| Home | << 1 2 3 4 5 6 >> |
|
Touma, C., Sachser, N., Mostl, E., & Palme, R. (2003). Effects of sex and time of day on metabolism and excretion of corticosterone in urine and feces of mice. Gen Comp Endocrinol, 130(3), 267–278.
Abstract: Non-invasive techniques to monitor stress hormones in small animals like mice offer several advantages and are highly demanded in laboratory as well as in field research. Since knowledge about the species-specific metabolism and excretion of glucocorticoids is essential to develop such a technique, we conducted radiometabolism studies in mice (Mus musculus f. domesticus, strain C57BL/6J). Each mouse was injected intraperitoneally with 740 kBq of 3H-labelled corticosterone and all voided urine and fecal samples were collected for five days. In a first experiment 16 animals (eight of each sex) received the injection at 9 a.m., while eight mice (four of each sex) were injected at 9 p.m. in a second experiment. In both experiments radioactive metabolites were recovered predominantly in the feces, although males excreted significantly higher proportions via the feces (about 73%) than females (about 53%). Peak radioactivity in the urine was detected within about 2h after injection, while in the feces peak concentrations were observed later (depending on the time of injection: about 10h postinjection in experiment 1 and about 4h postinjection in experiment 2, thus proving an effect of the time of day). The number and relative abundance of fecal [3H]corticosterone metabolites was determined by high performance liquid chromatography (HPLC). The HPLC separations revealed that corticosterone was extensively metabolized mainly to more polar substances. Regarding the types of metabolites formed, significant differences were found between males and females, but not between the experiments. Additionally, the immunoreactivity of these metabolites was assessed by screening the HPLC fractions with four enzyme immunoassays (EIA). However, only a newly established EIA for 5alpha-pregnane-3beta,11beta,21-triol-20-one (measuring corticosterone metabolites with a 5alpha-3beta,11beta-diol structure) detected several peaks of radioactive metabolites with high intensity in both sexes, while the other EIAs showed only minor immunoreactivity. Thus, our study for the first time provides substantial information about metabolism and excretion of corticosterone in urine and feces of mice and is the first demonstrating a significant impact of the animals' sex and the time of day. Based on these data it should be possible to monitor adrenocortical activity non-invasively in this species by measuring fecal corticosterone metabolites with the newly developed EIA. Since mice are extensively used in research world-wide, this could open new perspectives in various fields from ecology to behavioral endocrinology.
|
|
Reynhout, I. C., Cornelissen, J. J. L. M., & Nolte, R. J. M. (2007). Self-assembled architectures from biohybrid triblock copolymers. J Am Chem Soc, 129(8), 2327–2332.
Abstract: The synthesis and self-assembly behavior of biohybrid ABC triblock copolymers consisting of a synthetic diblock, polystyrene-b-polyethylene glycol (PSm-b-PEG113), where m is varied, and a hemeprotein, myoglobin (Mb) or horse radish peroxidase (HRP), is described. The synthetic diblock copolymer is first functionalized with the heme cofactor and subsequently reconstituted with the apoprotein or the apoenzyme to yield the protein-containing ABC triblock copolymer. The obtained amphiphilic block copolymers self-assemble in aqueous solution into a large variety of aggregate structures. Depending on the protein and the polystyrene block length, micellar rods, vesicles, toroids, figure eight structures, octopus structures, and spheres with a lamellar surface are formed.
|
|
Boucher, J. M., Hanosset, R., Augot, D., Bart, J. M., Morand, M., Piarroux, R., et al. (2005). Detection of Echinococcus multilocularis in wild boars in France using PCR techniques against larval form. Vet Parasitol, 129(3-4), 259–266.
Abstract: Recently, new data have been collected on the distribution and ecology of Echinococcus multilocularis in European countries. Different ungulates species such as pig, goat, sheep, cattle and horse are known to host incomplete development of larval E. multilocularis. We report a case of E. multilocularis portage in two wild boars from a high endemic area in France (Department of Jura). Histological examination was performed and the DNA was isolated from hepatic lesions then amplified by using three PCR methods in two distinct institutes. Molecular characterisation of PCR products revealed 99% nucleotide sequence homology with the specific sequence of the U1 sn RNA gene of E. multilocularis, 99 and 99.9% nucleotide sequence homology with the specific sequence of the cytochrome oxydase gene of Echinococcus genus and 99.9% nucleotide sequence homology with a genomic DNA sequence of Echinococcus genus for the first and the second wild boar, respectively.
Keywords: Animals; Base Sequence; DNA, Helminth/chemistry/genetics; Echinococcosis/parasitology/pathology/*veterinary; Echinococcus multilocularis/*isolation & purification; Electron Transport Complex IV/chemistry/genetics; France; Histocytochemistry/veterinary; Liver/parasitology/pathology; Male; Molecular Sequence Data; Polymerase Chain Reaction/veterinary; Sequence Alignment; Sus scrofa/*parasitology; Swine Diseases/*parasitology/pathology
|
|
Traversa, D., Giangaspero, A., Iorio, R., Otranto, D., Paoletti, B., & Gasser, R. B. (2004). Semi-nested PCR for the specific detection of Habronema microstoma or Habronema muscae DNA in horse faeces. Parasitology, 129(Pt 6), 733–739.
Abstract: Habronema microstoma and Habronema muscae (Spirurida: Habronematidae) are parasitic nematodes which infect the stomach and/or skin of equids. The accurate diagnosis of gastric habronemosis is central to studying its epidemiology, but data on its distribution and prevalence are lacking, mainly due to the limitations of clinical and coprological diagnosis in live horses. To overcome this constraint, a two-step, semi-nested PCR-based assay was validated (utilizing genetic markers in the nuclear ribosomal DNA) for the specific amplification of H. microstoma or H. muscae DNA from the faeces from horses (n = 46) whose gastrointestinal parasite status had been determined at autopsy and whose faeces were examined previously using a conventional parasitological approach. Of these horses examined at autopsy, some harboured adults of either H. microstoma (n= 19) or H. muscae (n =4), and others (n = 7) harboured both species. Most of them were also infected with other parasites, including strongylid nematodes (subfamilies Cyathostominae and Strongylinae), bots and/or cestodes; there was no evidence of metazoan parasites in 2 horses. Larvated spirurid eggs were detected in the faeces of 1 of the 30 horses (3.3 %) shown to be infected with Habronema at autopsy. For this set of 46 samples, the PCR assay achieved a diagnostic specificity of 100 % and a sensitivity of approximately 97 % (being able to specifically detect as little as approximately 0.02 fg of Habronema DNA). The specificity of the assay was also tested using a panel of control DNA samples representing horse, the gastric spirurid Draschia megastoma and 26 other species of parasites from the alimentary tract of the horse. H. microstoma, H. muscae and D. megastoma could be readily differentiated from one another based on the sizes of their specific amplicons in the PCR. The results of this study showed that the performance of the PCR for the diagnosis of gastric habronemosis was similar to that of autopsy but substantially better than the traditional coprological examination procedure used. The ability to specifically diagnose gastric habronemosis in equids should have important implications for investigating the epidemiology and ecology of H. microstoma and H. muscae.
|
|
Abbruzzetti, S., Viappiani, C., Small, J. R., Libertini, L. J., & Small, E. W. (2001). Kinetics of histidine deligation from the heme in GuHCl-unfolded Fe(III) cytochrome C studied by a laser-induced pH-jump technique. J Am Chem Soc, 123(27), 6649–6653.
Abstract: We have developed an instrumental setup that uses transient absorption to monitor protein folding/unfolding processes following a laser-induced, ultrafast release of protons from o-nitrobenzaldehyde. The resulting increase in [H(+)], which can be more than 100 microM, is complete within a few nanoseconds. The increase in [H(+)] lowers the pH of the solution from neutrality to approximately 4 at the highest laser pulse energy used. Protein structural rearrangements can be followed by transient absorption, with kinetic monitoring over a broad time range (approximately 10 ns to 500 ms). Using this pH-jump/transient absorption technique, we have examined the dissociation kinetics of non-native axial heme ligands (either histidine His26 or His33) in GuHCl-unfolded Fe(III) cytochrome c (cyt c). Deligation of the non-native ligands following the acidic pH-jump occurs as a biexponential process with different pre-exponential factors. The pre-exponential factors markedly depend on the extent of the pH-jump, as expected from differences in the pK(a) values of His26 and His33. The two lifetimes were found to depend on temperature but were not functions of either the magnitude of the pH-jump or the pre-pulse pH of the solution. The activation energies of the deligation processes support the suggestion that GuHCl-unfolded cyt c structures with non-native histidine axial ligands represent kinetic traps in unfolding.
|
|
Elsaesser, F., Klobasa, F., & Ellendorff, F. (2001). ACTH stimulation test for the determination of salivary cortisol and of cortisol responses as markers of the training status/fitness of warm-blooded sports horses]. Dtsch Tierarztl Wochenschr, 108(1), 31–36.
Abstract: Previous work (Marc et al., 2000) suggested that plasma cortisol responses to treadmill exercise or ACTH injection are a reliable marker for performance evaluation in warmblood horses. For practical purposes blood sample collections and treadmill exercise tests are somewhat troublesome and time consuming. The goal of this study was thus to evaluate the use of saliva for cortisol determination (by direct EIA) as a marker for performance and to investigate the reliability and repeatability of plasma cortisol responses to a single i.v. injection of ACTH (50 micrograms or 250 micrograms). Furthermore, the effect of training horses for 8 weeks 3 times per week covering the same distance (increasing from 3.5 km during the first week to 8 km during the last week) either by trotting (approximately 240 m/min) or by cantering (375 m/min) was investigated. For this purpose initially ten four-year-old Hannovarian geldings, all reared in the same State stud, were used. Mean overall correlation between salivary cortisol and plasma cortisol concentrations was 0.64 when samples of various points of time were used. However, in spite of attempts to standardize saliva sample collection, correlation between salivary cortisol levels and plasma cortisol levels at distinct points of time in different tests were low and significant (r = 0.85, p < 0.02) only in one test. Thus, salivary cortisol measurements for diagnostic purposes are not reliable or useful. The repeatability of plasma cortisol responses to ACTH for untrained and trained horses were r = 0.86 and r = 0.8 respectively (p < or = 0.01 and p < or = 0.05 respectively). Training horses either by trotting or cantering did not affect the cortisol response either to treadmill exercise or to stimulation by ACTH. It is concluded that the relationship between salivary cortisol levels and plasma cortisol levels is not close enough to allow the use of salivary cortisol determination as marker of the training status/fitness of horses. The repeatability of the cortisol response to ACTH is similar to the cortisol response to treadmill exercise. Based on plasma cortisol responses to ACTH or treadmill exercise training horses by cantering at low speed is not superior to training by trotting for the fitness of horses.
|
|
Turner, J. W. J., Liu, I. K., & Kirkpatrick, J. F. (1996). Remotely delivered immunocontraception in free-roaming feral burros (Equus asinus). J Reprod Fertil, 107(1), 31–35.
Abstract: Regulation of local overpopulations of free-roaming feral equids is in demand worldwide for ecological balance and habitat preservation. Contraceptive vaccines have proven effective in feral horses, which breed seasonally, but no data are available for equids such as the burro, which is reproductively active all year round. In the present study, 27 individually identified female feral burros (Equus asinus) roaming free in Virgin Islands National Park (St John, US Virgin Islands; Lesser Antilles) were remotely treated with pig zonae pellucidae (PZP) vaccine. Between January and May, 16 burros were darted with a 1 ml emulsion of PZP plus Freund's adjuvant. Ten to twelve months later each treated burro was given a single booster injection of PZP plus adjuvant to maintain contraception through a second year. Eleven adult untreated jennies served as controls. Beginning one year after initial vaccination, these burros were monitored for pregnancy and foal production. Collection of data to determine treatment effect was not begun until 12 months after initial treatment to ensure that pregnancies existing before vaccination were not included. Pregnancy was assessed using previously validated methods for steroid metabolite measurement in fresh faecal samples. None of the PZP-treated burros produced foals between 0 and 12 months after the last inoculation. One PZP-treated burro tested positive for pregnancy at 10 months after the final inoculation. During this same period, six of 11 untreated burros tested pregnancy-positive, and four were observed with foals. There was no difference in pregnancy rates among treated, control and randomly sampled jennies between 12 and 24 months after the last inoculation. The results demonstrate that, in free-roaming feral burros that are reproductively active all year round: (1) burros can be accessed for remotely delivered PZP vaccination; (2) PZP contraception is effective; (3) PZP contraception is reversible; and (4) pregnancy can be reliably detected by faecal steroid analysis.
|
|
Uzawa, T., Akiyama, S., Kimura, T., Takahashi, S., Ishimori, K., Morishima, I., et al. (2004). Collapse and search dynamics of apomyoglobin folding revealed by submillisecond observations of alpha-helical content and compactness. Proc. Natl. Acad. Sci. U.S.A., 101(5), 1171–1176.
Abstract: The characterization of protein folding dynamics in terms of secondary and tertiary structures is important in elucidating the features of intraprotein interactions that lead to specific folded structures. Apomyoglobin (apoMb), possessing seven helices termed A-E, G, and H in the native state, has a folding intermediate composed of the A, G, and H helices, whose formation in the submillisecond time domain has not been clearly characterized. In this study, we used a rapid-mixing device combined with circular dichroism and small-angle x-ray scattering to observe the submillisecond folding dynamics of apoMb in terms of helical content (f(H)) and radius of gyration (R(g)), respectively. The folding of apoMb from the acid-unfolded state at pH 2.2 was initiated by a pH jump to 6.0. A significant collapse, corresponding to approximately 50% of the overall change in R(g) from the unfolded to native conformation, was observed within 300 micros after the pH jump. The collapsed intermediate has a f(H) of 33% and a globular shape that involves >80% of all its atoms. Subsequently, a stepwise helix formation was detected, which was interpreted to be associated with a conformational search for the correct tertiary contacts. The characterized folding dynamics of apoMb indicates the importance of the initial collapse event, which is suggested to facilitate the subsequent conformational search and the helix formation leading to the native structure.
|
|
Ballew, R. M., Sabelko, J., & Gruebele, M. (1996). Direct observation of fast protein folding: the initial collapse of apomyoglobin. Proc. Natl. Acad. Sci. U.S.A., 93(12), 5759–5764.
Abstract: The rapid refolding dynamics of apomyoglobin are followed by a new temperature-jump fluorescence technique on a 15-ns to 0.5-ms time scale in vitro. The apparatus measures the protein-folding history in a single sweep in standard aqueous buffers. The earliest steps during folding to a compact state are observed and are complete in under 20 micros. Experiments on mutants and consideration of steady-state CD and fluorescence spectra indicate that the observed microsecond phase monitors assembly of an A x (H x G) helix subunit. Measurements at different viscosities indicate diffusive behavior even at low viscosities, in agreement with motions of a solvent-exposed protein during the initial collapse.
|
|
McCutcheon, L. J., & Geor, R. J. (2000). Influence of training on sweating responses during submaximal exercise in horses. J Appl Physiol, 89(6), 2463–2471.
Abstract: Sweating responses were examined in five horses during a standardized exercise test (SET) in hot conditions (32-34 degrees C, 45-55% relative humidity) during 8 wk of exercise training (5 days/wk) in moderate conditions (19-21 degrees C, 45-55% relative humidity). SETs consisting of 7 km at 50% maximal O(2) consumption, determined 1 wk before training day (TD) 0, were completed on a treadmill set at a 6 degrees incline on TD0, 14, 28, 42, and 56. Mean maximal O(2) consumption, measured 2 days before each SET, increased 19% [TD0 to 42: 135 +/- 5 (SE) to 161 +/- 4 ml. kg(-1). min(-1)]. Peak sweating rate (SR) during exercise increased on TD14, 28, 42, and 56 compared with TD0, whereas SRs and sweat losses in recovery decreased by TD28. By TD56, end-exercise rectal and pulmonary artery temperature decreased by 0.9 +/- 0.1 and 1.2 +/- 0.1 degrees C, respectively, and mean change in body mass during the SET decreased by 23% (TD0: 10.1 +/- 0.9; TD56: 7.7 +/- 0.3 kg). Sweat Na(+) concentration during exercise decreased, whereas sweat K(+) concentration increased, and values for Cl(-) concentration in sweat were unchanged. Moderate-intensity training in cool conditions resulted in a 1.6-fold increase in sweating sensitivity evident by 4 wk and a 0.7 +/- 0.1 degrees C decrease in sweating threshold after 8 wk during exercise in hot, dry conditions. Altered sweating responses contributed to improved heat dissipation during exercise and a lower end-exercise core temperature. Despite higher SRs for a given core temperature during exercise, decreases in recovery SRs result in an overall reduction in sweat fluid losses but no change in total sweat ion losses after training.
|