Keith T. Oldham

Evidence for tumor necrosis factor-induced pulmonary microvascular injury after intestinal ischemia-reperfusion injury

Acute lung injury characterized by increased microvascular permeability Is one feature of multiple-organ system failure and the adult respiratory distress syndrome. Intestinal ischemia–reperfusion injury has been linked to this type of acute lung injury. The purpose of these experiments was to examine the pathogenic mediators that link the two processes, with particular emphasis on the roles of endotoxln and tumor necrosis factor α (TNFα). Previously described characteristics of the acute lung Injury in this rat model of Intestinal ischemia-reperfusion include pulmonary neutrophil sequestration, depletion of lung tissue ATP, alveolar endotnelial cell disruption, and increased microvascular permeability. Plasma levels of TNF In the systemic circulation of sham-operated animals and those with intestinal ischemlc injury less than 60 minutes In duration were very low or undetectable. Intestinal ischemia for 120 minutes was associated with TNF elevation to 1.19 ± 0.50 U/mL. Reperfusion for periods of 15 and 30 minutes generated 5− to 10-fold Increases In circulating TNF levels (6.61 ± 3.11 U/mL, p > 0.05 and 10.41 ± 5.41 U/mL, p.= 0.004 compared to sham); however this increase in circulating TNF was transient and largely cleared within 60 minutes after Initiating reperfusion. Portal vein endotoxln levels were found to increase significantly before the appearance of TNF in systemic plasma, suggesting that gut-derived endotoxin may Induce TNF release from hepatic macrophages into the systemic circulation. Antl-TNF antibody attenuated the increase in pulmonary microvascular permeability In this preparation but did not prevent pulmonary, neutrophil sequestration. These observations suggest that endotoxin and TNF have pathogenic roles in this acute lung Injury, but that mechanisms of adherence of neutrophils to endotnelial cells Independent of TNF may be Involved. The accumulation of neutrophils in the lung but the prevention of a vascular permeability increase in the presence of antibody to TNF may imply an in vivo role for TNF in the process of neutrophil activation. These studies provide additional evidence of the Importance of the endogenous inflammatory mediators in the development of systemic injury in response to local tissue Injury.

The production of tumor necrosis factor alpha and the development of a pulmonary capillary injury following hepatic ischemia/reperfusion.

The large mass of fixed macrophages resident in the liver make it a potentially rich source of cytokines. We have previously demonstrated that an isolated and severe ischemia/reperfusion injury to the liver results in cytokine release, specifically tumor necrosis factor alpha, and that TNF is then involved in the development of pulmonary pathology. This study was designed to determine the kinetics of TNF release following varying periods of hepatic ischemia and to further investigate the acute lung injury that follows. Suprahepatic blood samples were obtained at serial time points following a 45-, 60-, 75-, or 90-min ischemic insult to a segment of the rat liver with subsequent reperfusion. Using a bioassay based on the WEHI 164 cell line, plasma TNF levels were measured in all experimental animals; sham-operated control animals had undetectable levels. Changes in pulmonary capillary permeability were then measured using a standard 125I-labeled albumin washout technique following a 90-min ischemic insult with subsequent reperfusion. A significant increase in the mean permeability index was observed 9 to 12 hr following hepatic reperfusion (.601 +/- 102 as compared with .114 +/- .085 in sham-operated controls, P less than 0.005). Animals treated with anti-TNF antiserum prior to the induction of hepatic ischemia had a significantly reduced pulmonary capillary leak compared to animals pretreated with rabbit serum without TNF-blocking properties (.184 +/- .029 versus .694 +/- 052 for the control serum, P less than 0.005). TNF release follows both moderate and severe ischemic injury to the liver and the results reported here implicate TNF as an important mediator of increased pulmonary capillary permeability. These experiments confirm previous histologic studies that demonstrated pulmonary edema and intra-alveolar hemorrhage following hepatic ischemia/reperfusion, with subsequent blockade of the histologic injury by pretreatment with anti-TNF antiserum.

Pediatric surgery and urology:long term outcomes

Part I: Introduction. Introduction. U.S. Historical Perspective. European Historical Perspective. Outcome in the Pediatric Surgical Patient. Long-Term Psychological Impact of Pediatric Surgical Interventions. Principles of Outcomes Analysis. Neonatal Intensive Care: Perinatal Mortality and Morbidity. Part II: Head and Neck. Cleft-Lip and Cleft Palate. Lymphangiomas. Thyroid and Parathyroid Disorder. Salivary Disease. Head and Neck Tumours. Part III: Thorax. Chest Wall Defects. Congenital Diaphragmic Hernia. Surgically Managed Pediatric Airway Obstruction. Pulmonary Resection and Thoracotomy. Esophageal Atresia: Surgical Aspects. Esophageal Atresia: Nutrition, Growth & Respiratory Function. Anti-Reflux Surgery. Esophageal Replacement and Corrosive Injury. Esophageal Achalasia. Part IV: Abdomen. Abdominal Surgery (Gastrointestinal Resections/Stomas & Adhesions). Short Bowel. Abdominal Wall Defects. Inguinal Hernia Repair. Infantile Hypertrophic Pyloric Stenosis. Small Bowel Disorders. Cystic Fibrosis: Surgical Considerations. Necrotizing Enterocilitis. Inflammatory Bowel Disease. Appendicitis. Hirshprungs Disease: Duhamel Pull-Through. Hirshprungs Disease: Endorectal Pull-Through. Hirshprungs Disease: Primary Pull-Through. Anorectal Malformations: Overview. Experience with the Posterior Sagittal Approach. Psychological Aspects. Splenectomy (Including Trauma). Biliary Atresia. Choledochal Cysts. Portal Hypertension. Sclerotherapy. Portosystemic Shunts. Benign Pancreatic Disorders. Biliary Stones. Liver and Biliary Trauma. Part V: Urology. Introduction. Antenatal Uropathies. Posterior Urethral Valves. Upper Tract Dilatation. Vesicoureteral Reflux: Endoscopic. Surgical. Conservative. Bladder Exstrophy. Genitoplasty in Genital Ambiguities. Hypospadias. Genitoplasty in Extrophy Patients. Neuropathic Bladder: Surgery for Incontinence. Augmentation and Substitution Cystoplasty. Gastrocystoplasty. Undescended Testes. Circumcision. The Single Kidney. Multicystic Kidney. Part VI: Oncology. Late Effects of Chemo

Neutrophil-dependent, oxygen-radical mediated lung injury associated with acute pancreatitis.

Cerulein-induced acute pancreatitis in rats is associated with a reversible lung injury that is characterized by alveolar capillary endothelial-cell injury, increased microvascular permeability, interstitial edema formation, and intraalveolar hemorrhage and fibrin deposition. The role of mediators in this injury was analyzed using gravimetric data, microvascular permeability indices, electron microscopy, and a quantitative morphometric analysis. Neutrophil depletion induced by a specific antibody was highly protective against lung injury. Interruption of the complement pathway (using low dose Naja naja cobra venom factor) also protected against lung injury. Catalase and superoxide dismutase were also protective. The iron chelator deferoxamine and the hydroxyl radical scavenger, dimethylsulfoxide, were not protective against acute lung injury. These data suggest that complement, neutrophils, and neutrophil-derived (H2O2-dependent) oxygen products mediate lung injury that occurs secondary to cerulein-induced pancreatitis. In contrast to other models of neutrophil-dependent, oxygen-radical-mediated lung injury, this lung injury does not appear to be an iron-dependent and hydroxyl-radical mediated injury. We postulate that the process of acute pancreatitis leads to complement activation followed by neutrophil recruitment, sequestration, and adherence to alveolar capillary endothelial cells. Ultimately lung injury appears to result from local endothelial-cell injury secondary to neutrophil-generated oxygen products that may be myeloperoxidase dependent.

L-4F, an Apolipoprotein A-1 Mimetic, Dramatically Improves Vasodilation in Hypercholesterolemia and Sickle Cell Disease

Background—Hypercholesterolemia and sickle cell disease (SCD) impair endothelium-dependent vasodilation by dissimilar mechanisms. Hypercholesterolemia impairs vasodilation by a low-density lipoprotein (LDL)–dependent mechanism. SCD has been characterized as a chronic state of inflammation in which xanthine oxidase (XO) from ischemic tissues increases vascular superoxide anion (O2·−) generation. Recent reports indicate that apolipoprotein (apo) A-1 mimetics inhibit atherosclerosis in LDL receptor–null (Ldlr−/−) mice fed Western diets. Here we hypothesize that L-4F, an apoA-1 mimetic, preserves vasodilation in hypercholesterolemia and SCD by decreasing mechanisms that increase O2·− generation. Methods and Results—Arterioles were isolated from hypercholesterolemic Ldlr−/− mice and from SCD mice that were treated with either saline or L-4F (1 mg/kg per day). Vasodilation in response to acetylcholine was determined by videomicroscopy. Effects of L-4F on LDL-induced increases in endothelium-dependent O2·− generation were determined on arterial segments via the hydroethidine assay and on stimulated endothelial cell cultures via superoxide dismutase–inhibitable ferricytochrome c reduction. Effects of L-4F on XO bound to pulmonary arterioles and content in livers of SCD mice were determined by immunofluorescence. Hypercholesterolemia impaired vasodilation in Ldlr−/− mice, which L-4F dramatically improved. L-4F inhibited LDL-induced increases in O2·− in arterial segments and in stimulated cultures. SCD impaired vasodilation, increased XO bound to pulmonary endothelium, and decreased liver XO content. L-4F dramatically improved vasodilation, decreased XO bound to pulmonary endothelium, and increased liver XO content compared with levels in untreated SCD mice. Conclusions—These data show that L-4F protects endothelium-dependent vasodilation in hypercholesterolemia and SCD. Our findings suggest that L-4F restores vascular endothelial function in diverse models of disease and may be applicable to treating a variety of vascular diseases.

Surgical repair of pectus excavatum markedly improves body image and perceived ability for physical activity: multicenter study.

OBJECTIVE. This study evaluated changes in both physical and psychosocial quality of life reported by the parent and child after surgical repair of pectus excavatum. METHODS. As part of a multicenter study of pectus excavatum, a previously validated tool called the Pectus Excavatum Evaluation Questionnaire was administered by the research coordinator, via telephone, to parents and patients (8–21 years of age) before and 1 year after surgery. Eleven North American childrens hospitals participated. From 2001 to 2006, 264 patients and 291 parents completed the initial questionnaire, and 247 patients and 274 parents completed the postoperative questionnaire. Responses used a Likert-type scale of 1 to 4, reflecting the extent or frequency of a particular experience, with higher values conveying less-desirable experience. RESULTS. Preoperative psychosocial functioning was unrelated to objective pectus excavatum severity (computed tomographic index). Patients and their parents reported significant positive postoperative changes. Improvements occurred in both physical and psychosocial functioning, including less social self-consciousness and a more-favorable body image. For children, the body image component improved from 2.30 ± 0.62 (mean ± SD) to 1.40 ± 0.42 after surgery and the physical difficulties component improved from 2.11 ± 0.82 to 1.37 ± 0.44. For the parent questionnaire, the childs emotional difficulties improved from 1.81 ± 0.70 to 1.24 ± 0.36, social self-consciousness improved from 2.86 ± 1.03 to 1.33 ± 0.68, and physical difficulties improved from 2.14 ± 0.75 to 1.32 ± 0.39. Ninety-seven percent of patients thought that surgery improved how their chest looked. CONCLUSIONS. Surgical repair of pectus excavatum can significantly improve the body image difficulties and limitations on physical activity experienced by patients. These results should prompt physicians to consider the physiologic and psychological implications of pectus excavatum just as they would any other physical deformity known to have such consequences.

Reversal of mortality for congenital diaphragmatic hernia with ECMO.

Extracorporeal Membrane Oxygenation (ECMO) has been available to neonates with respiratory failure at the University of Michigan School of Medicine since June 1981. In order to evaluate the impact of this type of pulmonary support, a retrospective analysis of 50 neonates with posterolateral congenital diaphragmatic hernia (CDH) who were symptomatic during the first hour of life and were treated between June 1974 and December 1987 was carried out. The patients were divided into two groups, those treated before June 1981 (16 patients) and those treated after June 1981 (34 patients). Overall survival improved from 50% (eight of 16 patients) during the pre-ECMO era to 76% (26 of 34 patients) during the post-ECMO period (p = 0.06). During the period after June 1981, 21 neonates were unresponsive to conventional therapy and were therefore considered for ECMO. Failure of conventional therapy was defined as acute clinical deterioration with an expected mortality of greater than 80% based on an objective formula previously reported. Six patients were excluded on the basis of specific contraindications to ECMO. Thirteen of 15 infants (87%) supported with ECMO survived. Three patients treated before 1981 met criteria for ECMO; all three died while receiving treatment using conventional therapy. These survival differences are significant (p less than 0.01). In addition, the survival of 87% for the infants treated with ECMO versus the expected mortality of greater than 80% for these same patients when treated with conventional therapy is highly significant (p less than 0.005). Based on this data, ECMO appears to be a successful, reliable, and safe method of respiratory support for selected, critically ill infants with CDH.

Comparative proteomic analysis of PAI-1 and TNF-alpha-derived endothelial microparticles

Endothelium‐derived microparticles (EMPs) are small vesicles released from endothelial cells in response to cell injury, apoptosis, or activation. Elevated concentrations of EMPs have been associated with many inflammatory and vascular diseases. EMPs also mediate long range signaling and alter downstream cell function. Unfortunately, the molecular and cellular basis of microparticle production and downstream cell function is poorly understood. We hypothesize that EMPs generated by different agonists will produce distinct populations of EMPs with unique protein compositions. To test this hypothesis, different EMP populations were generated from human umbilical vein endothelial cells by stimulation with plasminogen activator inhibitor type 1 (PAI‐1) or tumor necrosis factor‐alpha (TNF‐α) and subjected to proteomic analysis by LC/MS. We identified 432 common proteins in all EMP populations studied. Also identified were 231 proteins unique to control EMPs, 104 proteins unique to PAI‐1 EMPs and 70 proteins unique to TNF‐α EMPs. Interestingly, variations in protein abundance were found among many of the common EMP proteins, suggesting that differences exist between EMPs on a relative scale. Finally, gene ontology (GO) and KEGG pathway analysis revealed many functional similarities and few differences between the EMP populations studied. In summary, our results clearly indicate that EMPs generated by PAI‐1 and TNF‐α produce EMPs with overlapping but distinct protein compositions. These observations provide fundamental insight into the mechanisms regulating the production of these particles and their physiological role in numerous diseases.

L-4F, an Apolipoprotein A-1 Mimetic, Restores Nitric Oxide and Superoxide Anion Balance in Low-Density Lipoprotein-Treated Endothelial Cells

Background—Low-density lipoprotein (LDL) impairs endothelial cell function by uncoupling endothelial nitric oxide synthase (eNOS) activity, which allows superoxide anion (O2·−) to be generated rather than nitric oxide (·NO). Recent reports indicate that apolipoprotein (apo) A-1 mimetics inhibit the development of atherosclerotic lesions in LDL receptor-null mice. Here we hypothesize that L-4F, an apoA-1 mimetic that inhibits atherosclerosis induced by hypercholesterolemia, protects endothelial cell function by preventing LDL from uncoupling eNOS activity. Methods and Results—Bovine aortic endothelial cells were incubated with LDL±L-4F, and changes in A23187-stimulated ·NO and O2·− generation were determined by ozone chemiluminescence and superoxide dismutase-inhibitable ferricytochrome c reduction, respectively. Western analysis of eNOS immunoprecipitates was used to determine effects of LDL and L-4F on heat shock protein 90 (hsp90) interactions with eNOS. LDL decreased ·NO production and increased eNOS-dependent O2·− generation. Pretreatment of LDL with L-4F increased ·NO and decreased O2·− generation. By itself, L-4F had no effect on O2·− but did increase ·NO generation. Stimulation of endothelial cells incubated with LDL decreased the association of hsp90 with eNOS. Pretreatment of LDL with L-4F prevented a decrease in hsp90 association with eNOS and often enhanced association on stimulation. Conclusions—These data demonstrate that L-4F protects endothelial cell function by preventing LDL from uncoupling eNOS activity. L-4F allows endothelial cell to maintain coupled eNOS activity to generate ·NO even in the face of atherogenic concentrations of LDL.

Pediatric American College of Surgeons National Surgical Quality Improvement Program: feasibility of a novel, prospective assessment of surgical outcomes

PURPOSE The American College of Surgeons (ACS) National Surgical Quality Improvement Program (NSQIP) provides validated assessment of surgical outcomes. This study reports initiation of an ACS NSQIP Pediatric at 4 childrens hospitals. METHODS From October 2008 to June 2009, 121 data variables were prospectively collected for 3315 patients, including 30-day outcomes and tailoring the ACS NSQIP methodology to childrens surgical specialties. RESULTS Three hundred seven postoperative complications/occurrences were detected in 231 patients representing 7.0% of the study population. Of the patients with complications, 175 (75.7%) had 1, 39 (16.9%) had 2, and 17 (7.4%) had 3 or more complications. There were 13 deaths (0.39%) and 14 intraoperative occurrences (0.42%) detected. The most common complications were infection, 105 (34%) (SSI, 54; sepsis, 31; pneumonia, 13; urinary tract infection, 7); airway/respiratory events, 27 (9%); wound disruption, 18 (6%); neurologic events, 8 (3%) (nerve injury, 4; stroke/vascular event, 2; hemorrhage, 2); deep vein thrombosis, 3 (<1%); renal failure, 3 (<1%); and cardiac events, 3 (<1%). Current sampling captures 17.5% of cases across institutions with unadjusted complication rates ranging from 6.8% to 10.2%. Completeness of data collection for all variables exceeded 95% with 98% interrater reliability and 87% of patients having full 30-day follow-up. CONCLUSION These data represent the first multiinstitutional prospective assessment of specialty-specific surgical outcomes in children. The ACS NSQIP Pediatric is poised for institutional expansion and future development of risk-adjusted models.