Changes in serum and intestinal diamine oxidase (DAO) activity after proximal enterectomy in rats

To determine whether serum and mucosal DAO activity reflects quantitative changes in the small bowel mucosal mass, we have chosen an experimental model of mucosal hyperplasia which is known to occur in the rat after enterectomy. A 50% proximal enterectomy or a single transection was performed in 20 growing rats weighing 145–160 g. Ten days following surgery, we determined mucosal mass parameters (weight, protein, and DNA content), sucrase activity, and DAO activity in the duodenum (segment A), proximal ileum (segment B), and distal ileum (segment C) of the remaining small intestine. Mucosal hyperplasia was demonstrated by the finding that in each segment, mucosal weight, protein, and DNA content per centimeter of gut length were significantly (P<0.01) higher (+38 to+78%) in the resected group than in transected controls. In segments B and C of resected rats, the changes in DAO activity expressed per gram of mucosa paralleled the changes in mucosal mass, the activity being increased by +69% and +49% (P<0.05) compared to the values recorded in transected controls. Expressed per centimeter of gut length, total DAO activity was also enhanced by +141% in segment B (P<0.05 vs controls) and by +87% in segment C(P>0.01 vs controls) of resected rats. In the duodenum, the changes in DAO activity were small (+36%) and not significant. In the ileum (segment C), significant correlations were established between total DAO activity and either mucosal weight (r=0.75,N=20,P<0.01) or mucosal DNA concentration (r=0.78, N=20, P<0.01) per centimeter of gut length, but there was no correlation between DAO activity and sucrase activity. Compared to control rats with transection, proximal enterectomy produced marked changes in the serum activity of DAO. Ten days following surgery, the mean value of serum DAO was fivefold higher (P< it0.005) in the resected group than in the transected group. These data indicate that after jejunectomy (1) the intestinal activity of DAO reflects accurately quantitative changes of the mucosal mass in the remaining ileum but not in the duodenum, and (2) circulating levels of DAO could be used as a marker of ileal mucosal adaptation after proximal enterectomy.     

A comparative phenotypical analysis of rheumatoid nodules and rheumatoid synovium with special reference to adhesion molecules and activation markers

OBJECTIVES—(1)To analyse the in situ expression of adhesion molecules in rheumatoid nodules. (2) To compare the endothelial expression of adhesion molecules in synovial tissue and subcutaneous nodules obtained from the same patients. (3) To compare the expression of adhesion molecules and activation markers on T cell lines from nodules and synovium.
METHODS—(1) Immunohistochemical analysis by APAAP technique of E selectin, CD44, ICAM-1, PECAM-1, and VCAM-1 was performed on 10 rheumatoid nodules from seven patients with rheumatoid arthritis (RA); nodules and synovium were simultaneously analysed from three patients. (2) T cell lines were generated from RA nodules (n=7) and synovium (n=7) by interleukin 2 expansion, and subsequently characterised by flow cytometry for surface expression of αEβ7, α4β7, CD44, L selectin, LFA-1a, PECAM-1, and CD30.
RESULTS—(1) In rheumatoid nodules, the palisading layer strongly stains for ICAM-1 and PECAM-1, but less pronounced for CD44. VCAM-1 staining was usually negative. ICAM-1 is upregulated in the vessels surrounding the central zone of fibrinoid necrosis. The immunohistological picture in different nodules derived from the same patient was similar. (2) The endothelial expression of adhesion molecules is comparable in RA nodules and synovium on an individual level, except for E selectin, which is overexpressed in nodule endothelium. (3) T cell lines from nodules and synovium display similar adhesion molecule profiles. However, the expression of CD30, a T cell activation marker linked with Th2 subsets, is higher in nodules compared with synovium.
CONCLUSION—These data support a recirculation hypothesis of T cells between articular and extra-articular manifestations in RA, although the activation state of the T cells in each of these localisations may differ.

Keywords: T cells; adhesion molecules; rheumatoid nodules; rheumatoid synovium     

Fatty acid profiles in Leishmania spp. isolates with natural resistance to nitric oxide and trivalent antimony

Fatty acids, especially those from phospholipids (PLFA), are essential membrane components that are present in relatively constant proportions in biological membranes under natural conditions. However, under harmful growth conditions, such as diseases, environmental changes, and chemical exposure, the fatty acid proportions might vary. If such changes could be identified and revealed to be specific for adverse situations, they could be used as biomarkers. Such biomarkers could facilitate the identification of virulence and resistance mechanisms to particular chemotherapeutic agents. Therefore, specific biomarkers could lead to better therapeutic decisions that would, in turn, enhance treatment effectiveness. The objective of this study was to compare the fatty acid profiles of trivalent antimony and nitric oxide (NO)-resistant and -sensitive Leishmania chagasi and Leishmania amazonensis isolates. Fatty acid methyl esters (FAMEs) were obtained from total lipids (MIDI), ester-linked lipids (ELFA), and ester-linked phospholipids (PLFA). FAMEs were analyzed by chromatography and mass spectrometry. Species- or resistance-associated differences in FAME profiles were assessed by nonmetric multidimensional scaling, multiresponse permutation procedures, and indicator species analyses. The isolate groups had different MIDI-FAME profiles. However, neither the ELFA nor PLFA profiles differed between the sensitive and resistant isolates. Levels of the fatty acid 18:1 Δ9c were increased in sensitive isolates (p?<?0,001), whereas the fatty acid 20:4 Δ5,8,11,14 showed the opposite trend (p?<?0.01). We conclude that these two fatty acids are potential biomarkers for NO and antimony resistance in L. chagasi and L. amazonensis and that they could be helpful in therapeutic diagnoses.     

Copper is an endogenous modulator of neural circuit spontaneous activity

For reasons that remain insufficiently understood, the brain requires among the highest levels of metals in the body for normal function. The traditional paradigm for this organ and others is that fluxes of alkali and alkaline earth metals are required for signaling, but transition metals are maintained in static, tightly bound reservoirs for metabolism and protection against oxidative stress. Here we show that copper is an endogenous modulator of spontaneous activity, a property of functional neural circuitry. Using Copper Fluor-3 (CF3), a new fluorescent Cu⁺ sensor for one- and two-photon imaging, we show that neurons and neural tissue maintain basal stores of loosely bound copper that can be attenuated by chelation, which define a labile copper pool. Targeted disruption of these labile copper stores by acute chelation or genetic knockdown of the CTR1 (copper transporter 1) copper channel alters the spatiotemporal properties of spontaneous activity in developing hippocampal and retinal circuits. The data identify an essential role for copper neuronal function and suggest broader contributions of this transition metal to cell signaling.The foundation of cellular signal transduction relies on intricate chemical messenger systems that operate through the dynamic spatial and temporal regulation of elements, ions, and molecules. Nowhere is this concept better illustrated than in the brain, which extensively regulates fluxes of alkali and alkaline earth metals such as sodium, potassium, and calcium for a diverse array of signaling processes. Interestingly, the brain also accumulates among the highest levels of transition metals in the body (1–3), including redox-active copper. This high-redox metal load, in combination with the brain''s disproportionately active oxygen metabolism (4), makes this organ particularly susceptible to oxidative stress (4–6). As such, copper has been historically regarded as a tightly sequestered cofactor that must be buried within protein active sites to protect against reactive oxygen species generation and subsequent free radical damage chemistry. Indeed, elegant work continues to identify molecular players that maintain copper homeostasis in the brain (7, 8) and related organs (9–11), and loss of this strict regulation is implicated in neurotoxic stress (12–14) and a variety of neurodegenerative and neurodevelopmental disorders including Menkes (15, 16) and Wilson''s (17) diseases, familial amyotrophic lateral sclerosis (18, 19), Alzheimer''s (6, 14, 20–22) and Huntington''s (23, 24) diseases, and prion-mediated encephalopathies (14, 25, 26).Despite this long-held paradigm, emerging data also link pools of labile copper (defined as dynamic and loosely bound stores that undergo facile ligand exchange relative to static, tightly bound pools buried within protein active sites) to neurophysiology. Included are observations of ⁶⁴Cu efflux from stimulated neurons (12, 27), reversible trafficking of ATP7A from the perinuclear trans-Golgi to neuronal processes by NMDA receptor activation (12), effects of copper chelation on olfactory response to thiol odorants (28), and direct X-ray fluorescence imaging of copper translocation in neurons from somatic cell bodies to peripheral processes upon depolarization (29). Against this backdrop, we have initiated a program aimed at exploring the potential contributions of loosely bound forms of redox-active metals like copper in cell signaling. In this report, we identify a role for copper in the brain as a modulator of spontaneous activity, a fundamental property of developing neural circuits. The design and synthesis of Copper Fluor-3 (CF3), a fluorescent copper sensor based on a hydrophilic and tunable rhodol scaffold, along with Control Copper Fluor-3 (Ctrl-CF3), a matched control dye based on an identical fluorophore but lacking responsiveness to copper, enabled the visualization of loosely bound Cu⁺ in dissociated neurons and neural tissue by one- and two-photon microscopy. Disruption of Cu⁺ stores by acute application of a copper chelator or genetic knockdown of the copper ion channel CTR1 altered the spatiotemporal properties of spontaneous activity. In dissociated hippocampal cultures, these manipulations increased the correlation of spontaneous calcium transients, whereas in retina, both cell participation and frequency of correlated calcium transients increased.     

First 24-h SNAP-II score and highest PaCO2 predict the need for ECMO in congenital diaphragmatic hernia

Background/Purpose

Early clinical predictors for the use of ECMO in patients with congenital diaphragmatic hernia (CDH) are lacking. We sought to evaluate the first 24-h SNAP-II score and highest PaCO₂ as predictors of ECMO support and in-hospital mortality in neonates with CDH.

Methods

Retrospective review of 47 consecutive neonates with CDH admitted to our institution from January 2007 to December 2010 was performed. Covariates of ECMO use including SNAP-II score and highest PaCO₂ within the first 24 h of NICU admission were evaluated.

Results

Of the 47 infants in this study, 24 patients were supported with ECMO. The ECMO group had a higher incidence of pulmonary hypertension, higher PaCO₂, and higher 24-h SNAP-II scores. Only the SNAP-II score and not highest PaCO₂ predicted mortality following multivariate adjustment.

Conclusions

The first 24-h SNAP-II score and highest PaCO₂ may provide some prognostic value in identifying neonates who undergo ECMO support; however neither measure was independently associated with the use of therapy. Only the SNAP-II score was associated with in-hospital mortality following multivariate adjustment. Additional study is needed to validate these results in a larger data set.     

Single‐Center Experience With Extracorporeal Life Support in 103 Nonpostcardiotomy Patients

Extracorporeal membrane oxygenation (ECMO) has been successfully used to support patients with cardiac arrest failing to respond to conventional cardiopulmonary resuscitation (CPR). Preimplant factors being indicative for success are unknown up to now. The study describes single center experience with special focus on differences between survivors and nonsurvivors. Between 2002 and 2009, 103 patients were supported within the scope of CPR by means of ECMO. Besides primary diagnosis, duration, and outcome, pH, lactate, mean arterial pressure, aspartate aminotransferase, bilirubin, catecholamine dosage, and oxygenation ratio before ECMO, after 2 h, 1 day, and at explantation were analyzed. One hundred three patients (51.2 ± 16 years, 35 women, 68 men) were analyzed. Primary cardiac failure led to CPR in 54%. Duration of support was 4.8 ± 0.6 days. Twenty‐nine (28.1%) patients survived to hospital discharge. On ECMO support, pH, lactate, and mean arterial pressure improved significantly. Catecholamine dosage was significantly reduced after ECMO implantation. Demographic data and primary diagnosis revealed no significant influence on outcome. pH, lactate, creatinine, and bilirubin differed significantly between survivors and nonsurvivors in the course of ECMO support. ECMO support during CPR reliably improves the circulatory and respiratory situation. Considering observed survival critical patient selection is mandatory. Although there are several significant differences between surviving patients and patients with fatal outcome, patient selection turns out to be difficult as clinically relevant factors show only limited predictive value. Future research should focus on better defining a population that may be best of all suited for the use of ECMO support in CPR.     

Impairment of brain redox homeostasis caused by the major metabolites accumulating in hyperornithinemia–hyperammonemia–homocitrullinuria syndrome in vivo

Ornithine, ammonia and homocitrulline are the major metabolites accumulating in hyperornithinemia-hyperammonemia-homocitrullinuria syndrome, a genetic disorder characterized by neurological regression whose pathogenesis is still not understood. The present work investigated the in vivo effects of intracerebroventricular administration of ornithine and homocitrulline in the presence or absence of hyperammonemia induced by intraperitoneal urease treatment on a large spectrum of oxidative stress parameters in cerebral cortex from young rats in order to better understand the role of these metabolites on brain damage. Ornithine increased thiobarbituric acid-reactive substances (TBA-RS) levels and carbonyl formation and decreased total antioxidant status (TAS) levels. We also observed that the combination of hyperammonemia with ornithine resulted in significant decreases of sulfhydryl levels, reduced glutathione (GSH) concentrations and the activities of catalase (CAT) and glutathione peroxidase (GPx), highlighting a synergistic effect of ornithine and ammonia. Furthermore, homocitrulline caused increases of TBA-RS values and carbonyl formation, as well as decreases of GSH concentrations and GPx activity. Hcit with hyperammonemia (urease treatment) decreased TAS and CAT activity. We also showed that urease treatment per se was able to enhance TBA-RS levels. Finally, nitric oxide production was not altered by Orn and Hcit alone or in combination with hyperammonemia. Our data indicate that the major metabolites accumulating in hyperornithinemia-hyperammonemia-homocitrullinuria syndrome provoke lipid and protein oxidative damage and a reduction of the antioxidant defenses in the brain. Therefore, it is presumed that oxidative stress may represent a relevant pathomechanism involved in the brain damage found in patients affected by this disease.