Caspase activation in the absence of mitochondrial changes in granulocyte apoptosis

Abstract

SUMMARY Eosinophils and neutrophils are two different types of granulocytes evolved from a common haematopoetic precursor in the bone marrow. Eosinophils are mainly involved in parasitic infection and allergic inflammation while neutrophils mainly participate in the defence against bacterial infections. Prolongation of granulocyte life span by inhibition of apoptosis may lead to tissue load of cells, and this has been detected in different inflammatory reactions. The molecular mechanisms and the potential role of the mitochondria in granulocyte apoptosis are poorly understood. In the present study we have characterized further the role of the mitochondria in granulocyte-apoptosis by studying the sequence of mitochondrial permeability transition (MPT) induction, loss of mitochondrial membrane potential (Δψm) and release of cytochrome c. This was made possible by applying tributyltin (TBT), a well-characterized apoptotic stimulus and MPT-inducer. We also studied potential differences in apoptosis-susceptibility between eosinophils and neutrophils. Ten minutes of TBT-exposure resulted in a substantial caspase-3 activity in both eosinophils and neutrophils, followed by phosphatidylserine (PS)-exposure after 30–120 min. Interestingly, caspase-3 activity was not preceded by MPT-induction, loss of Δψm or by cytochrome c-release in either eosinophils or neutrophils. In conclusion, we have demonstrated an extremely rapid induction of caspase-3 activity and apoptosis in human blood granulocytes without prior mitochondrial changes, including loss of mitochondrial membrane potential and release of cytochrome c. Our results open the possibility for a mitochondrial-independent activation of caspase 3 and subsequent apoptosis in granulocytes.

Tributyltin (TBT) induces ultra-rapid caspase activation independent of apoptosome formation in human platelets

“…Our data in human platelets are in line with the recent findings in human eosinophils and neutrophils, where a rapid activation of caspase-3 by TBT was suggested to occur independent of mitochondrial alterations (Nopp et al, 2002). Using exogenous cytochrome c and dATP, we could confirm previous studies demonstrating that apoptosome formation could be induced in human platelets that contain all elements of the mitochondrial apoptosis pathway including Apaf-1, cytochrome c, caspase-9 and caspase-3 (Wolf et al, 1999).…”

Apoptotic related biochemical changes in human amnion cells induced by tributyltin

“…By means of flow cytometric method, Annexin V-FITC and propidium iodide (PI) were used to distinguish intact, dead and apoptotic cells, according to the change of cell membranes, for the phosphatidylserine (PS) can translocate to the outer leaflet of the cell membrane during the early stage of apoptosis (Nopp et al, 2002). The density of cell suspension was adjusted to 5 × 10 5 cell/mL and 195 L of the cell suspension was transferred into the microtubes.…”

“…TBT has been shown to induce apoptosis in various cells and tissues (Nopp et al, 2002;Reader et al, 1999;Stridh et al, 2001;Aw et al, 1990), and it is clearly known that in some cell type, the mechanisms responsible for TBT-induced apoptosis mainly involve the damage to mitochondrial function, the increase of the cytosolic free calcium concentration, the production of ROS, the release of cytochrome C from the mitochondrial membrane into the cytosol and the activation of caspase (Chow et al, 1992;Gennari et al, 2000;Tiano et al, 2003). However, the mechanisms are still not fully elucidated, such as: how are the three apoptotic pathways (mitochondrial pathway, death receptor pathway and ER stress pathway) involved in the TBT-induced apoptosis?…”

Lower apoptosis rate and higher CD69 expression in neutrophils from atopic individuals

“…This might be one explanation for the low caspase 3 expression observed in neutrophils. Another plausible explanation might be that neutrophils inactivate caspase 3 [10] due to their high content of reactive oxygen metabolites [6].…”

“…In contrast, several studies indicate that the effect of steroids on neutrophil number is the opposite [7,9]. These results resulted in theories of differences in the molecular mechanism of apoptosis between eosinophils and neutrophils [7,9,10].…”