Specific contrast ultrasound using sterically stabilized microbubbles for early diagnosis of thromboembolic disease in a rabbit model

Specific contrast ultrasound is widely applied in diagnostic procedures on humans but remains underused in veterinary medicine. The objective of this study was to evaluate the use of microbubble-based contrast for rapid ultrasonographic diagnosis of thrombosis in small animals, using male New Zealand white rabbits (average weight about 3.5 kg) as a model. It was hypothesized that the use of microbubble-based contrast agents will result in a faster and more precise diagnosis in our model of thrombosis. A pro-coagulant environment had been previously established by combining endothelial denudation and external vessel wall damage. Visualization of thrombi was achieved by application of contrast microbubbles [sterically stabilized, phospholipid-based microbubbles filled with sulfur hexafluoride (SF₆) gas] and ultrasonography. As a result, rapid and clear diagnosis of thrombi in aorta abdominalis was achieved within 10 to 30 s (mean: 17.3 s) by applying microbubbles as an ultrasound contrast medium. In the control group, diagnosis was not possible or took 90 to 180 s. Therefore, sterically stabilized microbubbles were found to be a suitable contrast agent for the rapid diagnosis of thrombi in an experimental model in rabbits. This contrast agent could be of practical importance in small animal practice for rapid diagnosis of thrombosis.     

Biochemical responses and oxidative stress in Francisella tularensis infection: a European brown hare model

Background

The aim of the present study was to investigate biochemical and oxidative stress responses to experimental F. tularensis infection in European brown hares, an important source of human tularemia infections.

Methods

For these purposes we compared the development of an array of biochemical parameters measured in blood plasma using standard procedures of dry chemistry as well as electrochemical devices following a subcutaneous infection with a wild Francisella tularensis subsp. holarctica strain (a single dose of 2.6 × 10⁹ CFU pro toto).

Results

Subcutaneous inoculation of a single dose with 2.6 × 10⁹ colony forming units of a wild F. tularensis strain pro toto resulted in the death of two out of five hares. Plasma chemistry profiles were examined on days 2 to 35 post-infection. When compared to controls, the total protein, urea, lactate dehydrogenase, aspartate aminotransferase and alanine aminotransferase were increased, while albumin, glucose and amylase were decreased. Both uric and ascorbic acids and glutathione dropped on day 2 and then increased significantly on days 6 to 12 and 6 to 14 post-inoculation, respectively. There was a two-fold increase in lipid peroxidation on days 4 to 8 post-inoculation.

Conclusions

Contrary to all expectations, the present study demonstrates that the European brown hare shows relatively low susceptibility to tularemia. Therefore, the circumstances of tularemia in hares under natural conditions should be further studied.     

Effect of the gonadotrophins treatment on morphological alterations in ovary and peripheral plasma concentrations of steroid hormones in gilts

The present study was designed to examine the influence of gonadotrophins treatment on the ovarian morphology changes and plasma concentrations of steroid hormones in peripheral blood. The experiment was performed on sexually pubertal gilts (Large White x Landrace) of similar age (7-8 months) and body mass (100-110 kg) with two controlled subsequent estrous cycles. The animals were randomly divided into four groups: two control consisting of pigs with the luteal phase (n = 9, the 10th day of the estrous cycle) and the follicular phase (n = 6, the 20th day of the estrous cycle) and two experimental ones consisting of animals with both mentioned periods (n = 7 and n = 9) treated with gonadotrophins (PMSG and hCG). The gilts in the luteal phase were injected (s.c.) with gonadotrophins at a daily dose of PMSG 400 and hCG 200 IU from the 16th to the 27th day (the 6th day of the next estrous cycle). The gilts in the follicular phase, were injected with the same dose of gonadotrophins but from the 8th to the 19th day of the estrous cycle. Plasma concentrations of P4, A4, T, E1, E2 and metabolite of PGF2 alpha-PGFM were determined by radioimmunoassay (RIA) method. Injections of PMSG and hCG in both experimental groups produced several times enlarged: weight, size and volume of ovaries and alterations in a number of structural elements as compared with those found in the control animals. The morphological elements presented in ovaries: corpora haemorrhagica, corpora lutea, regular and atretic follicles and first of all cysts by distinctly differentiation thickness of the walls are characteristic for cystic ovarian degeneration. Plasma concentrations all determined hormones after gonadotrophins treatment in experimental groups were increased except E1 (insignificant decrease) in luteal phase as compared with those found in the control groups. Statistically significant increase (p < 0.001) in plasma concentrations of P4, A4, and T in both experimental groups and E2 (p < 0.001) in luteal phase were noted. In peripheral plasma concentrations increase of E1 and E2 in follicular phase of the estrous cycle were insignificant.     

Bilateral thyroarytenoid cartilage lateralization and vocal fold excision with mucosoplasty for treatment of idiopathic laryngeal paralysis: 67 dogs (1998-2005)

Objective— To evaluate combined bilateral thyroarytenoid cartilage lateralization, vocal fold excision, and mucosoplasty technique (BTAL) through ventral median laryngotomy for treatment of laryngeal paralysis in dogs.
Design— Retrospective study.
Animals— Dogs (n=67) with laryngeal paralysis.
Methods— Medical records were reviewed for dogs with idiopathic laryngeal paralysis that had BTAL between January 1998 and March 2005. Retrieved data included signalment, history, physical and laryngoscopic examination findings, clinicopathologic tests, and results of recheck examination findings.
Results— BTAL was performed by a single surgeon. Short-term (<6 months) follow-up information was available for 67 dogs and long-term (>12 months) for 40 dogs. Major postoperative complications were surgical failures (13; 7 short term, 6 long term) and aspiration pneumonia (1). Mean recurrence of clinical signs was at 19 weeks (range, 2–30 weeks). Minor complications occurred in 22 (33%) dogs including occasional coughing or gagging, stridorous breathing during exercise, panting, noisy or heavy breathing, and aspiration pneumonia (3 dogs) that did not require hospitalization. All owners reported an improved quality of life and had no regrets with surgical outcome.
Conclusions— BTAL is seemingly an effective procedure for treatment of laryngeal paralysis.
Clinical Relevance— BTAL is associated with a low incidence of aspiration pneumonia; however, there is substantial risk of recurrence of clinical signs associated with narrowing of the glottis. Consequently, unilateral arytenoid lateralization currently represents the accepted approach to the treatment of laryngeal paralysis.     

The impact of zearalenone on the level of the selected estrogens in blood serum of sexually immature gilts

The aim of the study was to evaluate the influence of low dose (LOEL - lowest observed effect level) of zearalenone (200 microg/kg b.w.), applied per os for 7 days (short-term intoxication), on sexual behavior, concentration of the examined xenobiotic and its metabolite and selected estrogens in sexually immature gilts: ovariohysterectomised (group D1) and intact (group D2) animals. Clinical signs of oestrus (reddening, oedema and hyperaemia of the vulva and serorhoea from the reproductive tract--lack of standing reflex) were obserwed in group D1 on day 6 and in group D2 on day 4 of the experiment. Laboratory analyses of blood plasma were carried out determine the presence of zearalenone and alpha-zearalenole. They revealed an increase in the level of alpha-zearalenol before the oestrus, decrease in total amount of both examined substances on day when the oestrus appeared and increase in the level of both examined xenobiotics in the post oestrus period together with the higher share of zearalenone. Medium concentrations of estrone and estradiol within the borders of method determination in the majority of periods examined. Higher levels of estrone (32.0 pg/ml) were found on day 4, in the group D2 and estradiol (6.5 pg/ml) on day 6 in the D2 group. The presents study revealed that zearalenone applied per os at LOEL dose causes the incidence of apparent sexual readiness (without standing reflex) in sexually immature gilts with the somatically immature reproductive system.     

Cryptosporidium scrofarum n. sp. (Apicomplexa: Cryptosporidiidae) in domestic pigs (Sus scrofa)

We describe the morphological, biological, and molecular characteristics of Cryptosporidium pig genotype II and propose the species name Cryptosporidium scrofarum n. sp. to reflect its prevalence in adult pigs worldwide. Oocysts of C. scrofarum are morphologically indistinguishable from C. parvum, measuring 4.81–5.96 μm (mean = 5.16) × 4.23–5.29 μm (mean = 4.83) with a length to width ratio of 1.07 ± 0.06 (n = 400). Oocysts of C. scrofarum obtained from a naturally infected pig were infectious for 8-week-old pigs but not 4-week-old pigs. The prepatent period in 8-week-old Cryptosporidium-naive pigs was 4–6 days and the patent period was longer than 30 days. The infection intensity of C. scrofarum in pigs was generally low, in the range 250–4000 oocysts per gram of feces. Infected pigs showed no clinical signs of cryptosporidiosis and no pathology was detected. Cryptosporidium scrofarum was not infectious for adult SCID mice, adult BALB/c mice, Mongolian gerbils (Meriones unguiculatus), southern multimammate mice (Mastomys coucha), yellow-necked mice (Apodemus flavicollis), or guinea pigs (Cavia porcellus). Phylogenetic analyses based on small subunit rRNA, actin, and heat shock protein 70 gene sequences revealed that C. scrofarum is genetically distinct from all known Cryptosporidium species.     

Acrosomal and viability status of bovine spermatozoa evaluated by two staining methods

Artificial insemination with frozen-thawed spermatozoa is commonly used in cattle breeding. A simple and fast procedure is needed for routine evaluation of the acrosomal status of frozen-thawed bovine sperm. Therefore, the purpose of this study was to test two staining procedures used to determine the viability and integrity of acrosome of frozen-thawed bovine spermatozoa. Double staining and Hoechst/FITC-Pisum sativum agglutinin (FITC-PSA) labelling were tested for evaluating the viability and acrosome reaction induced by calcium ionophore of bull spermatozoa. In our experiments no significant differences were detected in the frequency of acrosome-reacted sperm either by double staining (37.98%) or by FITC-PSA labelling (39.33%). The viability of sperm stained by the double staining method was 67.17%, and a higher portion of viable sperm (82.67%) was observed by staining with the Hoechst procedure (P < 0.01). On the basis of the results obtained it is concluded that both methods can be used for detecting the acrosome reaction of frozen-thawed bovine spermatozoa.     

Lipopolysaccharide and cytokines modulate leukotriene (LT)B4 and LTC4 production by porcine endometrial endothelial cells

Uterine inflammatory response is mediated by inflammatory mediators including eicosanoids and cytokines produced by immune and endometrial cells. Interactions between lipopolysaccharide (LPS) and cytokines, and leukotrienes (LTs) in endothelium, important for the host defence during the inflammation, are unknown. We studied the effect of LPS, tumour necrosis factor (TNF)‐α, interleukin (IL)‐1β, IL‐4 and IL‐10 on 5‐lipooxygenase (5‐LO), LTA₄ hydrolase (LTAH) and LTC₄ synthase (LTCS) mRNA and protein expression, LTB₄ and LTC₄ release from porcine endometrial endothelial cells, and cell viability. For 24 hr, cells were exposed to LPS (10 or 100 ng/ml of medium) and cytokines (each 1 or 10 ng/ml). 5‐LO mRNA/protein expression augmented after incubation with larger doses of LPS, TNF‐α, IL‐4 and IL‐10 and smaller dose of IL‐1β. Larger dose of TNF‐α, smaller doses of LPS and IL‐1β and both doses of IL‐10 increased LTAH mRNA/protein expression. LTAH protein content was up‐regulated by larger dose of LPS, but it was reduced in response to both doses of IL‐4. LTCS mRNA expression was elevated by larger doses of LPS, IL‐4 and IL‐10 or both doses of TNF‐α and IL‐1β. LTCS protein level increased after treatment with both doses of IL‐1β, IL‐4 and IL‐10, smaller dose of LPS and larger dose of TNF‐α. Both doses of LPS and larger doses of TNF‐α and IL‐10 increased LTB₄ release. LPS, IL‐1β and IL‐10 at smaller doses, or TNF‐α and IL‐4 at larger doses stimulated LTC₄ release. Smaller doses of TNF‐α and IL‐1β or both doses of IL‐4 enhanced the cell viability. This work provides new insight on the participation of LPS, TNF‐α, IL‐1β, IL‐4 and IL‐10 in LTB₄ and LTC₄ production/release from porcine endometrial endothelial cells, and the effect of above factors on these cells viability. The used cellular model gives the possibility to further establish the interactions between inflammatory mediators.     

Effects of α-lipoic and ascorbic acid on the muscle and brain fatty acids and antioxidant profile of the South American pacu Piaractus mesopotamicus

The effects of dietary α-lipoic acid (LA) and vitamin C on the fatty acid (FA) composition in the brain and muscle and vitamins E and C levels in the brain were studied in the fish Piaractus mesopotamicus. A two-factorial design, where diets were devoid or supplemented with ascorbate (500 mg AA kg^− 1) and/or lipoic acid (1000 mg kg^− 1), was used. The levels of eicosapentaenoic acid (20:5n − 3, EPA) increased (P < 0.01) in muscle polar lipids (PL) in LA groups (6.93% ± 0.43 vs. 5.83% ± 0.40 and 6.68% ± 0.53 vs. 6.00% ± 0.39), and the same trend was also seen in the brain, however not significant. These changes are suggested to be caused by a change in lipid metabolism rather than being a direct effect of protection by LA against lipid peroxidation. No interaction of vitamin C and LA neither effects of LA on vitamin E (15.1-19.2 mg α-tocopherol g^− 1 tissue) or vitamin C (total AA, 41.7-89.8 μg g^− 1 tissue) in brain was detected.