Mitochondrial respiratory activity and superoxide radical generation in the liver, brain and heart after chronic ethanol intake

Peritoneal macrophages were elicited in rats by using casein as a stimulus; when stimulated with phorbol 12-myristate 13-acetate (PMA) they produced O2.-. Nearly 60% of the total cytochrome b had a low Em,7.0 of -247 mV, typical of the cytochrome b component found in the NADPH-dependent O2(.-)-generating oxidase of neutrophils. The rate of O2.- generation by macrophages was 1.23 mol of O2.-/s per mol of cytochrome b. Treatment of intact macrophages with diphenyleniodonium (DPI) at 0.9 microM caused 50% inhibition of PMA-induced O2.- generation, with little effect on mitochondrial respiratory activity; KCN inhibited respiratory activity without affecting PMA-induced O2.- generation. A similar specificity of inhibition was found for di-2-thienyliodonium (50% inhibition of O2.- generation at 0.5 microM) and, at higher concentrations, for diphenyl iodonium. When macrophage suspensions were incubated with [125I]DPI followed by autoradiography of SDS/polyacrylamide-gel-electrophoresis-separated polypeptides, radioactivity was most strongly associated with a band of Mr 45,000, similar to that found in neutrophils [Cross & Jones (1986) Biochem. J. 237, 111-116]. The O2(.-)-generating oxidase of macrophages appears to have components in common with the NADPH oxidase of neutrophils, despite differences in activity. Its sensitivity to DPI suggests that selective prevention of radical generation by macrophages in vivo is possible.

Download Full-text

Clock-genes and mitochondrial respiratory activity: Evidence of a reciprocal interplay

Biochimica et Biophysica Acta (BBA) - Bioenergetics ◽

10.1016/j.bbabio.2016.03.035 ◽

2016 ◽

Vol 1857 (8) ◽

pp. 1344-1351 ◽

Cited By ~ 17

Author(s):

Rosella Scrima ◽

Olga Cela ◽

Giuseppe Merla ◽

Bartolomeo Augello ◽

Rosa Rubino ◽

...

Keyword(s):

Clock Genes ◽

Respiratory Activity ◽

Mitochondrial Respiratory Activity ◽

Mitochondrial Respiratory

Download Full-text

Detection of Mitochondrial Respiratory Dysfunction in Circulating Lymphocytes Using Resazurin

Archives of Pathology & Laboratory Medicine ◽

10.5858/2005-129-1295-domrdi ◽

2005 ◽

Vol 129 (10) ◽

pp. 1295-1298

Author(s):

Khaled K. Abu-Amero ◽

Thomas M. Bosley

Keyword(s):

Normal Control ◽

Respiratory Activity ◽

Mitochondrial Protein ◽

Mitochondrial Disorders ◽

Control Group ◽

Respiratory Dysfunction ◽

Control Subjects ◽

Mitochondrial Fractions ◽

Mitochondrial Respiratory Activity ◽

Mitochondrial Respiratory

Abstract Context.—Laboratory methods currently available for detecting mitochondrial respiratory dysfunction are labor-intensive, require large amounts of isolated mitochondrial protein, invasive (require a skeletal muscle biopsy), and usually produce conflicting results. Objective.—To develop a rapid, reliable, and noninvasive method for detecting oxidative phosphorylation activity without the need to isolate mitochondrial fractions. Design.—Lymphocytes from 6 patients with mitochondrial disorders (3 with mitochondrial myopathy and 3 with Leber hereditary optic neuropathy) and 51 normal control subjects were incubated with 6μM resazurin without and with mitochondrial inhibition by amiodarone (200μM), and the fluorescence intensity resulting from resazurin reduction was monitored spectrofluorometrically over time. Mitochondrial respiratory activity was calculated as the difference between uninhibited and inhibited measurements. Results.—Mitochondrial respiratory activity was established for 51 normal control subjects and was decreased in all 6 patients with mitochondrial syndromes. Mitochondrial respiratory activity values for patients 1 through 6 compared to the control group after 240 minutes' incubation with resazurin were 55%, 71%, 49%, 61%, 68%, and 59%, respectively (mean mitochondrial respiratory activity of patients, 13.6% or 60.5% of control mean; P < .001). Conclusion.—This resazurin-based technique proved to be a fast and reproducible method for quantifying mitochondrial activity and identifying respiratory functional defects in patients with mitochondrial disorders.

Download Full-text