Role of oxygen radicals generated by NADPH oxidase in apoptosis induced in human leukemia cells.

Abstract

We have used a human leukemia cell line that, after homologous recombination knockout of the gp91-phox subunit of the phagocyte respiratory-burst oxidase cytochrome b -558, mimics chronic granulomatous disease (X-CGD) to study the role of oxygen radicals in apoptosis. Camptothecin (CPT), a topoisomerase I inhibitor, induced significantly more apoptosis in PLB-985 cells than in X-CGD cells. Sensitivity to CPT was enhanced after neutrophilic differentiation, but was lost after monocytic differentiation. No difference between the two cell lines was observed after treatment with other apoptosis inducers, including etoposide, ultraviolet radiation, ionizing radiation, hydrogen peroxide, or 7-hydroxystaurosporine. After granulocytic differentiation of both cell lines, CPT still induced apoptosis, suggesting independence from replication in fully differentiated and growth-arrested cells. Pyrrolidine dithiocarbamate (an antioxidant inhibitor of NF-B) and catalase partially inhibited CPT-induced DNA fragmentation in granulocytic-differentiated PLB-985 cells, but had no effect in X-CGD cells. Flow cytometry analysis revealed that reactive oxygen intermediates were generated in CPT-treated PLB-985 cells. These data indicate that oxygen radicals generated by NADPH oxidase may contribute directly or indirectly to CPT-induced apoptosis in human leukemia and in neutrophilic-differentiated cells. ( J. Clin. Invest. 1998Invest. . 102:1961Invest. -1968

Induction of Neutrophil Apoptosis by thePseudomonas aeruginosaExotoxin Pyocyanin: A Potential Mechanism of Persistent Infection

“…ROI have been shown to accelerate neutrophil apoptosis (23,31). Production of hydrogen peroxide was measured by flow cytometry, using DCF (20). Pyocyanin treatment of neutrophils was found to induce rapid ROI generation, detectable within 5 min of pyocyanin addition (Fig.…”

“…The generation of intracellular ROI was measured using the cell-permeable molecule 2Ј,7Ј-dichloro-dihydrofluorescein diacetate (DCF; Sigma-Aldrich), which becomes fluorescent upon reaction with hydrogen peroxide, as previously described (20). Neutrophils were preincubated with DCF for 30 min before treatment with appropriate stimulus.…”

NF-κB activation by combinations of NEMO SUMOylation and ATM activation stresses in the absence of DNA damage

“…Inhibition of the ATM kinase activity was monitored by the inhibition of CHK2 phosphorylation, a downstream target of ATM (Sarkaria et al, 1998). While DSBs are the key lesions induced, the above DNA damaging agents can also cause other stress events including single-stranded nicks, protein-DNA adducts, modified bases, stalled replication forks, transcriptional perturbations and/or oxidative stress (Hsiang et al, 1985;Hsiang et al, 1989;Wu and Liu, 1997;Hiraoka et al, 1998;Strumberg et al, 2000). Thus, it is unclear whether DSBs per se are sufficient to initiate the NF-kB activation pathway.…”

Mechanisms Involved in Spontaneous and Viscum album Agglutinin-I-Induced Human Neutrophil Apoptosis: Viscum album Agglutinin-I Accelerates the Loss of Antiapoptotic Mcl-1 Expression and the Degradation of Cytoskeletal Paxillin and Vimentin Proteins Via Caspases

“…20) were cultured in RPMI 1640 supplemented with 10% FCS (Sigma-Aldrich), at a cell concentration not exceeding 1.5 ϫ 10 6 cells/ml. Neutrophils were isolated from the venous blood of healthy volunteers by dextran sedimentation followed by centrifugation over Ficoll-Hypaque (Pharmacia Biotech, Baie d'Urfie, Canada), as previously described (16,21).…”

“…We thank Dr. Yves Pommier (National Cancer Institute, Bethesda, MD) for donation of the PLB-985 and X-CGD cell lines (20) and Mary Gregory for reading the manuscript.…”