Biomedical subjects

Patricia L Foley

Publications and source records attributed to Patricia L Foley.

9 recordsLinked to original sources

PubMed2005

Regulatory considerations for marker vaccines and diagnostic tests in the U.S.

Marker vaccines and diagnostic tests can prove to be invaluable in disease eradication and control programs, as was found in the pseudorabies (Aujeszky's Disease) virus eradication program in the U.S. During that campaign, numerous gene-deleted vaccines and companion diagnostic test kits were used to differentiate infected animals from vaccinated animals, in a strategy that ultimately led to eradication of the disease in commercial swine herds. The United States Department of Agriculture played a key role in delivery of that success by developing biologics policy, evaluating each product, and ensuring that the conditions of licensure were met. What was most critical in the overall eradication effort, however, was the detailed and dedicated interaction among key players: the biologics regulators, manufacturers, Federal, State, and local regulatory partners, veterinary researchers, industry associations, and animal owners. A good disease control program has to include all of these. The regulatory requirements for licensure of marker vaccines and diagnostic test kits are not different from that for other products. There are several mechanisms for vaccine approval, some more rapid than others, but only a few that could apply to these products. Generally, the platforms that might support marker vaccines and companion diagnostic kits are those based on genetic engineering or protein manipulation. If the product is derived from the application of biotechnology, then additional regulatory considerations are applicable. Most important of these are the considerations found in the National Environmental Policy Act (NEPA), wherein deliberate release of any organism containing recombinant DNA into the environment is subject to review and approval by appropriate federal agencies. Environmental release and NEPA compliance are discussed.

Animal Diseases↗

PubMed2004

Outbreak of otitis media caused by Burkholderia gladioli infection in immunocompromised mice.

An athymic nude mouse with severe head tilt due to otitis media was identified. Within weeks of identification of this first case, immune-deficient mice of various genotypes from the same facility were similarly affected, and cases from other facilities were found within two months. Culture of ear exudate specimens from affected mice yielded bacteria that were initially identified as Burkholderia cepacia, a plant pathogen considered an important opportunistic pathogen in persons with cystic fibrosis or chronic granulomatous disease. Several of these isolates, however, were subsequently identified as B. gladioli on the basis of results of biochemical analysis and a species-specific polymerase chain reaction (PCR) assay. Genotyping analysis revealed clonality among the isolates, indicating a shared strain among affected mice. A 16S rDNA-based PCR assay specific for the genera Burkholderia and Ralstonia, and a selective culture medium were used in efforts to characterize the epidemiology of this outbreak. In addition to culture of specimens from the oropharyngeal cavity of affected mice, samples were obtained from the environment, feces, sipper tubes, drinking water, and soiled bedding from cages of affected individuals. Burkholderia gladioli was most consistently detected in oropharyngeal swab specimens from affected mice. The PCR assay was equivalent to selective culture in identifying mice in the carrier state that did not have clinical signs of infection. However, neither detection method had sufficient sensitivity to reliably identify all carrier mice, causing the organism to persist at low levels unless entire colonies of immune-deficient mice were removed. The organism was highly resistant to antibiotic therapy. The source and epidemiology of this organism remain unknown. This epizootic serves as an important reminder that immunocompromised rodent colonies may harbor important human opportunistic pathogens.

Animals↗

PubMed2003

B cell complement receptor 2 transfer reaction.

The B cell C receptor specific for C3dg (CR2) shares a number of features with the primate E C receptor (CR1). Previously, we have demonstrated, both in vitro and in animal models, that immune complexes (IC) bound to primate E CR1, either via C opsonization or by means of bispecific mAb complexes, can be transferred to acceptor macrophages in a process that also removes CR1 from the E. We have now extended this paradigm, the transfer reaction, to include B cell CR2. We used both flow cytometry and fluorescence microscopy to demonstrate that IC bound to Raji cell CR2, either via C opsonization or through the use of an anti-CR2 mAb, are transferred to acceptor THP-1 cells. This reaction, which appears to require Fc recognition of IgG bound to Raji cell CR2, also leads to transfer of CR2. Additional support for the B cell transfer reaction is provided in a prototype study in a monkey model in which IC bound to B cell CR2 are localized to the spleen. These findings may have important implications with respect to defining the role of C in IC handling during the normal immune response.

Animals↗

PubMed2002

Concerted clearance of immune complexes bound to the human erythrocyte complement receptor: development of a heterologous mouse model.

Experiments in primates have demonstrated that immune complexes (IC) bound to erythrocytes (E) via complement receptor 1 (CR1) are cleared to the liver in a process which removes CR1, but otherwise spares the E. Human E are stabilized for >1 h in the circulation of the mouse if the terminal complement pathway is blocked, and we used this paradigm to examine clearance in a mouse model. Human E were opsonized with an anti-CR1 mAb cross-linked to dsDNA (antigen-based heteropolymer, AHP), and then incubated with systemic lupus erythematosus (SLE) plasmas containing IgG anti-dsDNA to form IC in situ. These IC stably bind to E CR1 in the complete absence of complement, thus allowing analysis in a model which does not require human C3b to facilitate E binding. Dual label experiments, based on RIA, flow cytometry and fluorescence microscopy, were employed to monitor separately E and IC. When opsonized E-IC were injected into A/J mice, >90% of the IC were rapidly removed from the E coincident with loss of CR1. The E remained in the circulation while IC were localized to the liver, mainly to Kupffer cells. Preliminary experiments in NZB/W mice, which spontaneously develop IgG anti-dsDNA, indicated that infusion of E-AHP led to rapid binding of murine IgG to the E-AHP, followed by removal of the nascent IC from E, and loss of CR1 in a concerted reaction. These studies provide additional evidence that E CR1 functions as a privileged site for IC clearance, and that the key step in clearance requires removal of CR1 from E to release bound IC for uptake by acceptor macrophages. This model can be extended to genetically altered mice to investigate the role of specific Fc gamma receptors as well as complement receptors in IC clearance.

Animals↗

PubMed2002

T cell requirement for development of chronic ulcerative dermatitis in E- and P-selectin-deficient mice.

C57BL/6 mice deficient in E- and P-selectin (E(-/-)P(-/-)) kept under specific pathogen-free barrier conditions have high circulating neutrophil counts and develop hypercellular cervical lymph nodes with substantial plasma cell infiltrates, severe ulcerative dermatitis, conjunctivitis, and lung pathology, which eventually lead to premature death. To test the hypothesis that the pathology in E(-/-)P(-/-) mice may be caused by dysfunctional lymphocyte activity, we crossed E(-/-)P(-/-) mice with recombination activation gene (Rag)-1(-/-) mice to generate E(-/-)P(-/-)Rag-1(-/-) mice lacking mature T and B lymphocytes. E(-/-)P(-/-)Rag-1(-/-) mice had circulating neutrophil counts and plasma G-CSF levels similar to E(-/-)P(-/-) mice. Remarkably, none of the E(-/-)P(-/-)Rag-1(-/-) mice developed conjunctivitis or ulcerative dermatitis typical of E(-/-)P(-/-) mice. These mice were overall healthier in appearance than E(-/-)P(-/-) mice, and histopathologic changes in the lung were reduced. Cervical lymph nodes in E(-/-)P(-/-)Rag-1(-/-) mice were much smaller than those of E(-/-)P(-/-) mice, containing few mononuclear cells and no plasma cells. These data show that the severe disease phenotype of E(-/-)P(-/-) mice depends on lymphocyte function. We conclude that a dysregulated immune response in E(-/-)P(-/-) mice causes disease development, but is not necessary for elevated neutrophil counts.

Animals↗

PubMed2002

An anti-C3b(i) mAb enhances complement activation, C3b(i) deposition, and killing of CD20+ cells by rituximab.

We investigated deposition of the complement protein fragment C3b and its breakdown products (collectively designated as C3b(i)) on CD20-positive cells treated with rituximab (RTX) in the presence of normal human serum (NHS). Radioimmunoassay (RIA) demonstrates that about 500 000 C3b(i) molecules deposit per cell, and fluorescence microscopy reveals that C3b(i) colocalizes with bound RTX. Use of mAb 3E7, specific for C3b(i) bound to substrates, enhances C3b(i) deposition; > 1 million C3b(i) deposit when cells are incubated with NHS, RTX and mAb 3E7. Treatment of Raji cells in NHS plus RTX leads to robust cell killing (95%) after 24 to 48 hours, and mAb 3E7 significantly enhances RTX-mediated killing of Raji and DB cells. A cynomolgus monkey model based on intravenous infusion of RTX followed by mAb 3E7 demonstrated that RTX rapidly binds to B cells and promotes complement activation and C3b(i) deposition; fluorescence microscopy analyses revealed the same pattern of colocalization of C3b(i) on cell-bound RTX in vivo as observed in vitro. Preliminary in vitro studies with blood samples from patients with chronic lymphocytic leukemia lead to similar findings. These experiments suggest that complement plays a key role in the mechanism of action of RTX; moreover, the in vivo molecular form of RTX (and possibly other antitumor mAbs) in the circulation or in tissues may include C3b(i) molecules covalently bound to the therapeutic mAb, thus allowing it to interact with cells containing both Fc and complement receptors.

Animals↗

PubMed2002

Severely reduced neutrophil adhesion and impaired host defense against fecal and commensal bacteria in CD18-/-P-selectin-/- double null mice.

Leukocyte recruitment to sites of inflammation requires the functions of selectins and integrins. P-selectin null (CD62P-/-) mice show a mild and CD18 null (CD18-/-) mice a more severe neutrophil recruitment defect in some inflammatory models. To investigate the possible cooperative interactions between CD18 integrins and P-selectin in mediating neutrophil recruitment, we generated CD18-/-CD62P-/- double null mice. CD18-/-CD62P-/- mice were apparently normal at weaning and fertile but later failed to gain weight, showed increased susceptibility to infection by fecal and commensal bacteria, and survived only 5-6 months. Some CD18-/-CD62P-/- mice showed severe spontaneous skin lesions; most showed neutrophil infiltration in the lungs and liver, and positive bacterial cultures from internal organs. The number and velocity of rolling leukocytes in tumor necrosis factor alpha treated venules of CD18-/-CD62P-/- mice was similar to those in wild-type mice, but neutrophil adhesion was severely reduced. Only 25% of adhered leukocytes were neutrophils in CD18-/-CD62P-/- mice vs. >90% in wild-type, CD62P-/-, and CD18-/- single mutants. Our data show that removing both P-selectin and CD18 integrins from mice leads to severe neutrophil recruitment defects and spontaneous pathology.

Animals↗

PubMed2002

Effect of covalently bound heparin coating on patency and biocompatibility of long-term indwelling catheters in the rat jugular vein.

Many physiologic and pharmacologic studies require long-term vascular access for repeated substance infusion and/or blood sample collection. The study reported here was undertaken to determine whether a functionally active heparin coating would improve long-term patency of venous catheters in rats. Uncoated or coated catheters were surgically placed in the jugular vein, and patency was evaluated twice weekly for a total of 30 days. Culturing of blood and catheters, and histologic examination were performed for all rats. All heparin-coated catheters remained patent for the study duration, with patency defined as ability to infuse saline and withdraw a blood sample. Median patency for uncoated catheters was 17.5 days, with a range of three to 30 days. Histologic evaluation of vessels revealed more advanced and severe lesions in rats with uncoated, compared with coated catheters. Furthermore, uncoated catheters had increased association with bacteremia (3/8), compared with coated (0/9) catheters. Taken together, these results indicate that coating catheters with covalently bound heparin molecules can significantly prolong patency and cause less pathologic damage to the catheterized vessel.

Animals↗

PubMed2002

Comparison of an espB gene fecal polymerase chain reaction assay with bacteriologic isolation for detection of Citrobacter rodentium infection in mice.

PURPOSE: To develop a polymerase chain reaction (PCR) assay for specific detection of Citrobacter rodentium in fecal samples of mice and to compare this assay with bacterial isolation and identification methods. METHODS: The target sequence of the PCR assay was the espB gene encoding a secreted virulence factor. To facilitate visual identification during primary isolation on MacConkey agar containing ampicillin, C. rodentium ATCC type strain 51459 was transformed by use of a plasmid encoding the enhanced green fluorescent protein (EGFP) and ampicillin resistance. The EGFP-C. rodentium was inoculated into Swiss Webster (SW) mice to study the time course of detection of the organism by use of fecal PCR analysis, bacterial isolation, and development of colonic hyperplasia by light microscopy. Lactose-fermenting fluorescent bacterial colonies identified during primary isolation of fecal bacteria on MacConkey-ampicillin agar were identified by use of biochemical typing. RESULTS: Mice inoculated with EGFP-transformed C. rodentium developed colonic mucosal hyperplasia, characterized by a three-fold increase in colonic crypt height that peaked at post-inoculation day (PID) 14. The espB PCR assay detected as little as 0.3 colony-forming units of C. rodentium. The PCR assay was specific in that it did not detect the espB gene of Escherichia coli 0157. Results of in vivo studies in SW mice indicated that EGFP-C. rodentium could be detected by use of espB fecal PCR analysis in 100% of inoculated mice tested on PID 1, 3, 7, and 8, in 60% on PID 9, and in 20% on PID 10 (n = 5). Bacterial isolation from the same fecal samples detected the organism in 100% of the inoculated mice on PID 7, in 50% on PID 8, and in none on subsequent PID 9-14. The ability of the PCR assay to detect C. rodentium in fresh feces of inoculated mice was significantly better than that of bacterial isolation methods (Fisher-Irwin exact test, P < 0.01). At the time of peak colonic hyperplasia, the organism could no longer be cultivated or detected in mice by use of fecal PCR analysis. CONCLUSIONS: The EGFP-C. rodentium was capable of inducing transmissible murine colonic hyperplasia similar to that previously reported in SW mice. The PCR assay for detection of the espB gene sequence of C. rodentium in total fecal DNA was a more sensitive diagnostic assay than was bacterial isolation.

Animals↗