Establishment of THP-1 Monocytes with Compromised Mitochondrial Functions

Abstract

The process of inflammation is important for both normal health and in a number of diseases, such as metabolic disorders, autoimmune diseases, and neurodegenerative diseases. Mitochondria are vital for the functioning of all cells. It had been implicated as a key player in inflammatory processes, especially through reactive oxygen species as signals of various immune responses. This study aimed to establish a THP-1 cell line with compromised mitochondrial functions, using antimycin A as a Complex III inhibitor, and to investigate the role of mitochondrial stress, as monitored by the expression of Hsp60, in inflammatory processes. High concentrations of antimycin A (100 and 200 μM) were cytotoxic to THP-1 monocytes that they were rapidly killed within 48 hours of exposure. Lower concentrations of antimycin A (5, 10, 25 and 50 μM) gave growth inhibition effects to THP-1 monocytes. Pyruvate and uridine were used with an intention of rescuing the THP-1 cell growth at lower antimycin A concentrations. The THP-1 monocytes treated with antimycin A with uridine and pyruvate showed more growth compared to the ones without uridine and pyruvate supplement. Yet this difference is insignificant statistically. The expressions of Hsp60 and TNF-α at the mRNA level was monitored using reverse transcriptase polymerase chain reaction. Hsp60 expression from THP-1 monocytes only showed some minor fluctuations in different antimycin A concentrations, regardless of uridine and iii pyruvate supplement, indicating the stress mitochondrial response was unobvious. On the other hand, TNF-α expression was dramatically down-regulated in THP-1 monocytes treated with antimycin A only compared to the untreated control and ones supplemented with uridine and pyruvate. These results suggest that antimycin A may have inhibition effect towards TNF-α expression, and uridine and pyruvate could also have other functions in THP-1 monocytes apart from redox rescue compounds. Yet the mitochondrial stress response shown by Hsp60 induction still remains to be further investigated.