Application of intestinal xenografts to the study of enteropathogenic infectious disease.

We have developed, characterized and utilized paired segments of fetal intestine subcutaneously transplanted into heterogenic nude or SCID mice as a model system for the study of viral, bacterial and protozoal pathogens. The xenografted intestine matures in the recipient mouse and is biochemically and anatomically comparable to intestine from age-matched, whole-animal controls. The grafted tissue is free of ingesta, intestinal flora, extra-intestinal secretions and host immune functions. The transplanted intestine is long-lived and easily accessible to manipulation and harvest. Tissue from a single fetal donor can be used to create numerous xenografts allowing for tightly controlled experiments. Xenografts enable the study of species-specific intestinal pathogens in the homologous intestinal tissue thus preserving biological applicability of results. Xenografts can be used to study pathogenesis, pathophysiology and therapeutics of enteric disease in situations where such study might otherwise be prohibitively expensive or confounded by intercurrent variables inherent to whole animals. Xenografts have important advantages over in vitro models that may not approximate the in vivo biology of the intestine in the disease process.

Monitoring pH of otitis media effusion in chinchillas using fluorescence spectroscopy.

We have developed a fiber optic fluorometer to measure fluorescent signal intensities across an epithelium barrier. As a medically relevant example, we have measured the pH of the effusion formed during Hemophilus influenzae induced otitis media infection in the chinchilla, the classical animal model for human middle ear disease. Because the choice of antibiotic used in clinical therapy is dependent on the pH of the effusion, a noninvasive method of measuring pH is highly desirable. Using the fluorescent pH probe carboxy-seminapthorhodafluor, we were able to detect pH changes of 0.15 units in the pH range around 7.0. The development and resolution of the otitis media was followed with magnetic resonance imaging to confirm the presence of the effusion formed during the infection.

Annual changes in serum concentrations of prolactin in captive male black bears (Ursus americanus).

Prolactin may be involved in the regulation of reproduction in black bears (Ursus americanus) as it is a mediator of photoperiodic changes in a number of species. The objectives of this study were to validate a radioimmunoassay to measure prolactin in bear serum and to describe seasonal changes in serum prolactin concentrations in captive male bears. Serum samples were obtained nine times during a year from three captive male black bears that were denning between November and March and active during the other months. The heterologous prolactin radioimmunoassay, using pig 125I-labelled prolactin and goat anti-pig prolactin as a primary antibody, was validated. Injection of thyrotrophin-releasing hormone into the three male bears in June resulted in a rapid increase in serum concentrations of prolactin (t = 0, 11.4-14.8 ng ml-1; t = 15-30 min, 18.4-28.7 ng ml-1). The sensitivity of the assay was 0.08 ng per tube. Intra- and interassay coefficients of variation were 5.5% (n = 6) and 5.7% (n = 6), respectively. Serum concentrations of prolactin changed seasonally, with the lowest concentrations in December (mean +/- SD = 1.1 +/- 0.1 ng ml-1); this was followed by a gradual increase between January (2.6 +/- 0.6 ng ml-1) and April (6.4 +/- 1.2 ng ml-1) and the highest concentrations in May (17.6 +/- 4.7 ng ml-1), preceding peak testosterone concentrations in June. The observation that prolactin secretion increased with increasing daylength suggests that photoperiod may be an external regulator. The presence of high concentrations of prolactin before peak testosterone concentrations suggests that prolactin may play a role in regulating seasonal changes in the testes.

An intestinal xenograft model for Cryptosporidium parvum infection.

Paired segments of near-term fetal rabbit small intestine were transplanted subcutaneously into athymic nude mice. At 5 weeks postsurgery, the xenografts were inoculated intraluminally with Cryptosporidium parvum sporozoites. Parasites rapidly and reliably infected the xenograft mucosal epithelium. Lesions typical of cryptosporidiosis were readily apparent by light microscopy and scanning and transmission electron microscopy. Xenografts are well suited to the study of the early events of C. parvum infection and are of potential value in the evaluation of anticryptosporidial chemotherapeutic agents.

Development, characterization, and utilization of an intestinal xenograft model for infectious disease research.

Paired xenografts of near-term fetal rabbit jejunum were subcutaneously implanted in the backs of athymic nude (nu/nu) mice. At 3 to 4 weeks post-implantation, the grafts had histologic, ultrastructural, and biochemical (lactase, sucrase, alkaline phosphatase, leucine aminopeptidase) parameters comparable to age-matched control rabbits. Four weeks post-transplantation the xenografts were intraluminally inoculated with various strains of lapine attaching and effacing E. coli or group A rotavirus. Infection with 2 strains of E. coli resulted in typical light microscopic and ultrastructural lesions of attachment and effacement. Immunohistochemical analysis of rotavirus-infected xenografts demonstrated rotavirus antigen within enterocytes. These lesions are comparable to those in conventional rabbits. Intestinal xenografts are a novel, highly controlled, and reproducible model which may have unique applications in the study of enteric diseases. The model provides anatomically and biochemically correct intestinal mucosal epithelium uncomplicated by variables such as enteric flora, host immune response, gastric, hepatic, and pancreatic secretions and is susceptible to infection by specific enteropathogens. Xenografts, therefore, may be a viable alternative in certain investigations where whole animals, ligated intestinal loops, organ cultures, or cell cultures might otherwise be chosen.