bluuebunny/crossref_metadata_2025 huggingface dataset:
Exploring relationships between mimic configuration, peptide conformation and biological activity in indolizidin‐2‐one amino acid analogs of gramicidin S*
Authors: Roy S. , Lombart H.‐G. , Lubell W. D. , Hancock R. E. W. , Farmer S. W.
Published: March 2003

Abstract: Indolizidin‐2‐one amino acids (I2aas, 6S‐ and 6R‐1) possessing 6S‐ and 6R‐ring‐fusion stereochemistry were introduced into the antimicrobial peptide gramicidin S (GS) to explore the relationships between configuration, peptide conformation and biological activity. Solution‐phase and solid‐phase techniques were used to synthesize three analogs with I2aa residues in place of the d‐Phe‐Pro residues at the turn regions of GS: [(6S)‐I2aa4−5,4′−5′]GS (2), [Lys2,2′,(6S)‐I2aa4−5,4′−5′]GS (3) and [(6R)‐I2aa4−5,4′−5′]GS (4). Although conformational analysis of [I2aa4−5,4′−5′]GS analogs 2−4 indicated that both ring‐fusion stereoisomers of I2aa gave peptides with CD and NMR spectral data characteristic of GS, the (6S)‐I2aa analogs 2 and 3 exhibited more intense CD curve shapes, as well as greater numbers of nonsequential NOE between opposing Val and Leu residues, relative to the (6R)‐I2aa analog 4, suggesting a greater propensity for the (6S)‐diastereomer to adopt the β‐turn/antiparallel β‐pleated sheet conformation. In measurements of antibacterial and antifungal activity, the (6S)‐I2aa analog 2 exhibited significantly better potency than the (6R)‐I2aa diastereomer 4. Relative to GS, [(6S)‐I2aa4−5,4′−5′]GS (2) exhibited usually 1/2 to 1/4 antimicrobial activity as well as 1/4 hemolytic activity. In certain cases, antimicrobial and hemolytic activities of GS were shown to be dissociated through modification at the peptide turn regions with the (6S)‐I2aa diastereomer. The synthesis and evaluation of GS analogs 2−4 has furnished new insight into the importance of ring‐fusion stereochemistry for turn mimicry by indolizidin‐2‐one amino acids as well as novel antimicrobial peptides.
Relative hydrophobicity of di‐ to hexapeptides as measured by aqueous two‐phase partitioning
Authors: Gulyaeva N. , Zaslavsky A. , Chait A. , Zaslavsky B.
Published: March 2003

Abstract:  Partitioning of 153 di‐ to hexapeptides in an aqueous dextran–PEG two‐phase system containing 0.15 m NaCl in 0.01 m sodium phosphate buffer, pH 7.4 was examined. The relative hydrophobicity of the peptides was estimated and expressed in equivalent numbers of methylene units. Analysis of the data shows that the additivity principle applies for the relative hydrophobicity of up to hexapeptides. The relative hydrophobicities of Trp, Glu, and Asp residues in heterooligopeptides are noticeably different from those in corresponding homooligopeptides. The relative hydrophobicity of peptides can be calculated and used as a structural descriptor in quantitative structure–activity relationship analysis. The peptide bitterness threshold is shown to be quantitatively related to the peptide structure described as a combination of the relative hydrophobicity and lipophilicity (logD) of peptides.
Dermorphin‐based potential affinity labels for µ‐opioid receptors*
Authors: Choi H. , Murray T. F. , Aldrich J. V.
Published: March 2003

Abstract: Dermorphin and [Lys7]dermorphin, selective µ‐opioid receptor ligands originating from amphibian skin, have been modified with various electrophiles in either the ‘message’ or ‘address’ sequences as potential peptide‐based affinity labels for µ‐receptors. Introduction of the electrophilic isothiocyanate and bromoacetamide groups on the para position of Phe3 and Phe5 was accomplished by incorporating Fmoc‐Phe(p‐NHAlloc) into the peptide followed by selective deprotection and modification. The corresponding amine‐containing peptides were also prepared. The pure peptides were evaluated in radioligand binding experiments using Chinese hamster ovary (CHO) cells expressing µ‐ and δ‐opioid receptors. In dermorphin, introduction of the electrophilic groups in the ‘message’ domain lowered the binding affinity by > 1000‐fold; only [Phe(p‐NH2)3]dermorphin retained nanomolar affinity for µ‐receptors. Modifications in the ‘address’ region of both dermorphin and [Lys7]dermorphin were relatively well tolerated. In particular, [Phe(p‐NH2)5,Lys7]dermorphin showed similar affinity to dermorphin, with almost 2‐fold higher selectivity for µ‐receptors. [Phe(p‐NHCOCH2Br)5]‐ and [Phe(p‐NHCOCH2Br)5,Lys7]dermorphin exhibited relatively high affinity (IC50 = 27.7 and 15.1 nm, respectively) for µ‐receptors. However, neither of these peptides inhibited [3H]DAMGO binding in a wash‐resistant manner.
Simultaneous induction of specific immunoglobulin A–producing cells in major and minor salivary glands after tonsillar application of antigen in rabbits
Authors: Inoue H. , Fukuizumi T. , Tsujisawa T. , Uchiyama C.
Published: March 2003

The immunoglobulin A (IgA)–producing cells in the stroma of major salivary glands are induced by antigenic stimulation of the mucosal immune system. Whether such cells also are induced in minor salivary glands by this stimulation remains to be determined. After application of sheep red blood cells to the palatine tonsils every 3 days for 6 weeks, anti–sheep red blood cell IgA was detected in saliva both by agglutination tests and by enzyme‐linked immunosorbent assay. Using enzyme‐linked immunospot assay, an increase in the number of anti–sheep red blood cell IgA‐producing cells was found in minor as well as in major salivary glands of the sixth week of application; such cells constituted 4.9% to 5.9% of the total number of IgA‐producing cells in these tissues. Tonsillar application of whole cells of formalin‐killed Streptococcus sobrinus induced anti–S. sobrinus IgA in saliva. The number of anti–S. sobrinus IgA‐producing cells in the above glands simultaneously increased over 6 weeks, and reached 5.2–5.6% of the total number of IgA‐producing cells.
Actinobacillus actinomycetemcomitans may utilize either actin‐dependent or actin‐independent mechanisms of invasion
Authors: Brissette C. A. , Fives‐Taylor P. M.
Published: March 2003

Actinobacillus actinomycetemcomitans is an important pathogen implicated in juvenile and adult periodontal diseases. An important virulence factor of A. actinomycetemcomitans is the ability to invade human oral epithelial cells. A clinical isolate, A. actinomycetemcomitans SUNY 465, has previously been shown to enter epithelial cells by an actin‐dependent mechanism. The internalized bacteria are surrounded by an actin halo upon entry. These data are consistent with the mode of entry associated with many enteric pathogens. We tested the effects of cytochalasin D, an inhibitor of the actin microfilament network, on bacterial entry to determine whether this mode of entry was common to other A. actinomycetemcomitans clinical isolates. Cytochalasin D was added prior to infection. A. actinomycetemcomitans SUNY 523 and A. actinomycetemcomitans 4065 exhibited enhanced ability to enter epithelial cells in the presence of cytochalasin D. Immunofluorescent labeling of bacteria and host cell actin confirmed that actin was not being mobilized by the entry of A. actinomycetemcomitans SUNY 523. Inhibitors of receptor‐mediated endocytosis inhibited invasion of A. actinomycetemcomitans SUNY 523 and A. actinomycetemcomitans 4065. Microtubule effectors did not inhibit invasion of A. actinomycetemcomitans. A. actinomycetemcomitans SUNY 523, but not A. actinomycetemcomitans 4065, was deficient in exit from epithelial cells as determined by the absence of organisms in the assay medium. These data suggest that A. actinomycetemcomitans strains utilize at least two distinct mechanisms for entry into epithelial cells, and that A. actinomycetemcomitans SUNY 523 may be defective in exit and cell‐to‐cell spread.
Intracellular flux of glucose metabolism in streptococcal cells by simultaneous monitoring of fluorescence dependent on reduced nicotinamide adenine nucleotide and acid excretion under strictly anaerobic conditions
Authors: Iwami Y. , Yamada T.
Published: March 2003

Reduced nicotinamide adenine nucleotide (NADH)–dependent fluorescence and acid excretion during glucose pulse to washed Streptococcus mutans cells were monitored simultaneously at pH 7.0 with a fluorescence spectrophotometer and a pH‐stat. Acid excretion started with addition of glucose. At the same time, the fluorescence dropped quickly to a minimum level and increased to a plateau level, suggesting that pyruvate metabolism started immediately after addition of glucose, then the rate of the pyruvate metabolism became almost equal to the rate of glycolysis. When the acid excretion stopped, the fluorescence increased rapidly from the plateau to the maximum level, suggesting that the pyruvate metabolism stopped first, and then began to decrease to the original level. The system used in this study for simultaneously monitoring the level of NADH and acid excretion gives us a crucial tool to clarify a biochemical mechanism of the control of sugar metabolism by streptococci.
Salivary mucin as related to oral Streptococcus mutans in elderly people
Authors: Baughan L. W. , Robertello F. J. , Sarrett D. C. , Denny P. A. , Denny P. C.
Published: March 2003

MG1 (MUC5b and MUC4 ) and MG2 (MUC7), predominant mucins in human whole saliva, provide lubrication and antimicrobial protection for oral tissues. This study examines potential relationships between Streptococcus mutans titers in the oral cavity and the following: mucin concentrations; unstimulated and stimulated whole saliva flow rates; decayed, missing, and filled tooth surfaces; and age of 24 elderly patients. S. mutans titers were determined using Denticult SM. Mucin concentrations were determined using Stains‐all, sodium dodecyl sulfate–polyacrylamide gel electrophoresis. Logistic regression was used to identify potential relationships between the above variables. S. mutans classification served as the dependent variable. The remaining variables were possible predictor variables. The best model for predicting S. mutans category contained log MG2 as a predictor variable for all of its parameter estimates. No other set of parameter estimates were statistically significant. These results suggest that elevated S. mutans titers are significantly associated with diminished concentrations of MG2 in unstimulated whole saliva, as quantified in mucin‐dye binding units.
Importance of Dialister pneumosintes in human periodontitis
Authors: Contreras A. , Doan N. , Chen C. , Rusitanonta T. , Flynn M. J. , Slots J.
Published: March 2003

Anaerobic bacteria play important roles in the pathogenesis of human periodontitis. This study examined the relationship between a potentially new periodontopathic bacterium Dialister pneumosintes and periodontal disease. A total of 73 women and 62 men aged 18 to 86 years participated in the study. Using a 16S rRNA polymerase chain reaction identification method, the presence of D. pneumosintes was determined in paper‐point samples from periodontal pockets of 105 periodontitis and 30 gingivitis patients. D. pneumosintes was detected in 83% of patients having severe periodontitis and in 19% of patients having slight periodontitis. We suggest adding D. pneumosintes to the group of suspected periodontal pathogens.
Histatin 5 inhibits inflammatory cytokine induction from human gingival fibroblasts by Porphyromonas gingivalis
Authors: Imatani T. , Kato T. , Minaguchi K. , Okuda K.
Published: March 2003

Porphyromonas gingivalis is a gram‐negative rod associated with the progression of human periodontal disease. It has been demonstrated that outer‐membrane proteins as well as lipopolysaccharides from P. gingivalis ATCC 53977 can induce interleukin 6 (IL‐6) and IL‐8 from the cells of the periodontium in vitro. But, they cannot induce IL‐1 and tumor necrosis factor‐α from the cells. In the present study, we studied the effects of salivary protein on cytokine induction from human gingival fibroblasts by P. gingivalis outer‐membrane protein. Histatin 5 suppressed the IL‐6 and IL‐8 induction by P. gingivalis outer‐membrane protein. This activity was more effective when outer‐membrane protein was incubated with histatin 5 before addition to the cell culture. The present study indicates that histatin 5 restrains induction of inflammatory cytokines by periodontal pathogens and that histatin is one of the salivary proteins responsible for this activity.
Identification of an antigenic protein Pga30 from Porphyromonas gingivalis W50
Authors: Hendtlass A. , Dashper S. G. , Reynolds E. C.
Published: March 2003

Porphyromonas gingivalis is a black‐pigmented, gram‐negative bacterium that has been implicated as a major pathogen in the development of adult periodontitis. In an approach to identify a P. gingivalis antigen uniquely seroreactive with healthy subjects, we produced a surface and periplasmic extract of P. gingivalis, separated that extract into 36 fractions using anion‐exchange chromatography and screened each fraction for reactivity in an enzyme‐linked immunosorbent assay (ELISA) using sera from eight periodontitis patients and eight age‐ and sex‐matched healthy controls. All of the diseased subjects harboured subgingival P. gingivalis by DNA probe analysis and exhibited similar seroreactivity profiles to the anion exchange fractions. However, only two of the healthy subjects (C10 and C15) were seroreactive with the fractions. The highest reactivity for all the seropositive subjects was with the same anion‐exchange fractions 13–15. The anion exchange fraction (
14) with the highest seroreactivity was subjected to gel filtration chromatography, and fraction 22 from this chromatography exhibited the highest eactivity with all the seropositive subjects. However, fraction 27 was found to be uniquely seroreactive with healthy subject C10, as it was not recognized by sera from any of the diseased or the other healthy subjects. This fraction was further purified by reversed‐phase high‐pressure liquid chromatography and shown to contain a 30‐kDa protein as determined by SDS‐PAGE. Control subject C10 had no pocket depths greater than 3 mm and no sites that bled on gentle probing; however, P. gingivalis was detected in subgingival plaque samples at a level of 105–106 cells per site in two of the ten sites sampled. This subject was also unusual in that he exhibited a seroreactivity profile similar to that of diseased subjects, which was not characteristic of the healthy control subjects. The unique reactivity of the 30‐kDa antigen, designated Pga30, with subject C10 serum was confirmed in a Western blot with the purified antigen. N‐Terminal sequence analysis of Pga30 produced a single, unambiguous protein sequence confirming the purity of the protein. A search of the database using the N‐terminal sequence obtained did not reveal any significant sequence similarity. In conclusion, we have identified a P. gingivalis antigen that was uniquely reactive in an ELISA and Western blot with serum from a subject with no clinical signs of periodontitis who harbored P. gingivalis in subgingival plaque.
Establishment of Porphyromonas gingivalis–specific T‐cell lines from atherosclerosis patients
Authors: Choi J. , Chung S.‐W. , Kim S.‐J. , Kim S.‐J.
Published: March 2003

Porphyromonas gingivalis–specific T‐cell lines were established from atherosclerotic lesions from two patients. T‐cell lines were found to be a mixture of CD4+ and CD8+ T‐cells producing variable amount of interferon‐gamma, interleukin‐4 and interleukin‐10. The two patients have had a periodontal disease whose anti–P. gingivalis immunoglobulin G titers were significantly elevated, and P. gingivalis was the one of the predominantly cultivable microorganisms. This is the first report of the successful establishment of P. gingivalis–specific T‐cell lines from atherosclerotic patients.
Functional characteristics of antibodies induced by Arg‐gingipain (HRgpA) and Lys‐gingipain (Kgp) from Porphyromonas gingivalis
Authors: Nakagawa T. , Sims T. , Fan Q. , Potempa J. , Travis J. , Houston L. , Page R. C.
Published: March 2003

Arginine‐specific gingipain (HRgpA) and lysine‐specific gingipain (Kgp), enzymes produced by Porphyromonas gingivalis, may be candidates for an anti–P. gingivalis vaccine. The purpose of our study was to determine whether HRgpA and Kgp have opsonic target sites and whether these sites are available and accessible on intact P. gingivalis cells. Rabbits were used to generate polyclonal antibodies to both proteins. Animals were immunized and immunoglobulin G (IgG) fractions were isolated from preimmune and immune sera. Functional characteristics of the antibodies were assessed by determining antibody titers by enzyme‐linked immunosorbent assay (ELISA), generating Western immunoblots, and measuring antibody enhancement of P. gingivalis opsonization, phagocytosis and killing by polymorphonuclear leukocytes (PMN) of intact cells of strains of P. gingivalis representative of the four serotypes. Strains studied included 33277 (serotype A), A7A1–28 (serotype B), W50 (serotype C) and 381 (serotype D). Both HRgpA and Kgp induced high titers of IgG antibody. Anti‐HRgpA and anti‐Kgp bound to both HRgpA and Kgp demonstrating a large proportion of shared antigenic epitopes. The two antibodies bound equally well to all four P. gingivalis serotypes with titers ranging from 77 to 205 ELISA units when compared to preimmune IgG set at 1 ELISA unit. The immunoblot patterns of binding of the two antibodies to HRgpA and Kgp and to sonicates of the four P. gingivalis serotypes were virtually identical. Both antibodies detected components in HRgpA at 27, 35 and 45 kDa and in Kgp at 27, 32, 35, 40 and 55 kDa. The antibodies also detected components at or near these same positions in addition to multiple high molecular mass components in the cell sonicates of P. gingivalis. Both proteins induced antibodies that significantly enhanced opsonization as assessed by chemiluminescence, with values ranging from 130 mV to 375 mV for anti‐HRgpA IgG and from 240 mV to 475 mV for anti‐Kgp IgG. Both antibodies significantly enhanced PMN‐mediated bacterial killing of the four P. gingivalis serotypes, although the percentage of killing varied among the serotypes (24–81% for anti‐HRgpA and 37–89% for anti‐Kgp). Thus, both HRgpA and Kgp express opsonic target sites and induce high titers of antibodies that opsonize and enhance killing of all four serotypes of P. gingivalis. These two proteins appear to be potential candidate antigens for an anti–P. gingivalis vaccine.
Actinobacillus actinomycetemcomitans in a rural adult population in southern Thailand
Authors: Dahlén G. , Widar F. , Teanpaisan R. , Papapanou P. N. , Baelum V. , Fejerskov O.
Published: March 2003

The prevalence of Actinobacillus actinomycetemcomitans isolates was examined in a rural population of southern Thailand. Sixty individuals aged 30–39 and 50–59 years were randomly selected from a group of 363 persons, living in four villages, who had been clinically examined previously. A subgingival plaque sample was taken with a curette from the mesial aspect of the two upper and lower first molars. Each sample was dispersed in 3.3 ml of VMGA III transport medium and spread onto Trypticase Soy Broth with Bacitracin and Vancomycin (TSBV)‐agar plates on the same day. After incubation in 10% CO2 for 5 days the plates were examined for typical A. actinomycetemcomitans colonies which were tested for catalase activity. Each strain was further tested for biochemical characteristics, serotyped against serotype‐specific antisera a–e and ribotyped after DNA digestion using the restriction endonucleases HindIII and EcoRI. For 53 (88%) of the 60 individuals, A. actinomycetemcomitans was present in at least one subgingival sample, which is considerably higher than the prevalence in Western European adults. In 11 individuals, two or three different strains were found. Serotypes a and c were the most prevalent, and serotype b was found only once among 46 tested isolates. Eleven ribotypes were found among the 46 strains. While the same ribotype could be found among individuals of the same village, no ribotype of A. actinomycetemcomitans was unique for individuals of any one village. The study demonstrated a high prevalence of A. actinomycetemcomitans among adults of the rural population of southern Thailand and indicates that this species is present as part of the resident oral flora in this population.
Persistence of oral Candida albicans carriage in healthy Portuguese schoolchildren followed for 3 years
Authors: Starr J. R. , White T. C. , Leroux B. G. , Luis H. S. , Bernardo M. , Leitao J. , Roberts M. C.
Published: March 2003

Little is known about carriage of Candida albicans, the predominant pathogenic yeast in oral infection, in children. We cultured buccal mucosal and gingival swabs from 150 Portuguese children to investigate the prevalence of C. albicans at baseline (before dental treatment), post‐treatment, and 12, 24, and 36 months post‐baseline. The children, aged 8 to 11 years at baseline, had no systemic disease or clinical symptoms of oral candidiasis. At each successive visit, respectively, 47, 32, 21, 27, and 28% of children were C. albicans positive, resulting in an almost 50% reduction in prevalence from baseline to post‐treatment (P < 0.0005). Children who carried C. albicans at one visit had 3 to 20 times greater odds of carrying C. albicans at another visit. C. albicans was cultured from 12 children at all time‐points and from 10 children at four time‐points. Children with oral C. albicans frequently maintained carriage over time, even with regular dental care.
Oral microbiota associated with hyposalivation of different origins
Authors: AlmståhI A. , Wikström M. , Stenberg I. , Jakobsson A. , Fagerberg‐Mohlin B.
Published: March 2003

We analysed and compared the oral microbial flora in four groups with hyposalivation caused by radiation therapy (RT), primary Sjögren's syndrome (pSS), medication or unknown factors (Unknown), or neuroleptic treatment (Neuro). A control group with normal salivary secretion was also included. The subjects included were 54 ± 8 years old and had 25 ± 4 teeth. We analysed their microflora in rinsing samples using a cultivation technique. A marked increase in Lactobacillus spp. and Candida albicans was characteristic of the RT group. In the pSS group, 85% of subjects had high numbers of mutans streptococci despite good oral hygiene, frequent dental visits and fluoride use. The Unknown group had an oral flora similar to that of the controls. In the Neuro group, with a stimulated secretion rate similar to that of the Unknown group, the numbers of aciduric and acidogenic microorganisms were close to those in the pSS group. The results indicate that changes in the oral microflora associated with hyposalivation are related to the reason for the hyposalivation rather than to the magnitude of the decrease in the salivary secretion rate.
Comparison of different techniques of quantitative PCR for determination of Streptococcus mutans counts in saliva samples
Authors: Rupf S. , Merte K. , Kneist S. , Al‐Robaiy S. , Eschrich K.
Published: March 2003

Saliva samples from 16 children with current caries activity were investigated for Streptococcus mutans using three different PCR techniques, and the results were compared with those of selective cultivation on mitis salivarius agar with bacitracin (MSB) (I, II: LightCycler – competitive PCR end‐point analysis; III: LightCycler – kinetic real‐time analysis; IV, V: block cycler – competitive PCR end‐point analysis; VI: cultivation on MSB agar). In groups I, III, IV and VI the saliva samples were analyzed directly. A DNA preparation before PCR with added competitors was carried out in groups II and V to exclude the influence of PCR inhibitors. The coefficients of correlation ranged from 0.97 to 0.98 among the competitive PCR methods, 0.8 to 0.85 for competitive vs. real‐time PCR and 0.5 to 0.65 for PCR vs. cultivation methods. Competitive PCR on the real‐time instrument was found to be more rapid than, comparably sensitive to, but less reproducible than competitive PCR on a block cycler.
Defective tumor necrosis factor‐α production in infants with cow’s milk allergy
Authors: Österlund Pamela , Järvinen Kirsi‐marjut , Laine Seppo , Suomalainen Hanna
Published: March 2003

As an aid to clarifying the role of immune mechanisms in the development of cow’s milk allergy (CMA) in suckling infants, we studied the capacity of peripheral blood mononuclear cells (PBMC) to produce tumor necrosis factor‐α (TNF‐α) in vitro. The study population consisted of 43 infants, aged 0.12–11.2 months; of these, 31 had challenge‐proven cow’s milk allergy manifested with either skin or gastrointestinal symptoms or both. In addition, 12 healthy infants were studied as controls. The spontaneous, unstimulated and mitogen‐induced production of TNF‐α and interferon‐γ (IFN‐γ) by isolated peripheral blood leukocytes was evaluated. TNF‐α and IFN‐γ production of PBMC was significantly lower in infants with cow’s milk allergy than in healthy children. Our results indicate that, in infants with CMA, the function of TNF‐α‐producing cells is defective. This might disturb the development of oral tolerance and thereby lead to cow’s milk allergy. These results may help to clarify the etiopathology of CMA.
Allergy to an occupational allergen (Sapelli wood) in a child
Authors: Matheu Victor , Huertas Julio , Martínez‐molero Isabel , Zubeldia Jose Manuel , Zapatero Lidia , Baeza Maria Luisa
Published: March 2003

The case is presented of a child who developed rhinoconjunctivitis, angioedema and asthma by sensitization to Sapelli wood, which was used in his father's carpentry. Positive skin‐prick test, high levels of specific immunoglobulin E by ELISA and a positive conjunctival challenge test suggest a type I hypersensitivity mechanism to this wood. This is the first case report of sensitization to Sapelli wood and it confirms that occupational allergen exposure as Sapelli wood may also cause sensitization in a child.
Anti‐neutrophil cytoplasmic antibodies directed against the bactericidal/permeability‐increasing protein (BPI) in pediatric cystic fibrosis patients do not recognize N‐terminal regions important for the anti‐microbial and lipopolysaccharide‐binding activity of BPI
Authors: Schultz Hendrik , Csernok Elena , Schuster Antje , Schmitz Thomas S. , Ernst Martin , Gross Wolfgang L.
Published: March 2003

This study was performed to examine the prevalence and clinical correlates of bactericidal/permeability‐increasing protein anti‐neutrophil cytoplasmic antibodies (BPI‐ANCA) in pediatric cystic fibrosis (CF) patients and to elucidate their possible role in CF pulmonary pathology. Sera of 27 CF patients were tested for ANCA by indirect immunofluorescence (IFT) and by enzyme‐linked immunosorbent assay (ELISA) for ANCA sub‐specificities. BPI‐ANCA were examined by using standard ELISA for BPI, lipopolysaccharide‐binding protein (LBP), and BPI/LBP fusion proteins to epitope map the main binding sites and look for cross‐reactivity with LBP. Pulmonary function and serum concentrations of total immunoglobulin G (IgG) were measured and infections were diagnosed. In addition, release of reactive oxygen species (ROS) by neutrophil granulocytes was measured after stimulation with monoclonal BPI‐ANCA. Using IFT, two patients showed atypical ANCA staining, six patients exhibited perinuclear ANCA staining, and no cytoplasmic ANCA staining was detected. Of 27 patients, 13 (48%) were BPI‐ANCA (IgG) positive, and three were also immunoglobulin A (IgA) BPI‐ANCA positive; one patient had ANCA against lactoferrin; and no proteinase 3 ANCA was detected in any of the patients. All BPI‐ANCA bound to the C‐terminal region of the molecule; none bound to the N‐terminus or to LBP. There was no significant correlation between clinical data and the occurence of BPI‐ANCA in this cross‐sectional study. Release of ROS from granulocytes was induced by monoclonal BPI‐ANCA. Activation of neutrophils and possible modulation of BPI‐mediated opsonophagocytosis and disposal of Gram‐negative bacteria and lipopolysaccharides by BPI‐ANCA raise the possibility that they contribute to pulmonary pathology in pediatric CF patients but intervention longitudinal studies in large groups of patients are needed to establish a causative association.
Definition of respiratory symptoms and disease in early childhood in large prospective birth cohort studies that predict the development of asthma
Authors: Koopman Laurens P. , Brunekreef Bert , De Jongste Johan C. , Neijens Herman J.
Published: March 2003

We have reviewed the prospective value of early respiratory symptoms for determining the risk of development of asthma later in life by using data from studies based on the general population, hospital population, and general practices. Although ‘wheezing’ in infancy generally has a good prognosis, it is an important risk factor for the development of asthma later in life. The prognostic value of ‘coughing’ and ‘shortness of breath’ in infancy for the later development of asthma is less clear. Despite the fact that no internationally accepted criteria for the definition of asthma in early childhood are available, many studies have been performed on this topic. We also investigated the outcome variables that were used to describe respiratory symptoms and disease in early childhood in the publications of nine large prospective birth cohort studies on the development of asthma. From seven of these studies, we reviewed the original questionnaires. We found that various studies used different outcome variables, but the data actually collected were similar. This is an important observation because it implies that comparisons between studies can be markedly improved by data sharing among investigators.
Developmental follow‐up in 15‐month‐old infants of asthmatic vs. control mothers
Authors: Schatz Michael , Harden Kathleen , Kagnoff Marcia , Zeiger Robert S. , Chilingar Linda
Published: March 2003

The purpose of this study was to evaluate mental and psychomotor development in infants of mothers whose asthma was actively managed during pregnancy and to compare the results with those froms infants of non‐asthmatic mothers. Bayley Scales were assessed at age 15 ± 3 months in 379 infants of asthmatic mothers and 376 control infants. Relationships were assessed between developmental indices and asthma severity, socioeconomic status, and infant prematurity. No significant differences in developmental indices were observed between infants of asthmatic mothers and control infants. No relationships were identified between developmental indices and maternal asthma severity. In the infants of both asthmatic and control mothers, a lower mean psychomotor developmental index was associated with birth weight < 2,500 g, and a lower mental developmental index with lower socioeconomic status. Hence, infants of asthmatic mothers whose asthma has been actively managed during pregnancy have developmental outcomes at 15 months of age that are similar to those of control infants.
Placebo‐controlled study of the mite allergen‐reducing effect of tannic acid plus benzyl benzoate on carpets in homes of children with house dust mite sensitization and asthma
Authors: Lau Susanne , Wahn Julia , Schulz Gabriele , Sommerfeld Christine , Wahn Ulrich
Published: March 2003

We studied the effect of a spray containing 1% benzyl benzoate, an acaricide, and 1% tannic acid (‘Lowal’; a protein‐denaturing substance), on concentrations of major allergens from house dust mite (HDM) species Dermatophagoides pteronyssinus and D. farinae (Der p 1 and Der f 1, respectively) in carpets. In a double‐blind, placebo‐controlled study with crossover design, 30 homes of children with HDM sensitization and asthma were included. All houses showed ≥ 400 ng/g of Der p 1 + Der f 1 in carpet dust. The first treatment was performed on day 0 (group 1 active treatment, n = 15; group 2 placebo treatment, n = 15). After 2 and 8 weeks, dust samples were collected for quantification of mite allergens. After a 2‐week washout period, the second treatment was performed (group 1 placebo treatment; group 2 active treatment). Again, carpet dust was collected after 2 and 8 weeks. Twenty‐two of 30 families completed the trial: 14/15 in group 1 and eight of 15 in group 2. On day 0, there was no significant difference in mite allergen exposure between group 1 and group 2 (1,498 vs. 2,239 ng/g of Der p 1 + Der f 1, respectively). In group 1, the geometric mean for the difference of mite allergen concentration comparing day 0 and week 6 was 196 ng/g (95% CI: −7,161 and 8,401) for the first treatment (active) and 15 ng/g (95% CI: −1,079 and 1,292) for the second treatment (control). In group 2, the difference was 66 ng/g (95% CI: −398 and 1,515) for the first treatment (control) and 609 ng/g (95% CI: 186 and 9,264) for the second treatment (active). Comparing placebo and active treatment in total, there was a significant decrease following placebo treatment after 14 days (p = 0.026). After 8 weeks, active treatment was superior to placebo treatment (p = 0.049), but the allergen reduction achieved was < 20% (median 1,500 ng/g on day 0 vs. 1,250 ng/g after 8 weeks). We conclude that the slight mite allergen reduction on carpets achieved by the treatment with ‘Lowal’ is unlikely to achieve worthwhile clinical benefit either in the treatment of mite‐sensitive patients or in primary or secondary prophylaxis.
Auxiliary partial orthotopic liver transplantation for acute liver failure in two children
Authors: Rodeck Burkhard , Kardorff Rüdiger , Melter Michael , Schlitt Hans J. , Oldhafer Karl J.
Published: March 2003

Abstract: Acute liver failure in children and adults is associated with a high mortality rate. At present the treatment of choice is orthotopic whole‐liver transplantation. However, allogeneic liver transplantation necessitates lifelong immunosuppressive therapy, which is associated with substantial risks to the patient. Temporary auxiliary partial orthotopic liver transplantation has been developed recently as an alternative, enabling the native liver to regenerate while avoiding the risks of long‐term immunosuppressive treatment. Here we describe two cases of partial orthotopic liver transplantation in children. Auxiliary partial orthotopic liver transplantation was performed in two boys (5 and 6 years old) suffering from acute liver failure of unknown origin. The native left lateral liver lobes (segment II and II) were removed and replaced by left lateral liver grafts from young blood‐group‐compatible adults. In the first child the native liver, which was 80% necrotic at time of transplantation, showed regeneration within two weeks and the partially necrotic graft could be surgically removed on day 15 after auxiliary transplantation. Four years after transplantation, the child is in excellent condition with normal liver function and does not require any treatment. In the second case the native liver (90% necrotic at time of transplantation) regenerated within 6 weeks of transplantation, at which time the transplanted liver was removed. The patient developed aplastic anemia and died 2 months after transplantation from candida sepsis. The conclusion was that auxiliary partial liver transplantation in childhood provides a valuable option to maintain liver function in acute liver failure until functional recovery of the native liver. The main advantage over whole‐liver transplantation is the chance to avoid lifelong immunosuppression. However, there is a higher surgical risk. Therefore, auxiliary transplantation should be considered carefully in every case of acute liver failure in children.
Long‐term results of pre‐emptive liver transplantation in primary hyperoxaluria type 1
Authors: Nolkemper Daniela , Kemper Markus J. , Burdelski Martin , Vaismann Irith , Rogiers Xavier , Broelsch Christoph E. , Ganschow Rainer , Müller‐Wiefel Dirk E.
Published: March 2003

Abstract: In primary hyperoxaluria type 1 (PH 1), deficiency or mis‐targeting of hepatic alanine glyoxylate aminotransferase (AGT) results in over‐production of oxalate and hyperoxaluria, leading to nephrocalcinosis and development of end‐stage renal disease (ESRD) in the majority of patients. Renal transplantation (Tx) alone carries a high risk of disease recurrence as the metabolic defect is not cured. Therefore, combined liver/kidney Tx is recommended for patients with ESRD. An alternative approach is to cure PH 1 by pre‐emptive isolated liver Tx (PLTx) before ESRD has occurred, but this approach has been carried out only occasionally and there are no uniformly accepted recommendations concerning the timing of this procedure. We report follow‐up 3–5.7 yr after performing successful PLTx in four children (at the age of 3–9 yrs) with PH 1 prior to the occurrence of ESRD (glomerular filtration rate [GFR] range 27–98 mL/min/1.73 m2). There was no mortality or long‐term morbidity associated with the Tx procedure. Plasma and urinary oxalate levels normalized rapidly within 4 weeks, and renal function did not deteriorate under immunosuppression, even in one patient with advanced chronic renal failure (GFR 27 mL/min/1.73 m2) who showed a stable course for more than 5.7 yrs. Although treatment must be individualized in this severe metabolic disorder, and PLTx has to be regarded as an invasive procedure, we consider that PLTx should be offered and considered early in the course of PH 1. PLTx cures the metabolic defect in PH 1 and can help to prevent, or at least delay, the progression to ESRD and systemic oxalosis.
Granulocyte colony‐stimulating factor alone at 20 µg/kg vs. 10 µg/kg for peripheral blood stem cell mobilization in children
Authors: Halle P. , Kanold J. , Rapatel C. , Boiret N. , Berger M. , Stephan J. L. , Al Buisson E. , Tournilhac O. , Bonhomme J. , Deméocq F.
Published: March 2003

Abstract: Mobilization of peripheral blood stem cells (PBSC) by granulocyte colony‐stimulating factor (G‐CSF), at 10 µg/kg/day vs. 20 µg/kg/day (in 42 and 29 patients, respectively), was compared in children with solid tumors or leukemias. During mobilization, differences were noted in the peak values of CD34+ cells in peripheral blood (PB) in these two groups (median 28 × 106/L for 10 µg/kg/day vs. 61 × 106/L for 20 µg/kg/day; p = 0.025). Similar numbers of progenitor cells were harvested for the two concentrations of G‐CSF. However, similar CD34+ cell levels in the leukapheresis product were obtained after only the third dose of G‐CSF at 20 µg/kg/day compared with the fourth dose of G‐CSF at 10 µg/kg/day (1.7 and 1.2 × 106 CD34+ cells/kg/one patient's blood volume processed, respectively). Of note is the impact of diagnosis on PB CD34+ cell levels. We conclude that, in children, mobilization with G‐CSF at 20 µg/kg/day could minimize the duration of priming but not reduce the number of leukaphereses. Thus, the impact on outcome, clinical practice, bed utilization, and health economics is uncertain.
Mycophenolate, tacrolimus and post‐transplant lymphoproliferative disorder: A report of the North American Pediatric Renal Transplant Cooperative Study
Authors: Dharnidharka Vikas R. , Ho Ping‐Leung , Stablein Donald M. , Harmon William E. , Tejani Amir H.
Published: March 2003

Abstract: Tacrolimus (FK506) and mycophenolate mofetil (MMF) have been reported to increase PTLD risk. The NAPRTCS registry database now has several years of data on FK506 and MMF use in pediatric kidney transplantation. We analyzed the data registry to determine if the risk of PTLD was enhanced by the use of MMF or tacrolimus in initial immunosuppression. Data on day 30 therapy in the PTLD group were compared to corresponding data in patients who did not develop PTLD. Data were analyzed using SAS software and a log‐rank test for significance. As of October 2000, there were 108 cases of PTLD in 6720 total transplants(1.60%). The use of MMF at day 30 was not a significant risk factor (0. 78% PTLD rate vs. 1.78% in cohort, RR = 1.05, p = 0.89). The relationship of FK506 with PTLD was linked to transplant era, 1987–95 or 1996–2000. In the earlier era, use of FK506 at day 30 was associated with PTLD (seen in 7/15 patients given FK506, RR = 47.7, p < 0.001). However, in the more recent era, there was no such significant association (seen in 3/313 patients given FK506, RR = 1.28, p = 0.692). There have been no cases of PTLD in 197 patients who received both FK506 and MMF at day 30. We conclude that FK506 and MMF use are not currently associated with increased risk of PTLD in pediatric kidney transplants.
Efficacy and pharmacokinetics of tacrolimus oral suspension in pediatric liver transplant recipients
Authors: Reding Raymond , Sokal Etienne , Paul Killick , Janssen Magda , Evrard Veerle , Wilmotte Léon , Chardot Christophe , Otte Jean‐Bernard , Wallemacq Pierre
Published: March 2003

Abstract: The use of tacrolimus in small pediatric graft recipients may require the availability of a suspension formulation for appropriate dose titration and easier administration. The pharmacokinetics (Pk) of an extemporaneously prepared oral suspension of tacrolimus (OST) was investigated in 15 pediatric liver transplant recipients, and was compared with the corresponding data with tacrolimus capsules (TC). Graft and patient survival rates were 100%. Acute rejection and steroid‐resistant rejection were encountered in 9/15 and 3/15 patients, respectively. Comparison of Pk data showed a lower oral absorption of OST when compared with TC. No significant correlation could be made between the Pk parameters and rejection. Despite the lower bioavailability of OST when compared with TC, the rejection incidence was similar with both formulations (60% vs. 55%, respectively). Accordingly, the use of OST may constitute an alternative option for tacrolimus administration in low body weight organ recipients, to allow dosage titration in the early post‐transplant weeks.
Adventitious rooting in hypocotyls of sunflower (Helianthus annuus) seedlings. IV. The role of changes in endogenous free and conjugated indole‐3‐acetic acid
Authors: Liu Jin‐Hao , Reid David M.
Published: March 2003

We have studied the role of endogenous auxin on adventitious rooting in hypocotyls of derooted sunflower (Helianthus annuus L. var. Dahlgren 131) seedlings. Endogenous free and conjugated indole‐3‐acetic acid (IAA) were measured in three segments of hypocotyls of equal length (apical, middle, basal) by using gas chromatography‐mass spectrometry with [13C6]‐IAA as an internal standard. At the time original roots were excised (0 h), the free IAA level in the hypocotyls showed an acropetally decreasing gradient, but conjugated IAA level increased acropetally; i.e. free to total IAA ratio was highest in the basal portion of hypocotyls. The basal portion is the region where most of root primordia were found. Some primordia were seen in this region within 24 h after the roots were excised. The quantity of free IAA in the middle portion of the hypocotyl increased up to 15 h after excision and then decreased. In this middle region there were fewer root primordia, and they could not be seen until 72 h. In the apical portion the amount of free IAA steadily increased and no root primordia were seen by 72 h.

Surgical removal of various parts of the hypocotyl tissues caused adventitious root formation in the hypocotyl regions where basipetally transported IAA could accumulate. Reduction in the basipetal flow of auxin by N‐1‐naphthylphthalamic acid and 2,3,5‐tri‐iodobenzoic acid resulted in fewer adventitious roots. The fewest root primordia were seen if the major sources of endogenous auxin were removed by decapitation of the cotyledons and apical bud. Exogenous auxins promoted rooting and were able to completely overcome the inhibitory effect of 2,3,5‐tri‐iodobenzoic acid. Exogenous auxins were only partially able to overcome the inhibitory effect of decapitation. We conclude that in sunflower hypocotyls endogenously produced auxin is necessary for adventitious root formation. The higher concentrations of auxin in the basal portion may be partially responsible for that portion of the hypocotyl producing the greatest number of primordia. In addition to auxins, other factors such as wound ethylene and lowered cytokinin levels caused by excision of the original root system cuttings must also be important.
Somatic embryo development in carrot is associated with an increase in levels of S‐adenosylmethionine, S‐adenosylhomocysteine and DNA methylation
Authors: Munksgaard Dorte , Mattsson Ole , Okkels Finn Thyge
Published: March 2003

Almost homogeneous populations representing different developmental stages of somatic embryos (globular, torpedo‐shaped, plantlets) and vacuolated cells were obtained from a cell suspension culture of carrot. The concentrations of S‐adenosylmethionine (SAM), S‐adenosylhomocysteine (SAH) and methylated DNA were determined in embryos at different developmental stages and were found to increase during somatic embryogenesis. The highest increase during embryogenesis was a 5‐fold increase in the level of SAM. A considerable increase in the methylation index (SAM/SAH ratio) was also found. We propose that the levels of SAM and SAH may be involved in the control of somatic embryogenesis by affecting the level of DNA methylation, which in turn might cause differential changes in gene activation. An increase in the level of SAM may be a prerequisite for progression of embryogenesis and the development of complete embryos.
Stress‐induced degradation of the photosynthetic apparatus is accompanied by changes in thylakoid protein turnover and phosphorylation
Authors: Dannehl Heidrun , Herbik Alexandra , Godde Doris
Published: March 2003

Development of chlorosis and loss of PSII were compared in young spinach plants suffering under a combined magnesium and sulphur deficiency. Loss of chlorophyll could be detected already after the first week of deficiency and preceded any permanent functional inhibition of PSII as detected by changes in the chlorophyll fluorescence parameter Fv/Fm. A substantial decrease in Fv/Fm was observed only after the second week of deficiency. After 4 weeks, the plants had lost about 70% of their original chlorophyll content, but fluorescence data indicated that 80% of the existing PSII centers were still capable of initiating photosynthetic electron transport. The degradation of the photosynthetic apparatus without loss of PSII activity was due to changes in protein turnover, especially of the PSII D1 reaction center protein. Already by day 7 of deficiency, a 1.4‐fold increase in D1 protein synthesis was observed measured as incorporation of 14C‐leucine. Immunological determination by western‐blotting did not reveal a change in D1 protein content. Thus, D1 protein was also degraded more rapidly. The increased turnover was high enough to prevent any loss or inhibition of PSII. After 3 weeks, D1 protein synthesis on a chlorophyll basis was further increased by a factor of 2. However, this was not enough to prevent a net loss of D1 protein of about 70%. Immunological determination revealed that together with the D1 protein also other polypeptides of PSII became degraded. This process prevented a large accumulation of photo‐inactivated PSII centers. However, it initiated the breakdown of the other thylakoid proteins, especially of LHCII, resulting in the observed chlorosis. Together with the change in protein turnover and stability, a characteristic change in thylakoid protein phosphorylation was observed. In the deficient plants steady state phosphorylation of both LHCII and PSII proteins was increased in the dark. In the light phosphorylation of PSII proteins was stimulated and after 3 weeks of deficiency was even higher in the deficient leaves than in the control plants. In contrast, the phosphorylation level of LHCII decreased in the light and could hardly be detected after 3 weeks of deficiency. Phosphorylation of the reaction center polypeptides presumably increased their stability against proteolytic attack, whereas phosphorylated LHCII seems to be the substrate for proteolysis.