Document Type

Dissertation - University Access Only

Award Date

2003

Degree Name

Doctor of Philosophy (PhD)

Department / School

Veterinary and Biomedical Sciences

First Advisor

Christopher C. L. Chase

Abstract

The mononuclear phagocytic system (monocytes, tissue macrophages and dendritic cells) plays an essential role in the innate immune response and in initiating acquired immune responses. Many viruses infect the mononuclear phagocytic system cells and interfere with their inflammatory functions. Bovine viral diarrhea virus (BVDV) is a major cattle pathogen. BVDV strains exist in two biotypes: cytopathic (cp BVDV) and noncytopathic (ncp BVDV). Cp and/or ncp BVDV infect macrophages and cause transient and persistent infection. Only ncp BVDV strains cause transient and persistent infection, while cp BVDV strains cause any transient infection. The specific cellular and molecular mechanisms of BVDV-macrophages interaction are not completely defined. The objective of this study was to determine the outcome of BVDV on macrophage inflammatory functions and to investigate the molecular mechanisms underlying BVDV associated immunosuppression. To study the outcome of BVDV infection of macrophages, a simple reproducible method for the generation of functional bovine macrophages from peripheral blood monocytes was established by culturing monocytes on gelatin/autologous fresh plasma coated surfaces and in slightly acidic media containing fresh autologous serum for 4-6 days in vitro. Monocytes-derived macrophages (MDM) were 99% homologous in morphology and in non-specific esterase expression and more than 97% of the cells phagocytoze yeast, bacteria or beads. MDM had much higher killing activity for yeast or bacteria than monocyte and expressed CD14, CDllb, MHC I, MHC II or CDl l c. This culture system provides a convenient method to obtain large number of homologous macrophages to be used in immunological and microbiological studies. MDM were susceptible to BVDV infection and supported BVDV protein expression, but did not release infectious virus. Cp BVDV strains triggered cytopathic effects (CPE), detachment, cell fusion, and death in infected culture, while ncp BVDV did not. Only the most virulent BVDV biotypes caused impairment in MDM inflammatory functions, including: phagocytic capacity, microbicidal activities (bactericidal and /or fungicidal), CD14, MHC I or MHC II surface expre sion by approximately 35-60%, and ability to produce nitric oxide (NO). Inactivated BVDV failed to reduce these functions in MDM confirming the requirement of replicating virus as a determinant of BVDV pathogenesis. Purified supernatants from cp BVDV or from high virulent ncp BVDV were able to reduce uninfected MDM phagocytic capacity, bactericidal activity or surface markers expression. As a possible cause for viral pathogenicity, only cp BVDV strains caused apoptosis in macrophages culture using the p53/Bax-Bcl2 pathway. Interestingly, Madin Darby Bovine Kidney Cells (MDBK) treated with supernatants from high virulent ncp BVDV or cp BVDV strains also under went apoptosis. These results indicated that BVDV might induce MDM to release soluble factor(s). This soluble factor(s) might be a determinate for BVDV infection and host cell interactions.

Library of Congress Subject Headings

Bovine viral diarrhea.
Cattle -- Virus diseases.
Macrophages.
Inflammation -- Immunological aspects.

Publisher

South Dakota State University

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Rights Statement

In Copyright