Functional distinction of rat liver natural killer cells from spleen natural killer cells under normal and acidic conditions in vitro.
BACKGROUND
The microenvironment within solid tumors has often been shown to exhibit an acidic extracellular pH. Although the morphologic and functional differences in natural killer (NK) cells of the liver and spleen have been reported previously under physiological conditions, the difference under acidic conditions is still unclear. This study was to investigate the differences in the morphological and functional characteristics between rat liver and spleen NK cells under normal and acidic conditions in vitro.


METHODS
Liver and spleen NK cells were isolated and purified from Sprague-Dawley rats by density gradient centrifugation and the Dynabeads(®) FlowComp(TM) Flexi system, and stimulated for 4 days with or without IL-2 or treated with low pH or control for different times. Morphology was examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), cell death and proliferation assays were performed by flow cytometry, IFN-gamma production was tested by ELISA, and cytotoxic activity was evaluated by lactate dehydrogenase (LDH) release assay.


RESULTS
Liver NK cells had significantly higher levels of cytotoxic activity than spleen NK cells under normal and acidic conditions, and the maximum difference was observed at pH 5.6. Further analysis revealed that the cytotoxic activity of NK cells was correlated with morphology, cell death, proliferative activity and IFN-gamma production. By TEM, liver NK cells contained a greater number of electron-dense granules per cell at pH 5.6. Moreover, a modest elevation of cell death and reduction of proliferation of liver NK cells occurred within a range of 5.6-7.2. Interestingly, an acidic extracellular pH only marginally, and not significantly, suppressed IFN-gamma production by liver NK cells.


CONCLUSIONS
The sharp morphological and functional differences shown by the two types of NK cells in vitro indicate that liver NK cells are unexpectedly resistant to pH shock.
Department of Hepatobiliary Surgery, Sun Yat-Sen University, Guangzhou, China.
Li-Hong
LH
Lv
Jian-Dong
JD
Yu
Guo-Lin
GL
Li
Tian-Zhu
TZ
Long
Wei
W
Zhang
Ya-Jin
YJ
Chen
Jun
J
Min
Yun-Le
YL
Wan
D000818:Animals; D016923:Cell Death; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D060833:Cellular Microenvironment; D002499:Centrifugation, Density Gradient; D003601:Cytotoxicity Tests, Immunologic; D003602:Cytotoxicity, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006863:Hydrogen-Ion Concentration; D007371:Interferon-gamma / Q000378:metabolism; D007376:Interleukin-2 / Q000378:metabolism; D007694:Killer Cells, Natural / Q000276:immunology* / Q000648:ultrastructure; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000276:immunology* / Q000648:ultrastructure; D008297:Male; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D051381:Rats; D017207:Rats, Sprague-Dawley; D013154:Spleen / Q000276:immunology* / Q000648:ultrastructure; D013997:Time Factors
D007376:Interleukin-2; D007371:Interferon-gamma; D007770:L-Lactate Dehydrogenase