Young and aging human embryonic lung diploid cell line WI-38: A comparison of mitochondrial content, relative amount of mtDNA, and mitochondrial functions / 第二军医大学学报

ABSTRACT

Objective: To compare the mitochondrial content, the relative amount of mtDNA, and mitochondrial functions between the young and aging WI-38 cells, so as to investigate the correlation between mitochondrial and aging. Methods: Human embryonic lung diploid cell line WI-38 was cultured and its viability was assayed by MTT assay; the content of mitochondrial protein was determined using BCA-100 Protein Quantitative Analysis Kit after mitochondria were fractionated by differential centrifugation; mtDNA relative content was measured by a competitive polymerase chain reaction (PCR) method; mitochondrial membrane potential was measured by flow cytometry; and NADH oxidase activity was measured by spectrophotometry. Results: Compared with the young cells, the aging cells had a longer time to form a monolayer, an obviously decreased cell viability and mitochondrial membrane potential (by 50%), and a decreased NADH oxidase activity, with the maximal reaction speed declining from 66.73 nmol/(mg protein · min) to 36. 01 nmol/(mg protein · min). Mitochondrial content in the aging cells([0.78 ± 0.02] mg/ml) was higher than that in the young cells([0.56 ± 0.03] mg/ml). Using 18S rDNA of nuclear as an internal reference, the relative amount of mtDNA in the aging cells (1.557 ± 0.072) was found to be obviously higher than that in the young cells (1.292 ± 0.068). Conclusion: The increase of mitochondrial contents and mtDNA relative amounts in aging cells may be one of the compensatory mechanisms for decreased mitochondrial function, which may provide an evidence for studying the correlation between mitochondrial and aging.