Investigations on 5-HT₄ receptor expression and effects of tegaserod on human platelet aggregation in vitro.
A meta-analysis of 29 clinical studies on tegaserod revealed an imbalance of cardiovascular ischemic events in patients treated with drug versus placebo. Because increased platelet activity is known to attribute to cardiovascular events, we examined the presence of serotonin receptor type 4 (5-HT₄) receptors and the effects of tegaserod on in vitro aggregation of human platelets. Blood samples were obtained from 20 healthy volunteers and 20 subjects with irritable bowel syndrome with constipation. Samples of whole blood-citrate mixtures were incubated with different tegaserod concentrations mimicking human Cmax values (10 nM), 3.3 times, and 10 times Cmax for at least 1 hour. Conventional plasma platelet aggregation was induced by adenosine diphosphate, collagen, thrombin receptor activating peptide, epinephrine, and serotonin plus adenosine diphosphate. Gene expression analyses targeting 5-HT₄ and serotonin receptor type 2 receptors were carried out using human platelet RNA. The presence of 5-HT₄ receptor protein was investigated by Western blot analysis using membrane fractions from human platelets. Preincubation with tegaserod resulted in mild but statistically significant increases in platelet aggregation induced by adenosine diphosphate, collagen, epinephrine, or serotonin, mostly at supratherapeutic concentrations of tegaserod. The effects were comparable using thrombocytes obtained from healthy volunteers and patients with irritable bowel syndrome with constipation. Expression analysis revealed that mRNA encoding both 5-HT₄ and serotonin receptor type 2 receptors was present in human platelets. The expression of 5-HT₄ receptor mRNA was approximately eightfold lower than serotonin receptor type 2 receptor mRNA. Results from Western blot analyses examining the presence of 5-HT₄ receptor protein in human platelets were in agreement with the findings of the mRNA expression analysis. In platelets harvested from normal persons and patients with irritable bowel syndrome with constipation and exposed in vitro to tegaserod, we detected small but statistically significant concentration-dependent increases in induced platelet aggregation. The relationship of these in vitro effects to clinical cardiovascular ischemic events is presently unclear. Western blot analysis findings suggest the presence of 5-HT₄ receptor protein on human platelets. Further investigations on the potential role of 5-HT₄ receptors in human platelets may be warranted.
HeartDrug™ Research Laboratories, Johns Hopkins University Baltimore, MD, USA. Heartdrug@aol.com
Victor L
VL
Serebruany
Mohamed
M
El Mouelhi
Hans-Jürgen
HJ
Pfannkuche
Kristine
K
Rose
Martin
M
Marro
Dominick J
DJ
Angiolillo
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000328:Adult; D000368:Aged; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004837:Epinephrine / Q000494:pharmacology; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology*; D008297:Male; D008875:Middle Aged; D010974:Platelet Aggregation / Q000187:drug effects*; D012333:RNA, Messenger / Q000096:biosynthesis; D044348:Receptors, Serotonin, 5-HT2 / Q000096:biosynthesis / Q000235:genetics; D044422:Receptors, Serotonin, 5-HT4 / Q000096:biosynthesis* / Q000235:genetics; D012701:Serotonin / Q000494:pharmacology; D058828:Serotonin 5-HT4 Receptor Agonists / Q000494:pharmacology*
D007211:Indoles; D012333:RNA, Messenger; D044348:Receptors, Serotonin, 5-HT2; D058828:Serotonin 5-HT4 Receptor Agonists; D044422:Receptors, Serotonin, 5-HT4; D012701:Serotonin; C105050:tegaserod; D000244:Adenosine Diphosphate; D004837:Epinephrine