Restitution of superoxide generation in autosomal cytochrome-negative chronic granulomatous disease (A22(0) CGD)-derived B lymphocyte cell lines by transfection with p22phax cDNA

The respiratory burst oxidase of phagocytes and B lymphocytes is a multicomponent enzyme that catalyzes the one-electron reduction of oxygen by NADPH. It is responsible for the O2-production that occurs when these cells are exposed to phorbol 12-myristate 13-acetate or physiologic stimuli, such as phagocytosis in phagocytes or cross- linking of surface immunoglobulin in B lymphocytes. The activity of this enzyme is greatly diminished or absent in patients with chronic granulomatous disease (CGD), an inherited disorder characterized by a severe defect in host defense against bacteria and fungi. In every CGD patient studied so far, an abnormality has been found in a gene encoding one of the four components of the respiratory burst oxidase: the membrane proteins p22phox or gp91phox which together form the cytochrome b558 protein, or the cytosolic proteins p47phox or p67phox. Autosomal recessive cytochrome-negative CGD (A22(0) CGD) is associated with mutations in the gene coding for p22phox. We report here that the capacity for O2- production and cytochrome b558 protein expression were restored to Epstein-Barr virus-transformed B lymphocytes from two A22(0) CGD patients by transfection with an expression plasmid containing a p22phox cDNA. No detectable O2- was generated by untransfected p22phox-deficient lymphocytes. The genetic reconstitution of the respiratory burst in A22(0) CGD B lymphocytes by transfer of the wild-type p22phox cDNA represents a further step towards somatic gene therapy for this subgroup of A22(0) CGD. This system will also be useful for expression of genetically engineered mutant p22phox proteins in intact cells, facilitating the structure-function analysis of cytochrome b558

Reactive oxygen species in phagocytic leukocytes

Phagocytic leukocytes consume oxygen and generate reactive oxygen species in response to appropriate stimuli. The phagocyte NADPH oxidase, a multiprotein complex, existing in the dissociated state in resting cells becomes assembled into the functional oxidase complex upon stimulation and then generates superoxide anions. Biochemical aspects of the NADPH oxidase are briefly discussed in this review; however, the major focus relates to the contributions of various modes of microscopy to our understanding of the NADPH oxidase and the cell biology of phagocytic leukocytes

Superoxide production by phagocytic leukocytes: the scientific legacy of Bernard Babior

It was 32 years ago that Bernard Babior, Ruby Kipnes, and I submitted a paper to the JCI reporting that polymorphonuclear leukocytes produce superoxide (O(2)(–)) during phagocytosis and that this highly reactive oxygen radical might function as a microbicidal agent. The story of how our lab came to this discovery is one of a special relationship between a student and his brilliant mentor

Biological Defense Mechanisms. THE PRODUCTION BY LEUKOCYTES OF SUPEROXIDE, A POTENTIAL BACTERICIDAL AGENT

As a highly reactive substance produced in biological systems by the one-electron reduction of oxygen, superoxide (O(2)(-)) seemed a likely candidate as a bactericidal agent in leukocytes. The reduction of cytochrome c, a process in which O(2)(-) may serve as an electron donor, was found to occur when the cytochrome was incubated with leukocytes. O(2)(-) was identified as the agent responsible for the leukocyte-mediated reduction of cytochrome c by the demonstration that the reaction was abolished by superoxide dismutase, an enzyme that destroys O(2)(-), but not by boiled dismutase, albumin, or catalase. Leukocyte O(2)(-) production doubled in the presence of latex particles. The average rate of formation of O(2)(-) in the presence of these particles was 1.03 nmol/10(7) cells per 15 min. This rate, however, is only a lower limit of the true rate of O(2)(-) production, since any O(2)(-) which reacted with constituents other than cytochrome c would have gone undetected. Thus. O(2)(-) is made by leukocytes under circumstances which suggest that it may be involved in bacterial killing