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.
