Heat preconditioning ameliorates hepatocyte viability after cold preservation and rewarming, and modulates its immunoactivity.
BACKGROUND
Heat preconditioning significantly preserved liver graft function after cold preservation in animal experimental model. The elevation of heat shock protein 70 (HSP70) was claimed to play a critical role in protecting grafts against cold preservation-induced hepatocyte apoptosis. However, little is known about whether HSP70 also plays an immunomodulatory role in cold preserved cells. This study aimed at investigating the relationship between HSP70 protein and the immunoactivity in response to lipopolysaccharide (LPS) stimulation.


RESULTS
A normal rat hepatocyte cell line was preserved with University of Wisconsin (UW) solution, Ringer's lactate solution (RL), and phosphate-buffered saline (PBS) at 4 degrees C. No significant morphological alteration was noted in UW-preserved cells after 24 h through phase-contrast microscopic observation and fluorescent viability stain. Western blotting showed a two-fold increase in the ratio of HSP70/Bax proteins in cells after 24 h of UW preservation. Heat preconditioning significantly enhanced the recovery of lactate dehydrogenase (LDH) activity in both RL- and UW-preserved cells that were stored for a period of 12 h or less. Moreover, heat preconditioning promoted HSP70 and NF-kappaB p50 nuclear translocation and suppressed the LPS-induced nuclear p50 accumulation in cells before UW preservation. Immunofluorescent stain revealed that the LPS-induced p50 protein redistribution to nuclear membrane might contribute to NF-kappaB activation, while heat preconditioning and UW cold preservation completely abrogated the p50 intranuclear redistribution. Thus NF-kappaB p50 might be responsible for the endotoxin tolerance induction.


CONCLUSIONS
These findings strongly suggest that heat preconditioning not only preserves hepatocyte viability after cold preservation and rewarming, but also ameliorates its immunoactivity.
Kao
Ying-Hsien
YH
Liver Transplantation Program, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, 123 Ta-Pei Rd., Niao-Sung, Kaohsiung 833, Taiwan.
Goto
Shigeru
S
Jawan
Bruno
B
Nakano
Toshiaki
T
Hsu
Li-Wen
LW
Lin
Yu-Chun
YC
Pan
Mei-Chun
MC
Lai
Chia-Yun
CY
Sun
Cheuk-Kuan
CK
Cheng
Yu-Fan
YF
Tai
Ming-Hong
MH
Huang
Hung-Tu
HT
Chen
Chao-Long
CL
D000818:Animals; D002470:Cell Survival; D003080:Cold Temperature; D015925:Cryopreservation; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology / Q000276:immunology / Q000502:physiology*; D006358:Hot Temperature; D020672:I-kappa B Proteins / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology; D000072000:NF-KappaB Inhibitor alpha; D009926:Organ Preservation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D017694:Rewarming; D000097782:NF-kappaB-Inducing Kinase
D018840:HSP70 Heat-Shock Proteins; D020672:I-kappa B Proteins; D007770:L-Lactate Dehydrogenase; D008070:Lipopolysaccharides; D000072000:NF-KappaB Inhibitor alpha; D017346:Protein Serine-Threonine Kinases; D000097782:NF-kappaB-Inducing Kinase; C000606206:Nfkbia protein, rat