bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular cloning, characterization and tissue expression of prophenoloxidase cDNA from the mosquito Armigeres subalbatus inoculated with Dirofilaria immitis microfilariae
Authors: Cho W. L. , Liu H. S. , Lee C. H. , Kuo C. C. , Chang T. Y. , Liu C. T. , Chen C. C.
Published: March 2003

A cDNA encoding mosquito Armigeres subalbatus pro‐phenol oxidase (As‐pro‐PO) was obtained by rapid amplification of cDNA ends‐polymerase chain reaction (RACE‐PCR) after Dirofilaria immitis inoculation. The 2205 bp As‐pro‐PO cDNA contains a 32 bp 5′‐noncoding region, a 2055 bp open reading frame (685 amino acids), and a 118 bp 3′‐noncoding region. Hydrophobic signal peptide for the endoplasmic reticulum targeting is not found in the NH2‐terminal region. Two potential copper‐binding domains, amino acids 197–245 and 345–412, are highly homologous to those of the other insect pro‐POs. A 2.2 kb As‐pro‐PO transcript was identified by Northern blot analysis using D. immitis microfilariae‐inoculated A. subalbatus. Both in situ hybridization and Northern blot analysis demonstrated that As‐pro‐PO mRNA was synthesized in mosquito haemocytes but not in other tissues, i.e. fat bodies, midguts and ovaries, etc.
A member of the hsp70 gene familyfrom the Mediterranean fruit fly, Ceratitis capitata
Authors: Thanaphum S. , Haymer D. S.
Published: March 2003

A gene from the Mediterranean fruit fly (Medfly) Ceratitis capitata, belonging to the hsp70 family of genes, has been cloned and characterized. This gene appears to be an example of a heat shock cognate (hsc) gene based on a variety of structural and functional criteria. The heat shock cognate genes serve critical roles in chaperoning of cellular proteins, and they are expressed in a more constitutive manner compared to the inducible heat shock genes. We determined the sequence of the entire coding region of this gene (designated Cerhsc1) together with 5′ and 3′ flanking regions. The coding region exhibits specific structural features that are characteristic of the subclass of heat shock genes producing proteins localized to the cytosol. The 5′ regulatory region of Cerhsc1 contains only the minimal array of canonical heat shock element sequences, a phenomenon commonly seen in heat shock cognates. Abundant transcripts are detected from this gene in adult tissue compared to the larval stage. In larval tissue, heat shock appears to produce only a slight increase in the level of transcripts detected. Based on overall structure and function, this Medfly gene appears to be an ortholog of heat shock cognate gene 4 (Drohsc4) of Drosophila melanogaster.
Characterization of the soluble guanylyl cyclase β‐subunit gene in the mosquito Anopheles gambiae
Authors: Caccone A. , Garcia B. A. , Mathiopoulos K. D. , Min G.‐S. , Moriyama E. N. , Powell J. R.
Published: March 2003

Genomic DNA corresponding ot the soluble guanylyl cyclase β‐subunit (GCSβ) gene was cloned and sequenced form Anopheles gambiae. The sequence was 8103 bp long and presumably included the entire coding region. The deduced amino acid sequence was 71% and 62% similar to previously known Drosophila and vertebrate GCSβ, while the C‐terminus of A. gambiae GCSβ was shorter. Because of the conserved characteristics in each functional domain, the high G+C % in the third codon positions compared to the introns, the lack of internal stop codons, and the fact that we identified the gene from a cDNA, we conclude that this A. gambiae gene is functional. This is the first detailed description of a guanylyl cyclase gene structure (e.g. intron–exon boundaries). Interstingly, within the fifth intron we found high similarity to the flanking regions of the Pegasus‐27 transposable element and other noncoding regions of the A. gambiae genome.
Isolation and characterization of insect PC2‐like prohormone convertase cDNA
Authors: Mentrup B. , Londershausen M. , Spindler K.‐D. , Weidemann W.
Published: March 2003

Endoproteolytic processing of large precursor molecules at basic amino acid residues plays an important role in the maturation of many hormones, neuropeptides and other regulatory proteins. Enzymes performing these reactions are designated as prohormone or proprotein convertases and belong to the subtilisin family of serine proteases. The screening of a larval cDNA library of the sheep blowfly Lucilia cuprina resulted in the isolation of two cDNAs encoding a PC2‐like prohormone convertase. The predicted 675 amino acid preproprotein (LcuPC2) exhibits its highest identity to invertebrate and vertebrate prohormone convertase 2 homologues, and a noticeably lower identity to the so far known insect furin‐like prohormone convertases of Drosophila melanogaster and Aedes aegypti. In Northern blot experiments a signal at 2.5 kb could be detected.
Semliki Forest virus as an expression vector in insect cell lines
Authors: Pettigrew M. M. , O'neill S. L.
Published: March 2003

Studies were undertaken to determine if replication‐deficient Semliki Forest virus expression vectors could be successfully used to express foreign gene constructs in insect cell lines. Using green fluorescent protein (GFP) as a marker we recorded infection levels of nearly 100% in the Aedes albopictus cell lines C6/36 and Aa23T, as well as in the Ae. aegypti cell line MOS20. The virus was capable of infecting an Anopheles gambiae cell line MOS55. The amount of GFP protein produced in each cell line was quantified. Northern analysis of viral transcription revealed the presence of novel transcripts in Aa23T, C6/36, and MOS55 cell lines, but not in the BHK or MOS20. The initial characterization of these transcripts is described.
Mariner (Mos1) transposase and genomic integration of foreign gene sequences in Bombyx mori cells
Authors: Wang W. , Swevers L. , Iatrou K.
Published: March 2003

Widespread occurrence in insects and the capacity to transpose in the absence of host‐derived factors means that mariner‐like elements are considered to be attractive candidates for the development of a universal insect genetic transformation system. Here we show that the Mos1 mariner element of Drosophila mauritiana is capable of mediating excision and transposition events in a silkmoth (Bombyx mori) derived tissue culture cell line (Bm5 cells). Plasmid rescue assays, in combination with Southern hybridization and polymerase chain reaction (PCR) analyses, confirm that the Mos1 transposase can mediate excision of DNA sequences, inserted between terminally repeated sequences recognized by the transposase, and integration into the chromosomal DNA of the Bm5 cells. In addition to chromosomal integration events, inter‐ and intraplasmid transposition and target element excision events were also detected. Approximately 50% of the plasmids recovered from plasmid rescue assays were found to contain the ‘signature’ of Mos1‐specific excision and/or integration events, indicating that the mariner transposase functions efficiently in the Bombyx cells. Because mariner‐induced excision and integration events are strictly dependent on the presence of a co‐transfected Mos1 transposase expression vector, it is clear that the multiple copies of endogenous mariner‐like elements (Bmmar1) that exist in the Bombyx genome are neither functional nor do they interfere with the efficiency of the transposition process. Thus, the Mos1 element and, probably, mariner elements, in general, hold great promise for the development of genetic transformation systems for lepidopteran insects.
Evidence for extensive genetic differentiation among populations of the malaria vector Anopheles arabiensis in Eastern Africa
Authors: Donnelly M. J. , Townson H.
Published: March 2003

We describe the geographical population structure of the malaria vector Anopheles arabiensis in Eastern Africa. Allelic variation at eight microsatellite loci was scored in samples from nine localities along a 4500 km transect from Sudan to Mozambique. Highly significant differences in genotype frequencies were found between all populations separated by more than 200 km. Populations within Malawi separated by 191 km were indistinguishable, as were those within Sudan separated by 134 km. FST and ρST gave significant estimates of isolation by distance. These data, lead us to conclude that there are extensive barriers to gene flow in this region. The high estimates of Nm (9.4 from FST and 5.2 from ρST) indicate recent range expansion in this species rather than extensive contemporary gene flow.
Genetic transformation of the housefly Musca domestica with the lepidopteran derived transposon piggyBac
Authors: Hediger M. , Niessen M. , Wimmer E. A. , Dübendorfer A. , Bopp D.
Published: March 2003

The piggyBac transposable element was successfully used for stable genetic transformation of the housefly Musca domestica. The construct contains the EGFP marker under the control of Pax‐6 binding sites, which can drive eye‐specific expression in insect species as distantly related as Drosophila melanogaster and Tribolium castaneum[Berghammer, A.J., Klingler, M. and Wimmer, E.A. (1999)Nature 402: 370–371]. We obtained seven independent integration events among 41 fertile G0Musca flies. Most of the transformed lines contained two or more chromosomal insertions of the EGFP marker which were stably inherited over more than 15 generations. piggyBac‐mediated transposition was verified by identifying the characteristic TTAA duplication at the insertion sites. This first report of stable transmission of a genetic marker in Musca confirms the use of this vector‐marker system for effective gene transfer in a broad range of insect species.
The juvenile hormone binding protein of silkworm haemolymph: gene and functional analysis
Authors: Vermunt A. M. W. , Kamimura M. , Hirai M. , Kiuchi M. , Shiotsuki T.
Published: March 2003

A cDNA fragment of haemolymph juvenile hormone binding protein (hJHBP) from larvae of Bombyx mori was amplified by RT‐PCR using degenerate primers based on the N‐terminal amino acid sequence of purified hJHBP and a conserved region near the C‐terminus of other lepidopteran hJHBPs. 5′‐ and 3′‐ends were amplified by RACE to yield cDNAs, hJHBP1 and hJHBP2, encoding 225 amino acids with three substitutions. hJHBP‐mRNA levels in the fat body were constant in the 4th instar, but decreased in the 5th. JHBP protein was constant until wandering, then declined. Recombinant hJHBP1 expressed in E. coli migrated on SDS‐PAGE with a Mr of 32 kDa and showed a Kd of 4.5 × 10−7 M with JH III, both similar to those of native hJHBP.
Louse (Insecta: Phthiraptera) mitochondrial 12S rRNA secondary structure is highly variable
Authors: Page R. D. M. , Cruickshank R. , Johnson K. P.
Published: March 2003

Lice are ectoparasitic insects hosted by birds and mammals. Mitochondrial 12S rRNA sequences obtained from lice show considerable length variation and are very difficult to align. We show that the louse 12S rRNA domain III secondary structure displays considerable variation compared to other insects, in both the shape and number of stems and loops. Phylogenetic trees constructed from tree edit distances between louse 12S rRNA structures do not closely resemble trees constructed from sequence data, suggesting that at least some of this structural variation has arisen independently in different louse lineages. Taken together with previous work on mitochondrial gene order and elevated rates of substitution in louse mitochondrial sequences, the structural variation in louse 12S rRNA confirms the highly distinctive nature of molecular evolution in these insects.
Arrangement and structural conservation of the mitochondrial control region of two species of Plecoptera: utility of tandem repeat‐containing regions in studies of population genetics and evolutionary history
Authors: Schultheis A. S. , Weigt L. A. , Hendricks A. C.
Published: March 2003

Low levels of primary sequence similarity across insect taxa have led to the suggestion of conserved structural elements in the insect mitochondrial control region. Our aim was to determine whether previously described motifs and secondary structures exist in stoneflies (Plecoptera). Several motifs and structural elements previously described in Orthoptera and Diptera were found, including a conserved ‘hairpin’ structure that may play a role in the initiation of mtDNA replication. The repeat region had the highest percentage similarity, lowest A‐T content and highest transition to transversion ratio, suggesting a unique evolutionary pattern for the repeats. Finally, we discuss the usefulness of the control region in population genetic and evolutionary studies.
Vertical root fractures in adjacent maxillary premolars: an endodontic‐prosthetic perplexity
Authors: Tamse  , Zilburg  , Halpern
Published: March 2003

Vertical root fractures in endodontically treated teeth occur frequently in teeth or roots in which their mesiodistal dimension is narrow, such as the maxillary upper premolars. Two cases of vertical root fractures in two adjacent maxillary reconstructed teeth are presented. As in many cases of vertical root fractures, the primary diagnosis was of endodontic treatment failure. The final diagnosis of the fractures was made either by the radiograph showing separation of root segments in one case, or by surgical exploration in the other. In both cases, tooth extraction was unavoidable.
Antimicrobial effects of various endodontic irrigants on selected microorganisms
Authors: Ayhan H. , Sultan N. , Çirak M. , Ruhi M. Z. , Bodur H.
Published: March 2003

Aim This study was undertaken to determine the antimicrobial effect of various endodontic irrigants against six selected microorganisms

Methodology Staphylococcus aureus, Enterococcus faecalis, Streptococcus salivarius, Str. pyogenes, Escherichia coli and Candida albicans were included in the study. Pre‐sterilized Whatman paper discs, 6 mm in diameter and soaked with the test solution, were prepared and placed onto the previously seeded agar Petri plates. Each plate was incubated aerobically. A zone of inhibition was recorded for each plate and the results were analysed statistically.

Results 5.25% NaOCl was effective against all test microorganisms with a substantial zone of inhibition. Saline was always ineffective. Decreased concentration of NaOCl significantly reduced its antimicrobial effect. Cresophene showed a significantly larger (P > 0.05) average zone of inhibition compared to the other experimental irrigants. Alcohol had smaller but not significantly different zones of inhibition than chlorhexidine.

Conclusions 5.25% NaOCl was superior in its antimicrobial abilities compared with other irrigants used. A reduced concentration of NaOCl (0.5%) resulted in significantly decreased antimicrobial effects. When compared with 21% alcohol, 0.5% NaOCl and 2% chlorhexidine, paramonochlorophenol (cresophene) showed a greater antimicrobial effect.
Micro‐computed tomography: a new tool for experimental endodontology
Authors: Rhodes J. S. , Ford T. R. Pitt , Lynch J. A. , Liepins P. J. , Curtis R. V.
Published: March 2003

Aim Micro‐computed tomography (MCT) using cone‐beam geometry is a method of producing true 3D images of the structure of small samples. A prototype MCT unit was adapted for imaging teeth to examine whether it could be used to quantify the instrumentation of root canals.

Methodology Ten mandibular first molar teeth that had intact crowns and fully formed roots were scanned using MCT at a resolution of 0.081mm and 3D‐rendered images created; root canals were segmented from this. Reproducibility of MCT was verified for root canal shape and size. Access cavities were prepared into the pulp space and root canals enlarged to a continuously tapering preparation using a crowndown technique. Each tooth was scanned again to allow comparison of pre‐ and post‐instrumentation images. The roots were then sectioned at five predeterminedhorizontal levels for video‐digitized measurement of dimensions of roots and root canals. The video images had a resolution of 0.025mm. Video‐digitized images of the physical cut surfaces were compared with equivalent MCT reconstructed images. The total area of the root canals (internal) and root (external) at each level were calculated from both MCT reconstructions and video‐digitized images, and compared.

Results There was a highly significant correlation between MCT and video images for both external and internal areas (r = 0.94). Rendered 3D images were constructed to show the root canal systems of teeth. The total volumes of the apical 7.5mm of root canals were calculated from rendered images of nine teeth before and after instrumentation. The mean amount of dentine removed by instrumentation was 3.725mm3, which was 28% of the original canal volume.

Conclusions Micro‐computed tomography was shown to be accurate for experimental endodontology.
Cleaning efficacy of two apical preparation regimens following shaping with hand files of greater taper
Authors: Lumley P. J.
Published: March 2003

Aim

The aim of this investigation was to assess canal cleaning following shaping with hand files of greater taper.

Methodology

Thirty mesial canals and 30 distal canals in mandibular molars were prepared with .08 and .10 hand files of greater taper, respectively. Following initial preparation, 0.02 instruments were used incrementally shorter in a stepback manner. In half the canals instrumentation was performed to size 35 1 mm short of the canal terminus. In the other half the series was continued through to size 60 using stepback increments of half to one millimetre as appropriate (foramen size was maintained at size 20 in all groups). Sodium hypochlorite (4.5%) and REDTA (17%) were used as irrigants for all groups. Cleaning efficacy was evaluated by scoring the amount of remaining debris using a light microscope (×50) and calibrated eyepiece micrometer (range 0–3).

Results

Canals stepped back through to a size 60 were significantly cleaner than those instrumented to a size 35 only (Mann–Whitney P < 0.05).

Conclusion

Refining the apical region of canal preparation by stepping back through to a size 60 reduced remaining debris following shaping with files of greater taper.
Cleaning effectiveness of root canal irrigation with electrochemically activated anolyte and catholyte solutions: a pilot study
Authors: Solovyeva A. M. , Dummer P. M. H.
Published: March 2003

Aim

The aim of this study was to evaluate the potential of electrochemically activated (ECA) anolyte and catholyte solutions to clean root canals during conventional root canal preparation.

Methodology

Twenty extracted single‐rooted human mature permanent teeth were allocated randomly into four groups of five teeth. The pulp chambers were accessed and the canals prepared by hand with conventional stainless steel endodontic instruments using a double‐flared technique. One or other of the following irrigants was used during preparation: distilled water, 3% NaOCl, anolyte neutral cathodic (ANC) (300 mg L−1 of active chlorine), and a combination of anolyte neutral cathodic (ANC) (300 mg L−1 of active chlorine) and catholyte. The teeth were split longitudinally and the canal walls examined for debris and smear layer by scanning electron microscopy. SEM photomicrographs were taken separately in the coronal, middle and apical parts of canal at magnification of ×800 to evaluate the debridement of extra‐cellular matrix and at a magnification of ×2500 to evaluate the presence of smear layer.

Results

Irrigation with distilled water did not remove debris in the apical part of canals and left a continuous and firm smear layer overlying compressed low‐mineralized predentine. All chemically active irrigants demonstrated improved cleaning potential compared to distilled water. The quality of loose debris elimination was similar for NaOCl and the anolyte ANC solution. The combination of anolyte ANC and catholyte resulted in improved cleaning, particularly in the apical third of canals. The evaluation of smear layer demonstrated that none of the irrigants were effective in its total removal; however, chemically active irrigants affected its surface and thickness. Compared to NaOCl, the ECA solutions left a thinner smear layer with a smoother and more even surface. NaOCl enhanced the opening of tubules predominantly in the coronal and middle thirds of canals, whereas combination of ANC and catholyte resulted in more numerous open dentine tubules throughout the whole length of canals.

Conclusions

Irrigation with electrochemically activated solutions cleaned root canal walls and may be an alternative to NaOCl in conventional root canal treatment. Further investigation of ECA solutions for root canal irrigation is warranted.
Some factors affecting the concentration of available chlorine in commercial sources of sodium hypochlorite
Authors: Frais S. , Ng Y‐L. , Gulabivala K.
Published: March 2003

Aims To evaluate the availability, effect of storage, dilution and heating on some commercial sources of sodium hypochlorite (NaOCl).

Methodology All pharmacies in a Regional Health Authority were telephone surveyed to establish the proprietary and nonproprietary NaOCl products available, their cost and concentration. Ninety‐six freshly produced bottles of a commercially available thin household bleach (J. Sainsbury’s) were randomly stored in four different modes (full 1 L bottles in dark at room temperature, half‐empty 1 L bottles under the same conditions, full 200 mL bottles under the same conditions, full 200 mL bottles stored at 37 °C) up to 6 months. The available chlorine in the bleach solutions was evaluated at specific time intervals (0, 1, 2 weeks, 1, 2, 3, 4, 6 months) using iodometric titration. Four brands of commercially available bleaches (J. Sainsbury’s Thin Household Bleach, Tesco’s Value Bleach, Tesco’s Red Label Bleach, Safeway’s Savers Bleach) were diluted (by 1 : 1, 1 : 2 or 1 : 3 dilution factors) and samples taken from each dilution for titration to test predictability. NaOCl solutions (5% w/w) were heated in covered or uncovered beakers up to 60–85 °C. Samples were taken from each beaker at time 0 and each h up to 4 h, for titration.

Results Of the 116 pharmacies, 33% could supply a proprietary product and 53% a nonproprietary product. In all, 78% were able to supply a NaOCl product. There was no significant difference (P < 0.05) in available chlorine at baseline between the different modes of storage. After storage at room temperature for 6 months, there were no significant differences (3% of original; P > 0.05) between the different modes of storage. When the storage temperature was 37 °C, however, there was a significant difference (38% of original; P < 0.01) at 6 months. Comparison of baseline with the concentrations of available chlorine at 6 months showed significant differences (P < 0.01) for all conditions. The available chlorine concentration in four brands of ‘thin’ supermarket bleaches could be predictably reduced according to the dilution factor. When 5% NaOCl was heated to 60–85 °C for up to 4 h, the available chlorine was increased to 6% in covered solutions and 9% in uncovered solutions.

Conclusions Both proprietary and nonproprietary solutions of NaOCl should be relatively easily obtainable in the UK. Stored under appropriate conditions, commercially available thin bleaches and nonproprietary solutions of NaOCl may be diluted to obtain predictable concentrations. Heating solutions of NaOCl may cause unpredictable changes to the concentration, depending upon conditions.
Polymerase chain reaction detection of Treponema denticola in endodontic infections within root canals
Authors: Siqueira J. F. , Rôças I. N. , Favieri A. , Oliveira J. C. M. , Santos K. R. N.
Published: March 2003

Aim

A 16S rDNA‐based polymerase chain reaction (PCR) method was used to detect the occurrence of Treponema denticola in root canals.

Methodology

Samples were collected from 54 single‐rooted teeth having carious lesions and necrotic pulps. DNA extracted from the samples was amplified using the PCR assay, which yielded a specific fragment of T. denticola 16S rDNA.

Results

T. denticola was detected in 10 of 29 asymptomatic cases (34.5%), eight of 15 root canals that were tender to percussion (53.3%), and five of 10 teeth with acute periradicular abscess (50%). In general, T. denticola was found in 23 of 54 cases (42.6%).

Conclusions

Findings suggest that T. denticola can be involved in the pathogenesis of periradicular lesions of endodontic origin.
Incidence of root fractures and methods used for post removal
Authors: Abbott P. V.
Published: March 2003

Aim This study was undertaken to determine the incidence of root fractures during post removal, as well as the methods used and the success rates achieved with various post removal devices and techniques.

Methodology Patient records from a specialist endodontic practice were examined. A total of 1600 teeth from which posts were removed were identified. These teeth had no preoperative signs or symptoms to suggest root fractures. The incidence of root fractures was determined and a subset of 234 patient records were examined in detail to determine the methods used to remove different types of posts.

Results Only one tooth fractured during post removal (0.06%) and all posts were successfully removed, typically in about 3 min. Most cast posts were removed with an Eggler post remover but some dislodged when the crown was removed; ultrasonics was used occasionally. Most parallel‐sided posts were removed with ultrasonics and threaded posts were unscrewed. The Masserann kit and ultrasonics were combined to remove fractured cast posts and some parallel‐sided posts.

Conclusions This study indicates that, with good case selection, post removal is a predictable procedure. If appropriate techniques and devices are used for the particular type of post being removed, then root fracture is a rare occurrence.
Developmental changes and regional differences in histochemical localization of hyaluronan and versican in postnatal molar dental pulp
Authors: Shibata S. , Yoneda S. , Yanagishita M. , Yamashita Y.
Published: March 2003

Aim The main aim of this study was to investigate the developmental changes in the distribution patterns of hyaluronan (HA) and versican in postnatal rat molar dental pulp, in order to confirm the hypothesis that the distribution of both molecules can vary with physiological conditions in the dental pulp.

Methodology Thirty postnatal Sprague‐Dawley rats, 1, 7, 14, 21, 28, 35, 42 and 49 days old, were used for this study. Immunohistochemistry for versican with monoclonal antibodies 12C5 and CS‐56 and histochemical staining for HA with HA‐binding protein were applied to paraffin sections of the mandibular first molars at each age.

Results At day 1, both molecules were evenly distributed in the interior parts of the pulp, but strong reactions for both molecules appeared in the subodontoblastic layer of the coronal pulp by the completion of crown formation. However, a strong reaction for HA and a weak reaction for versican were seen in the subodontoblastic layer of the radicular pulp. Furthermore, a versican‐deficient, low‐HA area first appeared in the interior of the coronal pulp at day 42 and expanded at day 49.

Conclusions Distribution of hyaluronan and versican in the dental pulp varied with age and also showed regional differences between the coronal and the radicular pulp, and this supports the hypothesis described above.
In vivo performance of the new non‐instrumentation technology (NIT) for root canal obturation
Authors: Lussi A. , Suter B. , Fritzsche A. , Gygax M. , Portmann P.
Published: March 2003

Aim The aim of this in vivo study was to compare the radiographic quality of root fillings performed by the NIT‐obturation method versus conventional mechanical obturation.

Methodology

Sixty‐six patients needing root canal treatment participated in this study. The treatments were performed by three private practitioners. The root canals were instrumented with K‐Flexofiles to a master apical file between sizes 25 and 60, followed by step‐back flaring up to size 70. Copious irrigation was used throughout the instrumentation procedure with NaOCl (3%). The teeth were obturated either by lateral condensation, the McSpadden technique (control) or by the new non‐instrumentation technology (NIT) with and without using gutta‐percha points. In the NIT method, a low pressure was created within the tooth, and AH 26 sealer was sucked into the root canal system. Radiographs of the root‐filled teeth were analysed and the length of the root filling, the presence of voids and the area of any other fillings determined.

Results

The root canal fillings of the control group (0.1 ± 0.1 mm) and those of the NIT/gutta‐percha group (0.3 ± 0.1 mm) were both overextended when taking the apical constriction as a reference point. Root canal fillings of the NIT/gutta‐percha group were statistically (P < 0.05) significantly longer than those of the NIT without gutta‐percha group. The latter showed slightly underextended root canal fillings (–0.14 ± 0.1 mm).

Conclusions The present investigation demonstrated the performance of the NIT‐obturation method in vivo. Root canals filled by the reduced‐pressure‐method using sealer combined with gutta‐percha cones exhibited equivalent radiographic quality compared to conventionally filled canals.
Monitoring pulp vitality after transplantation of teeth with mature roots: a case report
Authors: Siers M. L. , Willemsen W. L. , Gulabivala K.
Published: March 2003

Aim To initiate discussion on the value of routine root canal treatment for transplanted teeth.

Summary Autotransplantation is an accepted treatment option to replace missing teeth. It is generally considered that revascularization of the pulp following such a procedure is more favourable in teeth with immature roots. In teeth with closed apices root canal treatment is considered necessary. This paper presents a case of pulp revascularization in a transplanted tooth with mature roots and casts doubt on whether root canal treatment is essential in such situations. An alternative treatment protocol is proposed.

Key learning points

Following transplantation original pulp tissue may survive the operation.

Teeth with obliterated pulp space do not become necrotic more often than those without obliteration.

Monitoring the tooth is an acceptable alternative to automatic root canal treatment for transplanted teeth.

Root canal treatment should be undertaken only upon occurrence of pathological signs.
Interstitial cell tumour and germ cell tumour with carcinoma in situ in rabbit testes
Authors: Veeramachaneni  , Vandewoude
Published: March 2003

Simultaneous occurrence of a well‐demarcated interstitial cell tumour and an intratubular seminoma‐like tumour, which was beginning to invade peritubular areas, in the contralateral testes of a 3‐year‐old Dutch‐Belted rabbit is described. Morphological hallmarks of carcinoma in situ, which have not been reported previously for the rabbit, were observed in association with the seminoma. These observations indicate that carcinoma in situ, preceding a seminoma‐like tumour, occurs in the rabbit and that the rabbit may serve as a practically useful animal model for studying testicular germ cell neoplasia.
Imipramine for successful treatment of retrograde ejaculation caused by retroperitoneal surgery
Authors: OCHSENKÜHN  , KAMISCHKE  , NIESCHLAG
Published: March 2003

Retrograde ejaculation is a known complication following retroperitoneal lymph node dissection. Without further therapy, achieving paternity may be impossible. We evaluated the use of the tricyclic antidepressant imipramine in a new ovarian cycle‐dependent dose regime for reversal of retrograde ejaculation in 11 patients desiring fertility. Ten patients with retroperitoneal lymph node dissection performed because of testicular cancer, and one with aortic surgery (thromboendarterectomy) were treated in an open, uncontrolled study with imipramine given at a daily oral dose increasing from 25 to 50 mg for 7 days prior to the planned ejaculation or the expected ovulation of the female partner. In all 11 patients, antegrade ejaculation could be induced (sperm counts: 3.9–276.0 × 106/mL). Major side‐effects were not observed, but half of the patients complained of minor degrees of dizziness, weakness, nausea or sweating during medication. Under treatment, two patients with normal sperm concentrations induced a spontaneous pregnancy. One ICSI cycle each was performed in 2 patients, with successful fertilization, but no pregnancy. In conclusion, temporary oral intake of imipramine is an effective and safe treatment to re‐establish antegrade ejaculation in patients with retrograde ejaculation following retroperitoneal surgery, thus providing possibilities for paternity either through intercourse or by assisted fertilization.
The effects of isolated lipoproteins and triglyceride, combined oxidized low density lipoprotein (LDL) plus triglyceride, and combined oxidized LDL plus high density lipoprotein on the contractile and relaxation response of rabbit cavernous smooth muscle
Authors: Kim Sae Chul , Seo Kyung Kuen , Kim Hyun Woo , Lee Moo Yeol
Published: March 2003

The aim of this study was to investigate the effects of isolated lipoproteins and triglyceride (TG), and the effects of combined oxidized low density lipoprotein (LDL) plus TG and the combined oxidized LDL plus high density lipoprotein (HDL) on the contractility and relaxation response of rabbit cavernous smooth muscle. Cavernous muscle strips from New Zealand White rabbits were studied in organ chambers for isometric tension measurement. The strips were exposed to HDL, LDL, oxidized LDL, TG, combined oxidized LDL plus TG and combined oxidized LDL plus HDL for 30 min. Both HDL and LDL did not affect contraction and relaxation responses of the cavernous muscles. The oxidized LDL did not affect norepinephrine (NE)‐induced contractility of the strips, but significantly (p < 0.05) decreased the relaxation response to endothelium‐dependent agonist, acetylcholine (Ach). Non‐specific NO synthase inhibitor (L‐NAME) completely inhibited the relaxation response to Ach, and L‐arginine partially improved the diminished relaxation. TG did not significantly change the relaxation responses to Ach, but decreased the contractility of cavernous muscle to NE. Neither the combined oxidized LDL plus TG nor oxidized LDL plus HDL had significant synergistic or detoxication effects on the contractility and relaxation responses. In conclusion, oxidized LDL may have acute toxic effects on the endothelium‐dependent, NO‐mediated relaxation, but not on the contractility, of rabbit cavernous smooth muscle. TG may decrease contractility of the cavernous muscle. There may be neither synergistic nor detoxication effects on the contractility and relaxation response when TG or HDL is added to the oxidized LDL.
Proteasomes in human spermatozoa
Authors: Wojcik  , Benchaib  , Lornage  , Czyba  , Guerin
Published: March 2003

In the present study we describe the localization of proteasomes in human spermatozoa by means of immunolabelling with different monoclonal and polyclonal antibodies detected by confocal microscopy. Western blotting confirmed the specificity of the antibodies and has shown that proteasomes are present in spermatozoa and in seminal fluid. In spermatozoa proteasomes are concentrated in the neck region where the centrioles are located. Some labelling was also detected at the periphery of the head, but no proteasomal antigens were detected in either the nucleus or associated with the flagellum. Proteasome inhibitors did not affect the motility of the spermatozoa, acrosome reaction nor zona binding. It is hypothesized that paternal proteasomes enter the oocyte during fertilization in tight association with the centrioles and may serve a special function during further development which can be associated with the function of a hypothetical proteolysis centre.
The advantages of standardized evaluation of male infertility
Authors: Pierik Frank H. , Van Ginneken Astrid M. , Dohle Gert R. , Vreeburg Jan T. M. , Weber Robertus F. A.
Published: March 2003

Healthcare can be improved by standardization and by evaluation of diagnostic methods and treatments. In the field of andrology, in which large patient numbers are required for the evaluation of diagnostic procedures and treatments, structured data collection and multicentre studies are especially warranted. Concomitant with routine clinical practice, a large amount of clinical data are collected that may be used to evaluate andrological care. Structuring and electronic storage of data holds promise in terms of clarity and accessibility of the data and its use for validation studies. The aim of the present work was to study the merits of routine collection of a common dataset in a computer‐based patient record (CPR) for standardization, quality of data and clinical research. It was studied whether the data were of sufficient quality and accessibility for much needed studies on aetiology, interventions and diagnostics in andrology.

Data collection in a structured CPR promoted complete and comprehensive data. We describe the advantages, pitfalls and solutions with this approach. Data on the uniform examination of 1549 infertile men became readily accessible. Population characteristics, basal associations and original studies were enabled and provided insight into the efficiency of clinical practice. In 66% of men, a cause for their infertility was identified, which provides a better rationale for treatment than semen parameters alone. For more than 30% of the patients, a rational andrological treatment was available, which could be deployed before assisted reproductive technologies were resorted to. However, most treatments have not been properly validated. The thorough diagnostic evaluation identifies subgroups that require an evidence base for treatment and further study on aetiology and diagnosis.

Structured collection of uniform patient data through a CPR was feasible and facilitated the evaluation of diagnostic and therapeutic modalities. The reported advantages, pitfalls and solutions with this approach may help other centres to decide on how to implement a CPR. Conscientious collection of a standard data set in infertility centres facilitates pooling of data and evidence‐based multicentre research.
Accuracy of the normal sperm morphology value by Sperm Quality Analyzer IIC: comparison with the strict criteria
Authors: Shibahara Hiroaki , Suzuki Tatsuya , Obara Hiromi , Hirano Yuki , Onagawa Tamaho , Taneichi Akiyo , Takamizawa Satoru , Sato Ikuo
Published: March 2003

The aim of the present study is to investigate the accuracy of the normal sperm morphology value by Sperm Quality Analyzer IIC (SQA IIC), which was developed to provide a rapid and low‐cost quantitative evaluation of semen quality. Normal sperm morphology was assessed using SQA IIC in comparison with that by the strict criteria in 62 semen samples. Normal sperm morphology value by SQA IIC was based on the studies of three traditional sperm parameters from over 4000 fresh, untreated semen samples, while the strict criteria was based on the method by Kruger et al. The mean ± SD of percent normal morphology by SQA IIC and the strict criteria were 37.6 ± 10.9% (range 15–52) and 19.9 ± 8.2 (range 1–34), respectively. There was a significant correlation of the sperm morphology assessment between the two methods (r=0.454, p < 0.001). Using the cut‐off value of >30% normal morphology by SQA IIC, the positive predictive value and the negative predictive value of the `normal' strict criteria were 79.6% (39/49) and 46.2% (6/13), respectively. These results indicate that SQA IIC might be used as an initial screening test for the evaluation of sperm morphology. However, sperm morphological assessment by the strict criteria should be performed in order to make decisions in planning strategies for the treatment of infertile couples.
Capacitation status and in vitro fertility of boar spermatozoa: effects of seminal plasma, cumulus‐oocyte‐complexes‐conditioned medium and hyaluronan
Authors: Suzuki K. , Asano A. , Eriksson B. , Niwa K. , Nagai T. , Rodriguez‐Martinez H.
Published: March 2003

In the present study, the effects of seminal plasma (SP), cumulus–oocyte‐complexes (COCs) conditioned medium (CCM) and hyaluronan (HA) on functional changes and in vitro fertilizing ability of porcine spermatozoa were examined. In in vitro fertilization (IVF) experiments, 10% (v/v) of exogenous SP in the fertilization medium prevented sperm penetration (using fresh‐extended and frozen–thawed ejaculated spermatozoa). Analysis of frozen–thawed CCM revealed a HA content to levels of 30 ng/mL per incubated COC. Presence of frozen–thawed CCM did not, however, prove effective to increase (furthermore decreasing) oocyte penetration in vitro, and neither did supplementation with exogenous HA at the same concentration as that present in the CCM (secreted by COCs). Analysis of sperm capacitation using the chlortetracycline (CTC) assay showed that frozen‐thawed CCM had no elevating effect on `B‐pattern' spermatozoa (implying capacitation‐like changes) and that addition of 10% (v/v) SP held spermatozoa in the `F‐pattern' (intact) status. Dose of 500 μg/mL HA and freshly prepared CCM increased, however, the frequency of capacitated spermatozoa (B‐pattern) without resulting in increased rates of `AR‐pattern' (acrosome‐reacted) spermatozoa, compared with controls. The present results confirm the decapacitating effect of SP and suggest capacitating actions of HA (dose‐related) and CCM (freshly prepared) on boar spermatozoa in vitro. The unclear effects of frozen‐thawed CCM and a low dose of HA on penetration rates of boar spermatozoa call for further researches of their function in vivo.
IL‐10 is highly expressed in the cryptorchid cryptepididymal epithelium: a probable mechanism preventing immune responses against autoantigenic spermatozoa in the epididymal tubule
Authors: Veräjänkorva E. , Pöllänen P. , Hänninen A. , Martikainen M. , Sundström J. , Antola H.
Published: March 2003

The expression of several immunoregulatory adhesion proteins and cytokines was studied in the normal epididymis, cryptorchid cryptepididymis, the epididymis of oestrogen‐treated mice and the epididymis of non‐obese diabetic (NOD) mice at the protein level to see which of these immunoregulatory proteins may be involved in lymphocyte regulation in the normal or pathological epididymis and if cytokine balance in this organ is on the cellular or humoral side. The aim of the study was to characterize the immunological microenvironment of the epididymis to explain the survival of the autoantigenic spermatozoa in this site. In the 6‐week‐old BALB/c or NOD mouse epididymis there were some CD18 and CD44 expressing cells in the interstitial tissue. There were no differences between these strains in the expression of the studied antigens, except that some CD4 positive cells were present in the interstitial tissue of BALB/c mice. In the cryptorchid cryptepididymis CD4, CD8, CD18, CD44, CD54 and CD106 expressing cells were occasionally present in the connective tissue surrounding the epididymal tubule. In the epididymis of the oestrogen‐treated mice these antigens were not expressed. In the cryptorchid cryptepididymis the epithelial cells expressed IL‐10 highly and the myoid peritubular cells IL‐6. The present results suggest that the epididymal epithelial IL‐10 suppressing TH0, TH1 and TH2 immune responses may be involved in the protection of autoantigenic spermatozoa from immune destruction.
Association of oestrogen receptor α polymorphisms and androgen receptor CAG trinucleotide repeats with male infertility: a study in 109 Greek infertile men
Authors: Kukuvitis A. , Georgiou I. , Bouba I. , Tsirka A. , Giannouli CH. , Yapijakis C. , Tarlatzis B. , Bontis J. , Lolis D. , Sofikitis N. , Papadimas J.
Published: March 2003

This study was performed to examine the contribution of genetic polymorphism of oestrogen and androgen receptor (AR) genes in male infertility. We have studied in total 173 Greek men, 109 infertile patients and 64 controls (group A). Patients were divided in to three subgroups: group B (n=29) with idiopathic moderate oligospermia, group C (n=42) with azoospermia or idiopathic severe oligospermia and group D (n=38) with azoospermia or oligospermia of various known aetiologies. All patients and controls were genotyped for two polymorphisms of the oestrogen receptor α (ERα) gene and also for the (CAG)n repeat length polymorphism of the X‐linked androgen receptor (AR)gene. The control group had statistically significant difference from group C regarding the XbaI polymorphism of ERα gene. Despite the fact that we did not observe any statistically significant differences in the mean and range of the CAG repeat number, the frequency of the higher repeats of the nucleotide repeat sequence (CAG)n of the AR gene was 2–4 times higher in groups B and C compared with the control group A. Our results indicate that both ERα and AR gene play significant role in male fertility. It is possible that a synergy may exist between unfavourable genotypes of these two genes in male infertility.
Semen analysis and external quality control schemes for semen analysis need global standardization
Authors: Cooper T. G. , Björndahl L. , Vreeburg J. , Nieschlag E.
Published: March 2003

One semen analysis laboratory [the Institute of Reproductive Medicine (IRM), Münster, Germany] was enrolled in three external quality assurance programmes in Europe (United Kingdom External Quality Assurance Scheme, European Academy of Andrology, European Society of Human Reproduction) that control for the assessment of sperm concentration, sperm motility and sperm morphology. Agreement between the participating laboratory and the sperm concentrations designated by all three programmes was good. Disagreement between two quality control (QC) programmes providing motility assessment was particularly noticeable in the distinction between motility grades a and b. For the identification of normal sperm morphology, marked differences between the standards set by different QC programmes were apparent. One programme was stricter in its categorization of normal forms, such that an overestimation of normal forms was diagnosed at IRM, whereas agreement with the other programmes was achieved. Variation of results from technicians in the IRM internal QC programme was <13%. The discrepancies between external quality control (EQC) centres demonstrated here are challenging problems to be overcome partly by the andrology laboratories and partly by the providers of EQC services. The introduction of objective, computer‐aided sperm assessment in providing designated values may help to rectify this situation. Until this is achieved, EQC programmes should develop an internal programme to monitor their materials and methods for assessment.