Pulmonary function in ventilated infants with bronchiolitis.
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Bronchiolitis↗
Biomedical subjects
J Hammer
Publications and source records attributed to J Hammer.
Excessive activation of tyrosine kinases leads to inhibition of proliferation in a thyroid carcinoma cell line.
Autocrine stimulation of growth is a hallmark of many tumor cell lines. In this work we investigated the synthesis and secretion of growth factors and the expression of their corresponding receptors in HTC-TSHr thyroid carcinoma cells. These cells synthesize epidermal growth factor (EGF) receptors and platelet-derived growth factor beta (PDGF beta) receptors and in addition transforming growth factor alpha (TGF alpha), PDGF-A and PDGF-B chains, respectively. Addition of EGF or PDGF-BB to the culture medium resulted in growth inhibition of HTC-TSHr cells. In contrast, treatment of the cells with low concentrations of neutralizing anti-TGF alpha antibodies or tyrosine kinase inhibitors led to stimulation of cell proliferation. Low concentrations of neutralizing anti-PDGF-B antibodies did not affect growth of the cells. As expected, cell proliferation was inhibited when high concentrations of either neutralizing anti-TGF alpha antibodies or anti-PDGF-B antibodies were applied. PDGF-AA did not influence growth of HTC-TSHr cells. We conclude that growth of HTC-TSHr thyroid carcinoma cells is influenced by two autocrine loops between TGF alpha and EGF receptors and between PDGF-B and PDGF beta receptors. However, our data suggest that excessive activation of tyrosine kinase receptors in these cells results in a relative inhibition rather than stimulation of growth.
Cell Division↗
Substrate selection by transporters associated with antigen processing occurs during peptide binding to TAP.
Presentation of antigenic peptides by major histocompatibility complex (MHC) class I molecules depends on translocation of cytosolic peptides into the endoplasmic reticulum (ER) by transporters associated with antigen processing (TAP). Peptide transport by TAP is thought to include at least two steps: initial binding of peptide to TAP, and its subsequent translocation requiring ATP hydrolysis. These events can be monitored in peptide binding and transport assays. Previous studies have shown that the efficiency of peptide transport by human, mouse and rat transporters varies according to the C-terminals of peptide substrates in an allele and species-specific manner. However, it has not been clear during which step of peptide interaction with TAP selection occurs. We used an assay monitoring the peptide binding step to study the binding affinity of a library of 199 peptides for human TAP and the two major allelic rat TAP complexes. We observed a dominant influence of the C-terminus on peptide binding affinity for all transporters, and highly restrictive selection of peptides with aliphatic and aromatic C-terminals by rat TAP1/TAP2u complexes. The selectivity of peptide binding to rat TAP complexes is in full accordance with published data on selective peptide transport and on control of antigen presentation by rat TAP. These results strongly suggest that (i) peptide selection by TAP occurs exclusively in the initial binding step; (ii) all factors involved in peptide selection by TAP are present in insect cells.
Alleles↗
Gastroduodenal permeability in Crohn's disease.
BACKGROUND: Gastric permeability was prospectively investigated by determination of sucrose excretion in 100 patients with Crohn's disease. Results were compared with histological findings and the lactulose-mannitol ratio as a measure of intestinal permeability. METHODS: All subjects underwent oesophagogastroduodenoscopy with biopsies of all investigated parts. Thirty-two Helicobacter pylori-positive patients were excluded from further analyses. RESULTS: Gastroduodenal permeability was significantly higher in patients with Crohn's disease than in control subjects (P < 0.00001). Sucrose excretion alone did not predict microscopic inflammation of the upper gastrointestinal tract. Increased gastroduodenal permeability with a concomitant rise in intestinal permeability predicted histological upper gastrointestinal involvement of Crohn's disease with a likelihood of 86%. The negative predictive value was 43%. CONCLUSION: In parallel with findings in the small intestine, gastroduodenal permeability is increased in a high proportion of patients with Crohn's disease. In patients with an increased lactulose-mannitol ratio, elevated sucrose excretion is highly predictive of histological gastroduodenal involvement.
Adolescent↗
Prediction of MHC class II-binding peptides using an evolutionary algorithm and artificial neural network.
MOTIVATION: Prediction methods for identifying binding peptides could minimize the number of peptides required to be synthesized and assayed, and thereby facilitate the identification of potential T-cell epitopes. We developed a bioinformatic method for the prediction of peptide binding to MHC class II molecules. RESULTS: Experimental binding data and expert knowledge of anchor positions and binding motifs were combined with an evolutionary algorithm (EA) and an artificial neural network (ANN): binding data extraction --> peptide alignment --> ANN training and classification . This method, termed PERUN, was implemented for the prediction of peptides that bind to HLA-DR4(B1*0401). The respective positive predictive values of PERUN predictions of high-, moderate-, low- and zero-affinity binders were assessed as 0.8, 0.7, 0.5 and 0.8 by cross-validation, and 1.0, 0.8, 0.3 and 0.7 by experimental binding. This illustrates the synergy between experimentation and computer modeling, and its application to the identification of potential immunotherapeutic peptides. AVAILABILITY: Software and data are available from the authors upon request. CONTACT: vladimir@wehi.edu. au
Algorithms↗
Accelerated right colonic emptying after simulated upper gut hemorrhage.
OBJECTIVE: Upper gastrointestinal hemorrhage delays gastric emptying. Our aim was to evaluate the effect of gastrointestinal hemorrhage on small intestinal, ileocecal, and proximal colonic transit. METHODS: Healthy volunteers were randomized to receive either a duodenal infusion of heparinized autologous blood (n = 7) or egg white, acting as a control substance with similar composition; 1.5 mCi99mTc-DTPA was added to the infused substances. We infused 30 ml at an infusion rate of 1 ml/min. Gamma-camera images were taken for 4 h or until all radioactivity had entered the colon. Arrival of radiolabels in the colon and also counts in the ascending and transverse colon were quantified. RESULTS: Small intestinal and ileocecal transit were not significantly different between blood and egg white infusions. However, ascending colonic emptying was significantly faster after blood infusion compared to egg white. Four hours after the start of blood infusion a median of 30% of counts were in the transverse colon (11-50%; 25th-75th percentile) versus 0% (0-7%) after egg white infusion (p < 0.001). CONCLUSION: We concluded that simulated upper gastrointestinal bleeding hastens proximal colonic transit, but does not alter small intestinal transit and colonic filling.
Adult↗
Lipids infused into the jejunum accelerate small intestinal transit but delay ileocolonic transit of solids and liquids.
BACKGROUND: Various nutrients are known to alter small intestinal motility patterns although their effect on transit of fluids and solids in man is not clear. AIMS: To determine small intestinal transit of solids and liquids during perfusion with lipids, protein, and non-energy solutions. METHODS: Twenty eight healthy volunteers received a jejunal infusion (1 ml/minute for 30 minutes) of one of four solutions: a lipid or a protein solution (4.18 J/ml), a non-absorbable electrolyte solution containing polyethylene glycol, or 0.9% sodium chloride. As solid phase marker 1 g of amberlite resin pellets labelled with 111InCl3 was added; 99mTc DTPA was used as a fluid phase marker. Images were obtained on a gamma camera at 10 minute intervals for four hours or until all radiolabel was detected in the colon. RESULTS: Intestinal transit of solids and liquids from the duodenojejunal junction to the caecum was simultaneous, and independent of the energy content of the solution infused. Lipid infusion accelerated transit through the small intestine but delayed transport of chyme along the ileocolonic junction. After protein small intestinal transit was slowest; ileocolonic transit on the other hand was fastest with protein. Transit of the non-energy solutions was in between that of the nutrient solutions. CONCLUSIONS: Transit times through the small intestine and the ileocolonic junction were influenced by the luminal contents. In the small intestine fat induced significantly faster transit compared with proteins, but delayed ileocolonic transit. Once in the small intestine, solids and liquids transit the small bowel together, independent of the luminal content.
Adolescent↗
Intraluminal capsaicin does not affect fluid and electrolyte absorption in the human jejunum but does cause pain.
BACKGROUND: Stimulation of sensory nerves with capsaicin regulates ion transport in the small intestine in animal experiments. AIM: To investigate whether sensory nerves that are stimulated by capsaicin administration influence fluid and electrolyte absorption in the human jejunum in vivo. METHOD: Intestinal perfusion studies were performed in 12 healthy subjects using a four lumen tube with a proximal occlusion balloon and a plasma-like electrolyte solution. After an initial control period, 5 (n = 3), 10 (n = 8), or 50 (n = 1) micrograms/ml capsaicin was added to the perfusate, and this was followed by a final control period. Rates of absorption of water, sodium, potassium, chloride, and bicarbonate were determined in a 30 cm segment of jejunum using a non-absorbable volume marker. RESULTS: At all three concentrations of capsaicin there were no significant changes in water and electrolyte absorption as compared with control periods. Two subjects who received 10 micrograms/ml and the subject receiving 50 micrograms/ml experienced crampy abdominal pain. CONCLUSION: The results do not support the hypothesis that capsaicin sensitive afferent nerves are involved in the physiological regulation of net absorption or secretion across the human jejunal mucosa. Chemical stimulation of these nerves, however, gives rise to abdominal pain.
Adult↗
Total lung capacity by N2 washout from high and low lung volumes in ventilated infants and children.
Although there is a strong rationale for the assessment of the subdivisions of lung volume, lung function testing has focused on the measurement of FRC alone in ventilated infants and children. To assess the feasibility, reproducibility, and accuracy of measurements of total lung capacity (TLC), FRC, and their ratio, we determined both lung volumes in 50 critically ill, intubated, and paralyzed infants (mean age [SEM]), 19.9 [4.6] mo) with a variety of lung diseases, by a modified N2 washout technique from end-exhalation and from +40 cm H2O inspiratory pressure, respectively. In the same infants, we also defined TLC by adding inspiratory capacity, measured by pneumotachograph during a passive exhalation from +40 cm H2O to FRC measured by N2 washout. Respiratory mechanics were measured by single-breath occlusion, and the patients were classified according to clinical picture and lung function into groups without lung disease or with restrictive or obstructive disease. The TLC data obtained by both methods showed good agreement for the infants without lung disease or restrictive disease (limits of agreement [LOA]: -3.8/4.6 and -2.9/3.2 ml/kg, respectively). The agreement was less in the infants with airflow obstruction where the N2 washout gave slightly higher values (LOA: -7.1/11.3 ml/kg). Mean FRC/TLC was significantly elevated in the obstructive group, whereas mean FRC alone did not differ from the group without lung disease. Our results suggest that TLC can be measured by both methods in intubated infants, but with limited agreement in obstructive disease. FRC/TLC ratios allow an estimation of the degree of pulmonary hyperinflation.
Child, Preschool↗
A novel approach to human anti-HLA mABs production: use of phage display libraries.
The production of human monoclonal antibodies was previously limited to very laborious and time-consuming processes involving EBV-transformation and/or hybridoma generation. Due to the development of molecular cloning techniques, it is now possible to produce human monoclonal antibody fragments quickly by panning phage display libraries against predefined antigenic specificities. Therefore, we tested this technology for producing human single chain Fv fragments (scFvs) against HLA-DR1 purified molecules immobilized on solid phase. Enrichment of DR1-specific phages was measured through five selection rounds of a synthetic library and revealed a 100-fold amplification. Soluble antibody fragments were then expressed and 7 out of 48 clones were found to secrete scFvs which specifically bind to DR1 molecules in ELISA. Further analysis revealed binding of the scFvs also to DR3 but not to DR5 or DR7 molecules correlating with the presence of particular polymorphic aminoacid residues in the DR beta chain. Western blot analysis indicated that the 7 scFvs react with the DR1 alpha/beta-dimer but not with free alpha- or beta- chains. This study shows that the innovative approach of phage display libraries can efficiently provide scFv fragments as useful reagents for the identification and dissection of HLA polymorphic epitopes.
Antibodies, Monoclonal↗
Absence of functional relevance of human transporter associated with antigen processing polymorphism for peptide selection.
The polymorphic TAP (TAP1 and TAP2) genes code for a heteromeric complex that translocates cytosolic peptides across the membrane of the endoplasmic reticulum where they associate with MHC class I molecules. In the rat, extensive TAP2 polymorphism modifies the spectrum of peptides presented by MHC class I molecules on the cell surface. Although polymorphism of human TAP proteins is more limited, it has been proposed to be associated with susceptibility to autoimmune diseases. To analyze the peptide selectivity and transport efficiency of human TAP alleles, we have generated the most frequent TAP1 and TAP2 alleles using site-directed mutagenesis. Alleles were cloned in the baculovirus expression system and six TAP1/2 allelic combinations were overexpressed in Sf9 insect cells. Microsomes containing TAP1/2 dimers were isolated, and peptide binding assays were performed using random sequence peptide libraries of variable length (7-18 mer) and a 9-mer polyalanine peptide substituted with various amino acids in positions 1, 2, 3, 7, and 9. All studied allelic combinations selected peptides with identical substitutions and length. Furthermore, transport assays showed similar efficiency and kinetics of peptide translocation by the different TAP alleles. These data suggest that the major human TAP alleles transport identical peptide sets and are therefore unlikely to contribute to predisposition to autoimmune diseases.
ATP Binding Cassette Transporter, Subfamily B, Mem↗
Binding of malaria T cell epitopes to DR and DQ molecules in vitro correlates with immunogenicity in vivo: identification of a universal T cell epitope in the Plasmodium falciparum circumsporozoite protein.
The efficacy of a malaria peptide vaccine would be enhanced by the inclusion of a parasite-derived universal T cell epitope to ensure that all vaccinees develop parasite-specific cellular and humoral immunity. Two circumsporozoite (CS) protein T cell epitopes, previously identified by CD4+ T cell clones derived from Plasmodium falciparum sporozoite-immunized volunteers, were studied to determine their HLA class II binding potential. One epitope, located in amino acid (aa) 326-345 of the P. falciparum (NF54 strain) CS protein, was "universal" in that it could bind to multiple DR and DQ molecules in vitro. In contrast, the second epitope, T1, which is located in the CS repeat region, was recognized by T cells in the context of DQ6 (DQB1*0603) and did not bind with high affinity to any of the class II molecules tested in the peptide binding assays. The in vitro patterns of peptide/HLA interactions correlated with immunogenicity in vivo. A multiple antigen peptide (MAP) containing the aa 326-345 epitope elicited responses in eight inbred strains (H-2(a,b,d,k,p,q,r,s)), while the T1 MAP was recognized by only a single haplotype, H-2b. The combination of the universal aa 326-345 T cell epitope and the T1 repeat in a di-epitope MAP overcame the genetic restriction to the P. falciparum CS repeat region and elicited antisporozoite Ab responses in all of the MAP-immunized mice. Synthetic peptide malaria vaccines containing the aa 326-345 universal T cell epitope would be expected to elicit parasite-specific immune responses in both sporozoite-primed and naive individuals of diverse genetic backgrounds.
Animals↗
Different modes of peptide interaction enable HLA-DQ and HLA-DR molecules to bind diverse peptide repertoires.
The role of HLA-DQ molecules in Ag presentation has, thus far, remained elusive. Here we report that two DQ allotypes, DQ7 (DQA1*0501/B1*0301) and DQ9 (DQA1*0201/B1*0303), are capable of binding peptide repertoires in complementarity with DR molecules. The results reflect fundamental differences in the binding modes of these two HLA class II isotypes, in that DQ7 and DQ9 but not DR molecules appear to have the capacity to bind peptide structures without type 1-like anchor residues. Consistent with this is our observation that none of the amino acid side chains of the class II-associated invariant chain peptides (CLIP) are required for association with DQ7 and DQ9, even though many of them are essential for CLIP-DR interaction. Together, these data reveal a functional complementarity of HLA-DR and -DQ molecules in Ag presentation.
Amino Acid Sequence↗
A peptide-binding motif for I-A(g7), the class II major histocompatibility complex (MHC) molecule of NOD and Biozzi AB/H mice.
The class II major histocompatibility complex molecule I-A(g7) is strongly linked to the development of spontaneous insulin-dependent diabetes mellitus (IDDM) in non obese diabetic mice and to the induction of experimental allergic encephalomyelitis in Biozzi AB/H mice. Structurally, it resembles the HLA-DQ molecules associated with human IDDM, in having a non-Asp residue at position 57 in its beta chain. To identify the requirements for peptide binding to I-A(g7) and thereby potentially pathogenic T cell epitopes, we analyzed a known I-A(g7)-restricted T cell epitope, hen egg white lysozyme (HEL) amino acids 9-27. NH2- and COOH-terminal truncations demonstrated that the minimal epitope for activation of the T cell hybridoma 2D12.1 was M12-R21 and the minimum sequence for direct binding to purified I-A(g7) M12-Y20/K13-R21. Alanine (A) scanning revealed two primary anchors for binding at relative positions (p) 6 (L) and 9 (Y) in the HEL epitope. The critical role of both anchors was demonstrated by incorporating L and Y in poly(A) backbones at the same relative positions as in the HEL epitope. Well-tolerated, weakly tolerated, and nontolerated residues were identified by analyzing the binding of peptides containing multiple substitutions at individual positions. Optimally, p6 was a large, hydrophobic residue (L, I, V, M), whereas p9 was aromatic and hydrophobic (Y or F) or positively charged (K, R). Specific residues were not tolerated at these and some other positions. A motif for binding to I-A(g7) deduced from analysis of the model HEL epitope was present in 27/30 (90%) of peptides reported to be I-A(g7)-restricted T cell epitopes or eluted from I-A(g7). Scanning a set of overlapping peptides encompassing human proinsulin revealed the motif in 6/6 good binders (sensitivity = 100%) and 4/13 weak or non-binders (specificity = 70%). This motif should facilitate identification of autoantigenic epitopes relevant to the pathogenesis and immunotherapy of IDDM.
Amino Acid Sequence↗
Assessment of pulmonary function in the early phase of ARDS in pediatric patients.
Scant data are available on lung function in acute respiratory distress syndrome (ARDS) in pediatric patients. We measured respiratory mechanics by single-breath occlusion and maximum expiratory flow-volume curves by forced deflation in ten critically ill infants with clinical ARDS. Ten mechanically ventilated infants without lung disease served as the control group. To assess the severity of the lung injury in the infants with ARDS, we modified an adult scoring system that calculates a score (from 0 to 4; > 2.5 indicates severe lung injury) based on the extent of chest radiographic changes, degree of hypoxemia, amount of positive end-expiratory pressure (PEEP), and total respiratory system compliance. The lung injury scores of our patients were in the range of 2.75 to 3.75. The lung injury scores of the control group were zero. The predominant alteration in lung function was restrictive, as characterized by a significant decrease in total respiratory system compliance (0.41 +/- 0.13 ml/cmH2O/kg versus 1.12 +/- 0.16 ml/cmH2O/kg of controls; P < 0.001) and forced vital capacity (21.5 +/- 6.5 ml/kg versus 59.2 +/- 6.3 ml/kg of controls; P < 0.001). Maximum expiratory flow rates at 10% forced vital capacity were significantly increased (23.6 +/- 20.1 ml/kg/sec versus 8.4 +/- 2.5 ml/kg/sec of controls; P < 0.05), confirming the absence of any significant obstructive abnormalities. The passive expiratory flow-volume curves were curvilinear and convex in shape, indicating inhomogeneous lung pathology. The inhomogeneous distribution of lung injury in ARDS restricts the validity of respiratory mechanics measurements that rely on a single-compartment model. However, the forced deflation technique allows accurate spirometric assessments of the severity of restrictive (and obstructive) lung function changes in intubated infants with severe ARDS. Such measurements can be incorporated into lung injury scoring systems to classify the severity of the disease process for the purpose of outcome evaluation and to evaluate the effect of therapeutic interventions.
Case-Control Studies↗
Acute respiratory distress syndrome caused by respiratory syncytial virus.
Acute respiratory distress syndrome (ARDS) complicating severe respiratory syncytial virus (RSV) infection has been described in only a few infants. In contrast to the low mortality rates usually associated with RSV infections (< 5%), mortality rates in the range of 40-70% have been reported in pediatric patients with ARDS. However, studies on patients with ARDS are usually lumped with respect to causation, and the disease course of RSV-induced ARDS has not been previously studied. We examined the pulmonary function abnormalities of 37 infants with RSV-induced respiratory failure who were admitted to our pediatric intensive care unit for assisted ventilation. Measurements included respiratory mechanics, maximum expiratory flow-volume curves, and lung volumes. These allowed the calculation of a Murray lung injury score (modified for pediatric use) in which radiographic findings, ventilator settings, lung compliance, and blood gas results were considered. We identified ten infants with severe restrictive lung disease who fulfilled the clinical criteria for classification as ARDS. All had lung injury scores above 2.5, compatible with a diagnosis of ARDS. Twenty-seven infants had obstructive patterns of lung function consistent with a clinical diagnosis of RSV bronchiolitis. The patients with RSV-induced ARDS were significantly younger, and had a longer time on assisted ventilation (P < 0.05) and a higher proportion of predisposing illnesses (P < 0.05, odds ratio = 6.67, two-tailed Fisher's exact test) when compared with the patients who had obstructive disease. Only one patient (who had immunodeficiency) died, and all others were successfully managed on conventional mechanical ventilation. We conclude that RSV-induced respiratory failure represents a relatively benign cause of ARDS in pediatric patients. Our observations support the notion of differentiating ARDS with respect to causation, especially when novel and experimental therapy is considered and mortality rates are analyzed.
Causality↗
Does the anchorage form and depth influence the pull-out strength of screws from bone cement? An experimental study.
The pull-out strengths of cortical screws inserted into soft, unpolymerised Refobacin Palacos bone cement (procedure S) and into hardened polymerised cement into which a hole had been drilled and tapped (procedure P) were compared. Cortical screws 58 mm in length, outer diameter 4.5 mm and inner diameter 2.95 mm were used. Screws were inserted into cement cylinders at 5 mm incremental depths between 10 and 30 mm. At a screw depth of less than 25 mm, the screws pulled out, and at a depth of greater than 25 mm, the screws broke in both procedures. There was no statistically significant difference in pull-out strength leading to burst or break between the two procedures for screws inserted to comparable depths, but there was a statistically significant difference regarding the screwing depth regardless of the procedure of screw insertion chosen. The average material stability (sigma) of the cortical screws used was calculated to be 1191 N/mm2, and the elasticity limit was 5137 N. This study demonstrated that the material stability and not the depth of screw insertion was the limiting parameter in screw anchorage in bone cement while static testing. To avoid screw breakage due to fatigue during continuous alternate loading, the screws should not be loaded above this value.
Analysis of Variance↗
[Strahlentherapie und Onkologie--the publishing organ of German-speaking radio-oncologists].
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Germany↗