The hydrogen sulfide donor 4-carboxyphenyl-isothiocyanate decreases blood pressure and promotes cardioprotective effect through reduction of oxidative stress and nuclear factor kappa B/matrix metalloproteinase (MMP)-2 axis in hypertension.

AIMS: Our aim was to evaluate whether the hydrogen sulfide (H2S) donor, 4-carboxyphenyl-isothiocyanate (4-CPI), exerts cardioprotective effect in the two kidney- one clip (2K-1C) rats through oxidative stress and MMP-2 activity attenuation and compare it with the classical H2S donor, Sodium Hydrosulfide (NaHS). MATERIALS AND METHODS: Renovascular hypertension (two kidneys-one clip; 2K-1C) was surgically induced in male Wistar rats. After two weeks, normotensive (2K) and hypertensive rats were intraperitoneally treated with vehicle (0.6 % dimethyl sulfoxide), NaHS (0.24 mg/Kg/day) or with 4-CPI (0.24 mg/Kg/day), for more 4 weeks. Systolic blood pressure (SBP) was evaluated weekly by tail-cuff plethysmography. Heart function was assessed by using the Millar catheter. Cardiac hypertrophy and fibrosis were evaluated by hematoxylin and eosin, and Picrosirius Red staining, respectively. The H2S was analyzed using WSP-1 fluorimetry and the cardiac oxidative stress was measured by lucigenin chemiluminescence and Amplex Red. MMP-2 activity was measured by in-gel gelatin or in situ zymography assays. Nox1, gp91phox, MMP-2 and the phospho-p65 subunit (Serine 279) nuclear factor kappa B (NF-κB) levels were evaluated by Western blotting. KEY FINDINGS: 4-CPI reduced blood pressure in hypertensive rats, decreased cardiac remodeling and promoted cardioprotection through the enhancement of cardiac H2S levels. An attenuation of oxidative stress, with inactivation of the p65-NF-κB/MMP-2 axis was similarly observed after NaHS or 4-CPI treatment in 2K-1C hypertension. SIGNIFICANCE: H2S is a mediator that promotes cardioprotective effects and decreases blood pressure, and 4-CPI seems to be a good candidate to reverse the maladaptive remodeling and cardiac dysfunction in renovascular hypertension.

Investigating autonomic nervous system dysfunction among patients with post-COVID condition and prolonged cardiovascular symptoms.

Heart Rate Variability (HRV) and arterial pressure (AP) variability and their responses to head-up tilt test (HUTT) were investigated in Post-COVID-19 syndrome (PCS) patients reporting tachycardia and/or postural hypotension. Besides tachycardia, PCS patients also showed attenuation of the following HRV parameters: RMSSD [square root of the mean of the sum of the squares of differences between adjacent normal-to-normal (NN) intervals] from statistical measures; the power of RR (beat-to-beat interval) spectra at HF (high frequency) from the linear method spectral analysis; occurrence of 2UV (two unlike variation) pattern of RR from the nonlinear method symbolic analysis; and the new family of statistics named sample entropy, when compared to control subjects. Basal AP and LF (low frequency) power of systolic AP were similar between PCS patients and control subjects, while 0 V (zero variation) patterns of AP from the nonlinear method symbolic analysis were exacerbated in PCS patients. Despite tachycardia and a decrease in RMSSD, no parameter of HRV changed during HUTT in PCS patients compared to control subjects. PCS patients reassessed after 6 months showed higher HF power of RR spectra and a higher percentage of 2UV pattern of RR. Moreover, the reassessed PCS patients showed a lower occurrence of 0 V patterns of AP, while the HUTT elicited HR (heart rate) and AP responses identical to control subjects. The HRV and AP variability suggest an autonomic dysfunction with sympathetic predominance in PCS patients. In contrast, the lack of responses of HRV and AP variability indices during HUTT indicates a marked impairment of autonomic control. Of note, the reassessment of PCS patients showed that the noxious effect of COVID-19 on autonomic control tended to fade over time.

Heart rate fragmentation is impaired in type 2 diabetes mellitus patients.

AIMS: To evaluate the heart rate fragmentation (HRF) of type 2 diabetes mellitus (T2DM) patients and its relationship with heart rate variability (HRV) indices. METHODS: One hundred sixty-four men, aged 47-57 years were retrospectively analyzed from a database. Participants were T2DM (n = 82) and apparently healthy (n = 82). R-R interval time series recorded by electrocardiogram were collected at the supine position for 10 to 15 min. From HRF, the percentage of inflection points (PIP), percentage of words with zero, one, two, or three inflections points (W0, W1, W2, W3), and percentage with only type hard, soft, or mixed inflections points type (WH, WS, WM) were quantified. RESULTS: T2DM presented higher PIP, WS, WM and W3, while WH and W1 was lower compared with healthy (p < 0.05). Moreover, a positive moderate correlation was found between WH and root mean square of the successive R-R differences (RMSSD) and high frequency (HF) indices. In contrast, a negative moderate correlation was found between WS and WM with RMSSD and HF indices. CONCLUSIONS: T2DM have increased fragmentation patterns, and words grouped by inflection type are more closely related to HRV. The HRF approach might be useful to assess heart rate dynamic abnormalities in males with type 2 diabetes.

Heart rate variability as a biomarker in patients with Chronic Chagas Cardiomyopathy with or without concomitant digestive involvement and its relationship with the Rassi score.

BACKGROUND: Dysautonomia plays an ancillary role in the pathogenesis of Chronic Chagas Cardiomyopathy (CCC), but is the key factor causing digestive organic involvement. We investigated the ability of heart rate variability (HRV) for death risk stratification in CCC and compared alterations of HRV in patients with isolated CCC and in those with the mixed form (CCC + digestive involvement). Thirty-one patients with CCC were classified into three risk groups (low, intermediate and high) according to their Rassi score. A single-lead ECG was recorded for a period of 10-20 min, RR series were generated and 31 HRV indices were calculated. The HRV was compared among the three risk groups and regarding the associated digestive involvement. Four machine learning models were created to predict the risk class of patients. RESULTS: Phase entropy is decreased and the percentage of inflection points is increased in patients from the high-, compared to the low-risk group. Fourteen patients had the mixed form, showing decreased triangular interpolation of the RR histogram and absolute power at the low-frequency band. The best predictive risk model was obtained by the support vector machine algorithm (overall F1-score of 0.61). CONCLUSIONS: The mixed form of Chagas' disease showed a decrease in the slow HRV components. The worst prognosis in CCC is associated with increased heart rate fragmentation. The combination of HRV indices enhanced the accuracy of risk stratification. In patients with the mixed form of Chagas disease, a higher degree of sympathetic autonomic denervation may be associated with parasympathetic impairment.

Aryl hydrocarbon receptor (AhR) activation contributes to high-fat diet-induced vascular dysfunction.

BACKGROUND AND PURPOSE: Metabolic and vascular dysfunction are common features of obesity. Aryl hydrocarbon receptor (AhR) regulates lipid metabolism and vascular homeostasis, but whether vascular AhR are activated in obesity or have a protective and/or harmful effects on vascular function in obesity are unknown. Our study addresses whether AhR activation contributes to obesity-associated vascular dysfunction and the mechanisms involved in these AhR effects. EXPERIMENTAL APPROACH: Male AhR KO (Ahr-/- ) and WT mice were fed either control or a HF (high-fat) diet for 10 weeks. Metabolic and inflammatory parameters were measured in serum and adipose tissue. Vascular reactivity (isometric force) was evaluated using a myography. Endothelial NOS (eNOS) and AhR protein expression was determined by western blot, Cyp1A1 and Nos3 gene expression by RT-PCR and.NO production was quantified by DAF fluorescence. KEY RESULTS: HF diet increased total serum HDL and LDL, as well as vascular AhR protein expression and proinflammatory cytokines in the adipose tissue. HF diet decreased endothelium-dependent vasodilation. AhR deletion protected mice from HF diet-induced dyslipidaemia, weight gain and inflammatory processes. HF diet-induced endothelial dysfunction was attenuated in Ahr-/- mice. Vessels from Ahr-/- mice exhibited a greater NO reserve. In cultured endothelial cells, lysophosphatidylcholine (LPC) a major component of LDL and oxidized LDL [oxLDL]) reduced Nos3 gene expression and NO production. Antagonism of the AhR inhibited LPC effects on endothelial cells and induced decreased endothelium-dependent vasodilation. CONCLUSION AND IMPLICATIONS: AhR deletion attenuates HF diet-induced dyslipidaemia and vascular dysfunction by improving eNOS/NO signalling. Targeting AhRs may prevent obesity-associated vascular dysfunction.

Correlation between heart rate variability and polysomnography-derived scores of obstructive sleep apnea.

Obstructive sleep apnea (OSA) is one of the most common sleep disorders and affects nearly a billion people worldwide. Furthermore, it is estimated that many patients with OSA are underdiagnosed, which contributes to the development of comorbidities, such as cardiac autonomic imbalance, leading to high cardiac risk. Heart rate variability (HRV) is a non-invasive, widely used approach to evaluating neural control of the heart. This study evaluates the relationship between HRV indices and the presence and severity of OSA. We hypothesize that HRV, especially the nonlinear methods, can serve as an easy-to-collect marker for OSA early risk stratification. Polysomnography (PSG) exams of 157 patients were classified into four groups: OSA-free (N = 26), OSA-mild (N = 39), OSA-moderate (N = 37), and OSA-severe (N = 55). The electrocardiogram was extracted from the PSG recordings, and a 15-min beat-by-beat series of RR intervals were generated every hour during the first 6 h of sleep. Linear and nonlinear HRV approaches were employed to calculate 32 indices of HRV. Specifically, time- and frequency-domain, symbolic analysis, entropy measures, heart rate fragmentation, acceleration and deceleration capacities, asymmetry measures, and fractal analysis. Results with indices of sympathovagal balance provided support to reinforce previous knowledge that patients with OSA have sympathetic overactivity. Nonlinear indices showed that HRV dynamics of patients with OSA display a loss of physiologic complexity that could contribute to their higher risk of development of cardiovascular disease. Moreover, many HRV indices were found to be linked with clinical scores of PSG. Therefore, a complete set of HRV indices, especially the ones obtained by the nonlinear approaches, can bring valuable information about the presence and severity of OSA, suggesting that HRV can be helpful for in a quick diagnosis of OSA, and supporting early interventions that could potentially reduce the development of comorbidities.

Short-term effect of ligature-induced periodontitis on cardiovascular variability and inflammatory response in spontaneously hypertensive rats.

BACKGROUND: We previously reported that periodontal disease (PD) induces high arterial pressure variability (APV) consistent with sympathetic overactivity and elicits myocardial inflammation in Balb/c mice. However, it is unknown whether PD can change APV and heart rate variability (HRV) in spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats. This study aimed to evaluate the hemodynamic level, HRV, and APV associating with myocardial inflammation and plasma concentrations of oxide nitric (NO) in SHR and WKY rats with PD. METHODS: Three weeks after bilateral ligation of the first mandibular molar, or Sham operation, the rats received catheters into the femoral artery and had their arterial pressure (AP) recorded the following day. Subsequently, plasma, heart, and jaw were collected. The NO was quantified by the chemiluminescence method in plasma, and the myocardial IL-1ß concentrations were evaluated by ELISA. In the jaw was evaluated linear alveolar bone loss induced by PD. RESULTS: The linear alveolar bone loss in jaws of SHR with PD was higher than in all other groups. AP and heart rate were higher in SHR than in their WKY counterparts. SHR with PD showed lower AP than control SHR. HRV and APV were different between SHR and WKY rats; however, no differences in these parameters were found between the animals with PD and their control counterparts. Plasma NO and myocardial IL-1ß concentrations were higher in SHR with PD as compared to control WKY. A significant correlation was found between linear alveolar bone loss and plasma NO and myocardial IL-1ß concentrations. CONCLUSION: Our results demonstrated that short-term PD lowered the AP in SHR, which might be due to the higher levels of plasma NO. Even though PD did not affect either HRV or APV, it did induce myocardial inflammation, which can determine cardiovascular dysfunction in long-term PD.

Heart rate fragmentation, a novel approach in heart rate variability analysis, is altered in rats 4 and 12 weeks after myocardial infarction.

An approach recently proposed to assess ultra-rapid patterns of heart rate variability, namely heart rate fragmentation (HRF), is increased in aging and coronary disease. We aimed to evaluate and to correlate HRF with cardiac functional parameters in a rat model of myocardial infarction (MI). Wistar rats were submitted to MI (n = 18) or sham operation (n = 20), and after 4 or 12 weeks, their arterial pressure was recorded. Subsequently, cardiac function was evaluated by echocardiography. From pulse interval series, HRF patterns with zero, one, two, or three inflection points were estimated, as well as the total percentage of inflection points (PIP). Cardiac function was reduced in MI rats. Ejection fraction was smaller 4 (28 ± 3 vs 68 ± 2%, p < 0.0001) and 12 weeks after MI (38 ± 3 vs 70 ± 3%, p < 0.0001). Fractional shortening was also smaller 4 (13 ± 2 vs 41 ± 2%, p < 0.0001) and 12 weeks after MI (20 ± 2 vs 41 ± 3%, p < 0.0001). PIP was increased in MI rats 4 (74 ± 2 vs 69 ± 1%, p = 0.03) and 12 weeks after surgery (70 ± 2 vs 63 ± 1%, p = 0.02). We found a significant negative correlation between cardiac functional parameters and HRF at both 4 and 12 weeks after MI. These findings reveal that MI increases HRF, reinforcing the importance of this approach to explore pathophysiological conditions. Evaluation of heart rate fragmentation (HRF) in a rat model of myocardial infarction (MI). Wistar rats were submitted to MI (n = 18) or sham operation (n = 20), and after 4 or 12 weeks, their arterial pressure was recorded. Cardiac function was evaluated by echocardiography. From pulse interval series, HRF patterns with zero (W0), one (W1), two (W3), or three (W3) inflection points were estimated, as well as the total percentage of inflection points (PIP). Cardiac function was reduced, while PIP was increased in all MI rats. Fluent patterns (W0 and W1) decreased in MI rats after 12 weeks. Altogether, the findings reveal that MI increases HRF, reinforcing the potential of this approach to explore pathophysiological conditions.

MMP inhibition attenuates hypertensive eccentric cardiac hypertrophy and dysfunction by preserving troponin I and dystrophin.

PURPOSE: Cardiac transition from concentric (C-LVH) to eccentric left ventricle hypertrophy (E-LVH) is a maladaptive response of hypertension. Matrix metalloproteinases (MMPs), in particular MMP-2, may contribute to tissue remodeling by proteolyzing extra- and intracellular proteins. Troponin I and dystrophin are two potential targets of MMP-2 examined in this study and their proteolysis would impair cardiac contractile function. We hypothesized that MMP-2 contributes to the decrease in troponin I and dystrophin in the hypertensive heart and thereby controls the transition from C-LVH to E-LVH and cardiac dysfunction. METHODS: Male Wistar rats were divided into sham or two kidney-1 clip (2K-1C) hypertensive groups and treated with water (vehicle) or doxycycline (MMP inhibitor, 15 mg/kg/day) by gavage from the tenth to the sixteenth week post-surgery. Tail-cuff plethysmography, echocardiography, gelatin zymography, confocal microscopy, western blot, mass spectrometry, in silico protein analysis and immunofluorescence were performed. RESULTS: 6 out of 23 2K-1C rats (26%) had E-LVH followed by reduced ejection fraction. The remaining had C-LVH with preserved cardiac function. Doxycycline prevented the transition from C-LVH to E-LVH. MMP activity is increased in C-LVH and E-LVH hearts which was inhibited by doxycycline. This effect was associated with an increase in troponin I cleavage products and a decline in dystrophin in the left ventricle of E-LVH rats, which was prevented by doxycycline. CONCLUSION: Hypertension causes increased cardiac MMP-2 activity which proteolyzes troponin I and dystrophin, contributing to the transition from C-LVH to E-LVH and cardiac dysfunction.

The ability of baroreflex activation to improve blood pressure and resistance vessel function in spontaneously hypertensive rats is dependent on stimulation parameters.

Baroreflex activation by electric stimulation of the carotid sinus (CS) effectively lowers blood pressure. However, the degree to which differences between stimulation protocols impinge on cardiovascular outcomes has not been defined. To address this, we examined the effects of short- and long-duration (SD and LD) CS stimulation on hemodynamic and vascular function in spontaneously hypertensive rats (SHRs). We fit animals with miniature electrical stimulators coupled to electrodes positioned around the left CS nerve that delivered intermittent 5/25 s ON/OFF (SD) or 20/20 s ON/OFF (LD) square pulses (1 ms, 3 V, 30 Hz) continuously applied for 48 h in conscious animals. A sham-operated control group was also studied. We measured mean arterial pressure (MAP), systolic blood pressure variability (SBPV), heart rate (HR), and heart rate variability (HRV) for 60 min before stimulation, 24 h into the protocol, and 60 min after stimulation had stopped. SD stimulation reversibly lowered MAP and HR during stimulation. LD stimulation evoked a decrease in MAP that was sustained even after stimulation was stopped. Neither SD nor LD had any effect on SBPV or HRV when recorded after stimulation, indicating no adaptation in autonomic activity. Both the contractile response to phenylephrine and the relaxation response to acetylcholine were increased in mesenteric resistance vessels isolated from LD-stimulated rats only. In conclusion, the ability of baroreflex activation to modulate hemodynamics and induce lasting vascular adaptation is critically dependent on the electrical parameters and duration of CS stimulation.

Electrocardiographic changes in the acute hyperkalaemia produced by intragastric KCl load in rats.

NEW FINDINGS: What is the central question of this study? This study presents a new model for studying the rapid onset of severe, acute hyperkalaemia in rats with intact kidney function by administering an intragastric KCl load. What is the main finding and its importance? This new model of intragastric KCl load produces a reliable and reproducible model for studying the rapid onset of severe, acute hyperkalaemia in rats with intact kidney function. We report unprecedented rapid changes (30 min) in ECG, blood pressure and various arterial blood analyses with this new model, providing a solid foundation for future experiments in this field. ABSTRACT: A variety of animal models have been proposed to study hyperkalaemia, but most of them have meaningful limitations when the goal is to study the effect of potassium overload on healthy kidneys. In this study, we aimed to introduce a new approach for induction of hyperkalaemia in a reliable and reproducible animal model. We used intragastric administration of potassium chloride [KCl 2.3 M, 10 ml/(kg body weight)] to male Holtzman rats (300-350 g) to induce hyperkalaemia. The results showed that this potassium load can temporarily overwhelm the renal and extrarenal handling of this ion, causing an acute and severe hyperkalaemia that can be useful to study the effect of potassium imbalance in a variety of scenarios. Severe hyperkalaemia (>8 meqiv/l) and very profound ECG alterations, characterized by lengthening waves and intervals, were seen as early as 30 min after intragastric administration of KCl in rats. In addition, a transient increase in arterial blood pressure and time-dependent bradycardia were also seen after the KCl administration. No metabolic acidosis was present in the animals, and the potassium ion did not increase proportionally to chloride ion in the blood, leading to an increased anion gap. In conclusion, the results suggest that intragastric KCl loading is a reliable model to promote rapid and severe hyperkalaemia that can be used for further research on this topic.

PyBioS: A freeware computer software for analysis of cardiovascular signals.

BACKGROUND AND OBJECTIVE: Several software applications have been proposed in the past years as computational tools for assessing biomedical signals. Many of them are focused on heart rate variability series only, with their strengths and limitations depending on the necessity of the user and the scope of the application. Here, we introduce new software, named PyBioS, intended for the analysis of cardiovascular signals, even though any type of biomedical signal can be used. PyBioS has some functionalities that differentiate it from the other software. METHODS: PyBioS was developed in Python language with an intuitive, user-friendly graphical user interface. The basic steps for using PyBioS comprise the opening or creation (simulation) of signals, their visualization, preprocessing and analysis. Currently, PyBioS has 8 preprocessing tools and 15 analysis methods, the later providing more than 50 metrics for analysis of the signals' dynamics. RESULTS: The possibility to create simulated signals and save the preprocessed signals is a strength of PyBioS. Besides, the software allows batch processing of files, making the analysis of a large amount of data easy and fast. Finally, PyBioS has plenty of analysis methods implemented, with the focus on nonlinear and complexity analysis of signals and time series. CONCLUSIONS: Although PyBioS is not intended to overcome all the necessities from users, it has useful functionalities that may be helpful in many situations. Moreover, PyBioS is continuously under improvement and several simulated signals, tools and analysis methods are still to be implemented. Also, a new module is being implemented on it to provide machine learning algorithms for classification and regression of data extracted from the biomedical signals.

Electrical stimulation of the carotid sinus lowers arterial pressure and improves heart rate variability in L-NAME hypertensive conscious rats.

We evaluated the effects of long-term (48 h) electrical stimulation of the carotid sinus (CS) in hypertensive rats. L-NAME-treated (10 days) Wistar rats were implanted with a catheter in the femoral artery and a miniaturized electrical stimulator attached to electrodes positioned around the left CS, encompassing the CS nerve. One day after implantation, arterial pressure (AP) was directly recorded in conscious animals for 60 min. Square pulses (1 ms, 3 V, 30 Hz) were applied intermittently (20/20 s ON/OFF) to the CS for 48 h. After the end of stimulation, AP was recorded again. Nonstimulated rats (control group) and rats without electrodes around the CS (sham-operated) were also studied. Next, the animals were decapitated, and segments of mesenteric resistance arteries were removed to study vascular function. After the stimulation period, AP was 16 ± 5 mmHg lower in the stimulated group, whereas sham-operated and control rats showed similar AP between the first and second recording periods. Heart rate variability (HRV) evaluated using time and frequency domain tools and a nonlinear approach (symbolic analysis) suggested that hypertensive rats with electrodes around the CS, stimulated or not, exhibited a shift in cardiac sympathovagal balance towards parasympathetic tone. The relaxation response to acetylcholine in endothelium-intact mesenteric arteries was enhanced in rats that underwent CS stimulation for 48 h. In conclusion, long-term CS stimulation is effective in reducing AP levels, improving HRV and increasing mesenteric vascular relaxation in L-NAME hypertensive rats. Moreover, only the presence of electrodes around the CS is effective in eliciting changes in HRV similar to those observed in stimulated rats.

Cardiac sympathetic drive is increased in cafeteria diet-fed rats independent of impairment in peripheral baroreflex and chemoreflex functions.

BACKGROUND AND AIMS: Consumption of a high caloric diet induces autonomic imbalance, which can lead to cardiovascular disease. Impaired arterial baroreflex control is suggested to play an important role in cardiovascular autonomic imbalance, often seen in obesity. We previously demonstrated that cafeteria diets increase the sympathetic drive to white and brown adipose tissue. METHODS AND RESULTS: After feeding a cafeteria diet to rats for 26 days, we evaluated: (i)heart rate (HR) and arterial pressure (AP); (ii)baroreflex and chemoreflex function; and (iii) autonomic modulation of the heart and vessels, measured through pulse interval (PI) and systolic arterial pressure (SAP) variability analyses and following administration of autonomic blockers. The cafeteria diet increased body fat mass and serum insulin, leptin, triacylglycerol and cholesterol levels. Baseline HR (15%) was also increased, accompanied by increased power in the low frequency band (60%) and in the low frequency/high frequency ratio (104%) in the PI spectra. Nonlinear analysis revealed an increased occurrence of 0V (39%) and decreased occurrence of 2UV (18%) patterns. Following administration of autonomic blockers, we observed an increase in cardiac sympathetic tone (425%) in cafeteria diet-fed rats. The cafeteria diet had no effect on AP, SAP variability, baroreflex and chemoreflex control. CONCLUSION: Our findings suggest that consumption of a cafeteria diet increases sympathetic drive to the heart but not to the blood vessels, independent of impairment in baroreflex and chemoreflex functions. Other mechanisms may be involved in the increased cardiac sympathetic drive, and compensatory vascular mechanisms may prevent the development of hypertension in this model of obesity.

Verapamil decreases calpain-1 and matrix metalloproteinase-2 activities and improves hypertension-induced hypertrophic cardiac remodeling in rats.

AIMS: Increased activity of calpain-1 and matrix metalloproteinase (MMP)-2 was observed in different models of arterial hypertension and contribute to thicken the left ventricle (LV) walls and to hypertrophy cardiac myocytes. MMP-2 activity may be regulated by calpain-1 via bioactive molecules activation such as transforming growth factor (TGF)-ß in cardiovascular diseases. This study analyzed whether calpain-1 causes cardiac hypertrophy and dysfunction by modulating the expression and activity of MMP-2 in renovascular hypertension. MAIN METHODS: Male Wistar rats were submitted to two kidneys, one clip (2K1C) model of hypertension or sham surgery and were treated with verapamil (VRP, 8 mg/kg/bid) by gavage from the second to tenth week post-surgery. Systolic blood pressure (SBP) was weekly assessed by tail-cuff plethysmography and morphological and functional parameters of LV were analyzed by echocardiography. MMP-2 activity was analyzed by in situ and gelatin zymography, while calpain-1 activity by caseinolytic assay. MMP-2, calpain-1, TGF-ß and MMP-14/TIMP-2 levels were identified in the LV by western blots. Fluorescence assays were performed to evaluate oxidative stress, MMP-2 and calpain-1 levels. KEY FINDINGS: SBP increased in 2K1C rats and was unaltered by VRP. However, VRP notably ameliorated hypertension-induced increase in LV thickness. VRP decreased hypertension-induced enhances in calpain-1 and MMP-2 activities, oxidative stress and mature TGF-ß levels. Treatment with VRP also decreased the accentuated MMP-14/TIMP-2 levels in 2K1C. SIGNIFICANCE: Treatment with VRP decreases calpain-1 and MMP-2 activities and also reduces TGF-ß and MMP-14/TIMP-2 levels in the LV of hypertensive rats, thus contributing to ameliorate cardiac hypertrophy.

Revisiting the Sequence Method for Baroreflex Analysis.

The sequence method is an important approach to assess the baroreflex function, mainly because it is based on the spontaneous fluctuations of beat-by-beat arterial pressure (for example, systolic arterial pressure or SAP) and pulse interval (PI). However, some studies revealed that the baroreflex effectiveness index (BEI), calculated through the sequence method, shows an intriguing oscillatory pattern as function of the delay between SAP and PI. It has been hypothesized that this pattern is related to the respiratory influence on SAP and/or PI variability, limiting the SAP ramps to 3 or 4 beats of length. In this study, this hypothesis was tested by assessing the sequence method using raw (original) and filtered series. Results were contrasted to the well-established transfer function, estimated between SAP and PI. Continuous arterial pressure recordings were obtained from healthy rats (N = 61) and beat-by-beat series of SAP and PI were generated. Low-pass (LP) and high-pass (HP) filtered series of SAP and PI were created by filtering the original series with a cutoff frequency of 0.8 Hz. Original series were analyzed by either the sequence method or cross-spectral analysis (transfer function) at low- (LF) and high- (HF) frequency bands, while filtered series were evaluated only by the sequence method. Baroreflex sensitivity (BRS) and BEI of original series, calculated by sequence method, was highly (85-90%) determined by HP series, with no significant association between original and LP series. A high correlation (>0.7) was found between the BRS estimated from original series (sequence method) and HF band (transfer function), as well as for LP series (sequence method) and LF band (transfer function). These findings confirmed the hypothesis that the sequence method quantifies only the high-frequency components of the baroreflex, neglecting the low-frequency influences, such as the Mayer waves. Therefore, we propose using both the original and LP filtered time series for a broader assessment of the baroreflex function using the sequence method.

Interaction between baroreflex and chemoreflex in the cardiorespiratory responses to stimulation of the carotid sinus/nerve in conscious rats.

Electrical stimulation of the carotid baroreflex has been thoroughly investigated for treating drug-resistant hypertension in humans. However, a previous study from our laboratory, performed in conscious rats, has demonstrated that electrical stimulation of the carotid sinus/nerve (CS) activated both the carotid baroreflex as well as the carotid chemoreflex, resulting in hypotension. Additionally, we also demonstrated that the carotid chemoreceptor deactivation potentiated this hypotensive response. Therefore, to further investigate this carotid baroreflex/chemoreflex interaction, besides the hemodynamic responses, we evaluated the respiratory responses to the electrical stimulation of the CS in both intact (CONT) and carotid chemoreceptors deactivated (CHEMO-X) conscious rats. CONT rats showed increased ventilation in response to electrical stimulation of the CS as measured by the respiratory frequency (fR), tidal volume (VT) and minute ventilation (VE), suggesting a carotid chemoreflex activation. The carotid chemoreceptor deactivation abolished all respiratory responses to the electrical stimulation of the CS. Regarding the hemodynamic responses, the electrical stimulation of the CS caused hypotensive responses in CONT rats, which were potentiated by the carotid chemoreceptors deactivation. Heart rate (HR) responses did not differ between groups. In conclusion, the present study showed that the electrical stimulation of the CS, in conscious rats, activates both the carotid baroreflex and the carotid chemoreflex driving an increase in ventilation and a decrease in AP. These findings further contribute to our understanding of the electrical stimulation of CS.

Acute Increase in O-GlcNAc Improves Survival in Mice With LPS-Induced Systemic Inflammatory Response Syndrome.

Sepsis is a systemic inflammatory response syndrome (SIRS) resulting from a severe infection that is characterized by immune dysregulation, cardiovascular derangements, and end-organ dysfunction. The modification of proteins by O-linked N-acetylglucosamine (O-GlcNAcylation) influences many of the key processes that are altered during sepsis, including the production of inflammatory mediators and vascular contractility. Here, we investigated whether O-GlcNAc affects the inflammatory response and cardiovascular dysfunction associated with sepsis. Mice received an intraperitoneal injection of lipopolysaccharide (LPS, 20 mg/Kg) to induce endotoxic shock and systemic inflammation, resembling sepsis-induced SIRS. The effects of an acute increase in O-GlcNAcylation, by treatment of mice with glucosamine (GlcN, 300 mg/Kg, i.v.) or thiamet-G (ThG, 150 µg/Kg, i.v.), on LPS-associated mortality, production and release of cytokines by macrophages and vascular cells, vascular responsiveness to constrictors and blood pressure were then determined. Mice under LPS-induced SIRS exhibited a systemic and local inflammatory response with increased levels of interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor (TNF-α), as well as severe hypotension and vascular hyporesponsiveness, characterized by reduced vasoconstriction to phenylephrine. In addition, LPS increased neutrophil infiltration in lungs and produced significant lethality. Treatment with GlcN and ThG reduced systemic inflammation and attenuated hypotension and the vascular refractoriness to phenylephrine, improving survival. GlcN and ThG also decreased LPS-induced production of inflammatory cytokines by bone marrow-derived macrophages and nuclear transcription factor-kappa B (NF-κB) activation in RAW 264.7 NF-κB promoter macrophages. Treatment of mice with ThG increased O-glycosylation of NF-κB p65 subunit in mesenteric arteries, which was associated with reduced Ser536 phosphorylation of NF-κB p65. Finally, GlcN also increased survival rates in mice submitted to cecal ligation and puncture (CLP), a sepsis model. In conclusion, increased O-GlcNAc reduces systemic inflammation and cardiovascular disfunction in experimental sepsis models, pointing this pathway as a potential target for therapeutic intervention.

Lack of scarring is not always a sign of cardiac health: Functional and molecular characterization of the rat heart's following chronic reperfusion.

Even though the coronary reperfusion process is the most important tool to preserve cardiac function, after myocardial infarction, reperfusion of acutely ischemic myocardium can induce injury. We aimed to evaluate the functional and molecular aspects 4 weeks after myocardial ischemia-reperfusion (IR) in rats. Male Wistar rats (N = 47) were subjected to myocardial IR by short-term (30 min) ligation and subsequent reperfusion of the left descending coronary artery. Control rats (N = 7) underwent the same surgical maneuver without coronary ligation. After 4 weeks, rats had their cardiac function examined by ventricular pressure recording under basal condition or pharmacological stress. Myocardial fibrosis and molecular mediators of IR injury (reactive oxygen species, tumor necrosis factor-alpha and matrix-metalloproteinase-2) were assessed as well. Most of the rats subjected to IR did not show macroscopic signs of infarct, while only 17% of these animals showed large myocardial infarction scars. Of note, all animals submitted to IR presented the functional and molecular parameters altered when compared with the control subjects. Cardiac function was attenuated in all animals submitted to IR, regardless the presence or size of macroscopic cardiac scars. Interstitial fibrosis, matrix-metalloproteinase-2 activity and the expression of tumor necrosis factor-alpha were higher in the myocardium of all IR rats as compared to the control subjects (p<0.05). Myocardium superoxide anion and hydrogen peroxide were increased in rats without or with mild cardiac scars. These results show that IR leads to myocardial injury in rats. Besides, even the animals with an apparent healthy myocardium (without infarct scar) presented cardiac dysfunction and molecular changes that may contribute to the development of heart failure over time.

Baroreflex stimulation attenuates central but not peripheral inflammation in conscious endotoxemic rats.

We previously reported that activation of the baroreflex, a critical physiological mechanism controlling cardiovascular homeostasis, through electrical stimulation of the aortic depressor nerve attenuates joint inflammation in experimental arthritis. However, it is unknown whether baroreflex activation can control systemic inflammation. Here, we investigate whether baroreflex activation controls systemic inflammation in conscious endotoxemic rats. Animals underwent sham or electrical aortic depressor nerve stimulation initiated 10 min prior to a lipopolysaccharide (LPS) challenge, while inflammatory cytokine levels were measured in the blood, spleen, heart and hypothalamus 90 min after LPS treatment. Baroreflex activation did not affect LPS-induced levels of pro-inflammatory (tumor necrosis factor, interleukin 1ß and interleukin 6) or anti-inflammatory (interleukin 10) cytokines in the periphery (heart, spleen and blood). However, baroreflex stimulation attenuated LPS-induced levels of all these cytokines in the hypothalamus. Notably, these results indicate that the central anti-inflammatory mechanism induced by baroreflex stimulation is independent of cardiovascular alterations, since aortic depressor nerve stimulation that failed to induce hemodynamic changes was also efficient at inhibiting inflammatory cytokines in the hypothalamus. Thus, aortic depressor nerve stimulation might represent a novel therapeutic strategy for neuroprotection, modulating inflammation in the central nervous system.