Rabies Virus-Neutralizing Antibodies in Free-Ranging Invasive Wild Boars (Sus scrofa) from Brazil.

Rabies, one of the most lethal global zoonoses, affects all mammals. It remains circulating worldwide in sylvatic cycles through terrestrial and airborne reservoirs, and in Brazil, bats are currently the main reservoirs and source of transmission. Wild boars, an important invasive alien species in Brazil, are a proven food source for hematophagous bats and may participate in the Brazilian sylvatic cycle of rabies. We evaluated the presence of this pathogen in hunted wild boars from the São Paulo state using histopathology, the direct fluorescent antibody test (DFA), viral isolation in cell culture (VICC), the rapid fluorescent focus inhibition test (RFFIT), and quantitative reverse transcription polymerase chain reaction (RT-qPCR). The results of histopathological, DFA, VICC, and RT-qPCR analysis were negative for all samples; seven serum samples tested positive in the RFFIT, and titers ranged from 0.13 IU/mL to 0.5 IU/mL. The presence of rabies virus-neutralizing antibodies in the studied wild boars suggests the circulation of the virus in these animals. Educative actions directed at hunters should include information on the prevention of this important zoonosis.

Treatment With Methotrexate Associated With Lipid Core Nanoparticles Prevents Aortic Dilation in a Murine Model of Marfan Syndrome.

In Marfan syndrome (MFS), dilation, dissection, and rupture of the aorta occur. Inflammation can be involved in the pathogenicity of aortic defects and can thus be a therapeutic target for MFS. Previously, we showed that the formulation of methotrexate (MTX) associated with lipid nanoparticles (LDE) has potent anti-inflammatory effects without toxicity. To investigate whether LDEMTX treatment can prevent the development of aortic lesions in the MFS murine model. MgΔloxPneo MFS (n = 40) and wild-type (WT, n = 60) mice were allocated to 6 groups weekly injected with IP solutions of: (1) only LDE; (2) commercial MTX; (3) LDEMTX (dose = 1mg/kg) between 3rd and 6th months of life. After 12 weeks of treatments, animals were examined by echocardiography and euthanatized for morphometric and molecular studies. MFS mice treated with LDEMTX showed narrower lumens in the aortic arch, as well as in the ascending and descending aorta. LDEMTX reduced fibrosis and the number of dissections in MFS but not the number of elastic fiber disruptions. In MFS mice, LDEMTX treatment lowered protein expression of pro-inflammatory factors macrophages (CD68), T-lymphocytes (CD3), tumor necrosis factor-α (TNF-α), apoptotic factor cleaved-caspase 3, and type 1 collagen and lowered the protein expression of the transforming growth factor-ß (TGF-ß), extracellular signal-regulated kinases ½ (ERK1/2), and SMAD3. Protein expression of CD68 and CD3 had a positive correlation with an area of aortic lumen (r 2 = 0.36; p < 0.001), suggesting the importance of inflammation in the causative mechanisms of aortic dilation. Enhanced adenosine availability by LDEMTX was suggested by higher aortic expression of an anti-adenosine A2a receptor (A2a) and lower adenosine deaminase expression. Commercial MTX had negligible effects. LDEMTX prevented the development of MFS-associated aortic defects and can thus be a candidate for testing in clinical studies.

Influence of VistaScan image enhancement filters on diagnosis of simulated periapical lesions on intraoral radiographs.

OBJECTIVE:: To evaluate whether image enhancement filters of VistaScan system improve the diagnostic accuracy of simulated periapical lesions. METHODS:: 10 sockets were prepared on bovine ribs to fit a bovine tooth. Bone defects were created and successively enlarged providing four groups (n = 10): Group 0, absence of lesions; Group 1, periapical lesions with 1.6 mm in diameter; Group 2, with 1.8 mm in diameter; and Group 3, with 2.1 mm in diameter. Periapical radiographs were taken using a photostimulable storage phosphor plate and DBSWIN software. VistaScan filters were applied and the images were allocated into seven groups: Nonfiltered, Fine, Caries 1, Caries 2, Endodontic, Periodontal and Noise Reduction. All the 280 images were assessed about the presence or absence of periapical lesions. Pixel intensities standard deviation were compared between nonfiltered and filtered images. Two-Way Analysis of Variance and the post hoc Tukey's test were used to compare area under the ROC curve, sensitivity and specificity. RESULTS:: VistaScan filters showed no significant difference for area under receiver operating characteristic curve (p = 0.124), sensitivity (p = 0.835) and specificity (p = 0.832). Area under receiver operating characteristic curve (p = 0.000) and sensitivity (p = 0.000) in 2.1 mm lesions size were significantly higher than in 1.6 mm and 1.8 mm lesions size. Pixel intensities standard deviation was significantly changed in the filtered images compared to nonfiltered ones (p < 0.01), except for Fine in the bone region (p > 0.05). CONCLUSION:: VistaScan enhancement filters do not influence the diagnostic accuracy of simulated periapical lesions. On the other hand, larger lesions were more frequently detected. The filters change the pixel intensities reducing or intensifying the differences between similar regions.

Anti-inflammatory effects of intravenous methotrexate associated with lipid nanoemulsions on antigen-induced arthritis.

OBJECTIVE: To test the hypothesis that intravenous use of methotrexate associated with lipid nanoemulsions can achieve superior anti-inflammatory effects in the joints of rabbits with antigen-induced arthritis compared with commercial methotrexate. METHODS: Arthritis was induced in New Zealand rabbits sensitized with methylated bovine serum albumin and subsequently intra-articularly injected with the antigen. A nanoemulsion of methotrexate labeled with 3H-cholesteryl ether (4 mg/kg methotrexate) was then intravenously injected into four rabbits to determine the plasma decaying curves and the biodistribution of the methotrexate nanoemulsion by radioactive counting. Additionally, the pharmacokinetics of the methotrexate nanoemulsion were determined by high-pressure liquid chromatography. Twenty-four hours after arthritis induction, the animals were allocated into three groups, with intravenous injection with saline solution (n=9), methotrexate nanoemulsion (0.5 µmol/kg methotrexate, n=7), or commercial methotrexate (0.5 µmol/kg, n=4). The rabbits were sacrificed 24 h afterward. Synovial fluid was then collected for protein leakage and cell content analyses and synovial membranes were collected for histopathological analysis. RESULTS: The methotrexate nanoemulsion was taken up mainly by the liver and the uptake by arthritic joints was two-fold greater than that by control joints. The methotrexate nanoemulsion treatment reduced leukocyte influx into the synovial fluid by nearly 65%; in particular, mononuclear and polymorphonuclear cells were reduced by 47 and 72%, respectively. In contrast, cell influx was unaffected following treatment with commercial methotrexate. Protein leakage into the arthritic knees of the rabbits was also more limited following methotrexate nanoemulsion treatment than following commercial methotrexate treatment. CONCLUSIONS: The intravenous methotrexate nanoemulsion showed anti-inflammatory effects on the synovia of arthritic joints that were clearly superior to the effects of a commercial methotrexate preparation. This result is conceivably due to greater methotrexate uptake by the joints when the drug is associated with a nanoemulsion.

Endotoxemic myocardial dysfunction: subendocardial collagen deposition related to coronary driving pressure.

Sepsis impairs the autoregulation of myocardial microcirculatory blood flow, but whether this impairment is correlated with myocardial remodeling is unknown. This study investigated the role of coronary driving pressure (CDP) as a determinant of microcirculatory blood flow and myocardial fibrosis in endotoxemia and sepsis. The study is composed of two parts: a prospective experimental study and an observational clinical study. The experimental study was performed on male Wistar rats weighing 300 to 320 g. Endotoxemia was induced in rats by lipopolysaccharide (LPS) injection (10 mg·kg intraperitoneally). Hemodynamic evaluation was performed 1.5 to 24 h after LPS injection by measuring the mean arterial pressure, CDP, left ventricular end-diastolic pressure, dP/dtmax, and dP/dtmin. Microspheres were also infused into the left ventricle to measure myocardial blood flow, and myocardial tissue was histologically assessed to analyze collagen deposition. The CDP, mean arterial pressure, and myocardial blood flow were reduced by 55%, 30%, and 70%, respectively, in rats 1.5 h after LPS injection compared with phosphate buffer saline injection (P < 0.05). The CDP was significantly correlated with subendocardial blood flow (r = 0.73) and fibrosis (r = 0.8). Left ventricular function was significantly impaired in the LPS-treated rats, as demonstrated by dP/dtmax (6,155 ± 455 vs. 3,746 ± 406 mmHg·s, baseline vs. LPS; P < 0.05) and dP/dtmin (-5,858 ± 236 vs. -3,516 ± 436 mmHg·s, baseline vs. LPS; P < 0.05). The clinical study was performed on 28 patients with septic shock analyzed for CDP. The CDP data and histological slices were collected from septic patients. In addition, the clinical data demonstrated fibrosis and 45% CDP reduction in nonsurvivors compared with survivors. In conclusion, the left ventricular subendocardial blood flow was positively correlated with CDP, and higher CDP was negatively correlated with myocardial collagen deposition. Thus, early reductions in myocardial blood flow and CDP facilitate late myocardial fibrosis in rats and likely in humans.

Role of focal adhesion kinase in lung remodeling of endotoxemic rats.

Despite significant advances in the care of critically ill patients, acute lung injury continues to be a complex problem with high mortality. The present study was designed to characterize early lipopolysaccharide (LPS)-induced pulmonary injury and small interfering RNA targeting focal adhesion kinase (FAK) as a possible therapeutic tool in the septic lung remodeling process. Male Wistar rats were assigned into endotoxemic group and control group. Total collagen deposition was performed 8, 16, and 24 h after LPS injection. Focal adhesion kinase expression, interstitial and vascular collagen deposition, and pulmonary mechanics were analyzed at 24 h. Intravenous injection of small interfering RNA targeting FAK was used to silence expression of the kinase in pulmonary tissue. Focal adhesion kinase, total collagen deposition, and pulmonary mechanics showed increased in LPS group. Types I, III, and V collagen showed increase in pulmonary parenchyma, but only type V increased in vessels 24 h after LPS injection. Focal adhesion kinase silencing prevented lung remodeling in pulmonary parenchyma at 24 h. In conclusion, LPS induced a precocious and important lung remodeling. There was fibrotic response in the lung characterized by increased amount in total and specific-type collagen. These data may explain the frequent clinical presentation during sepsis of reduced lung compliance, oxygen diffusion, and pulmonary hypertension. The fact that FAK silencing was protective against lung collagen deposition underscores the therapeutic potential of FAK targeting by small interfering RNA.

MEF2C silencing attenuates load-induced left ventricular hypertrophy by modulating mTOR/S6K pathway in mice.

BACKGROUND: The activation of the members of the myocyte enhancer factor-2 family (MEF2A, B, C and D) of transcription factors promotes cardiac hypertrophy and failure. However, the role of its individual components in the pathogenesis of cardiac hypertrophy remains unclear. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated whether MEF2C plays a role in mediating the left ventricular hypertrophy by pressure overload in mice. The knockdown of myocardial MEF2C induced by specific small interfering RNA (siRNA) has been shown to attenuate hypertrophy, interstitial fibrosis and the rise of ANP levels in aortic banded mice. We detected that the depletion of MEF2C also results in lowered levels of both PGC-1alpha and mitochondrial DNA in the overloaded left ventricle, associated with enhanced AMP:ATP ratio. Additionally, MEF2C depletion was accompanied by defective activation of S6K in response to pressure overload. Treatment with the amino acid leucine stimulated S6K and suppressed the attenuation of left ventricular hypertrophy and fibrosis in the aforementioned aortic banded mice. CONCLUSION/SIGNIFICANCE: These findings represent new evidences that MEF2C depletion attenuates the hypertrophic responses to mechanical stress and highlight the potential of MEF2C to be a target for new therapies to cardiac hypertrophy and failure.

A queda da pressão de perfusão coronariana está associada com remodelamento subendocárdico e disfunção ventricular esquerda na fístula aorto-cava / Low coronary driving pressure is associated with subendocardial remodeling and left ventricular disfunction in aortocaval

Os objetivos deste estudo foram investigar o papel da pressão de perfusão coronariana (PPC) no remodelamento subendocárdico (SE) e os possíveis efeitos sobre a função cardíaca em um modelo de fístula aorto-cava (FAC). Ratos Wistar, foram submetidos ao modelo de FAC ou à cirurgia fictícia (SH) com seguimento de 8 semanas. Comparado ao grupo SH, FAC apresentou PPC, +dP/dt, –dP/dt menores. O fluxo miocárdico foi menor em SE. As metaloproteinases-2 (MMP-2), os níveis de IL-1 e IL-6 e a fibrose apresentaram maiores em SE. A PPC está associada com a fibrose SE, com a +dP/dt e com a –dP/dt. Na fístula, a queda da PPC está associada a dano em SE, representado por isquemia, estresse oxidativo, aumento citocinas e de MMP-2, com desenvolvimento de fibrose. O remodelamento de SE interfere negativamente na função do ventrículo esquerdo...

The aim of this study was to investigate the role of coronary driving pressure (CDP) in the development of subendocardial (SE) remodeling and the conceivable effects on cardiac function, using a rat model of aortocaval fistula (ACF). Wistar rats were submitted to ACF or sham (SH) operations with follow-up of 8 weeks. Compared with SH, ACF showed lower CDP, +dP/dt and –dP/dt. ACF showed lower myocardial blood flow in SE. Metalloproteiase-2 (MMP-2), levels of IL-6 and IL-1 and fibrosis were predominated in SE of ACF. CDP was associated with SE fibrosis, +dP/dt and –dP/dt. Low CDP early in the course of ACF is associated with SE damage characterized by ischemia, oxidative stress, increase in cytokines and MMP-2, the development of fibrosis and by this mechanism interferes negatively in left ventricular function...

Low coronary perfusion pressure is associated with endocardial fibrosis in a rat model of volume overload cardiac hypertrophy.

UNLABELLED: Left ventricular hypertrophy following volume overload is regarded as an example of cardiac remodeling without increased fibrosis accumulation. However, infarction is associated with increased fibrosis within the noninfarcted, hypertrophied myocardium, particularly in the subendocardial regions. It is conceivable to suppose that, as also occurs postinfarction, low coronary driving pressure may also interfere with accumulation of myocardial fibrosis following aortocaval fistula. PURPOSE: To investigate the role of acute hemodynamic changes in subsequent deposition of cardiac fibrosis in response to aortocaval fistula. METHOD: Aortocaval fistula were created in 4 groups of Wistar rats that were followed over 4 and 8 weeks: aortocaval fistula 4 and aortocaval fistula 8 (10 rats each) and their respective controls (sham-operated controls - Sh), Sh4 and Sh8 (8 rats each). Hemodynamic measurements were performed 1 week after surgery. Hypertrophy and fibrosis were quantified by myocyte diameter and collagen volume fraction at the end of follow up. RESULT: Compared with Sh4 and Sh8, pulse pressure, left ventricular end-diastolic pressure, and +dP/dt were higher in aortocaval fistula 4 and aortocaval fistula 8, but -dP/dt was similar. Coronary driving pressure (mm Hg), used as an estimate of perfusion pressure, was lower in aortocaval fistula 8 (52.6 +/- 4.1) than in Sh8 (100.8 +/- 1.3), but comparable between aortocaval fistula 4 (50.0 +/- 8.9) and Sh4 (84.8 +/- 2.3). Myocyte diameter was greater in aortocaval fistula 8, whereas interstitial and subendocardial fibrosis were greater in aortocaval fistula 4 and aortocaval fistula 8. Coronary driving pressure correlated inversely and independently with subendocardial fibrosis (r(2) = .86, P <.001), whereas left ventricular systolic pressure (r(2) = 0.73, P = .004) and end-diastolic pressure (r(2) = 0.55, P = 012) correlated positively and independently with interstitial fibrosis. CONCLUSION: Coronary driving pressure falls and ventricular pressures increase early after aortocaval fistula and are associated with subsequent myocardial fibrosis deposition.

Low coronary perfusion pressure is associated with endocardial fibrosis in a rat model of volume overload cardiac hypertrophy

No remodelamento que se segue às sobrecargas de volume não é descrito o aumento de fibrose miocárdica. Após o infarto, entretanto, há hipertrofia do miocárdio remoto com acúmulo de fibrose, particularmente no subendocárdio. Na fístula aorto-cava, tal como no infarto, é possível que a queda da pressão de perfusão coronariana interfira com a fibrose cardíaca. OBJETIVO: Investigar o papel das mudanças hemodinâmicas agudas sobre a fibrose cardíaca na fístula aorto-cava. MÉTODO: Ratos Wistar submetidos a fístula aorto-cava, seguidos por 4 e 8 semanas, constituíram 4 grupos, fístula aorto-cava 4 e fístula aorto-cava 8 (10 ratos cada) e seus respectivos controles (sham-operated controls - Sh), Sh4 e Sh8 (8 ratos cada). A hemodinâmica foi realizada 1 semana após a cirurgia. A hipertrofia e a fibrose foram quantificadas ao final do seguimento pelo diâmetro dos miócitos e pela fração de volume do colágeno. RESULTADOS: Comparados com Sh4 e Sh8, a pressão de pulso, a pressão diastólica final do ventrículo esquerdo e a +dP/dt foram maiores em fístula aorto-cava 4 e fístula aorto-cava 8, enquanto a -dP/dt foi similar. A pressão estimada da perfusão coronariana (mmHg) foi menor em fístula aorto-cava 8 (52,6±4,1) do que em Sh8 (100,8±1,3), mas comparável entre fístula aorto-cava 4 (50,0±8,9) e Sh4 (84,8±2,3). O diâmetro dos miócitos foi maior em fístula aorto-cava 8 e a fibrose intersticial e subendocárdica maior em fístula aorto-cava 4 e fístula aorto-cava 8. Houve correlação inversa e independente da pressão de perfusão coronariana com a fibrose subendocárdica (r2=0,86; p<0,0001) e das pressões sistólica (r2=0,73; p=0,0035) e diastólica final do ventrículo esquerdo (r2=0,55; p=0.0124) com a fibrose intersticial. CONCLUSÃO: A queda precoce da pressão de perfusão coronariana e o aumento das pressões ventriculares após a fístula aorto-cava associam-se com fibrose miocárdica subseqüente.