risks of chronic diseases associated with the increasing consumption of fructose-laden foods are amplified by the lack of regular physical activity and have become a serious public health issue worldwide. Moreover, childhood eating habits are strongly related to metabolic syndrome in adults. Thus, we aimed to investigate the preventive role of exercise training undertaken concurrently with a high fructose diet on cardiac function, hemodynamics, cardiovascular autonomic modulation and oxidative stress in male rats after weaning. Male Wistar rats were divided into 4 groups (n = 8/group): Sedentary control (SC), Trained control (TC), Sedentary Fructose (SF) and Trained Fructose (TF). Training was performed on a treadmill (8 weeks, 40-60% of maximum exercise test). Evaluations of cardiac function, hemodynamics, cardiovascular autonomic modulation and oxidative stress in plasma and in left ventricle (LV) were performed. Chronic fructose overload induced glucose intolerance and an increase in white adipose tissue (WAT) weight, in myocardial performance index (MPI) (SF:0.42±0.04 vs. SC:0.24±0.05) and in arterial pressure (SF:122±3 vs. SC:113±1 mmHg) associated with increased cardiac and vascular sympathetic modulation. Fructose also induced unfavorable changes in oxidative stress profile (plasmatic protein oxidation- SF:3.30±0.09 vs. SC:1.45±0.08 nmol/mg prot; and LV total antioxidant capacity (TRAP)- SF: 2.5±0.5 vs. SC:12.7±1.7 uM trolox). The TF group showed reduced WAT, glucose intolerance, MPI (0.35±0.04), arterial pressure (118±2mmHg), sympathetic modulation, plasmatic protein oxidation and increased TRAP when compared to SF group. Therefore, our findings indicate that cardiometabolic dysfunctions induced by fructose overload early in life may be prevented by moderate aerobic exercise training.

Impact of aging on cardiac function in a female rat model of menopause: role of autonomic control, inflammation, and oxidative stress.

OBJECTIVE: The aim of this study was to evaluate the effects of aging on metabolic, cardiovascular, autonomic, inflammatory, and oxidative stress parameters after ovarian hormone deprivation (OVX). METHODS: Female Wistar rats (3 or 22 months old) were divided into: young controls, young ovariectomized, old controls, and old ovariectomized (bilateral ovaries removal). After a 9-week follow-up, physical capacity, metabolic parameters, and morphometric and cardiac functions were assessed. Subsequently, arterial pressure was recorded and cardiac autonomic control was evaluated. Oxidative stress was measured on the cardiac tissue, while inflammatory profile was assessed in the plasma. RESULTS: Aging or OVX caused an increase in body and fat weight and triglyceride concentration and a decrease in both insulin sensitivity and aerobic exercise capacity. Left ventricular diastolic dysfunction and increased cardiac overload (myocardial performance index) were reported in old groups when compared with young groups. Aging and OVX led to an increased sympathetic tonus, and vagal tonus was lower only for the old groups. Tumor necrosis factor-α and interleukin-6 were increased in old groups when compared with young groups. Glutathione redox balance (GSH/GSSG) was reduced in young ovariectomized, old controls, and old ovariectomized groups when compared with young controls, indicating an increased oxidative stress. A negative correlation was found between GSH/GSSG and tumor necrosis factor-α (r=-0.6, P<0.003). Correlations were found between interleukin-6 with adipose tissue (r=0.5, P<0.009) and vagal tonus (r=-0.7, P<0.0002); and among myocardial performance index with interleukin-6 (r=0.65, P<0.0002), sympathetic tonus (r=0.55, P<0.006), and physical capacity (r=-0.55, P<0.003). The findings in this trial showed that ovariectomy aggravated the impairment of cardiac and functional effects of aging in female rats, probably associated with exacerbated autonomic dysfunction, inflammation, and oxidative stress.

Positive effect of combined exercise training in a model of metabolic syndrome and menopause: autonomic, inflammatory, and oxidative stress evaluations.

It is now well established that after menopause cardiometabolic disorders become more common. Recently, resistance exercise has been recommended as a complement to aerobic (combined training, CT) for the treatment of cardiometabolic diseases. The aim of this study was to evaluate the effects of CT in hypertensive ovariectomized rats undergoing fructose overload in blood pressure variability (BPV), inflammation, and oxidative stress parameters. Female rats were divided into the following groups (n = 8/group): sedentary normotensive Wistar rats (C), and sedentary (FHO) or trained (FHOT) ovariectomized spontaneously hypertensive rats undergoing and fructose overload. CT was performed on a treadmill and ladder adapted to rats in alternate days (8 wk; 40-60% maximal capacity). Arterial pressure (AP) was directly measured. Oxidative stress and inflammation were measured on cardiac and renal tissues. The association of risk factors (hypertension + ovariectomy + fructose) promoted increase in insulin resistance, mean AP (FHO: 174 ± 4 vs. C: 108 ± 1 mmHg), heart rate (FHO: 403 ± 12 vs. C: 352 ± 11 beats/min), BPV, cardiac inflammation (tumor necrosis factor-α-FHO: 65.8 ± 9.9 vs. C: 23.3 ± 4.3 pg/mg protein), and oxidative stress cardiac and renal tissues. However, CT was able to reduce mean AP (FHOT: 158 ± 4 mmHg), heart rate (FHOT: 303 ± 5 beats/min), insulin resistance, and sympathetic modulation. Moreover, the trained rats presented increased nitric oxide bioavailability, reduced tumor necrosis factor-α (FHOT: 33.1 ± 4.9 pg/mg protein), increased IL-10 in cardiac tissue and reduced lipoperoxidation, and increased antioxidant defenses in cardiac and renal tissues. In conclusion, the association of risk factors promoted an additional impairment in metabolic, cardiovascular, autonomic, inflammatory, and oxidative stress parameters and combined exercise training was able to attenuate these dysfunctions.

Loss of prolyl carboxypeptidase in two-kidney, one-clip goldblatt hypertensive mice.

It is well documented that angiotensin (Ang) II contributes to kidney disease progression. The protease prolyl carboxypeptidase (PRCP) is highly expressed in the kidney and may be renoprotective by degrading Ang II to Ang-(1-7). The aim of the study was to investigate whether renal PRCP protein expression and activity are altered in two-kidney, one-clip (2K1C) Goldblatt hypertensive mice. Left renal artery was constricted by using 0.12 mm silver clips. Blood pressure was measured using telemetry over the eleven weeks of study period and revealed an immediate increase in 2K1C animals during the first week of clip placement which was followed by a gradual decrease to baseline blood pressure. Similarly, urinary albumin excretion was significantly increased one week after 2K1C and returned to baseline levels during the following weeks. At 2 weeks and at the end of the study, renal pathologies were exacerbated in the 2K1C model as revealed by a significant increase in mesangial expansion and renal fibrosis. Renal PRCP expression and activity were significantly reduced in clipped kidneys. Immunofluorescence revealed the loss of renal tubular PRCP but not glomerular PRCP. In contrast, expression of prolyl endopeptidase, another enzyme capable of converting Ang II into Ang-(1-7), was not affected, while angiotensin converting enzyme was elevated in unclipped kidneys and renin was increased in clipped kidneys. Results suggest that PRCP is suppressed in 2K1C and that this downregulation may attenuate renoprotective effects via impaired Ang II degradation by PRCP.

Novel role of aminopeptidase-A in angiotensin-(1-7) metabolism post myocardial infarction.

Aminopeptidase-A (APA) is a less well-studied enzyme of the renin-angiotensin system. We propose that it is involved in cardiac angiotensin (ANG) metabolism and its pathologies. ANG-(1-7) can ameliorate remodeling after myocardial injury. The aims of this study are to (1) develop mass spectrometric (MS) approaches for the assessment of ANG processing by APA within the myocardium; and (2) investigate the role of APA in cardiac ANG-(1-7) metabolism after myocardial infarction (MI) using sensitive MS techniques. MI was induced in C57Bl/6 male mice by ligating the left anterior descending (LAD) artery. Frozen mouse heart sections (in situ assay) or myocardial homogenates (in vitro assay) were incubated with the endogenous APA substrate, ANG II. Results showed concentration- and time-dependent cardiac formation of ANG III from ANG II, which was inhibited by the specific APA inhibitor, 4-amino-4-phosphonobutyric acid. Myocardial APA activity was significantly increased 24 h after LAD ligation (0.82 ± 0.02 vs. 0.32 ± 0.02 ρmol·min(-1)·µg(-1), MI vs. sham, P < 0.01). Both MS enzyme assays identified the presence of a new peptide, ANG-(2-7), m/z 784, which accumulated in the MI (146.45 ± 6.4 vs. 72.96 ± 7.0%, MI vs. sham, P < 0.05). Use of recombinant APA enzyme revealed that APA is responsible for ANG-(2-7) formation from ANG-(1-7). APA exhibited similar substrate affinity for ANG-(1-7) compared with ANG II {Km (ANG II) = 14.67 ± 1.6 vs. Km [ANG-(1-7)] = 6.07 ± 1.12 µmol/l, P < 0.05}. Results demonstrate a novel role of APA in ANG-(1-7) metabolism and suggest that the upregulation of APA, which occurs after MI, may deprive the heart of cardioprotective ANG-(1-7). Thus APA may serve as a potentially novel therapeutic target for management of tissue remodeling after MI.

Sexual behavior and dendritic spine density of posterodorsal medial amygdala neurons in oxytocin knockout female mice.

Central oxytocin (OT) and arginine-vasopressin (AVP) have been shown to play an important role in sexual behavior and neuroendocrine secretion in rodents. The results of exogenous OT administration on sexual behaviors in male and female mice are controversial. This study aimed to analyze the role of OT in sexual behavior, the number of oocytes and the density of dendritic spines in the posterodorsal medial amygdala (MePD) of female mice with selective deletion of the OT gene (OTKO). Female C57BL/6 mice were genotyped and divided into control (WT) and OTKO groups (n=11 each). All experiments were performed in the proestrus phase. Compared to WT data, our results showed that the OTKO group had a significant increase in the latency for the display of lordosis behavior (490.8 ± 113.8 and 841.9 ± 53.9, respectively) and a decrease in both the frequency (6.3 ± 2.4 and 0.5 ± 0.4) and duration (49.3 ± 19.9 and 7.2 ± 7.1) of lordosis and a reduction in the number of oocytes (12.2 ± 0.8 and 9.9 ± 0.6). However, the OTKO group showed a higher density of proximal dendritic spines in the MePD compared to the WT group (2.4 ± 0.1 and 1.9 ± 0.1 spines/dendritic µm, respectively). No significant difference was observed in the plasma levels of AVP between the groups (OTKO: 617.1 ± 96.0 and WT: 583.3 ± 112.0 pg/mL). Our data suggest that OT plays a crucial role in the sexual behavior display, number of released oocytes and density of dendritic spines in the MePD of female mice. The AVP plasma concentration was not affected in the OTKO animals.

Identification of prolyl carboxypeptidase as an alternative enzyme for processing of renal angiotensin II using mass spectrometry.

Angiotensin-converting enzyme 2 (ACE2) catalyzes conversion of ANG II to ANG-(1-7). The present study uses newly established proteomic approaches and genetic mouse models to examine the contribution of alternative renal peptidases to ACE2-independent formation of ANG-(1-7). In situ and in vitro mass spectrometric characterization showed that substrate concentration and pH control renal ANG II processing. At pH ≥6, ANG-(1-7) formation was significantly reduced in ACE2 knockout (KO) mice. However, at pH <6, formation of ANG-(1-7) in ACE2 KO mice was similar to that in wild-type (WT) mice, suggesting alternative peptidases for renal ANG II processing. Furthermore, the dual prolyl carboxypeptidase (PCP)-prolyl endopeptidase (PEP) inhibitor Z-prolyl-prolinal reduced ANG-(1-7) formation in ACE2 KO mice, while the ACE2 inhibitor MLN-4760 had no effect. Unlike the ACE2 KO mice, ANG-(1-7) formation from ANG II in PEP KO mice was not different from that in WT mice at any tested pH. However, at pH 5, this reaction was significantly reduced in kidneys and urine of PCP-depleted mice. In conclusion, results suggest that ACE2 metabolizes ANG II in the kidney at neutral and basic pH, while PCP catalyzes the same reaction at acidic pH. This is the first report demonstrating that renal ANG-(1-7) formation from ANG II is independent of ACE2. Elucidation of ACE2-independent ANG-(1-7) production pathways may have clinically important implications in patients with metabolic and renal disease.

Oxytocin modulates social interaction but is not essential for sexual behavior in male mice.

Recently, several studies have shown different conclusions regarding the effect of oxytocin (OT) on the social behaviors of male mice. Most of these studies used exogenous OT, but currently, investigations of the neural bases of social behavior are increasingly employing gene inactivation. This study aimed to analyze the role of OT in the modulation of social behaviors (i.e., sexual and social interaction behaviors) in male mice with selective deletions of the OT gene (OTKO) and the influence of this deletion in basal vasopressin (AVP) plasma concentrations. Our results showed that in the social interaction test, OTKO mice exhibited lower levels of social behaviors and higher levels of non-social behaviors compared to the wild type (WT) group. Additionally, the OTKO group showed a decrease in the number of agonistic behaviors delivered, and consequently, their dominance score was lower than that of the WT group. In the ethological analysis, the OTKO group had a lower aggressive performance and increased social investigation than the WT group. No significant differences were observed in the sexual behavior between groups. Finally, we found lower AVP plasma concentrations in the OTKO compared with the WT group. In conclusion, our data suggest that OT modulates social investigation behavior and the aggressiveness of male mice. The decrease in AVP concentrations in the OTKO group allows us to infer that AVP is physiologically relevant to these behavioral modulations. However, sexual behaviors do not seem to be affected by the lack of OT or by a decrease in the AVP concentration.

Angiotensin 1-7 and Mas decrease thrombosis in Bdkrb2-/- mice by increasing NO and prostacyclin to reduce platelet spreading and glycoprotein VI activation.

Bradykinin B2 receptor-deleted mice (Bdkrb2(-/-)) have delayed carotid artery thrombosis times and prolonged tail bleeding time resulting from elevated angiotensin II (AngII) and angiotensin receptor 2 (AT2R) producing increased plasma nitric oxide (NO) and prostacyclin. Bdkrb2(-/-) also have elevated plasma angiotensin-(1-7) and messenger RNA and protein for its receptor Mas. Blockade of Mas with its antagonist A-779 in Bdkrb2(-/-) shortens thrombosis times (58 ± 4 minutes to 38 ± 4 minutes) and bleeding times (170 ± 13 seconds to 88 ± 8 seconds) and lowers plasma nitrate (22 ± 4 µM to 15 ± 5 µM), and 6-keto-PGF1α (259 ± 103 pg/mL to 132 ± 58 pg/mL). Bdkrb2(-/-) platelets express increased NO, guanosine 3',5'-cyclic monophosphate, and cyclic adenosine monophosphate with reduced spreading on collagen, collagen peptide GFOGER, or fibrinogen. In vivo A-779 or combined L-NAME and nimesulide treatment corrects it. Bdkrb2(-/-) platelets have reduced collagen-related peptide-induced integrin α2bß3 activation and P-selectin expression that are partially corrected by in vivo A-779, nimesulide, or L-NAME. Bone marrow transplantations show that the platelet phenotype and thrombosis time depends on the host rather than donor bone marrow progenitors. Transplantation of wild-type bone marrow into Bdkrb2(-/-) hosts produces platelets with a spreading defect and delayed thrombosis times. In Bdkrb2(-/-), combined AT2R and Mas overexpression produce elevated plasma prostacyclin and NO leading to acquired platelet function defects and thrombosis delay.

Low-dose sarin exposure produces long term changes in brain neurochemistry of mice.

Sarin is a toxic organophosphorus (OP) nerve agent that has been reported to cause long-term alterations in behavioral and neuropsychological processes. The present study was designed to investigate the effect of low dose sarin exposure on the monoamine neurotransmitter systems in various brain regions of mice. The rationale was to expand our knowledge about the noncholinergic neurochemical alterations associated with low dose exposure to this cholinesterase inhibitor. We analyzed the levels of monoamines and their metabolites in different brain areas after exposure of male C57BL/6 mice to a subclinical dose of sarin (0.4 LD50). Mice did not show any signs of cholinergic toxicity or pathological changes in brain tissue. At 1, 4 and 8 weeks post-sarin exposure brains were collected for neurochemical analysis. A significant decrease in the dopamine (DA) turnover, as measured by the metabolite to parent ratio, was observed in the frontal cerebral cortex (FC) at all time points tested. DA turnover was significantly increased in the amygdala at 4 weeks but not at 1 or 8 weeks after exposure. The caudate nucleus displayed a decrease in DA turnover at 1 week but no significant change was observed at 4 and 8 weeks suggesting a reversible effect. In addition to this, serotonin (5-HT) levels were transiently altered at various time points in all the brain regions studied (increase in FC, caudate nucleus and decrease in amygdala). Since there were no signs of cholinergic toxicity or cell death after sarin exposure, different non-cholinergic mechanisms may be involved in regulating these effects. Our results demonstrate that non-symptomatic dose of OP nerve agent sarin has potent long-term, region-specific effects on the monoaminergic neurotransmitter systems. Data also suggests differential effects of sarin on the various DA projections. These neurochemical alterations could be associated with long term behavioral and neuropsychological changes associated with low dose OP exposure.

Enhanced angiotensin II-induced cardiac and aortic remodeling in ACE2 knockout mice.

Angiotensin-converting enzyme 2 (ACE2) is present in the heart and thought to exert protective functions. We conducted studies in ACE2 deficient mice to determine whether enzyme loss would exacerbate the cardiac and vascular pathological responses to chronic subcutaneous (sc) angiotensin II (Ang II) infusion. Eight-week-old male ACE2 knockout (KO) and wild type (WT) mice were infused with Ang II (1000 ng/kg per min, 4 weeks) using mini-osmotic pumps. Blood pressure (radiotelemetry), cardiac function (echocardiography, echo), cardiac/aortic structure (histology, collagen, and oxidative stress), and vascular inflammation were examined. Before Ang II infusion, ACE2 KO mice showed unaltered cardiac function and blood pressure. After 4 weeks of Ang II infusion, the mean arterial pressure (MAP) increased from 96 ± 2 to 136 ± 17 mm Hg (∼40%) in WT and from 104 ± 5 to 141 ± 13 mm Hg (∼ 35%) in ACE2 KO. While there were no differences in MAP between groups, the ACE2 KO responded differently to the hypertensive stimulus. Echo analysis revealed severe myocardial dysfunction in Ang II-infused ACE2 KO (Ang ACE2 KO). Ejection fraction was lower (39% versus 50%) as was fractional shortening (27% versus 38%) in ACE2 KO versus WT, respectively. Cardiac dysfunction was associated with hypertrophic cardiomyopathy shown by increased left-ventricular wall thickness, average cardiomyocyte cross-sectional area, and heart weight/body weight ratio. Collagen staining in the myocardium and aorta revealed increased collagen in Ang ACE2 KO, suggestive of remodeling. Results also showed enhanced oxidative stress in the myocardium and aorta of Ang ACE2 KO. There was a 3-fold elevation in macrophage inflammatory protein 1α (MIP 1α) in the aorta of ACE2 KO. Studies in the ACE2 KO model reveal the importance of ACE2 in the maladaptive cardiac and aortic responses to Ang II stimulation, seen as enhanced remodeling using physiological, structural, and biochemical markers. Results document a cardio- and vascular-protective role of ACE2 under pathological conditions.