Safety of the etonogestrel-releasing implant during the immediate postpartum period: a pilot study

Background: The effects of etonogestrel (ETG)-releasing contraceptive implant during the immediate postpartum period on maternal safety are unknown. Study design: Forty healthy women exclusively breastfeeding were randomized to receive either ETG-releasing implant 24-48 h after delivery (n=20) or depot medroxyprogesterone acetate (DMPA group; n=20) at the sixth week postpartum. We measured blood pressure, maternal and neonatal weight, body mass index (BMI; kg/m(2)), waist circumference (WC), complete blood count, C-reactive protein, interleukin-6, tumor necrosis factor (TNF-alpha), lipid profile, fasting serum glucose and maintenance of exclusive lactation up to the 12th week postpartum. Results: Decreases in mean maternal weight, BMI (kg/m(2)) and WC were significantly greater in the ETG-releasing implant group than in the MPA group during the first 6 weeks postpartum (-4.64 +/- 2.71 kg vs. -2.6 +/- 2.45 kg mean +/- SD, p=.017; -1.77 +/- 1.06 kg/m(2) vs. -0.97 +/- 0.95 kg/m(2), p=.026; -15.3 +/- 6.72 cm vs. -9.05 +/- 5.84 cm, p=.003, respectively). In addition, total cholesterol and HDL, were lower in DMPA users, and TNF-alpha and leukocytes were higher in DMPA users compared to in the implant group, between 6 and 12 weeks after delivery. The newborns of implant users showed a trend towards gaining more weight, as compared with the infants of the DMPA mothers during the first 6 weeks of life (implant group: +1460.50 +/- 621.34 g vs. DMPA group: +1035.0 +/- 562.43 g, p=.05). The remaining variables, including the duration of exclusive breastfeeding, were similar between the groups. Conclusion: The insertion of ETG-releasing contraceptive implant during the immediate postpartum period was not associated with deleterious maternal clinical effects or with significant maternal metabolic alterations or decreased infant weight gain. (C) 2009 Elsevier Inc. All rights reserved.

Treatment with cannabidiol reverses oxidative stress parameters, cognitive impairment and mortality in rats submitted to sepsis by cecal ligation and puncture

Oxidative stress plays an important role in the development of cognitive impairment in sepsis. Here we assess the effects of acute and extended administration of cannabidiol (CBD) on oxidative stress parameters in peripheral organs and in the brain, cognitive impairment, and mortality in rats submitted to sepsis by cecal ligation and perforation (CLP). To this aim, male Wistar rats underwent either sham operation or CLP. Rats subjected to CLP were treated by intraperitoneal injection with ""basic support"" and CBD (at 2.5, 5, or 10 mg/kg once or daily for 9 days after CLP) or vehicle. Six hours after CLP (early times), the rats were killed and samples from lung, liver, kidney, heart, spleen, and brain (hippocampus, striatum, and cortex) were obtained and assayed for thiobarbituric acid reactive species (TBARS) formation and protein carbonyls. On the 10th day (late times), the rats were submitted to the inhibitory avoidance task. After the test, the animals were killed and samples from lung, liver, kidney, heart, spleen, and brain (hippocampus) were obtained and assayed for TBARS formation and protein carbonyls. The acute and extended administration of CBD at different doses reduced TBARS and carbonyl levels in some organs and had no effects in others, ameliorated cognitive impairment, and significantly reduced mortality in rats submitted to CLP. Our data provide the first experimental demonstration that CBD reduces the consequences of sepsis induced by CLP in rats, by decreasing oxidative stress in peripheral organs and in the brain, improving impaired cognitive function, and decreasing mortality. (C) 2010 Elsevier B.V. All rights reserved.

Effect of BCG stimulus on proinflammatory cytokine production in laryngeal cancer

Background Evaluate the production of TNF and IL-6 in the supernatant of peripheral blood mononuclear cell (PBMC) cultures of patients with supraglottic laryngeal cancer before and after surgical treatment. Materials and methods Adherent cell cultures were stimulated with LPS and BCG. Fourteen patients with advanced supraglottic laryngeal cancer were studied. Cytokine concentration was determined by ELISA in supernatants of mononuclear cell cultures. Results In non-stimulated cultures, lower TNF cytokine levels were detected during the late postoperative (LP) period compared to control (P = 0.02). LP TNF and IL-6 levels were high in cultures stimulated with LPS compared with the preoperative period (PREOP) (P = 0.007; P = 0.008, respectively). Stimulation with BCG led to increased levels of TNF and IL-6 during the LP period compared to control (P = 0.001; P = 0.04, respectively). Conclusions BCG is able to modulate the immune response of patients with advanced supraglottic laryngeal cancer in vitro, increasing the secretion of TNF and IL-6 by macrophages during the postoperative period.

Risk stratification in neonates and infants submitted to cardiac surgery with cardiopulmonary bypass: A multimarker approach combining inflammatory mediators, N-terminal pro-B-type natriuretic peptide and troponin I

Low cardiac output syndrome (LCOS) is a common problem following cardiac surgery with cardiopulmonary bypass (CPB) in neonates and infants, and its early recognition remains a challenging task. We aimed to test whether a multimarker approach combining inflammatory and cardiac markers provides complementary information for prediction of LCOS and death in children submitted to cardiac surgery with CPB. Forty-six children younger than 18 months with congenital heart defects were prospectively enrolled. No intervention was made. Blood samples were collected pre-operatively, during CPB and post-operatively (PO) for measurement of interleukin (IL)-6, IL-8, IL-10, tumor necrosis factor (TNF)-alpha, cardiac troponin I (cTnI) and N-terminal pro-B-type natriuretic peptide (NT-proBNP). Clinical data and outcome variables were recorded. Logistic regression was used to identify predictors of LCOS and death. Multivariate logistic regression identified pre-operative NT-proBNP and IL-8 4 h PO as independent predictors of LCOS, while cTnI 4 h PO and CPB length were independent predictors of death. The use of inflammatory and cardiac markers in combination improved sensitivity, negative predictive value and accuracy of the models. In conclusion, the combined assessment of inflammatory and cardiac biochemical markers can be useful for identifying young children at increased risk for LCOS and death after heart surgery with CPB. (C) 2008 Elsevier Ltd. All rights reserved.

Effects of Caffeoylquinic Acid Derivatives and C-Flavonoid from Lychnophora ericoides on in vitro Inflammatory Mediator Production

In this study we aimed at evaluating the effect of the major polar constituents of the medicinal plant Lychnophora ericoides on the production of inflammatory mediators produced by LPS-stimulated U-937 cells. The 6,8-di-C-beta-glucosylapigenin (vicenin-2) presented no effect on tumor necrosis factor (TNF)-alpha production, but inhibited, in a dose-dependent manner, the production of prostaglandin (PG) E(2) without altering the expression of cyclooxygenase (COX) -2 protein. 3,5-Dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid, at lower concentrations, had small but significant effects on reducing PG E, levels; at higher doses these compounds stimulated PGE(2) and also TNF-alpha production by the cells. All the caffeoylquinic acid derivatives, in a dose-dependent fashion, were able to inhibit monocyte chemoattractant protein-3 synthesis/release, with 4,5-DCQ being the most potent at the highest tested concentration. These results add important information on the effects of plant natural polyphenols, namely vicenin-2 and caffeoylquinic acid derivatives, on the production of inflammatory mediators by cultured cells.

Clinical and Immunological Insights on Severe, Adverse Neurotropic and Viscerotropic Disease following 17D Yellow Fever Vaccination

Yellow fever (YF) vaccines (17D-204 and 17DD) are well tolerated and cause very low rates of severe adverse events (YEL-SAE), such as serious allergic reactions, neurotropic adverse diseases (YEL-AND), and viscerotropic diseases (YEL-AVD). Viral and host factors have been postulated to explain the basis of YEL-SAE. However, the mechanisms underlying the occurrence of YEL-SAE remain unknown. The present report provides a detailed immunological analysis of a 23-year-old female patient. The patient developed a suspected case of severe YEL-AVD with encephalitis, as well as with pancreatitis and myositis, following receipt of a 17D-204 YF vaccination. The patient exhibited a decreased level of expression of Fc-gamma R in monocytes (CD16, CD32, and CD64), along with increased levels of NK T cells (an increased CD3(+) CD16(+/-) CD56(+/-)/CD3(+) ratio), activated T cells (CD4(+) and CD8(+) cells), and B lymphocytes. Enhanced levels of plasmatic cytokines (interleukin-6 [IL-6], IL-17, IL-4, IL-5, and IL-10) as well as an exacerbated ex vivo intracytoplasmic cytokine pattern, mainly observed within NK cells (gamma interferon positive [IFN-gamma(+)], tumor necrosis factor alpha positive [TNF-alpha(+)], and IL-4 positive [IL-4(+)]), CD8(+) T cells (IL-4(+) and IL-5(+)), and B lymphocytes (TNF-alpha(+), IL-4(+), and IL-10(+)). The analysis of CD4(+) T cells revealed a complex profile that consisted of an increased frequency of IL-12(+) and IFN-gamma(+) cells and a decreased percentage of TNF-alpha(+), IL-4(+), and IL-5+ cells. Depressed cytokine synthesis was observed in monocytes (TNF-alpha(+)) following the provision of antigenic stimuli in vitro. These results support the hypothesis that a strong adaptive response and abnormalities in the innate immune system may be involved in the establishment of YEL-AND and YEL-AVD.

Inflammation and Overweight in Peritoneal Dialysis: Is There an Association?

More than 30% of the patients on peritoneal dialysis show chronic systemic inflammatory activity with high levels of C-reactive protein. The purpose of this cross-sectional study was to investigate the influence of the inflammatory state on clinical and nutritional markers in patients on peritoneal dialysis. Twenty-seven patients were included: mean age was 57.6 +/- 19 years, 48% were male, and median time on peritoneal dialysis was 16.0 (8.3; 35.8) months. Clinical, dialytic, laboratory, anthropometric and electric bioimpedance data were collected with the sample stratified for C-reactive protein. In patients, the levels of Interleukin-6 and tumor necrosis factor-a were higher, while adiponectin levels were lower than in healthy individuals (p <= 0.001), indicating the presence of inflammatory activity in the sample. When compared to patients with C-reactive protein < 1 mg/dL, those with = 1mg/dL showed higher body mass index (29.4 +/- 6.1 vs. 24.4 +/- 4.5 kg/m(2); p = 0.009), percent of standard body weight (124.5 +/- 25.4 vs. 106.8 +/- 17.9 %; p = 0.012), and percent of body fat as assessed by both anthropometry (31.3 +/- 9.9 vs. 23.9 +/- 9.1%; p = 0.056) and bioimpedance (38.9 +/- 6.3 vs. 26.2 +/- 12.6 %; p < 0.001). Patients with C-reactive protein = 1mg/dL also exhibited higher levels of ferritin (701 +/- 568 vs. 532 +/- 356 ng/mL; p = 0.054) and lower total lymphocyte count (median 1838 vs. 1638 mm(3); p = 0.001). In conclusion, higher body mass index and body fat markers were associated with C-reactive protein = 1mg/dL, and higher C-reactive protein was associated with immunocompetence impairment evidenced by the lower total lymphocyte count. Our findings confirm the relationship between inflammation, body fat, and immunocompetence, which may be superimposed potentializing the inflammatory status.

Inhibition of COX 1 and 2 prior to Renal Ischemia/Reperfusion Injury Decreases the Development of Fibrosis

Ischemia and reperfusion injury (IRI) contributes to the development of chronic interstitial fibrosis/tubular atrophy in renal allograft patients, Cyclooxygenase (COX) 1 and 2 actively participate in acute ischemic injury by activating endothelial cells and inducing oxidative stress. Furthermore, blockade of COX I and 2 has been associated with organ improvement after ischemic damage. The aim of this study was to evaluate the role of COX I and 2 in the development of fibrosis by performing a COX I and 2 blockade immediately before IRI We subjected C57BI/6 male mice to 60 min of unilateral renal pedicle occlusion, Prior to surgery mice were either treated with indomethacin (IMT) at days -1 and 0 or were untreated. Blood and kidney samples were collected 6 wks after IRI. Kidney samples were analyzed by real-time reverse transcription-poly me rase chain reaction for expression of transforming growth factor beta (TGF-beta), monocyte chemoattractant protein 1 (MCP-1), osteopontin (OPN), tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-1 beta, IL-10, heme oxygenose 1 (HO-1), vimentin, connective-tissue growth factor (CTGF), collagen 1, and bone morphogenic protein 7 (BMP-7), To assess tissue fibrosis we performed morphometric analyses and Sirius red staining. We also performed immunohistochemical analysis of anti-actin smooth muscle, Renal function did not significantly differ between groups. Animals pretreated with IMT showed significantly less interstitial fibrosis than nontreated animals. Gene transcript analyses showed decreased expression of TGF-beta, MCP-1,TNF-alpha, IL-1-beta, vimentin, collagen 1, CTGF and IL-10 mRNA (all P < 0.05), Moreover, HO-I mRNA was increased in animals pretreated with IMT (P < 0.05) Conversely, IMT treatment decreased osteopontin expression and enhanced BMP-7 expression, although these levels did rot reach statistical significance when compared with control expression levels, I he blockade of COX 1 and 2 resulted in less tissue fibrosis, which was associated with a decrease in proinflammatory cytokines and enhancement of the protective cellular response.

The Role of Toll-Like Receptor 2 in the Recognition of Aggregatibacter actinomycetemcomitans

Background: Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans) is a Gram-negative bacterium present in the oral cavity and is usually associated with localized aggressive periodontitis. Isolated antigens from A. actinomycetemcomitans can activate innate immune cells through Toll-like receptors (TLRs), which are molecules that recognize structural components conserved among microorganisms. In this study, we evaluate the role of TLR2 in the recognition of A. actinomycetemcomitans. Methods: Macrophages and neutrophils from knockout mice with targeted disruption of TLR2 (TLR2(-/-) mice) and wild-type mice were collected and used for the subsequent assays. The production of cytokines and chemokines was evaluated by enzyme-linked immunosorbent assay (ELISA), and the presence of apoptotic cells was determined by flow cytometry. In addition, the mechanisms that modulate the outcome of A. actinomycetemcomitans-induced periodontal disease in TLR2(-/-) mice were examined. Results: The results show that TLR2-deficient mice developed more severe periodontitis after A. actinomycetemcomitans infection, characterized by significantly higher bone loss and inflammatory cell migration to periodontal tissues. The inflammatory cell influx into the peritoneal cavities of TLR2(-/-) mice was three-fold lower than that observed for the littermate controls. A significantly diminished production of the cytokines tumor necrosis factor-alpha and interleukin-1 beta as well as the chemokine CC-ligand-5 in the peritoneal cavities of TLR2(-/-) mice was observed. In addition, a high frequency of apoptotic cells in the inflammatory exudates from TLR2(-/-) mice was observed. Phagocytosis and nitric oxide production was diminished in cells from TLR2(-/-) mice, facilitating the dissemination of the pathogen to the spleen. Conclusion: The results of this study highlight the involvement of TLR2 in recognizing A. actinomycetemcomitans and its essential role in controlling A. actinomycetemcomitans infection. J Periodontot 2009,80:2070-2019.

The Potential Role of Suppressors of Cytokine Signaling in the Attenuation of Inflammatory Reaction and Alveolar Bone Loss Associated with Apical Periodontitis

Inflammatory cytokines contribute to periapical tissue destruction. Their activity is potentially regulated by suppressors of cytokine signaling (SOCS), which down-regulate signal transduction as part of an inhibitory feedback loop. We investigated the expression of the cytokines tumor necrosis factor alpha (TNF-alpha); interleukin (IL)-10 and RANKL; and SOCS-1, -2, and -3 by real-time polymerase chain reaction in 57 periapical granulomas and 38 healthy periapical tissues. Periapical granulomas exhibited significantly higher SOCS-1, -2, and -3, TNF-alpha, IL-10, and RANKL messenger RNA levels when compared with healthy controls. Significant positive correlations were found between SOCS1 and IL-10 and between SOCS3 and IL-10. Significant inverse correlations were observed between SOCS1 and TNF-alpha, SOCS1 and RANKL, and SOCS3 and TNF-alpha. Increased SOCS-1, -2, and -3 messenger RNA levels in periapical granulomas may be related to the downregulation of inflammatory cytokines in these lesions; therefore, SOCS molecules may play a role in the dynamics of periapical granulomas development. (J Endod 2008;34:1480-1484)

Nitric oxide and tissue destruction

Nitric oxide (NO) is a free radical which has complex roles in both health and disease. It is now recognized that NO is essential for a vast spectrum of intracellular and extracellular events in a wide variety of tissues. NO has also been implicated in the pathogenesis of numerous inflammatory and autoimmune diseases. In this review we consider the roles of NO generally and in particular the implications for periodontal diseases.

Influence of Antiplatelet Drugs in the Pathogenesis of Experimental Periodontitis and Periodontal Repair in Rats

Background: Platelets contain an array of biologic mediators that can modulate inflammation and repair processes including proinflammatory mediators and growth factors. Previous studies have shown that periodontitis and periodontal repair are associated with platelet activation. We hypothesized that drug-induced platelet inactivation may interfere in the processes of inflammation and repair in experimental periodontitis in rats by suppressing the release of biologic mediators from platelets to the site of injury. Methods: To measure the effects on periodontitis, ligatures were placed around first molars, and aspirin (Asp, 30 mg/kg) or clopidogrel (Clo, 75 mg/kg) was given intragastrically once daily for 15 days. Interleukin-6 (IL-6), tumor necrosis factor-a (TNF-alpha), and thromboxane A(2) levels were measured by enzyme-linked immunosorbent assay. To evaluate the effects of antiplatelet drugs on periodontal repair, ligatures were removed after 15 days of periodontitis induction, and Asp or Clo were administered beginning the following day for 15 days. Periodontal repair was assessed by microcomputed tomography. Results: On periodontitis phase, Asp and Clo significantly reduced levels of TNF-alpha and II-6 (P < 0.05), but only Asp decreased thromboxane A(2) (P < 0.05). Asp and Clo decreased inflammatory infiltration; however, this reduction was more pronounced with Clo treatment (P < 0.05). Histometric analysis showed that Asp and Clo impaired alveolar bone resorption. During the repair phase and after removal of the ligatures, microcomputed tomography analysis demonstrated that treatment with Asp and Clo did not impair alveolar bone repair. Conclusion: Systemic administration of Asp and Clo attenuates the inflammation associated with periodontitis without affecting the repair process when stimulus is removed. J Periodontol 2011;82:767-777.

Neonatal endotoxin exposure changes neuroendocrine, cardiovascular function and mortality during polymicrobial sepsis in adult rats

Our aim was to investigate whether neonatal LPS challenge may improve hormonal, cardiovascular response and mortality, this being a beneficial adaptation when adult rats are submitted to polymicrobial sepsis by cecal ligation and puncture (CLP). Fourteen days after birth, pups received an intraperitoneal injection of lipopolysaccharide (LPS; 100 mu g/kg) or saline. After 8-12 weeks, they were submitted to CLP, decapitated 4,6 or 24 h after surgery and blood was collected for vasopressin (AVP), corticosterone and nitrate measurement, while AVP contents were measured in neurohypophysis, supra-optic (SON) and paraventricular (PVN) nuclei. Moreover, rats had their mean arterial pressure (MAP) and heart rate (HR) evaluated, and mortality and bacteremia were determined at 24 h. Septic animals with neonatal LPS exposure had higher plasma AVP and corticosterone levels, and higher c-Fos expression in SON and PVN at 24 h after surgery when compared to saline treated rats. The LPS pretreated group showed increased AVP content in SON and PVN at 6 h, while we did not observe any change in neurohypophyseal AVP content. The nitrate levels were significantly reduced in plasma at 6 and 24 h after surgery, and in both hypothalamic nuclei only at 6 h. Septic animals with neonatal LPS exposure showed increase in MAP during the initial phase of sepsis, but HR was not different from the neonatal saline group. Furthermore, neonatally LPS exposed rats showed a significant decrease in mortality rate as well as in bacteremia. These data suggest that neonatal LPS challenge is able to promote beneficial effects on neuroendocrine and cardiovascular responses to polymicrobial sepsis in adulthood. (C) 2011 Elsevier B.V. All rights reserved.

Role of circumventricular organs in pro-inflammatory cytokine-induced activation of the hypothalamic-pituitary-adrenal axis

1. The past 15 years has seen the emergence of a new field of neuroscience research based primarily on how the immune system and the central nervous system can interact. A notable example of this interaction occurs when peripheral inflammation, infection or tissue injury activates the hypothalamic- pituitary-adrenal axis (HPA). 2. During such assaults, immune cells release the pro- inflammatory cytokines interleukin (IL)-1, IL-6 and tumour necrosis factor-alpha into the general circulation. 3. These cytokines are believed to act as mediators for HPA axis activation. However, physical limitations of cytokines impede their movement across the blood-brain barrier and, consequently, it has been unclear as to precisely how and where IL-1beta signals cross into the brain to trigger HPA axis activation. 4. Evidence from recent anatomical and functional studies suggests two neuronal networks may be involved in triggering HPA axis activity in response to circulating cytokines. These are catecholamine cells of the medulla oblongata and the circumventricular organs (CVO). 5. The present paper examines the role of CVO in generating HPA axis responses to pro-inflammatory cytokines and culminates with a proposed model based on cytokine signalling primarily involving the area postrema and catecholamine cells in the ventrolateral and dorsal medulla.

CD40 and CD86 upregulation with divergent CMRF44 expression on blood dendritic cells in inflammatory bowel diseases

OBJECTIVE: Dendritic cells (DC) are the only antigen-presenting cells that can activate naive T lymphocytes and initiate a primary immune response. They are also thought to have a role in immune tolerance. DC traffic from the blood to peripheral tissue where they become activated. They then present antigen and the costimulating signals necessary to initiate an immune response. In this study, we investigated the number, subsets, and activation pattern of circulating and intestinal DC from patients with clinically mild ulcerative colitis (UC) or Crohn's disease. METHODS: Patients were recruited, if they were not taking immunosuppressive therapy, and were assessed for clinical severity of their disease using for UC, the Clinical Activity Index, and for Crohn's disease, the Crohn's Disease Activity Index. Blood CD11c(+) and CD11c(-) DC subsets, expression of costimulatory antigens, CD86 and CD40, and the early differentiation/activation antigen, CMRF44, were enumerated by multicolor flow cytometry of lineage negative (lin(-) = CD3(-), CD19(-), CD14(-), CD16(-)) HLA-DR+ DC. These data were compared with age-matched healthy and the disease control groups of chronic noninflammatory GI diseases (cGI), acute noninflammatory GI diseases (aGI), and chronic non-GI inflammation (non-GI). In addition, cryostat sections of colonoscopic biopsies from healthy control patients and inflamed versus noninflamed gut mucosa of inflammatory bowel disease (IBD) patients were examined for CD86(+) and CD40(+)lin(-) cells. RESULTS: Twenty-one Crohn's disease and 25 UC patients, with mean Crohn's Disease Activity Index of 98 and Clinical Activity Index of 3.1, and 56 healthy controls, five cGI, five aGI, and six non-GI were studied. CD11c(+) and CD11c(-) DC subsets did not differ significantly between Crohn's, UC, and healthy control groups. Expression of CD86 and CD40 on freshly isolated blood DC from Crohn's patients appeared higher (16.6%, 31%) and was significantly higher in UC (26.6%, 46.3%) versus healthy controls (5.5%, 25%) (p = 0.004, p = 0.012) and non-GI controls (10.2%, 22.8%) (p = 0.012, p = 0.008), but not versus cGI or aGI controls. CD86(+) and CD40(+) DC were also present in inflamed colonic and ileal mucosa from UC and Crohn's patients but not in noninflamed IBD mucosa or normal mucosa. Expression of the CMRF44 antigen was low on freshly isolated DC, but it was upregulated after 24-h culture on DC from all groups, although significantly less so on DC from UC versus Crohn's or healthy controls (p = 0.024). The CMRF44(+) antigen was mainly associated with CD11c(+) DC, and in UC was inversely related to the Clinical Activity Index (r = -0.69, p = 0.0002). CONCLUSIONS: There is upregulation of costimulatory molecules on blood DC even in very mild IBD but surprisingly, there is divergent expression of the differentiation/activation CMRF44 antigen. Upregulation of costimulatory molecules and divergent expression of CMRF44 in blood DC was also apparent in cGI and aGI but not in non-GI or healthy controls, whereas intestinal CD86(+) and CD40(+) DC were found only in inflamed mucosa from IBD patients. Persistent or distorted activation of blood DC or divergent regulation of costimulatory and activation antigens may have important implications for gut mucosal immunity and inflammation. (Am J Gastroenterol 2001;96:2946-2956. (C) 2001 by Am. Coll. of Gastroenterology).