The recruitment of extra-intestinal cells to the injured mucosa promotes healing in radiation enteritis and chemical colitis in a mouse parabiosis model.

Mucosal healing occurs through migration and proliferation of cells within injured epithelium, yet these processes may be inadequate for mucosal healing after significant injury where the mucosa is denuded. We hypothesize that extra-intestinal cells can contribute to mucosal healing after injury to the small and large intestine. We generated parabiotic pairs between wild-type and tdTomato mice, which were then subjected to radiation-induced enteritis and 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis. We now show that as compared with singleton mice, mice with a parabiotic partner were protected against intestinal damage as revealed by significantly reduced weight loss, reduced expression of pro-inflammatory cytokines, reduced enterocyte apoptosis, and improved crypt proliferation. Donor cells expressed CD45-, Sca-1+, c-kit+, and CXCR4+ and accumulated around the injured crypts but did not transdifferentiate into epithelia, suggesting that extra-intestinal cells play a paracrine role in the healing response, while parabiotic pairings with Rag1-/- mice showed improved healing, indicating that adaptive immune cells were dispensable for mucosal healing. Strikingly, ablation of the bone marrow of the donor parabionts removed the protective effects. These findings reveal that the recruitment of extra-intestinal, bone marrow-derived cells into the injured intestinal mucosa can promote mucosal healing, suggesting novel therapeutic approaches for severe intestinal disease.

Intestinal epithelial Toll-like receptor 4 prevents metabolic syndrome by regulating interactions between microbes and intestinal epithelial cells in mice.

Little is known about the pathogenesis of metabolic syndrome, although Toll-like receptor 4 (TLR4) has been implicated. We investigated whether TLR4 in the intestinal epithelium regulates metabolic syndrome by coordinating interactions between the luminal microbiota and host genes that regulate metabolism. Mice lacking TLR4 in the intestinal epithelium (TLR4ΔIEC), but not mice lacking TLR4 in myeloid cells nor mice lacking TLR4 globally, developed metabolic syndrome; these features were not observed in TLR4ΔIEC mice given antibiotics. Metagenomic analysis of the fecal microbiota revealed differences between TLR4ΔIEC and wild-type mice, while meta-transcriptome analysis of the microbiota showed that intestinal TLR4 affected the expression of microbial genes involved in the metabolism of lipids, amino acids, and nucleotides. Genes regulated by peroxisome proliferator-activated receptors (PPARs) and the antimicrobial peptide lysozyme were significantly downregulated in TLR4ΔIEC mice, suggesting a mechanism by which intestinal TLR4 could exert its effects on the microbiota and metabolic syndrome. Supportingly, antibiotics prevented both downregulation of PPAR genes and the development of metabolic syndrome, while PPAR agonists prevented development of metabolic syndrome in TLR4ΔIEC mice. Thus, intestinal epithelial TLR4 regulates metabolic syndrome through altered host-bacterial signaling, suggesting that microbial or PPAR-based strategies might have therapeutic potential for this disease.

Breast milk protects against the development of necrotizing enterocolitis through inhibition of Toll-like receptor 4 in the intestinal epithelium via activation of the epidermal growth factor receptor.

Breast milk is the most effective strategy to protect infants against necrotizing enterocolitis (NEC), a devastating disease that is characterized by severe intestinal necrosis. Previous studies have demonstrated that the lipopolysaccharide receptor Toll-like receptor 4 (TLR4) plays a critical role in NEC development via deleterious effects on mucosal injury and repair. We now hypothesize that breast milk protects against NEC by inhibiting TLR4 within the intestinal epithelium, and sought to determine the mechanisms involved. Breast milk protected against NEC and reduced TLR4 signaling in wild-type neonatal mice, but not in mice lacking the epidermal growth factor receptor (EGFR), whereas selective removal of EGF from breast milk reduced its protective properties, indicating that breast milk inhibits NEC and attenuates TLR4 signaling via EGF/EGFR activation. Overexpression of TLR4 in the intestinal epithelium reversed the protective effects of breast milk. The protective effects of breast milk occurred via inhibition of enterocyte apoptosis and restoration of enterocyte proliferation. Importantly, in IEC-6 enterocytes, breast milk inhibited TLR4 signaling via inhibition of glycogen synthase kinase-3ß (GSK3ß). Taken together, these findings offer mechanistic insights into the protective role for breast milk in NEC, and support a link between growth factor and innate immune receptors in NEC pathogenesis.

Diagnosis and outcome of Hirschsprung's disease: does age really matter?

Although Hirschsprung's disease (HD) typically presents in the newborn period, it is often diagnosed in older children, in whom the presentation and management remain poorly defined. We hypothesized that older patients with HD have a milder variant of the disease with an improved prognosis compared with those diagnosed earlier. Children with HD (1995-2001) were divided into Group I (diagnosis before 30 days) and Group II (after 30 days). Patients with total colonic disease were excluded. There were 66 patients; 47 in Group I and 19 in group II. Mean age at diagnosis was 7.1+/-1.3 days (range 1-30 days) versus 27+/-10 months (1.3 months-19 years). Older children differed mainly in the symptoms at presentation and the length of the involved segment of aganglionosis. Surgical strategies were applied equally in both groups. Complications, including postoperative enterocolitis, occurred equally, but the length of stay and costs were lower in Group II. The delayed diagnosis of HD does not worsen outcomes of older children with HD. This finding implies that these children have a milder form of the disease, perhaps because of adaptation to the aganglionic state.

Indirect role for COPI in the completion of FCgamma receptor-mediated phagocytosis.

Recent evidence suggests that extension of pseudopods during phagocytosis requires localized insertion of endomembrane vesicles. The nature of these vesicles and the processes mediating their release and insertion are unknown. COPI plays an essential role in the budding and traffic of membrane vesicles in intracellular compartments. We therefore assessed whether COPI is also involved in phagosome formation. We used ldlF cells, a mutant line derived from Chinese hamster ovary cells that express a temperature-sensitive form of epsilonCOP. To confer phagocytic ability to ldlF cells, they were stably transfected with Fc receptors type IIA (FcgammaRIIA). In the presence of functional COPI, FcgammaRIIA-transfected ldlF cells effectively internalized opsonized particles. In contrast, phagocytosis was virtually eliminated after incubation at the restrictive temperature. Similar results were obtained impairing COPI function in macrophages using brefeldin A. Notably, loss of COPI function preceded complete inhibition of phagocytosis, suggesting that COPI is indirectly required for phagocytosis. Despite their inability to internalize particles, COPI-deficient cells nevertheless expressed normal levels of FcgammaRIIA, and signal transduction appeared unimpeded. The opsonized particles adhered normally to COPI-deficient cells and were often found on actin-rich pedestals, but they were not internalized due to the inability of the cells to extend pseudopods. The failure to extend pseudopods was attributed to the inability of COPI-deficient cells to mobilize endomembrane vesicles, including a VAMP3-containing compartment, in response to the phagocytic stimulus.

Effects of other intra-abdominal injuries on the diagnosis, management, and outcome of small bowel trauma.

BACKGROUND: Difficulty with and delays in diagnosis are possible causes of increased morbidity and mortality in small bowel injuries. We assessed whether multiple intra-abdominal injuries led to earlier laparotomy and whether this resulted in improved outcome. METHODS: Patients with small bowel injuries between January 1993 and December 1997 from the trauma database at St. Michael's Hospital in Toronto were assessed after dividing them into those with isolated small bowel injury ("isolated") and those with small bowel injuries in association with other intra-abdominal injuries ("nonisolated"). Parameters compared were age, gender, length of stay, mortality, intra-abdominal complications, mechanism of injury, diagnostic time, and how the diagnosis was made. RESULTS: Of 1,207 patients, 244 sustained abdominal injuries, and 83 had small bowel injuries (30 patients in the isolated group and 53 in the nonisolated group). Groups were similar with respect to age and gender, yet differed significantly with respect to mechanism and mean Injury Severity Scores (isolated, 18 +/- 8 vs. nonisolated, 30 +/- 15). Outcome differed between groups, as mortality (isolated, 0 of 30 vs. nonisolated, 4 of 53 deaths), length of stay (isolated, 13 +/- 2 vs. nonisolated, 22 +/- 3 days), and patients with intra-abdominal complications (isolated, 5 of 30 vs. nonisolated, 14 of 53 patients) were significantly higher in the nonisolated group. Time to diagnosis was significantly less in the nonisolated group. Decision for laparotomy and diagnosis of small bowel injuries were based more on physical findings in the nonisolated group and on computed tomography in the isolated group. CONCLUSION: The presence of associated intra-abdominal injuries significantly affects presentation and outcome of patients with small bowel injuries and the selection of diagnostic modalities.

Role of COPI in phagosome maturation.

Phagosomes mature by sequentially fusing with endosomes and lysosomes. Vesicle budding is presumed to occur concomitantly, mediating the retrieval of plasmalemmal components and the regulation of phagosomal size. We analyzed whether fission of vesicles from phagosomes requires COPI, a multimeric complex known to be involved in budding from the Golgi and endosomes. The role of COPI was studied using ldlF cells, that harbor a temperature-sensitive mutation in epsilon-COP, a subunit of the coatomer complex. These cells were made phagocytic toward IgG-opsonized particles by heterologous expression of human FcgammaRIIA receptors. Following incubation at the restrictive temperature, epsilon-COP was degraded in these cells and their Golgi complex dispersed. Nevertheless, phagocytosis persisted for hours in cells devoid of epsilon-COP. Retrieval of transferrin receptors from phagosomes became inefficient in the absence of epsilon-COP, while clearance of the FcgammaRIIA receptors was unaffected. This indicates that fission of vesicles from the phagosomal membrane involves at least two mechanisms, one of which requires intact COPI. Traffic of fluid-phase markers and aggregated IgG-receptor complexes along the endocytic pathway was abnormal in epsilon-COP-deficient cells. In contrast, phagosome fusion with endosomes and lysosomes was unimpaired. Moreover, the resulting phagolysosomes were highly acidic. Similar results were obtained in RAW264.7 macrophages treated with brefeldin A, which precludes COPI assembly by interfering with the activation of adenosine ribosylation factor. These data indicate that neither phagosome formation nor maturation are absolutely dependent on COPI. Our findings imply that phagosomal maturation differs from endosomal progression, which appears to be more dependent on COPI-mediated formation of carrier vesicles.

Functional expression of Nramp1 in vitro in the murine macrophage line RAW264.7.

Mutations at the Nramp1 locus in vivo cause susceptibility to infection by unrelated intracellular microbes. Nramp1 encodes an integral membrane protein abundantly expressed in the endosomal-lysosomal compartment of macrophages and is recruited to the phagosomal membrane following phagocytosis. The mechanism by which Nramp1 affects the biochemical properties of the phagosome to control microbial replication is unknown. To devise an in vitro assay for Nramp1 function, we introduced a wild-type Nramp1(G169) cDNA into RAW 264.7 macrophages (which bear a homozygous mutant Nramp1(D169) allele and thus are permissive to replication of specific intracellular parasites). Recombinant Nramp1 was expressed in a membranous compartment in RAW264.7 cells and was recruited to the membrane of Salmonella typhimurium and Yersinia enterocolitica containing phagosomes. Evaluation of the antibacterial activity of RAW264.7 transfectants showed that expression of the recombinant Nramp1 protein abrogated intracellular replication of S. typhimurium. Studies with a replication-defective S. typhimurium mutant suggest that this occurs through an enhanced bacteriostatic activity. The effect of Nramp1 expression was specific, since (i) it was not seen in RAW264.7 transfectants overexpressing the closely related Nramp2 protein, and (ii) control RAW264.7 cells, Nramp1, and Nramp2 transfectants could all efficiently kill a temperature-sensitive, replication-defective mutant of S. typhimurium. Finally, increased antibacterial activity of the Nramp1 RAW264.7 transfectants was linked to increased phagosomal acidification, a distinguishing feature of primary macrophages expressing a wild-type Nramp1 allele. Together, these results indicate that transfection of Nramp1 cDNAs in the RAW264.7 macrophage cell line can be used as a direct assay to study both Nramp1 function and mechanism of action as well as to identify structure-function relationships in this protein.

The iron transport protein NRAMP2 is an integral membrane glycoprotein that colocalizes with transferrin in recycling endosomes.

The natural resistance associated macrophage protein (Nramp) gene family is composed of two members in mammals, Nramp1 and Nramp2. Nramp1 is expressed primarily in macrophages and mutations at this locus cause susceptibility to infectious diseases. Nramp2 has a much broader range of tissue expression and mutations at Nramp2 result in iron deficiency, indicating a role for Nramp2 in iron metabolism. To get further insight into the function and mechanism of action of Nramp proteins, we have generated isoform specific anti-Nramp1 and anti-Nramp2 antisera. Immunoblotting experiments indicate that Nramp2 is present in a number of cell types, including hemopoietic precursors, and is coexpressed with Nramp1 in primary macrophages and macrophage cell lines. Nramp2 is expressed as a 90-100-kD integral membrane protein extensively modified by glycosylation (>40% of molecular mass). Subcellular localization studies by immunofluorescence and confocal microscopy indicate distinct and nonoverlapping localization for Nramp1 and Nramp2. Nramp1 is expressed in the lysosomal compartment, whereas Nramp2 is not detectable in the lysosomes but is expressed primarily in recycling endosomes and also, to a lower extent, at the plasma membrane, colocalizing with transferrin. These findings suggest that Nramp2 plays a key role in the metabolism of transferrin-bound iron by transporting free Fe2+ across the endosomal membrane and into the cytoplasm.

Snow-related recreational injuries in children: assessment of morbidity and management strategies.

PURPOSE: The aim of this study was to investigate the causes, clinical course, and financial impact of snow-related sport injuries in children. METHODS: Reports of snow-related injuries (skiing, toboganning, snowboarding) occurring in 147 consecutive children (< or =16 years of age) admitted from 1991 through 1997 were collected prospectively and assessed retrospectively. During the last year of the study, outpatients treated and released from the emergency department (1996 through 1997) were examined in parallel (n = 101). Total financial impact was determined from the aggregate hospital, rehabilitation, and societal costs. RESULTS: One hundred thirty-seven patients (M:F, 2:1; mean age, 13 yrs) were admitted (toboggan [n = 74], ski [n = 59], snowboard [n = 16]), of which 66% occurred at licensed resorts, and 33% at parks or private property. There was one death. Although the pattern of injury was similar in all groups (head greater than long bone greater than intraabdominal injuries), mean injury severity scores (ISS) were significantly higher for snowboard injuries. Seventy-five percent of patients required at least one operation. Postdischarge, 15% of patients required institutional care. Of the 101 ambulatory patients (ski [n = 48], toboggan [n = 35], snowboard [n = 18]), 65% were injured at licensed resorts, and 56% required outpatient rehabilitation or home care. The per-patient costs were: hospital treatment, $27,936; outpatient services, $15,243; lost parental income, $1,500. CONCLUSIONS: Snow sport injuries, particularly snowboarding, cause severe childhood morbidity. Helmet usage, training requirements, and regulation of licensed resorts may reduce the morbidity and staggering costs.

Bioavailability of oral ciprofloxacin in early postsurgical patients.

OBJECTIVE: To evaluate the absorption of oral ciprofloxacin within 24 hours of laparotomy for major elective surgery or peritonitis. DESIGN: In this prospective trial, patients were given a 750-mg oral close the morning after major elective surgery (n=15) or surgery for peritonitis (n=7). Healthy volunteers served as controls (n=9). Serial urine and blood samples were drawn during the subsequent 12 hours, and pharmacokinetic measures were determined by standard high-performance liquid chromatography assay procedures. SETTING: Multicenter, university-affiliated hospitals. MAIN OUTCOME MEASURES: Drug absorption as determined by area under the concentration time curve, maximum concentration, and time to maximum concentration. RESULTS: Oral bioavailability was reduced in elective surgery and peritonitis patients compared with controls. Among the 15 elective surgery patients, 27% (4/15) showed no absorption. The remaining 73% (11/15) had an area under the curve comparable with that of controls (8.3+/-1.6 (mg/[L x h]). Among all patients, those who showed drug absorption vs those who showed no absorption did not differ with respect to malignant neoplasm, case type, age, or biochemistry. However, patients showing no absorption were significantly heavier than patients showing absorption (patients showing absorption, 15%+/-3% over ideal body weight vs patients showing no absorption, 29%+/-6% over ideal body weight; P<.05). When elective surgery patients were stratified by presence or absence of obesity (25% above ideal body weight), mean area under the curve in nonobese patients was 9.80+/-2.37 vs 0.91+/-0.56 (mg/(L x h) in obese patients (P<.05). CONCLUSIONS: Oral bioavailability was reduced for peritonitis surgery patients on the first day postoperatively, and for obese elective surgery patients. To achieve adequate serum levels requires continuation of intravenous antibiotics in patients with peritonitis, and adjustment of oral dosage in obese patients in the early period after elective surgery.

v-SNARE-dependent secretion is required for phagocytosis.

Phagosomes are generally believed to form by gradual apposition of the plasma membrane of leukocytes onto the surface of invading microorganisms. The internalization of the encapsulated particle is therefore predicted to reduce the surface area of the phagocyte. Contrary to this prediction, we observed that phagocytosis is associated with a net increase in cell surface area, suggesting the concomitant occurrence of exocytosis. Selective cleavage of components of the secretory machinery by microinjection or transfection of bacterial neurotoxins induced a pronounced inhibition of phagocytosis. These observations indicate that vesicle-soluble N-ethylmaleimide-sensitive factor attachment protein receptor-mediated exocytosis of endomembranes is essential for optimal completion of particle internalization during phagocytosis.

Host resistance to intracellular infection: mutation of natural resistance-associated macrophage protein 1 (Nramp1) impairs phagosomal acidification.

The mechanisms underlying the survival of intracellular parasites such as mycobacteria in host macrophages remain poorly understood. In mice, mutations at the Nramp1 gene (for natural resistance-associated macrophage protein), cause susceptibility to mycobacterial infections. Nramp1 encodes an integral membrane protein that is recruited to the phagosome membrane in infected macrophages. In this study, we used microfluorescence ratio imaging of macrophages from wild-type and Nramp1 mutant mice to analyze the effect of loss of Nramp1 function on the properties of phagosomes containing inert particles or live mycobacteria. The pH of phagosomes containing live Mycobacterium bovis was significantly more acidic in Nramp1- expressing macrophages than in mutant cells (pH 5.5 +/- 0.06 versus pH 6.6 +/- 0.05, respectively; P <0.005). The enhanced acidification could not be accounted for by differences in proton consumption during dismutation of superoxide, phagosomal buffering power, counterion conductance, or in the rate of proton "leak", as these were found to be comparable in wild-type and Nramp1-deficient macrophages. Rather, after ingestion of live mycobacteria, Nramp1-expressing cells exhibited increased concanamycin-sensitive H+ pumping across the phagosomal membrane. This was associated with an enhanced ability of phagosomes to fuse with vacuolar-type ATPase-containing late endosomes and/or lysosomes. This effect was restricted to live M. bovis and was not seen in phagosomes containing dead M. bovis or latex beads. These data support the notion that Nramp1 affects intracellular mycobacterial replication by modulating phagosomal pH, suggesting that Nramp1 plays a central role in this process.

Enterocolitis after the surgical treatment of Hirschsprung's disease: risk factors and financial impact.

BACKGROUND/PURPOSE: Enterocolitis (EC) represents a serious complication after the surgical correction of Hirschsprung's disease (HD). Although previous studies have identified risk factors associated with the development of this complication before definitive repair, the factors leading to EC after pull-through have not been examined. This study was therefore designed to determine risk factors for the development of post-pull-through EC. METHODS: Patients with HD treated from 1991 through 1996 at the Hospital for Sick Children in Toronto, Canada were assessed. Risk factors were examined in three areas: patient factors (gender, age at diagnosis, age and weight at pull-through), technical factors (type of repair, number of stages, location of transition zone, previous EC), and mechanical factors. RESULTS: In 105 consecutive patients, the incidence of postoperative EC was 32%. There was no mortality. The risk of postoperative EC was significantly increased by mechanical factors related to anastomotic complications (relative risk, 2.8) and intestinal obstruction (relative risk, 3.5). This finding was not attributable to the general occurrence of any postoperative complication because the incidence of postoperative complications was equally distributed in patients with and without EC. The presence of EC significantly increased the number of hospital admissions, mean length of stay, and total treatment cost. CONCLUSION: These findings suggest the use of measures to decrease mechanical obstruction so as to decrease the incidence and impact of this potentially devastating complication.

Host response to laparoscopic surgery: mechanisms and clinical correlates.

Minimal access surgery has revolutionized the treatment of a variety of surgical diseases, partly because it is associated with less patient morbidity than nonlaparoscopic surgical procedures. Emerging evidence suggests that alteration in the host response after laparoscopic procedures has significantly contributed to the improved postoperative course. Laparoscopy modulates both afferent stimuli (including tissue trauma, pain and wound size) and efferent responses (via neuroendocrine, metabolic, immunologic and cardiorespiratory systems). These effects lead to a decrease in postoperative pain, fever and disability. Laparoscopy mediates these effects through reduced wound size, the activities of endotoxin and immunomodulatory actions of the insufflated gas, resulting in impaired macrophage activity. Although clearly beneficial in reducing postoperative morbidity after elective surgery, this immunosuppression could increase the risk of complications during procedures for infection or neoplasia.

Regulation of phagosomal acidification. Differential targeting of Na+/H+ exchangers, Na+/K+-ATPases, and vacuolar-type H+-atpases.

Vacuolar-type (V) ATPases are thought to be the main determinant of phagosomal acidification. In phagosomes containing mycobacteria, which ostensibly impair the delivery of V-ATPases to the phagosomal membrane, the pH would be expected to be near neutral. This prediction was tested by microfluorescence ratio imaging using macrophages from mice susceptible to mycobacterial infection. Although less acidic than their counterparts containing dead bacteria, phagosomes containing live Mycobacteria bovis were nearly 1 pH unit more acidic than the cytosol, suggesting the existence of alternate H+ transport mechanisms. We therefore investigated whether Na+/H+ exchange (NHE) contributes to phagosomal acidification. Immunoblotting, reverse transcriptase-polymerase chain reaction, and pharmacological studies indicated that NHE1 is the predominant isoform of the exchanger in macrophages. Fractionation revealed that NHE1 is incorporated into the phagosomal membrane, and measurements of pH indicated that it is functional in this location. Nevertheless, acidification of the lumen of phagosomes containing either latex beads or live M. bovis was insensitive to (3-methylsulfonyl-4-piperidinobenzoyl)-guanidine methanesulfonate, a potent inhibitor of NHE1. This may have been due to the absence of an appropriate lumen to cytosol Na+ gradient, because the phagosomal membrane was found to be devoid of Na+/K+ pumps. Unexpectedly, the acidification of M. bovis phagosomes was fully reversed by specific inhibitors of the vacuolar H+-ATPase, suggesting that ATPases are present only transiently or in reduced quantities in the phagosomal membrane. Alternatively, acid equivalents accumulated in endosomes by V-ATPases may be delivered to the mycobacterial phagosome by carrier vesicles devoid of ATPases.

Rho is required for the initiation of calcium signaling and phagocytosis by Fcgamma receptors in macrophages.

Phagocytosis of bacteria by macrophages and neutrophils is an essential component of host defense against infection. The mechanism whereby the interaction of opsonized particles with Fcgamma receptors triggers the engulfment of opsonized particles remains incompletely understood, although activation of tyrosine kinases has been recognized as an early step. Recent studies in other systems have demonstrated that tyrosine kinases can in turn signal the activation of small GTPases of the ras superfamily. We therefore investigated the possible role of Rho in Fc receptor-mediated phagocytosis. To this end we microinjected J774 macrophages with C3 exotoxin from Clostridium botulinum, which ADP-ribosylates and inactivates Rho. C3 exotoxin induced the retraction of filopodia, the disappearance of focal complexes, and a global decrease in the F-actin content of J774 cells. In addition, these cells exhibited increased spreading and the formation of vacuolar structures. Importantly, inactivation of Rho resulted in the complete abrogation of phagocytosis. Inhibition of Fcgamma receptor-mediated phagocytosis by C3 exotoxin was confirmed in COS cells, which become phagocytic upon transfection of the FcgammaRIIA receptor. Rho was found to be essential for the accumulation of phosphotyrosine and of F-actin around phagocytic cups and for Fcgamma receptor-mediated Ca2+ signaling. The clustering of receptors in response to opsonin, an essential step in Fcgamma-induced signaling, was the earliest event shown to be inhibited by C3 exotoxin. The effect of the toxin was specific, since clustering and internalization of transferrin receptors were unaffected by microinjection of C3. These data identify a role for small GTPases in Fcgamma receptor-mediated phagocytosis by leukocytes.

Mechanism of decreased in vitro murine macrophage cytokine release after exposure to carbon dioxide: relevance to laparoscopic surgery.

OBJECTIVE: The objective of this study was to determine the effect of carbon dioxide (CO2) on the function of peritoneal macrophages. SUMMARY BACKGROUND DATA: Laparoscopic surgery is associated with minimal pain, fever, and low levels of inflammatory cytokines. To understand the mechanisms involved, the authors investigated the effect of different gases on murine peritoneal macrophage intracellular pH and correlated these alterations with alterations in LPS-stimulated inflammatory cytokine release. METHODS: Peritoneal macrophages were incubated for 2 hours in air, helium, or CO2, and the effect of the test gas on immediate or next day lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNF) and interleukin-1 release compared. Cytosolic pH of macrophages exposed to test gases was measured using single-cell fluorescent imaging. The in vivo effects of test gases were determined in anesthetized rats during abdominal insufflation. RESULTS: Macrophages incubated in CO2 produced significantly less TNF and interleukin-1 in response to LPS compared to incubation in air or helium. Cytokine production returned to normal 24 hours later. Exposure to CO2, but not air or helium, caused a marked cytosolic acidification. Pharmacologic induction of intracellular acidification to similar levels reproduced the inhibitory effect. In vitro studies showed that CO2 insufflation lowered tissue pH and peritoneal macrophage LPS-stimulated TNF production. CONCLUSIONS: The authors propose that cellular acidification induced by peritoneal CO2 insufflation contributes to blunting of the local inflammatory response during laparoscopic surgery.

Single-stage repair of Hirschsprung's disease: a comparison of 109 patients over 5 years.

Current management of Hirschsprung's disease (HD) typically involves staged therapy, which necessitates multiple hospital admissions and associated costs. The authors therefore investigated the course and outcome of treating such children using a single-staged (SS) approach, and compared them with those treated via multiple-staged (MS) therapy. The cases of one hundred nine consecutive patients who presented with HD from 1991 to 1996 were reviewed. Four patients were excluded (two unrelated deaths, two with small intestinal aganglionosis). Twenty-one of the remaining 105 patients underwent SS repair. Both groups were similar in gender, age at diagnosis, and frequency of comorbidities. Repair was possible in 100% of the SS patients. Complications, including enterocolitis, occurred in 63% of patients, and did not significantly differ between groups. The outcome in SS patients was unaffected by whether the repair was performed before or after 30 days of life. The outcome was unaffected by operative weight in either group. By contrast, the number of hospital admissions and total length of stay was significantly higher in the MS group, which resulted in a twofold increase in total costs associated with MS repair compared with SS repair. These data indicate that primary repair of HD is efficacious (even in the newborn), with morbidity equal to MS repair, and requires fewer hospital admissions. The significant savings to the patient and the health care system suggest that SS repair may be an improved strategy for treating HD.

Enteric duplication in the adult, derived from the foregut, midgut and hindgut: presentation, patterns and literature review.

Duplication of the alimentary tract may affect patients of all ages. Although they are relatively rare, the importance of these congenital lesions lies in the fact that they readily mimic other surgical disease processes and may result in significant morbidity if left untreated. Prompt recognition and treatment using combined radiologic and surgical management are generally associated with an excellent outcome. Three patients who presented with intestinal duplication arising from each of the major embryologic origins are reported. Their clinical histories reveal the spectrum of presentation associated with these lesions and provide a framework for a discussion of current management strategies.