The Role of Intestinal Flora in Endotoxin (LPS)-Induced Gut Injury and Group II Phospholipase A₂ (PLA₂-II) Activation in the Small Intestine

Abstract 256 Poster Session III, Monday, 5/3 (poster 109)

PLA₂-II is a 14kDa secretory PLA₂ which is involved in septic shock and inflammation. It is preferentially localized in the intestinal crypt cells and has bactericidal functions. PLA₂-II is upregulated by platelet activating factor (PAF), an inflammatory mediator, and LPS. We have previously shown that germ-free rats and antibiotic-treated rats (a mixture of polymyxin B, neomycin and metronidazole in drinking water for 3 days to reduce gut flora) were protected against PAF-induced shock, PLA₂-II activation and gut injury. However, they were not protected against PAF-induced hemoconcentration and leukopenia. Here, we examined the response of germ-free and antibiotic-treated rats to LPS. Anesthetized S.D. rats (germ-free, antibiotic-treated, and conventional) were injected with LPS (5mg/kg, iv), and the small bowel was removed at 2 hours for PLA₂-II assay and to assess injury. Blood pressure, hematocrit and white blood cell count were also obtained at baseline and at the end of the experiment. Each group had their own controls. We found that intestinal PLA₂-II activity increased 2 fold from baseline level after LPS in germ-free rats and conventional rats. In contrast, antibiotic-treated rats only increased 1.3 fold from baseline. This suggests that pretreatment with antibiotic mixture is protective against LPS-induced PLA₂-II activation. Neither antibiotic treatment nor germ-free environment diminished the basal level of PLA₂-II activity. We also found that LPS induced shock, leukopenia, hemoconcentration and gut injury in germ-free and antibiotic-treated rats, as well as in conventional rats when compared to their respective controls. However, antibiotic-treated rats had milder shock, leukopenia, hemoconcentration and gut injury when compared with germ-free and conventional rats. Germ-free rats had a mortality rate of 4/7, compared to 0/9 in conventional, and 0/7 in antibiotic-treated rats. This observation indicates that germ-free rats are more susceptible to LPS-induced injury. In conclusion, gut flora may play an important role in LPS-induced intestinal injury and in the regulation of gut PLA₂-II activity in vivo.

(Supported by NIH grant DK34574.)