The Role Of Kupffer Cells In Sepsis-Induced Organ Injury

Objective:This study is to investigate:1.the role of NF- κB / IκB pathway in Kupffer cell activation by in vitro stimulation with lipopolysaccharide (LPS);2. the effect of dexamethasone and heat shock response on endotoxin-induced cytokine expression by Kupffer cells;3. the effect of dexamethasone and heat shock response on endotoxemia-induced production of: NF- κB activity; TNF α , IL-6 and IL—10 mRNA levels; mRNA and protein expression of I κB α in murine liver.Method:Murine Kupffer cells were isolated with collagenase digestion. TNF α , IL-6 and IL-10 production by Kupffer cells after LPS stimulation was analyzed with ELISA. The mRNA levels of the above cytokines were assayed semiquantitatively by RT-PCR. NF- κ B activity in Kupffer cells was assayed by EMSA after LPS stimulation. I κ B α and I κ B β protein and mRNA expression were determined by Western blot analysis and RT-PCR, respectively. Pretreated with dexamethasone or sodium arsenite, Kupffer cells were tested, after LPS stimulation, for TNF α , IL-6, and IL-10 protein and mRNA levels, NF-κ B activity, and IκBα protein and mRNA levels. The mRNA stability of TNF α , IL-6, IL-10 and IκBα was tested against sodium arsenite. A bolus of sodium arsenite (6 mg/kg) was intravenously injected to produce heat shock response model. In endotoxemia mice produced by intraperitoneal injection of LPS(10mg/kg), the effect of either dexamethasone or heat shock was tested in the liver for NF κ B activity, TNF α , IL-6, and IL-...