Ryanodine receptor antagonism protects the ischemic liver and modulates TNF-alpha and IL-10.
BACKGROUND
Dantrolene is a ryanodine receptor and intracellular calcium antagonist. The ryanodine receptor (RyR) Ca(2+) release channel mobilizes Ca(2+) from internal stores to support a variety of cellular functions, including the inflammatory response after ischemia and reperfusion. The pharmacological mechanism of dantrolene is associated with the inhibition of the release of Ca(2+) from the skeletal muscle sarcoplasmic reticulum (SR). We hypothesized that dantrolene could exert a protective effect in our model of liver ischemia and reperfusion by modulating TNF-alpha and IL-10.


METHODS
Mice subjected to 90 min of partial (70 to 80%) hepatic ischemia and 3 h of reperfusion were divided into five groups (n = 6/group): sham, ischemic control, and the dantrolene 1 mg/kg group studied at three times of administration: 15 min before reperfusion (DAN-PRE), at the time of reperfusion (DAN-RP), and 15 min after reperfusion (DAN-POS). The parameters measured at 3 h of reperfusion included serum liver function tests alanine aminotransferase (ALT) and aspartate aminotransferase (AST), TNF-alpha, and IL-10 in serum and liver histology.


RESULTS
It was demonstrated that the RyR intracellular calcium antagonist dantrolene offered the most significant protection for the ischemic liver when given before reperfusion and at the time of reperfusion. AST significantly differed between the control group and the DAN-PRE and DAN-RP groups (P < 0.05). ALT showed a statistically significant decrease in the DAN-PRE treated group and a decrease, although not significant, in the DAN-RP. Histological examination demonstrated a significant decrease in vacuolization in the same both groups (P < 0.05). Necrosis was significantly diminished when dantrolene was used at the time of reperfusion; congestion decreased in the same groups but without statistical significant difference. The levels of TNF-alpha were significantly decreased in the DAN-RP group. There was a decrease in TNF-alpha in the DAN-PRE group but not statistically significant. IL-10 reflected the protection observed in necrosis and vacuolization in the histopathology with an increment at the time of reperfusion (P < 0.05). DAN-POS did not exert a protective effect in ALT, AST, liver histology, or cytokine response.


CONCLUSIONS
For the first time the ryanodine receptor antagonist dantrolene offered significant functional and structural protection of the ischemic liver when given at the time for reperfusion and partial protection when given prereperfusion. RyR inhibition approach down-regulated the expression of TNF-alpha and induced an increment of the protective cytokine IL-10 when administered at the time of reperfusion. There was no protective effect of dantrolene after reperfusion.
López-Neblina
Fernando
F
Trauma, Surgery Research, and Molecular Biology, Borgess Research Institute, Kalamazoo, Michigan 49048, USA.
Toledo-Pereyra
Luis H
LH
Toledo
Alexander H
AH
Walsh
Jon
J
D000410:Alanine Transaminase / Q000097:blood; D000818:Animals; D001219:Aspartate Aminotransferases / Q000097:blood; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D003620:Dantrolene / Q000494:pharmacology*; D016753:Interleukin-10 / Q000097:blood*; D019194:Ischemic Preconditioning / Q000379:methods*; D008099:Liver / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D015427:Reperfusion Injury / Q000276:immunology / Q000473:pathology / Q000517:prevention & control*; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000097:blood*
D002121:Calcium Channel Blockers; D019837:Ryanodine Receptor Calcium Release Channel; D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10; D001219:Aspartate Aminotransferases; D000410:Alanine Transaminase; D003620:Dantrolene; D002118:Calcium