Vivaria housing conditions expose sex differences in brain oxidation, microglial activation, and immune system states in aged hAPOE4 mice.

Apolipoprotein E ε4 allele (APOE4) is the predominant genetic risk factor for late-onset Alzheimer's disease (AD). APOE4 mouse models have provided advances in the understanding of disease pathogenesis, but unaccounted variables like rodent housing status may hinder translational outcomes. Non-sterile aspects like food and bedding can be major sources of changes in rodent microflora. Alterations in intestinal microbial ecology can cause mucosal barrier impairment and increase pro-inflammatory signals. The present study examined the role of sterile and non-sterile food and housing on redox indicators and the immune status of humanized-APOE4 knock-in mice (hAPOe4). hAPOE4 mice were housed under sterile conditions until 22 months of age, followed by the transfer of a cohort of mice to non-sterile housing for 2 months. At 24 months of age, the redox/immunologic status was evaluated by flow cytometry/ELISA. hAPOE4 females housed under non-sterile conditions exhibited: (1) higher neuronal and microglial oxygen radical production and (2) lower CD68+ microglia (brain) and CD8+ T cells (periphery) compared to sterile-housed mice. In contrast, hAPOE4 males in non-sterile housing exhibited: (1) higher MHCII+ microglia and CD11b+CD4+ T cells (brain) and (2) higher CD11b+CD4+ T cells and levels of lipopolysaccharide-binding protein and inflammatory cytokines in the periphery relative to sterile-housed mice. This study demonstrated that sterile vs. non-sterile housing conditions are associated with the activation of redox and immune responses in the brain and periphery in a sex-dependent manner. Therefore, housing status may contribute to variable outcomes in both the brain and periphery.

LP2, a stable lanthipeptide derived from cAng-(1-7), exerts myeloprotective action in mice.

OBJECTIVES: Linear unstable angiotensins stimulate hematopoiesis. Here we address: (1) Is cyclic angiotensin-(1-7) myeloprotective in mice? (2) Is cyclic angiotensin-(1-7) stable in rat? (3) Does LP2, a cyclic angiotensin-(1-7) with an N-terminal d-lysine, exert myeloprotective action in tumor-bearing mice? MATERIALS AND METHODS: Cyclic angiotensin-(1-7)'s capacity to restore levels of blood platelets and white blood cells was studied in gemcitabine-treated mice. The stability of cyclic angiotensin-(1-7) in rat was measured in blood samples taken after injection or infusion. The capacity of LP2 to restore total bone marrow cell levels in mice after treatment with 5-fluoruracil was measured. In addition, the capacity of LP2 to counter anemia in tumor-bearing mice treated with erlotinib was measured. RESULTS: Cyclic angiotensin-(1-7) dose-dependently restored blood platelet levels in gemcitabine-treated mice, whereas its capacity to restore levels of white blood cells was less. In vivo aminoterminal breakdown of cyclic angiotensin-(1-7) yielded cyclic angiotensin-(2-7) and cyclic angiotensin-(3-7). LP2 significantly (p < .0001 at 100 µg/kg/day) restored bone marrow cell counts in mice after treatment with 5-fluoruracil. LP2 also significantly (p < .05) countered anemia in tumor-bearing mice treated with erlotinib. CONCLUSIONS: LP2 exerts myeloprotective action with perspectives for continuation of its clinical development.

Reduced disease severity following therapeutic treatment with angiotensin 1-7 in a mouse model of multiple sclerosis.

Multiple Sclerosis (MS) is a chronic disease of the central nervous system (CNS) characterized by autoimmune and neurodegenerative pathologies for which there is no cure and no defined etiology. Although several, modestly effective, disease modifying drugs are available to treat MS, there are presently no treatments that offer neuroprotection and prevent clinical progression. Therapies are needed that control immune homeostasis, prevent disease progression, and stimulate regeneration in the CNS. Components of the renin-angiotensin-system (RAS) have recently been identified as chemical mediators in the CNS and in neurological disease. Here we show the beneficial effect of therapeutic treatment with the Mas receptor agonist and metabolite of the protective arm of RAS, angiotensin 1-7 (A(1-7)), in the experimental autoimmune encephalomyelitis (EAE) animal model of MS. Therapeutic treatment with A(1-7) caused a dose-dependent reduction both in clinical disease severity and progression, and was dependent on Mas receptor activation. Further analysis of the most optimal dose of A(1-7) treatment revealed that the reductions in clinical disease course were associated with decreased immune infiltration and demyelination, axonal loss and oxidative stress in the spinal cord. In addition A(1-7) treatment was also associated with increases in circulating alternatively activated monocytes/macrophages.

Angiotensin-(1-7) administration benefits cardiac, renal and progenitor cell function in db/db mice.

BACKGROUND AND PURPOSE: Diabetic patients are at an increased risk of cardiovascular disease, in part due to inflammation and oxidative stress. These two pathological mechanisms also affect other organs and cells including the kidneys and progenitor cells. Angiotensin-(1-7) [Ang-(1-7)] has previously been shown to counterbalance pathological effects of angiotensin II, including inflammation and oxidative stress. The aim of this study was to investigate the effects of short-term (2 weeks) Ang-(1-7) treatment on cardiovascular and renal function in a mouse model of type 2 diabetes (db/db). EXPERIMENTAL APPROACH: Eight- to nine-week-old db/db mice were administered either vehicle, Ang-(1-7) alone, or Ang-(1-7) combined with an inhibitor (losartan, PD123319, A-779, L-NAME or icatibant) daily for 14 days. KEY RESULTS: An improvement in physiological heart function was observed in Ang-(1-7)-treated mice. Ang-(1-7) also reduced cardiomyocyte hypertrophy, fibrosis and inflammatory cell infiltration of the heart tissue and increased blood vessel number. These changes were blocked by antagonists of the MAS1, AT2 and bradykinin receptors and inhibition of NO formation. Treatment with Ang-(1-7) reduced glomerular damage and oxidative stress in kidney tissue. Bone marrow and circulating endothelial progenitors, as well as bone marrow mesenchymal stem cells, were increased in mice treated with Ang-(1-7). CONCLUSIONS AND IMPLICATIONS: Short-term Ang-(1-7) treatment of young db/db mice improved heart function and reduced kidney damage. Treatment also improved bone marrow and circulating levels of endothelial and mesenchymal stem cells. All of this may contribute to improved cardiovascular and renal function.

Localization of follicle regulatory protein in the porcine ovary.

The granulosa cell secretes a protein (follicle regulatory protein: FRP) that affects the responsiveness of other follicles to gonadotropin stimulation. This protein was purified, partially characterized, and rabbit antisera as well as monoclonal antibodies were prepared against FRP. Fixed sections of porcine ovaries were prepared on slides and then incubated with the monoclonal antibody or polyclonal antisera and then incubated with either biotinylated mouse IgM or rabbit IgG antisera, respectively. These sections were then incubated with avidin conjugated to horseradish peroxidase, followed by substrate. Staining with both the monoclonal antibody and the antisera was present in the cytoplasm of granulosa cells of small- or medium-sized antral follicles. Staining distribution was localized preferentially to cells near the basal lamina; the antral granulosa cells of viable follicles did not stain. Neither primordial follicles nor pre-antral follicles (less than 300 microns in diameter) showed any positive staining. Thecal cells were not stained in follicles less than 5 mm in diameter, whereas some large follicles (greater than 5 mm) contained staining in the theca. In the latter, specific granulosa staining was only weakly positive with the polyclonal antibody and negative with the monoclonal antibody. Atretic follicles contained significant staining of all epithelial cells adjacent to the basal lamina by both the monoclonal and polyclonal antibody preparations. Staining of the luteal ovary by the monoclonal antibody was limited to the large luteal cells. These findings suggest that FRP is produced by the granulosa cells of porcine follicles at the stage of maturation corresponding to 0.5 mm in diameter. As the viable follicle increases in size, production of FRP in the granulosa is reduced below the detectable level when the follicle exceeds 5 mm in diameter. The main source of FRP during the luteal phase is the large cell of the corpus luteum.

Alteration of follicle regulatory protein levels in human reproductive disorders: anovulation.

Recently, a follicle regulatory protein was identified that suppresses ovarian response to gonadotropins. In this study, the serum levels of follicle regulatory protein were measured in five groups of women: reproductive age undergoing oophorectomy (N = 10), postmenopausal (N = 10), ovulatory (N = 13), anovulatory (N = 16), and anovulatory receiving clomiphene citrate therapy (N = 14). Follicle regulatory protein-related immunoreactivity was measured by a competitive enzyme-linked immunosorbent assay, while peripheral estradiol and progesterone levels were determined by established radioimmunoassay. Concentration of follicle regulatory protein in serum in all ovariectomized patients decreased significantly from preoperative levels. The postmenopausal women had significantly lower follicle regulatory protein levels than did ovulatory and anovulatory women. Patients with low levels of serum estradiol in the early follicular phase exhibited either significantly elevated or suppressed follicle regulatory protein levels compared with patients with normal estradiol concentrations, suggesting two different etiologies for ovarian dysfunction. Eleven to 12 and 22-23 days after the onset of the last menstrual period, patients with elevated follicle regulatory protein levels were found to be anovulatory. These observations suggest that elevated intraovarian levels of follicle regulatory protein may cause a disruption of follicular maturation that leads to anovulation and, in some cases, to resistance to clomiphene citrate therapy.

Inhibition of ovarian cancer cell proliferation in vivo and incorporation of 3H-thymidine in vitro after follicle regulatory protein administration.

Follicle regulatory protein immunoreactivity and biologic activity were measured in ascites from a patient with juvenile granulosa cell tumor. Microscopic examination of immunohistochemical staining of a juvenile granulosa cell tumor with anti-follicle regulatory protein antisera showed homogeneous cytosolic expression of follicle regulatory protein throughout the tumor. Tumor cells were injected subcutaneously into nude mice. Partially purified follicle regulatory protein (50 micrograms/day) was then injected daily for 10 days, or for 25 days once the tumor became palpable. Treatment with follicle regulatory protein significantly slowed the rate of tumor growth with both treatments. To test the tissue specificity of the effect, a metastatic, well-differentiated endometrial adenocarcinoma was also grown in nude mice. Follicle regulatory protein treatment did not alter the rate of tumor growth. An in vitro clonigenic assay confirmed these in vivo results. Partially purified follicle regulatory protein had a biphasic effect on the proliferation of juvenile granulosa tumor cell but did not affect the proliferation of endometrial adenocarcinoma cells. Clonigenic assays were performed on five ovarian adenocarcinomas passaged in vitro, and these tumor cells exhibited a biphasic response to follicle regulatory protein. Immunoneutralization studies showed that this biphasic response was due to impurities in the follicle regulatory protein preparations. The longer the exposure of the tumor cells to follicle regulatory protein, the greater the degree of inhibition of proliferation. In summary, administration of follicle regulatory protein slowed tumor growth through a direct effect on the tumor cell rather than an indirect effect on the hormonal or immune status of the host.

Symposium overview: alterations in cytokine receptors by xenobiotics.

A symposium entitled Alterations in Cytokine Receptors by Xenobiotics was held at the 37th Annual Meeting of the Society of Toxicology (SOT) in Seattle, Washington. The symposium was sponsored by the Immunotoxicology Specialty Section of SOT and was designed to present information on the effect of several different classes of xenobiotics on various aspects of receptor function (i.e., post-receptor signal transduction of receptor expression), or the involvement of cytokine receptors in the action of the toxicant under consideration. This symposium brought together scientists in the area of receptor immunobiology whose expertise in receptor modulation encompassed those major signaling agents involved in the normal immune response, i.e., proinflammatory cytokines, chemokines, interleukins, and interferons. The following is a summary of each of the individual presentations.

Reduction of retrosternal and pericardial adhesions with rapidly resorbable polymer films.

BACKGROUND: The formation of postoperative cardiac adhesions makes a repeat sternotomy time consuming and dangerous. Many attempts have been made to solve this problem by using either drugs to inhibit fibrinolytic activity or different types of pericardial substitutes. The results have not been satisfactory. METHODS: The efficacy of bioresorbable film prototypes made of polyethylene glycol (EO) and polylactic acid (LA) (EO/LA = 1.5, 2.5, and 3.0) in the prevention of adhesions after cardiac operations in canine models was tested. After desiccation and abrasion of the epicardium, a transparent bioresorbable film was placed over the heart. The pericardium was closed to allow intrapericardial adhesions (n = 32) or left open and attached to the chest wall to induce retrosternal adhesions (n = 17). Postoperative recovery was similar among the groups. Retrosternal and pericardial adhesions were evaluated at necropsy 3 weeks later by assessing area, tenacity, and density of the adhesions. RESULTS: In the control dogs, tenacious, dense adhesions were observed. In contrast, adhesion formation was reduced at all sites covered by the films. The bioresorbable films were efficacious in the reduction of adhesion formation between epicardium and pericardium or between epicardium and sternum after cardiac operation. The EO/LA 1.5 film most effectively prevented the early adhesions. CONCLUSIONS: The bioresorbable films (EO/LA = 1.5, 2.5, and 3.0) significantly reduced adhesion formation, with EO/LA = 1.5 (Repel CV) being optimal. As the barrier was rapidly resorbed, the capsule formation induced by permanent barriers was avoided.

Effect of tolmetin sodium dihydrate on adhesion formation by intraperitoneal administration of antineoplastic agents.

Antineoplastic agents are currently being administered through catheters placed intraperitoneally to treat cancer localized to the peritoneum. This route allows for high local concentrations of antineoplastic drug at the tumor site with low levels of the drug systemically, thereby reducing the systemic toxicity. However, there are complications with this mode of delivery, including a decrease in catheter patency and induction of adhesion formation, which leads to decreased drug dispersion and limits continuing drug administration. A model was developed in rats to mimic this method of antineoplastic drug administration that produced fibrin deposition around the catheter and adhesion formation involving bowel, intestines and liver. All antineoplastic agents tested, including Adriamycin, methotrexate, bleomycin, mitoxantrone and cisplatin, induced moderate to severe adhesion formation with varying effects on catheter patency. When an intraperitoneal bolus of tometin encapsulated in liposomes was tested with Adriamycin delivered via an osmotic minipump, a reduction in adhesion formation was observed. However, highly significant adhesion reduction was found when tolmetin was coadministered with the antitumor agents.

Reduction of adhesion formation by intraperitoneal administration of Arg-Gly-Asp-containing peptides.

OBJECTIVE: To evaluate the ability of a variety of peptides containing the Arg-Gly-Asp (RGD) sequence to reduce the formation of intraperitoneal adhesions in a rabbit model. DESIGN: Prospective, randomized, double-blinded study. SETTING: University-based laboratory. ANIMAL(S): New Zealand white rabbits. INTERVENTION(S): Administration of RGD-containing peptides. MAIN OUTCOME MEASURE(S): Reduction of adhesion information. RESULT(S): In initial studies, two RGD-containing peptides (3 or a 10 amino acid peptides) were administered via Alzet miniosmotic pump to the site of injury. Administration of either of these peptides significantly reduced adhesion formation, but the larger peptide was more efficacious and reduced variability in the response. Further studies then were conducted with RGD-containing peptides five to six amino acids in length. Administration of these peptides also significantly reduced adhesion formation in a standard rabbit model. Administration of three of these peptides in a viscous vehicle at the end of surgery was also effective in reducing adhesion formation. CONCLUSION(S): The most effective combination tested was RGD-containing peptide Gly-dser-Arg-Gly-Asp-Ser-Pro in a viscous, cremophor-containing vehicle. These studies demonstrate that administration of an RGD-containing peptide was effective in reducing adhesion formation in this model.

Reduction of adhesion formation by postoperative administration of ionically cross-linked hyaluronic acid.

OBJECTIVE: To examine the efficacy of various formulations of hyaluronic acid (HA), including HA ionically cross-linked with trivalent iron, in animal models of adhesion formation. DESIGN: Hyaluronic acid formulation of varying concentrations and cross-linked densities were prepared and evaluated in a rabbit uterine horn model and a rabbit sidewall model. SETTING: ETHICON, Inc., Somerville, New Jersey. SUBJECT(S): New Zealand White rabbits. INTERVENTION(S): Test formulations were applied as intraperitoneal instillates after surgery. MAIN OUTCOME MEASURE(S): Adhesion formation was assessed at 7 and 14 days (sidewall and uterine horn model, respectively). RESULT(S): Hyaluronic acid that was not ionically cross-linked was ineffective in reducing adhesions in these models even at high viscosity, whereas the ionically cross-linked formulations of HA with trivalent iron were highly effective. Efficacy improved with increased levels of ionic cross-linking. Flowable gels, which could be delivered readily by syringe and cannula, also were effective when administered at a site remote from injury and with saline present. CONCLUSION(S): Whereas previous studies show that HA was effective in reducing adhesions peripheral to the site of injury, HA ionically cross-linked with trivalent iron was effective in reducing adhesions at all sites. From these studies, a formulation of HA ionically cross-linked with trivalent iron, 0.5% Ferric Hyaluronate Gel (LUBRICOAT; ETHICON, Inc., Somerville, NJ), was identified for subsequent clinical evaluations.

Effect of oxiplex* films (PEO/CMC) on adhesion formation and reformation in rabbit models and on peritoneal infection in a rat model.

OBJECTIVE: To assess the efficacy of Oxiplex (FzioMed, Inc., San Luis Obispo, CA) barriers. DESIGN: Film of polyethylene oxide and carboxymethylcellulose (Oxiplex) were tested for strength and tissue adherence. Films were selected for evaluation in models for biocompatability and adherence. Three films were selected for evaluation in efficacy studies, and one was evaluated for effects on bacterial peritonitis. Handling characteristics of Oxiplex film were evaluated via laparoscopy. SETTING: University laboratory. PATIENT(S): Rabbits, rats, pigs. INTERVENTION(S): Placement of Oxiplex prototypes at the site of injury. MAIN OUTCOME MEASURE(S): Mechanical properties, biocompatibility, tissue adherence, adhesion development, infection potentiation, and device handling. RESULT(S): Mechanical tests indicated that tensile strength and elongation were inversely correlated. All films tested had excellent tissue adherence properties. Selected films, based on residence time and biocompatibility, prevented adhesion formation in all animals and were highly efficacious in preventing adhesion reformation. The optimal Oxiplex prototype prevented adhesion reformation in 91% of the animals. This Oxiplex film, dyed to allow visualization, prevented adhesion reformation and did not affect bacterial peritonitis. In a laparoscopic model, the Oxiplex film, delivered in FilmSert forceps, via a 5.0-mm trocar, rapidly unfurled and could be easily applied to tissue with strong adherence. CONCLUSION(S): These data show development of an adhesion prevention material that is tissue adherent, can be placed via laparoscopy, and does not affect host resistance.

Reduction of adhesion formation with hyaluronic acid after peritoneal surgery in rabbits.

OBJECTIVE: To examine the effect of hyaluronic acid, a high-molecular-weight glucosaminoglycan found in the extracellular matrix, on the formation of adhesions, a major source of postoperative complications. DESIGN: The ability of hyaluronic acid to reduce adhesion formation was evaluated using a standardized rabbit model. The material was administered i.p. at the end of surgery. SETTING: University laboratory. ANIMAL(S): New Zealand White female rabbits. INTERVENTION(S): Intraperitoneal administration of various formulations of hyaluronic acid at the end of surgery. MAIN OUTCOME MEASURE(S): One week after surgery, a second laparotomy was performed and the extent of adhesion formation was determined. RESULT(S): Five separate molecular weight ranges of hyaluronic acid representing eight viscosities between 1,000 and 12,000 centipoise (CPS) were shown to reduce adhesion formation in this model. All volumes, 1 to 30 mL, of hyaluronic acid tested reduced adhesion formation. In addition, the low-viscosity, low-molecular-weight hyaluronic acid significantly reduced adhesion formation when added to the trauma site or when injected at a site remote from the trauma area. CONCLUSION(S): This study showed that hyaluronic acid administered at the end of surgery reduced adhesion formation.

Effects of acute administration of O,S,S-trimethyl phosphorodithioate on the generation of cellular and humoral immune responses following in vitro stimulation.

The time course of immune modulation induced by acute treatment with O,S,S-trimethyl phosphorodithioate (OSS-TMP), an impurity in technical formulations of malathion, was examined in female C57BL/6 mice. The immune parameters studied included the generation of cytotoxic T lymphocytes (CTL) to alloantigen (H-2 incompatible) and antibody secreting cells to sheep red blood cells, proliferative response to the mitogens, and interleukin-2 (IL-2) production. Acute administration of the non-toxic doses of OSS-TMP, i.e. 20 or 40 mg/kg, led to an elevation in the generation of a CTL response on day 1 or 7, respectively. At 20 mg/kg OSS-TMP, the antibody response was elevated at day 3. However, at a dose of 40 mg/kg OSS-TMP, the antibody response was suppressed at day 1 following treatment. Following acute administration of 60 or 80 mg/kg OSS-TMP, the generation of an antibody and CTL responses was suppressed at all time points tested with 1 exception. One day following treatment at a dose of 60 mg/kg OSS-TMP, there was no change in the CTL response. At day 7 following treatment, the mitogenic responses to lipopolysaccharide and phytohemagglutinin were elevated at all doses of OSS-TMP administered. At this time point, however, the proliferative response to Concanavalin A was elevated in a dose dependent manner. IL-2 production was suppressed following acute administration of 60 or 80 mg/kg OSS-TMP at all time points tested and at all doses tested on day 5 following treatment. These data indicate that OSS-TMP, unlike its congener, O,O,S-trimethyl phosphorothioate, enhances the generation of humoral and cell mediated immune responses of C57BL/6 mice following administration of non-toxic doses.

Effects of O,S,S-trimethyl phosphorodithioate on immune function.

The effect of acute administration of 20-80 mg/kg O,S,S-trimethyl phosphorodithioate (OSS-TMP) to C57BL/6 female mice on the murine immune system was determined. The parameters examined to evaluate overt toxicity of the compound included body weight, plasma cholinesterase levels, splenic nucleated cell number and thymic weight and nucleated cell number. Acute administration of 60 or 80 mg/kg OSS-TMP led to a 75 or 63% decrease, respectively, in plasma cholinesterase levels and a decrease in thymic size. At a dose of 80 mg/kg OSS-TMP, the animals also exhibited some lethargy and body weight loss. Below 60 mg/kg OSS-TMP, no overt toxic manifestations were observed. These studies were carried further to determine the effect of OSS-TMP on the generation of in vivo primary and in vitro secondary cellular and humoral immune responses. At nontoxic doses of the compound, i.e. 20 and 40 mg/kg OSS-TMP, the in vivo generation of a primary cytotoxic T lymphocyte (CTL) response to alloantigen was significantly elevated, but this response was unaffected following restimulation of the splenocytes by alloantigen in vitro. The generation of an in vivo primary and in vitro secondary humoral responses to sheep red blood cells (SRBC) was elevated following a single dose of 40 mg/kg OSS-TMP. Administration of toxic doses of OSS-TMP, i.e. 60 and 80 mg/kg, did not alter the ability of splenocytes to generate a primary or secondary CTL response, but suppressed the generation of humoral immune responses. These results differ significantly from those observed in a similar system following acute administration of a structural analog, O,O,S-trimethyl phosphorothioate which was previously shown to have potent immunosuppressive activity at nontoxic doses.

Effects of subacute administration of O,S,S-trimethyl phosphorodithioate on cellular and humoral immune response systems.

The effects of 14-day treatment with low doses of O,S,S-trimethyl phosphorodithioate (OSS-TMP), an impurity in technical malathion, on the generation of cell-mediated and humoral immune responses were examined in female C57BL/6 mice. At a dose of 2.0 mg/kg per day OSS-TMP, the generation of antibody-secreting cells to sheep red blood cells, the generation of cytotoxic T lymphocytes (CTL) to alloantigen and the production of Interleukin-2 were elevated approximately 2-3 fold, while no changes were observed in the proliferative responses to the polyclonal activators, Concanavalin A, lipopolysaccharide, or phytohemagglutinin. In contrast, at 5.0 mg/kg per day OSS-TMP, both the CTL and specific antibody responses were suppressed, while all other immune parameters examined were unchanged. Data from cell separation and reconstitution experiments indicated that both T and B lymphocytes were affected by these treatment regimes. These data suggest that long-term exposure to low doses of OSS-TMP may enhance the ability of an animal to generate an immune response while higher doses of OSS-TMP may suppress the generation of an immune response.

Function of peritoneal exudate cells after abdominal surgery.

Peritoneal macrophages and polymorphonuclear neutrophils are key cells in the repair of postoperative injury. Increased numbers of macrophages migrate into the peritoneal cavity after operation and the function of these cells changes over the postoperative interval. Macrophage activities, such as respiratory burst, arachidonic acid metabolism, monokine secretion, and plasminogen activator inhibitory activity, are elevated by peritoneal operation. However, the secretion of plasminogen activator activity is decreased after operation. The kinetics with which each of these functions changes varies with the parameter examined, indicating a complex regulation of the differentiation of leukocytes after operation. In addition, the activity of postoperative macrophages can be modulated in vitro by exposure to cytokines and conditioned media from polymorphonuclear neutrophils and macrophages. Thus, cell-cell interactions and factors secreted within the peritoneal cavity may regulate the contribution of postoperative leukocytes to peritoneal repair after operation.

Kinetics of interleukin-1 and tumor necrosis factor secretion by rabbit macrophages recovered from the peritoneal cavity after surgery.

Macrophages play a crucial role in wound healing after surgical injury, both as scavenger cells responsible for wound debridement and as cells that secrete soluble factors such as interleukin-1 (IL-1) and tumor necrosis factor (TNF). IL-1 and TNF alter many of the biological activities of cells that appear in postsurgical wounds. In this study, we determined the kinetics of IL-1 and TNF production by rabbit macrophages harvested from postsurgical peritoneal exudate (postsurgical macrophages) at several time points after peritoneal surgery. To further characterize the level of functional activities of postsurgical macrophages, the IL-1 and TNF levels were determined with or without stimulating the cells with lipopolysaccharide (LPS) and phorbol myristate acetate (PMA). After surgery, the number of macrophages harvested by peritoneal lavage increased, reached peak levels on postsurgical day 3, and then decreased. IL-1 levels secreted by macrophages cultured without stimuli were elevated on postsurgical day 14 compared to the values on day 3 and 7. TNF concentrations peaked on days 1 and 14. In the conditioned culture media from LPS-PMA-stimulated macrophages, the levels of both IL-1 and TNF peaked on postsurgical days 3 and 14. These data suggest that the susceptibility of postsurgical macrophages to stimuli changes during the wound healing process with maximum sensitivity to the stimuli present during the early phase of peritoneal repair (day 3).

Reduction of adhesion formation by intraperitoneal administration of anti-inflammatory peptide 2.

Adhesion formation is a major source of postoperative morbidity and mortality. Therefore, the reduction of postoperative adhesion formation would be of clinical benefit. Various modalities have been shown to reduce adhesion formation, including fibrinolytic enzymes, nonsteroidal anti-inflammatory drugs, and barriers that reduce the apposition of sites of potential adhesion formation. This study examined the ability of a phospholipase A2 inhibitor, anti-inflammatory peptide 2 (antinflammin), to reduce the formation of intraperitoneal adhesions in two rabbit models of adhesion formation. In the sidewall model, antinflammin was administered via Alzet miniosmotic pump for the entire postoperative interval, and there was a dose-dependent reduction in the area of the sidewall injury that was involved in adhesions to the cecum and the bowel. In the double uterine horn model, antinflammin was administered via Alzet miniosmotic pump to the area of injury for either 1, 2, 3, or 7 days. Administration of antinflammin for as little as 24 h after surgery significantly reduced the extent of adhesion formation. Administration of the peptide for longer periods of time did not further increase the reduction in adhesion formation. These studies clearly demonstrate that postoperative administration of antinflammin to the site of injury reduced the formation of postoperative adhesions in two animal models.