Expression of Toll-like receptors (TLRs) in the lungs of an experimental sepsis mouse model.

Abstract

Sepsis is a condition characterized by high mortality rates and often accompanied by multiple-organ dysfunction. During sepsis, respiratory system may be affected and possibly result in acute respiratory distress syndrome (ARDS). Toll-like receptors (TLRs), as a first line defense against invading pathogens, seem to be highly expressed in septic states. Therefore, expression of TLRs in the lungs of a sepsis animal model could indicate the involvement of the respiratory system and appear as a severity index of the clinical course.A total of 72 C57BL/6J mice, aged 12-14 weeks, were studied. The animals were divided into 3 sepsis (S) groups (24h, 48h and 72h) and 3 control (C) groups (24h, 48h and 72h), each consisting of 12 mice. The S-groups were subjected to cecal ligation and puncture (CLP) while the C-groups had a sham operation performed. Blood samples were drawn from all groups. Total blood count analysis was performed along with the measurement of certain biochemical markers. Additionally, lung tissues were harvested and the expression of TLRs, namely TLR 2, TLR 3, TLR 4 and TLR 7 were evaluated by means of immunofluorescence (IF) and qRT-PCR (quantitative-Polymerase Chain Reaction). Statistical analysis was performed by using one-way ANOVA followed by student t-test. Results were considered statistically significant when p<0.05.WBCs and lymphocytes were decreased in all S-groups compared to the corresponding C-groups (p<0.05), while RBCs showed a gradual decline in S-groups with the lowest levels appearing in the S72 group. Only, monocytes were higher in S-groups, especially between S48-C48 (p<0.05) and S72-C72 (p<0.05). Creatinine, IL-10 and IL-6 levels were significantly increased in the S-groups compared to the corresponding C-groups (S24 vs C24, S48 vs C48 and S72 vs C72, p<0.05). IF showed that expression of TLRs 2, 3, 4 and 7 was increased in all S-groups compared to the time-adjusted C-groups (p<0.05). Similarly, qRT-PCR revealed that expression of all TLRs was higher in all S-groups compared to their respective C-groups in both lungs and intestine (p<0.05). Comparing lung and intestinal tissues from S-groups, TLRs 2 and 4 were found increased in the lung at 24, 48 and 72 hours (p<0.05), whereas TLR 3 was higher in the intestine at all time points examined (p<0.05). Finally, TLR 7 levels were significantly higher in the intestinal tissues at 24 hours (p<0.0001), while lungs predominated at 48 hours (p<0.0001).TLRs seem to be highly expressed in the lungs of septic mice, therefore suggesting a potential role in the pathogenesis of ARDS during sepsis. While more studies need to be conducted in order to completely understand the underlying mechanisms, TLRs may represent a promising target for establishing novel therapeutic strategies in the treatment of sepsis.