Clofazimine, but Not Isoniazid or Rifampicin, Augments Platelet Activation in vitro

Abstract

Although the inclusion of the cationic amphiphilic, anti-mycobacterial agent, clofazimine, in the chemotherapeutic regimens of patients with multidrug-resistant tuberculosis (TB) has contributed to improved outcomes, concerns remain about the cardiotoxic potential of this agent. Accordingly, the current study was undertaken with the primary objective of investigating the effects of clofazimine, on the reactivity of human platelets in vitro, a seemingly unexplored, mechanism of cardiotoxicity. Platelet-rich plasma (PRP) prepared from the blood of healthy, adult humans was treated with clofazimine (0.625–10 mg/L), or the primary anti-TB agents, isoniazid and rifampicin (at final concentrations of 5 and 10 mg/L), followed by addition of either adenosine 5′-diphosphate (ADP) or thrombin and measurement of platelet activation according to the magnitude of expression of CD62P (P-selectin), as well as the CD62P-mediated formation of heterotypic neutrophil:platelet (NP) aggregates, using flow cytometry. Clofazimine, but neither isoniazid nor rifampicin, caused dose-related potentiation of both ADP- and thrombin-activated expression of CD62P by platelets, achieving statistical significance at threshold concentrations of 0.625 and 2.5 mg/L, respectively, as well as significant formation of N:P aggregates. These stimulatory effects of clofazimine on platelet activation were partly attenuated by pre-treatment of PRP with the membrane-stabilizing agent, α-tocopherol, possibly consistent with a membrane-disruptive mechanism. In conclusion, clofazimine, at concentrations within the therapeutic range, augments platelet activation in vitro, probably by a mechanism linked to membrane destabilization. If operative in vivo, these pro-thrombotic activities of clofazimine may predispose for development of microvascular occlusion, exacerbating an already existing high risk for development of TB-associated cardiovascular disease.

ADP-Mediated Upregulation of Expression of CD62P on Human Platelets Is Critically Dependent on Co-Activation of P2Y1 and P2Y12 Receptors

“…After 15 min of incubation in the dark, the samples were analyzed on a Gallios flow cytometer (Beckman Coulter) and the results expressed as the percentage of activated platelets with 50,000 cells interrogated during each measurement. The gating and analytical strategies used in a typical experiment have been described elsewhere [54].…”

Anti-Inflammatory Effects of Salbutamol on Human Bronchial Epithelial Cells Stimulated by Cigarette Smoke Extract

“…Salbutamol inhibited the ROS production in RAW264.7 macrophages stimulated by lipopolysaccharide (26) . In neutrophils stimulated by N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), salbutamol reduced the generation of ROS (27) . Our results are in agreement with the previous studies: salbutamol reduced the ROS production when compared to CSE-stimulated BEAS-2B cells.…”

Role of non-cardiomyocytes in anticancer drug-induced cardiotoxicity: A systematic review

“…Clofazimine upregulated Cluster Of Differentiation 62 Platelet (CD62P) expression, enhanced ADP, and activation of thrombin-mediated human platelet in vitro , which appears to be a novel cardiotoxic mechanism ( Anderson et al., 2018 ). CD62P is widely recognized as a critical factor in activating endothelial cells and react situations of living blood platelet.…”