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Time-dependent and dose-dependent cellular and functional alterations in rat jejunal muscularis in response to lipopolysaccharide.
MK Eskandari, JC Kalff, KKW Lee, TR Billiar, AJ Bauer.
Departments of Surgery and Medicine/Gastroenterology, University of Pittsburgh Medical Center, Pittsburgh, PA.

Gastrointestinal (GI) dysfunction resulting in stasis, bacterial overgrowth, gut barrier failure, and bacterial translocation are known to occur during sepsis and multisystem organ failure. Currently, the mechanisms involved in the initiation of GI dysmotility during systemic infections has not been elucidated. The aim of this study was to identify both cellular and functional changes in rat jejunal muscularis after an intraperitoneal injection of lipopolysaccharide (LPS: 0, 10, 15, and 20mg/kg). Observations were made over 24 hours at various timepoints (1, 3, 6, 12, and 24 hours). Cellular infiltration of the muscularis was assessed by histology and immunohistochemistry (cell counts at 200X mag; *p<0.05). LPS (15mg/kg) caused a rapid activation of resident muscularis macrophages evident only one hour after injection (LFA1: control=2.45±0.55 vs LPS=49.60±2.03*). This was followed by a moderate patchy infiltration of neutrophils (control=0.70±0.16 vs LPS=4.10±1.31*) and mast cells (control=0.55±0.13 vs LPS=4.20±1.17*) occurring at 3 hours. A more gradual infiltration of the muscularis by monocytes (ED1: control=2.30±0.49 vs LPS=21.40±2.10*) was demonstrated, peaking at 12 hours after LPS. The predominant cell type throughout the 24 hour time period was the CD14+ resident macrophage. Contractile force was then assessed using mucosa-free strips of jejunal muscularis in a standard organ bath (n=4). LPS treatment resulted in a profound dose-dependent and time-dependent reduction in circular muscularis activity in the presence of the agonist bethanechol (0-300µM) (control=0.61±0.13; LPS 10mg/kg=0.50±0.12; LPS 15mg/kg=0.21±0.07*; LPS 20mg/kg=0.06±0.01*, normalized as gm/mm²}/min at 30µM bethanechol 24 hours after LPS). The time-dependent effect became statistically significant as early as 6 hours after LPS (15mg/kg). In contrast, these functional changes were absent in the longitudinal muscularis. This is the first study to demonstrate both cellular and subsequent functional hanges in small bowel motility in reponse to endotoxemia. The data support the theory that activated resident muscularis macrophages may play an integral role in the modulation of intestinal motility.