Human mature red blood cells express caspase-3 and caspase-8, but are devoid of mitochondrial regulators of apoptosis

Abstract

Although proteases of the caspase family are essential mediators of apoptosis in nucleated cells, in anucleate cells their presence and potential functions are almost completely unknown. Human erythrocytes are a major cell population that does not contain a cell nucleus or other organelles. However, during senescence they undergo certain morphological alterations resembling apoptosis. In the present study, we found that mature erythrocytes contain considerable amounts of caspase-3 and -8, whereas essential components of the mitochondrial apoptotic cascade such as caspase-9, Apaf-1 and cytochrome c were missing. Strikingly, although caspases of erythrocytes were functionally active in vitro, they failed to become activated in intact erythrocytes either during prolonged storage or in response to various proapoptotic stimuli. Following an increase of cytosolic calcium, instead the cysteine protease calpain but not caspases became activated and mediated fodrin cleavage and other morphological alterations such as cell shrinkage. Our results therefore suggest that erythrocytes do not have a functional death system. In addition, because of the presence of procaspases and the absence of a cell nucleus and mitochondria erythrocytes may be an attractive system to dissect the role of certain apoptosis-regulatory pathways. Cell Death and Differentiation (2001) 8, 1197 ± 1206.

Tissue transglutaminase (TG2) facilitates phosphatidylserine exposure and calpain activity in calcium-induced death of erythrocytes

“…So we decided to reinvestigate the potential role of m-calpain in regulating PS exposure during calcium-induced death of erythrocytes. In contrast to previous suggestions based on calpain inhibitory studies, 1 we found no difference in the kinetics of PS exposure, when wild type and m-calpain-null erythrocytes were exposed to either ionomycin treatment, oxidative or osmotic stress (data not shown). Altogether these observations suggest that TG2 using its crosslinking activity influences two independent cell death processes during calcium-induced death of RBCs: it facilitates the m-calpain-dependent proteolytic cleavage by directly activating m-calpain, and in addition it accelerates the PS exposure, which appears to be independent of m-calpain.…”

“…However, recently it has been revealed that treatment of erythrocytes with the Ca 2 þ ionophore ionomycin, 1 or their exposure to oxidative or osmotic stress, 2 situations that mimic red blood cell aging, leads to cell shrinkage, cell membrane blebbing and phosphatidylserine (PS) exposure, all typical features of apoptosis in other cell types. As macrophages are equipped with receptors recognizing PS, erythrocytes exposing PS at their cell surface will be rapidly recognized, engulfed and degraded.…”

The metabolites of glutamine prevent hydroxyl radical-induced apoptosis through inhibiting mitochondria and calcium ion involved pathways in fish erythrocytes

“…After incubation at 37°C for 7 h, the erythrocytes in positive control and control were collected for measurement of the activities of caspase-8, caspase-9 and caspase-3 and the levels of ROS and cytochrome c. After washing three times with PCS, the erythrocytes in positive control were suspended in PCS and PCS containing 100 μM of Ac-IETD-fmk or Z-LEHD-fmk, 1.40 mM of Ala, Cit or Pro, 100 μM of Ac-IETD-fmk and 1.40 mM of Ala, Cit or Pro, 100 μM of Ac-IETD-fmk and Z-LEHD-fmk, or 100 μM of Ac-IETD-fmk and Z-LEHD-fmk and 1.40 mM of Ala, Cit or Pro at a 1% hematocrit, respectively. Or, a cell-free assays were performed as described by Berg et al [43] with slight modifications. Briefly, after washing with PCS, 5 Â 10 7 erythrocytes in positive control were lysed in 100 μl of lysis buffer (Beyotime, Nantong, China) and lysis buffer containing 100 μM of Z-LEHD-fmk, 2.0 μg/ml of Anti-Cyt C [44] or 2.0 μg/ml of Anti-Cyt C and 1.40 mM of Ala, Cit or Pro, respectively.…”

Oxidation of proteins: Basic principles and perspectives for blood proteomics

“…Oxidative stress is also associated with protein degradation [158], band 3 clustering [159], phosphatidylserine externalisation [160], activation of caspases [160][161][162], and down-regulation of glycophorins [163], some of these being recognised as biomarkers of senescence or "apoptosis". In particular phosphatidylserine externalisation signals macrophages to recognise and degrade the RBCs.…”