The influence of ferrylhemoglobin and methemoglobin on the human erythrocyte membrane

Abstract

The aim of the study was to examine and compare the effects of methemoglobin (metHb) and ferrylhemoglobin (ferrylHb) on the erythrocyte membrane. Kinetic studies of the decay of ferrylhemoglobin (^*HbFe(IV)=O denotes ferryl derivative of hemoglobin present 5 min after initiation of the reaction of metHb with H₂O₂; ferrylHb) showed that autoredecay of this derivative is slower than its decay in the presence of whole erythrocytes and erythrocyte membranes. It provides evidence for interactions between ferrylHb and the erythrocyte membrane. Both hemoglobin derivatives induced small changes in the structure and function of the erythrocyte membrane which were more pronounced for ferrylHb. The amount of ferrylHb bound to erythrocyte membranes increased with incubation time and, after 2 h, was twice that of membrane-bound metHb. The incubation of erythrocytes with metHb or ferrylHb did not influence osmotic fragility and did not initiate peroxidation of membrane lipids in whole erythrocytes as well as in isolated erythrocyte membranes. Membrane acetylcholinesterase activity increased by about 10% after treatment of whole erythrocytes with both metHb and ferrylHb. ESR spectra of membrane-bound maleimide spin label demonstrated minor changes in the conformation of label-binding proteins in ferrylHb-treated erythrocyte membranes. The fluidity of the membrane surface layer decreased slightly after incubation of erythrocytes and isolated erythrocyte membranes with ferrylHb and metHb. In whole erythrocytes, these changes were not stable and disappeared during longer incubation.     

Cancer cell killing by Celecoxib: Reality or just in vitro precipitation‐related artifact?

Among NSAIDs Celecoxib is one of the most efficient in triggering in vitro cancer cell death, and from this perspective has been subject of numerous studies. However, it is still controversial whether this in vitro‐observed effect can also occur in vivo and contribute to the antitumor action of the drug. Moreover, besides common agreement on the involvement of COX‐independent pathways, the mechanisms underlying Celecoxib toxicity are still unclear. In an attempt to shed light on these mechanisms, I found that cell death only occurs at insoluble concentrations of the drug, and follows irreversible binding and damage of the plasmamembrane by precipitates. This evidence strongly suggests that Celecoxib is devoid of true molecular toxicity. Moreover, since plasma levels reached during therapy are far below the threshold of toxic precipitation, direct cytotoxicity by Celecoxib is unlikely to occur on tumor cells in vivo. Thus the antitumor effect might be only due to COX inhibition, which requires significantly lower levels of the drug. Nonetheless, direct cytotoxicity might not be confined to an in vitro artifact, but contribute to the upper gastrointestinal side effects of Celecoxib. Overall, these findings represent an important basis for further studies on Celecoxib, where true molecular actions of the drug should be discriminated from the precipitate‐dependent ones, and the relationship between in vitro and in vivo effects considered at the light of the precipitate‐dependent model. Moreover, remarkably, this article indicates a model of critical analysis that can be extended to other poorly soluble drugs. J. Cell. Biochem. 114: 1434–1444, 2013. © 2013 Wiley Periodicals, Inc.