Dietary iron and recovery from peritonitis in guinea pigs.

Ninety female Hartley guinea pigs underwent gastrostomy placement. One week later they underwent implantation of an osmotic pump, which allowed constant delivery of bacteria into the peritoneal cavity. Three days after pump implantation the animals were begun on enteral diets differing only in iron content (the None [no Fe], Low [1 X RDA], and High [10 X RDA] groups). When survivors were killed no differences were found in body, carcass, or organ weights among the three groups. Serum Fe and percent Fe-binding sites occupied were significantly lower in the None group although total Fe-binding capacity was similar. Mortality was not statistically different (p = 0.29): 18/32 in the None group (56%), 14/24 in the Low group (58%), and 25/34 in the High group (73%). We conclude that although deprivation of dietary sources of Fe does affect available circulating Fe, diet-induced hypoferremia does not alter mortality rates from bacterial peritonitis in the guinea pig.

Immunomodulators in the treatment of peritonitis in burned and malnourished animals.

Deficiencies in the immune system that lead to increased morbidity and mortality from infectious complications have been well documented in patients suffering from trauma, malnutrition, sepsis, and thermal injuries. We investigated the potential benefit of immune stimulation for preventing infection in such conditions in an animal model by evaluating three drugs: Corynebacterium parvum, thymopentin (TP-5), and CP-46,665. One-hundred eighty female guinea pigs were rendered immunodeficient by first inflicting a 30% total body surface burn and then placing the animals on diets with calories inadequate to maintain body weight. One half of the animals were then given one of the three immunomodulators on the first, third, and fifth days after burn injury, to try to reverse immunodeficiency. The remaining animals received saline solution injections. Animal responses were evaluated by inserting a clot containing Escherichia coli and Bacillus fragilis into their peritoneal cavity 6 days after burn injury. The animals were followed for 21 days after burn injury. Autopsies on those that died revealed peritonitis and/or pneumonia; autopsies on these that survived showed no pneumonia and there was consistent resolution of peritonitis. TP-5 and CP-46,665, but not C. parvum, significantly improved survival rates and mean survival time in those animals receiving 100 kcal/kg/day. TP-5 and CP-46,665 may be of benefit to the severely stressed, malnourished surgical patient who is at risk of bacterial infection.

Mechanisms of action of two new immunomodulators.

Despite antibiotics, infection remains a significant problem in surgical patients. The reasons are multiple, and include acquired immunologic deficiencies that are seen in malnutrition, sepsis, trauma, and burns. Two immunomodulators, thymopentin (TP-5) and CP-46,665, have been shown to improve survival in infectious animal models of such deficiencies. We investigated the mechanism of action in guinea pigs subjected to a burn of 30% of the total body surface area. These animals received 0.3 mg/kg of thymopentin, 0.3 mg/kg of CP-46,665, or saline solution. Neutrophils, macrophages, and serum samples were obtained from the animals and tested for their ability to phagocytose and kill Pseudomonas aeruginosa. The serum was tested for its ability to opsonize Escherichia coli. Thymopentin was found to improve neutrophil function on postburn days 2 and 4 and to improve macrophage function on postburn day 4. CP-46,665 was found to improve both macrophage function and opsonization on postburn day 2.

Metabolic and immune effects of dietary arginine supplementation after burn.

The effect of supplemental dietary arginine on metabolism and immunity was studied in 36 burned guinea pigs (30% of total body surface area) with previously placed catheter gastrostomies. The animals were randomized into four groups. After an initial three-day adaptation period, all groups received continuous isonitrogenous, isocaloric (175 kcal [735 kJ]/kg/d), and isovolemic intragastric tube feedings until postburn day (PBD) 14. Groups A, B, C, and D received 0%, 1%, 2%, and 4%, respectively, of total energy intake as arginine given in the form of crystalline arginine hydrochloride with 22%, 21%, 20%, and 18%, respectively, of total energy as whey protein. The average body weight after burn decreased equally in all groups. Resting metabolic expenditure on PBD 6 was higher in groups B (151% +/- 6% of preburn) and C (156% +/- 7%) than in groups A (131% +/- 4%) and D (136% +/- 3%). Ear-thickness response to dinitrofluorobenzene challenge on PBD 12 showed the best response in group C. The mortality rates of groups A, B, C, and D were 56%, 29%, 22%, and 56%, respectively. This study suggests that oral dietary arginine supplementation up to 2% of energy intake may be beneficial after burn injury.

Arginine supplementation and its effect on established peritonitis in guinea pigs.

To evaluate the efficacy of supplemental arginine with nutritional support in the presence of sepsis, eighty-eight gastrostomized female Hartley guinea pigs were implanted with osmotic pumps effusing an Eschericia coli/Staphylococcus aureus mixture. Animals were randomized and infused for two weeks with isocaloric and isovolumetric diets containing 0%, 2%, 4%, or 6% supplemental arginine as arginine hydrochloride. Survival was 12/22 (54%) in 0%, 9/22 (41%) in 2% and 4%, and 2/22 (9%) in 6%. Analysis by chi-square test of independence was significant (p = 0.0141) with 6% survival lower than the others. Median survival was 11 days in 0%, 8 days in 2% and 6%, and 9 days in 4%. Median survival was longer in 0% than in 2% or 6% (Kruskal-Wallis ANOVA: p = 0.02). Nitrogen balance was significantly lower in 6% compared to 0% on days 2 through 10, and lower than 2% and 4% on days 6 and 9. Nitrogen balance was higher in 0% than in 2% on days 4, 6, 10, and 13. Serum albumin and C3 were lower in all experimental groups than normal controls (ANOVA: p = 0.01). Comparison of liver, spleen, adrenals, gastrocnemius, and carcass weights, cell-mediated immunity as determined by contact sensitivity to DNFB, and transferrin showed no significant differences. There was a positive dose-response effect seen amongst the experimental groups for the amino acids arginine, ornithine, and citrulline in relation to the amount of supplemental arginine. This study suggests that dietary arginine supplementation does not enhance survival in a guinea pig model of established peritonitis.

The prolongation of skin allograft survival by topical use of cyclosporine A.

The absorption of CsA applied topically on normal or grafted skin was studied. When care was taken to prevent the rat from ingesting the CsA off their backs, the CsA blood levels was about 100 ng/mL or less, a suboptimal level to prevent rejection. The mean survival time of BUF grafts topically treated with 12 mg CsA/d was still significantly prolonged from 9.8 +/- 0.4 to 12.0 +/- 0.3 days. In bilateral grafting experiments where one graft was treated with CsA and one was not, the treated graft survived longer. Both grafts survived significantly longer than control animals with single grafts. These findings indicate that there is both a systemic and local immunosuppressive effect of topical CsA.

An evaluation of cyclophosphamide as an immunomodulator in multiple septic animal models.

Cyclophosphamide has shown promise as an immunomodulator in early animal trials involving mice and guinea pigs. To test the effectiveness of this agent in a rat model, 268 adult Lewis rats were subjected to a 30% total body surface area burn and were challenged with Pseudomonas aeruginosa 1244 administered intraperitoneally on postburn day 1 or 4, or painted on the burn wounds on postburn day 1. The animals were then followed up to determine mean survival times and survival rate percentages. Cyclophosphamide was administered to the animals on postburn days 0, 2, and 4 at a dose of either 0.5, 1.0, 5.0, or 15.0 mg/kg. A final group received saline. None of the doses significantly prolonged survival in any of the septic models, and the 15 mg/kg dose significantly shortened it in all three models. Thus, the beneficial immunomodulatory effects previously seen in mouse and guinea pig models were not seen in the rat model. Further animal investigations would appear to be indicated before human studies are undertaken.

A comparison of Biobrane vs. homograft for coverage of contaminated burn wounds.

An evaluation was made comparing homograft to Biobrane for the coverage of excised burn wounds using a rat model which had been contaminated with 1 x 10(5) Pseudomonas aeruginosa 1244. The homograft and Biobrane were then covered with either silver sulphadiazine cream, a 5 per cent mafenide acetate solution, or no topical antibacterial agent. Five days later the wounds were examined to determine the percentage of graft take. The homograft was found to have a superior graft take in this model. It is therefore recommended that a surgeon using Biobrane for the first time ensures the wound is at least as free of bacterial contamination as he would if he were using homograft.

Alterations in host defense associated with inhalation anesthesia and blood transfusion.

Anesthesia and blood transfusions have been demonstrated to impair immune function. The present study evaluated whether these impairments led to increased susceptibility to infectious complications in two animal models. Both transfusions and anesthesia (methoxyflurane) were found to increase susceptibility to peritoneal Escherichia coli infections. This susceptibility increased with time after the transfusions but decreased with time after anesthesia. Neither transfusions nor anesthesia altered susceptibility to an intravenous Escherichia coli challenge in rats.

A new model for studying nutrition in peritonitis. The adverse effect of overfeeding.

In guinea pigs fed ad libitum, controlled intraperitoneal infusion of bacteria by an implanted 7-day osmotic pump resulted in peritonitis or abscess formation with a 50% survival 14-18 days after pump implantation. Administration of 125 kcal/kg/day of a diet found to be optimal for burned guinea pigs by continuous pump controlled feedings via a previously placed gastrostomy was well-tolerated, with a 62.5% mortality by Day 17. Administration of only 100 kcal/kg/day caused weight loss of approximately 17% after 16 days, but fewer animals died (42.8%, p = NS). Feeding either 150 kcal/kg/day or 175 kcal/kg/day caused death in all 25 animals (p less than 0.001) and their survival time was slightly shortened (p = NS) when compared with animals receiving 100 or 125 kcal/kg/day. This is the first animal model of peritonitis that permits incisive dissection of the relative influences of dietary composition on outcome, because survival can be extended to 2 weeks or more in the presence of continuing sepsis.

Low protein diets improve survival from peritonitis in guinea pigs.

Enteral diets with different protein content were tested to determine their effect on outcome in a model of protracted bacterial peritonitis. Hartley guinea pigs were provided with gastrostomies, and 1 week later, osmotic pumps were implanted into the peritoneal cavity to allow for continuous release of live bacteria over the course of 1 week. Three days after pump implantation, the animals began receiving isocaloric enteral diets that contained 5%, 10%, 15%, or 20% of total calories as protein. After 2 weeks of observation, the survivors were killed. All animals lost weight during the 2-weeks period, but there was no difference in weight lost. Nitrogen balance correlated with dietary protein. The mortality rate was significantly higher in the groups that received 15% and 20% of total calories compared with the group that received 5% (p less than 0.05). Although dietary protein in the 5% group was insufficient for meeting the nutritional needs of the animal, survival was best in this group. Possible explanations are that protein restriction in this model may either augment host defence or impair bacterial virulence.

Combined use of topical cyclosporin A and silver sulphadiazine on allografts infected with Pseudomonas in burned rats.

In this study, we investigated the effects of a combination of topical Cyclosporin A (CyA) and silver sulphadiazine on infected and normal skin allografts in burned and unburned rats. Buffalo (Buf) allografts on unburned Lewis (Lew) recipients survived 16.8 +/- 0.9 days after a 10-day course of topical CyA and silver sulphadiazine, as compared to 7.4 +/- 1.1 days for untreated allografts and 18.6 +/- 0.9 days for those receiving CyA only for 10 days. Allografts infected with Pseudomonas aeruginosa and placed on burned recipients survived significantly longer (13.7 +/- 1.1 days) when treated with silver sulphadiazine and topical CyA for 7 days compared to a similar group of animals treated with silver sulphadiazine alone (8.4 +/- 1.0 days). The mortality rate for burned, infected and allografted animals receiving no medication or topical CyA was 50 per cent compared with zero in animals treated with silver sulphadiazine only or combined silver sulphadiazine and topical CyA.

Effect of blood transfusion and anesthesia on resistance to bacterial peritonitis.

Patients who undergo gastrointestinal operations and require prolonged anesthesia or blood transfusions have been reported to have a higher incidence of infectious complications. A rat peritonitis model was used to determine if the increased rate of infection was due to the severity of their underlying disease process or to possible immunosuppressive effects of transfusions and anesthesia. Four hundred adult Lewis rats were divided into four groups. Each group received either 0.5 ml of allogenic blood, 0.5 ml of syngeneic blood, metaphane anesthesia, or 1.5 ml of saline. They were challenged with either 1 X 10(8) or 1 X 10(7) Escherichia coli on the day of transfusion or 4 days after transfusion. Survival rates and mean survival times were determined. Syngeneic transfusions were found not to significantly impair survival. Anesthesia administration resulted in a moderate impairment in survival. Allogeneic blood transfusions caused the most severe impairment with a greater than 50% decrease in survival rates compared to controls in three of the four groups tested. Blood transfusions would thus appear to impair resistance to bacterial infections to an even greater degree than anesthesia. Unnecessary transfusions should therefore be avoided.

Effect of blood transfusions on macrophage-lymphocyte interaction in an animal model.

Blood transfusions have been reported over the last 2 decades to decrease allograft rejection, to increase the rate of tumor growth, and to increase susceptibility to infectious complications. The effect of transfusions on macrophages, specifically on their regulation of lymphocyte proliferation, was investigated. Both macrophages and their supernatants obtained from transfused rats impaired lymphocyte blastogenesis to a greater degree than those from nontransfused rats. This effect was greatest when the lymphocytes were subjected to mitogen stimulation. The immunosuppression was seen with macrophages from both allogeneically and syngeneically transfused rats. Blood transfusions exert their immunosuppressive effect at least in part by increasing macrophage suppression of lymphocyte response to stimuli.

Synthetic immunomodulators for prevention of fatal infections in a burned guinea pig model.

Individuals who have suffered severe trauma, such as burns, have a high incidence of infection associated with impaired host resistance. Nonspecific stimulators of host defense mechanisms, i.e., immunomodulators, may be of benefit in such situations. A small animal model (guinea pigs) was developed to study the efficacy of immunomodulators in burns. Anesthetized animals received a 20% total body surface area, full-thickness, scald burn. There was no mortality associated with this injury, but these animals were highly susceptible to challenge with Pseudomonas aeruginosa strain 1244 by direct injection into the burn wound within 24 hours of injury. This susceptibility persisted about 7 days. The standard model adopted was to injure animals, then challenge with 1 median lethal dose (LD50) of P. aeruginosa 96 hours after injury. Using this model, six synthetic immunomodulators were tested: CP-20,961, CP-46,665, muramyl dipeptide, thymopoietin pentapeptide (TP-5), levamisole, and lithium. Drug administration began 24 hours after injury and ended prior to challenge with P. aeruginosa at 96 hours. CP-20,961, muramyl dipeptide, levamisole, and lithium all had no beneficial effect on survival. A single dosage (0.3 mg/kg, I.V.) of CP-46,665, administered 24 hours postinjury, increased the survival rate from 50% to 85% and mean survival time (MST) from 8.2 days to 12.4 days. TP-5, given in four doses (0.1 mg/kg, I.V. each) every 24 hours, increased the survival rate from 40% to 80% and MST from 6.9 days to 11.6 days. These data show that immunomodulators could be of benefit in burns, but also that not all agents are effective in this particular situation.