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Characterization of ovine pattern recognition receptors expression

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NalubambaKS_2007redux.pdf (44.94Mb)
Date
2007
Author
Nalubamba, King Shimumbo.
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Abstract
 
 
The innate immune system senses pathogens via germ-line encoded proteins called pattern recognition receptors (PRRs), which include Toll-like receptors (TLRs) and C-type lectins. They are able to distinguish a vast range of microbial signatures or pathogen associated molecular patterns (PAMPs) and induce both adaptive and innate, pathogen specific/tailored immune responses. The repertoire of PRRs expressed by each cell type has a bearing on its ability to recognize a specific array of PAMPs on a pathogen and thus mount a specific immune response against such challenge. The cellular content of a tissue thus has a bearing on its ability to mount a competent immune response against pathogens. Age-related differences in immune competence are well documented. Neonates are known to have immature immune systems (and thus increased susceptibility to infections) and postulated to have lower PRR expressions. A discernment of the expression of PRRs in normal cells/tissues enables us understand the significance of these receptors play in immune response development and form a basis for understanding disease pathogenesis. Baseline data also allows deviations from normal to be identified in diseased states. Johne's disease (JD) is a chronic disease of ruminants caused by Mycobacterium avium paratuberculosis (Map) and has three clinical forms - asymptomatic, paucibacillary or multibacillary. The innate immune system is known to have a pivotal role in the control of the dissemination in mycobacterial diseases though the precise mechanisms are not well defined. 1 hypothesized that antigen recognition via PRRs has a definitive role in the development of the different forms of JD.
 
For part one of this study, blood and tissues were obtained from clinically healthy sheep for RNA extraction. Blood subsets were immuno-stained with specific monoclonal antibodies and DCs into CD172a~" populations. Definitive populations were obtained using Flow cytometry assisted cell sorting followed by RNA extraction.
 
To explore PRR expression in foetal immune system, second trimester foetal skins and spleens were collected for PRR mRNA expression determination. Archival, RNAlaterk stabilized ileum samples were used to investigate PRR expression profiles of different JD forms.
 
PRR specific mRNA expression was evaluated using reverse transcriptase quantitative real time PCR.
 
The spleen, lung and lymph nodes express all TLRs; the kidney expresses high levels of TLR1, 2, 3, 4, 5, 6, but very low levels of TLR8, 9 and 10. The skin expresses low levels of TLR5, 6, 9 and 10 mRNA. CD172a DCs express most PRRs and MyD88 while CD 172a" DCs express TLR2, 6 and MyD88.
 
Foetal spleens have comparable levels of PRRs except for CD 14. Significant differences were observed with TLR1, 4, 5, CD 14, CARD 15 and Dectin-1 between foetal and adult skin tissues.
 
Results from the present study show the importance of the following PRRs in ovine Johne's disease discrimination; TLR2, CD14, TLR8, CARD15, dectin-1 and dectin2. These findings provide an insight into one facet of Map innate immune recognition and help to elucidate new target genes for possible mutation analyses and disease genotyping.
 
URI
http://hdl.handle.net/1842/29909
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  • Biological Sciences thesis and dissertation collection

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