Pang N. Shek

Peritoneal cytokine concentrations and survival outcome in an experimental bacterial infusion model of peritonitis

Objective: To correlate the dynamics of peritoneal cytokines with systemic concentrations and survival outcome. Design: Randomized, controlled study using a recently developed rat model of peritonitis. Setting: Government research facility. Subjects: Male Sprague‐Dawley rats. Interventions: Infected animals (INF) received an intraperitoneal infusion of 6.5 × 108 colony‐forming units of Escherichia coli over 12 hrs, whereas control rats (CON) received a sterile inoculum. Peritoneal fluid and plasma samples were obtained from all rats at the end of the 12‐hr infusion period as well as from all animals that survived the 7‐day study (SURV). Measurements and Main Results: Interleukin (IL)‐1β concentration in the peritoneal fluid at 12 hrs tended to be higher in nonsurvivors (NONSURV) than in SURV. Tumor necrosis factor‐α and IL‐6 peritoneal concentrations at 12 hrs were significantly greater in NONSURV than in SURV. There were no significant differences in IL‐2 and IL‐4 peritoneal concentrations at 12 hrs between SURV and NONSURV. Although the concentrations of IL‐1β and tumor necrosis factor‐α in the peritoneal fluid of INF decreased gradually during the study, these concentrations remained significantly higher than those of CON at 7 days. In contrast, peritoneal IL‐2 concentrations remained lower in INF than in CON for most of the experiment. Peritoneal IL‐6 concentrations in INF were transiently elevated above those of CON for 12 hrs. Cytokine concentrations in the peritoneal fluid of INF were always higher than those in plasma, which remained relatively unchanged throughout the study. For most of the variables assessed, CON showed no significant changes compared with INF. Conclusions: This model of peritonitis is associated with a significant and prolonged peritoneal inflammatory response that is adversely correlated with survival outcome. Our data would suggest that to be effective, novel immunotherapies should target mainly the peritoneal compartment.

Development of in situ-forming hydrogels for hemorrhage control

We report the preparation of in situ-forming hydrogels, composed of oxidized dextran (Odex) and amine-containing polymers, for their potential use as a wound dressing to promote blood clotting. Dextran was oxidized by sodium periodate to introduce aldehyde groups to form hydrogels, upon mixing in solution with different polymers containing primary amine groups, including polyallylamine (PAA), oligochitosan and glycol chitosan. A series of experiments were conducted to identify the optimum gelation condition for the Odex-PAA system. The polymer concentration appeared to have a major effect on gelation time and the polymer weight ratio affected the resulting gel content and swelling. Other influencing factors included pH of the buffer used to dissolve each polymer, PAA molecular weight, and the type of individual material. The latter also contributed significantly to gel content and swelling. Thromboelastography was used to examine the effects of the in situ gelation on blood coagulation in vitro, where the Odex-PAA combination was found to be most pro-hemostatic, as indicated by a decrease in clotting time and an increase in clot strength. The results of this study demonstrated that in situ-forming hydrogels could promote clotting in vitro; however, further studies are required to determine if the same hydrogel formulations are effective in controlling hemorrhage in vivo.

A sustained release bacterial inoculum infusion model of intra-abdominal infection in conscious rats: bacteriology, metabolism, and histopathology.

This report describes the development of a rat peritonitis model that simulates a slow, sustained bacterial release from the gut. Septic animals (SEP) received an intraperitoneal infusion of a bacterial inoculum (6.5 x 10(8) colony forming units Escherichia coli) over 12 h, while control rats (CON) received a sterile inoculum. This model yielded a 52% mortality over 7 days in SEP, with deaths usually occurring 24-48 h after the onset of infusion. Septic rats showed greater febrile responses and body weight losses than those of CON, as well as mild hyperlactacidemia, hypoglycemia, and episodic bacteremia. Maximum bacterial counts in peritoneal fluid and several organs of SEP were observed at 36 h, with bacterial counts progressively decreasing by 7 days to levels similar to those observed at 12 h. Lung and spleen wet weights increased by 17% at 36 h and 7 days post-infection in SEP. Histological evaluation of random organ samples revealed mild to moderate morphological changes in SEP while CON showed no or minimal changes in the parameters measured during the study. This new model of chronic peritonitis in the rat reproduces many of the clinical features observed in human sepsis, and thus should prove to be a useful tool in further studies of the pathophysiology of peritonitis.

Efficacy of liposomal antibiotic therapy in a rat infusion model of Escherichia coli peritonitis

OBJECTIVE To compare the potential therapeutic effect of liposomal vs. free cefoxitin. DESIGN Randomized, controlled study, using a rat model of peritonitis. SETTING Government research facility. SUBJECTS Male Sprague-Dawley rats. INTERVENTIONS Rats were infused intraperitoneally with 6.5 x 10(8) colony forming units of Escherichia coli over 12 hrs. Animals were then randomized to receive intravenous saline, free cefoxitin, liposomal cefoxitin, or plain liposomes twice daily until they were killed. MEASUREMENTS AND MAIN RESULTS Free cefoxitin significantly reduced the number of E. coli after 24 hrs compared with saline treatment in both liver and spleen. However, liposomal cefoxitin further decreased the bacterial content by five-fold to ten-fold in these organs. Minimal bactericidal effect was observed in animals injected with plain liposomes. Although administration of liposomal cefoxitin for 7 days further reduced bacterial counts in liver and spleen, there was no apparent beneficial bactericidal effect of free cefoxitin over saline at 7 days. There was approximately a ten-fold reduction in bacterial content in the lungs after 24 hrs in all three treatments, but no further reduction was observed after 7 days. There was no difference in 7-day survival rate in animals treated with plain liposomes or saline (45% vs. 39%). Although survival tended to increase with free cefoxitin treatment (64%), this outcome was significantly improved with the use of liposomal cefoxitin (82%). CONCLUSIONS Liposomal cefoxitin enhanced bacterial killing in liver and spleen in this model of E. coli peritonitis. It also improved survival outcome relative to no treatment but not compared with free cefoxitin.

Changes in circulating lymphocyte subpopulations and mitogen-stimulated response in a rat infusion model of intra-abdominal infection

Objective To describe the alterations in circulating concentrations of immune cells as well as in in vitro mitogen-stimulated response in a recently developed rat model of intra-abdominal infection. Design Randomized, controlled study. Setting Government research facility. Subjects Male Sprague-Dawley rats. Interventions Infected animals received an intraperitoneal infusion of 6.0 × 108 colony forming units of Escherichia coli during 12 hrs, whereas control rats received a sterile inoculum. All experimental animals underwent laparotomy and peritoneal lavage at the end of the infusion period. Blood samples were obtained 12 hrs, 36 hrs, or 7 days after the onset of infusion. Splenocytes were concomitantly harvested and assayed for response to the mitogens phytohemagglutinin (PHA), concanavalin A (Con A), and lipopolysaccharides, as well as for production of interleukin (IL)-2. Measurements and Main Results Infected rats showed a marked leukopenia (−82% for 36 hrs), with leukocyte counts returning to normal at 7 days. They also developed a marked lymphocytopenia throughout the study; this was achieved through comparable reductions in circulating T and B cells. Con A responses in both groups were similar for 7 days. In contrast, splenocytes from infected animals showed reduced responses to lipopolysaccharides (−64%) and PHA (−30%) for 36 hrs compared with control splenocytes. IL-2 production from mitogen-stimulated splenocytes was suppressed in infected rats to 66% of that of control rats for 7 days. Suppressed PHA responses were not restored to control values in the presence of IL-2. For all of the parameters assessed, control animals showed either no significant changes or relatively fewer changes than infected rats. Conclusions This model of intra-abdominal infection is associated with changes in circulating concentrations of immune cells as well as with temporary functional defects in B and T cells, consistent with those often observed in patients with peritonitis. However, the role of IL-2 in limiting the adverse effects of infection in this experimental model seems to be limited. This model may be a useful tool in furthering our understanding of the pathophysiology of intra-abdominal infections and in assessing the efficacy of new therapeutic modalities.