bluuebunny/crossref_metadata_2025 huggingface dataset:
Convergence between CD98 and integrin‐mediated T‐lymphocyte co‐stimulation
Authors: Warren A. P. , Patel K. , Miyamoto Y. , Wygant J. N. , Woodside D. G. , Mcintyre B. W.
Published: March 2003

CD98 is a widely expressed cell surface heterodimeric glycoprotein, which is rapidly up‐regulated upon activation of T lymphocytes. Monoclonal antibody (mAb) 80A10 recognizes an epitope on CD98 and in combination with CD3 antibody causes proliferation of peripheral blood T lymphocytes. CD98 co‐stimulatory activity, mediated by either mAb 80A10 or 4F2, a well‐characterized CD98‐specific mAb, is blocked in the presence of the soluble β1 integrin antibody 18D3. Previously we have reported that co‐stimulatory activity of antibodies to integrins α4β1, α5β1, αLβ2 and α4β7 is inhibited by 18D3, whereas co‐stimulation mediated by non‐integrins was unaffected. Thus the non‐integrin CD98 is uniquely sensitive to the inhibitory effects of β1 integrin‐blocking antibodies, which may reflect convergent signalling mechanisms between integrins and CD98. This is consistent with recent reports suggesting that CD98 may regulate integrin‐mediated adhesive events.
CpG‐oligodeoxynucleotides enhance T‐cell receptor‐triggered interferon‐γ production and up‐regulation of CD69 via induction of antigen‐presenting cell‐derived interferon type I and interleukin‐12
Authors: Kranzer K. , Bauer M. , Lipford G. B. , Heeg K. , Wagner H. , Lang R.
Published: March 2003

Bacterial cytidine–phosphate–guanosine (CpG‐DNA) activates antigen‐presenting cells (APC) and drives T helper 1 (Th1)‐polarized immune responses in the mouse. Claims have been made that CpG‐DNA costimulates murine T cells. We examined the direct and indirect effects of CpG‐oligodeoxynucleotides (CpG‐ODN) on human T‐cell activation. CpG‐ODN failed to costimulate purified human T cells activated with α‐CD3 or α‐T‐cell receptor (TCR)αβ antibodies. In contrast, CpG‐ODN sequence‐specifically caused increased expression of CD69 on CD4 and CD8 T cells when peripheral blood mononuclear cells (PBMC) were stimulated via α‐CD3. CpG‐ODN and α‐CD3 stimulation synergized to induce interferon‐γ (IFN‐γ) in T cells and natural killer (NK) cells, as shown by intracellular fluorescence‐activated cell sorter (FACS) staining. These effects of CpG‐ODN on human T cells were caused by the release of IFN type I (IFN‐I) and interleukin‐12 (IL‐12) from PBMC. Enhancement of CD69 expression on α‐CD3‐triggered T cells could be reproduced in a coculture transwell system of purified T cells and PBMC, was inhibited by neutralizing antibodies to IFN‐I and could be mimicked by adding exogenous IFN‐I. Furthermore, neutralization of either IFN‐I or IL‐12 diminished, and in combination abolished, IFN‐γ production. These findings show that CpG‐ODN potentiate TCR‐triggered activation of human T cells in an APC‐dependent manner.
Native IgG2ab is barely antigenic to major histocompatibility complexclass II‐restricted T cells owing to inefficient internalization by professionalantigen‐presenting cells
Authors: Bartnes K. , Hannestad K.
Published: March 2003

Peptide epitopes derived from immunoglobulin variable regions represent tumour‐specific antigens on B‐cell neoplasms and can be recognized by syngeneic, major histocompatibility complex (MHC) class II‐restricted T cells. Immunoglobulin peptide/MHC class II complexes may also be involved in autoimmunity and CD4+ T‐cell‐mediated B‐cell regulation. Thus, the IgG2ab H‐chain allopeptide γ2ab 435–451 presented on I‐Ad mimics the epitope implicated in herpes simplex virus‐induced autoimmune stromal keratitis and is the target of T helper 1 (Th1) clones that suppress IgG2ab production in vivo. We here report that spleen and thymus cells constitutively present the autologous γ2ab epitope to a γ2ab 435–451/I‐Ad reactive T‐cell hybridoma as a function of the animal housing conditions (specific pathogen‐free or not) and the serum levels of IgG2ab. Constitutive presentation in the spleen was predominantly performed by dendritic cells. Whereas spleen cells poorly presented native IgG2ab to a γ2ab 435–451/I‐Ad reactive T‐cell hybridoma, IgG2ab in the form of immune complexes were presented > 200‐fold more efficiently owing to internalization via low‐affinity FcγR on macrophages. The antigenicity could also be improved by homotypic aggregation and by targeting IgG2ab to complement receptors on the A20 B‐cell lymphoma. Mice without detectable IgG2ab‐containing immune complexes typically exhibited minimal constitutive presentation. Nevertheless, native IgG2ab can sensitize antigen‐presenting cells in vivo, as mice that were devoid of immune complexes and carried an IgG2ab‐producing tumour did present constitutively, even at physiological IgG2ab serum levels. Whereas the amounts of IgG released from most B‐cell lymphomas may be too low to allow spontaneous priming of tumour‐specific MHC class II‐restricted T cells, administration of tumour immunoglobulin in aggregated form might improve the efficacy of idiotype vaccination.
Chronic restraint stress induces severe disruption of the T‐cell specific response to tetanus toxin vaccine
Authors: Tournier J.‐ N. , Mathieu J. , Mailfert Y. , Multon E. , Drouet C. , Jouan A. , Drouet E.
Published: March 2003

Chronic stress is known to induce immunological disorders. In the present study we examined the consequences of chronic restraint stress on the immune response to tetanus toxin in mice. We investigated the repartition of subsets of lymphoid cells in blood and spleen, the functional ability of lymphocytes to proliferate and to produce cytokines, and antibody titres against tetanus toxin following stress. We report discordance of the stimulation index of lymphocytes in the restraint group: the proliferating rate severely decreased following stimulation with a relevant antigen, whereas it increased with mitogen. Thus, we report a decrease in cytokine production with relevant antigen (interferon‐γ and interleukin‐10), without a T helper type 1 and 2 secretion imbalance. Moreover, we observed an alteration in the humoral response, including a delay in isotype maturation and an immunoglobulin G1/G2a imbalance.
Analysis of recombinant mycobacteria as T helper type 1 vaccines in an allergy challenge model
Authors: Janssen R. , Kruisselbrink A. , Hoogteijling L. , Lamb J. R. , Young D. B. , Thole J. E. R.
Published: March 2003

The potential for development of mycobacteria as T helper type 1 (Th1) vaccines capable of induction of Th1 responses to recombinant antigens was explored in a model system based on an immunodominant peptide from house dust mite. Different recombinant mycobacterial preparations were compared for their ability to induce a Th1 response to the peptidea. It was found that mycobacterial viability was not a prerequisite for Th1 immunogenicity. A dominant interferon‐γ (IFN‐γ) response to peptide was observed in splenocytes from C57BL/6J mice immunized with live or heat‐killed preparations of recombinant Mycobacterium vaccae or with live attenuated bacillus Calmette–Guèrin (BCG) vaccine expressing the antigen. Interleukin‐5 (IL‐5), a marker of a Th2 response, was detected only in mice receiving live M. vaccae. A similar pattern was observed in BALB/b mice, although the magnitude of the IFN‐γ response was much lower. Control and immunized mice were subsequently exposed to allergen using a Th2‐inducing challenge protocol. A significant shift from a Th2 to a Th1 response was observed in immunized mice, as judged by cytokine expression by splenocytes and by subclass of circulating antibody. The effect was seen in three inbred mouse strains differing in their innate bias towards Th1 or Th2 responses. It was dependent on the presence of specific antigen in the mycobacterial preparation and, under the immunization conditions tested, was more pronounced with dead M. vaccae than with live BCG as carrier vaccine. The results demonstrate the potency of killed mycobacteria as Th1 adjuvants and suggest a potential application for recombinant mycobacteria in antigen‐specific immune modulation.
Characterization of self‐T‐cell response and antigenic determinants of U1A protein with bone marrow‐derived dendritic cells in NZB × NZW F1 mice
Authors: Suen Jau‐Ling , Wu Chung‐Hsiun , Chen Ying‐Yu , Wu Wen‐Mein , Chiang Bor‐Luen
Published: March 2003

Systemic lupus erythematosus (SLE) is characterized by the existence of a heterogeneous group of autoantibodies directed against nuclear intact structures, such as nucleosomes and small nuclear ribonucleoproteins (snRNPs). Autoantibodies against snRNPs are of special interest because they are detectable in the majority of SLE patients. Although the B‐cell antigenic determinants have been well characterized, very limited data have been reported in regard to the T‐cell epitopes of snRNPs. Furthermore, several studies have demonstrated that determination of the auto‐T‐cell epitopes recognized by freshly isolated T cells is difficult from unprimed lupus mice when self‐antigen‐pulsed B cells or macrophages are used as antigen‐presenting cells (APCs) in vitro. In the present study, we showed a novel approach for determining the auto‐T‐cell epitopes, using bone marrow‐derived dendritic cells (BMDCs) pulsed with the murine U1A protein – an immunodominant antigen of the U1 snRNPs – which is capable of activating freshly isolated T cells from unprimed (NZB × NZW) F1 (BWF1) mice in vitro. The T‐cell epitope area was found to be located at the C‐terminus of U1A, overlapping the T‐cell epitope of human U1A that has been reported in human SLE. Identification of the autoreactive T‐cell epitope(s) in snRNPs will help to elucidate how reciprocal T–B determinant spreading of snRNPs emerges in lupus. The results presented here also indicate that it is feasible to use this approach to further explore strategies to design immunotherapy for patients with lupus.
Grass pollen immunotherapy for hayfever is associated with increases in local nasal but not peripheral Th1 : Th2 cytokine ratios
Authors: Wachholz Petra A. , Nouri‐Aria Kayhan T. , Wilson Duncan R. , Walker Samantha M. , Verhoef Adrienne , Till Stephen J. , Durham Stephen R.
Published: March 2003

Grass pollen immunotherapy is the only treatment for hayfever that is both effective and confers long‐term benefit. Immunotherapy may act by altering the local nasal mucosal T helper type 2 (Th2) to type 1 (Th1) cytokine balance either by down‐regulation and/or immune deviation of T‐lymphocyte responses. There is controversy as to whether these changes are detectable in peripheral blood. We therefore examined both local nasal and peripheral T‐cell responses to allergen exposure in the same subjects before and after immunotherapy. In a double‐blind trial of grass pollen immunotherapy, nasal biopsies were obtained at baseline and during the peak pollen season following 2 years of immunotherapy. Placebo‐treated patients showed a seasonal increase in CD3+ T cells (P = 0·02) and in interleukin‐5 (IL‐5) mRNA+ cells (P = 0·03) and no change in interferon‐γ (IFN‐γ ) mRNA+ cells (P = 0·2) in the nasal mucosa. In contrast, in the immunotherapy‐treated group, there were no changes in the number of CD3+ T cells (P = 0·3) and IL‐5 mRNA+ cells (P = 0·2) but a significant increase in the number of IFN‐γ mRNA+ cells (P = 0·03). Furthermore, clinical improvement in the immunotherapy‐treated group was accompanied by a seasonal increase in the ratio of IFN‐γ to IL‐5 mRNA+ cells in the nasal mucosa (P = 0·03). In contrast, there were no significant changes in peripheral T‐cell proliferative responses or cytokine production for IFN‐γ or IL‐5 in response to grass pollen either within or between the two treatment groups. We conclude that successful grass pollen immunotherapy was associated with an increase in the ratio of IFN‐γ to IL‐5 mRNA+ cells in the nasal mucosa, whereas these changes were not reflected by alterations in peripheral blood T‐cell proliferative responses or cytokine production before/after treatment.
Endotoxin, but not platelet‐activating factor, activates nuclear factor‐κB and increases IκBα and IκBβ turnover in enterocytes
Authors: De Plaen Isabelle G. , Qu Xiao‐Wu , Wang Hao , Tan Xiao‐Di , Wang Liya , Han Xin‐Bing , Rozenfeld Ranna A. , Hsueh Wei
Published: March 2003

Bacterial endotoxin (lipopolysaccharide; LPS) and platelet‐activating factor (PAF) are important triggers of bowel inflammation and injury. We have previously shown that LPS activates the transcription factor nuclear factor (NF)‐κB in the intestine, which up‐regulates many pro‐inflammatory genes. This effect partly depends on neutrophils and endogenous PAF. However, whether LPS and PAF directly activate NF‐κB in enterocytes remains controversial. In this study, we first investigated the effect of LPS and PAF on NF‐κB activation in IEC‐6 (a non‐transformed rat small intestinal crypt cell line) cells, by electrophoresis mobility shift assay and supershift, and found that LPS, but not PAF, activates NF‐κB mostly as p50–p65 heterodimers. The effect was slower than tumour necrosis factor (TNF). Both LPS and TNF induce the expression of the NF‐κB‐dependent gene inducible nitric oxide synthase (iNOS), which occurs subsequent to NF‐κB activation. We then examined the effect of LPS and TNF on the inhibitory molecules IκBα and IκBβ. We found that TNF causes rapid degradation of IκBα and IκBβ. In contrast, LPS did not change the levels of IκBα and IκBβ up to 4 hr (by Western blot). However, in the presence of cycloheximide, there was a slow reduction of IκBα and IκBβ, which disappeared almost completely at 4 hr. These observations suggest that LPS causes slow degradation and synthesis of IκBα and IκBβ and therefore activates NF‐κΒ via at least two mechanisms: initially, through an IκB‐independent mechanism, and later, via an increased turnover of the inhibitor IκB. NF‐κΒ activation precedes the gene expression of iNOS (assayed by reverse transcription–polymerase chain reaction), suggesting that LPS up‐regulates iNOS via this transcription factor.
Macrophage phagocytosis of foot‐and‐mouth disease virus may create infectious carriers
Authors: Rigden Rachael C. , Carrasco Carlos P. , Summerfield Artur , MCCullough Kenneth C.
Published: March 2003

Macrophages play critical roles in innate defences against virus infections, particularly pertinent to the rapid immune response required following emergency vaccination against foot‐and‐mouth disease virus (FMDV). Consequently, macrophage–FMDV interaction was studied in vitro, in the absence of specific antibodies, to mimic the animal early postvaccination. A gradual loss of infectivity and viral antigen was observed over 48 hr, and no evidence of productive virus replication was found. From the pathological viewpoint, an important observation was that the majority of macrophages carried infectious virus for at least 10 hr. Pronase and mild acid treatments showed the virus to be primarily on the cell surface during the first 4 hr. Thereafter, it became internalized (pronase‐ and pH resistant), but remained infectious for 10–24 hr. The internalization process was dependent on microfilament activity, while the survival of infectious virus related to live virus‐dependent inhibition of macrophage protein synthesis. Infectious centre assays demonstrated that this infectious virus – whether on the cell surface or internalized – was actually being released from the cells. This is interesting considering that FMDV is highly pH labile. Together, these characteristics suggest that the virus had been internalized by a process such as macropinocytosis, and fusion with endosomes was delayed or impaired. This mechanism whereby the virus could ‘piggyback’ on or in the macrophage, becoming internalized but not degraded for at least 10 hr, are important considerations in FMD pathogenesis. Such ‘virus‐transporting’ macrophages would be in a position to carry infectious FMDV to different sites in the body, where it could be released to infect other cells for replication.
Expression and regulation of the pattern recognition receptors Toll‐like receptor‐2 and Toll‐like receptor‐4 in the human placenta
Authors: Holmlund Ulrika , Cebers Gvido , Dahlfors Agneta R. , Sandstedt Bengt , Bremme Katarina , EkstrÖm EVA S. , Scheynius Annika
Published: March 2003

The placenta constitutes a physical and immunological barrier against invading infectious agents and has been suggested to be a pregnancy‐specific component of the innate immune system. The aim of this study was to investigate the presence and regulation of Toll‐like receptors‐2 and ‐4 (TLR2 and TLR4) in the human placenta, because these receptors are believed to be important for immune responses against pathogens. Twenty‐eight placentas from normal term pregnancies were analysed with immunohistochemistry, which showed a strong immunoreactivity for TLR2 and TLR4 in the villous and the intermediate trophoblasts. The regulation of TLR2 and TLR4 by microbial stimulus was assessed by incubating explants of term chorionic villi with zymosan or lipopolysaccharide (LPS) and analysed with real‐time reverse transcriptase–polymerase chain reaction. Stimulation with zymosan and LPS readily induced interleukin (IL)‐6 and IL‐8 cytokine production in the placenta cultures, whereas TLR2 and TLR4 mRNA and protein expression remained at the same high level as in unstimulated explants. These data suggests a novel mechanism for the fetoplacental unit to interact with micro‐organisms.
Cytokines in Lyme borreliosis: lack of early tumour necrosis factor‐α and transforming growth factor‐β1 responses are associated with chronic neuroborreliosis
Authors: Widhe Mona , Grusell Mattias , Ekerfelt Christina , Vrethem Magnus , Forsberg Pia , Ernerudh Jan
Published: March 2003

The clinical outcome of the tick born infection Lyme borreliosis seems to be influenced by the type of immune response mounted during the disease, as suggested by various animal models. Here we report the serum and cerebrospinal fluid levels of tumour necrosis factor‐α (TNF‐α), transforming growth factor β1 (TGF‐β1) and interleukin‐6 (IL‐6) in samples drawn at different disease intervals during the course of non‐chronic neuroborreliosis (n=10), chronic neuroborreliosis (n=15), erythema migrans (n=8, serum only) and controls (n=7). When comparing early neuroborreliosis cerebrospinal fluid samples, significantly higher levels of TNF‐α were found in non‐chronic patients than in chronic patients (P<0·05). Moreover, TGF‐β1 was increased in the early serum samples of non‐chronic patients, as compared to chronic patients (P<0·01). Elevated serum levels of TGF‐β1 were also found in erythema migrans as compared to neuroborreliosis and controls (P<0·05). The high TNF‐α levels noted in early cerebrospinal fluid samples of non‐chronic patients only, possibly reflects an ongoing pro‐inflammatory immune response in the central nervous system, which could be beneficial in eliminating disease. High serum levels of TGF‐β1 probably mirror an anti‐inflammatory response, which might play a role in controlling the systemic immune response.
Mycobacteria, an environmental enhancer of lupus nephritis in a mouse model of systemic lupus erythematosus
Authors: Hawke Christine G. , Painter Dorothy M. , Kirwan Paul D. , Van Driel Rosemary R. , Baxter Alan G.
Published: March 2003

Systemic lupus erythematosus (SLE) is a chronic systemic autoimmune disease characterized by the production of antibodies directed against self antigens. Immune complex glomerulonephritis (GN) is one of the most serious complications of this disorder and can lead to potentially fatal renal failure. The aetiology of SLE is complex and multifactorial, characterized by interacting environmental and genetic factors. Here we examine the nature of the renal pathology in mycobacteria‐treated non‐obese diabetic (NOD) mice, in order to assess its suitability as a model for studying the aetiopathogenesis of, and possible treatment options for, lupus nephritis (LN) in humans. Both global and segmental proliferative lesions, characterized by increased mesangial matrix and cellularity, were demonstrated on light microscopy, and lesions varied in severity from very mild mesangiopathic GN through to obliteration of capillary lumina and glomerular sclerosis. Mixed isotype immune complexes (IC) consisting of immunoglobulin G (IgG), IgM, IgA and complement C3c were detected using direct immunofluorescence. They were deposited in multiple sites within the glomeruli, as confirmed by electron microscopy. The GN seen in mycobacteria‐treated NOD mice therefore strongly resembles the pathology seen in human LN, including mesangiopathic, mesangiocapillary and membranous subclasses of LN. The development of spontaneous mixed isotype IC in the glomeruli of some senescent NOD mice suggests that mycobacterial exposure is accelerating, rather than inducing, the development of GN in this model.
Recognition of bovine respiratory syncytial virus proteins by bovine CD8+ T lymphocytes
Authors: Gaddum Ruth M. , Cook Roy S. , Furze Julie M. , Ellis Shirley A. , Taylor Geraldine
Published: March 2003

CD8+ T lymphocytes play a major role in the clearance of bovine respiratory syncytial virus (BRSV), an important respiratory pathogen of young calves that shares many of the epidemiological and pathological features of human respiratory syncytial virus (HRSV) in infants. Recombinant vaccinia virus (rVV) and recombinant fowlpox virus (rFPV), expressing individual BRSV proteins, were used to demonstrate that the F, N and M2 proteins were the major antigens recognized by bovine CD8+ T cells in major histocompatibility complex (MHC)‐defined cattle. BRSV protein recognition by CD8+ T cells was analysed using cytotoxic T lymphocyte (CTL) assays or by the production of interferon‐γ (IFN‐γ) following restimulation with BRSV proteins. Strong recognition of the G protein by CD8+ T cells was observed in cattle that had been vaccinated with rVV expressing this protein and subsequently challenged with BRSV. Although there is variation in the number of expressed MHC genes in cattle with different class I haplotypes, this did not appear to influence BRSV protein recognition by CD8+ T cells. Knowledge of the antigenic specificity of BRSV‐specific CD8+ T cells will facilitate the qualitative and quantitative analysis of BRSV‐specific CD8+ T‐cell memory in cattle and help to ensure that potential vaccines induce a qualitatively appropriate CD8+ T‐cell response.
Macrophages exposed to Mycobacterium tuberculosis release chemokines able to recruit selected leucocyte subpopulations: focus on γδ cells
Authors: Ferrero Elisabetta , Biswas Priscilla , Vettoretto Katuscia , Ferrarini Marina , Uguccioni Mariagrazia , Piali Luca , Leone Biagio Eugenio , Moser Bernhard , Rugarli Claudio , Pardi Ruggero
Published: March 2003

Granuloma is a typical feature of tuberculosis. We evaluated the chemotaxis of selected human leucocyte subsets induced by macrophages incubated with Mycobacterium tuberculosis (MT)‐derived products in vitro. The release of monocyte chemotactic protein 1 (MCP‐1) and interleukin‐8 (IL‐8) correlated with the specific induction of strong chemotaxis towards monocytes and polymorphonuclear leucocytes (PMNs). γδ and T helper type 1 (Th1) αβ lymphocytes were chemoattracted, while T‐resting, IL‐2‐activated and Th2 lymphocytes were unaffected. Activation with mycobacterium‐derived, phosphate‐containing components, modulated the chemokine receptor profile of γδ T lymphocytes as well as their pattern of cyto‐chemokine production, disclosing a potential for their active participation in granuloma formation. In particular, CXCR3 and IP‐10, which we found to be released by MT‐pulsed alveolar macrophages, seem to represent the receptor–counter‐receptor pair implicated in the chemotaxis of γδ lymphocytes. Immunohistochemical analysis and in situ hybridization revealed the in vivo presence of IL‐8, MCP‐1 and IL‐10 in lymph node and lung tuberculous granulomas. Our results underscore the role of MT extracts in the induction of macrophage‐derived chemokines responsible for the orchestrated recruitment of PMNs, monocytes, and Th1 and γδ T cells, as well as in the regulation of γδ function.
Genetic variation in geographical populations of western and Mexican corn rootworm
Authors: Szalanski A. L. , Roehrdanz R. L. , Taylor D. B. , Chandler L.
Published: March 2003

Genetic variation in the nuclear rDNA ITS1 region of western corn rootworm, Diabrotica virgifera virgifera (WCR), and Mexican corn rootworm, D. v. zeae (MCR) was studied. Two sites were detected which differentiated WCR and MCR in the 642‐base sequence. Polymerase chain reaction–restriction fragment length polymorphism (PCR‐RFLP) analysis of the first internal transcribed spacer region (ITS1) sequence revealed no variation within or among the twelve WCR and two MCR populations. PCR‐RFLP of 75% of the mitochondrial DNA genome detected one significant polymorphic site out of the approximately 190 restriction sizes observed in WCR. The polymorphism did not differentiate geographical populations of WCR and is not diagnostic for the subspecies. The low levels of variation observed in WCR suggests either high levels of gene flow or a recent geographical expansion from a relatively small base. Gene flow would facilitate the rapid spread of traits that could compromise control programmes, such as insecticide resistance or behavioural modifications. The minimal genetic differentiation between WCR and MCR raises questions about the evolutionary history of these subspecies and how the distinct phenotypes are maintained.
Sequence and evolution of a hexamerin from the ant Camponotus festinatus
Authors: Martínez T. , Burmester T. , Veenstra J. A. , Wheeler Diana
Published: March 2003

In the ant Camponotus festinatus, two different hexamerins accumulate stage‐specifically during the late larval period and at various times in adults. These hexamerins serve as storage proteins and play important roles in brood nourishment and colony founding. We report an analysis of the cDNA sequence of C. festinatus hexamerin 2 (CfeHex2). The native protein contains 732 amino acids, which are moderately enriched in aromatic amino acids, aspartate and asparagine. Phylogenetic analyses show a close relationship of CfeHex2 to a putative toxin of the braconid wasp, Bracon hebetor. The divergence of Formicidae and Braconidae hexamerins was calculated to have begun 187 MYA, an estimate consistent with currently accepted phylogeny of insect orders.
Cytochrome P450s CYP6D3 and CYP6D1 are part of a P450 gene cluster on autosome 1 in the house fly
Authors: Kasai S. , Scott J. G.
Published: March 2003

The P450 monooxygenases of insects are important in the metabolism of numerous endogenous and exogenous compounds. However, identity of the P450 isoform(s) involved in these reactions is rarely known. A critical first step in the identification of important P450s is the cloning and sequencing of their genes. Toward this goal we report the genomic sequence of a new cytochrome P450, termed CYP6D3, from the house fly, Musca domestica. CYP6D3 is part of a P450 gene cluster located on chromosome 1 and is located upstream of a related gene, CYP6D1. The similar genetic structures of CYP6D3 and CYP6D1 (5 exons and 4 introns of similar length) suggest one of these genes may have been the result of a duplication event. The CYP6D3 deduced amino acid sequence indicates a protein with 518 amino acids and a molecular weight of 59.3 kDa. The CYP6D3 protein is most similar to house fly CYP6D1 (78%) and Cyp6D2 (56%) from Drosophila melanogaster. The deduced amino acid sequences of CYP6D3 and CYP6D1 are identical at the Helix I and heme binding regions.
Molecular evolution of thecacophonyIVS6 region in sandflies
Authors: Lins R. M. M. A. , Oliveira S. G. , Souza N. A. , De Queiroz R. G. , Justiniano S. C. B. , Ward R. D. , Kyriacou C. P. , Peixoto A. A.
Published: March 2003

A number of insects produce acoustic signals during courtship. Genes involved in the control of the courtship song are particularly interesting from an evolutionary viewpoint because interspecific variation in this signal is potentially important as a reproductive isolation mechanism and, as a consequence, in the speciation process. Thecacophonygene was identified by a mutation affecting the ‘lovesong’ inDrosophila melanogaster. Phlebotomine sandflies (Diptera: Psychodidae) also produce acoustic stimuli during courtship and thereforecacophonycan be used as an interesting molecular marker in evolutionary studies in these important disease vectors. In this paper we have studied the molecular evolution of the IVS6 region ofcacophonyin sandflies. We compared the level of divergence in the exon sequences encoding this conserved domain inDrosophilaand Phlebotomines. We also analysed the high level of variation in an intron that is present in sandflies but that was lost inDrosophiladuring evolution. The availablecacophonysequences were also used for a phylogenetic analysis of some species of the Neotropical genusLutzomyia.
Differential expression and induction of two Drosophila cytochrome P450 genes near the Rst(2)DDT locus
Authors: Brandt A. , Scharf M. , Pedra J. H. F. , Holmes G. , Dean A. , Kreitman M. , Pittendrigh B. R.
Published: March 2003

Previous studies have shown that the major metabolic resistance locus in the insecticide‐resistant Drosophila line Rst(2)DDTWisconsin maps between the markers cn and vg on chromosome 2. Six cytochrome P450 genes exist in this region. We investigated the expression levels of these P450 genes in DDT‐resistant and ‐susceptible fly lines. We report: (i) DDT resistance is significant (> 30‐fold) and dominant, (ii) resistance is reduced by the cytochrome P450 inhibitor PBO, (iii) there is constitutive over‐expression relative to susceptible flies of two genes encoding cytochrome P450 enzymes within the cn‐vg region (CYP6G1 = 4.3‐fold; CYP12D1 = 6.0‐fold), and (iv) exposure to DDT results in an increased expression of only one of these two P450 genes (CYP12D1  ≥ 6‐fold above constitutive resistant fly baselines).
New perspectives on radicular cysts: do they heal?
Authors: Nair
Published: March 2003

During the past few decades several authors have perpetuated the notion that nearly half of all periapical lesions are radicular cysts. A few studies, based on meticulous serial sectioning of periapical lesions retrieved in toto, have shown that the actual incidence of radicular cyst is only about 15% of all periapical lesions. Equally significant was the discovery in 1980 and recent confirmation that radicular cysts exist in two structurally distinct classes namely, those containing cavities completely enclosed in epithelial lining (periapical true cysts) and those containing epithelium‐lined cavities that are open to the root canals (periapical pocket cysts). From a clinical point of view a periapical pocket cyst may heal after conventional root canal therapy whereas an apical true cyst is less likely to be resolved without surgical intervention.
Shaping ability of Quantec Series 2000 rotary nickel‐titanium instruments in simulated root canals: Part 2
Authors: Thompson  , Dummer
Published: March 2003

The aim of this laboratory based study was to determine the shaping ability of Quantec Series 2000 nickel‐titanium rotary instruments in simulated root canals. A total of 40 canals with four different shapes in terms of angle and position of curve were prepared with Quantec Series 2000 instruments using the technique recommended by the manufacturer. Part 2 of this report describes the efficacy of the instruments in terms of prevalence of canal aberrations, the amount and direction of canal transportation and overall post‐operative shape. Pre‐ and post‐operative images of the canals were taken using a video camera attached to a computer with image analysis software. The pre‐ and post‐operative views were superimposed to highlight the amount and position of material removed during preparation. Twenty‐one zips and elbows were created during preparation with a significant difference (P < 0.005) between canal shapes in terms of the incidence of aberrations. Four perforations were created, with significant differences (P < 0.005) between the canal shapes; three ledges were also created but no danger zones. Highly significant differences (P < 0.001) were apparent between the canal shapes in total canal width at specific points along the canal length and in the amount of resin removed from the inner and outer aspects of the curve. Canal transportation at the end‐point of preparation was most frequently directed towards the outer aspect of the curve, and without exception in canals with 40° curves. At the beginning of the curve, transportation became more evenly balanced between the inner and outer aspect of the curve, although predominated towards the outer. Transportation was generally directed towards the outer at the orifice, especially in canals with 40° curves. Mean absolute transportation at the various measurement points was less than 0.11 mm; significant differences occurred between canal shapes at the end‐point of preparation (P < 0.0001), at the zips (P < 0.005), at the apex (P < 0.0001) and beginning of the curve (P < 0.05) and at the orifice (P < 0.0001). Under the conditions of this study, Quantec Series 2000 rotary nickel‐titanium instruments created a relatively large number of aberrations including four perforations. The aberrations were created by the larger instruments implying that these should be used with caution at the full working distance. Scanning electron micrographs of these instruments revealed sharp instrument tips which appeared likely to predispose to transportation and the creation of defects along the outer aspect of severely curved canals.
Surgical extrusion of crown‐root‐fractured teeth: a clinical review
Authors: Çalisşkan M. K. , Türkün M. , Gomel M.
Published: March 2003

Aim In this clinical study, combined surgical and endodontic treatment was performed in 20 cases of crown‐root fracture and the outcomes reviewed.

Methodology Surgical treatment involved a conventional extraction and stabilization technique. Root canal treatment using calcium hydroxide was performed. Before root canal obturation, a calcium hydroxide dressing was maintained for 3 months.

Results Follow‐up examinations, which varied between 6 and 36 (mean 14.5) months, showed that there were no radiographic and clinical signs of progressive root resorption, marginal bone loss or periapical disease in all except one case.

Conclusions The favourable results of this study demonstrate that surgical extrusion in teeth with crown‐root fractures may be an alternative treatment to orthodontic extrusion.
Detection of bacterial virulence genes associated with infective endocarditis in infected root canals
Authors: Bate A. L. , Ma J. K‐C. , Ford T. R. Pitt
Published: March 2003

Aim  The aim of this study was to examine whether bacteria associated with root canals possess genes that might predispose to bacterial colonization of the endocardium.

Methodology  Oligonucleotides were designed from DNA sequences encoding the functional binding regions of streptococcal fibronectin‐binding protein (FnBP) and staphylococcal fibrinogen‐binding protein (FgBP). The specificity and cross‐reactivity of the oligonucleotide primers were investigated; streptococcal primers were tested for recognition of FnBP genes in other strains of streptococci, and the staphylococcal primers for detection of FgBP from other staphylococci. Interspecies specificity of these primers was also investigated. In a pilot clinical study, the pulp chambers of 16 nonvital teeth without sinus tracts, were opened aseptically. Root canal samples were collected, along with samples from the gingival sulcus and anterior nares. From these samples DNA was extracted, subjected to polymerase chain reaction (PCR) and analysed by agarose gel electrophoresis.

Results  Using the streptococcal FnBP primers, PCR bands were amplified from eight root canal samples, eight gingival samples and three nasal samples. With the staphylococcal primers, PCR bands were amplified from seven root canals, 11 gingival and nine nasal samples. This study showed that PCR could be used to detect bacteria in root canals that possess genes with homology to functional regions of those encoding FnBP or FgBP.

Conclusions  If bacteria in root canals possess FnBP or FgBP, they may have the potential to cause infective endocarditis.
Effect of sodium hypochlorite on mechanical properties of dentine and tooth surface strain
Authors: Sim T. P. C. , Knowles J. C. , Ng Y‐L. , Shelton J. , Gulabivala K.
Published: March 2003

Aim

The aim of this study was to test the null hypothesis that sodium hypochlorite irrigation of root canals does not alter the properties of dentine and contribute to the weakening of root‐treated teeth.

Methodology

The effect of two concentrations (0.5%, 5.25%) of sodium hypochlorite (NaOCl) and saline on (i) the elastic modulus and flexural strength of machined dentine bars, and (ii) changes in strain of ‘whole’ extracted human teeth were evaluated. One hundred standardized plano‐parallel dentine bars (> 11.7 × 0.8 × 0.8 mm) were randomly divided into the three groups, immersed for 2 h in the respective solutions and then subjected to a three‐point bend test. Changes in strain of each of 10 teeth on cyclical nondestructive occlusal loading were measured using electrical resistance strain gauges bonded to the cervical aspects. Each tooth had its crown and enamel reduced and root canal prepared. These were irrigated sequentially in a series of four separate, 30‐minute regimes; initial‐saline, 0.5% NaOCl, 5.25% NaOCl and final‐saline. The changes in strains after each irrigation regime were compared.

Results

There was a significant decrease in elastic modulus of the dentine bars immersed in 5.25% NaOCl compared with the saline group (P < 0.01). There was also a significant decrease in flexural strength of the dentine bars in the 5.25% NaOCl group compared to both the saline and 0.5% NaOCl groups (P < 0.01). The strain data from the nondestructive tooth loading tests revealed significant increases in tensile strain between the initial‐saline and the final‐saline stages (P < 0.01). Significant increases in compressive strains were also found between initial‐saline and 5.25% NaOCl; and between 0.5% NaOCl and 5.25% NaOCl stages (P < 0.01).

Conclusions

The null hypothesis was rejected, 5.25% NaOCl reduced the elastic modulus and flexural strength of dentine. Irrigation of root canals of single, mature rooted premolars with 5.25% NaOCl affected their properties sufficiently to alter their strain characteristics when no enamel was present.
Dye penetration in dry and water‐filled gaps along root fillings
Authors: Kontakiotis E. G. , Georgopoulou M. K. , Morfis A. S.
Published: March 2003

Aim

The aim of this study was to investigate the influence of hydration in voids along root fillings on methylene blue penetration.

Methodology

A total of 80 human root canals were prepared using a step‐back technique and filled with a zinc oxide based sealer and gutta‐percha. Leakage along the fillings was measured by a transport fluid model and classified into three categories: gross leakage (GL), slight leakage (SL) and no leakage (NL). Specimens with NL and SL were immersed into methylene blue (MB) 2% for 24 h (group I). Specimens with GL which had wide gaps filled with water were randomly divided into two groups (II, III). Transport air was applied to remove water from gaps only in specimens of group III. All tested specimens from groups II and III were also immersed into MB 2% for 24 h. Each specimen was then split longitudinally and linear measurements of dye penetration were recorded.

Results

Group III (with dry gaps) showed significantly more dye penetration than group II. No significant difference was found between group I and group II.

Conclusions

Methylene blue penetrates along root fillings more easily in dry gaps than in water‐filled gaps.
A scanning electron microscopic study of debris and smear layer remaining following use of GT rotary instruments
Authors: Gambarini G. , Laszkiewicz J.
Published: March 2003

Aim The aim of the present study was to assess debris and smear layer remaining following canal preparation with GT rotary instruments.

Methodology Sixteen freshly extracted single‐rooted premolar teeth were instrumented with GT™ rotary instruments using a crown‐down preparation technique. All specimens were flushed with 2 mL of 5% NaOCl between each rotary instrument. At the end of instrumentation the following final irrigation sequence was repeated two times: 2 mL of EDTA + Cetrimide for 1 min (Largal Ultra, Septodont, France) and 2 mL of 5% NaOCl for 5 min. A final flush with saline solution was made to halt any chemical activity. Two other uninstrumented teeth that were not irrigated served as controls. All teeth were split longitudinally and prepared for SEM evaluation. The presence of debris and smear layer was evaluated from photomicrographs at ×200 and ×1000 magnification taken in the apical, middle and coronal thirds of the canals. Blind evaluation was performed by two trained observers and scores were compiled separately. A five category scoring system for debris and smear layer was used. Values obtained were tabulated and statistical analysis was carried out using a parametric chi‐squared test.

Results Statistical analysis showed that there was no significant difference between the three regions of the root canals (P > 0.05) for debris. Comparison of the removal of the smear layer between the three regions showed that there was a statistically significant difference between all parts, especially between the coronal and apical thirds (P < 0.001). Overall, the coronal sections were cleaner than the middle and apical sections. The uninstrumented canals showed walls completely covered with tissue, confirming that specimen preparation alone did not remove tissue.

Conclusions Under the conditions of the present study GT™ rotary instruments removed debris effectively, but left root canal walls covered with smear layer, particularly in the apical third.
In vitro comparison of different types of sodium perborate used for intracoronal bleaching of discoloured teeth
Authors: Arı H. , Üngör M.
Published: March 2003

Aim To compare the bleaching efficacy of three different types of sodium perborate (SP) commonly used for intracoronal bleaching of discoloured non‐vital teeth.

Methodology Sixty‐five extracted human maxillary central incisors with intact crowns were used. Following obturation with gutta‐percha and a root canal sealer using a lateral condensation technique, the coronal aspects of the root canal fillings were covered with a 1‐mm thick protective base placed to a level 1 mm apical to the labial cemento–enamel junction (CEJ). The teeth were than stained artificially with fresh blood over a period of 18 days. One or other of the bleaching materials (group 1: SP monohydrate + water, group 2: SP trihydrate + water, group 3: SP tetrahydrate + water, group 4: SP monohydrate + hydrogen peroxide (HP), group 5: SP trihydrate + HP, group 6: SP tetrahydrate + HP) were placed in the pulp chamber of the discoloured teeth and sealed with Cavit for 21 days. They were replaced with fresh preparations after 3, 7 and 14 days. The shades of the tooth crowns were evaluated at baseline and at 7, 14 and 21 days. Comparison of tooth shades was completed at each interval and analysed statistically using the chi‐squared test.

Results There was no statistically significant difference between the groups at 7, 14, 21 days. Period of bleaching significantly affected the outcome (P < 0.01). No colour changes were noted in the control teeth.

Conclusion The data obtained from this study demonstrate that sodium perborate can be used mixed with water rather than with hydrogen peroxide for bleaching discoloured teeth.
Recent advances in the microinsemination of laboratory animals
Authors: Ogura Atsuo , Inoue Kimiko , Ogonuki Narumi , Suzuki Osamu , Mochida Keiji , Matsuda Junichiro , Sankai Tadashi
Published: March 2003

Microinsemination techniques were applied to laboratory animals, including mice, Mongolian gerbils, mastomys, guinea pigs and cynomolgus monkeys. After micro‐insemination with spermatozoa or round spermatids, their oocytes were successfully fertilized and subsequently developed into 4‐cells to blastocysts, depending on species. The efficiencies of microinsemination techniques for producing fertilized oocytes were comparable with, or superior to those of IVF in these species.
Regulatory influence of germ cells on Sertoli cell function in the pre‐pubertal rat after acute irradiation of the testis
Authors: Guitton N. , Touzalin A. M. , Sharpe R. M. , Cheng C. Y. , Pinon‐Lataillade G. , Méritte H. , Chenal C. , Jégou B.
Published: March 2003

While germ cell regulation of Sertoli cells has been extensively explored in adult rats in vivo, in contrast, very little is known about germ cell influence on Sertoli cell function at the time when spermatogenesis begins and develops. In the present study various Sertoli cell parameters (number, testicular androgen binding protein (ABP) and testin, serum inhibin‐B and, indirectly, follicle‐stimulating hormone (FSH)) were investigated after the exposure of 19‐day‐old rats to a low dose of 3 Grays of γ‐rays. Differentiated spermatogonia were the primary testicular targets of the γ‐rays, which resulted in progressive maturation depletion, sequentially and reversibly affecting all germ cell classes. Testicular weight declined to a nadir when pachytene spermatocytes and spermatids were depleted from the seminiferous epithelium and complete or near complete recovery of spermatogenesis and testicular weight was observed at the end of the experiment. Blood levels of FSH and ABP were normal during the first 11 days after irradiation, when spermatogonia and early spermatocytes were depleted. While the number of Sertoli cells was not significantly affected by the irradiation, from days 11–66 after γ‐irradiation, ABP production declined and FSH levels increased when pachytene spermatocytes and spermatids were depleted and the recovery of these parameters was only observed when spermatogenesis was fully restored. Comparison of the pattern of change in serum levels of inhibin‐B and testicular levels of testin and of germ cell numbers strongly suggest a relationship between the disappearance of spermatocytes and spermatids from the seminiferous epithelium and the decrease in levels of inhibin‐B and increase in levels of testin from 7 to 36 days post‐irradiation. Levels of testin and inhibin‐B were restored before spermatogenesis had totally returned to normal. In conclusion, this in vivo study shows that pre‐pubertal Sertoli cell function is under the complex control of various germ cell classes. This control presents clear differences when compared with that previously observed in adult animals and depends on the Sertoli cell parameter of interest, as well as on the germ cell type.