👤 M Kluger

ECs lining arteries respond to LSS by suppressing pro-inflammatory changes, in part through the activation of MEK5, ERK5 and induction of KLF4. We examined if this anti-inflammatory pathway operates in human ECs lining microvessels, the principal site of inflammatory responses. We used immunofluorescence microscopy of human skin to assess ERK5 activation and KLF4 expression in HDMECs in situ. We applied LSS to or overexpressed MEK5/CA in cultured HDMECs and assessed gene expression by microarrays and qRT-PCR and protein expression by Western blotting. We assessed effects of MEK5/CA on TNF responses using qRT-PCR, FACS and measurements of HDMEC monolayer electrical resistance. We used siRNA knockdown to assess the role of ERK5 and KLF4 in these responses. ERK5 phosphorylation and KLF4 expression is observed in HDMECs in situ. LSS activates ERK5 and induces KLF4 in cultured HDMECs. MEK5/CA-transduced HDMECs show activated ERK5 and increased KLF4, thrombomodulin, eNOS, and ICAM-1 expression. MEK5 induction of KLF4 is mediated by ERK5. MEK5/CA-transduced HDMECs are less responsive to TNF, an effect partly mediated by KLF4. MEK5 activation by LSS inhibits inflammatory responses in microvascular ECs, in part through ERK5-dependent induction of KLF4. Show less

Apolipoprotein A-V (apoA-V) was discovered in 2001 both by comparative sequencing and as a liver regeneration protein. The gene is a located at the APOA1/C3/A4/A5 gene cluster on chromosome 11q23, a locus well known for playing a major role in regulating plasma cholesterol and triglyceride (TG) levels. ApoA-V is produced in the liver and has very low plasma concentrations (0.1-0.4 mug/ml). Mice lacking apoA-V have 4-fold increased TG levels, whereas apoA-V overexpression leads to 40% plasma TG reduction. Based on metabolic studies in vivo, apoA-V enhances the catabolism of TG rich lipoproteins rather than affecting their intestinal or hepatic production. By activating proteoglycans-bound lipoprotein lipase (LPL), apoA-V can accelerate TG hydrolysis from VLDL and chylomicrons independent from other apoproteins. Several variants at the APOA5 gene locus have been detected in humans. Some single nucleotide polymorphisms (SNPs) are associated with significantly higher plasma TG levels in patients (e.g., -1131T > C, S19W, G185C). In addition, these SNPs may affect fibrate response and obesity. However, data for a possible association of APOA5 variants with coronary heart disease are not consistent. Severe structural mutations (Q139X, Q148X, IVS3 + 3G > C) predispose to familial hypertriglyceridaemia and late-onset chylomicronaemia. Thus, despite its low plasma concentration, apoA-V is a major regulator of plasma TG metabolism in humans. However, the precise mechanism of its function is not yet clear. Show less

Apolipoprotein A5 (APOA5) is associated with differences in triglyceride levels and familial combined hyperlipidemia. In genetically engineered mice, apoAV plasma levels are inversely correlated with plasma triglycerides. To elucidate the mechanism by which apoAV influences plasma triglycerides, metabolic studies and in vitro assays resembling physiological conditions were performed. In human APOA5 transgenic mice (hAPOA5tr), catabolism of chylomicrons and very low density lipoprotein (VLDL) was accelerated due to a faster plasma hydrolysis of triglycerides by lipoprotein lipase (LPL). Hepatic VLDL and intestinal chylomicron production were not affected. The functional interplay between apoAV and LPL was further investigated by cross-breeding a human LPL transgene with the apoa5 knock-out and the hAPOA5tr to an lpl-deficient background. Increased LPL activity completely normalized hypertriglyceridemia of apoa5-deficient mice; however, overexpression of human apoAV modulated triglyceride levels only slightly when LPL was reduced. To reflect the physiological situation in which LPL is bound to cell surface proteoglycans, we examined hydrolysis in the presence or absence of proteoglycans. Without proteoglycans, apoAV derived either from triglyceride-rich lipoproteins, hAPOA5tr high density lipoprotein, or a recombinant source did not alter the LPL hydrolysis rate. In the presence of proteoglycans, however, apoAV led to a significant and dose-dependent increase in LPL-mediated hydrolysis of VLDL triglycerides. These results were confirmed in cell culture using a proteoglycan-deficient cell line. A direct interaction between LPL and apoAV was found by ligand blotting. It is proposed, that apoAV reduces triglyceride levels by guiding VLDL and chylomicrons to proteoglycan-bound LPL for lipolysis. Show less