Herding the U.S. cattle industry toward a paradigm shift in parasite control.

Contemporary management of nematode parasitism in cattle relies heavily on a single class of drugs, the macrocyclic lactones (MLs). The potency and convenience of the MLs, along with the low cost of generic formulations, have largely supplanted the need for critical thinking about parasite control, and rote treatment has become the default 'strategy'. This approach to parasite control has exerted substantial pressure to select populations of nematodes that can survive recommended dosages of ML products. Although macrocyclic lactones have been available for over 30 years, putative ML resistance in U.S. cattle was not reported until fairly recently. This pattern begs the question, "Is this a new, emergent problem, or an old issue that is finally commanding some attention?" The implications of bovine anthelmintic resistance should stimulate a paradigm shift for U.S. cattle producers and their advisors. However, there are significant obstacles to changes in current thinking. It is anticipated that cattle producers will be extremely reluctant to abandon historical practices unless they can be convinced of the value of alternatives that are communicated through targeted education, practical demonstrations, economic analyses, and scientific evidence. Historically, the management advice of practitioners has not relied strongly on parasite epidemiology, and practitioners may not have the knowledge to implement evidence-based recommendations. Pharmaceutical companies could play a significant role in helping to shape and shift the thinking about sustainable use of anthelmintics. However, their primary responsibility is to stockholders, and they have strong economic incentives for maintaining the status quo. It is complicated and difficult to change attitudes and practices, and it will take more than logic or fear to shift the parasite control paradigm in the U.S. cattle industry. Achieving that goal will require collaboration among stakeholders, a consistent, straightforward and understandable message about resistance, and recommendations that are practical as well as effective. But if we hope to ultimately influence producers and their advisors, we need to be conscious of how individuals and groups change their minds.

Spontaneous high-grade glial intramedullary tumor of the spine in a rhesus macaque (Macaca mulatta).

BACKGROUND: A 4-year-old rhesus macaque presented with acute, progressive paresis of the extremities. METHODS: A complete blood count, serum biochemical analysis, neurologic exam and necropsy were performed. RESULTS: The clinical, histopathological, and immunohistochemical findings confirmed a high-grade intramedullary glial tumor of the spinal cord that was most consistent with an ependymoma. CONCLUSIONS: We describe a case of a naturally occurring spontaneous spinal cord neoplasia in a non-human primate.

The envelope glycoprotein of simian immunodeficiency virus contains an enterotoxin domain.

By the use of a mouse model, the enteropathic effects of the simian immunodeficiency virus (SIV) surface unit (SU) envelope glycoprotein were explored. Purified SU (0.01-0.45 nmol) was administered intraperitoneally to 6- to 8-day-old mouse pups and induced a dose-dependent diarrheal response. Surgical introduction of SU into adult mouse intestinal loops revealed fluid accumulation without histological alterations and SU-treated unstripped intestinal mucosa induced chloride (Cl(-)) secretory currents in Ussing chambers. Similarly to rotavirus NSP4, the first described viral enterotoxin, SU induced a transient increase in intracellular calcium levels and increased inositol 1,4,5-triphosphate (IP(3)) levels in HT-29 cells. These data indicate the calcium response is mediated by IP(3). The presence of diarrhea and fluid accumulation within intestinal loops in the absence of histological alterations and induction of Cl(-) secretory currents demonstrate that SIV contains an enterotoxic domain localized within SU and is the second viral enterotoxin described.

Transdermal extraction of interstitial fluid by low-frequency ultrasound quantified with 3H2O as a tracer molecule.

The transdermal extraction of interstitial fluid by low-frequency ultrasound offers a potential minimally invasive method of obtaining a fluid sample for at-home blood glucose monitoring. Here we show that the application of low-frequency ultrasound (20 kHz) enhances the transdermal transport of interstitial fluid across hairless rat skin. Using 3H2O as a tracer injected intravenously, a measurable amount of water (>1 microL) was extracted without producing any histologic evidence of injury, even after repeated exposures. Piezoelectric transducers were imbedded in the extraction chamber and used to correlate ultrasound spectral properties to the amount of fluid extracted. Results indicate that the highest amount of water extracted occurs when the acoustic coupling media on the surface of the skin is cavitating, resulting in mild ablation of the stratum corneum and a reduction in its resistance to water mass transfer.

Cellular uptake and intracellular trafficking of long chain fatty acids.

While aspects of cellular fatty acid uptake have been studied as early as 50 years ago, recent developments in this rapidly evolving field have yielded new functional insights on the individual mechanistic steps in this process. The extremely low aqueous solubility of long chain fatty acids (LCFA) together with the very high affinity of serum albumin and cytoplasmic fatty acid binding proteins for LCFA have challenged the limits of technology in resolving the individual steps of this process. To date no single mechanism alone accounts for regulation of cellular LCFA uptake. Key regulatory points in cellular uptake of LCFA include: the aqueous solubility of the LCFA; the driving force(s) for LCFA entry into the cell membrane; the relative roles of diffusional and protein mediated LCFA translocation across the plasma membrane; cytoplasmic LCFA binding protein-mediated uptake and/or intracellular diffusion; the activity of LCFA-CoA synthetase; and cytoplasmic protein mediated targeting of LCFA or LCFA-CoAs toward specific metabolic pathways. The emerging picture is that the cell has multiple, overlapping mechanisms that assure adequate uptake and directed intracellular movement of LCFA required for maintenance of physiological functions. The upcoming challenge is to take advantage of new advances in this field to elucidate the differential interactions between these pathways in intact cells and in tissues.

Spontaneous and engineered mutant mice as models for experimental and comparative pathology: history, comparison, and developmental technology.

During the last half-century pathologists have explored the biologic mechanisms associated with inherited human and veterinary diseases by using inbred and inbred mutant (spontaneous) strains of mice. The first successful gene transfer to mice by pronuclear injection of the herpes simplex virus thymidine kinase gene and rabbit and human beta-globulin genes was achieved in the early 1980s. This accomplishment was followed a few years later with the creation of a mouse bearing a disrupted hypoxanthine phosphoribosyl transferase (hrpt) gene (targeted mutation based on ES cell blastocyst injection). Since then, hundreds of genetically engineered models of biomedical importance have been created. The unprecedented scale and scope of development of engineered models present great opportunities as well as experimental challenges to the investigator. The aim of the present review is to provide a framework of information on engineered mouse models from the perspective of experimental and comparative pathology research. Sections include: 1) a brief historical account of the development of mouse models of disease, with increasing progression of genetic refinement as represented by inbred (spontaneous) and congenic (targeted) mutant strains of mice; 2) a synopsis of spontaneous and targeted mutations, with anecdotal examples of expression of individual genes and interactions between multiple mutant genes; 3) selected examples of targeted mutations of interest to developmental and cancer biologists and immunologists; 4) an overview of the technology of development of transgenic mice; and 5) an introduction to on-line database resources of current multi-species genomic information.

A polygenic mouse model of psoriasiform skin disease in CD18-deficient mice.

Previously, a hypomorphic mutation in CD18 was generated by gene targeting, with homozygous mice displaying increased circulating neutrophil counts, defects in the response to chemically induced peritonitis, and delays in transplantation rejection. When this mutation was backcrossed onto the PL/J inbred strain, virtually all homozygous mice developed a chronic inflammatory skin disease with a mean age of onset of 11 weeks after birth. The disease was characterized by erythema, hair loss, and the development of scales and crusts. The histopathology revealed hyperplasia of the epidermis, subcorneal microabscesses, orthohyperkeratosis, parakeratosis, and lymphocyte exocytosis, which are features in common with human psoriasis and other hyperproliferative inflammatory skin disorders. Repetitive cultures failed to demonstrate bacterial or fungal organisms potentially involved in the pathogenesis of this disease, and the dermatitis resolved rapidly after subcutaneous administration of dexamethasone. Homozygous mutant mice on a (PL/J x C57BL/6J)F1 background did not develop the disease and backcross experiments suggest that a small number of genes (perhaps as few as one), in addition to CD18, determine susceptibility to the disorder. This phenotype provides a model for inflammatory skin disorders, may have general relevance to polygenic human inflammatory diseases, and should help to identify genes that interact with the beta2 integrins in inflammatory processes.

Human adenovirus type 9 E4 open reading frame 1 encodes a cytoplasmic transforming protein capable of increasing the oncogenicity of CREF cells.

The induction of estrogen-dependent rat mammary tumors by human adenovirus type 9 (Ad9) requires the Ad9 E4 open reading frame 1 (9ORF1) protein, which alone can transform that rat embryo fibroblast cell line CREF in vitro. In the present study, independent pools of both 9ORF1-expressing and control CREF cells were generated by selection with G418 and compared with respect to transformed properties. Indirect immunofluorescence analyses revealed that more than 99% of the cells that made up the 9ORF1-transfected pools expressed 9ORF1 protein and, together with confocal laser scanning microscopy, indicated that this E4 protein was located predominantly within the cytoplasm of cells. With regard to transformation, cells of the 9ORF1-expressing pools differed from those of control pools by forming foci, displaying morphological alterations, growing more efficiently in soft agar, and reaching higher saturation densities. Following injection into immunocompetent syngeneic rats, the 9ORF1-expressing pool cells exhibited greatly enhanced oncogenicity compared with control pool cells. These results show that 9ORF1 protein (i) localizes predominantly within the cytoplasm, (ii) confers multiple general transformed characteristics to CREF cells in vitro, and (iii) increases the tumorigenic properties of these cells in vivo.

Effects of genetic background on tumorigenesis in p53-deficient mice.

Mice with disrupted germline p53 alleles have been engineered by us and others and have been shown to have enhanced susceptibility to spontaneous tumors of various types. We monitored a large number of p53-deficient mice (p53+/- and p53-/-) and their wild-type littermates (p53+/+) of two different genetic backgrounds (129/Sv and mixed C57BL/6 x 129/Sv) up to 2 yr of age. p53+/- and p53-/- 129/Sv mice show accelerated tumorigenesis rates compared with their p53-deficient counterparts of mixed C57BL/6 x 129/Sv genetic background. The tumor spectra of the two strains of mice are similar except that almost half of 129/Sv p53-/- males develop malignant teratomas, whereas these tumors are rarely observed in C57BL/6 x 129/Sv mice and never in 129/Sv p53+/- males. In the study reported here, we further characterized the lymphomas that arose in the p53-nullizygous mice and found that over three-quarters of the lymphomas were of thymic origin and contained primarily immature (CD4+/CD8+) T-cells, whereas the remainder originated in the spleen and peripheral lymph nodes and were of B-cell type. The high incidence of early-onset lymphomas in the nullizygous mice makes these animals a good lymphoma model, whereas the heterozygous mice may be a useful model for Li-Fraumeni syndrome, a human inherited cancer predisposition.

Disruption of the adenosine deaminase gene causes hepatocellular impairment and perinatal lethality in mice.

We have generated mice with a null mutation at the Ada locus, which encodes the purine catabolic enzyme adenosine deaminase (ADA, EC 3.5.4.4). ADA-deficient fetuses exhibited hepatocellular impairment and died perinatally. Their lymphoid tissues were not largely affected. Accumulation of ADA substrates was detectable in ADA-deficient conceptuses as early as 12.5 days postcoitum, dramatically increasing during late in utero development, and is the likely cause of liver damage and fetal death. The results presented here demonstrate that ADA is important for the homeostatic maintenance of purines in mice.

A rapid test for endotoxin in whole blood.

A rapid whole blood agglutination test has been developed for the detection of endotoxin. The test reagent consists of polymyxin B (PmB) conjugated to the Fab fragment of the anti-glycophorin antibody 1C3/86. After addition of reagent to whole blood, red cell agglutination occurs within two minutes in samples from endotoxaemic patients or with the addition of either whole Gram negative bacteria, supernatants from Gram negative bacterial cultures or purified endotoxin. In clinical samples there was a strong correlation (r = 0.83) between the strength of agglutination and the level of endotoxin measured by the Limulus amaebolysate test (LAL). The prospect of a rapid and accurate test for endotoxin may enable better clinical management of Gram negative sepsis.

Spontaneous and carcinogen-induced tumorigenesis in p53-deficient mice.

Using gene targeting techniques, mice that have been generated with two germ-line p53 null alleles (homozygotes) develop normally but are highly susceptible to early onset spontaneous tumours. Here, we show that mice with a single null p53 allele (heterozygotes) produced in the same way are also susceptible to spontaneous tumours, but with a delayed onset compared to homozygotes. The most frequent tumour type in homozygotes was malignant lymphoma; in heterozygotes, osteosarcomas and soft tissue sarcomas predominated. Heterozygous mice treated with a liver carcinogen, dimethylnitrosamine, showed a decreased survival time in comparison to treated wild type mice, suggesting that the p53-deficient mice may be useful for some in vivo carcinogenesis assays.

Genetic background alters the spectrum of tumors that develop in p53-deficient mice.

Using gene targeting in embryonic stem cells, we have generated mice with one or two null p53 germ line alleles. Mice with both p53 alleles inactivated are developmentally normal but highly susceptible to the early development of spontaneous tumors. Initial studies were performed in mice with a mixed inbred genetic background (75% C57BL/6 and 25% 129/Sv) (Donehower et al., Nature (London) 356, 215-221, 1992). To study the effect of genetic background on tumorigenesis in p53-deficient mice, we have put the p53 null allele into a pure 129/Sv background and monitored tumor development. 129/Sv mice with two p53 null alleles developed tumors sooner than the mixed genetic background p53-deficient animals. The most frequently observed tumor in p53 null mice of both genetic backgrounds was a malignant lymphoma. Because the 129/Sv strain has a low incidence of lymphoma, the frequent occurrence of lymphomas in all p53 null mice suggests that this particular tumor type may be a direct result of p53 loss and not a result of a particular genetic background. In addition to malignant lymphomas, the 129/Sv p53-deficient mice showed an increased incidence of aggressive teratocarcinomas (8 of 18 tumor-bearing males), a tumor type rare in virtually all inbred mice except for 129 strains. Thus, it appears that loss of p53 may accelerate a prior tumor predisposition and that genetic background can play a role in mediating both the rate and spectrum of tumor development in these mice.

Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours.

Mutations in the p53 tumour-suppressor gene are the most frequently observed genetic lesions in human cancers. To investigate the role of the p53 gene in mammalian development and tumorigenesis, a null mutation was introduced into the gene by homologous recombination in murine embryonic stem cells. Mice homozygous for the null allele appear normal but are prone to the spontaneous development of a variety of neoplasms by 6 months of age. These observations indicate that a normal p53 gene is dispensable for embryonic development, that its absence predisposes the animal to neoplastic disease, and that an oncogenic mutant form of p53 is not obligatory for the genesis of many types of tumours.

Reality therapy with rape victims.

Rape is a violent act that inflicts injury on the very essence of the self. Reality therapy offers an applicable conceptual framework for the treatment of rape victims. Reality therapy groups minimize attention to the concept of mental illness. The warm, friendly atmosphere of the reality therapy group provides a supportive arena where the victim can tell her story, diminish her desire to withdraw from others, and recognize control over her behavior. Through her involvement she can begin to fulfill her needs to love and be loved, and feel worthwhile to herself and others.

Juvenile-type granulosa theca cell tumour producing precocious pseudopuberty. A case report.

The juvenile-type of granulosa theca cell tumour in a young girl is reported. The child presented with pain, vaginal bleeding, distension of the abdomen and enlarged breasts and exhibited pubic and axillary hair, i.e. signs of oestrogenic and androgenic effects and overproduction of luteinizing hormone--an unusual phenomenon. Discussion includes the differences between the juvenile and adult types of this tumour. Salpingo-oophorectomy of the affected side is recommended in such cases. A recurrence many years after surgery has been reported, so long-term follow-up is envisaged in this case.