Is the Wistar Rat a more Suitable Normotensive Control for SHR to Test Blood Pressure and Cardiac Structure and Function?

Abstract Background: There are divergences in the literature regarding the experimental model (Wistar-WIS or Wistar Kyoto-WKY) to be used as a Spontaneously Hypertensive Rat (SHR) control. The characterization of these models in terms of cardiovascular parameters provides researchers with important tools at the time of selection and application in scientific research. Objective: The aim of this study was to evaluate the use of WIS and WKY as a Spontaneously Hypertensive Rat (SHR) control by assessing the long-term behavior of blood pressure and cardiac structure and function in these strains. Methods: To this end, WIS, WKY, and SHR underwent longitudinal experiments. Blood pressure and body mass were measured every two weeks from the 8th to the 72nd. Echocardiographic analysis was performed in all groups with 16, 48, and 72 weeks of life. After having applied the normality test, the Two-Way ANOVA of repeated measures followed by the Tukey post hoc test was used. A significance level of 5% was established. Results: The WIS group showed higher body mass (p<0.05), while the WKY and SHR presented higher body mass variation over time (p<0.05). SHR exhibited increased values of systolic, diastolic, and mean blood pressure when compared to WKY and WIS, whereas the WKY generally showed higher values than WIS (p<0.05). Regarding the cardiac function, SHR showed reduced values, while the WKY presented an early decrease when compared to WIS with aging (p<0.05). Conclusion: WIS is a more suitable normotensive control for SHR than WKY in experiments to test blood pressure and cardiac structure and function.

Core temperature circadian rhythm across aging in Spontaneously Hypertensive Rats.

The purpose of this study was to evaluate the circadian rhythm of core temperature (Tcore) across aging in Spontaneously Hypertensive Rats (SHR) with comparison to the two rat strains often used as their normotensive control animals, namely, Wistar (WIS) and Wistar Kyoto (WKY). METHODS: WIS, WKY and SHR rats were subdivided into three different groups according their age: WIS16, WIS48, WIS72, WKY16, WKY48, WKY72, SHR16, SHR48 and SHR72 weeks-old. Body mass and blood pressure were periodically measured along the experiments. All animal group had their circadian rhythm of Tcore evaluated over three consecutive days (72 h) by telemetry using an implanted temperature sensor. The Tcore circadian rhythm was averaged in 1-h blocks and analyzed using the cosinor method. RESULTS: Sixteen-week-old SHR (SHR16) presented higher Tcore than WIS16 (from 06am to 06pm) and WKY16 (from 07am to 06pm). Both normotensive groups exhibited increases in Tcore during circadian rhythm with aging. The cosinor analysis showed no differences between strains and ages for the acrophase. An age effect on the SHR strain (SHR16 < SHR72) was observed regarding the amplitude. SHR16 had higher values regarding MESOR compared to WIS16 and WKY16. In addition, WIS72 and WKY72 showed higher values than WIS16 and WKY16, respectively. Finally, no differences were observed in the strength rhythm analysis. CONCLUSIONS: SHR presented impaired thermoregulatory control at only 16 weeks of age when showing a higher body temperature during the activity phase, while other circadian rhythm parameters showed no differences across aging. Therefore, in taking our results as a whole we can conclude that WIS and WKY are appropriate Wistar strains to be used as normotensive controls for SHR.

Effects of chia (Salvia hispanica L.) on oxidative stress and inflammation in ovariectomized adult female Wistar rats.

The present study investigated the influence of chia consumption on inflammation, oxidative stress, and lipid profiles in adult female ovariectomized rats fed a high-fat diet. Forty ovariectomized and 40 intact (SHAM) rats were allocated into 8 groups (n = 10), and each rat received one of the following four diets: standard diet (ST); standard diet + chia (STC); high-fat diet (HF); and high-fat diet + chia (HFC) for 126 days. Biochemical parameters and biomarkers of lipid peroxidation, inflammation, and oxidative stress were evaluated. The mRNA expression levels of PPAR-α, NFκB, TNF-α and Zn-SOD1 were analyzed, as well as those of TNF-α and IL-1ß. Chia intake increased HDL cholesterol (HDL-c) and reduced LDL cholesterol (LDL-c) levels. Plasma catalase activity was elevated in the STC group. Concentrations of TBARS were higher in all groups fed HF. PPAR-α mRNA expression was elevated, and levels of NFκB mRNA expression were reduced in the STC group. mRNA expression and protein levels of TNF-α were lower in rats fed the standard diet. Protein levels of IL-1ß were reduced in rats fed the standard diet, and the high fat diet with chia. In general, ovariectomy did not influence the inflammatory and oxidative stress parameters. Chia intake improved antioxidant activity by increasing SOD expression, PPAR-α expression, catalase activity, and HDL-c levels. In addition, chia consumption decreased the concentrations of the inflammatory markers IL-1ß and LDL-c.

Does aerobic exercise associated with tryptophan supplementation attenuates hyperalgesia and inflammation in female rats with experimental fibromyalgia?

The objective of this study was to verify the effects of aerobic exercise associated with tryptophan (TRP) supplementation on hyperalgesia, as well as on cortisol, IL-6 and TNF concentrations in female rats with experimental fibromyalgia (FM). Female Wistar rats (initial body weight: ~ 350 g; age: 12 months) were randomly divided into 5 groups: CON (Control); F (Fibromyalgia induced); FE (Fibromyalgia induced plus exercise); FES (Fibromyalgia induced plus exercise and TRP supplementation) and FS (Fibromyalgia induced plus TRP supplementation). Fibromyalgia was induced with two injections (20 µL) of acidic saline (pH 4.0) into the right gastrocnemius muscle with a 3-day interval. Control animals received the same doses of neutral saline (pH 7.4). The exercised animals underwent progressive low-intensity aerobic exercise (LIAE) on a treadmill (10-12 m/min, 30-45 min/day, 5 days/week) for three weeks. During this period, the supplemented animals received a TRP supplemented diet (210 g/week), while the others received a control diet. Mechanical hyperalgesia was evaluated weekly and serum cortisol and muscle IL-6 and TNF concentrations were assessed after three weeks of interventions. Experimental FM caused bilateral hind paw hyperalgesia and augmented serum cortisol and muscle IL-6 concentrations. After 3 weeks of interventions, LIAE alone reduced hyperalgesia (151%) and reduced serum cortisol concentrations (72%). Tryptophan supplementation itself diminished hyperalgesia (57%) and reduced serum cortisol concentrations (67%). Adding TRP supplementation to LIAE did not further reduce hyperalgesia significantly (11%), which was followed by an important decrease in muscle IL-6 concentrations (68%), though reduction in serum cortisol pulled back to 45%. Muscle TNF concentrations were not affected. In conclusion, the association of TRP supplementation to LIAE does not potentiate significantly the reduction of bilateral mechanical hyperalgesia promoted by LIAE in female rats with experimental FM, however an important decrease in IL-6 is evident.

Meniscal allograft transplantation in rabbit.

This study evaluated the technique for meniscal allograft transplantation using allografts preserved in glycerin 98% in rabbits. Euthanasia was performed at 70 days to compare the transplanted (TM1 to TM16) versus the contralateral meniscus (OM1 to OM16). Sixteen menisci, 8 transplanted and 8 contralateral, were submitted to gross examination, histomorphometric analysis for identification and quantification of cellular type, and for quantification and distribution of collagen fibers. A revascularization study was conducted in all of the other samples. Lengths of the OM varied from 0.9 to 1.0 cm and two TM were smaller. All TM were completely attached to the synovial membrane, except for one case that presented partial fixation. Both, TM and OM had similar amounts of chondrocytes, fibroblasts and fibrocytes, and at the horns, chondrocytes were predominant. The collagen fibers in TM were well organized throughout the body, and disorganized at the horns. These fibers in OM were organized. The amounts of collagen type I and III, and the vascularization of the perimeniscal tissue and of the edge were similar in OM and TM. These results demonstrated graft integration and thus this transplantation technique and preservation method may be recommended.

Acute and chronic observations of complete atrioventricular block in rats.

The mechanisms of production, and gross, microscopic and electrocardiograhic findings of surgically-induced complete heart block (CHB) in the adult rat are presented. This is an effective in vivo model for establishing alternative methods to electronic pacemakers and for providing detailed information aimed at replacement, reduction and refinement of the technique. Sternal thoracotomy was employed to identify the epicardial fat pad by the aortic root, used as a landmark for cauterization of the atrioventricular (AV) node. Stable CHB was produced in 60 rats with a 70% survival rate. The best survival rate was observed in 8-week-old animals weighing 221 ± 27.6 g. Heart rate before cauterization was 387 ± 55 bpm, reduced after cauterization to 126 ± 40 bpm in the survival and to 65 ± 19 bpm in the non-survival groups. At 30 days findings were: elevated left ventricular end-diastolic pressure (21 ± 5.4 mmHg, P < 0.05); maximal rate of rise of left ventricular pressure (LVP) during isovolumetric contraction (2192 ± 235 mmHg/s, P < 0.05); maximal rate of decrease of LVP (-1658 ± 191 mmHg/s, P < 0.05); isovolumetric relaxation constant (5.7 ± 0.8 ms, P < 0.05) with wet-to-dry lung-weight ratio (78.1 ± 0.4, P < 0.05); heart weight/body weight (0.6 ± 0.1, P < 0.05); heart volume (1.8 ± 0.3 mL, P < 0.05); longitudinal diameter (20.2 ± 1.91 mm, P < 0.05); and transversal diameter (17.0 ± 1.4 mm, P < 0.05) with supported dilated cardiomyopathy which culminated in chronic heart failure. CHB hearts had increased preload and replacement of myofibrils by collagen. CHB was achieved reproducibly by cauterization of the rat AV node and/or His bundle. This led to electrophysiological, hemodynamic, and structural remodeling, and could be useful in long-term cardiac remodeling assessments and potential therapy development.

Diferenciação de células-tronco mesenquimais derivadas do tecido adiposo em cardiomiócitos / Differentiation of adipose tissue-derived mesenchymal stem cells into cardiomyocytes

FUNDAMENTO: O potencial de renovação e proliferação dos cardiomiócitos, in vivo, é pequeno, e por isso, o músculo cardíaco apresenta limitada capacidade de repor células perdidas. Na tentativa de minimizar os danos oriundos de lesões hipóxico-isquêmicas e daquelas que acometem o sistema de condução do coração, a terapia celular com células-tronco mesenquimais (MSC) vem sendo utilizada, inclusive com cardiomiócitos diferenciados a partir de MSC. OBJETIVO: O presente trabalho comparou três protocolos distintos de indução de diferenciação objetivando a sugestão de um método viável para a diferenciação de maior número de células funcionais que expressem fenótipo cardiomiogênico. MÉTODOS: Culturas de MSC obtidas de tecido adiposo de ratos jovens da linhagem Lewis transgênicos para proteína verde fluorescente (GFP) foram submetidos a três diferentes meios de diferenciação cardiogênica: Planat-Bérnard, 5-azacitidina e meio Planat-Bérnard + 5-azacitidina e observadas quanto a expressão de marcadores celulares cardíacos. RESULTADOS: Nos três protolocos utilizados observou-se formação da proteína alfa-actinina sarcomérica no citoesqueleto das células submetidas à diferenciação, expressão de conexina 43 na membrana nuclear e citoplasmática e formação de gap junctions, necessárias para a propagação do impulso elétrico no miocárdio, contudo, em nenhum protocolo foi observada contração espontânea das células submetidas à diferenciação cardiogênica. CONCLUSÃO: A indução com 5-azacitidina proporcionou diferenciação celular cadiomiogênica efetiva e similar à encontrada com o meio Planat-Bénard e, por ser um protocolo mais simples, rápido e com menor custo torna-se o método de eleição.

BACKGROUND: Cardiomyocytes have small potential for renovation and proliferation in vivo. Consequently, the heart muscle has limited capacity of self-renewal. Mesenchymal stem cells (MSC) therapy, as well as MSC differentiated into cardiomyocytes, has been used in the attempt to minimize the effects of ischemic-hypoxic lesions and those affecting the electrical conduction system of the heart. OBJECTIVE: The present study compared three distinct protocols for induced differentiation of MSC into cardiomyocytes aimed at finding a viable method for producing a large number of functional cells expressing cardiomyogenic phenotype. METHODS: Mesenchymal stem cells were obtained from the adipose tissue of young transgenic Lewis rats expressing green fluorescent protein (GFP), and submitted to three distinct differentiation-inducing media: 1) Planat-Bérnard, 2) 5-azacytidine, and 3) Planat-Bérnard + 5-azacytidine; further, these cells were identified based on the expression of cardiac cell markers. RESULTS: All three protocols detected the expression of sarcomeric-alpha-actinin protein in the exoskeleton of cells, expression of connexin-43 in the nuclear and cytoplasmic membrane, and formation of gap junctions, which are necessary for electrical impulse propagation in the myocardium. However, no spontaneous cell contraction was observed with any of the tested protocols. CONCLUSION: Induction with 5-azacytidine provided an effective cadiomyogenic cellular differentiation similar to that obtained with Planat-Bénard media. Therefore, 5-azacytidine was the method of choice for being the simplest, fastest and lowest-cost protocol for cell differentiation.

Differentiation of adipose tissue-derived mesenchymal stem cells into cardiomyocytes.

BACKGROUND: Cardiomyocytes have small potential for renovation and proliferation in vivo. Consequently, the heart muscle has limited capacity of self-renewal. Mesenchymal stem cells (MSC) therapy, as well as MSC differentiated into cardiomyocytes, has been used in the attempt to minimize the effects of ischemic-hypoxic lesions and those affecting the electrical conduction system of the heart. OBJECTIVE: The present study compared three distinct protocols for induced differentiation of MSC into cardiomyocytes aimed at finding a viable method for producing a large number of functional cells expressing cardiomyogenic phenotype. METHODS: Mesenchymal stem cells were obtained from the adipose tissue of young transgenic Lewis rats expressing green fluorescent protein (GFP), and submitted to three distinct differentiation-inducing media: 1) Planat-Bérnard, 2) 5-azacytidine, and 3) Planat-Bérnard + 5-azacytidine; further, these cells were identified based on the expression of cardiac cell markers. RESULTS: All three protocols detected the expression of sarcomeric-alpha-actinin protein in the exoskeleton of cells, expression of connexin-43 in the nuclear and cytoplasmic membrane, and formation of gap junctions, which are necessary for electrical impulse propagation in the myocardium. However, no spontaneous cell contraction was observed with any of the tested protocols. CONCLUSION: Induction with 5-azacytidine provided an effective cadiomyogenic cellular differentiation similar to that obtained with Planat-Bénard media. Therefore, 5-azacytidine was the method of choice for being the simplest, fastest and lowest-cost protocol for cell differentiation.

Processo de reparação de lesões da córnea e a membrana amniótica na oftalmologia / Repair process of corneal damage and the amniotic membrane in ophthalmology

Os eventos que fazem parte do processo de reparação de lesões da córnea ocorrem simultaneamente e envolvem proliferação, migração, diferenciação e apoptose celular, além da comunicação intercelular. Vários fatores solúveis, além de proteínas da matriz mesenquimal, proteoglicanos, enzimas proteolíticas e alguns tipos celulares são abordados nesta revisão, na qual explicam-se os processos de reparação de lesões superficiais ou penetrantes da córnea. A membrana amniótica, muito utilizada na cirurgia oftálmica, foi estudada por apresentar funções que colaboram com o processo de reparação. Entretanto, tais funções poderão ser perdidas quando tal tecido for submetido à conservação. Assim, torna-se importante conhecer o processo de reparação de lesões que envolvem, ou não, a córnea em toda a sua espessura e escolher a melhor forma de utilização da membrana amniótica quando ela for indicada na terapia para estas lesões.

The events included in the process of repair of corneal damage occur simultaneously and involve proliferation, migration, differentiation, cell apoptosis and intercellular communication. Several soluble factors, mesenchymal matrix proteins, proteoglycans, proteolytic enzymes and some cell types are covered in this review, which explains the processes of repair of corneal wounds, either superficial or penetrating. The amniotic membrane, used in ophthalmic surgery, was studied because of the contribution of its functions to the repair process. However, these functions may be lost when the amniotic membrane is subjected to conservation. Therefore, it is important to understand the repair process of lesions involving or not the entire thickness of the cornea, and choose the best use of the amniotic membrane, when it is indicated for the treatment of these lesions.

Ambulatory esophageal pHmetry in healthy dogs with and without the influence of general anesthesia.

This research aimed to determine the value of esophageal pH in awake and anesthetized dogs, to evaluate the esophageal pH value in awake dogs, in different body positions, as well as to study the occurrence of gastroesophageal reflux episodes in these positions. Thus, 40 healthy male and female adult dogs with mean body weight of 15.5 ± 4.6 kg were used. Esophageal pHmetry was conducted by inserting a catheter through the oropharynx in 30 dogs (stage 1) anesthetized with acepromazine, propofol and isoflurane, submitted to elective ovariosalpingohysterectomy. In addition, 8-h esophageal pHmetry was carried out transnasally in 10 awake dogs (stage 2), allowed to move and change body positions (lateral and sternal decubitus, and standing position), which were recorded. The mean esophageal pH value was lower (p < 0.01) in the anesthetized dogs (7.3 ± 0.82) than in the awake dogs (8.2 ± 0.3). Only four anesthetized dogs (13.33%) suffered reflux episodes. Reflux was not observed in the awake dogs and no esophageal pH differences were found between the body positions studied. Compared to the alert state, general anesthesia in dogs submitted to the previously mentioned anesthesia protocol causes esophageal pH reduction and predisposes to the occurrence of gastroesophageal reflux episodes. Transnasal pHmetry of 8 h in healthy awake dogs reveals that the esophageal pH value is alkaline and does not vary according to body position. In these animals, decubitus position is not a determining factor for reflux episodes to occur.