Interspecies comparison of cellular localization of the cyanide metabolizing enzyme rhodanese within olfactory mucosa

The observation of high levels of xenobiotic metabolizing enzyme activity in the olfactory mucosa has produced speculation on the functional significance of these enzymes in the nose. Hypothesized roles include protection of the nasal epithelium, lung, and other downstream tissues, and termination or modification of olfactory responses. The enzyme rhodanese metabolizes cyanide, which is a commonly inhaled toxicant and an odorant and therefore of interest to both toxicologists and olfactory neurobiologists. The cellular localization of this enzyme within the olfactory mucosa will have important consequences for its ability to protect specific cells, as well as its ability to alter the concentration of inhaled cyanide at receptors, and therefore could provide clues as to its function in this tissue. We have compared the distribution of this enzyme in two species, the rat and the cow, using immunohistochemical localization techniques employing species-specific polyclonal antisera raised in our laboratory. In the rat, rhodanese-like immunoreactivity was greatest within the apical portion of the sustentacular cells, the basal cells, and the duct cells of Bowman's glands. Very little to no reaction was observed in the acinar cells of Bowman's glands. In the cow, however, the acinar cells and duct cells of Bowman's glands showed intense immunoreactivity with little to no reaction observed in the sustentacular or basal cells. The differences in localization of rhodanese in these two species may have important implications for cell types at risk during inhalation of cyanide or organonitrile compounds metabolized to cyanide within the nasal mucosa. In addition, the difference in distribution in the two species emphasizes the importance of considering enzyme activity and localization in the determination of an appropriate animal model for study of both nasal toxicology and olfactory responsiveness in humans.     

Streptococci from root canals in teeth with apical periodontitis receiving endodontic treatment.

OBJECTIVES: The object of this study was to investigate the diversity among streptococcal species isolated from root canals in conjunction with endodontic therapy and to characterize their production of extracellular proteins. STUDY DESIGN: Consecutive root canal samples (RCS) taken as bacteriological controls during root canal treatment of teeth with apical periodontitis were analyzed in a total of 100 clinical cases. Bacteria were isolated and classified by selective media and gas liquid chromatography. Streptococcal strains were identified by carbohydrate fermentation, hydrolysis of aesculin/arginine, and production of enzymes. Releases of extracellular proteins by streptococci and Enterococcus spp in fluid culture media were examined with SDS-PAGE and 2-dimension gel electrophoresis (2 DE). Extracellular proteins produced were quantified and qualitatively analyzed. Specific proteins were targeted with Western immunoblot assays. Comparisons were made with type strains. RESULTS: Of a total of 241 bacterial strains recovered in the first samples submitted, Streptococcus gordonii, S anginosus, and S oralis were the most frequently isolated streptococci. In 49 of 89 resubmitted samples showing bacterial growth, S gordonii and S oralis still predominated among streptococci. Other common bacterial isolates were Enterococcus spp, Lactobacillus paracasei, and Olsenella uli. Quantitative and qualitative differences in extracellular protein production were observed among clinical isolates and laboratory streptococcal strains. In similar conditions for growth, S intermedius, S anginosus, S oralis, and S gordonii were strong producers of extracellular proteins (>3.0 microg/mL), while Enterococcus spp and S mutans were weak. Whole cell protein extracts showed a different profile from that of extracellular proteins. The chaperone protein DnaK was recognized to be produced extracellularly by S gordonii, S oralis, S anginosus, and S parasanguis. CONCLUSIONS: Being strong producers of extracellular proteins and by virtue of common presence in teeth undergoing endodontic therapy, S gordonii, S anginosus, and S oralis may be of pathogenic significance in posttreatment apical periodontitis.     

Basal cell carcinoma. An epidemiologic study in a defined population.

BACKGROUND. The incidence of basal cell carcinoma (BCC) in a defined population is unknown. METHODS. The incidence of histologically diagnosed cases of BCC in a defined population of a city in southern Sweden was studied. Four 2-year periods were examined from 1970 to 1986. Cases from 1970 and 1980 were reexamined microscopically. RESULTS. From 1970 to 1986, the age-standardized incidence of BCC was doubled. BCC was equally common in male and female patients. It was possible to trace another BCC in the medical history of 41% of the patients. The risk of development of BCC increased rapidly with age greater than 55 years. Approximately two-thirds of the tumors were found in the head and neck region. There was a possible increase in superficial multicentric tumors on the trunk but otherwise an essentially unchanged pattern in type and distribution. CONCLUSIONS. The age-standardized incidence of histologically diagnosed BCC was doubled from 1970 to 1986. The rapid increase should be taken into consideration when planning allocation of medical resources.     

WHERE ARE THE GATES IN GAP JUNCTION CHANNELS?

1. In the formation and function of gap junction channels two types of gates ought to be discriminated: the docking gate and the channel gates proper. The docking gate is involved in the transformation of a closed hemichannel to a patent gap junction channel. By definition the trigger mechanism for this gate and maybe even the gate itself is contained within the extracellular loops of the gap junction proteins, the connexins. The channel gates proper determine the open and closed states of the complete gap junction channels. 2. Probing the docking gate by mutagenesis of connexins and by synthetic peptides indicates that this gate is the consequence of complex interactions between a large fraction of the amino acids comprising the extracellular loops. Probably both inter- and intra-molecular interactions are involved, and disulfide exchange may be entailed in the stabilization of the open and closed states. 3. Of the various effectors on the channel gate(s) the voltage effects have obtained the most scrutiny to date. The response of gap junction channels and hemichannels is diverse, the various channels respond differently to transjunctional and membrane potential. No equivalent to the S4 segment representing the voltage sensor in other voltage dependent ion channels is present in the connexin sequences, instead mutations in various segments of connexins have been reported to affect the voltage dependence of gap junction channels. To understand the complexity of voltage effects on gap junction channels, non-connexin peptides may need to be considered as voltage sensors or as modifiers thereof.     

Case-control study of risk factors for cervical neoplasia in Denmark. II. Role of sexual activity,reproductive factors,and venereal infections

Sexual, reproductive and venereal risk factors for cervical neoplasia were investigated in a population-based case-control study of 586 women with histologically verified, cervical squamous-cell carcinoma in situ, and 59 women with invasive squamous-cell cervical cancer, diagnosed from 1985 to 1986 in Copenhagen. Cases were identified from the computerized Danish Cancer Registry. An age-stratified control group (n=614) was drawn at random from the female population in the study area by means of the Danish Central Population Register. A structured questionnaire was mailed to cases as well as controls. Increasing number of sexual partners exerted a significant effect on the risk both for carcinoma in situ, and invasive cancer, independently of age at first intercourse and other potential confounders. Conversely, the association with early age at first intercourse became statistically insignificant after allowance for other risk factors, although an increasing risk was still observed with decreasing age at sexual debut. Early age at first episode with genital warts was a significant risk factor for carcinoma in situ, perhaps indicating a possible increased susceptibility of the cervix epithelium during adolescence. A history of genital warts was a good predictor of risk for carcinoma in situ, whereas a history of previous gonorrhea was associated with an increased risk for invasive carcinoma. Women with multiple births had a significantly increased adjusted risk, especially for carcinoma in situ, although some association was also observed with invasive cervical cancer. The study supports the hypothesis of cervical neoplasia being a sexually transmitted disease, and that carcinoma in situ and invasive cervical carcinoma, to a high degree, have similar patterns of risk factors.Drs Kjaer, Engholm, and Lynge are with the Danish Cancer Registry. Dr Dahl is with the Department of Surgery, Slagelse Hospital, Denmark. Dr Bock is with the Department of Gynecology, Rigshospitalet, Copenhagen, Denmark. Dr Jensen, formerly with the Danish Cancer Registry, is deceased. Address correspondence to Dr Kjaer, Danish Cancer Registry, Institute of Cancer Epidemiology, Danish Cancer Society, Rosenvengets Hovedvej 35, Box 839, Copenhagen. Denmark. The Danish Cancer Society supported this study through grants.     

Effect of thoracic epidural etidocaine 1.5% on somatosensory evoked potentials, cortisol and glucose during cholecystectomy

The effect of thoracic (T7-8) epidural etidocaine 1.5%, 9 ml, and continuous per- and postoperative epidural infusion of etidocaine 1.5%, 4 ml/h, on early (less than 500 ms) somatosensory evoked potentials (SEPs), and cortisol and glucose in plasma during cholecystectomy, was examined in ten patients. Spread of analgesia (pin-prick) was T3 (T1-T3) to L2 (T11-L3) 35 min after injection of etidocaine, and T3 (T2-T4) to T12 (T8-L4) 3 h after surgical incision (median (range)). Before operation, epidural etidocaine had no significant effects on peak-to-peak amplitude of SEPs to electrical stimulation at the L1, T10 or T6 dermatomal level (P greater than 0.09). SEPs were abolished in only two patients at T6, and no patient had SEPs abolished at T10 or L1. The plasma concentrations of cortisol and glucose were significantly increased 20 min after surgical incision and remained increased throughout the study. No correlation was found between the block-induced decrease in the peak-to-peak amplitude at T6 or T10 and increase in plasma cortisol, except for a negative correlation at T10 and the initial increase in cortisol (Rs = 0.72, P = 0.03). In conclusion, thoracic epidural administration of 9 ml of etidocaine 1.5% does not provide total afferent somatic blockade assessed by SEP and the stress response to cholecystectomy.     

An Assay for Mannan-Binding Lectin-Associated Serine Protease 3, MASP-3

In analogy to DNA microarrays, protein microarrays offer a new distinct possibility to perform sensitive high-throughput global proteome analysis. However, the development of the protein microarray technology will place high demands upon the design of both probes and solid supports. The analysis of thousands of heterogeneous proteins on a single microarray requires the use of uniform probes, such as antibodies, directly designed for protein microarray applications. We have recently generated a human recombinant single-chain Fv antibody library, genetically constructed around one framework, the nCoDeR-library, containing 2 × 1010 clones. Single framework antibody fragments (sinFabs) selected from this library were successfully applied as probes for microarrays providing sensitive detection in the 600 attomol (mass spectrometry) and the 300 zeptomole range (fluorescence). However, the choice of framework is critical. We have shown that the selected nCoDeR framework displayed excellent functional on-chip stability and arrayed dehydrated probes retained their activity for several months. Furthermore, we have addressed the issues of biocompatibility of the solid support and immobilization strategies for our microarray setup. An in-house-designed substrate, macroporous silicon coated with nitrocellulose (MAP3-NC7), displayed properties equal to, or better than, those of five commercially available supports used as reference surfaces. We have also evaluated different coupling strategies, such as adsorption, covalent coupling, diffusion and affinity coupling. Using a novel affinity tag, the double-(his)6-tag, we increased the binding efficiency of sinFab-molecules to Ni2⁺-coated solid supports, thereby allowing nonpurified probes to be directly applied.     

Prejunctional alpha-adrenoceptor-mediated inhibition of norepinephrine release in blood-perfused skeletal muscle in situ

The influence of alpha-adrenoceptor antagonists on the overflow of endogenous norepinephrine (NE) and vasoconstrictor responses elicited by sympathetic nerve stimulation (1-4 Hz, 2 min) was investigated in desipramine-pretreated canine blood-perfused skeletal muscle in situ. The nonselective alpha-adrenoceptor antagonist phentolamine enhanced stimulation-evoked NE overflow and reduced vasoconstrictor responses concentration-dependently. Similar effects were obtained with phenoxybenzamine (irreversible alpha-adrenoceptor antagonist). Desipramine pretreatment attenuated the enhancement of stimulation-evoked NE overflow produced by phenoxybenzamine, indicating that phenoxybenzamine also inhibits neuronal uptake. The enhancement by phenoxybenzamine was independent of the stimulation frequency, suggesting a similar engagement of prejunctional alpha-adrenoceptor-mediated inhibition of transmitter release over the frequency range studied here. The alpha 2-selective adrenoceptor antagonist yohimbine enhanced nerve stimulation-evoked NE overflow at concentrations similar to those required to antagonize vasoconstrictor responses to exogenous NE; 10-fold higher concentrations were required, however, to antagonize nerve stimulation-induced vasoconstriction. The concept of a quantitatively important prejunctional alpha 2-adrenoceptor-mediated feedback inhibition of NE release in vivo is supported by our findings in the skeletal muscle vasculature. Postjunctional alpha 2-adrenoceptors appear to be preferentially activated by circulating catecholamines but also seem to be involved in the nervous control of vascular tone.     

Depression and anxiety disorders associated with headache frequency. The Nord‐Trøndelag Health Study

The aim of this large cross‐sectional population‐based study was to examine the association between migraine, non‐migrainous headache and headache frequency with depression, and anxiety disorders. From 1995 to 1997, all 92 566 inhabitants aged 20 years and above in Nord‐Trøndelag County in Norway were invited to participate in the Nord‐Trøndelag Health Study (‘Helseundersøkelsen i Nord‐Trøndelag’ = HUNT‐2). A total of 64 560 participated, whereof 51 383 subjects (80%) completed a headache questionnaire that was included. Of these 51 383 individuals, 47 257 (92%) completed the depression subscale items and 43 478 (85%), the anxiety subscale items of the Hospital Anxiety and Depression Scale (HADS). Associations were assessed in multivariate analyses, estimating prevalence odds ratios (OR) with 95% confidence intervals (CI). Depression and anxiety disorders as measured by HADS, were significantly associated with migraine (OR = 2.7, 95% CI 2.3–3.2; OR = 3.2, 95% CI 2.8–3.6) and non‐migrainous headache (OR = 2.2, 95% CI 2.0–2.5; OR = 2.7, 95% CI 2.4–3.0) when compared with headache‐free individuals. The association was stronger for anxiety disorders than for depression. The ORs for depression and anxiety disorders amongst both migraine and non‐migrainous sufferers increased with increasing headache frequency. Depression and anxiety disorders are associated with both migraine and non‐migrainous headache, and this association seems more dependent on headache frequency than diagnostic category.