Brief reportsMelatonin is not stored in platelets
Introduction
Over 300 bioactive components have been detected in platelet releasates [1,2]. Once secreted these bioactive components act in an autocrine or paracrine way to modulate cell signaling.
One of these bioactive components was considered to be melatonin [3]. This is of relevance given the increasing interest in the potential roles of melatonin in the human body. It is already known that melatonin is a regulator of the circadian rhythm. Deregulation of the circadian rhythm and of circulating melatonin levels are associated with increased incidence of cancer, neurological, cardiovascular and metabolic diseases, and epigenetic abnormalities [4,5].
The finding of platelet melatonin was based on an ELISA assay [3]. We developed a refined method for melatonin detection to facilitate detailed melatonin research [6]. Therefore, we also wanted to confirm the storage of melatonin in platelets using a sensitive liquid chromatography in combination with isotope dilution tandem mass spectrometry (LC-MS/MS). Platelet-rich plasma (PRP) and platelet-poor plasma (PPP) served as proxy for platelet levels of melatonin, because isolation of platelets is prone to errors [7].
Section snippets
Material and methods
Twenty healthy volunteers over 18 years of age were included. All gave written informed consent. The study was waived by the Medical Ethical Review Committee of the University Medical Center Groningen. Blood (two 10 mL EDTA tubes per volunteer) was drawn at 8:00 AM using a butterfly needle. Blood was immediately processed to minimise platelet activation [7]. To obtain PRP, one EDTA tube per volunteer was centrifuged at 120g for 30 min at room temperature. Of this, 500 μL served to determine the
Results
Comparison of the LC-MS/MS and the ELISA method showed poor agreement for melatonin concentrations in PRP (slope = 2.35 [0.65 to18]; 95% CI, intercept = −100 [−1015 to −7.3]; 95% CI) and in PPP (slope = 1.68 [1.04–2.71]; 95% CI, intercept = −61 [−123 to −27]; 95% CI) (Fig. 1).
With both techniques, we found no evidence for storage of melatonin in platelets, as no difference between PRP and PPP was detected using the Wilcoxon signed rank test, P = .89 and P = .53 for the LC-MS/MS and ELISA
Discussion and conclusion
These results show that indeed plasma contains melatonin, but melatonin is not stored in platelets.
Our results are in contrast with a study where the same ELISA was used to demonstrate presence of melatonin in platelets of only three healthy individuals [3]. This difference can be explained by the following differences, first we studied a larger group of twenty subjects. Secondly, we used the difference between PRP and PPP as proxy for platelets. Isolation of platelets is prone to errors, as it
Author contributions
Study concept and design IK, SO, EdV, acquisition, analysis, and interpretation of data MvF, MP; drafting and critical revision of the manuscript MvF, MP, AW, EdV, SO, IK.
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These authors contributed equally.