INTRACELLULAR CALCIUM ALTERATIONS AND FREE-RADICAL FORMATION EVALUATED BY FLOW-CYTOMETRY IN ENDOTOXIN-TREATED RAT-LIVER KUPFFER AND ENDOTHELIAL-CELLS

During endotoxic shock, the liver exerts an endotoxin (lipopolysacchar ide, LPS) clearance function with the participation of both sinusoidal (mainly Kupffer and endothelial cells) and parenchymal cells. In orde r to determine the specificity and diversity of response of each liver cell type, the effect of Escherichia coli 0111:B4 endotoxin (LPS) on intracellular Ca2+ content and reactive oxygen metabolite production i n rat liver Kupffer, endothelial and parenchymal cells, was evaluated by Bow cytometry during short treatment times (from 0-2 min) with a lo w dose of LPS (10 mu g/ml). Concerning sinusoidal cells, LPS produced a significant increase of intracellular Ca2+ in both endothelial and K upffer cells. The LPS effect on Kupffer cells was higher than on endot helial cells. When intracellular reactive oxygen metabolite production was evaluated in LPS-treated sinusoidal cells, a fast and significant increase of Kupffer cells in activated state (cells with a high react ive oxygen intermediate production) was observed. However, endothelial cells did not show LPS-induced changes in their intracellular reactiv e oxygen metabolite content. All these results support a rapid activat ion of liver Kupffer cells by endotoxin consistent with the major role of this cellular type as active first line of defense during endotoxi c shock. The liver endothelial cells are also involved in the first st eps of the cell damage showing intracellular Ca2+ alterations. Liver p arenchymal cells did not show any response at these experimental condi tions (short treatment time and low LPS dose) indicating that longer t reatment times are needed for LPS binding and action in agreement with previous studies.