Unveiling mutation effects on the structural dynamics of the main protease from SARS-CoV-2 with hybrid simulation methods.

The main protease of SARS-CoV-2 (called Mpro or 3CLpro) is essential for processing polyproteins encoded by viral RNA. Several Mpro mutations were found in SARS-CoV-2 variants, which are related to higher transmissibility, pathogenicity, and resistance to neutralization antibodies. Macromolecules adopt several favored conformations in solution depending on their structure and shape, determining their dynamics and function. In this study, we used a hybrid simulation method to generate intermediate structures along the six lowest frequency normal modes and sample the conformational space and characterize the structural dynamics and global motions of WT SARS-CoV-2 Mpro and 48 mutations, including mutations found in P.1, B.1.1.7, B.1.351, B.1.525 and B.1.429+B.1.427 variants. We tried to contribute to the elucidation of the effects of mutation in the structural dynamics of SARS-CoV-2 Mpro. A machine learning analysis was performed following the investigation regarding the influence of the K90R, P99L, P108S, and N151D mutations on the dimeric interface assembling of the SARS-CoV-2 Mpro. The parameters allowed the selection of potential structurally stable dimers, which demonstrated that some single surface aa substitutions not located at the dimeric interface (K90R, P99L, P108S, and N151D) are able to induce significant quaternary changes. Furthermore, our results demonstrated, by a Quantum Mechanics method, the influence of SARS-CoV-2 Mpro mutations on the catalytic mechanism, confirming that only one of the chains of the WT and mutant SARS-CoV-2 Mpros are prone to cleave substrates. Finally, it was also possible to identify the aa residue F140 as an important factor related to the increasing enzymatic reactivity of a significant number of SARS-CoV-2 Mpro conformations generated by the normal modes-based simulations.

Chronic Mercury Exposure in Prehypertensive SHRs Accelerates Hypertension Development and Activates Vasoprotective Mechanisms by Increasing NO and H2O2 Production.

Mercury is a heavy metal associated with cardiovascular diseases. Studies have reported increased vascular reactivity without changes in systolic blood pressure (SBP) after chronic mercury chloride (HgCl2) exposure, an inorganic form of the metal, in normotensive rats. However, we do not know whether individuals in the prehypertensive phase, such as young spontaneously hypertensive rats (SHRs), are susceptible to increased arterial blood pressure. We investigated whether chronic HgCl2 exposure in young SHRs accelerates hypertension development by studying the vascular function of mesenteric resistance arteries (MRAs) and SBP in young SHRs during the prehypertensive phase. Four-week-old male SHRs were divided into two groups: the SHR control group (vehicle) and the SHR HgCl2 group (4 weeks of exposure). The results showed that HgCl2 treatment accelerated the development of hypertension; reduced vascular reactivity to phenylephrine in MRAs; increased nitric oxide (NO) generation; promoted vascular dysfunction by increasing the production of reactive oxygen species (ROS), such as hydrogen peroxide (H2O2); increased Gp91Phox protein levels and in situ levels of superoxide anion (O2·-); and reduced vasoconstrictor prostanoid production compared to vehicle treatment. Although HgCl2 accelerated the development of hypertension, the HgCl2-exposed animals also exhibited a vasoprotective mechanism to counterbalance the rapid increase in SBP by decreasing vascular reactivity through H2O2 and NO overproduction. Our results suggest that HgCl2 exposure potentiates this vasoprotective mechanism against the early establishment of hypertension. Therefore, we are concluding that chronic exposure to HgCl2 in prehypertensive animals could enhance the risk for cardiovascular diseases.

Ultra-short-term heart rate variability during resistance exercise in the elderly.

Despite the appeal of ultra-short-term heart rate variability (HRV) methods of analysis applied in the clinical and research settings, the number of studies that have investigated HRV by analyzing R-R interval (RRi) recordings shorter than 5 min is still limited. Moreover, ultra-short-term HRV analysis has not been extensively validated during exercise and, currently, no indications exist for its applicability during resistance exercise. The aim of the present study was to compare ultra-short-term HRV analysis with standard short-term HRV analysis during low-intensity, dynamic, lower limb resistance exercise in healthy elderly subjects. Heart rate (HR) and RRi signals were collected from 9 healthy elderly men during discontinuous incremental resistance exercise consisting of 4-min intervals at low intensities (10, 20, 30, and 35% of 1-repetition maximum). The original RRi signals were segmented into 1-, 2-, and 3-min sections. HRV was analyzed in the time domain (root mean square of the of differences between adjacent RRi, divided by the number of RRi, minus one [RMSSD]), RRi mean value and standard deviation [SDNN] (percentage of differences between adjacent NN intervals that are greater than 50 ms [pNN50]), and by non-linear analysis (short-term RRi standard deviation [SD1] and long-term RRi standard deviation [SD2]). No significant difference was found at any exercise intensity between the results of ultra-short-term HRV analysis and the results of standard short-term HRV analysis. Furthermore, we observed excellent (0.70 to 0.89) to near-perfect (0.90 to 1.00) concordance between linear and non-linear parameters calculated over 1- and 2-min signal sections and parameters calculated over 3-min signal sections. Ultra-short-term HRV analysis appears to be a reliable surrogate of standard short-term HRV analysis during resistance exercise in healthy elderly subjects.

Anaerobic Threshold by Mathematical Model in Healthy and Post-Myocardial Infarction Men.

The aim of this study was to determine the anaerobic threshold (AT) in a population of healthy and post-myocardial infarction men by applying Hinkley's mathematical method and comparing its performance to the ventilatory visual method. This mathematical model, in lieu of observer-dependent visual determination, can produce more reliable results due to the uniformity of the procedure. 17 middle-aged men (55±3 years) were studied in 2 groups: 9 healthy men (54±2 years); and 8 men with previous myocardial infarction (57±3 years). All subjects underwent an incremental ramp exercise test until physical exhaustion. Breath-by-breath ventilatory variables, heart rate (HR), and vastus lateralis surface electromyography (sEMG) signal were collected throughout the test. Carbon dioxide output (V˙CO2), HR, and sEMG were studied, and the AT determination methods were compared using correlation coefficients and Bland-Altman plots. Parametric statistical tests were applied with significance level set at 5%. No significant differences were found in the HR, sEMG, and ventilatory variables at AT between the different methods, such as the intensity of effort relative to AT. Moreover, important concordance and significant correlations were observed between the methods. We concluded that the mathematical model was suitable for detecting the AT in both healthy and myocardial infarction subjects.

Lactate and heart rate variability threshold during resistance exercise in the young and elderly.

This purpose of this study was to: 1) determine the intensity corresponding to anaerobic threshold (AT) during a discontinuous resistance exercise protocol in healthy young and elderly subjects by analyzing heart rate variability (HRV) and blood lactate (BL) and 2) investigate the effect of aging on these variables. A total of 28 individuals, 14 young and 14 elderly healthy men underwent one-repetition maximum (1RM) testing to determine maximum load on the leg press. Discontinuous resistance exercise testing was initiated at 10% of the 1RM with subsequent increases of 10%. The load corresponding to AT was approximately 30% 1RM in both groups. The determination of AT by HRV was associated with BL responses (p<0.01). While HRV indexes decreased with increasing of loads in both groups, the elderly had lower values at loads below AT (p<0.05). Additionally, BL increased sharply after the load corresponding to AT in both groups, although elderly subjects showed the lowest values (p<0.05). In conclusion, HRV is an effective tool for determining AT, which was approximately 30% 1RM under the testing procedures included in the present study. Furthermore, there was a marked change in autonomic function, with gradual vagal withdrawal followed by sympathetic activation. These responses were lower in elderly subjects.

MMP(-2) expression in skeletal muscle after strength training.

The aim of this study was to assess the effects of resistance training on ladders (RTL) on MMP(-2) expression and blood lactate concentration [La-]. 30 male (3 months of age), albino rats were divided into 3 groups: sedentary control (SC, n=10), low resistance exercise training (Low-IntRT, n=10) and high-intensive exercise training (High-IntRT, n=10). Animals of High-IntRT were submitted to a progressively increasing overload in relation to body weight until exhaustion, while the Low-IntRT group performed the same exercise regimen with no external load. The program had a frequency of 3 times per week over 8 weeks. MMP(-2) expression of tibialis anterior muscle and [La-] were measured. While there was a significant increase of MMP(-2) (pro-form) in both groups, only High-IntRT significantly increased MMP(-2) in active-form (p<0.05). Both trained groups exhibited an increase in [La-] when compared to controls, however, the increase in [La-] was significantly higher in the High-IntRT compared to Low-IntRT (p<0.05). Strong correlation was found between MMP(-2) (active form) and [La-] in High-IntRT (r=0.91). RTL in using low and high-intensity exercise can serve as a model to demonstrate different responses of MMP(-2) expression in an animal model. It appears active form expression of MMP(-2) is modulated by exercise intensity.

Cardiac autonomic modulation during progressive upper limb exercise by patients with coronary artery disease

The purpose of this study was to investigate the behavior of heart rate (HR) and HR variability (HRV) during different loads of resistance exercise (incline bench press) in patients with coronary artery disease (CAD) and healthy sedentary controls. Ten healthy men (65 ± 1.2 years, control group, CG) and 10 men with clinically stable CAD (66 ± 2.4 years, CADG) were recruited. A discontinuous progressive protocol was applied with an initial load of 10 percent of the maximum load achieved in the 1RM (1 repetition maximum) with increases of 10 percent until 30 percent 1RM was reached, which was followed by subsequent increases of 5 percent 1RM until exhaustion. HRV was analyzed by linear and non-linear methods. There was a significant reduction in rMSSD (CG: 20 ± 2 to 11 ± 3 ms; CADG: 19 ± 3 to 9 ± 1 ms) and SD1 indexes (CG: 14 ± 2 to 8 ± 1 ms; CADG: 14 ± 2 to 7 ± 1 ms). An increase in HR (CG: 69 ± 5 to 90 ± 5 bpm; CADG: 62 ± 4 to 75 ± 4 bpm) and in systolic blood pressure (CG: 124 ± 3 to 138 ± 3 mmHg; CADG: 122 ± 6 to 126 ± 9 bpm) were observed (P < 0.05) when comparing pre-effort rest and 40 percent 1RM in both groups. Furthermore, an increase in RMSM index was also observed (CG: 28 ± 3 to 45 ± 9 ms; CADG: 22 ± 2 to 79 ± 33 ms), with higher values in CADG. We conclude that loads up to 30 percent 1RM during incline bench press result in depressed vagal modulation in both groups, although only stable CAD patients presented sympathetic overactivity at 20 percent 1RM upper limb exercise.

Cardiac autonomic modulation during progressive upper limb exercise by patients with coronary artery disease.

The purpose of this study was to investigate the behavior of heart rate (HR) and HR variability (HRV) during different loads of resistance exercise (incline bench press) in patients with coronary artery disease (CAD) and healthy sedentary controls. Ten healthy men (65 ± 1.2 years, control group, CG) and 10 men with clinically stable CAD (66 ± 2.4 years, CADG) were recruited. A discontinuous progressive protocol was applied with an initial load of 10% of the maximum load achieved in the 1RM (1 repetition maximum) with increases of 10% until 30% 1RM was reached, which was followed by subsequent increases of 5% 1RM until exhaustion. HRV was analyzed by linear and non-linear methods. There was a significant reduction in rMSSD (CG: 20 ± 2 to 11 ± 3 ms; CADG: 19 ± 3 to 9 ± 1 ms) and SD1 indexes (CG: 14 ± 2 to 8 ± 1 ms; CADG: 14 ± 2 to 7 ± 1 ms). An increase in HR (CG: 69 ± 5 to 90 ± 5 bpm; CADG: 62 ± 4 to 75 ± 4 bpm) and in systolic blood pressure (CG: 124 ± 3 to 138 ± 3 mmHg; CADG: 122 ± 6 to 126 ± 9 bpm) were observed (P < 0.05) when comparing pre-effort rest and 40% 1RM in both groups. Furthermore, an increase in RMSM index was also observed (CG: 28 ± 3 to 45 ± 9 ms; CADG: 22 ± 2 to 79 ± 33 ms), with higher values in CADG. We conclude that loads up to 30% 1RM during incline bench press result in depressed vagal modulation in both groups, although only stable CAD patients presented sympathetic overactivity at 20% 1RM upper limb exercise.

A PK/PD approach on the effects of clarithromycin against oral and nasal microbiota of healthy volunteers.

OBJECTIVE: To assess the pharmacokinetics of clarithromycin (CLR) and its effects on oral and nasal microbiota in healthy volunteers in an open, randomized, two-period crossover design. METHODS: A single 500 mg oral dose of CLR (Group 1: Merck; Group 2: Klaricid) was administered observing a 1-week interval between doses. Blood samples were collected from pre-dose to 24 h. Plasmatic concentrations of CLR were quantified by the LC-MS-MS method. Saliva and nasal mucosa swabs were obtained previously and after 1.33, 2, 6 and 12 h of drug administration. Pharmacokinetics and PK/PD (t > MIC, %t > MIC and AUC0-24/MIC ratio) parameters were estimated. The microorganism counts were obtained on different culture media. RESULTS: No statistically significant differences were observed between the two formulations (p > 0.05) regarding the pharmacokinetic parameters. Total microorganisms, staphylococci and streptococci counts did not show statistical differences (p > 0.05) between the two groups during each sampling time. Considering the microorganisms of each group, no statistically significant differences were found after drug administration, but all differed from pre-dose counts (p < 0.05). The observed t > MIC ranged from 14.45 h (+/- 1.69) to 1.19 h (+/- 2.17) considering MICs of 0.25 microg/ml and 2.0 microg/ml, respectively. There was no correlation between any t > MIC, %t > MIC or AUC0-24 and bacterial reduction (between 0- and 12-h periods). However, the profile of reduction of microorganisms in both saliva and nasal samples were compatible with high values of t > MIC verified for both clarithromycin formulations. CONCLUSION: Both formulations of clarithromycin had similar pharmacokinetics and efficacy.

Atividade autonômica em uma adolescente com ventrículo único submetida à intervenção fisioterapêutica: relato de caso / Autonomic activity in an adolescent with a single ventricle who underwent physical therapy intervention: case report

OBJETIVO: Avaliar a atividade autonômica de uma adolescente com ventrículo único (VU), sem correção cirúrgica, participante de um programa de fisioterapia. MATERIAIS E MÉTODOS: Paciente do sexo feminino, 14 anos, com diagnóstico de VU tipo esquerdo, sem correção cirúrgica. A atividade autonômica foi avaliada pela variabilidade da freqüência cardíaca (VFC) nas posições supina e sentada, e pela manobra para acentuar a arritmia sinusal respiratória (M-ASR) no início do primeiro (A1), segundo (A2) e terceiro anos (A3) de tratamento fisioterapêutico cardiovascular (TFC) ambulatorial. Os intervalos RR e a freqüência cardíaca batimento a batimento foram calculados e armazenados para posterior análise. A VFC foi avaliada pelos índices RMSSD e RMSM e no domínio da freqüência pelas bandas de alta e baixa freqüência em unidades normalizadas (AFun e BFun, respectivamente) e pela razão BF/AF. O TFC constou de exercícios respiratórios associados a exercícios ativos e resistidos gerais, durante dois anos. RESULTADOS: De A1 para A3, o RMSM reduziu (14,5 por cento), a banda de BFun (42,2 por cento) e a razão BF/AF aumentou (117,0 por cento), e a banda AFun diminuiu (35,2 por cento). Em todas as situações, a banda BFun foi maior e a AFun foi menor na posição sentada. Além disso, a resposta parassimpática à M-ASR aumentou da situação A1 para A3 em 7,4 e 47,3 por cento, respectivamente. CONCLUSÕES: Concluímos que, na paciente estudada, a redução da VFC parece estar associada com o avanço da doença, porém, não houve prejuízos frente à mudança postural. Adicionalmente, o TFC proposto incrementou a resposta parassimpática durante a M-ASR.

OBJECTIVE: To evaluate the autonomic activity of an adolescent with a single ventricle without surgical correction who participated in a physical therapy program. METHODS: This was a 14-year-old female patient with a left-type single ventricle who had not undergone any surgical intervention. The autonomic activity was evaluated according to the heart rate variability in the supine and seated positions and by means of the respiratory sinus arrhythmia accentuation maneuver (RSA-M) in the beginning of the first, second and third years of outpatient cardiovascular physical therapy treatment. The RR intervals and beat-to-beat heart rate were calculated and stored for subsequent analysis. The heart rate variability was evaluated by means of the RMSSD and RMSM indexes and in the frequency domain with high and low frequency bands in normalized units (HFnu and LFnu, respectively) and using the LF/HF ratio. The cardiovascular physical therapy consisted of respiratory exercises associated with active and general resistance exercises, for two years. RESULTS: From the first to the third year, the RMSM decreased (14.5 percent), LFnu band increased (42.2 percent), LF/HF ratio increased (117.0 percent) and HFnu decreased (35.2 percent). In all of the situations, the LFnu band was higher and HFnu was lower in the seated position. Additionally, the parasympathetic response to RSA-M increased from the first to the third years in 7.4 percent and 47.3 percent, respectively. CONCLUSIONS: We concluded that, for the patient studied, the reduction in heart rate variability seemed to be associated with the advance of the disease. Nevertheless, there was no damages related to postural change. In addition, the cardiovascular physical therapy proposed increased the parasympathetic response during RSA-M.

Comparative bioavailability of clarithromycin formulations in healthy brazilian volunteers.

OBJECTIVE: To compare the bioavailability of clarithromycin 500 mg tablets (Merck S.A Industrias Quimicas, Sao Paulo, SP, Brazil, used as test formulation) and Klaricid (Abbott Laboratórios do Brasil Ltda, Sao Paulo, SP, Brazil, used as reference formulation) in 24 healthy volunteers. MATERIAL AND METHODS: The study was conducted using an open, randomized, two-period crossover design with one-week interval between doses. Blood samples were collected at pre-dose, 0.33, 0.66, 1, 1.33, 1.66, 2, 2.5, 3, 4, 6, 8, 10, 12, 16, 20 and 24 hours after the administration. AUC was calculated by the trapezoidal rule extrapolation method. Cmax and tmax were compiled from the plasmatic concentration-time data. Analysis of variance was carried out using logarithmically transformed AUC(0-inf), AUC(0-24 h), Cmax and untransformed tmax. RESULTS: Intraindividual coefficient of variation (CV%) values were 14.25% and 12.62%, respectively for Cmax and AUC(0-24 h). The geometric mean values (+/- SD) for AUC(0-24 h) (microg x h/ml), AUC(0-inf) (microg x h/ml), and Cmax (microg/ml) for test medication were 18.56 (+/- 6.87), 18.8 (+/- 5.70) and 2.45 (+/- 0.88); the obtained values for reference medication were 18.29 (+/- 5.39), 19.10 (+/- 7.21) and 2.5 (+/- 0.69). 90% Cl for clarithromycin geometric mean of AUC(0-24 h), AUC(0-inf) and Cmax ratios (test/reference) were: 93.6-105.9%, 93.8-106.2% and 89- 103.2%. CCONCLUSION The test medication was considered bioequivalent to the reference medication based on the rate and extent of absorption.

Antimicrobial activity of garlic, tea tree oil, and chlorhexidine against oral microorganisms.

OBJECTIVE: To compare the antimicrobial activity of tea tree oil, garlic, and chlorhexidine solutions against oral microorganisms. METHOD: The five-week study consisted of thirty subjects. The first week was considered baseline. All subjects used a control solution (second week), and were randomly divided into the three groups (third week): G1-0.12% chlorhexidine; G2 - 2.5% garlic (Allium sativum, L.); and G3 - 0.2% tea tree oil (Melaleuca alternifolia). Dishes containing blood agar and Mitis Salivarius Bacitracin agar (MSB) were inoculated with the subjects' saliva (collected twice a week). Total microorganisms and mutans streptococci were counted in blood agar and MSB, respectively. RESULTS: Chlorhexidine and garlic groups showed antimicrobial activity against mutans streptococci, but not against other oral microorganisms. The tea tree oil group showed antimicrobial activity against mutans streptococci and other oral microorganisms. Maintenance of reduced levels of microorganisms was observed only for garlic and tea tree oil during the two consecutive weeks (fourth and fifth). Unpleasant taste (chlorhexidine 40%, tea tree oil 30%, garlic 100%), burning sensation (chlorhexidine 40%, tea tree oil 60%, garlic 100%), bad breath (chlorhexidine 40%, tea tree oil 20%, garlic 90%), and nausea (chlorhexidine 0%, tea tree oil 10%, garlic 30%) were reported. CONCLUSION: Garlic and tea tree oil might be an alternative to chlorhexidine.

[Jaccoud's syndrome. A case report]. / Síndrome de Jaccoud. Relato de caso.

The study concerns a 44 year-old male patient with a history of intermittent episodes of rheumatic activity; the physical examination showed mitral valve stenosis and metacarpophalangeal joint deformities. The joint deformities were bilateral and symmetrical, similar to that of rheumatoid arthritis. Nevertheless, the ulnar deviation of the hands was reversible and there were no typical signs of rheumatoid arthritis in the X-ray. In view of the facts, the diagnosis of Jaccoud's Artrhopathy (JA) was made. The pathogenesis, the clinical features and the association between mitral valve stenosis and joints deformities of JA are discussed. This disease has a good prognosis and it is sparsely mentioned in the cardiologic literature. Many patients with JA are diagnosed as having rheumatoid arthritis, which is a chronic and disabling disease.