Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2014 Sep 4;14:5.
doi: 10.1186/s12899-014-0005-1.

MAPK-activated Protein Kinase 2-deficiency Causes Hyperacute Tumor Necrosis Factor-Induced Inflammatory Shock

Affiliations
Free PMC article

MAPK-activated Protein Kinase 2-deficiency Causes Hyperacute Tumor Necrosis Factor-Induced Inflammatory Shock

Benjamin Vandendriessche et al. BMC Physiol. .
Free PMC article

Abstract

Background: MAPK-activated protein kinase 2 (MK2) plays a pivotal role in the cell response to (inflammatory) stress. Among others, MK2 is known to be involved in the regulation of cytokine mRNA metabolism and regulation of actin cytoskeleton dynamics. Previously, MK2-deficient mice were shown to be highly resistant to LPS/d-Galactosamine-induced hepatitis. Additionally, research in various disease models has indicated the kinase as an interesting inhibitory drug target for various acute or chronic inflammatory diseases.

Results: We show that in striking contrast to the known resistance of MK2-deficient mice to a challenge with LPS/D-Gal, a low dose of tumor necrosis factor (TNF) causes hyperacute mortality via an oxidative stress driven mechanism. We identified in vivo defects in the stress fiber response in endothelial cells, which could have resulted in reduced resistance of the endothelial barrier to deal with exposure to oxidative stress. In addition, MK2-deficient mice were found to be more sensitive to cecal ligation and puncture-induced sepsis.

Conclusions: The capacity of the endothelial barrier to deal with inflammatory and oxidative stress is imperative to allow a regulated immune response and maintain endothelial barrier integrity. Our results indicate that, considering the central role of TNF in pro-inflammatory signaling, therapeutic strategies examining pharmacological inhibition of MK2 should take potentially dangerous side effects at the level of endothelial barrier integrity into account.

Figures

Figure 1
Figure 1
Effect of MK2-deficiency on morbidity and mortality in systemic inflammation and sepsis. (A-B) Body temperature and mortality for WT (n = 3) and MK2−/− (n = 5) mice after i.v. injection of 10 mg/kg LPS. (C) Body temperature for WT (n = 5) and MK2−/− (n = 9) mice after i.v. injection of 50 or 125 μg/kg TNF. Dose groups were pooled together because statistical comparison within one genotype revealed no differences. (D) Mortality for WT (n = 10) and MK2−/− (n = 13) mice after i.v. injection of 10, 50, 125 or 250 μg/kg TNF. Dose groups were pooled together because statistical comparison within one genotype revealed no differences. (E) Mortality for WT (n = 9) and MK2−/− (n = 10) mice after CLP surgery. Survival curves of different genotypes were compared via log-rank test. ****, p ≤ 0.0001; **, p ≤ 0.01.
Figure 2
Figure 2
IL-1β, IL-6, hexosaminidase and lactate dehydrogenase levels. (A-B) Plasma IL-1β and IL-6 levels for WT and MK2−/− mice, 2.5 h after i.v. injection of 450 μg/kg TNF (nPBS (WT and MK2−/−) = 10, nTNF (WT) = 5, nTNF (MK2−/−) = 5). (C-D) Plasma hexosaminidase and lactate dehydrogenase plasma levels for WT and MK2−/− mice, 2.5 h after i.v. injection of 450 μg/kg TNF. Pooled data from 2 separate experiments is shown (nPBS (WT and MK2−/−) = 17, nTNF (WT) = 10, nTNF (MK2−/−) = 8). Comparisons were made between baseline (PBS) and TNF challenged animals (#), and between genotypes (*) via one-way ANOVA with Sidak’s multiple comparisons test. Error bars indicate SD. ****, p ≤ 0.0001; **, p ≤ 0.01; *, p ≤ 0.05.
Figure 3
Figure 3
Peroxide equivalents, NO x levels, and effect of antioxidant treatment on mortality. (A) Plasma peroxide equivalents for WT and MK2−/− mice, 2.5 h after i.v. injection of 125 μg/kg TNF. Pooled data from 2 separate experiments is shown (nPBS (WT and MK2−/−) = 13, nTNF (WT) = 11, nTNF (MK2−/−) = 12). (B) Plasma NOx levels for WT and MK2−/− mice, 2.5 h after i.v. injection of 125 μg/kg TNF. Pooled data from 2 separate experiments is shown (nPBS (WT and MK2−/−) = 12, nTNF (WT) = 7, nTNF (MK2−/−) = 9). Comparisons were made between baseline (PBS) and TNF challenged animals (#), and between genotypes (*) via one-way ANOVA with Sidak’s multiple comparisons test. (C) Mortality for TNF challenged MK2−/− mice after pre- and post-treatment with tempol or SOD. Survival curves of different treatment groups were compared to controls via log-rank test (*). Error bars indicate SD. ****, p ≤ 0.0001; ***, p ≤ 0.001; **, p ≤ 0.01; *, p ≤ 0.05.
Figure 4
Figure 4
Assessment of cytoskeletal integrity via quantification of F-actin structures and density in liver endothelial cells. (A) Representative liver z-stacks stained for F-actin (green) and DAPI (blue), with a focus on the endothelial cell lining of a small vessel for WT and MK2−/− mice, 2.5 h after i.v. injection of PBS or 450 μg/kg TNF; (nPBS (WT and MK2−/−) = 10 × 3 vessels/animal, nTNF (WT) = 5 × 3, nTNF (MK2−/−) = 5 × 3). For every vessel, part of the endothelial cell sheet was defined as region of interest (ROI). (B) Actin density, expressed as the percentage of F-actin positive voxels per ROI. (C) The number of segmented F-actin positive structures per 100 000 voxels. (D) The average size of the actin structures, expressed as the total number of F-actin positive voxels per actin structure, normalized over the number of discrete actin structures. Comparisons were made between baseline (PBS) and TNF challenged animals (#), and between genotypes (*) via one-way ANOVA with Sidak’s multiple comparisons test. Error bars indicate SD. *, p ≤ 0.05.
Figure 5
Figure 5
Vascular permeability assessed by extravasation of FITC-Dextran (4 kDa). FITC-Dextran quantification for WT and MK2−/− mice, 2.5 h after i.v. injection of 450 μg/kg TNF for liver (A), kidney (B), lung (C), heart (D), and spleen (E); (n = 5 × 2 for liver, kidney and lungs; n = 5 for heart and spleen). Comparisons were made between baseline (PBS) and TNF challenged animals (#), and between genotypes (*) via one-way ANOVA with Sidak’s multiple comparisons test. Error bars indicate SD. ***, p ≤ 0.001; **, p ≤ 0.01; *, p ≤ 0.05.

Similar articles

See all similar articles

Cited by 3 articles

References

    1. Ronkina N, Kotlyarov A, Gaestel M. MK2 and MK3–a pair of isoenzymes? Front Biosci. 2008;13:5511–5521. doi: 10.2741/3095. - DOI - PubMed
    1. Cuadrado A, Nebreda AR. Mechanisms and functions of p38 MAPK signalling. Biochem J. 2010;429:403–417. doi: 10.1042/BJ20100323. - DOI - PubMed
    1. Kotlyarov A, Gaestel M. Is MK2 (mitogen-activated protein kinase-activated protein kinase 2) the key for understanding post-transcriptional regulation of gene expression? Biochem Soc Trans. 2002;30(Pt 6):959–963. - PubMed
    1. Ben-Levy R, Hooper S, Wilson R, Paterson HF, Marshall CJ. Nuclear export of the stress-activated protein kinase p38 mediated by its substrate MAPKAP kinase-2. Curr Biol. 1998;8:1049–1057. doi: 10.1016/S0960-9822(98)70442-7. - DOI - PubMed
    1. Anderson P. Post-transcriptional control of cytokine production. Nat Immunol. 2008;9:353–359. doi: 10.1038/ni1584. - DOI - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources

Feedback