Coupling Between Respiratory and Sympathetic Activities as a Novel Mechanism Underpinning Neurogenic Hypertension

Enhanced sympathetic outflow to the heart and resistance vessels greatly contributes to the onset and maintenance of neurogenic hypertension. There is a consensus that the development of hypertension (clinical and experimental) is associated with an impairment of sympathetic reflex control by arterial baroreceptors. More recently, chronic peripheral chemoreflex activation, as observed in obstructive sleep apnea, has been proposed as another important risk factor for hypertension. In this review, we present and discuss recent experimental evidence showing that changes in the respiratory pattern, elicited by chronic intermittent hypoxia, play a key role in increasing sympathetic activity and arterial pressure in rats. This concept parallels results observed in other models of neurogenic hypertension, such as spontaneously hypertensive rats and rats with angiotensin II-salt-induced hypertension, pointing out alterations in the central coupling of respiratory and sympathetic activities as a novel mechanism underlying the development of neurogenic hypertension.

Lineage-Negative Bone Marrow Cells Protect Against Chronic Renal Failure

Progressive renal failure continues to be a challenge. The use of bone marrow cells represents a means of meeting that challenge. We used lineage-negative (Lin(-)) cells to test the hypothesis that Lin(-) cell treatment decreases renal injury. Syngeneic Fischer 344 rats were divided into four groups: sham ( laparotomy only, untreated); Nx (five-sixth nephrectomy and untreated); NxLC1 (five-sixth nephrectomy and receiving 2 x 10(6) Lin(-) cells on postnephrectomy day 15); and NxLC3 (five-sixth nephrectomy and receiving 2 x 10(6) Lin(-) cells on postnephrectomy days 15, 30, and 45). On postoperative day 16, renal mRNA expression of interleukin (IL)-1 beta, tumor necrosis factor-alpha, and IL-6 was lower in NxLC rats than in Nx rats. On postnephrectomy day 60, NxLC rats presented less proteinuria, glomerulosclerosis, anemia, renal infiltration of immune cells, and protein expression of monocyte chemoattractant protein-1, as well as decreased interstitial area. Immunostaining for proliferating cell nuclear antigen showed that, in comparison with sham rats, Nx rats presented greater cell proliferation, whereas NxLC1 rats and NxLC3 rats presented less cell proliferation than did Nx rats. Protein expression of the cyclin-dependent kinase inhibitor p21 and of vascular endothelial growth factor increased after nephrectomy and decreased after Lin(-) cell treatment. On postnephrectomy day 120, renal function (inulin clearance) was significantly better in Lin(-) cell-treated rats than in untreated rats. Lin(-) cell treatment significantly improved survival. These data suggest that Lin(-) cell treatment protects against chronic renal failure. STEM CELLS 2009; 27: 682-692

Efeitos das formulações nanoestruturadas de doxorrubicina e cisplatina em dispersão de óxido de grafeno reduzido no tratamento da progressão do câncer de bexiga Não-músculo invasivo

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Development and characterization of a rodent model of immune-mediated cholangitis.

The cholangiopathies are a group of hepatobiliary diseases in which intrahepatic bile duct epithelial cells, or cholangiocytes, are the target for a variety of destructive processes, including immune-mediated damage. We tested the hypothesis that cholangitis could be induced in rodents by immunization with highly purified cholangiocytes. Inbred Wistar rats were immunized with purified hyperplastic cholangiocytes isolated after bile duct ligation from either syngeneic Wistar or allogeneic Fischer 344 rats; control rats were immunized with bovine serum albumin (BSA) or hepatocytes. After immunization with cholangiocytes, recipient animals developed histologic evidence of nonsuppurative cholangitis without inflammation in other organs; groups immunized with BSA or hepatocytes showed no cholangitis. Immunohistochemical studies revealed that portal tract infiltrates around bile ducts consisted of CD3-positive lymphocytes, some of which expressed major histocompatibility complex class II antigen; B cells and exogenous monocytes/macrophages were essentially absent. Transfer of unfractionated ConA-stimulated spleen cells from cholangiocyte-immunized (but not BSA-immunized) rats into recipients also caused nonsuppurative cholangitis. Moreover, these splenocytes from cholangiocyte-immunized (but not BSA-immunized) rats were cytotoxic in vitro for cultured rodent cholangiocytes; no cytotoxicity was observed against a rat hepatocyte cell line. Also, a specific antibody response in sera of cholangiocyte-immunized rats was demonstrated by immunoblots against cholangiocyte proteins. Finally, cholangiograms in cholangiocyte-immunized rats showed distortion and tortuosity of the entire intrahepatic biliary ductal system. This unique rodent model of experimental cholangitis demonstrates the importance of immune mechanisms in the pathogenesis of cholangitis and will prove useful in exploring the mechanisms by which the immune system targets and damages cholangiocytes.

Thymic lymphomas in Wistar rats exposed to N-methyl-N-nitrosourea (MNU)

The Brazilian Agency for the Environment (IBAMA) recently adopted an alternative medium-term multiple-organ assay system with the Wistar rat strain for detection of the carcinogenic potential of pesticides. Originally, this initiation-promotion protocol was established in Japan with the isogenic Fischer 344 male rat. Among the initiating agents used in that assay, N-methyl-N-nitrosourea (MNU) rapidly induces malignant lymphoma and leukemia and early mortality of rats from different strains. This study was developed to evaluate whether the outbred Wistar rats are also similarly susceptible to MNU. Particularly, it aimed to evaluate the dose-response relationship and to register the MNUinduced pre-neoplasia and neoplasia that may develop in the lympho-hematopoietic system (LHS) of the Wistar rat within a medium-term period. Four groups of male Wistar rats were treated during 2 weeks with vehicle or with MNU (80, 160 or 240 mg/kg body weight, i.p.). After sacrifice at the 12th and 20th weeks, the thymus, spleen, bone marrow, cervical and mesenteric lymph nodes and liver were collected for analysis. At the 20th week, LHS malignant tumors and benign vascular tumors occurred only in the high- and intermediate-dose MNU-treated animals. Four animals treated with 240 mg/kg developed diffuse thymic lymphomas; two others, treated respectively with 240 mg/kg and 160 mg/kg, developed spleen hemangiomas. The present observations indicate that the Wistar strain is as susceptible as other strains to the early development of MNU-induced LHS (pre)neoplasia. Therefore, this strain seems suitable to be used as test system in bioassay protocols that adopt MNU as an initiating agent for carcinogenesis.

Expression of neuromuscular junction messenger RNAs and protein in aged rats

The loss of skeletal muscle mass is believed to be the dominant reason for reduced strength in aging humans. The purpose of this investigation was to gain some information as to why skeletal muscles lose mass as we age. Since nervous system innervation is essential for skeletal muscle fiber viability, incomplete regional reinnervation during normal synaptic junction turnover has been hypothesized to result in selective muscle fiber loss. Examined here was the age-related association in skeletal muscle between atrophy and the expression of mRNAs encoding the γ- and ϵ-subunits of the nicotinic acetylcholine receptor, myogenin, and muscle specific receptor kinase (MuSK). Gastrocnemius and biceps brachii muscles were collected from young (2 month), adult (18 month), and old (31 month) Fischer 344 cross brown Norway F 1 male rats. In the gastrocnemius, muscles of old vs. young and adult rats, lower muscle mass was accompanied by significantly elevated acetylcholine receptor γ-subunit, myogenin, and MuSK mRNA levels. In contrast, the biceps brachii muscle in the same animals exhibited neither atrophy nor a change in acetylcholine receptor γ-subunit, myogenin, or MuSK mRNA levels. Expression of the acetylcholine receptor ϵ-subunit mRNA did not change with age in either gastrocnemius or biceps brachii muscles. Since acetylcholine receptor γ-subunit, myogenin, and MuSK mRNA levels are upregulated in surgically denervated skeletal muscles of young rats while expression of the acetylcholine receptor ϵ-subunit does not change, the findings of the current investigation suggest that a select fiber population within atrophied skeletal muscles of old rats may be in a denervated-like state. I speculate that increases in γ-subunit, myogenin, and MuSK mRNA levels in atrophied muscles of old rats are compensatory responses to nerve terminal retraction. Indeed, a prolongation of denervation in these muscle fibers would subsequently result in their atrophy and death, ultimately leading to a decline in the number of force generating elements present in the muscle. ^

Morphology and morphometry of the vagus nerve in male and female spontaneously hypertensive rats

The vagus nerve is an important component of the efferent arm of the baroreflex. Blood pressure levels as well as baroreflex control of circulation are significantly different in male and female spontaneously hypertensive rats (SHR). We proposed to investigate the morphometric differences between genders using the vagus nerve of SHR. Adult animals (20 weeks old) were anesthetized and had their arterial pressure (AP) and heart rate (HR) recorded by a computerized system. The rats were then systemically perfused with a fixative solution and had their cervical vagi nerves prepared for light microscopy. Proximal and distal segments of the left and right vagi nerves were evaluated for morphometric parameters including fascicle area and diameter, myelinated fiber number, density, area and diameter. Comparisons were made between sides and segments on the same gender as well as between genders. Differences were considered significant when p<0.05. Male SHR had significantly higher AP and HR. Morphometric data showed no differences between the same levels of both sides and between segments on the same side for male and female rats. In addition, no significant morphometric differences were observed when genders were compared. This is the first description of vagus nerve morphometry in SHR indicating that gender differences in AP and HR cannot be attributed to dissimilarities in vagal innervation of the heart. These data provide a morphological basis for further studies involving functional investigations of the efferent arm of the baroreflex in hypertension. (C) 2007 Elsevier B.V. All rights reserved.

Bone marrow mesenchymal stem cells stabilize already-formed aortic aneurysms more efficiently than vascular smooth muscle cells in a rat model.

PURPOSE: Abdominal aortic aneurysms (AAAs) expand because of aortic wall destruction. Enrichment in Vascular Smooth Muscle Cells (VSMCs) stabilizes expanding AAAs in rats. Mesenchymal Stem Cells (MSCs) can differentiate into VSMCs. We have tested the hypothesis that bone marrow-derived MSCs (BM-MSCs) stabilizes AAAs in a rat model. MATERIAL AND METHODS: Rat Fischer 344 BM-MSCs were isolated by plastic adhesion and seeded endovascularly in experimental AAAs using xenograft obtained from guinea pig. Culture medium without cells was used as control group. The main criteria was the variation of the aortic diameter at one week and four weeks. We evaluated the impact of cells seeding on inflammatory response by immunohistochemistry combined with RT-PCR on MMP9 and TIMP1 at one week. We evaluated the healing process by immunohistochemistry at 4 weeks. RESULTS: The endovascular seeding of BM-MSCs decreased AAA diameter expansion more powerfully than VSMCs or culture medium infusion (6.5% ± 9.7, 25.5% ± 17.2 and 53.4% ± 14.4; p = .007, respectively). This result was sustained at 4 weeks. BM-MSCs decreased expression of MMP-9 and infiltration by macrophages (4.7 ± 2.3 vs. 14.6 ± 6.4 mm(2) respectively; p = .015), increased Tissue Inhibitor Metallo Proteinase-1 (TIMP-1), compared to culture medium infusion. BM-MSCs induced formation of a neo-aortic tissue rich in SM-alpha active positive cells (22.2 ± 2.7 vs. 115.6 ± 30.4 cells/surface units, p = .007) surrounded by a dense collagen and elastin network covered by luminal endothelial cells. CONCLUSIONS: We have shown in this rat model of AAA that BM-MSCs exert a specialized function in arterial regeneration that transcends that of mature mesenchymal cells. Our observation identifies a population of cells easy to isolate and to expand for therapeutic interventions based on catheter-driven cell therapy.

Time-course effects of aerobic exercise training on cardiovascular and renal parameters in 2K1C renovascular hypertensive rats

Exercise training (Ex) has been recommended for its beneficial effects in hypertensive states. The present study evaluated the time-course effects of Ex without workload on mean arterial pressure (MAP), reflex bradycardia, cardiac and renal histology, and oxidative stress in two-kidney, one-clip (2K1C) hypertensive rats. Male Fischer rats (10 weeks old; 150–180 g) underwent surgery (2K1C or SHAM) and were subsequently divided into a sedentary (SED) group and Ex group (swimming 1 h/day, 5 days/week for 2, 4, 6, 8, or 10 weeks). Until week 4, Ex decreased MAP, increased reflex bradycardia, prevented concentric hypertrophy, reduced collagen deposition in the myocardium and kidneys, decreased the level of thiobarbituric acid-reactive substances (TBARS) in the left ventricle, and increased the catalase (CAT) activity in the left ventricle and both kidneys. From week 6 to week 10, however, MAP and reflex bradycardia in 2K1C Ex rats became similar to those in 2K1C SED rats. Ex effectively reduced heart rate and prevented collagen deposition in the heart and both kidneys up to week 10, and restored the level of TBARS in the left ventricle and clipped kidney and the CAT activity in both kidneys until week 8. Ex without workload for 10 weeks in 2K1C rats provided distinct beneficial effects. The early effects of Ex on cardiovascular function included reversing MAP and reflex bradycardia. The later effects of Ex included preventing structural alterations in the heart and kidney by decreasing oxidative stress and reducing injuries in these organs during hypertension.

Soya phytoestrogens change cortical and hippocampal expression of BDNF mRNA in male rats

Adult male hooded Lister rats were either fed a diet containing 150 microg/g soya phytoestrogens or a soya-free diet for 18 days. This concentration of phytoestrogens should have been sufficient to occupy the oestrogen-beta, but not the oestrogen-alpha, receptors. Using in situ hybridisation, significant reductions were found in brain-derived neurotrophic factor (BDNF) mRNA expression in the CA3 and CA4 region of the hippocampus and in the cerebral cortex in the rats fed the diet containing phytoestrogens, compared with those on the soya-free diet. No changes in glutamic acid decarboxylase-67 or glial fibrillary acidic protein mRNA were found. This suggests a role for oestrogen-beta receptors in regulating BDNF mRNA expression.

Exercise reduces inhibitory neuroactivity and protects myenteric neurons from age-related neurodegeneration

The practice of regular exercise is indicated to prevent some motility disturbances in the gastrointestinal tract, such as constipation, during aging. The motility alterations are intimately linked with its innervations. The goal of this study is to determine whether a program of exercise (running on the treadmill), during 6 months, has effects in the myenteric neurons (NADH- and NADPH-diaphorase stained neurons) in the colon of rats during aging. Male Wister rats 6 months (adult) and 12 months (middle-aged) old were divided into 3 different groups: AS (adult sedentary), MS (middle-aged sedentary) and MT (middle-aged submitted to physical activity). The aging did not cause a decline significant (p > 0.05) of the number of NADH-diaphorase stained neurons in sedentary rats (AS vs. MS group). In contrast, a decline of 3 1% was observed to NADPH-diaphorase stained neurons. Thus, animals that underwent physical activity (AS vs. MT group) rescued neurons from degeneration caused by aging (total number, density and profile of neurons did not change with age - NADH-diaphorase method). On the other hand, physical activity augmented the decline of NADPH-diaphorase positive neurons (total number, density and profile of neurons decreased). Collectively, the results show that exercise inhibits age-related decline of myenteric neurons however, exercise augments the decline of neurons with inhibitory activity (nitric oxide) in the colon of the rats. (c) 2008 Elsevier B.V. All rights reserved.

Low caloric value of ethanol itself increases alveolar bone loss in ligature-induced periodontitis in male rats

This study aimed at morphometrically evaluating the influence of variable caloric values of ethanol consumption on alveolar bone loss in periodontitis in male rats. Thirty-six male rats were randomized into four groups of nine rats each, as follows: Test group A (low) - rats were fed an ethanol-containing liquid diet (ethanol representing 22% of total caloric value); Control group A - rats were fed a pair-fed control diet (ethanol replaced by isocaloric amounts of carbohydrate); Test group B (high) - rats were fed an ethanol-containing liquid diet (ethanol representing 36% of total caloric value); Control group B - rats were fed a pair-fed control diet for Test B. Following anesthesia, cotton ligatures were placed around the cervix of the right upper second molar. At eight weeks, the maxillary bones were removed and alveolar bone loss was analyzed by measuring the distance between the cementoenamel junction and the alveolar bone crest at buccal and palatal sites of the upper second molar. The unligated groups showed no significant differences between the bone loss values observed for the low and high caloric values of ethanol (p > 0.05). In the ligated groups, the rats receiving low caloric values of ethanol showed significantly greater bone loss compared to the isocaloric rats (p < 0.05); however, the rats receiving high caloric values of ethanol showed no significant differences compared to the controls. Analysis of the results demonstrated that, in male rats, ethanol itself affected ligature-induced bone loss when representing a low value in the total caloric value.

Brazilian experience with the medium-term multi-organ bioassay: Scientific and regulatory developments

In 1996 the Brazilian Institute for the Environment (IBAMA) officially adopted a variation of the multiorgan initiation-promotion DMBDD bioassay as a valid source of evidence of the carcinogenic potential of pesticides. The protocol adopted by IBAMA was a modification of the one originally proposed by researchers led by Nobuyuki Ito, from the Nagoya City University Medical School. Among the modifications established in the Brazilian protocol were the use of both sexes of the outbreed Wistar strain of rats and two positive control test chemicals. The adoption of the modified DMBDD protocol was instrumental during the last decade for qualifying technical people and to spread knowledge on chemical carcinogenesis in Brazil.

Spatial Learning and Stress Response of Male Rats Prenatally Exposed to Dexamethasone

Dexamethasone is routinely administered to women at risk for a preterm birth in order to enhance fetal lung development and reduce uterine contractions. Research has demonstrated possible behavioral abnormalities in adulthood as a result of dexamethasone treatment. Using nonlinear mixed effects modeling, this study found thatprenatal dexamethasone treatment impaired spatial learning and memory of adult male Sprague-Dawley rats. Prenatal dexamethasone treatment also led to more anxiety related behaviors on Elevated Plus Maze testing 1.5 hours after a stress challenge. Because theassumptions underlying the independent samples t-test were violated, the randomization test was used to compare groups on the Elevated Plus Maze.

Genetic variation in vulnerability to the behavioral effects of neonatal hippocampal damage in rats.

We explored how two independent variables, one genetic (i.e., specific rat strains) and another environmental (i.e., a developmental excitotoxic hippocampal lesion), contribute to phenotypic variation. Sprague-Dawley (SD), Fischer 344 (F344), and Lewis rats underwent two grades of neonatal excitotoxic damage: small and large ventral hippocampal (SVH and LVH) lesions. Locomotion was tested before puberty [postnatal day 35 (P35)] and after puberty (P56) following exposure to a novel environment or administration of amphetamine. The behavioral effects were strain- and lesion-specific. As shown previously, SD rats with LVH lesions displayed enhanced spontaneous and amphetamine-induced locomotion as compared with controls at P56, but not at P35. SVH lesions in SD rats had no effect at any age. In F344 rats with LVH lesions, enhanced spontaneous and amphetamine-induced locomotion appeared early (P35) and was exaggerated at P56. SVH lesions in F344 rats resulted in a pattern of effects analogous to LVH lesions in SD rats--i.e., postpubertal onset of hyperlocomotion (P56). In Lewis rats, LVH lesions had no significant effect on novelty- or amphetamine-induced locomotion at any age. These data show that the degree of genetic predisposition and the extent of early induced hippocampal defect contribute to the particular pattern of behavioral outcome. These results may have implications for modeling interactions of genetic and environmental factors involved in schizophrenia, a disorder characterized by phenotypic heterogeneity, genetic predisposition, a developmental hippocampal abnormality, and vulnerability to environmental stress.