Effect of IL-1beta on survival and energy metabolism of R28 and RGC-5 retinal neurons.
OBJECTIVE
Interleukin-(IL)1beta expression is increased in the retina during a variety of diseases involving the death of retinal neurons and contributes to neurodegenerative processes through an unknown mechanism. This study was conducted to examine the effects of IL-1beta on the metabolism and viability of RGC-5 and R28 retinal neuronal cells.


METHODS
Cellular reductive capacity was evaluated using WST-1 tetrazolium salt. Mitochondrial transmembrane potential was determined by JC-1 fluorescence. Cellular ATP levels were measured with a luciferase assay. Caspase-3/7 activation was detected with a DEVDase activity assay. Cell death and lysis was evaluated by measuring release of lactate dehydrogenase (LDH). Glycolysis was assessed by measuring glucose disappearance and lactate appearance in cell culture medium. Cellular respiration was followed polarographically.


RESULTS
IL-1beta treatment caused a pronounced decrease in cellular reductive potential. IL-1beta caused depletion of intracellular ATP, loss of mitochondrial transmembrane potential, caspase-3/7 activation, and LDH release. IL-1beta treatment increased rates of glucose utilization and lactate production. The cells were partially protected from IL-1beta toxicity by ample ambient glucose. However, glucose did not block the ability of IL-1beta to cause a decline in mitochondrial transmembrane potential or ATP depletion. IL-1beta decreased oxygen consumption of the R28 cells by nearly half, but did not lower cytochrome c oxidase activity.


CONCLUSIONS
The present results suggest that IL-1beta inhibits mitochondrial energy metabolism of these retinal neuronlike cells.
Abcouwer
Steve F
SF
Department of Surgery, Penn State Diabetic Retinopathy Center, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania 17033-0850, USA. sabcouwer@psu.edu
Shanmugam
Sumathi
S
Gomez
Paul F
PF
Shushanov
Sain
S
Barber
Alistair J
AJ
Lanoue
Kathryn F
KF
Quinn
Patrick G
PG
Kester
Mark
M
Gardner
Thomas W
TW
D000255:Adenosine Triphosphate / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology; D003576:Electron Transport Complex IV / Q000378:metabolism; D004734:Energy Metabolism; D004789:Enzyme Activation; D005947:Glucose / Q000378:metabolism; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects; D012165:Retinal Ganglion Cells / Q000187:drug effects* / Q000378:metabolism
D053583:Interleukin-1beta; D019344:Lactic Acid; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D003576:Electron Transport Complex IV; D053148:Caspase 3; D053179:Caspase 7; D005947:Glucose