Age-related differences in hepatic ischemia/reperfusion: gene activation, liver injury, and protective effect of melatonin.
BACKGROUND
Ischemia/reperfusion (I/R) causes functional and structural damage to liver cells, this being more pronounced with increasing age of the tissue. Melatonin is a pineal indole that has been shown to play an important role as a free radical scavenger and anti-inflammatory molecule.


METHODS
The age-dependent responses to I/R were compared in 2-mo-old and 14-mo-old male Wistar rats. After 35 min of hepatic ischemia followed by 36 h of reperfusion, rats were sacrificed. Sham-operated control rats underwent the same protocol without real vascular occlusion. Animals were intraperitoneally injected with 10 mg/kg melatonin 24 h before the operation, at the time of surgery, and 12 and 24 h after it. The tissues were submitted to histopathologic evaluation. The levels of ALT and AST were analyzed in plasma. The expression of TNF-α, IL-1β, IL-10, MCP-1, IFN-γ, iNOS, eNOS, Bad, Bax, Bcl2, AIF, PCNA, and NFKB1 genes were detected by RT-PCR in hepatic tissue.


RESULTS
I/R was associated with significant increases in the expression of pro-inflammatory and pro-apoptotic genes in liver. Older rats submitted to I/R were found to respond with increased liver damage as compared with young rats, with serum ALT and AST levels significantly higher than in young animals. Mature rats also showed more evident increases in expression of pro-inflammatory cytokines (IL-1β, MCP-1, and IFN-γ) as well as a decrease in the mRNA expression of IL-10 as compared with young animals. Pro-apoptotic genes (Bax, Bad, and AIF) were significantly enhanced in liver after I/R, without differences between young and mature animals. However, the expression of Bcl2 gene did not show any change. Melatonin treatment was able to lower the expression of pro-inflammatory cytokines and pro-apoptotic genes and to improve liver function, as indicated by normalization of plasma AST and ALT levels and by reduction of necrosis and microsteatosis areas.


CONCLUSIONS
Melatonin treatment was able to reduce the I/R-stimulated pro-inflammatory and pro-apoptotic genes in the rat liver. Since older animals showed a more marked increase in inflammation and in liver injury, the treatment was more effective in those subjects.
Department of Physiology, Medical School, University Complutense of Madrid, Madrid, Spain. rakireev@gmail.com
Roman A
RA
Kireev
Sara
S
Cuesta
Carolina
C
Ibarrola
Teresa
T
Bela
Enrique
E
Moreno Gonzalez
Elena
E
Vara
Jesus A F
JA
Tresguerres
D000367:Age Factors; D000818:Animals; D016753:Interleukin-10 / Q000235:genetics; D053583:Interleukin-1beta / Q000235:genetics; D008099:Liver / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology; D008297:Male; D008550:Melatonin / Q000627:therapeutic use*; D052002:NF-kappa B p50 Subunit / Q000235:genetics; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D015533:Transcriptional Activation
D053583:Interleukin-1beta; D052002:NF-kappa B p50 Subunit; D018809:Proliferating Cell Nuclear Antigen; D016753:Interleukin-10; D052247:Nitric Oxide Synthase Type II; D008550:Melatonin