Kruppel-like factors and vascular inflammation: implications for atherosclerosis.

Myocardial infarction, peripheral arterial disease, and stroke are highly morbid and mortal conditions [1] that are consequent to the progression of atherosclerotic plaque lesions in the intimal component of vascular wall. Atherosclerosis silently develops over decades of life from sub-endothelial fatty streaks deposition into established lesions compromising blood flow in the arterial vessel that may rupture causing an acute luminal occlusion and organ ischemia [2]. In spite of the higher prevalence reported in modern industrialized societies [1], atherosclerosis has impacted human health for centuries as evidenced by the recent documentation of lesions in ancient Egyptian mummies [3]. Clinical, pathologic, and experimental observations have fueled the current zeitgeist that atherosclerosis is a chronic inflammatory disease state. This central feature of atherosclerosis is under enormous investigative efforts to dissect the pro-and anti-inflammatory cellular and molecular mechanisms governing disease development [4]. A number of cellular pathways have been implicated in modulating this pathologic inflammatory process [5]. Studies utilizing high throughput technologies coupled with enormous advances in our understanding of the human genome have led to the discovery of many atheroregulatory candidates. Amongst the most exciting developments of the past decade has been the identification of a family of transcriptional regulators termed Kruppel-like Factors (KLFs) as athero-protective factors [6]. Indeed, studies from our group and others have demonstrated the key role of this family of transcription factors (KLFs) in governing cell activation and inflammatory responses in many cellular components of the vascular wall and plaque lesion. The nomenclature of KLFs came from the German word kruppel (for “cripple”), a protein first described in Drosophila [7]. Embryos with mutated kruppel had a lethal developmental defect in body patterning and a “crippling” appearance. In 1993, the first mammalian homolog to kruppel was identified in red blood cell lineage, therefore named erythroid Kruppel-like factor (EKLF/KLF-1) [8]. Over the last two decades, a total of 17 members were identified in mammalians constituting a large family of KLFs that broadly regulate numerous physiological and pathological processes in many cell types and organ systems [.9], from fetal development to circadian regulation of cardiac repolarization [10, 11]. KLFs represent a subgroup of the zinc finger family of DNA-binding transcription factors, and they are distinguished by several features: i) three Cysteine2/Histidine2 containing zinc fingers located at the extreme C-terminus of the protein, ii) a highly conserved seven residue sequence between zinc fingers, TGEKP(Y/F)X, and iii) DNA-binding to specific sequences such as the “CACCC” sequence or the “GT box” [12–14]. The highly divergent non-DNA-binding regions mediate protein-protein interactions, and modulate transactivation and transrepression. In this review we summarize the emerging role of certain members of the KLF transcription factors in the endothelial, smooth muscle, and immune cells biology with a focus on inflammation and implications for atherosclerosis (summarized in Table 1). Table 1 Summary of KLFs role in different cell types relevant to vascular homeostasis Endothelial Cells Endothelial cell dysfunction plays a central role in the development of vascular disease states such as atherosclerosis and thrombosis [15]. The endothelium consists of a single layer of cells that line the blood vessel lumen and therefore is uniquely positioned to transduce and integrate both biochemical (e.g., inflammatory cytokines) and biomechanical (e.g., shear stress) stimuli from the blood to the underlying blood vessel wall [16] (book chapter). Inflammatory cytokines and non-laminar blood flow induce endothelial dysfunction and confer a pro-adhesive and pro-thrombotic phenotype. Accumulating evidence from our group and others shows that members of the KLF family critically regulate endothelial function and vascular homeostasis. Among the KLF family, KLF2, 4, and 6 have emerged as particularly important in endothelial biology. KLF2 KLF2 is highly expressed in ECs and has been implicated as a “molecular switch” regulating endothelial function in health and disease (reviewed in Atkins and Jain [15]). KLF2 is upregulated by laminar shear stress, as well as by 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (i.e., statins), and inhibited by turbulent shear stress, inflammatory cytokines, and oxidative stress. In turn, KLF2 differentially regulates the expression of factors that confer anti-inflammatory, antithrombotic, and antiproliferative effects in endothelial cells. The expression of KLF2 can be modulated at multiple levels, including transcriptional, posttranscriptional via miR-92a, and posttranslational modifications [17]. KLF2 has been shown to be regulated by MEF2 and MEK5/ERK5/MEF2 flow pathway [15] and more recently identified AMPK/ERK5/MEF2 pathway [18], thereby establishing a molecular link between flow and its downstream effects in endothelial cells. Our group was the first to establish KLF2 as an antiatherogenic factor in vivo, demonstrating that KLF2 hemizygous mice have increased plaque burden in the ApoE model of atherosclerosis. KLF2 strongly upregulates eNOS and thrombomodulin and inhibits cytokine mediated induction of cell adhesion molecules. KLF2 regulates endothelial inflammation by inhibiting expression of VCAM-1 and E-selectin in response to various proinflammatory cytokines, which correlated with decreased T cell attachment and rolling on endothelial monolayers [19]. KLF2 potently inhibits thrombin-mediated induction of multiple cytokines/chemokines (e.g., monocyte chemotactic protein 1, IL-6, and IL-8) by inhibiting expression of its principal receptor protease-activated receptor 1 [20]. Recent studies provide evidence that KLF2 plays an essential role in maintaining endothelial barrier integrity via differential effects on the expression of key junction protein occludin and modification of a signaling molecule myosin light chain [21]. Constitutive and endothelial-restricted KLF2−/− mice die at embryonic day 10.5 owing to lack of vascular tone, bleeding and cardiac dysfunction secondary to vascular complications [22, 23]. KLF2 is an inhibitor of HIF-1α expression and function and, therefore, potent inhibitor of angiogenesis [24]. KLF2 is also a regulator of the key VEGF receptor VEGFR2 [25]. A recent exciting finding may provide a promising strategy to combat atherosclerosis. KLF2-transduced cells release vesicles with a selective enrichment of miR-143/145 that regulates SMC gene expression and provide an atheroprotective effect in endothelial cells [26].