What is the pathophysiological role of ADMA?

Cardiovascular diseases are the major cause of death in North America and in the Western European countries.
Traditional cardiovascular risk factors like hypercholesterolemia, hypertension, smoking, and diabetes mellitus can explain up to 80% of coronary events occurring in the population of these countries. In some patient groups with extraordinarily high coronary event rates, like hemodialysis patients, an even larger portion of cardiovascular events remains unexplained by traditional risk factors [12].
Intense research into the molecular and cellular mechanisms underlying atherogenesis has led to the understanding that the vascular endothelium plays a crucial role for early functional changes in the vascular wall which finally initiate and promote the atherogenic process.

Figure 4. The ground-breaking experiment by Robert Furchgott and co-workers from the year 1980, for which he was awarded the Nobel Prize 18 years later, for the first time demonstrated that endothelial cells secrete a soluble factor (which was later identified as nitric oxide (NO)) which causes vasodilation. Stimulation of the left arterial segment (of which the endothelium is left intact) with acetylcholine causes NO release, which in turn causes relaxation of the right arterial segment of which the endothelium was mechanically rubbed off. Direct stimulation of the right arterial segment with acetylcholine would cause vasoconstriction due to direct action of acetylcholine on vascular smooth muscle cells.

There are numerous experimental data showing that the vascular endothelium plays a central role in the maintenance of physiological vascular tone and vascular structure [13]. One of the major mediators that are released by healthy endothelial cells is nitric oxide (NO) [14]. NO is formed by the enzyme NO synthase from the amino acid precursor L-arginine. NO is involved in a vast number of regulatory processes within the cardiovascular system. Its potent vasodilatory effect is most widely known, and this is the one that has led to the discovery, in the early 1980`s, of an "endothelium-derived relaxing factor (EDRF)" by Robert Furchgott and co-workers [15] (Robert Furchgott received the Nobel Prize for Medicine and Physiology in 1998 for this discovery) (Figure 4).

Besides its potent vasodilatory effects, NO also acts as an endogenous inhibitor of platelet aggregation. Furthermore, NO inhibits the adhesion of monocytes and leukocytes at the healthy vascular endothelium - an effect that, once disturbed, precedes the immigration of inflammatory cells into the vascular wall at sites that later become plaques. It inhibits the proliferation of vascular smooth muscle cells - this might be of great importance during the development of restenosis after angioplasty. Moreover, NO reduces the vascular release of superoxide radicals (O2-), radicals that are involved in inflammatory and cytotoxic processes, and it inhibits LDL oxidation. These numerous salutary actions of NO in the vascular system have led to its name as an "endogenous anti-atherogenic molecule" (Figure 5).

Figure 5. Schematic representation of the manifold physiologically relevant actions of NO as an endogenous anti-atherogenic molecule (from [16] with kind permission of the publishers).

What is the importance of ADMA in this context? Under experimental conditions that lead to sub-optimal L-arginine concentrations or to a relative deficiency of essential co-factors for NO synthase, the activity of this enzyme is "un-coupled". This means that the oxidation of L-arginine to NO is not complete [17-19]. Normally, five electrons are being transferred in two steps of a coupled reduction-oxidation reaction by the two domains of NO synthase from molecular oxygen to L-arginine, resulting in the release of L-citrulline and NO (Figure 6a).

Under suboptimal conditions like those mentioned above, the electron flow within the two domains of NO synthase is disturbed, and molecular oxygen acts as an electron acceptor. This makes NO synthase a superoxide (O2-) radical-producing enzyme (Figure 6b).

Figure 6. Mechanism of the biochemical reaction catalyzed by NO synthase. The enzyme consists of two subunits. Physiologically, electrons are transferred from molecular oxygen to L-arginine via several essential co-factors (A). Under pathophysiological conditions the NO synthase reaction mechanism is "uncoupled", i.e. the flow of electrons from the reductase domain to the oxidase domain is interrupted - and thereby the oxidation of L-arginine. In this situation molecular oxygen can be an electron acceptor - making NO synthase a superoxide radical producing enzyme (B) (from [20] with kind permission of the publishers).

Interestingly, cultured human endothelial cells produce O2- in the presence of ADMA. This led to the hypothesis that ADMA may interrupt the NO-producing activity of NO synthase and "un-couple" the enzyme, which results in a "switch" of the enzymatic activity from NO to O2-. This in turn will lead to activation of Redox-sensitive transcription factors, to subsequent upregulation of endothelial adhesion molecules and, thereby, to increased adhesiveness of monocytes to the vascular lining - an early step in the initiation and progression of atherosclerosis (Figure 7) [21].

Figure 7. Adhesion of monocytes to cultured human endothelial cells under "normal" cell culture conditions (A) as well as in the presence of ADMA (B). ADMA induces oxidative stress within the endothelial cells - probably via the mechanisms described in detail in the text. This causes upregulation of endothelial adhesion molecules. Monocyte adhesion is regarded as the major initial step leading to the development of atherosclerotic plaques, which are understood to be locations of local vascular inflammation.

Under experimental conditions, the expression of adhesion molecules is indeed upregulated and leukocyte adhesion is increased in cultured human endothelial cells in the presence of high ADMA concentrations. A similar phenomenon can be observed when monocytes are isolated from peripheral blood of patients with cardiovascular risk factors and are co-incubated with cultured human endothelial cells: Monocytes from hypercholesterolemic patients adhere more strongly to the endothelium than monocytes from normocholesterolemic controls. In this context it is interesting to note that monocyte hyperadhesiveness in hypercholesterolemic subjects can be normalized by supplemental L-arginine [22]. This also points in favour of a competitive displacement of endogenous L-arginine (by ADMA) as the cause of these pathophysiological changes (Figure 8).

Figure 8. Adhesion of monocytes that were isolated from peripheral venous blood of hypercholesterolemic patients or normocholesterolemic controls to cultured human endothelial cells ex vivo. During daily intake of supplemental L-arginine monocyte adhesion is reduced, as L-arginine can diminish the effects of elevated ADMA on the endothelium (Data from [22]).