Half Of Animals Research Articles

Uso do abate comparativo na determinação da exigência de energia de mantença de gado de corte pastejando capim-elefante: descrição da metodologia e dos resultados

Determinou-se o requerimento líquido de energia de mantença de novilhos mantidos em regime exclusivo de pastejo, em capim-elefante (Pennisetum purpureum, Schum) cv. Napier. O experimento teve duração de 175 dias, utilizando-se nove piquetes, estabelecidos em terreno plano, com área individual de 0,5 ha, sob pastejo rotacionado. Foram utilizados 36 novilhos ¾Gir-Holandês com 332 ± 37 kg e média de 20 meses de idade inicial. Metade dos animais teve o tempo de pastejo restrito a quatro horas por dia, de forma a se limitar o consumo de energia a pouco acima do nível de mantença, e metade teve acesso irrestrito à pastagem. Seis novilhos foram abatidos no início do experimento, para se determinar a composição corporal inicial (grupo referência) e seis (três de cada tratamento) a intervalos sucessivos de 35 dias. A composição corporal dos animais abatidos foi determinada por análise química de amostras representativas de todo o corpo. As exigências líquidas de energia para mantença foram estimadas como o calor produzido em nível zero de ingestão de energia metabolizável, ajustando-se equações do logaritmo da produção de calor em função do consumo de energia metabolizável, por dia e por unidade de tamanho metabólico (kg0,75). Foi ajustada uma equação, calculando-se o tamanho metabólico em função do peso vivo (PV) e outra em função do peso de corpo vazio (PCVZ). O consumo e a digestibilidade da forragem ingerida (extrusa) foram determinados mensalmente. O tempo de pastejo limitado a quatro horas diárias assegurou ingestão de energia em nível pouco acima da mantença, conforme indicaram os ganhos de peso individuais, evidenciando que este procedimento pode ser recomendado para trabalhos futuros. A exigência de energia de mantença encontrada foi de 57 kcal/kg PV0,75 ou 63,27 kcal/kg PCVZ0,75. Os valores obtidos estão dentro da faixa de resultados obtidos no Brasil, sendo mais baixos que os valores recomendados no exterior, para gado europeu.

Supplement to the Carcinogenic Potency Database (CPDB): Results of Animal Bioassays Published in the General Literature through 1997 and by the National Toxicology Program in 1997–1998

The Carcinogenic Potency Database (CPDB) is a systematic and unifying resource that standardizes the results of chronic, long-term animal cancer tests which have been conducted since the 1950s. The analyses include sufficient information on each experiment to permit research into many areas of carcinogenesis. Both qualitative and quantitative information is reported on positive and negative experiments that meet a set of inclusion criteria. A measure of carcinogenic potency, TD(50) (daily dose rate in mg/kg body weight/day to induce tumors in half of test animals that would have remained tumor-free at zero dose), is estimated for each tissue-tumor combination reported. This article is the ninth publication of a chronological plot of the CPDB; it presents results on 560 experiments of 188 chemicals in mice, rats, and hamsters from 185 publications in the general literature updated through 1997, and from 15 Reports of the National Toxicology Program in 1997-1998. The test agents cover a wide variety of uses and chemical classes. The CPDB Web Site (http://potency.berkeley.edu/) presents the combined database of all published plots in a variety of formats as well as summary tables by chemical and by target organ, supplemental materials on dosing and survival, a detailed guide to using the plot formats, and documentation of methods and publications. The overall CPDB, including the results in this article, presents easily accessible results of 6153 experiments on 1485 chemicals from 1426 papers and 429 NCI/NTP (National Cancer Institute/National Toxicology program) Technical Reports. A tab-separated format of the full CPDB for reading the data into spreadsheets or database applications is available on the Web Site.

Chronic Developmental Lead Exposure Reduces Neurogenesis in Adult Rat Hippocampus but Does Not Impair Spatial Learning

The dentate granule cell (DG) layer of the hippocampal formation has the distinctive property of ongoing neurogenesis that continues throughout adult life. Although the function of these newly generated neurons and the mechanisms that control their birth are unknown, age, activity, diet and psychosocial stress have all been demonstrated to regulate this type of neurogenesis. Little information on the impact of environmental insults on this process has appeared to date. Developmental lead (Pb) exposure has been well documented to impair cognitive function in children and animals and reduce activity-dependent synaptic plasticity in the hippocampus of rodents. Therefore, we examined the effects of this classic environmental neurotoxicant on hippocampal-dependent learning and adult neurogenesis in the hippocampus. Pregnant rats were exposed to a low level of Pb-acetate (0.2%) via the drinking water from late gestation (GD 16) until weaning on postnatal day 21 (PN 21). At weaning, half of the Pb-exposed animals were weaned to control drinking water and the remainder were maintained on Pb water until termination of the study. Animals were paired- housed and on PN 75 were administered a series of injections of a thymidine analog bromodeoxyuridine (BrdU), a marker of DNA synthesis that labels proliferating cells and their progeny. At 12-h intervals for 12 days, rats received an ip injection of BrdU (50 mg/kg). Subjects were sacrificed and perfused 24 h and 28 days after the last injection. Spatial learning was assessed in an independent group of animals beginning on PN 110 using a Morris water maze. No Pb-induced impairments were evident in water maze learning. Immunohistochemistry for the detection of BrdU-labeled cells was performed on 40-microm coronal sections throughout the hippocampus. Continuous exposure to Pb (Life) reduced the total number of BrdU-positive cells at 28 days without affecting the total number of labeled cells evident 24 h after the last injection. No differences in the number of progenitor cells labeled or surviving were seen between control and treated animals whose Pb exposure was terminated at weaning. Double labeling with BrdU and the glial specific marker, glial acidic fibrillary protein (GFAP) indicated that the bulk of the surviving cells were of a neuronal rather than a glial phenotype. These data reveal that chronic low-level Pb exposure reduces the capacity for neurogenesis in the adult hippocampus. Despite deficits in synaptic plasticity previously reported from our laboratory, and now structural plasticity, no significant impact on spatial learning was detected.

The FoxO-subclass in Xenopus laevis development

Transcription factors of the Fox ( fork head b ox) family are involved in cellular specification and determination processes. Here, we report on the isolation and first characterisation of two members of the FoxO subclass in Xenopus laevis, xFoxO1 and xFoxO3. These sequences exhibit 68% (67%) and 69% (70%) identity to their mouse (human) orthologues, respectively. Serine and threonine residues, which are phosphorylated upon insulin signalling, are evolutionarily conserved from frogs to mammals. xFoxO1 and xFoxO3 genes are maternally transcribed, but transcripts disappear during early cleavage stages. Zygotic transcription of both genes starts at the late neurula stages and transcripts accumulate at the end of organogenesis. While maternal transcripts of both genes are found within the animal half of the early embryo, zygotic transcripts show distinct patterns. xFoxO1 expression is observed in the pronephros, within head mesenchyme in front of the eye, within the branchial arches and in the liver primordium. At the late neurula, xFoxO3 is found to be specifically expressed in the anterior neural plate and in neural crest cells. Later, expression of xFoxO3 is observed in a variety of organs and tissues, like the head, the branchial arches and the somites.

Negative regulation of Smad2 by PIASy is required for properXenopusmesoderm formation

Mesoderm induction and patterning are primarily regulated by the concentration of locally expressed morphogens such as members of the TGFbetasuperfamily. Smad2 functions as a transcription factor to regulate expression of mesodermal genes downstream of such morphogens. We have identified Xenopus PIASy (XPIASy), a member of the PIAS family, by yeast two-hybrid screening using Xenopus Smad2 (XSmad2) as a bait. During mesoderm induction, XPIASy is expressed in the animal half of embryos with a ventral high-dorsal low gradient at the marginal zone. XPIASy expression is positively and negatively regulated by activities of the XSmad2 and Wnt pathways, respectively. Interestingly, inhibition of XPIASy by morpholinos induces elongation of animal caps with induction of mesoderm genes even in the absence of their morphogen-mediated activation. In addition, their introduction into the ventral marginal zone results in a secondary axis formation. Gain-of-function analysis revealed that XPIASy inhibits mesoderm induction by specific and direct downregulation of XSmad2 transcriptional activity. These observations indicate that XPIASy functions as an essential negative regulator of the XSmad2 pathway to ensure proper mesoderm induction at the appropriate time and in the appropriate region, and suggest that both the initial step of morphogen-mediated activation of the XSmad2 pathway and regulation of the final downstream transcription step have crucial roles in mesoderm induction and patterning.

Ultrastructural alterations in capillaries of the diabetic hypertensive rat retina: protective effects of ACE inhibition.

The ACE inhibitor cilazapril was administered to diabetic hypertensive rats to evaluate its ability to influence the development of retinal capillary alterations. Normotensive (strain: Wistar Kyoto) and genetically hypertensive (strain: spontaneously hypertensive) rats were rendered diabetic by intravenous injections of streptozotocin. Half of the diabetic animals received cilazapril with their daily food. At 20 weeks of diabetes, endothelial cells, pericytes and extracellular matrix were assessed by ultrastructural morphometry. Each experimental group consisted of seven animals. Cilazapril normalised systolic arterial pressure in diabetic hypertensive rats (137+/-2 mm Hg compared with 188+/-16 mm Hg in non-medicated diabetic hypertensive rats, p<0.001). The number of endothelial intercellular junctions was reduced in untreated diabetic hypertensive rats (0.15+/-0.05, p<0.02, vs 0.47+/-0.20 in non-diabetic normotensive rats). In diabetic hypertensive animals treated with cilazapril, this loss was attenuated (0.32+/-0.16, p<0.05). The significant thickening of the basement membrane observed in the diabetic normotensive (132.8+/-19.4 nm) and diabetic hypertensive (150.3+/-20.2 nm) groups was decreased by cilazapril in the diabetic hypertensive group (116.7+/-11.0 nm, p<0.01), but was unaffected in the normotensive (131.9+/-17.3 nm) group. No protective effect of the drug was observed in either group on pericytes. Long-term administration of an effective antihypertensive therapy normalises endothelial alterations and basement membrane thickness in diabetic hypertensive conditions, and thus may account for the well-known improvement of the blood-retinal barrier observed during antihypertensive treatment.

Therapeutic doses of L-dopa reverse hypersensitivity of corticostriatal D2-dopamine receptors and glutamatergic overactivity in experimental parkinsonism

Levodopa (l-dopa) therapy is still considered the gold-standard in the treatment of Parkinson's disease. However, the synaptic and cellular mechanisms involved in the amelioration of motor symptoms during this treatment are still unclear. To address this issue, we analysed the physiological and pharmacological properties of striatal glutamatergic and GABAergic synaptic transmission in an experimental model of Parkinson's disease. Single-cell recordings were performed in sham-operated rats, in 6-hydroxydopamine-lesioned animals and in rats receiving chronic l-dopa treatment following dopamine (DA) denervation. We utilized a dose of l-dopa (10 mg/kg, twice daily for 21 days) able to reverse motor deficits in about half of parkinsonian animals. In the striatum of parkinsonian animals showing therapeutic benefits following l-dopa treatment, we observed a reversal of glutamatergic overactivity and of the hypersensitivity of presynaptic D2 DA receptors controlling glutamate release from corticostriatal terminals. Conversely, no change was detected in the sensitivity of presynaptic D2 DA receptors modulating striatal GABA transmission in both parkinsonian and l-dopa-treated rats. We suggest that the reversal of striatal glutamatergic overactivity and the normalization of hypersensitive D2 DA receptors modulating excitatory transmission might underlie some of the therapeutic actions of l-dopa in Parkinson's disease.

Ehrlichia ruminantium seroprevalence in domestic ruminants in Ghana : I. Longitudinal survey in the Greater Accra Region

Serum samples collected monthly over a 34-month period from cattle, sheep and goats in the Greater Accra Region of Ghana were tested for antibodies to Ehrlichia (previously Cowdria) ruminantium, the causative agent of heartwater, by polyclonal competitive ELISA (PC-ELISA). Maternal antibodies, detected in about half of animals followed from under 1 month old, declined to negative levels within 2–4 months. Amblyomma variegatum tick vectors were present on livestock in rural areas throughout the year, and first seroconversion occurred at any age, although the majority of calves seroconverted between 1 and 10 months old, sheep by 11 months, and goats by 7 months. All the cattle in the study became seropositive by 20 months of age, except one animal which subsequently died of heartwater. Following seroconversion, 25% of bovine sera tested negative in the PC-ELISA. Just over half the sheep in the survey seroconverted before or during the study period; following seroconversion, less than 3% of ovine sera became PC-ELISA negative. About a quarter of the goats seroconverted, and 34% of their post-seroconversion sera tested negative in the PC-ELISA. Overall, the serology indicated that virtually all cattle on the survey farms were exposed to E. ruminantium without suffering disease, but that a substantial proportion of sheep and goats escaped exposure and thus formed a susceptible population. E. ruminantium was detected in brains of 14, 36 and 4% of cattle, sheep and goats submitted for post mortem at the Accra Veterinary Laboratory, indicating that sheep were most at risk from heartwater disease.

Neurodéveloppement chez les enfants exposés aux antiépileptiques et aux psychotropes en anténatal

Attention deficit/hyperactivity disorder (ADHD) is encountered among patients with epilepsy at a significantly higher rate than in the general population. Mechanisms of epilepsy–ADHD comorbidity remain largely unknown. We investigated whether a model of chronic epilepsy in rats produces signs of ADHD, and thus, whether it can be used for studying mechanisms of this comorbidity. Epilepsy was induced in male Wistar rats via pilocarpine status epilepticus. Half of the animals exhibited chronic ADHD-like abnormalities, particularly increased impulsivity and diminished attention in the lateralized reaction-time task. These impairments correlated with the suppressed noradrenergic transmission in locus coeruleus outputs. The other half of animals exhibited depressive behavior in the forced swimming test congruently with the diminished serotonergic transmission in raphe nucleus outputs. Attention deficit/hyperactivity disorder and depressive behavior appeared mutually exclusive. Therefore, the pilocarpine model of epilepsy affords a system for reproducing and studying mechanisms of comorbidity between epilepsy and both ADHD and/or depression.

Vitelline envelope gps 63 and 75 specifically bind sperm in "in vitro" assays in Discoglossus pictus.

In Xenopus, conflicting data related to sperm-vitelline envelope (VE) binding suggest that further experiments should be performed to study the role of VE glycoproteins in sperm binding. In this article, we studied the VE of Discoglossus pictus, where gp63, the product of the Dp ZP2 gene, has high molecular identity to Xenopus gp69/64 and to mouse ZP2 and only A23187-treated sperm bind to VE. Sperm bind to VE all over the egg, yet a sperm tuft was found only in the animal half of the egg, where the dimple, the site of fertilization, is located and an intense immunostain was detected in VE by an antiserum directed against gp69/64. The same antiserum inhibited sperm binding to VE. Sperm binding to beads coated with gp63, gp40, or gp75 was in the range of 62-70% for gp63-beads, 67-75% for 75 beads, and about 20% for BSA beads and gp40-coated beads. Soluble purified gp63 and gp75 competitively inhibited binding of sperm to gp63-coated beads. Similarly, the same glycoproteins inhibited sperm binding to gp75-coated beads. SDS-polyacrylamide gels (PAGE) of FE and comparison of VE and FE peptide maps showed that gp63 undergoes a minor shift to about 62 kDa in FE. In sperm binding assays with beads coated with FEs gp62, there was no binding. Following fertilization, in the region of the dimple, an F-layer is formed as well as an alteration of the VE structure. Lectin blots of the FE showed that the FE and in particular gp62 acquires a stronger affinity to Maackia amurensis agglutinin (MAA) with respect to VEs gp63. These results indicate that gps 63 and 75 are the sperm binding glycoproteins of D. pictus VE, where major post-fertilization changes occur as in other anuran species.

Prepubertal propagation of transgenic cloned goats by laparoscopic ovum pick-up and in vitro embryo production.

The use of laparoscopic ovum pick-up (LOPU) followed by in vitro embryo production was evaluated in the early propagation of cloned goats. Ten kinder goats produced by somatic cell nuclear transfer technology were used as oocyte donors. Half of the donor animals were subjected to LOPU at 2-3 months of age (prior to induction of lactation), whereas the other five goats were subjected to LOPU at 6-7 months of age (following induction to lactation). They were stimulated with 80 mg NIH-FSH-P1 (Folltropin, Vetrepharm, Canada) together with 300 IU eCG (Novormon, Vetrepharm, Canada) administered intramuscularly 36 h prior to LOPU. The number of follicles aspirated and oocytes recovered was higher in the younger group of donors (57 +/- 7 and 41 +/- 4 vs. 28 +/- 2 and 25.8 +/- 2, p < 0.05), however, oocytes from animals in the late prepubertal age showed higher developmental capacity resulting in higher transferable embryo yield (81.4% vs. 67.8%, p < 0.01), pregnancy rate (80% vs. 40%, p < 0.05) and total kids born (27 vs. 15, p < 0.01). In conclusion, LOPU in combination with in vitro embryo production techniques is an efficient method for the early propagation of valuable goats produced by somatic cell nuclear transfer.

Isolation and developmental expression of Xenopus FoxJ1 and FoxK1.

xFoxJ1 and xFoxK1, two novel members of the fork head/winged helix family of transcription factors in Xenopus, have been identified by sequence homology to their mammalian orthologues. We here report the complete amino acid sequences of xFoxJ1, its pseudo-allelic version xFoxJ1', and of xFoxK1, the latter also comprising a fork-head-associated (FHA) domain. We have further analysed the temporal and spatial expression of both genes. xFoxJ1 transcripts were detected during gastrulation in the dorsal blastopore lip and in the animal half. A predominant feature of xFoxJ1 expression until the tailbud stage is its spotty pattern within the epidermis. xFoxK1 is maternally expressed and localized within the animal half during early cleavage stages. Zygotic transcription starts during neurulation and is initially restricted to neuroectoderm. As development proceeds, xFoxK1 is expressed in ectoderm- and mesoderm-derived tissues, like the branchial arches, brain, eye, otic vesicle, pronephros, somites and abdominal muscle precursors.

An Engineered Biopolymer Prevents Mucositis Induced by 5-Fluorouracil in Hamsters

Oral mucositis is a common, treatment-limiting, and costly side effect of cancer treatments whose biological underpinnings remain poorly understood. In this study, mucositis induced in hamsters by 5-fluorouracil (5-FU) was observed after cheek-pouch scarifications, with and without administration of RGTA (RG1503), a polymer engineered to mimic the protective effects of heparan sulfate. RG1503 had no effects on 5-FU-induced decreases in body weight, blood cell counts, or cheek-pouch and jejunum epithelium proliferation rates, suggesting absence of interference with the cytotoxic effects of 5-FU. Extensive mucositis occurred in all of the untreated animals, and consisted of severe damage to cheek pouch tissues (epithelium, underlying connective tissue, and muscle bundles). Only half of the RG1503-treated animals had mucositis, over a mean area 70% smaller than in the untreated animals. Basement membranes were almost completely destroyed in the untreated group but was preserved in the RG1503 group. RG1503 blunted or abolished the following 5-FU-induced effects: increases in matrix metalloproteinase (MMP)-2, MMP-9, and plasmin, and decreases in tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2. These data indicate that mucositis lesions are related to massive release of proteolytic enzymes and are improved by RG1503 treatment, this effect being ascribable in part to restoration of the MMP-TIMP balance. RG1503 given with cancer treatment might protect patients from mucositis.

Dietary Lutein/Zeaxanthin Decreases Ultraviolet B-Induced Epidermal Hyperproliferation and Acute Inflammation in Hairless Mice

Lutein and zeaxanthin are carotenoids found in green leafy vegetables with interesting antioxidant properties. They are present in high concentrations in the fovea centralis of the human retina and their role in the prevention of age-related macula degeneration has been reported. We have investigated the effect of orally administered lutein and zeaxanthin in the cutaneous response to ultraviolet B irradiation. Female hairless SKh-1 mice receiving 0.4% and 0.04% lutein plus zeaxanthin-enriched diet for 2 wk were exposed to single doses of ultraviolet B radiation. Skin biopsies were taken at 24 and 48 h after irradiation and analyzed for the presence of apoptotic cells, proliferating cells, and expression of proliferating cell nuclear antigen. Our results show a clear ultraviolet-induced dose-dependent inflammatory response. Orally administered 0.4% lutein and zeaxanthin decreased significantly the edematous cutaneous response (p<0.01) as determined by the reduction of the UVB-induced increase of ear bifold thickening. Additionally, dietary carotenoids were efficient in reducing the ultraviolet B-induced increases in the percentage of proliferating cell nuclear antigen (p<0.05), bromodeoxyuridine (p<0.05), and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling-positive cells (p<0.01). These data demonstrate that oral supplementation of lutein and zeaxanthin diminishes the effects of ultraviolet B irradiation by reducing acute inflammatory responses and ultraviolet-induced hyperproliferative rebound.

Analysis of repeated tests for interferon-gamma (IFN-γ) response and faecal excretion for diagnosis of subclinical paratuberculosis in Danish cattle

A total of 315 cattle were tested for infection with Mycobacterium avium subsp. paratuberculosis ( M. paratuberculosis) at three consecutive samplings, using the interferon-gamma (IFN-γ) test on whole blood and bacteriological culture of faecal samples. Of 205 cattle from 10 infected herds 99 (48%) were positive in the IFN-γ test on at least one sampling using “IDEXX-criteria” for interpretation, and of 110 cattle from five non-infected herds three (3%) were positive. Forty-four animals from infected and one from non-infected herds tested positive at all three samplings. Although support for the specificity of the IFN-γ test was provided by these results, they also indicate problems with false positives. Approximately half of the positive animals did not give the same result at all three samplings, indicating that repeated testing increases the chance of detecting reactors. Changing, or fluctuating, IFN-γ test results occurred most frequently in animals younger than 1 year, indicating that the IFN-γ test should be applied only to animals 1 year and older. M. paratuberculosis was isolated from 16 (4%) of 371 cattle, all from infected herds. Fifteen culture-positive cattle tested positive at least once in the IFN-γ test. It was not possible to predict from the IFN-γ test result the number of animals that would eventually develop disease. However, the test may be useful to detect animals that have been exposed to M. paratuberculosis earlier in their lives, and the testing of young cattle could be included in a control program to check for the effectiveness of preventing transmission of infection to calves and to identify animals at risk of developing disease later in their lives.

Nuclear envelope breakdown in starfish oocytes proceeds by partial NPC disassembly followed by a rapidly spreading fenestration of nuclear membranes.

Breakdown of the nuclear envelope (NE) was analyzed in live starfish oocytes using a size series of fluorescently labeled dextrans, membrane dyes, and GFP-tagged proteins of the nuclear pore complex (NPC) and the nuclear lamina. Permeabilization of the nucleus occurred in two sequential phases. In phase I the NE became increasingly permeable for molecules up to ∼40 nm in diameter, concurrent with a loss of peripheral nuclear pore components over a time course of 10 min. The NE remained intact on the ultrastructural level during this time. In phase II the NE was completely permeabilized within 35 s. This rapid permeabilization spread as a wave from one epicenter on the animal half across the nuclear surface and allowed free diffusion of particles up to ∼100 nm in diameter into the nucleus. While the lamina and nuclear membranes appeared intact at the light microscopic level, a fenestration of the NE was clearly visible by electron microscopy in phase II. We conclude that NE breakdown in starfish oocytes is triggered by slow sequential disassembly of the NPCs followed by a rapidly spreading fenestration of the NE caused by the removal of nuclear pores from nuclear membranes still attached to the lamina.

Effects of a macrolide antibiotic on enamel formation in rat incisors--primary lesion of ameloblast at the transition stage.

A novel macrolide antibiotic was administered orally to 5-week-old Jcl:Wistar rats at a dose of 5,000 mg/kg/day for 5 weeks, and then a half of animals were maintained without any treatment for 10 weeks. A white discolored lesion with horizontal stripes developed on the surface of the upper and lower incisors after dosing for 4 weeks, and these macroscopical incisal lesions disappeared with the eruption in 4 weeks after stop of administration. Histopathologically, increase in number of karyopycnosis of ameloblast at the transitional stage, vacuolar degeneration of ameloblast and cystic change in the maturation stage, and impaired iron pigment secretion at the pigmentation stage were observed. Microradiography, calcio-traumatic zones, which means hypocalcification, were observed on the superficial layer of enamel. These results suggest that the primary lesion induced by a novel macrolide antibiotic is the increased karyopycnosis of ameloblast at the transitional stage, and followed by later stage.

Experimental analysis of second cleavage in the mouse.

Mammalian conceptuses typically have an approximately regular tetrahedral shape at the 4-cell stage. In the rabbit, this has been attributed to both 2-cell blastomeres dividing meridionally, but with the animal-vegetal axis of the second blastomere to divide rotating through roughly 90 degrees before or during cytokinesis. The aim of the present study was to ascertain whether this was also true for the mouse. First, the distribution in regular tetrahedral 4-cell conceptuses of fluorescent microspheres applied to the vegetal polar region of one or both blastomeres at the 2-cell stage was analysed. Second, the ability of 2-cell stages to form regular tetrahedral 4-cell conceptuses after the previtelline space had been gelated to prevent blastomeres from rotating was also investigated. Neither experiment yielded evidence supporting blastomere rotation during second cleavage. Rather, the findings were consistent with the regular tetrahedral form of 4-cell conceptus resulting from meridional division of one blastomere and approximately equatorial division of the other. Second cleavage in the mouse typically yields 4-cell conceptuses with three distinct types of blastomere. While both products of the meridional division include all axial levels of the zygote, those of the equatorial division acquire only its vegetal or animal half.

Nitric Oxide in Obstructive Uropathy: Role of Endothelial Nitric Oxide Synthase

Nitric oxide (NO) ameliorates fibrosis in experimental obstructive uropathy. Previously, we demonstrated that renal fibrosis was decreased after 2 weeks of unilateral ureteral obstruction in inducible nitric oxide synthase (iNOS) knock-out mice given L-arginine supplemented drinking water. We proposed that the 2 constitutive isoforms of nitric oxide synthase (NOS) mediated down-regulation of renal fibrosis in response to prolonged ureteral obstruction. To determine the specific role of endothelial NOS (eNOS) versus neuronal NOS in modulating renal fibrosis due to obstructive uropathy, we evaluated renal injury following unilateral ureteral obstruction in C57BL/6J mice subjected to biochemical inhibition of the constitutive isoforms of NOS and in eNOS knockout mice.Four groups of C57BL/6J mice were studied. Complete unilateral ureteral obstruction was created by ligating the right ureter at age 8 weeks. A single daily intraperitoneal injection of 30 mg./kg. S-methyl-L-thiocitrulline (SMLT), a selective neuronal and endothelial NOS inhibitor was started 24 hours before ureteral obstruction and administered to half of the study animals. SMLT treated mice and control animals were further subdivided to receive either regular tap water or 1% L-arginine (weight/volume) supplemented water after unilateral ureteral obstruction. Animals were sacrificed on postoperative day 3, 7 or 14. In addition, eNOS knockout mice with unilateral ureteral obstruction were given tap water or L-arginine supplemented water to drink and sacrificed after 14 days. Urine specimens from the bladder and the obstructed renal pelvis along with serum were collected. Nitrite level in each fluid was determined. Renal morphology and cortical thickness were assessed in the normal and obstructed kidneys. Interstitial fibrosis was evaluated using trichrome stain.SMLT was well tolerated by C57BL/6J mice. Serum nitrite levels and nitrite excretion in bladder urine were similar in all SMLT treated groups throughout the duration of unilateral ureteral obstruction. A reduction of pelvic urine nitrite levels by 89%, 68% and 48% versus bladder urine nitrite levels was observed after 3, 7 and 14 days of unilateral ureteral obstruction (p <0.05). Administration of SMLT resulted in a significant increase in bladder urine nitrite level at 7 days and in pelvic urine nitrite levels at 14 days. No significant histological differences in the obstructed kidney were seen after 3, 7 or 14 days of unilateral ureteral obstruction in SMLT treated versus control mice regardless of whether they received tap water or L-arginine supplemented drinking water. In eNOS knockout mice with unilateral ureteral obstruction for 14 days L-arginine supplementation had no effect on pelvic urine nitrite levels and did not alter renal histopathology or cortical thickness.NO production is decreased in the obstructed kidney in mice with unilateral ureteral obstruction. Biochemical inhibition of constitutive NOS did not modulate renal injury after 14 days of unilateral ureteral obstruction. In contrast to previous findings with iNOS knockout mice, dietary supplementation with L-arginine had no effect on the degree of fibrosis in the obstructed kidney in SMLT treated C57BL/6J or eNOS knockout mice. We conclude that NO derived from eNOS within the kidney has a pivotal role in protecting against renal fibrosis in response to unilateral ureteral obstruction.

Comparison of telithromycin, a new ketolide, with erythromycin and clarithromycin for the treatment of Haemophilus influenzae pneumonia in suckling, middle aged and senescent mice

We studied the efficacy of telithromycin in Haemophilus influenzae pneumoniae in three different age groups of mice. Pneumonia was produced by endotracheal instillation of 1×10 4 CFU/ml of bacteria and treatment was initiated with saline for control and compared with two different doses, 50 and 100 mg/kg per BID telithromycin twice a day for 1 week. For comparison, we used erythromycin (ERY) and clarithromycin (CLA), both given twice a day at 50 mg/kg per BID. Some animals were euthanized on the third or 7th day of therapy and their lung tissue was cultured for bacteria. Presence of bacteria was considered a failure and a sterile lung was considered cured. As expected, about one half of middle-aged animals (8–10 months) were cured on saline. In contrast, almost none of the young (2–3 weeks) and old animals (18–20 months) were cured without antibiotic therapy. Among the young, the cure rates with telithromycin, ERY and CLA were 81, 50 and 33%, respectively. Of the senescent mice, the cure rate with ERY was 50% whereas the rates with CLA and telithromycin (50 mg/kg) were 62 and 75%, respectively. In conclusion, telithromycin is effective against H. influenzae at both extremes of life.