p38α MAP kinase inhibition in atherosclerosis – a new therapeutic tool?

Atherosclerotic cardiovascular disease is the leading cause of mortality worldwide. Selective recruitment of monocytes into the arterial intima and their transformation into macrophage-derived foam cells is one of the earliest events in the pathogenesis of atherosclerosis. Foam cell formation is induced by modifications of low density lipoprotein (LDL). It has recently been shown that important events during atherogenesis like endothelial apoptosis and expression and secretion of IL-8 by human monocyte–derived macrophages are induced by enzymatically modified LDL (eLDL) via mitogen-activated protein-kinase (MAPKs), especially p38MAPK. Furthermore, p38MAPK is activated by oxidized LDL (oxLDL) in vascular smooth muscle cells, and its activation is required for foam cell formation of macrophages exposed to oxLDL and is involved in oxLDL-induced macrophage proliferation. Thus, the control of the MAPK cascade could be targeted as a potential treatment of atherosclerosis. Selective p38 inhibitors could facilitate elucidating the impact of the complex network of p38 MAPK signaling on atherogenesis. Now, the first ATP-competitive p38α MAPK/MAPK14 inhibitor with excellent in vivo efficacy and selectivity, skepinone-L, is available. In the present study, we investigated the impact of selective p38α MAPK/MAPK14 inhibition on eLDL-stimulated human monocytes and its implications for atherosclerosis.

Fig. 1. A) mRNA expression of p38 MAPK isoforms in human PBMCs from 14 healthy donors. Expression levels are given as RQ of target genes (p38a MAPK/MAPK14, p38b MAPK/MAPK11, p38d MAPK/MAPK13, p38g MAPK/MAPK12) normalized to reference gene b-actin. B) Skepinone-L inhibits eLDL-induced CD36 expression. THP-1 cells were incubated with skepinone-L at 100 nM for 2 h and further exposed to 40 and 80 mg/ml protein eLDL for 6 h. mRNA expression levels are demonstrated as RQ of CD36 normalized to reference gene GAPDH. Untreated cells were used as control. Data are means 6 SD of 6 independent experiments. *P , 0.05, **P , 0.01, statistically significant difference between eLDL-treated cells and untreated control (by unpaired Student’s t test with Welch’s correction).

We used expression analysis, histochemical and immunohistochemical stainings, fluoremetric quantification measurements and Western Blot analysis of human monocytes and macrophages from peripheral blood mononuclear cells (PBMCs) of 14 healthy donors, sequential sections from human carotid endarterectomy specimens from patients undergoing carotid endarterectomy for high-grade stenosis of the internal carotid artery, as well as the human acute monocytic leukemia cell line THP-1 with and without incubation with eLDL and skepinone-L to investigate the impact of the selective p38α MAPK/MAPK14 inhibitor skepinone-L on atherosclerosis.

First of all, we could show that p38α/MAPK14 is the predominantly expressed p38MAPK isoform in human monocytes and macrophages (Fig. 1A). The activation of the p38 MAPK pathway was actually induced by eLDL. This activation could be inhibited by the selective p38α MAPK/MAPK14 inhibitor skepinone-L. Skepinone-L also inhibited eLDL induced expression of the class B scavenger receptor CD36 which is known to be involved in eLDL uptake (Fig. 1B). Finally, skepinone-L had a cell- and time-dependent effect on eLDL-triggered apoptosis, and inhibited eLDL stimulated secretion of IL-8 and MIP-1β/CCL4.

Thus, inhibition of a key signaling molecule of the p38 MAPK pathway, p38α MAPK/MAPK14, by selective inhibitors like skepinone-L with multifaceted effects on foam cell formation, apoptosis, and cytokine induction, facilitates elucidation of the impact of the complex network of p38 MAPK signaling on atherogenesis and might provide a promising therapeutic tool to prevent inflammatory cascades in atherosclerosis, not least because the potency of skepinone-L has been recently demonstrated.

Laura Twardowski
Department of Laboratory Medicine, Robert-Bosch-Hospital, Stuttgart, Germany

 

Publication

Selective p38α MAP kinase/MAPK14 inhibition in enzymatically modified LDL-stimulated human monocytes: implications for atherosclerosis.
Cheng F, Twardowski L, Fehr S, Aner C, Schaeffeler E, Joos T, Knorpp T, Dorweiler B, Laufer S, Schwab M, Torzewski M
FASEB J. 2017 Feb

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