Endothelin antibodies

Studies in animal models using endothelin antibodies are now being reported. Miyamori et al. demonstrated in rabbits [163] that endothelin antibody administration failed to show any appreciable changes in systemic or renal haemodynamic parameters suggesting that endothelin in the circulation may not contribute to blood pressure maintenance under normal conditions. However, its role as a local modulator is as yet not defined. 

In spontaneously hypertensive rats, endothelin has been shown, by studies using endothelin antibodies, to play an important role in the modulation of systemic blood pressure and renal function [164], but there were no significant effects in Wistar Kyoto rats which may therefore be less sensitive to endothelin [165]. 

Endothelin has been implicated in myocardial infarction, coronary vasospasm and re-stenosis subsequent to percutaneous transluminal coronary angioplasty. The potent, prolonged coronary vasoconstrictor and mitogenic actions of endothelin are well-documented (see earlier section) and thus endothelin is certainly able to produce the biological effects seen in these disease states. Although a reduction in infarct size has been demonstrated in a rat model of myocardial infarction with an endothelin antibody [185], and with phosphoramidon [186], the argument for a role for endothelin in ischaemic heart disease has been based mainly on the finding of increased plasma endothelin levels in patients with myocardial infarction [187-190], coronary vasospasm [191, 192] and re-stenosis post-PTCA [193-195]. 

The immunosuppressant agents, cyclosporin A (cys A) and FK506 both have the potential to induce acute renal failure and endothelin has been proposed as the mediator. Cys A and FK506 have both been shown to stimulate ET-1 release from cultured kidney cells [200]. An endothelin antibody prevented the reduction in function induced by cys A in a rat kidney perfusion model [201]. In the same model, BQ 123 also prevented the detrimental renal effects of cys A [202]. 

L20. Lundblad, R and Giercksky, K. E Effect of volume support, antibiotic therapy, and monoclonal antiendotoxin antibodies on mortality rate and blood concentrations of endothelin and other mediators in fulminant intra-abdominal sepsis in rats. Crit. Care Med. 23, 1382-1390 (1995). 

The endothelin B receptor is an example of characterization of a homogeneous, affinity purified protein (Roos et al., 1998). Significant progress has been made in the development of techniques for more high-throughput identification of phosphorlyation events. Analysis of large sets of phosphorylated proteins is facilitated by the availability of affinity purification methods such as anti-phosphotyrosine or anti-phosphoserine antibodies or metal affinity chromatography (Neubauer and Mann, 1999 Soskic et al., 1999). These methods are not specific to a particular protein but rather are used to fractionate all proteins that are phosphorylated. 

Information on receptors, using antibodies, was first provided by Kondoh et al. This group prepared monoclonal antibodies which were reactive with endothelin receptors by the immunization of mice with rat lung membranes [167]. Four clones were isolated and these demonstrated that there was more than one receptor as they exhibited different affinities to two ET receptors. One of these receptors was of 32 kDa and the other 45 kDa. 

The increasing availability of antibodies should facilitate studies into whether or not ET-1 is synthesized and released at or near the site of action and make it possible to determine the levels of endothelin in pathological tissue samples and correlate this to the state of a disease. 

Endothelin converting enzyme (ECE) has yet to be fully characterized. It is a membrane-located enzyme found in the vascular endothelium. It is essential in the production of endothelin in the body since it converts the inactive precursor big ET-1 to endothelin-1. It is an unusual enzyme because it cleaves at a Tyr-Val link. Like endopeptidase-24.11, it is inhibited by phosphoramidon (so is a metalloproteinase), and these two enzymes are often colocated. Furthermore, monoclonal antibody co-precipitation studies indicate they share a common epitope, and a homology to the extent of about 39%, ECE is also similar to the bacterial metalloprotease thermolysin, but is more specific. If a specific inhibitor is discovered that can act in vivo, then clearly such a drug could modulate endothelin production throughout the body, which would have important consequences (e.g. in antihypertensive therapy). 

The wells of a microtiter plate were coated with a purified mouse anti-Endothelin-1 monoclonal antibody and goat anti-rabbit IgG antibody. The wells were then post-coated by adding 1 % water-soluble gelatin solution containing... 

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