Selection of rabbit CD4- CD8- T cell receptor-gamma/delta cells by in vitro transformation with human T lymphotropic virus-I.

In vitro transformation of rabbit peripheral blood mononuclear cells (PBMC) with human T lymphotropic virus-I (HTLV)-infected human or rabbit cells resulted in CD4- CD8- cell lines, some of which caused acute leukemia when injected into rabbits. Structural analyses of the proviruses from cell lines with diverse pathogenic effects provided no clear correlation with lethality. The rabbit lines were provisionally designated T cells because they express interleukin 2R (IL-2R) and CD5 and lack surface immunoglobulin, but none express functional T cell receptor (TCR) alpha or beta transcripts. A more detailed characterization of the HTLV-I-infected cells was required to determine cell lineage and its potential influence on pathogenic consequences. Probes for rabbit TCR gamma and delta genes were derived and used to detect gamma and delta TCR RNA transcripts, identifying the in vitro transformed lines as gamma/delta T cells. CD4+ and CD8+ lines were derived from PBMC of HTLV-I-infected rabbits and CD4+ TCR-alpha/beta HTLV-I lines were derived from rabbit thymus, eliminating the possibility that the HTLV-I isolates used here transform only CD4- CD8- TCR-gamma/delta cells. The percentage of gamma/delta cells in rabbit PBMC is relatively high (23% in adult rabbits); this with diminution of CD4+ and CD8+ cells in IL-2-supplemented PBMC or thymocyte cultures may account for selection of rabbit HTLV-I-infected gamma/delta T cell lines in vitro. The availability of well-characterized T cell lines with diverse in vivo effects in the rabbit HTLV-I disease model allows evaluation of roles played by cell type in HTLV-I-mediated disease.

Autoradiography in fetal golden hamsters treated with tritiated diethylnitrosamine.

Tritiated diethylnitrosamine was administered to female Syrian golden hamsters on each of the last 4 days (days 12-15) of pregnancy. The distribution of bound radioactivity was monitored by light microscopic autoradiography of fetal tracheas and livers, the placentas, and the maternal livers. In the trachea, the fetal target organ, bound radioactivity was restricted to the respiratory epithelium, where diethylnitrosamine-induced tracheal tumors arise. Mucous cells and nonciliated stem cells were identified as the principal sites of binding; other cell types within the tracheal epithelium contained only small amounts of bound radioactivity. The level of binding observed in the fetal trachea increased steadily from day 12 to day 15, which correlated well with the levels of differentiation of this tissue during this period. This observation also agrees with the previously reported observation that tumor incidence increases from 40 to 95% in Syrian golden hamsters between days 12 and 15.

Electron microscopic autoradiography of the pancreas in the hamster treated with tritiated N-nitroso-2,6-dimethylmorpholine.

Syrian golden hamsters were given a single dose of [3H]-N-nitroso-2,6-dimethylmorpholine and killed 8 hr later. The pancreas was processed for electron microscopic autoradiography to detect binding of radioactivity to cellular constituents. The pancreatic acinar cells and duct epithelia were found to be labeled, while islet cells, centroacinar cells, and all nonepithelial elements were not. Acinar cells active in secreting zymogen had a high concentration of grains over the zymogen granules and the rough endoplasmic reticulum. Their nonsecreting counterparts contained abundant bound material in the nuclei and rough endoplasmic reticulum. Labeling was lower in the duct epithelia than in acinar cells, with the majority of grains associated with the heterochromatin. Our findings suggest that the acinar cells are the principle site of metabolic activation in this organ.

Autoradiographic study of the distribution of bound radioactivity in the respiratory tract of Syrian hamsters given N-[3H]nitrosodiethylamine.

Adult Syrian golden hamsters received a single intragastric dose of N-[3H]nitrosodiethylamine. Their tracheas, extrapulmonary stem bronchi, and lungs were processed for high-resolution light-microscopic autoradiography to monitor the distribution of bound radioactivity. In the trachea and extrapulmonary stem bronchi, mucous cells contained the most bound radioactivity, while in the lobar and segmental bronchi and bronchioles, Clara cells were the major site of binding. In conjunction with earlier conducted studies on the pathogenesis of N-[3H]nitrosodiethylamine-induced respiratory tract tumors, these findings indicate that metabolic competence and a preexisting capacity for proliferation are important factors in determining the target cell types of this compound.

Replication of HIV type 1 in rabbit cell lines is not limited by deficiencies in tat, rev, or long terminal repeat function.

HIV-1 infection has been documented in rabbits, but infection proceeds slowly in this species. Human and rabbit cell lines were compared in order to identify barriers to efficient HIV-1 infection of rabbit cells. A direct comparison of human and rabbit CD4 as receptor for HIV-1 indicated that the rabbit CD4 homolog did not function well even when expressed by human cells. Examination of viral RNA production indicated that the major HIV transcripts were produced in HIV-infected rabbit cells, but were present at levels significantly lower than those found for human cells. Ability of HIV-1 LTRs to direct protein expression in human and rabbit cells was compared using gene constructs with the chloramphenicol acetyltransferase (cat) gene flanked by HIV-1 LTRs. Chloramphenicol acetyltransferase protein expression was equivalent in rabbit and human cell lines transfected with the HIV-1/CAT constructs and cotransfections with the HIV-1 tat gene led to similar increases in CAT expression. Subsequent transfections with an infectious molecular HIV clone yielded approximately equal levels of HIV protein expression in rabbit and human cell lines, suggesting that major barriers to virus production in rabbit lines exist at steps prior to transcription of the viral genome. Because HTLV-I replicates with high efficiency in rabbit cells, a chimeric virus clone was constructed consisting of the 5' portion of HIV-1 through the nef coding sequence followed by the 3' HTLV-I LTR. Transfection of most rabbit cell lines with the chimera produced levels of p24gag protein higher than those transfected with the parent HIV-1 clone. By contrast, the unmodified HIV clone replicated more efficiently in all human cell lines tested.

A macaque adherent cell line that expresses human CD4 is susceptible to SIV: utility for assessing neutralizing antibody.

A macaque CD4 + adherent cell line was generated by stable expression of the human CD4 gene in a rhesus macaque mammary tumor cell line, CMMT. The resulting cell line CMMT/CD4 expressed surface CD4 and was sensitive to infection by a wide range of isolates of simian immunodeficiency virus (SIV) of different subgroups, but was not susceptible to infection with HIV-1. The CMMT/CD4 cell line was used to develop a microassay for measurement of neutralizing antibody in plasma of SIV-infected or immunized animals. Single infected cells could be detected in a monolayer of CMMT/CD4 by immunoperoxidase and a 90% reduction in the number of positive cells was used as a measure of neutralizing activity of two-fold plasma dilutions. This assay had comparable sensitivity to methods based upon detecting a reduction in reverse transcriptase activity of SIV, reduction of viral antigen, or inhibition of cytopathic effect.

The application of hybridoma technology to the study of bovine immunoglobulins.

Studies are described in which hybridoma technology is used to produce a variety of reagents for the characterization and manipulation of the bovine humoral immune system. Selected members of a set of murine monoclonal antibodies (MAb) specific for each of four major isotypes of bovine Ig constant regions, one specific for anti-bovine Ig constant regions as well as one specific for anti-bovine light chains are discussed. Interspecific fusion of bovine lymphocytes with the established mouse cell line, SP2/0 was used to produce a collection of stable hybridomas among which were found secretors of bovine IgG1, IgG2, IgM, IgA and bovine light chain. Interspecific fusion of SP2/0 with lymphocytes from a multiparous Holstein four days post immunization with Streptococcus agalactiae yielded MAb with specificity for the immunizing antigen. One of these hybridomas, LHRB 19.17, which displayed a particularly stable secretory phenotype, was used as an immunogen for the production of a library of murine monoclonal anti-idiotype antibodies. Competitive antigen binding analysis showed that 15 of the 24 anti-LHRB 19.17 idiotype antibodies isolated blocked the binding of the idiotype to its nominal antigen and so were candidates for evaluation as antigen mimics. Some of the ways in which monoclonal anti-idiotypes in particular, and monoclonal in general, might be of use in problems of animal disease are discussed.

A district epilepsy service, with community-based specialist liaison nurses and guidelines for shared care.

In order to improve epilepsy services, Doncaster Hospital doctors and general practitioners agreed upon a collaborative approach. A district epilepsy service was inaugurated around specialist services based in a hospital clinic. Guidelines were produced to clarify respective roles and to assist non-specialist hospital doctors and general practitioners in epilepsy management. A novel feature of the new service was the appointment of a community-based specialist liaison nurse. The new service is perceived to be effective, and the contribution of the specialist nurse to represent an important advance in epilepsy care. It has been possible to supervise complicated changes in medication successfully at home with a reduction in the need to attend clinic or GP. Counselling and support for patients and families, previously lacking, has been a major contribution. Evaluation to determine the extent to which the service meets the needs of those in the local population with epilepsy is planned. This document describes the first 5 years of the service and the scope and content of the specialist liaison nurse's role.

Effects of high-dose gentamicin sulfate on neuromuscular blockade in halothane-anesthetized horses.

OBJECTIVE: To evaluate effects of a single high dose of gentamicin on neuromuscular function in horses anesthetized with halothane. ANIMALS: 6 healthy adult horses. PROCEDURE: Halothane-anesthetized horses were positioned in left lateral recumbency, and the right hind limb was immobilized in a reusable fiberglass cast fixed to a steel frame. The hoof was attached to a force transducer, and resting tension of 0.93 +/- 0.16 kg was maintained. A supramaximal train-of-four stimulus of 2 Hz for a duration of 0.25 millisecond was applied to the superficial peroneal nerve every 20 seconds by a square-wave stimulator. The force of the evoked digital extensor tension was recorded to determine first muscle twitch tension, compared with the baseline value (T1%) and the ratio of the force of the fourth twitch to the first twitch (T4/T1). Data were recorded at 5, 10, 15, 30, and 60 minutes after i.v. administration of vehicle or gentamicin (6 mg/kg of body weight). RESULTS: There was a significant (P = 0.04) treatment-time interaction for the effect of gentamicin on T1%; T1% associated with vehicle decreased from 100% to 92% during the 60- minute study period, but no decrease was associated with gentamicin. For T4/T1, there was no significant effect of treatment or time or treatment-time interaction between gentamicin and vehicle. CONCLUSIONS: Gentamicin did not cause a decrease in initial muscular strength, nor did it impair the muscles' ability to sustain strength. CLINICAL RELEVANCE: A single high dose of gentamicin does not cause significant neuromuscular blockade when administered alone to healthy horses anesthetized with halothane.

Effects of postoperative peritoneal lavage on pharmacokinetics of gentamicin in horses after celiotomy.

OBJECTIVE: To evaluate the effect of peritoneal lavage on pharmacokinetics of gentamicin sulfate in healthy horses after experimental celiotomy. ANIMALS: 13 clinically normal horses. PROCEDURE: Horses were randomly assigned to control or experimental groups. All horses received gentamicin (6.6 mg/kg of body weight, IV, q 24 h) before surgery, underwent experimental abdominal surgery, and had abdominal drains placed percutaneously. Horses of the experimental group received postoperative peritoneal lavage; horses of the control group did not receive peritoneal lavage. The day after surgery, 24 hours after the preoperative dose of gentamicin, a second dose of gentamicin was administered. Three and 15 hours after this second dose of gentamicin, horses of the experimental group received peritoneal lavage. Venous blood was obtained, for determination of concentration of gentamicin, immediately before and at specified intervals during the 24-hour period after the second dose of gentamicin. RESULTS: There were no differences in any of the pharmacokinetic values of gentamicin between horses of the control and experimental groups. CONCLUSIONS: Peritoneal lavage had no effect on pharmacokinetics of gentamicin in healthy horses after abdominal surgery, in which localized nonseptic peritonitis was induced. CLINICAL RELEVANCE: Peritoneal lavage in horses with localized nonseptic peritonitis or for the prevention of intra-abdominal adhesions should not necessitate alteration of the dosage of gentamicin to maintain predictable serum concentrations.

Tension band plating of an olecranon fracture in a bull.

A 1,018-kg (2,240-lb), 6-year-old bull was admitted for evaluation of a left forelimb lameness of 7 days' duration. Radiographic views of the proximal portion of the left forelimb revealed a type-III olecranon fracture that was minimally displaced. Conservative treatment, including stall confinement and administration of anti-inflammatory drugs, was attempted. Subsequent radiographic views, 27 days later, revealed displacement of the fracture with minimal callus formation. Open reduction and internal fixation was performed by placing a 4.5-mm, 10-hole broad dynamic-compression on the caudal surface of the olecranon. Good anatomic reduction was achieved, returning the bull to full weightbearing. Eight months following surgery the owners reported the bull was sound at a walk and able to breed cows.

Incomplete urethral obstruction associated with dilatation of the urethra, cystitis, and pyelonephritis in a Vietnamese pot-bellied pig.

A 2-year-old castrated male vietnamese pot-bellied pig examined because of stranguria was found to have severe dilatation and inflammation of the pelvic portion of the urethra accompanied by cystitis and pyelonephritis. The pig had apparently had chronic inflammation of the urinary tract that eventually progressed to incomplete obstruction of the urethra. However, the initial cause of the urinary tract inflammation could not be determined. Possible causes included chronic bacterial infection, traumatic injury during urethral catheterization, and injury to the urethral mucosa secondary to extreme acidification of the urine.

Treatment response and athletic outcome of foals with tarsal valgus deformities: 39 cases (1988-1997).

OBJECTIVE: To evaluate the response to various treatments and long-term outcome of foals with tarsal valgus deformities. DESIGN: Retrospective study. ANIMALS: 39 foals with tarsal valgus deformities. PROCEDURE: Data collected from medical records, included signalment, history, reason for admission, and clinical findings. Radiographic views of the tarsus were evaluated for incomplete ossification of tarsal bones and were classified as normal in appearance, type-I incomplete ossification, or type-II incomplete ossification. Treatment and athletic outcome were documented for each foal. RESULTS: Radiographic assessment revealed that 22 of 39 foals (56%) had concomitant tarsal valgus deformities and incomplete ossification of the tarsal bones. Eight of 19 foals with tarsal valgus deformities that were treated with periosteal stripping responded favorably. Foals < or = 60 days old were significantly more likely to respond to periosteal stripping than older foals. Five of 8 foals with tarsal valgus deformities that were treated with growth plate retardation responded favorably. Eleven of 21 foals with long-term follow-up performed as intended. Compared with foals with type-II incomplete ossification, foals with tarsal bones that had a normal radiographic appearance or type-I incomplete ossification were significantly more likely to perform as intended. CONCLUSIONS AND CLINICAL RELEVANCE: Foals with tarsal valgus deformities should have lateromedial radiographic views of the tarsus obtained to assess the tarsal bones for incomplete ossification, which will affect athletic outcome. Because foals with type-II incomplete ossification of the tarsal bones respond poorly to periosteal stripping alone, treatment by growth-plate retardation is recommended.

Deep digital flexor tenotomy as a treatment for chronic laminitis in horses: 35 cases (1988-1997)

OBJECTIVE: To determine long-term prognosis for horses with laminitis treated by deep digital flexor (DDF) tenotomy and to identify factors affecting success of the surgical procedure. DESIGN: Retrospective study. ANIMALS: 35 horses with laminitis treated by DDF tenotomy between 1988 and 1997. PROCEDURE: Information was obtained from individual medical records and follow-up telephone interviews with owners and referring veterinarians. Cumulative proportions of horses that survived 6 months and 2 years after tenotomy were determined. Effect of Obel grade of lameness on 6-month and 2-year survival and effect of distal phalangeal rotation on survival and future performance were evaluated by chi 2 analysis. Body weights of horses that survived > or = 2 years were compared with those of horses that survived < 2 years by ANOVA. RESULTS: 27 of the 35 (77%) horses survived > or = 6 months, and 19 of 32 (59%) survived > 2 years. Obel grade of lameness and body weight at time of surgery had no effect on 6-month or 2-year survival. Degree of distal phalangeal rotation had no effect on 2-year survival or the ability of horses to be used for light riding. Twenty-two of the 30 (73%) owners interviewed indicated they would have the procedure repeated on their horses given similar circumstances. CLINICAL IMPLICATIONS: DDF tenotomy is a viable alternative for horses with laminitis refractory to conventional medical treatment. In some instances, the procedure may be effective in returning horses to light athletic use.

Rectal stricture in an ostrich (Struthio camelus).

Exploratory celiotomy of an 18-mo-old female ostrich (Struthio camelus) with anorexia, lethargy, and constipation of 5 days' duration showed mesenteric volvulus and a focal narrowing of the rectum approximately 28 cm cranial to the cloaca. The prognosis was poor and the animal was euthanized. Necropsy revealed a fibrinonecrotic proctitis and a 3-cm-long circumferential stricture of the rectum. Histologically, the rectal wall at the stricture was deeply effaced by fibrovascular connective tissue with vascular thrombosis and necrotizing vasculitis, and it resembled the lesions in feeder pigs with fibrinonecrotic colitis.

Evidence for disparity detecting neurones in the human visual system.

1. It is known that adaptation to a grating pattern causes a rise in the contrast threshold for test gratings of similar spatial frequency and orientation.2. We find this after-effect also to be disparity-specific. Adaptation to a grating at zero horizontal disparity (at the same distance as the fixation point) causes a greater elevation of threshold for patterns at the same disparity than for those at nearby disparities, closer or more distant than the fixation point.3. Adaptation to a grating at some disparity other than zero causes a disparity-specific elevation of threshold centred on the adapting disparity.4. This finding also applies if the observer adapts to a grating but single bright bars are used as the test stimuli.5. The disparity-specific ;tuning curves' revealed by these techniques are quite broad, having a half-width at half-amplitude of several min of disparity.6. Adaptation to a grating at one disparity causes an apparent change in the distance of test gratings at nearby disparities.7. We compare these psychophysical experiments with the properties of disparity-selective binocular neurones in the visual cortex of cats and monkeys.

A morphometric study of the pulmonary Clara cell in normal and nitrosoheptamethyleneimine-treated European hamsters.

Previous studies have identified the bronchial Clara cell as the progenitor of the lung tumors induced in European hamsters by nitrosoheptamethyleneimine (NHMI). Using stereological methods, we compared the ultrastructure of Clara cells from untreated animals and lung tumor cells, induced by NHMI, in the European hamster. The composition and volume of the cytoplasm was significantly altered in the tumor cells whereas the nuclei did not show any measurable changes in size or structure when compared with the controls.