Related Experiment Videos
Inhibition of target cell mitochondrial electron transfer by tumor necrosis factor
J R Lancaster1, S M Laster, L R Gooding
1Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30322.
Abstract:
Using digitonin permeabilization to assay mitochondrial electron transfer, we have found that respiratory activity (succinoxidase and cytochrome oxidase) in three mouse fibroblast lines is completely eliminated by incubation with human recombinant tumor necrosis factor-alpha (hrTNF). As with cytotoxicity, hrTNF-induced mitochondrial dysfunction occurs in resistant cells upon inhibition of protein synthesis, whereas sensitive cells exhibit spontaneous respiratory inhibition. In C3HA cells, inhibition is detectable 1.5-2 h after hrTNF addition, preceding cell lysis by at least 5 h (as measured by dye exclusion), and is approximately coincidental with morphological changes we have previously reported for this cell line. LM cells also exhibit inhibition of electron transfer, coincidental with morphological changes. These results suggest that bioenergetic dysfunction may be involved in the cytotoxic mechanism of TNF.
Insights
Human recombinant tumor necrosis factor-alpha (hrTNF) completely eliminates mitochondrial respiratory activity in mouse fibroblast cells. This bioenergetic dysfunction, observed in sensitive and resistant cells, suggests a role in TNF-induced cytotoxicity.
Area of Science:
- Cell Biology
- Biochemistry
- Immunology
Background:
- Tumor necrosis factor-alpha (TNF) is a cytokine involved in inflammation and immunity.
- The precise mechanisms underlying TNF-induced cytotoxicity are not fully understood.
- Mitochondrial function plays a critical role in cellular health and survival.
Purpose of the Study:
- To investigate the effect of human recombinant tumor necrosis factor-alpha (hrTNF) on mitochondrial respiratory activity in mouse fibroblast cells.
- To determine if mitochondrial dysfunction is an early event in TNF-induced cytotoxicity.
Main Methods:
- Digitonin permeabilization was used to assay mitochondrial electron transfer.
- Respiratory activity (succinoxidase and cytochrome oxidase) was measured in three mouse fibroblast lines.
- The role of protein synthesis inhibition in hrTNF-induced mitochondrial dysfunction was examined.
Main Results:
- Incubation with hrTNF completely eliminated respiratory activity in all three fibroblast lines.
- Sensitive cells showed spontaneous respiratory inhibition, while resistant cells required protein synthesis inhibition.
- Mitochondrial dysfunction was detected 1.5-2 hours after hrTNF addition, preceding cell lysis.
- Inhibition of electron transfer coincided with morphological changes in LM cells.
Conclusions:
- Mitochondrial dysfunction, specifically the elimination of respiratory activity, is a key early event in hrTNF-induced cytotoxicity.
- Bioenergetic impairment may be a critical component of the cytotoxic mechanism of TNF.
- These findings provide insights into the cellular pathways affected by TNF.