BackgoundIt has been reported that Toll-like receptor 4 (TLR4) deficiency reduces infarct size after myocardial ischemia/reperfusion (MI/R). However, measurement of MI/R injury was limited and did not include cardiac function. In a chronic closed-chest model we assessed whether cardiac function is preserved in TLR4-deficient mice (C3H/HeJ) following MI/R, and whether myocardial and systemic cytokine expression differed compared to wild type (WT).ResultsInfarct size (IS) in C3H/HeJ assessed by TTC staining after 60 min ischemia and 24h reperfusion was significantly smaller than in WT. Despite a smaller infarct size, echocardiography showed no functional difference between C3H/HeJ and WT. Left-ventricular developed pressure measured with a left-ventricular catheter was lower in C3H/HeJ (63.0 ± 4.2 mmHg vs. 77.9 ± 1.7 mmHg in WT, p < 0.05). Serum cytokine levels and myocardial IL-6 were higher in WT than in C3H/HeJ (p < 0.05). C3H/HeJ MI/R showed increased myocardial IL-1β and IL-6 expression compared to their respective shams (p < 0.05), indicating TLR4-independent cytokine activation due to MI/R.ConclusionThese results demonstrate that, although a mutant TLR4 signaling cascade reduces myocardial IS and serum cytokine levels, it does not preserve myocardial function. The change in inflammatory response, secondary to a non-functional TLR-4 receptor, may contribute to the observed dichotomy between infarct size and function in the TLR-4 mutant mouse.

Our findings suggest that TLR4 plays a key role for regulating the expression of relevant cytokines within the lung during endotoxic shock.

Toll-like receptor 4 (TLR4) mediates innate immune responses following endotoxemia and myocardial ischaemia -reperfusion (I/R) injury. Pre-treatment with the major TLR4 ligand lipopolysaccharide (LPS) reduces infarct size. Matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) play a crucial role in endotoxemia possibly also determining I/R injury. Aims: We investigated the influence of TLR4 on infarct size and assessed the influence of MMP and TIMP regulation on I/R injury. Methods: Left anterior descending artery (LAD) occlusion was performed on wild-type (C3H/HeN) and TLR4-deficient (C3H/HeJ) mice. Animals were stimulated with LPS (1 mg/kg) or PBS 16 h ahead of 60 min LAD ligation. After 24 h of reperfusion, triphenyltetrazolium chloride staining was performed and infarct size was measured by planimetry. MMP-and TIMP-mRNA expression were determined by RPA after 3 h of reperfusion. MMP zymographic activity was monitored 6 h after occlusion. Results: TLR4-deficient mice and LPS-treated wild-type mice showed significantly reduced infarct areas. LPS-stimulation significantly increased the overall MMP/TIMP mRNA expression ratio due to elevated MMP-3, -8, -9, and TIMP-1 in wild-type mice. I/R overall reduced the MMP/TIMP ratio due to increased MMP-1, TIMP-1, and -3 mRNA expression. Conclusions: LPS pre-treatment and TLR4-deficiency led to a decreased infarct size. However, infarct area and MMP/TIMP ratio were not correlated. This means that in TLR4-deficient mice MMP/TIMP ratios are not determining the infarct size.

TAC modulates innate immunity by regulating the expression of CD14 and changes the myocardial tissue to respond more powerful to LPS.

Objective-Aim was to assess whether lipopolysaccharide (LPS)-induced decrease of total peripheral resistance depends on Toll-like receptor (TLR)4 signaling and whether it is sensitive to NO-synthase or TLR4 antagonists. Methods and Results-C3H/HeN mice (control), expressing a functional, and C3H/HeJ mice, expressing a nonfunctional TLR4, were compared. LPS (20 mg/kg) was injected i.p. 6 hours before hemodynamic measurements. L-NAME and SMT, inhibitors of NO production, and Eritoran, a TLR4 antagonist, were tested for their impact on vascular contractility. Aortic rings were incubated for 6 hours with or without LPS (1 g/mL), or with LPSϩEritoran (2 g/mL) and their phenylephrine-induced contractility was measured using a myograph. The expression of cytokines in aortic tissue was examined by real-time polymerase chain reaction. In control mice LPS induced a significant decrease of blood pressure and an increase of heart rate, whereas C3H/HeJ remained unaffected. LPS induced an increase of cytokine expression and a depression of vascular contractility only in control mice but not in C3H/HeJ. L-NAME and SMT increased contractility in all rings and restored LPS-dependent depression of contractility. Eritoran prevented LPS-induced loss of contractility. Key Words: blood pressure Ⅲ sepsis Ⅲ toll-like receptor 4 antagonism Ⅲ vascular contractility S epsis and septic shock are responsible for about 1400 deaths per day on noncardiac intensive care units in the United States. 1,2 During bacterial infection virulence factors are released and the immune system reacts by generating inflammatory cytokines 3 leading to a lowered total peripheral resistance (TPR) of the circulatory system. This is compensated by an increase in heart rate and cardiac output. In septic shock cardiac output cannot balance the loss in peripheral resistance any longer resulting in a drastic drop in blood pressure and disturbed microcirculation. 4 It has been shown that different members of the Toll-like receptor (TLR) family specifically bind different virulence factors from a wide variety of pathogens. Those virulence factors are very important for the pathogenesis of sepsis. Therefore, competitive inhibition of virulence factors at the Toll-like receptor level might be a potential therapeutic option to treat pathogen-induced sepsis and septic shock. LPS is an important virulence factor of Gram-negative bacteria, which binds to and signals via the TLR4/CD14 complex, thereby inducing inflammatory mediators. 3,5 Recently it has been demonstrated that a synthetic Lipid-A-analog called Eritoran (E5564, Eisai, currently also used in different clinical trials for the treatment of sepsis) competitively antagonizes LPS at the TLR4/CD14 complex and inhibits the synthesis and induction of sepsis relevant mediators. In addition, Eritoran was able to prevent the negative inotropic effect of LPS on isolated cardiac myocytes. 6,7 In addition nitric oxide (NO) and cytokines or adrenomedullin (ADM) appear to be increased during sepsis and play an important role in ...

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The phylogenetic status of Afrotropical galerucines was investigated with molecular and morphological analyses. The taxon sample analysed comprised 15 species within Monolepta, three within Afrocandezea, two each within Afrocrania and Barombiella and one Pseudocrania species; all were originally placed in “Monoleptites”. Further galerucines outside the “Monoleptites” are Diacantha sp., Exosoma polita, Exosoma sp., Galerudolphia tenuicornis, and Parasbecesta ruwensorica. The chrysomeline Linaeidea nubila was included as outgroup. 35 morphological characters including 16 characters on genital morphology were analysed. A 540 bp mitochondrial DNA NADH dehydrogenase subunit 1 (ND1) fragment and the entire second internal transcribed spacer region ITS2 (519–709 bp) of the nuclear ribosomal DNA were sequenced from 22 and 24 taxa, respectively. Both molecular data sets were characterized by a high average A-T content of 86.4% (ND1) and 62.7% (ITS2). Trees of separate and combined data sets were reconstructed with Maximum Parsimony (MP) and Maximum Likelihood (ML) approaches. The congruent tree topologies of both morphological and molecular data sets strongly support the monophyly of Monolepta, Afrocrania and Afrocandezea with regard to recently revised Afrotropical representatives. Barombiella emerged as polyphyletic, on species showing close relationship to Galerudolphia tenuicornis, which is traditionally placed in the “Scelidites”. “Monoleptites” is most likely polyphyletic since its decisive character, the elongated metatarsus, obviously evolved more than once in the Galerucinae. Understanding of the phylogenetic position and delimitation of the taxa primarily based on morphological characters could be significantly improved by molecular data.