bluuebunny/crossref_metadata_2025 huggingface dataset:
Conserved TCR β chain usage in reactive arthritis; evidence for selection by a putative HLA‐B27‐associated autoantigen
Authors:   E. May,   N. Dulphy, Frauendorf E. ,   R. Duchmann,   P. Bowness, Lopez de Castro J.A. ,   A. Toubert, Märker‐Hermann E.
Published: March 2003

Abstract: Previous work suggested that expanded CD8+ T‐cell clones in the synovial fluid (SF) of HLA‐B27+ patients with reactive arthritis (ReA) preferentially use the T‐cell receptor variable region (TCRBV) 1, similar CDR3 sequences, and joining region (BJ) 2S3. To determine the range of conservation and disease‐specificity of CDR3‐sequences, we analyzed the TCRBV1‐J2S3 repertoire from 33 healthy HLA‐B27+ individuals, patients with various types of spondyloarthropathies (SpA), and with rheumatoid arthritis (RA) by CDR3‐spectratyping. After collection and database submission of all available TCRB‐CDR3 from HLA‐B27‐restricted or SpA‐derived T cells, we systematically screened the entire human sequence database for sequences similar to the B27/SpA‐related CDR3. Spectratyping revealed expanded T cell clones using conserved TCRBV1J2S3 in the SF from 5/6 of the patients with acute ReA but not among the controls. In database searches, 50 HLA‐B27 or SpA‐related CDR3‐sequences generated similar clusters of matched sequences, and matched reciprocally. Identical or closely related sequences were identified in 15 different individuals and a canonical ReA‐associated TCRB was defined [BV1‐CASSVG(V/I/L)(Y/F)STDTQYF‐J2S3]. All but one patient‐derived conserved sequences originated from acute stage ReA‐patients, and were not present among ∼3800 other human TCRB sequences in the database. Five of the conserved sequences originated from T cell clones that recognized uninfected cells in an HLA‐B27‐restricted fashion, implying a role of HLA‐B27‐restricted CD8+ T cells specific for a ubiquitous self‐ or cross‐reactive microbial determinant in the early phase of ReA. Related sequences were independently identified in four different laboratories. The consensus TCRB motif could be a helpful diagnostic marker in HLA‐B27‐associated ‘undifferentiated arthritis’.
Identification of clusterin sequences mediating renal tubular cell interactions
Authors: Silkensen J.R. , Skubitz K.M. , Rosenberg M.E. , Skubitz A. P. N.
Published: March 2003

Abstract: Expression of the glycoprotein clusterin is markedly increased following tissue injury. One function of clusterin is to promote cell interactions which are perturbed in these pathologic settings. Clusterin causes cell aggregation and adhesion in vitro yet the molecular mechanism for this effect is not known. In order to identify the active site(s) of clusterin, 34 peptides, each 15 amino acid residues in length, were synthesized from hydrophilic regions of human clusterin. When studied individually, none of the peptides caused aggregation of LLC‐PK1 cells, a porcine renal epithelial cell line. However, two out of the 34 peptides inhibited clusterin‐induced cell aggregation in a dose‐dependent manner. Scrambled versions of these two ‘active’ peptides did not inhibit cell aggregation. Seven peptides promoted cell adhesion. In conclusion, these findings provide evidence for novel amino acid sequences mediating clusterin‐induced renal cell interactions.
The metal binding properties of the CCCH motif of the 50S ribosomal protein L36 from Thermus thermophilus
Authors: Boysen R.I. , Hearn M. T. W.
Published: March 2003

Abstract: The TthL36 protein of the 50S ribosomal proteins from Thermus thermophilus has been found to contain the rare C(Xaa)2C(Xaa)12C(Xaa)4H (CCCH) sequence motif, a zinc finger binding motif, which for other zinc finger proteins is known to cleave RNA hairpins. In order to investigate the metal‐binding properties of this T. thermophilus TthL36 protein, the core 26‐mer polypeptide containing this CCCH motif was prepared by solid‐phase peptide synthesis methods using Fmoc chemistry, purified by preparative RP‐HPLC and characterized by circular dichroism, high‐performance capillary zone electrophoresis and electrospray ionization mass spectrometry. Reaction of the acetamidomethyl (Acm)‐protected polypeptide with iodine under acidic conditions resulted in the formation of the fully de‐protected polypeptide. Of interest, the results demonstrate that the standard Acm‐deprotection method with the synthetic TthL36 polypeptide using mercuric acetate in the presence of a large excess of 2‐mercaptoethanol resulted in preferential formation of a very stable mercuro‐polypeptide complex. The properties of the Acm‐deprotected polypeptide in the presence of different metal ions were also investigated by spectroscopic methods. The results confirm that this TthL36 polypeptide containing the CCCH motif binds metal ions with different affinities, namely in the order Co(II) > Hg(II) > Zn(II).
Synthesis, conformational studies and biological activity of Να‐mono‐biotinylated rat relaxin*
Authors: Mathieu M.N. , Wade J.D. , Tregear G.W. , Bond C.P. , Summers R.J. , Catimel B. , Nice E.C. , Otvos L.
Published: March 2003

Abstract: Biotin−avidin immobilization can be a useful tool in structure–function studies of hormone receptors. A crucial step is the preparation of a specifically biotinylated hormone that is able to bind to its receptor while leaving the biotin group free for interaction with avidin. The receptor for relaxin, an ovarian peptidic hormone produced during pregnancy, has not yet been isolated. We therefore undertook to prepare a specifically monobiotinylated rat relaxin for use in ligand‐searching strategies. Rat relaxin is a convenient analogue because reliable bioassays exist, thus allowing assessment of the effect of N‐biotinylation on bioactivity. To help improve the yield of the two‐chain, three‐disulfide bond rat relaxin, 2‐hydroxy‐4‐methoxybenzyl (Hmb) backbone protection was used during the solid‐phase assembly of the B‐chain to help prevent any possible chain aggregation. As a final step, while the protected peptide was still on the resin, the biotin label was introduced at the N‐terminus of the B‐chain using standard coupling protocols. The chain combination with the A‐chain was accomplished in reasonable yield. Secondary structural measurements demonstrated that the biotin caused the starting B‐chain to adopt a more ordered conformation. The labelled synthetic relaxin exhibited similar circular dichroism spectra to native and synthetic single B‐chain peptides. In addition, the biotinylated relaxin showed no significant difference in its chronotropic activity in the rat isolated heart assay compared with the native peptide. Biosensor studies showed that antibody recognition was retained upon attachment of the synthetic relaxin to the streptavidin‐derivatized surface.
The development of highly efficient onium‐type peptide coupling reagents based upon rational molecular design
Authors: Li P. , Xu J.C.
Published: March 2003

Abstract  Novel and highly efficient immonium‐, pyridinium‐ and thiazolium‐type peptide coupling reagents, such as BOMI, BDMP, BPMP, BEP, FEP, BEPH, FEPH and BEMT, were developed by rational modifying of the molecular structures of commonly used uronium‐type reagents. The high efficiency of these onium salts has been evaluated and proven by model reaction tests and the successful synthesis of various oligopeptides and biologically active peptides, both in solution and in the solid‐phase, for example Leu‐enkephalin, the pentapeptide moiety of Dolastatin 15 and the immunosuppressive undecapeptide cyclosporin O. Based upon these results, the relationship between the molecular structure and the capability of onium‐type peptide coupling reagents was studied. A preliminary guideline for the molecular design of onium‐type coupling reagents was developed.
Spermidine as a potential biosynthetic precursor to the 1,5‐diazabicyclo[4:3:0]nonene residue in the efrapeptins
Authors: Uma M.V. , Balaram P. , Sudha R.
Published: March 2003

Abstract: Efrapeptins are a group of microheterogeneous polypeptide antibiotics produced by the fungus Tolypocladium niveum, which are potent inhibitors of mitochondrial F1‐ATPase. Efrapeptins contain an unusual 1,5‐diazabicyclo[4:3:0]nonene (DBN) residue at the C‐terminus. This study is driven by the hypothesis that the DBN residue could, in principle, arise by oxidative cyclization of a spermidine moiety. Electrospray ionization mass spectrometry of the peptide antibiotics ‘elvapeptins’ from T. niveum establishes the presence of a C‐terminal spermidine residue. Conversion of elvapeptins to efrapeptins by CuCl/pyridine demonstrates the transformation of the spermidine residue to the 1,5‐diazabicyclo[4:3:0]nonene system by oxidative cyclization.
Antimicrobial peptides from the skin of the Japanese mountain brown frog, Rana ornativentris
Authors: Kim J.B. , Conlon J.M. , Iwamuro S. , Knoop F.C.
Published: March 2003

Abstract: Six peptides with antimicrobial activity were isolated from an extract of freeze‐dried skin of the Japanese mountain brown frog Rana ornativentris. Two structurally related peptides (brevinin‐20a GLFNVFKGALKTAGKHVAGSLLNQLKCKVSGGC, 11 nmol/g dried tissue, and brevinin‐20b GIFNVFKGALKTAGKHVAGSLLNQLKCKVSGEC, 170 nmol/g) belong to the brevinin‐2 family, previously identified in Asian and European, but not North American, Ranid frogs. Four peptides (temporin‐1Oa FLPLLASLFSRLL.NH2, 13 nmol/g; temporin‐1Ob FLPLIGKILGTI L.NH2, 350 nmol/g; temporin‐1Oc FLPLLASLFSRLF.NH2, 14 nmol/g; and temporin‐1Od FLPLLASLFSGLF.NH2, 8 nmol/g) are members of the temporin family first identified in the European common frog Rana temporaria but also found in the skins of North American Ranids. The brevinin‐2 peptides showed broad‐spectrum activity against the gram‐positive bacterium, Staphylococcus aureus, the gram‐negative bacterium, Escherichia coli and the yeast Candida albicans, whereas the temporins showed potent activity only against S. aureus. The brevinins and temporins belong to the class of cationic antimicrobial peptides that adopt an amphipathic α‐helical conformation but it is significant that temporin‐1Od, which lacks a basic amino acid residue, is still active against S. aureus (minimum inhibitory concentration=13 µm compared with 2 µm for temporin‐1Oa). This suggests that strong electrostatic interaction between the peptide and the negatively charged phospholipids of the cell membrane is not an absolute prerequisite for antimicrobial activity.
Synthesis and biological activities of new side chain and backbone cyclic bradykinin analogues
Authors: Schumann C , Seyfarth L , Greiner G , Reissmann S , Paegelow I.
Published: March 2003

Abstract:A series of conformationally constrained cyclic analogues of the peptide hormone bradykinin (BK, Arg‐Pro‐Pro‐Gly‐Phe‐Ser‐Pro‐Phe‐Arg) was synthesized to check different turned structures proposed for the bioactive conformation of BK agonists and antagonists. Cycles differing in the size and direction of the lactam bridge were performed at the C‐ and N‐terminal sequences of the molecule. Glutamic acid and lysine were introduced into the native BK sequence at different positions for cyclization through their side chains. Backbone cyclic analogues were synthesized by incorporation ofN‐carboxy alkylated andN‐amino alkylated amino acids into the peptide chain. Although the coupling of Fmoc‐glycine to theN‐alkylated phenylalanine derivatives was effected with DIC/HOAt in SPPS, the dipeptide building units with more bulky amino acids were pre‐built in solution. For backbone cyclization at the C‐terminus an alternative building unit with an acylated reduced peptide bond was preformed in solution. Both types of building units were handled in the SPPS in the same manner as amino acids. The agonistic and antagonistic activities of the cyclic BK analogues were determined in rat uterus (RUT) and guinea‐pig ileum (GPI) assays. Additionally, the potentiation of the BK‐induced effects was examined. Among the series of cyclic BKagonistsonly compound3with backbone cyclization between positions 2 and 5 shows a significant agonistic activity on RUT. To study the influence of intramolecular ring closure we used an antagonistic analogue with weak activity, [d‐Phe7]‐BK. Side chain as well as backbone cyclization in the N‐terminus of [d‐Phe7]‐BK resulted in analogues with moderate antagonistic activity on RUT. Also, compound18in which a lactam bridge between positions 6 and 9 was achieved via an acylated reduced peptide bond has moderate antagonistic activity on RUT. These results support the hypothesis of turn structures in both parts of the molecule as a requirement for BK antagonism. Certain active and inactive agonists and antagonists are able to potentiate the bradykinin‐induced contraction of guinea‐pig ileum.
A new B‐chain mutant of insulin: comparison with the insulin crystal structure and role of sulfonate groups in the B‐chain structure
Authors: Dupradeau F.‐Y. , Le Flem G. , Richard T. , Monti J.‐P. , Oulyadi H. , Prigent Y.
Published: March 2003

Abstract: The solution structure of a new B‐chain mutant of bovine insulin, in which the cysteines B7 and B19 are replaced by two serines, has been determined by circular dichroism, 2D‐NMR and molecular modeling. This structure is compared with that of the oxidized B‐chain of bovine insulin [Hawkins et al. (1995) Int. J. Peptide Protein Res.46, 424–433]. Circular dichroism spectroscopy showed in particular that a higher percentage of helical secondary structure for the B‐chain mutant is estimated in trifluoroethanol solution in comparison with the oxidized B‐chain. 2D‐NMR experiments confirmed, among multiple conformations, that the B‐chain mutant presents defined secondary structures such as a α‐helix between residues B9 and B19, and a β‐turn between amino acids B20 and B23 in aqueous trifluoroethanol. The 3D structures, which are consistent with NMR data and were obtained using a simulated annealing protocol, showed that the tertiary structure of the B‐chain mutant is better resolved and is more in agreement with the insulin crystal structure than the oxidized B‐chain structure described by Hawkins et al. An explanation could be the presence of two sulfonate groups in the oxidized insulin B‐chain. Either by their charges and/or their size, such chemical groups could play a destructuring effect and thus could favor peptide flexibility and conformational averaging. Thus, this study provides new insights on the folding of isolated B‐chains.
Synthesis and conformational analysis of a coumarinic acid‐based cyclic prodrug of an opioid peptide with modified sensitivity to esterase‐catalyzed bioconversion
Authors: Ouyang H. , Borchardt R.T. , Siahaan T.J. , Vander Velde D.G.
Published: March 2003

Abstract: The coumarinic acid‐based cyclic DADLE (H‐Tyr‐D‐Ala‐Gly‐Phe‐D‐Leu‐OH) prodrug 1a exhibited more favorable physicochemical properties than did DADLE for permeation across the intestinal mucosa. However, prodrug 1a, whose bioconversion to DADLE was slow, was subject to extensive biliary clearance when administered to rats in vivo. To increase the rate of esterase‐catalyzed bioconversion of prodrug 1a, thus decreasing its biliary clearance, the oxymethyl‐modified prodrug 1, in which an aldehyde equivalent is inserted between the phenolic group of the promoiety and the carboxylic acid group of the peptide, was synthesized from benzofuran‐2‐carboxylic acid 16 via a nine‐step procedure. Briefly, phenacyl‐protected 3‐(2‐hydroxyphenyl)‐propynoic acid 17 was coupled with Boc‐d‐Leu‐OCH2I 5 to give the intermediate 18, which was further elaborated and conjugated with tetrapeptide 4 to give linear precursor 2. Precursor 2 was then deprotected and cyclized to obtain compound 1 using a high dilution technique. In an attempt to investigate the effect of the physicochemical properties and the conformation of prodrug 1 on its permeation characteristics, we calculated its physicochemical parameters and determined its solution conformation using spectroscopic techniques (CD and NMR) and molecular dynamics simulations. Prodrug 1 showed a cLogP value and a molecular size similar to that of prodrug 1a. The deconvoluted CD spectra indicated that prodrug 1 has more random component (71%) than prodrug 1a (42%). 2D‐NMR studies of prodrug 1 showed no signals for amide–amide hydrogen interactions and few ROE cross‐peaks in ROESY spectra. Using distance restraints constructed from ROESY spectra, molecular dynamics simulations of prodrug 1 generated five conformation families. One family satisfied most of the distance restraints and all of the dihedral angles measured by NMR coupling constants. In summary, prodrug 1 showed favorable physicochemical properties for permeation of the intestinal mucosa. Prodrug 1 adopted a more random conformation in solution than prodrug 1a. These differences in solution conformation could affect the permeation of the prodrugs across the intestinal mucosa by passive diffusion and/or their ability to interact with efflux transporter(s) that would limit their transcellular permeation.
Enhanced antitumor activity and selectivity of lactoferrin‐derived peptides
Authors: Yang N. , Rekdal Ø. , Stensen W. , Svendsen J. S.
Published: March 2003

Abstract:  A number of peptide analogs derived from the N‐terminal α‐helical region of bovine lactoferrin (LFB 14–31), were designed in order to investigate how deviating numbers and positions of positively charged residues and numbers of aromatic residues affected their activity against prokaryotic, normal and transformed eukaryotic cells. Most of the LFB derivatives were highly active against both Escherichia coli and Staphylococcus aureus. The peptides were more active against the tumor cell lines MethA, HT‐29 and MT‐1 than normal eukaryotic cells. The peptides that were most active against the tumor cell lines had all cationic residues concentrated in one sector of the helical structure. These peptides were less selective against the tumor cell lines than against normal fibroblasts. Quantitative structure−activity relationship studies showed that certain structural parameters affected toxicity against the tumor cell lines more than against fibroblasts. Peptides encompassing these parameters were slightly less active against tumor cells, but gained significant selectivity.
Productive human immunodeficiency virus‐1 infection of epithelial cell lines of salivary gland origin
Authors: Han Y. , Ventura C. L. , Black K. P. , Cummins J. E. , Hall S. D. , Jackson S.
Published: March 2003

To ascertain whether epithelial cells of oral cavity origin may be infected with human immunodeficiency virus (HIV‐1), a study to determine susceptibility to infection of salivary gland epithelial cell lines (HSY and HSG) was undertaken. Because of the potential for oral‐genital transmission, an endometrial cell line, HEC‐1, was also studied. Epithelial cell monolayers were infected with cell‐free HTLVIIIB or a primary HIV‐1 isolate. Several lines of evidence indicated that inoculation of these cell lines with HIV‐1 led to productive infection: 1) p24 antigen was present in supernatants, with levels peaking on days 3–4; 2) provirus was found in cells by polymerase chain reaction; 3) virions present in supernatants were infectious as confirmed by coculture with the T‐lympho‐

blastoid line CEM‐NKr. Following a period of virus production, HIV‐1 entered a latency phase over 10 weeks. All epithelial cell lines were positive for galactosylceramide (GalC) and CXCR4. HSY was weakly positive for surface CD4, and also expressed mRNA for CD4 and CCR5, as did HEC‐1. Blocking studies indicated that anti‐GalC, but not anti‐CD4, significantly reduced productive infection, and that regulated on activation normal T cell expressed and secreted (RANTES) but not stromal cell–derived factor (SDF‐1) could partially block infection of the M‐tropic primary isolate. These results suggest that epithelial cells in the oral cavity and the genital tract might be targets of HIV‐1 and potentially serve as a mediator of systemic infection.
Polarization of Porphyromonas gingivalis–specific helper T‐cell subsets by prior immunization with Fusobacterium nucleatum
Authors: Choi J.‐I. , Borrello M. A. , Smith E. S. , Zauderer M.
Published: March 2003

Antigen‐specific T‐cell clones were obtained from mice immunized with Fusobacterium nucleatum ATCC 10953 and/or Porphyromonas gingivalis 381. 10 BALB/c mice per group were immunized with F. nucleatum followed by P. gingivalis, or with P. gingivalis alone by intraperitoneal injection of viable microorganisms. Spleen T cells were isolated and stimulated in vitro with viable P. gingivalis cells to establish P. gingivalis–specific T‐cell clones. T‐cell phenotypes and cytokine profiles were determined along with T‐cell responsiveness to F. nucleatum or P. gingivalis. Serum immunoglobulin G antibody titers to F. nucleatum or P. gingivalis were also determined by enzyme‐linked immunosorbent assay. All the T‐cell clones derived from mice immunized with F. nucleatum followed by P. gingivalis demonstrated Th2 subsets, while those from mice immunized with P. gingivalis alone demonstrated Th1 subsets based on the flow cytometric analysis and cytokine profiles. All T‐cell clones from both groups were cross‐reactive to both P. gingivalis and F. nucleatum antigens. Phenotypes of T‐cell clones were all positive for CD4. Mean post‐immune serum IgG antibody levels to F. nucleatum or P. gingivalis were significantly higher than the pre‐immune levels (P<0.05, P<0.01, respectively). There were no significant differences in the antibody titers between the two groups. It was concluded that P. gingivalis–specific T cells initially primed by cross‐reactive F. nucleatum antigens were polarized to Th2 subset, while T cells stimulated with P. gingivalis alone maintained the profile of Th1 subset.
Subgingival strains of Candida albicans in relation to geographical origin and occurrence of periodontal pathogenic bacteria
Authors: Hannula J. , Dogan B. , Slots J. , Ökte E. , Asikainen S.
Published: March 2003

Clonal diversity of subgingival yeast strains was determined in relation to geographical location and coexistence of selected periodontal pathogenic bacteria. A total of 60 dental patients from Finland, the United States and Turkey each contributed five Candida albicans isolates. C. albicans isolates were serotyped using slide agglutination and genotyped using polymerase chain reaction (PCR) amplification and a random sequence primer. In general, each study subject yielded C. albicans isolates belonging to the same serotype and genotype. C. albicans serotype A occurred more frequently in subjects from Finland and Turkey than in subjects from the United States. A total of 27 PCR‐based C. albicans genotypes were identified. One C. albicans genotype occurred with particularly high frequency in subjects from Turkey and another genotype in subjects from the United States. Relationships were identified between C. albicans serotypes and genotypes. Further studies are needed to determine environmental factors of importance for subgingival colonization and persistence of C. albicans.
Salivary calprotectin levels are raised in patients with oral candidiasis or Sjögren’s syndrome but decreased by HIV infection
Authors: Sweet S. P. , Denbury A. N. , Challacombe S. J.
Published: March 2003

Calprotectin levels were determined in whole saliva from patients predisposed to oral candidiasis due to HIV infection or Sjögren’s syndrome and from patients with candidiasis associated with various oral disorders (e.g. lichen planus, oral ulceration). Mean calprotectin levels were higher in whole saliva (2 μg/ml) than in parotid saliva (0.3 μg/ml). Oral candidiasis was associated with raised whole saliva calprotectin levels in all groups studied. HIV infection was associated with lower levels of salivary calprotectin, in the presence of high or low salivary Candida counts, although CD4+ lymphocyte counts did not significantly correlate with calprotectin concentrations. Calprotectin levels were elevated in saliva from Sjögren’s syndrome patients with oral candidiasis, consistent with mucosal transudation of calprotectin from inflamed mucosa and limited dilution due to decreased salivary flow rates. This study indicates that oral candidiasis is associated with raised calprotectin levels secondary to mucosal inflammation, but that diminution of this candidacidal factor due to HIV infection may be a predisposing factor in the aetiology of oral candidiasis.
Incidence of β‐lactamase production and antimicrobial susceptibility of anaerobic gram‐negative rods isolated from pus specimens of orofacial odontogenic infections
Authors: Kuriyama T. , Karasawa T. , Nakagawa K. , Yamamoto E. , Nakamura S.
Published: March 2003

The incidence of β‐lactamase production in anaerobic gram‐negative rods isolated from 93 pus specimens of orofacial odontogenic infections and the antimicrobial susceptibility of these isolates against 11 antibiotics were determined. A total of 191 anaerobic gram‐negative rods were isolated from the specimens. β‐Lactamase was detected in 35.6% of the black‐pigmented Prevotella and 31.9% of the nonpigmented Prevotella. However, no strains among the other species isolated produced β‐lactamase. Ampicillin, cefazolin and cefotaxime showed decreased activity as regards β‐lactamase‐positive Prevotella strains, whereas the activity of ampicillin/sulbactam, cefmetazole, and imipenem continued to be effective against such strains. All tested β‐lactam antibiotics were effective against Porphyromonas and Fusobacterium. Erythromycin showed decreased activity against nonpigmented Prevotella and Fusobacterium. Clindamycin, minocycline and metronidazole were powerful antibiotics against which anaerobic gram‐negative rods could be tested. The present study showed that β‐lactamase‐positive strains were found more frequently in the Prevotella strains than in any of the other species of anaerobic gram‐negative rods. The effectiveness of adding sulbactam to ampicillin was demonstrated, as well as the difference in cephalosporin activity against β‐lactamase‐positive strains.
The effect of amino acid spacers on the antigenicity of dimeric peptide–inducing cross‐reacting antibodies to a cell surface protein antigen of Streptococcus mutans
Authors: Oishi Y. , Onozuka A. , Kato H. , Shimura N. , Imai S. , Nisizawa T.
Published: March 2003

In the course of developing a synthetic peptide vaccine for dental caries, we identified a unique 13‐mer peptide named PAc(365–377), TYEAALKQYEADL, as a minimum peptide inducing cross‐inhibiting antibodies to a cell surface protein antigen (PAc) of Streptococcus mutans. However, the peptide could hardly induce the production of antibody in the absence of adjuvant. Thus using this peptide as a unit peptide, tandem constructs of dimeric unit peptide with or without spacer amino acid residues were synthesized, and their antigenicities were examined in B10.D2 mice. Significant augmentation of antigenicity was obtained in all of the dimeric unit peptides with spacers, especially for lysine spacers. In addition, analysis for cross‐reactivity of anti‐construct antibodies against a set of double valine‐substituted analogues of the unit peptide revealed that the di‐lysine spacer might be more effective in inducing the cross‐reacting antibodies to rPAc.
Increase ofCandidacell virulence by anticancer drugs and irradiation
Authors: Ueta E. , Tanida T. , Yoneda K. , Yamamoto T. , Osaki T.
Published: March 2003

The influence of anticancer drugs and irradiation onCandidacell proliferation, adherence to HeLa cells and susceptibility to antifungal drugs (amphotericin B and miconazole) and neutrophils were examined using twoCandida albicansstrains. After treatment with 5‐fluorouracil (25 μg/ml to 250 μg/ml), cis‐diammine‐dichloroplatinum (10 μg/ml to 100 μg/ml), peplomycin (0.5 μg/ml to 5 μg/ml) or137Cs (20 Gy to 40 Gy) for 3 days or more, survivingCandidacells proliferated more rapidly than did untreated control cells. Anticancer agent‐pretreatedCandidacells revealed an increased adhesion to HeLa cells corresponding to an increase of binding to the lectins. The concentration of half limited colony formation (IC50) of amphotericin B and miconazole was increased to near two‐fold that of the control by pretreatment ofCandidacells with the anticancer agents, except peplomycin, which only weakly increased IC50. In addition, the enolase andCandidaacid proteinase activities in the culture supernatants were increased by pretreatment with the drugs and irradiation. Correspondingly, survivingCandidacells after these treatments were resistant to neutrophils, with a reduction to half of the killing. These results indicate that anti‐cancer drugs and irradiation potentiate the virulence ofCandidacells, or they eliminateCandidacells with low virulence, thereby enhancing the risk of oral and systemic candidiasis.
Inhibitory effect of synthetic histatin 5 on leukotoxin from Actinobacillus actinomycetemcomitans
Authors: Murakami Y. , Xu T. , Helmerhorst E. J. , Ori G. , Troxler R. F. , Lally E. T. , Oppenheim F. G.
Published: March 2003

Actinobacillus actinomycetemcomitans is a gram‐negative bacterium strongly implicated in the pathogenesis of juvenile periodontitis. This periodontal pathogen synthesizes a leukotoxin that destroys human polymorphonuclear leukocytes (PMNs), and this toxin is thought to be responsible for the virulence of A. actinomycetemcomitans. It was therefore of interest to assess whether major virulence factors of periodontal pathogens were neutralized by salivary components. This study focuses on the effect of histatins, components of the nonimmune oral defense system, on leukotoxin activity. Leukotoxin was extracted with polymyxin B from freshly grown anaerobic cultures of A. actinomycetemcomitans strain Y4. PMNs isolated from blood of healthy human volunteers were incubated in a cytotoxicity assay containing PMNs (107 cells/ml) and leukotoxin preparation (0–500 µg/ml) in Hanks' balanced salt solution at 37°C for 0–120 min with or without synthetic histatin 5 (0–500 µM). Cytotoxicity was measured by release of lactate dehydrogenase (LDH) at different time intervals. Histatin 5 neutralized the toxic effect of the leukotoxin preparation in a concentration‐dependent manner, with an IC50 value of 150 µM. When PMNs were preincubated with histatin 5 (300 µM), washed and subsequently exposed to leukotoxin, no protective effect was observed. This observation suggests a mechanism of inhibition whereby histatin 5 either directly neutralizes the leukotoxin or interferes with the leukotoxin–PMN interaction. The inhibitory effect of histatin 5 on leukotoxic activity may suggest a new biological function of histatins in the oral cavity as a naturally occurring secondary antibiotic.
Characterization and expression of a novel Porphyromonas gingivalis outer membrane protein, Omp28
Authors: Slakeski N. , Margetts M. , Moore C. , Czajkowski L. , Barr I. G. , Reynolds E. C.
Published: March 2003

We report the characterization of a Porphyromonas gingivalis gene, designated omp28, encoding a protein that we have previously purified and characterized as a 28‐kDa outer membrane protein. The deduced amino acid sequence of the omp28 open reading frame displayed an outer membrane leader sequence and lipoprotein attachment site but did not exhibit any significant overall sequence identity with protein sequences in the databases. A small stretch of amino acids (19 residues) exhibits 50% sequence identity with a segment of a fimbrial protein from Dichelobacter nodosus involved in adhesion, suggesting that Omp28 may be a surface adhesin/receptor of P. gingivalis. Using the pET‐24 vector we expressed recombinant Omp28 (rOmp28) in Escherichia coli. Western blot analyses of purified rOmp28 with rabbit antisera to a P. gingivalis outer membrane preparation, protective rat anti‐whole P. gingivalis antisera and pooled human sera from chronic periodontitis patients showed that the recombinant was recognized by all antisera. Further, anti‐rOmp28 antisera exhibited strong reactivity with a panel of four laboratory strains and 10 clinical isolates of P. gingivalis from the United States, Sudan, Romania and Norway. These results suggest that Omp28 is expressed by a wide distribution of P. gingivalis strains.
Cytokine production induced by a 67‐kDa fimbrial protein from Porphyromonas gingivalis
Authors: Hamada N. , Watanabe K. , Arai M. , Hiramine H. , Umemoto T.
Published: March 2003

Fimbriae have been reported to play an important role in the adherence of Porphyromonas gingivalis to oral surfaces and possibly in triggering host responses. P. gingivalis ATCC 33277 has two distinctly different fimbriae expressed on the cell surface. The 67‐kDa fimbriae differ in size and antigenicity from the earlier reported FimA, a major 41‐kDa fimbrial component of P. gingivalis. Expression of the 67‐kDa fimbriae on the cell surface of a fimA mutant was investigated by electron microscopy. The 67‐kDa fimbrial protein was purified from the fimA mutant by sonication, precipitation, and chromatography on a DEAE Sepharose CL‐6B column. The N‐terminal amino acid sequence of the 67‐kDa fimbrillin was distinct from that of the 41‐kDa fimbrillin. Moreover, we have found that the 67‐kDa fimbrial protein from P. gingivalis ATCC 33277 induced IL‐1α, IL‐β, IL‐6 and TNFα cytokine expression in mouse peritoneal macrophages. These results suggest that P. gingivalis 67‐kDa fimbriae may play a part in the inflammatory response during the development of periodontal diseases.
Susceptibility of Porphyromonas gingivalis in biofilms to amoxicillin, doxycycline and metronidazole
Authors: Larsen T.
Published: March 2003

The susceptibility of Porphyromonas gingivalis to amoxicillin, doxycycline and metronidazole was determined by a standardized method taking into account the biofilm mode of growth of subgingival bacteria. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of 48‐h biofilms of P. gingivalis established on membrane filters in a Modified Robbins Device were determined by agar dilution. The results were compared to (i) conventional MIC determinations, (ii) the susceptibility of planktonic cultures with cell numbers equal to those of the biofilms, and (iii) results for detached biofilm cells. The MICs of the biofilms of the six reference strains and clinical isolates containing 107−8 cells/filter were much higher than the conventional MIC values. However, the MIC of planktonic cultures of equal cell numbers also increased, indicating that an inoculum effect is part of the explanation of the increased resistance of biofilms. Still, the MBCs of biofilms were 2–8 times, and those for doxycycline up to 64 times, greater than the MBC values for planktonic cultures.
Clinical pharmacology of the H1‐receptor antagonists cetirizine and loratadine in children
Authors: Simons F. Estelle R. , Johnston Lana , Simons Keith J.
Published: March 2003

H1‐receptor antagonists are widely used in children but are not as well‐studied in children as they are in adults. Our objective was to determine the onset and duration of action and the relative potency of the H1‐receptor antagonists cetirizine and loratadine in children. We performed a prospective, randomized, placebo‐controlled, double‐blind, crossover, single‐dose study of cetirizine and loratadine using suppression of the histamine‐induced wheal and flare as the primary outcome. In 15 allergic children, mean age 9 years, compared with baseline, cetirizine (10 mg) suppressed the wheals and flares significantly from 0.25 to 24 h, achieving nearly 100% of flare suppression from 2 to 24 h, inclusive, and loratadine (10 mg) suppressed the wheals and flares significantly from 0.75 to 24 h, inclusive. Cetirizine suppressed the wheals and flares significantly more than loratadine from 0.25 to 1 h, inclusive, and at 0.5, 1, 2, 3, 5, 6, 7, and 24 h, respectively. Placebo also suppressed the wheal and flare significantly at some assessment times. Cetirizine and loratadine both have excellent antihistaminic activity in children, with a rapid onset of action and a 24‐h duration of action in this population.
Progression from allergic sensitization to asthma
Authors: Warner J. O. , Pohunek P. , Marguet C. , Clough J. B. , Roche W. R.
Published: March 2003

Asthma is one of the atopic diseases strongly associated with allergy. High aeroallergen exposure in the immediate postnatal period has been associated with higher risk of sensitization and chronic asthma. It is proposed that following in utero allergen sensitization, postnatal high dose allergen exposure localizes inflammation to the airways. In association with adjuvantizing effects of some virus infections, eosinophils and neutrophils are recruited which contribute to epithelial damage and the initiation of the remodelling process. Eventually, the latter processes lead to sufficient airway narrowing to manifest as the first symptoms of asthma. Thus, the immunopathology of asthma is fully established by the time of first symptoms and future strategies will need to identify those at risk of developing the disease before irreversible changes in the airways are established.
Vitamin A status in children with asthma
Authors: Arora Praveen , Kumar Virendra , Batra Sanjay
Published: March 2003

Low vitamin A levels have been found in a number of diseases in children. The aim of this study was to examine the vitamin A status in children with asthma and to correlate the changes with severity of disease. Serum levels of vitamin A, retinol‐binding protein (RBP), and albumin were estimated in 35 asthmatic children (24 males) in the age group of 2–12 years (mean 5.89 years) and 29 controls (19 males). Both study and control groups were similar with respect to age, sex, and overall nutritional status. Twenty‐four children in the study group (68.6%) had moderate to severe persistent asthma and eight children had mild persistent asthma. Only three patients suffered from mild intermittent asthma. Vitamin A levels in children with asthma (mean ± SD 22.14 ± 5.38 µg/dl) were found to be significantly lower than their controls (mean ± SD 27.54 ± 4.83 µg/dl) (p = 0.0001). Age, age of onset of asthma, and gender had no correlation with serum vitamin A levels. Low serum vitamin A levels (< 20 µg/dl) were observed four times more commonly in the study group (28.6%) than controls (6.9%). Severity of asthma had a negative correlation with serum vitamin A levels (r = − 0.61, p = 0.0001). Children with severe persistent asthma had markedly low serum vitamin A levels (mean ± SD 13.42 ± 5.19 µg/dl) as compared with mild intermittent asthma (mean ± SD 24.61 ± 2.32 µg/dl). Therapeutic trials are needed to prove whether low vitamin A levels contribute to asthma severity and the clinical utility of vitamin A supplementation in asthmatic children.
Allergic conditions in 5–8‐year‐old Maltese schoolchildren: Prevalence, severity, and associated risk factors [ISAAC]
Authors: Montefort Stephen , Muscat Hugo Agius , Caruana Simone , Lenicker Herbert
Published: March 2003

Allergic conditions, especially asthma, seem to be increasingly common worldwide. The International Study of Asthma and Allergies in Childhood (ISAAC) was the first study carried out worldwide using standardized questionnaires in order to create a reliable global map of childhood allergy. The Maltese Islands were one of the centres that participated in this study and in this article the data obtained from 3,506 5–8‐year‐old children from 24 state schools (78.5% response rate), and also data obtained from some added ‘local’ questions addressed to the same children, were analyzed in order to evaluate the problem of allergic conditions in Maltese schoolchildren. Of the participants, 19.1% were wheezers ‘ever,’ while 8.8% were current wheezers. Of the latter, 15.9% experienced nocturnal wheezing at least once a week and 13.3% had a wheezing episode of sufficient severity to limit speech. Nasal problems were present in 23.4% of these children, and in 20.7% of all respondents these symptoms persisted up to the year of answering the questionnaire. Hay fever had been diagnosed in 14.7% of all the children. Seven per cent of respondents had a recurrent, itchy rash (suggestive of eczema) for at least 6 months of their lives and 5.5.% had it currently. The prevalence of wheezing and eczema were slightly lower than the global mean, unlike rhinitis which in Malta was commoner than the world average. Multiple variables, such as gender, breast‐feeding, passive smoking, family history of atopy, pets, soft furnishings, and living next to busy roads, were factors that affected the prevalence and severity of the allergic conditions studied. In conclusion, allergic conditions are very common in Maltese schoolchildren and cause great hardship to these same youngsters. The results of this study should serve as a stimulus to try to decrease this suffering through better management of these conditions, measures to control identified detrimental factors (such as passive smoking), and further research on asthma, allergic rhinitis and eczema.
Functional symptoms confused with allergic disorders in children and adolescents
Authors: Niggemann Bodo
Published: March 2003

The diagnosis of a functional respiratory disorder is sometimes difficult and time‐consuming, because the symptoms often resemble those of organic diseases. The most common entities are hyperventilation syndrome, psychogenic cough, sighing dyspnea, and vocal cord dysfunction. Typical signs are heavy breathing or dyspnea, cough or sneezing, various breathing sounds, tightness of the throat or chest, pain, and fear. Criteria for differentiation include the lack of nocturnal symptoms, the sudden occurrence, no typical trigger factors, the variable duration, a quick regression, and that symptoms do not respond to adequate pharmacotherapy and finally normal results of diagnostic work‐up. Therapeutic options comprise psychological intervention (by reassurance, relaxation techniques, and behaviour therapy) and physiotherapy (e.g. breathing therapy, voice training). Intensive efforts should be made to diagnose functional symptoms at an early stage because this will prevent stigmatization and fixation of symptoms and disease, and also prevent children from undergoing unnecessary and potentially harmful therapies.
Cumulative incidence of asthma and allergy in north‐Norwegian schoolchildren in 1985 and 1995
Authors: Selnes Anders , Bolle Roald , Holt Jan , Lund Eiliv
Published: March 2003

The prevalence of asthma and allergy in children is increasing. In order to investigate time trends, follow‐up studies conducted several years apart and with identical study designs are essential. We compared two identical, cross‐sectional and questionnaire‐based studies of asthma and allergy in north‐Norwegian schoolchildren (7–13 years of age). The first study was conducted in 1985 (n = 10,093) and the second in 1995 (n = 8,676). The cumulative incidence was as follows: diagnosed asthma, 8.6% in 1995 vs. 5.1% in 1985, relative risk (RR) = 1.71 (95% CI: 1.53–1.90); allergic rhinoconjunctivitis, 22.1% in 1995 vs. 16.4% in 1985, RR = 1.39 (95% CI: 1.31–1.47); and atopic dermatitis, 19.7% in 1995 vs. 13.2% in 1985, RR = 1.48 (95% CI: 1.39–1.58). The cumulative incidence of allergic rhinoconjunctivitis and atopic dermatitis was higher in children of Sami ethnicity than Norse ethnicity in the 1985 study. Furthermore, although not statistically significant, there was a trend towards a greater increase in the cumulative incidence of diagnosed asthma, symptoms of asthma, allergic rhinoconjunctivitis, and atopic dermatitis from 1985 to 1995 in children of Sami ethnicity than Norse ethnicity. We conclude that there has been a marked increase in the cumulative incidence of asthma and allergy prevalence among schoolchildren in northern Norway from 1985 to 1995.
Wheezing requiring hospitalization in early childhood: Predictive factors for asthma in a six‐year follow‐up
Authors: Kotaniemi‐Syrjänen Anne , Reijonen Tiina M. , Korhonen Kaj , Korppi Matti
Published: March 2003

Although asthma is common after wheezing in early childhood, the risk factors for and the prevention of later asthma are poorly understood. During the present follow‐up study, a range of possible predictive factors for school‐age asthma was evaluated. The study group consisted of 82 children hospitalized for wheezing at age < 2 years in 1992–93. The baseline data were collected on entry to the study. In 1999, the children were re‐examined at the median age of 7.2 years. A structured questionnaire was applied to chart the symptoms suggestive of asthma, and the children were examined clinically. An exercise challenge test, as well as skin prick tests (SPT) to common inhalant allergens, was performed. Asthma was present in 33 (40%) children, 30 (91%) having continuous medication for asthma. The significant asthma‐predictive factors, present on entry to the study, were blood eosinophilia (p = 0.0008), atopic dermatitis (p = 0.0089), elevated total serum immunoglobulin E (IgE) (p = 0.0452), and a history of earlier episodes of wheezing in infancy (p = 0.0468). SPT positivity in early childhood was also associated with school‐age asthma (p = 0.002). In contrast, respiratory syncytial virus (RSV) identification during the index episode of wheezing played a minor role as a predictive factor for asthma. In conclusion, if hospitalization for wheezing occurs in infancy, more than every third child will suffer from asthma at early school age; the risk is significantly increased with recurrent wheezing in infancy and the development of allergic manifestations.
Endocrine complications of bone marrow transplantation in children
Authors: Legault L. , Bonny Y.
Published: March 2003

Abstract: Eighty‐seven patients had a bone marrow transplantation (BMT) at our institution between 1980 and 1992. We wished to study the endocrine complications that accompany this procedure as long‐term survival is now much more common. Forty‐three patients were retrospectively available for review and their records were examined for evidence of thyroid, pubertal, and growth complications. Fifteen per cent of the patients showed evidence of thyroid involvement. Pubertal delay or gonadal damage was almost universal in pubertal‐aged girls treated with busulfan/cyclophosphamide. Gonadal involvement was more frequent in girls than in boys (70% vs. 47%). Sixty per cent of children were shorter or grew at a slower rate. Sixty‐five per cent of the children presented with one or more endocrine complications. These are the combined effects of different treatment regimens (chemotherapy, radiotherapy, combined therapy). It is essential to know the natural history of these patients in order to offer proper guidance and treatment as survival rates are increasing.