Cortistatin—can it or can it not prevent vascular calcification by modulation of Wnt signalling?

In this issue of Acta Physiologica, a paper by Liu et al1 examines the mechanisms by which the neuropeptide cortistatin ameliorates calcification in animal and cell-based models of vascular calcification. Previous work by the authors established that cortistatin was able to inhibit medial vascular calcification and associated disease pathology, and the current work goes on to show a strong association between the beneficial effects of cortistatin and modulation of Wnt signalling pathways. Vascular calcification is recognized as one of the main causes of arterial stiffness. Arterial stiffness is a well-established independent risk factor for cardiovascular disease and of specific relevance is the clinical significance of aortic calcification, with the associated increase in stiffness resulting in a reduced capacity of the vascular system to buffer pulsatile changes in blood pressure, which puts vulnerable end organs at risk of microvascular damage, particularly the heart, kidney and brain.2 Vascular calcification can be classified as intimal atherosclerotic calcification or medial calcification (arteriosclerosis).3 In atherosclerotic calcification, subpopulations of vascular smooth muscle cells undergo mineralization in response to vascular injury associated with inflammation, oxidative stress and lipid deposition. Arteriosclerosis is characterized by calcification along the elastin fibres surrounding vascular smooth muscle cells and is a hallmark feature of the vascular pathology associated with type 2 diabetes, chronic kidney disease and ageing.4 The pathogenesis of vascular calcification in these disease states is complex but key to the process is a phenotypic switch of the vascular smooth muscle cell to osteoblast-like cells, associated with a change in expression of lineage markers from vascular to osteogenic. This switch is consistent with the fact that vascular smooth muscle cells and chondrocytes arise from a common progenitor mesenchymal stem cell and share the capacity to express osteochondrogenic markers. The Wnt/β-catenin signalling pathway is known to be a key regulator of osteo/chondrogenic differentiation and plays a critical role in bone metabolism.3 How this pathway can be impacted by corticostatin was specifically explored in the work by Liu et al1 Cortistatin is a neuropeptide that shares high structural and functional similarities with somatostatin (SST) and has been shown to bind to not only the SST G-protein coupled receptor family but also the GHSR1a ghrelin receptor. Cortistatin, named as such due to its predominant expression in the cortex and hippocampus, was first identified in the rat, with precortistatin cleaved into the active peptide cortistatin-14 (rat) or cortistatin-17 (human).5 It is important to note that the neuropeptide cortistatin should not be confused with the cortistatin steroidal alkaloids, identified from the marine sponge Corticium simplex. Like many other neuropeptides, in addition to its expression and function in the central nervous system, cortistatin has also been shown to be expressed outside of the central nervous system and have peripheral actions. These actions are of an anti-inflammatory nature and have been shown in various tissues including in the cardiovascular system.6 A number of different intracellular signalling pathways have been linked to cortistatin including the expression of cyclins, the cAMP/protein kinase A and p38-MAPK pathways, Akt and ERK activities and the GTPase Rac1.6 How cortistatin interacts with the different Wnt signalling pathways under pathological conditions has not been previously characterized. The work of Liu et al1 shows in a series of well controlled experiments that cortistatin interacts with the Wnt pathway at two points, namely through the canonical pathway by increasing the activity of glycogen synthase kinase 3 beta (GSK3-β) and thus reducing β-catenin levels, which would otherwise translocate to the nucleus and drive osteogenic differentiation, and through the Wnt/Ca2+ non-canonical pathway, increasing the expression of activated (phosphorylated) protein kinase C (p-PKC), which through the suppression of osteogenic differentiation and as an intrinsic regulator of bone resorption, has marked anti-calcification actions. In their first series of experiments, Liu et al confirmed that in both their animal model, being a rapid aortic calcification model induced by high-dose vitamin D3 injections over a period of 3 days, and their isolated vascular smooth muscle cell calcification model (driven by β-glycerophosphate), cortistatin was able to both promote an increase in the vascular smooth muscle cell lineage marker alpha smooth muscle actin, as well as inhibit the expression of the osteogenic lineage markers alkaline phosphatase and osteocalcin. Notably, both alkaline phosphatase and osteocalcin are not just markers but also likely active mediators of vascular calcification. They then specifically investigated the impact of corticostatin on the GSK3-β/β-catenin pathway, using western blots to show that cortistatin reduced β-catenin protein levels in both the animal and cell culture models, and prevented the relative increase in p-GSK3β in the cell culture model, pushing the balance towards active GSK3β, which functions as part of a complex with Axin and adenomatous polyposis coli (APC) to phosphorylate β-catenin, leading to its ultimate degradation.3 The authors did not confirm the increase in phosphorylated β-catenin but they did show that inhibition of GSK3β with lithium chloride in vitro prevented the protective effects of cortistatin, providing a strong foundation for their subsequent conclusions. The final part of the study examined if cortistatin was able to limit vascular calcification mechanisms via non-canonical Wnt signalling pathways. Using their in vitro model, the authors again used western blot, this time to assess if cortistatin influenced either the Wnt-c-Jun N-term inal kinase pathway or the Wnt/Ca2+ signalling pathway as measured by levels of JNK and PKC, respectively, in their inactive and phosphorylated forms. While cortistatin had no impact on JNK, it did act to increase levels of p-PKC. Similar to the studies using lithium chloride, Liu et al1 validated their finding by showing that Go6976, a PKC inhibitor, prevented the anti-calcification and osteogenic differentiation effects of cortistatin, and indeed, inhibition of PKC was able to induce a more severe ossification phenotype. This in and of itself is an important finding and presents the exciting possibility that modulation of PKC activity could be an important target for the prevention of vascular calcification. What was not determined in the study was the isoform of PKC involved, although the antagonist Go6976 is an inhibitor of the isoenzymes PKCα and PKCβ, nor were the downstream effectors investigated, and these will be important studies that hopefully follow. Vascular calcification is not the only cardiovascular pathology that cortistatin is able to limit. In a recent study by Delgado-Maroto et al7, treatment of hyperlipidemic ApoE-deficient mice with cortistatin had a significant beneficial effect in preventing the development of atherosclerotic plaques in the heart, carotid artery and aorta. Although a different model of vascular calcification, Delgado-Maroto et al7 demonstrated that cortistatin downregulated the vascular inflammatory and T-cell-mediated response, reducing the ability of endothelial cells to recruit monocytes and atherogenic T cells into the plaque, and reducing the formation of foam cells (by enhancing cholesterol efflux from macrophages). Of note, however, is that in both the study by Liu examining arteriosclerotic mechanisms1, and the work of Delgado-Maroto examining atherosclerotic lesions7, treatment regimens were initiated in acute onset animal models very early in the course of disease establishment. From a translational perspective, it remains to be established if cortistatin has any effect on established vascular lesions which have progressed over a longer time course, more consistent, for example, with that which occurs in humans with ageing, or in the presence of co-morbidities such as chronic kidney disease or diabetes. The author declares no conflict of interest.