Recruitment of mitochondrial uncoupling protein UCP2 after lipopolysaccharide induction.
Rat liver mitochondria contain a negligible amount of mitochondrial uncoupling protein UCP2 as indicated by 3H-GTP binding. UCP2 recruitment in hepatocytes during infection may serve to decrease mitochondrial production of reactive oxygen species (ROS), and this, in turn, would counterbalance the increased oxidative stress. To characterize in detail UCP2 recruitment in hepatocytes, we studied rats pretreated with lipopolysaccharide (LPS) or hepatocytes isolated from them, as an in vitro model for the systemic response to bacterial infection. LPS injection resulted in 3.3- or 3-fold increase of UCP2 mRNA in rat liver and hepatocytes, respectively, as detected by real-time RT-PCR on a LightCycler. A concomitant increase in UCP2 protein content was indicated either by Western blots or was quantified by up to three-fold increase in the number of 3H-GTP binding sites in mitochondria of LPS-stimulated rats. Moreover, H2O2 production was increased by GDP only in mitochondria of LPS-stimulated rats with or without fatty acids and carboxyatractyloside. When monitored by JC1 fluorescent probe in situ mitochondria of hepatocytes from LPS-stimulated rats exhibited lower membrane potential than mitochondria of unstimulated rats. We have demonstrated that the lower membrane potential does not result from apoptosis initiation. However, due to a small extent of potential decrease upon UCP2 recruitment, justified also by theoretical calculations, we conclude that the recruited UCP2 causes only a weak uncoupling which is able to decrease mitochondrial ROS production but not produce enough heat for thermogenesis participating in a febrile response.
Růzicka
Michal
M
Department of Membrane Transport Biophysics, No. 75 Institute of Physiology, Academy of Sciences of the Czech Republic, Vídenská 1083, 14220 Prague 4, Czech Republic.
Skobisová
Eva
E
Dlasková
Andrea
A
Santorová
Jitka
J
Smolková
Katarína
K
Spacek
Tomás
T
Zácková
Markéta
M
Modrianský
Martin
M
Jezek
Petr
P
D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D007473:Ion Channels; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism*; D051381:Rats; D020318:Rats, Long-Evans; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071246:Uncoupling Protein 2
D017931:DNA Primers; D007473:Ion Channels; D008070:Lipopolysaccharides; D026901:Membrane Transport Proteins; D024101:Mitochondrial Proteins; C000604391:Ucp2 protein, rat; D000071246:Uncoupling Protein 2