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PubMed2004

Tobacco use outcomes among patients with head and neck carcinoma treated for nicotine dependence: a matched-pair analysis.

BACKGROUND: The current study described tobacco use outcomes among patients with head and neck carcinoma who underwent treatment for nicotine dependence at the Mayo Clinic Nicotine Dependence Center (NDC; Rochester, MN). METHODS: Using a 1:1 matched-pair design, conditional logistic regression was employed to compare the 6-month tobacco abstinence outcomes of patients with head and neck carcinoma (n = 101) with controls (n = 101) from the general patient population treated for nicotine dependence between 1988 and 2001. The two groups were matched with regard to age, gender, date of treatment, and type of NDC treatment service. RESULTS: Baseline demographics were similar between both groups. However, patients with head and neck carcinoma smoked significantly more cigarettes per day (cpd) than controls (P = 0.003). The self-reported tobacco abstinence rate at the 6-month follow-up was 33% for patients with head and neck carcinoma compared with 26% for matched controls (P = 0.279; after adjusting for baseline cpd and stage of change, P = 0.205). Among patients with head and neck carcinoma, the tobacco abstinence rates were 47%, 22%, and 19%, respectively, for those receiving an NDC consult within 3 months, between 3 months and 5 years, and > 5 years after their diagnosis (P = 0.021). Furthermore, the patients with head and neck carcinoma treated within 3 months of diagnosis who received surgery (with or without radiation therapy) were more likely to be tobacco abstinent than those who received primary radiation therapy (P = 0.042). CONCLUSIONS: These findings suggested that nicotine dependence treatments were effective among patients with head and neck carcinoma, particularly when delivered shortly after initial diagnosis and for those who received surgery as their primary treatment.

Head and Neck Neoplasms↗
PubMed2005

Structure, protonation state and dynamics of catalase compound II.

Density functional calculations are performed to investigate the protonation state of the compound II intermediate (Cpd II) of the catalase reaction cycle. Several scenarios are considered, depending on the protonation state of the active center (heme) and the catalytic His residue. Only the form with a protonated Fe==O unit (i.e. Fe--OH) is in agreement with the recent high-resolution crystal structure, while the traditional description of Cpd II as an oxoferryl species corresponds to a configuration slightly higher in energy. The computed Fe--O stretch frequency is in agreement with the available experimental data. Molecular dynamics simulations show that the pocket water remains in the region between the His61 and Asn133 catalytic residues, but it occasionally tries to escape towards the main channel in a concerted motion with the Asn133 residue. A possible role for this residue in the process of ligand entry/escape from the binding pocket is proposed.

Catalase↗
PubMed2002

A simple and rapid scanning electron microscope preparative technique for delicate "gymnodinioid" dinoflagellates.

Light microscopy (LM) is routinely used to investigate delicate (unarmoured and lightly armoured) "gymnodinioid" dinoflagellate species but at this level of resolution, morphological features such as apical grooves, apical pores, thin thecal plates, and scales are often difficult to observe, thereby necessitating the use of scanning electron microscopy (SEM). Good results were obtained when harvested cells were fixed with osmium tetroxide (OsO(4)) as the primary fixative, adhered with poly-L-lysine to round glass coverslips, dehydrated in an ethanol series, and dried with hexamethyldisilazane (HMDS). Poly-L-lysine has in the past effectively been used to adhere biological material such as human red blood cells, mouse leukemic cells, and marine dinoflagellates to glass coverslips. HMDS has been used to substitute critical point drying (CPD) to dry soft insect tissues, rat hepatic endothelial cells, and the cilia of rat trachea. By combining and fine-tuning these two protocols in SEM studies of delicate "gymnodinioid" dinoflagellates, it is possible to overcome cell distortion such as shrinking and collapsing that result from centrifuging, filtering, and CPD. The combination of poly-L-lysine and HMDS not only produces good results but also requires limited expertise and equipment, is inexpensive, and is less time-consuming.

Animals↗
PubMed1990

Scanning electron microscopic study of immunogold-labeled human leukocytes.

For many years critical point drying (CPD) has been the method of choice for preparing cells for scanning electron microscopy (SEM). Described herein is a simple, efficient, inexpensive, reproducible, and safe procedure using Peldri II, a proprietary fluorocarbon compound that is solid at room temperature and a liquid above 25 degrees C, as a sublimation dehydrant for processing specimens for SEM. The utility of Peldri II was demonstrated in studies using leukocytes from the blood of healthy donors and patients with leukemia as well as from long-term lymphoblastoid cell lines. The application of the proposed Peldri II procedure was further documented in SEM studies in which the expression and distribution of the interleukin-2 receptor (IL-2R) on leukocyte surface membranes was imaged using colloidal gold-labeled antibodies (i.e., immunogold). When compared with current SEM preparation procedures using CPD, Peldri II is a useful alternative that is thought to offer several important advantages.

Cell Line↗
PubMed1991

Scanning electron microscopy of piliated Neisseria gonorrhoeae processed with hexamethyldisilazane.

Piliated Neisseria gonorrhoeae are virulent and attach readily to some human mucosal cells. The study of interactions between piliated Neisseria gonorrhoeae and surface structures of eukaryotic cells in tissue culture requires consistent high resolution imaging in scanning electron microscopy (SEM). The combination of the fixatives glutaraldehyde, osmium, tannic acid, and uranyl acetate improves preservation of pili and other delicate structures. Following the critical point drying (CPD) process, pili bundles remained intact, but charging produced image distortion in most of the specimens. The use of hexamethyldisilazane (HMDS) with air drying substantially reduced charging and image distortion. Less contrast and greater resolution of pili bundles and surface structures of bacteria or tissue culture cells were obtained at magnifications of 10,000 or higher. As an alternative to CPD, HMDS processing of cell culture monolayers was simple and was more efficient when a large number of samples was processed.

Fimbriae, Bacterial↗
PubMed1993

Ex vivo clonotype primer-directed gene amplification to identify malignant T cell repertoires.

A novel strategy that utilizes input genomic DNA and overcomes limitations encountered with traditional RNA reverse transcription-polymerase chain reaction (PCR) amplification methodology is described to screen for T cell receptor (TCR) repertoires. The methodology has been developed to identify individual T cell clonotypes with regard to their unique receptor beta chain variable/diversity/joining (VDJ) region gene rearrangement. The technique avoids preselection for a given antigen specificity and is therefore independent of artificial bias introduced by in vitro cell population expansion. This technique was used to detect and identify genetically of malignant clones from heterogeneous mononuclear cell populations from an array of hemato-oncological disorders, including mycosis fungoides/Sézary Syndrome, adult T cell leukemia, and large granular lymphoproliferative disease. An initial primary PCR, directed by a TCR-J beta generic primer and a complement of family-specific TCR-V beta primers, defines predominant T cell receptor variable gene usage. Use of a TCR-J beta generic primer supplants the use of a constant region primer anchor and thus eliminates the need to target mRNA. The process of variable gene screening also expedites gene sequencing. By sequencing through the VDJ juxtaposed region, i.e., the third complementarity determinant region, clonotype-specific primers are developed and used in a secondary clonotype primer-directed PCR (CPD-PCR) to detect, with extreme sensitivity and specificity, unique T cell clonal repertoires. Analysis of the products of the CPD-PCR permits the detection of a single malignant cell among one million polyclonal cells and supercedes the constraints of prior studies that provide a limited evaluation of family variable gene repertoire usage. This strategy may be applied in the detection of minimal residual disease, in surveillance after induction of disease-free states, and in analyzing the effectiveness of purging autologous bone marrow of malignant clones.

Base Sequence↗
PubMed2004

Can color doppler predict the uniformity of HIFU-induced prostate tissue destruction?

BACKGROUND: Tissue blood perfusion influences the results of some hyperthermia and thermotherapy procedures, but its role in the outcome of prostate cancer treatment by high-intensity focused ultrasound (HIFU) has not been evaluated yet. We evaluated preoperative prostate color Doppler as a predictor of the efficacy of HIFU treatment. METHODS: Thirty-five patients underwent pre- and post-contrast color Doppler examination of the prostate before HIFU treatment. Specific software was used to calculate, on color Doppler images, the color pixel density (CPD), and the specific flow (SF, i.e., mean velocity x CPD) in different regions of interest. Post-treatment sextant biopsies were obtained in 31 patients, 5.8 +/- 2.8 months after HIFU treatment. RESULTS: No significant correlation was found between the uniformity of HIFU-induced tissue destruction observed on control biopsies and the pre-treatment CPD/SF values in any region of interest, either before or after contrast injection. On the other hand, history of radiation therapy was significantly associated with homogeneous tissue destruction and history of hormone therapy was significantly associated with incomplete tissue destruction. CONCLUSIONS: Color Doppler cannot predict the uniformity of HIFU-induced tissue destruction. History of radiation therapy was found to be a factor of favorable prognosis and history of hormone therapy was found to be a factor of poor prognosis in our population.

Aged↗
PubMed1981

Morphological correlations with dimensional change during SEM specimen preparation.

We have previously reported details of the dimensional changes taking place during the processing of soft tissue specimens for scanning electron microscopy. Mouse embryo limbs were used for many of these measurements and the present paper deals with the associated morphological findings. Effects of fixation, dehydration and drying are considered. Freeze drying and critical point drying of glutaraldehyde and glutaraldehyde and osmium fixed samples give perfectly acceptable results for scanning electron microscopy. The best volume retention with freeze dried material is matched by the best morphological appearance of the specimen surface, except when ice crystal damage occurs due to a failure to freeze the tissue rapidly enough. For CPD tissues, perforation of the plasmalemma may occur in glutaraldehyde-only fixed tissue, this being prevented by post-osmication if the glutaraldehyde fixation is not unduly prolonged. This perforation may be due to the extraction of some plasmalemma component during dehydration or further solvent substitution on critical point drying. Solvent evaporation drying usually causes recognizable distortion due to shrinkage: this is minimal in the case of solvent evaporation drying in a nearly saturated atmosphere of the same solvent if this is a very volatile solvent. The examples of Freon 113 and diethyl ether are given here. Swelling during early stages of ethanol dehydration can be prevented by using 70% or 100% ethanol as the first step, with marginal reduction in the post CPD volume and no apparent differences in the SEM. The severe swelling causing sample disruption which can occur with GA + OsO4 fixed tissue can also be prevented by treating the sample with divalent cations, such as Ca++ or Cu++, at any stage.

Animals↗
PubMed1998

Evasion of UVC-induced apoptosis by photorepair of cyclobutane pyrimidine dimers.

Cyclobutyl pyrimidine dimer (CPD) photolyase is known to reverse pyrimidine dimers specifically under illumination with visible light. OCP13, a Medaka cell line showing a high level expression of the gene for CPD photolyase, completely reversed pyrimidine dimers induced by 20 J/m2 UVC by 1 h of photorepair. When OCP13 cells were irradiated with 20 J/m2 UVC, morphological changes such as shrinkage of cells, distorted nuclear shape, and decrease in the number of nucleoli appeared 2 to 4 h after UVC irradiation. Thereafter, the irradiated cells began to detach from the substratum, and DNA ladders were observed in the DNA extracted from detached cells. Thus, these changes in cells after UVC exposure were used to characterize the progression of UV-induced apoptosis in OCP13 cells. Although formation of DNA ladders and cell detachment were blocked by cycloheximide treatment prior to UVC exposure, the morphological changes were not. With photorepair treatment, even after the morphological changes appeared cells were still able to restore their normal morphological features and remained attached. On the other hand, the cell-cycle progression in UVC-irradiated cells was arrested even after photorepair of pyrimidine dimers. Thus, photorepair can rescue cells from UV-induced apoptosis, although DNA damage other than that of pyrimidine dimers, as well as additional non-DNA damage, possibly remained, and DNA replication was left inhibited. Among the various kinds of damage induced by UVC irradiation, the presence of pyrimidine dimers is proposed to be the major trigger for UVC-induced apoptosis.

Animals↗
PubMed1999

Characterization of a Neurospora crassa photolyase-deficient mutant generated by repeat induced point mutation of the phr gene.

We produced a photolyase-deficient mutant by repeat induced point mutation using the Neurospora crassa photolyase gene cloned previously. This mutation identified a new gene, phr, which was mapped on the right arm of linkage group I by both RFLP mapping and conventional mapping. To investigate the relationship between photoreactivation and dark repair processes, especially excision repair, double mutants of phr with representative repair-defective mutants of different types were constructed and tested for UV sensitivity and photoreactivation. The results show that the phr mutation has no influence on dark repair. Tests with CPD and TC(6-4) photoproduct-specific antibodies demonstrated that the phr mutant is defective in CPD photolyase and confirmed that there is no TC(6-4) photolyase activity in N. crassa. Furthermore, N. crassa photolyase is not a blue light receptor in the signal transduction that induces carotenoid biosynthesis.

Blotting, Southern↗
PubMed1998

Spatial and temporal distribution of proopiomelanotropin and proopiocortin mRNA during the life cycle of the sea lamprey: a qualitative and quantitative in situ hybridization study.

Two POMC-like pituitary prohormones proopiocortin (POC) and proopiomelanotropin (POM) have been characterized from adult sea lampreys (Petromyzon marinus). POC encodes a nasohypophysial factor (NHF), ACTH, an MSH, and beta-END; and POM encodes MSH-A, MSH-B, and beta-END. Two radiolabeled riboprobes, one encoding a unique portion of POC mRNA and the other encoding the MSH-B domain unique to POM mRNA, were generated in order to examine the expression of POC and POM during the life cycle of the sea lamprey by in situ hybridization. POC expression appears evenly distributed throughout most cells of the rostral pars distalis (RPD) during the entire life cycle. POC expression also occurs in scattered cells of the caudal (proximal) pars distalis (CPD) at stage 5 of metamorphosis. By the prespawner period, POC expression is mainly distributed in the dorsal aspect of this region. POM expression was completely confined to most cells of the pars intermedia (PI) at all periods examined. Quantitative, computer-assisted, image analysis of POM expression revealed high signal densities in all larvae which decreased by early metamorphosis, steadily increased and reached high levels by late metamorphosis (stages 6 and 7), and attained even higher levels in prespawners. Volumetric analysis revealed that the net volume of POM expressing cells is at its lowest in larvae and increases during subsequent development. Analysis of signal density and volumetric measurements of POC expression revealed that POC expression in the RPD is low in larvae and steadily increases during subsequent intervals of the life cycle reaching very high levels by the prespawning period. POC expression in the CPD, first visible at stage 5, increases steadily throughout the remainder of metamorphosis and reaches the highest levels of expression in prespawning animals. These results would implicate the role of POM and POC in some developmental processes but not in the initiation of metamorphosis. The very high levels of POM and POC expression in prespawner animals suggest that the two genes may have important roles at this time in the life cycle of lampreys.

Animals↗
PubMed1997

DNA repair of UV photoproducts and mutagenesis in human mitochondrial DNA.

The induction and repair of DNA photolesions and mutations in the mitochondrial (mt) DNA of human cells in culture were analysed after cell exposure to UV-C light. The level of induction of cyclobutane pyrimidine dimers (CPD) in mitochondrial and nuclear DNA was comparable, while a higher frequency of pyrimidine (6-4) pyrimidone photoproducts (6-4 PP) was detected in mitochondrial than in nuclear DNA. Besides the known defect in CPD removal, mitochondria were shown to be deficient also in the excision of 6-4 PP. The effects of repair-defective conditions for the two major UV photolesions on mutagenesis was assessed by analysing the frequency and spectrum of spontaneous and UV-induced mutations by restriction site mutation (RSM) method in a restriction endonuclease site, NciI (5'CCCGG3') located within the coding sequence of the mitochondrial gene for tRNALeu. The spontaneous mutation frequency and spectrum at the NciI site of mitochondrial DNA was very similar to the RSM background mutation frequency (approximately 10(-5)) and type (predominantly GC>AT transitions at G1 of the NciI site). Conversely, an approximately tenfold increase over background mutation frequency was recorded after cell exposure to 20 J/m2. In this case, the majority of mutations were C>T transitions preferentially located on the non-transcribed DNA strand at C1 and C2 of the NciI site. This mutation spectrum is expected by UV mutagenesis. This is the first evidence of induction of mutations in mitochondrial DNA by treatment of human cells with a carcinogen.

Base Sequence↗
PubMed1985

Spatial contrast sensitivity in macular disorder.

Spatial contrast sensitivity functions were evaluated in 30 patients with macular disorders and 16 age-matched control subjects. Eighteen of the affected eyes had macular holes and the rest had isolated macular problems. A loss in spatial contrast sensitivity in the central visual field indicated a relative scotoma resulting from macular dysfunctions. The nature and size of a localized scotoma such as the type caused by a macular lesion was estimated by measuring spatial contrast sensitivity as a function of retinal area centered on the fovea. Such spatial-contrast-threshold perimetry was found to be helpful in differential classification of macular holes. Spatial contrast sensitivity and visual resolution improved with increasing retinal area even in the eyes with macular holes, but never reached normal sensitivity for spatial frequencies above 5 cycles per degree (cpd) within our experimental conditions. The loss in sensitivity to low spatial frequencies (less than or equal to 5 cpd) was found to be apparent and a function of the retinal area stimulated. The summation curves (contrast sensitivity vs. stimulus size) were abnormal for all spatial frequencies. The 'critical' sizes (i.e. the stimulus size above which no increase in sensitivity occurred) were always larger in the affected eyes than those in the normal. Thus 'critical' areas of complete summation in the parafovea may be a better measure of functional integrity of central retina in the presence of a foveal lesion than the visual resolution that varies with the retinal area and location activated. Evaluation of contrast sensitivity of the parafoveal regions with nonfunctional fovea also indicated existence of similar mechanism of spatial vision in the fovea and the parafovea.

Aged↗
PubMed1982

The histochemical localization of steroid binding sites in the pituitary gland of a teleost (the platyfish).

Specific binding sites for estrogen, testosterone, and progesterone have been demonstrated in the pituitary gland of mature male and female platyfish (Xiphophorus maculatus). With a histochemical procedure, fluorescent-steroid-hormone conjugates were localized in the cytoplasm and nucleus of the gonadotrops of the caudal pars distalis (CPD) and in cells of the pars intermedia (PI) previously demonstrated to contain immunoreactive gonadotropin. The specificity of the response was confirmed by means of competitive binding analyses and by using fluoresceinated BSA not linked to steroids. The physiological significance of steroid binding in the PI, as well as in the CPD, is discussed in the light of other recent studies on the pituitary gland of the platyfish.

Animals↗
PubMed1984

The effect of contrast and spatial frequency on the visual evoked potential of the hooded rat.

Transient and steady-state visual evoked potentials, obtained with counterphase modulated grating patterns, were recorded through chronically implanted electrodes in lightly anesthetized and unanesthetized hooded rats. As a function of spatial frequency, the latency of the major positive wave (P1) of the transient VEP was minimal with a 0.1 cycles per degree (cpd) grating as stimulus. The amplitude of the P1-N2 component remained constant from 0.04 to 0.20 cpd and then decreased with higher spatial frequencies. The function relating amplitude of steady-state VEP to spatial frequency at high contrast depended on temporal frequency: It had a band-pass form at 5.0 Hz and a low-pass form at 8.4 Hz. As a function of contrast, the latency of the P1 wave linearly decreased and its amplitude increased with increments of contrast up to 55%. The contrast function of the amplitude of the second harmonic of the steady-state VEP (8.4 Hz) showed two linear segments: one for low contrasts with a shallow slope and one for high contrasts with a steep slope, suggesting two distinct contrast mechanisms.

Animals↗
PubMed1986

Hemodynamic oxygen transport and 2,3-diphosphoglycerate changes after transfusion of patients in acute respiratory failure.

The goals of management of patients with respiratory failure include improving arterial oxygenation with PEEP and red cell transfusion to maintain oxygen carrying capacity, both of which contribute to improving tissue oxygen delivery. However, standard CPD-stored blood is rapidly depleted of 2,3 diphosphoglycerate (2,3 DPG) and ATP, with resultant inadequacy of the red cell oxygen transport function. In 15 patients requiring mechanical ventilation with PEEP whose initial Hct less than or equal to 35%, we studied the effect of transfusion of 7 ml/kg of CPD-stored packed red blood cells on hemodynamic and oxygen delivery variables, pulmonary venous admixture (QA/QT), and erythrocytic P50, 2,3 DPG and ATP concentrations. Hemodynamics were not significantly altered by transfusion. 2,3 DPG decreased significantly from 14.5 +/- 1.1 to 13.1 +/- 1.5 mcmol/g Hb (mean +/- SD, p less than 0.05). There was no significant change in P50 or ATP. QA/QT rose significantly, from 20.1 +/- 7.8 to 28.9 +/- 12.3% (mean +/- SD, p less than 0.02). In our patients, an increase in arterial oxygen content obtained by transfusion was not followed by any associated decrease in cardiac work, as implied by solution of equations for oxygen delivery and oxygen consumption. The rise in QA/QT is undesirable in patients requiring PEEP, since it complicates management of their mechanical ventilatory support.

2,3-Diphosphoglycerate↗
PubMed1980

Treatment with lithium salts reduces ethanol dehydration shrinkage of glutaraldehyde fixed tissue.

Changes in the area of glutaraldehyde fixed 15 day p.c. mouse embryo limbs were recorded using a Quantimet 720 image analysing computer attached to a light microscope: during a period of treatment with an isotonic salt solution (mostly halides of the alkali or alkaline earth metals); a subsequent wash with distilled water; and dehydration through a 30, 50, 70, 80, 90, and 100% ethanol series. Pretreatment with NaCl, KCl, RbCl had no significant effect. Treatment with LiCl, LiNO3, LiF (0.03 M), CsF and CsCl caused an increase (relative to Na, K or Rb treated samples) in the specimen volume during dehydration, which persisted in 100% ethanol. Li treated samples showed the largest post-critical-point-drying (CPD) volumes, followed by Cs treated tissue. Pretreatment with Be, Mg, Ca, Sr and Ba chlorides caused shrinkage and the 100% ethanol and post-CPD volumes of these samples were all lower than those treated with the monovalent cation containing salts.

Aldehydes↗
PubMed1993

Hypogammaglobulinemia and fatal sepsis in an infant maintained on peritoneal dialysis.

Chronic peritoneal dialysis (CPD) is a common form of renal replacement therapy in children. Recent studies suggest that immunological abnormalities, in particular hypogammaglobulinemia, may develop in children and infants on peritoneal dialysis. We report an infant maintained on CPD who died of gram-negative sepsis. At post-mortem examination, he was noted to have severe panhypogammaglobulinemia.

Agammaglobulinemia↗