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

Abstract

BackgroundSepsis 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.Materials and methodsA 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.ResultsWBCs 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).ConclusionTLRs 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.

Increased expression of Toll-like receptors 2, 3, 4 and 7 mRNA in the kidney and intestine of a septic mouse model

“…studied the expression of liver function tests (LFTs) and the TLR expression in the hepatic and intestinal tissues after CLP induced peritonitis 27 , whereas Bakopoulos et al . concentrated on the expression of hematological and biochemical markers related to respiratory failure as well as TLR expression in the lung and intestine after the same experimental procedure 20 . We, on the other hand, measured AKI biochemical and specific immune biomarkers, in addition to TLR expression in the kidney and the intestine after CLP.…”

“…The protocol of the study has previously been published and the samples referenced in this study derive from the same 60 animals used in the study by Bakopoulos et al . 20 with the addition of 12 new animals (2 new animals/group/each time table). The control group mice underwent a sham surgery receiving a laparotomy without cecal ligation and puncture.…”

Neutrophils in the initiation and resolution of acute pulmonary inflammation: understanding biological function and therapeutic potential

“…Pathogen recognition receptors and their downstream signalling pathways drive chemotaxis, as well as priming and activating both intravascular and transmigrated neutrophils in order to fulfil their bactericidal functions . TLRs play an important role in regulating the response to pro‐inflammatory mediators and are rapidly upregulated in mouse models of sepsis‐related acute lung injury . In early sepsis‐related ARDS, downregulation of TLR1, TLR4 and TLR5 transcripts in mononuclear cells correlates with increased survival , whereas in a pulmonary contusion mouse model of lung injury, alveolar neutrophil recruitment is TLR4/MyD88‐dependent .…”

“…Sterile tissue injury, either in the context of direct injury to lung parenchyma or distant organ injury, results in necrotic cell death with the release of a range of DAMPs into the extracellular environment. These DAMPs serve to induce a pro‐inflammatory response, which drives neutrophil recruitment into the lung . A number of DAMPs have been described in ARDS, including high mobility group box 1 (HMGB1), heat shock proteins 60 and 72, hyaluronan and a range of mitochondrial‐derived factors, including DNA, formylated peptides and cardiolipin .…”

Toll-like receptors in sepsis-associated cytokine storm and their endogenous negative regulators as future immunomodulatory targets

“…The TLR2 and TLR4 expression in the lungs of mice subjected to the cecal-ligation and puncture (CLP)-induced polymicrobial sepsis increases within 24 h as compared to the sham group [58] . Whereas, TLR3 and TLR7 expression increases in their intestine within 24 h. This can be speculated as gut epithelial cells normally express a lower level of TLR2 (at apical surface) and TLR4, but a higher level of TLR3 and TLR5 [59] , [60] , [61] .…”