Capsular Polysaccharide is a Main Component of Mycoplasma ovipneumoniae in the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells

Figure 2

CPS stimulation induces the expression of TLRs in sheep bronchial epithelial cells. M. ovipneumoniae bacterial cells were preincubated with CPS antibody (1 : 100) at 4°C overnight. 4-week old ALI cultures of sheep bronchial epithelial cells were then apically treated with capsular polysaccharide (CPS) at 100 ng/mL or infected with M. ovipneumoniae (MO) at MOI of 30 for 48 h before samples were harvested for analysis. For each treatment, an additional nonspecific IgG (1 : 100) was supplemented as a control. (a) The induction of TLR1, TLR2, TLR4, and TLR6 transcripts was determined by a qRT-PCR assay. All TLR transcripts showed a significant increase in the cells stimulated with CPS or M. ovipneumoniae, in comparison with the controls. The expression levels of these mRNA were calculated by 2^−ΔΔCt method. (b) Immunoblotting assay also showed an evoked expression of TLR4 proteins in ALI cultures upon CPS/M. ovipneumoniae stimulation. (c) Densitometric values are shown for TLR4 proteins over β-actin. Values are mean ± SD for at least three independent experiments performed in triplicate. versus that of the control. Compared between indicated groups, ^▲▲.

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