bluuebunny/crossref_metadata_2025 huggingface dataset:
Development of a stereological method to measure levels of fluoropyrimidine metabolizing enzymes in tumor sections using laser scanning cytometry
Authors: Megyeri Attila , Bacsó Zsolt , Shields Anthony , Eliason James F.
Published: February 2005

Background

The enzymes thymidine phosphorylase (TP) and dihydropyrimidine dehydrogenase (DPD) influence the activities of fluoropyrimidine anticancer drugs. The sensitivity of cancer cells to capecitabine, which is an oral, tumor‐selective pre‐prodrug of 5‐fluorouracil may correlate better to the TP/DPD ratio than to levels of either enzyme alone. Our goal was to develop a quantitative immunofluorescent method for estimating the levels of TP, DPD, and their ratio in archival tumor sections.

Methods

Mouse anti‐TP and rat anti‐DPD monoclonal antibodies were used for parallel indirect immunofluorescent staining. The fluorescence was measured using a laser scanning cytometer (LSC; CompuCyte, Cambridge, MA) in single cells and in sections prepared from cell lines and a human tumor. The phantom contouring feature of the LSC provided a stereologic approach for collecting the fluorescence intensity data from sections.

Results

The relative fluorescence intensities measured in single cells or in sections of the cell lines, using single or double labeling, were similar, supporting the suitability of phantom contouring and two‐color staining. Sections of the T‐24 and ZR‐75‐1 cell lines placed on the same slide as the tumor section were used as internal standards for fluorescence measurements. The TP/DPD ratios measured in three cell lines correlated well with the cytotoxicity of 5′‐deoxy‐5‐fluorouridine measured in vitro, indicating that the measurements are related to the biological activity of the drug.

Conclusions

Plotting the data as contour maps of the topologic distribution of fluorescence intensities in tumor sections allows subsequent histopathologic examination, which may reveal features of the tumors leading to high or low ratios of these enzymes. In addition, this method can be used for any drug target/metabolic system where the key components are known and suitable antibodies are available. © 2005 Wiley‐Liss, Inc.
Cyclobutanone Approach to the Synthesis of Cardenolides
Authors: Błaszczyk Krzysztof , Koenig Hanna , Paryzek Zdzisław
Published: February 2005

17β‐(3‐Oxocyclobutyl)androstane, prepared by the thermal [2 + 2] cycloaddition of dichloroketene to 3β‐acetoxypregna‐5,20‐diene, is the key intermediate in the new, efficient synthesis of steroids bearing the 17β‐butenolide fragment that characterizes cardenolides. The six‐step synthesis of 3β‐tert‐butyldimethylsilyloxy‐14α‐carda‐5,20(22)‐dienolide was achieved with a total yield of 32%. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
Remote Participation‐Assisted Synthesis of β‐Mannosides
Authors: De Meo Cristina , Kamat Medha N. , Demchenko Alexei V.
Published: February 2005

The stereoselectivity of β‐mannosylation can be improved with the use of a participating moiety at C‐4 (O‐anisoyl, O‐thiocarbamoyl). This improvement was achieved in glycosidations of S‐ethyl and, especially, S‐benzoxazolyl (SBox) mannosides. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
Reactions of gem‐Aryl‐Disubstituted Methylenecyclopropanes with Diaryl Diselenide in the Presence of Iodosobenzene Diacetate
Authors: Shi Min , Wang Bao‐Yu , Li Jia
Published: February 2005

The reaction of gem‐aryl‐disubstituted methylenecyclopropanes 1 with diaryl diselenide produced the corresponding ring‐opened products 1,2‐bis(arylselanyl)‐3,3‐diarylcyclobut‐1‐ene 4 in the presence of iodosobenzene diacetate in moderate‐to‐good yields under mild conditions. The further transformation of 4 in the presence of MCPBA has also been examined. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
Phosphonium Acesulfamate Based Ionic Liquids
Authors: Pernak Juliusz , Stefaniak Filip , Węglewski Józef
Published: February 2005

We present a new class of solvents containing nonfluorous, well‐established, nontoxic organoanions that can be used as “green” alternatives to volatile organic solvents in a variety of applications such as solvent extraction, antielectrostatic agents, and as electrolytes in solar‐cell applications. The acesulfamate anion is an effective substitute for very expensive bis(trifluoromethylsulfonyl)imide (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
Practical P‐Chiral Phosphane Ligand for Rh‐Catalyzed Asymmetric Hydrogenation
Authors: Liu Duan , Zhang Xumu
Published: February 2005

A highly electron‐donating and conformationally rigid P‐chiral bis(trialkylphospholane) ligand 2 (DuanPhos) has been prepared in both enantiomeric forms through a concise syn‐thesis. Rh‐2 complex has exhibited remarkably high enantio‐selectivities (up to >99% ee) and reactivities (up to 10,000 TON) for the hydrogenation of a wide variety of functionalized prochiral alkenes (5 different types), which provides a very practical catalytic system for the preparation of various synthetically useful chiral compounds. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
A substantial proportion of microsatellite‐unstable colon tumors carry TP53 mutations while not showing chromosomal instability
Authors: Westra Jantine L. , Boven Ludolf G. , van der Vlies Pieter , Faber Hendrika , Sikkema Birgit , Schaapveld Michael , Dijkhuizen Trijnie , Hollema Harry , Buys Charles H. C. M. , Plukker John T. M. , Kok Klaas , Hofstra Robert M. W.
Published: February 2005

Chromosomal instability in colon tumors implies the presence of numerical and structural chromosome aberrations and is further characterized by the absence of microsatellite instability and the occurrence of KRAS and/or TP53 mutations. In a previous screening of 194 colon tumors for both microsatellite instability and TP53 mutation, we found 25 microsatellite‐unstable tumors, in 9 (36%) of which, presumed to be chromosomally stable, there were TP53 mutations. This prompted us to investigate whether a TP53 mutation in these microsatellite‐unstable tumors would be an indicator of chromosomal instability, that is, whether this would be a category of tumors showing both microsatellite and chromosomal instability. For chromosomal instability assessment, we performed array‐comparative genomic hybridization analysis of tumor and control DNA extracted from formalin‐fixed, paraffin‐embedded stage III colon tumor specimens. The array consisted of 435 subtelomere‐specific BACs. We compared all but one (whose DNA was of bad quality) of the microsatellite‐unstable TP53 mutation‐containing tumors (8) with a similarly sized group of microsatellite‐unstable tumors without TP53 mutation (11). Microsatellite‐unstable tumors with a TP53 mutation showed on average 0.9 aberrations (range 0–3) when assessed with this array system. Those without a TP53 mutation showed on average 0.7 aberrations (range 0–2). Thus, microsatellite‐unstable tumors showed few chromosomal abnormalities regardless of TP53 mutation status. Because, in our study, the microsatellite‐stable tumors had on average 3.4chromosomal abnormalities (range 0–7), a clear difference exists between microsatellite‐unstable and ‐stable tumors. Because a substantial proportion of microsatellite‐unstable colon tumors carry a TP53 mutation while showing relativelyfewchromosomal aberrations, a TP53 mutation in these tumors cannot be considered to be an indicator of chromosomal instability. © 2005 Wiley‐Liss, Inc.
Evolutionary‐based grouping of haplotypes in association analysis
Authors: Tzeng Jung‐Ying
Published: February 2005

Haplotypes incorporate more information about the underlying polymorphisms than do genotypes for individual SNPs, and are considered as a more informative format of data in association analysis. To model haplotypes requires high degrees of freedom, which could decrease power and limit a model's capacity to incorporate other complex effects, such as gene‐gene interactions. Even within haplotype blocks, high degrees of freedom are still a concern unless one chooses to discard rare haplotypes. To increase the efficiency and power of haplotype analysis, we adapt the evolutionary concepts of cladistic analyses and propose a grouping algorithm to cluster rare haplotypes to the corresponding ancestral haplotypes. The algorithm determines the cluster bases by preserving common haplotypes using a criterion built on the Shannon information content. Each haplotype is then assigned to its appropriate clusters probabilistically according to the cladistic relationship. Through this algorithm, we perform association analysis based on groups of haplotypes. Simulation results indicate power increases for performing tests on the haplotype clusters when compared to tests using original haplotypes or the truncated haplotype distribution. Genet. Epidemiol. © 2005 Wiley‐Liss, Inc.
Family history of cancer and risk of second malignancies in young cancer patients in Trieste, Italy
Authors: Brunetti Davide , Tamaro Paolo , Fanin Renato , Cavallieri Furio , Stanta Giorgio
Published: February 2005

We carried out a cohort study in the Italian province of Trieste (2001 population, 242,000) to ascertain whether the risk of a subsequent primary cancer among 265 individuals diagnosed with a first malignancy at ages up to 25 years between 1971 and 1993 differed from that in the general population and to evaluate the effect of cancer family history, quantified by the family risk index (FRI), on the occurrence of second primaries. During the follow‐up (median duration = 10 years; 25th–75th percentile = 2–16), 15 cohort members developed a second cancervs. 1.60 expected for a standardized incidence ratio (SIR) of 9.4 (p< 0.0001). The overall SIR fell to 4.7 (p= 0.004) after excluding the 8 patients with well‐known cancer‐predisposing conditions (SIR = 300.0;p< 0.0001) and the 50 with a positive family history (FRI ≥ 1.0) of malignant tumors (SIR = 20.0;p< 0.0001). Among 114 patients treated with radiotherapy and chemotherapy for their first neoplasms and not affected by predisposing disorders, 23 with a positive family history of cancer showed a 6.4‐fold excess risk of second primaries (p= 0.008) compared with 91 with a negative history (FRI < 1.0). It is imperative that clinicians carefully and regularly evaluate cancer family history of young patients with malignancies. This would enable them to identify possible individual and familial features in patients at higher risk of multiple primaries and to adopt more suitable preventive and therapeutic measures. © 2005 Wiley‐Liss, Inc.
Real‐time RT‐PCR detection of CK19, CK7 and MUC1 mRNA for diagnosis of lymph node micrometastases in non small cell lung carcinoma
Authors: Saintigny Pierre , Coulon Séverine , Kambouchner Marianne , Ricci Sylvie , Martinot Emmanuel , Danel Claire , Breau Jean‐Luc , Bernaudin Jean‐François
Published: February 2005

Metastatic lymph nodes (LNs) are the major prognostic factor in resected non small cell lung carcinoma (NSCLC). However, almost 50% of pN0 patients relapse, suggesting metastatic cells undetected by current staging procedures. A combination of markers [cytokeratins 19 and 7 (CK19, CK7) and mucin type 1 (MUC1) mRNAs] was therefore evaluated by real‐time RT‐PCR in order to detect occult cancer cells. Forty‐three NSCLC tumor samples, 4 micrometastatic, 6 metastatic and 84 histologically negative mediastinal LNs from 19 patients with NSCLC were evaluated as well as blood mononuclear cells from 29 healthy volunteers and 17 benign LNs. When tested on cell lines, RT‐PCR was particularly efficient for evaluation of CK19, CK7 and MUC1 mRNA expression. All tumor samples were positive for at least 1 marker and 74% of samples were positive for all 3 markers. CK7 and CK19 mRNA were not detected in benign LN and blood cells from healthy donors in contrast with MUC1 mRNA. Only CK7 and CK19 mRNA were therefore used for evaluation of mediastinal LNs: the 6 histologically metastatic and the 4 micrometastatic LNs were positive for at least one marker. Among the 84 histologically negative LNs, 6 (7%) were positive for at least one marker, potentially changing the stage of 2 out of 19 patients. In conclusion, in our feasibility study, parallel molecular detection of CK19 and CK7 mRNA can be considered a specific diagnostic tool for the assessment of microscopic lymphatic spread. Its prognostic impact remains to be evaluated in a prospective study. © 2005 Wiley‐Liss, Inc.
Metalloproteinases and their inhibitors in tumor angiogenesis
Authors: Handsley Madeleine M. , Edwards Dylan R.
Published: February 2005

Angiogenesis is the process by which new blood vessels are formed from preexisting vasculature. It is an essential feature of the female reproductive cycle, embryonic development and wound repair. Angiogenesis has also been identified as a causal or contributing factor in several pathologies, including cancer, where it is a rate‐limiting step during tumor progression. Matrix metalloproteinases (MMPs) are a family of soluble and membrane‐anchored proteolytic enzymes that can degrade components of the extracellular matrix (ECM) as well as a growing number of modulators of cell function. Several of the MMPs, in particular the gelatinases and membrane‐type 1 MMP (MT1‐MMP), have been linked to angiogenesis. Potential roles for these proteases during the angiogenic process include degradation of the basement membrane and perivascular ECM components, unmasking of cryptic biologically relevant sites in ECM components, modulation of angiogenic factors and production of endogenous angiogenic inhibitors. This review brings together what is currently known about the functions of the MMPs and the closely related ADAM (a
disintegrin and metalloproteinase domain) and ADAMTS (a
disintegrin and metalloproteinase with thrombospondin motifs) families in angiogenesis and considers how this information might be useful in manipulation of the angiogenic process, with a view to constraining tumor progression. © 2005 Wiley‐Liss, Inc.
Nasopharyngeal carcinoma–susceptibility locus is localized to a 132 kb segment containing HLA‐A using high‐resolution microsatellite mapping
Authors: Lu Cheng‐Chan , Chen Jung‐Chin , Tsai Sen‐Tien , Jin Ying‐Tai , Tsai Jui‐Chen , Chan Shih‐Huang , Su Ih‐Jen
Published: February 2005

Nasopharyngeal carcinoma (NPC) is an epithelial tumor uniquely prevalent in southern Chinese. HLA‐A2 is associated with NPC. In a previous study, we showed that the genes associated with susceptibility to NPC are primarily located within the HLA‐A locus in Taiwanese NPC patients. However, the pathogenic genes causing NPC susceptibility remain unknown. Here, 8 polymorphic microsatellite markers distributed over a 1 megabase region surrounding the HLA‐A locus were subjected to genetic analysis for the NPC‐susceptibility locus. Statistical studies of associated alleles detected on each microsatellite locus showed that the NPC‐ susceptibility genes are most likely located between the D6S510 and D6S211 markers within a 132 kb segment containing the HLA‐A locus. These results undoubtedly would facilitate the further positional cloning of the NPC‐susceptibility locus, which has been elusive for the past 30 years. © 2005 Wiley‐Liss, Inc.
Na+/H+ exchanger activity is increased in doxorubicin‐resistant human colon cancer cells and its modulation modifies the sensitivity of the cells to doxorubicin
Authors: Miraglia Erica , Viarisio Daniele , Riganti Chiara , Costamagna Costanzo , Ghigo Dario , Bosia Amalia
Published: February 2005

Multidrug resistant (MDR) tumor cells exhibit an altered pH gradient across different cell compartments, which favors a reduced intracellular accumulation of antineoplastic drugs and a decreased therapeutic effect. In our study, we have observed that the activity and expression of Na+/H+ exchanger (NHE), which is involved in the homeostasis of intracellular pH (pHi), are increased in doxorubicin‐resistant (HT29‐dx) human colon carcinoma cells in comparison with doxorubicin‐sensitive HT29 cells. The pHi was significantly higher in HT29‐dx cells, which accumulated less doxorubicin than HT29 cells. The NHE inhibitor 5‐(N‐ethyl‐N‐isopropyl)amiloride (EIPA) significantly reduced the pHi value and increased the intracellular accumulation of doxorubicin in both cell populations: in the presence of EIPA HT29‐dx cells accumulated as much drug as control HT29 cells. On the other hand, monensin, a Na+/H+ ionophore mimicking NHE activation, and phorbol 12‐myristate 13‐acetate (PMA), which stimulates NHE, significantly increased the pHi and decreased the drug accumulation in HT29 cells to values similar to those observed in control HT29‐dx cells. EIPA potentiated the cytotoxic effect of doxorubicin in HT29 cells, and made HT29‐dx cells as sensitive to the cytotoxic effect of the drug as control HT29 cells. Instead, PMA and monensin made HT29 cells as insensitive to doxorubicin as HT29‐dx cells. These results suggest that in MDR cells the higher cytosolic pH is likely to decrease drug accumulation, and that such resistance can be reverted by inhibiting the NHE activity. This result opens the possibility to revert MDR with the clinical use of NHE inhibitors. © 2005 Wiley‐Liss, Inc.
Retinoid‐related molecule AGN193198 potently induces G2M arrest and apoptosis in bladder cancer cells
Authors: Reitmair Armin , Shurland Dixie‐Lee , Tsang Kwok‐Yin , Chandraratna Roshantha A.S. , Brown Geoffrey
Published: February 2005

The novel synthetic retinoid‐related molecule 4‐[3‐(1‐heptyl‐4,4‐dimethyl‐2‐oxo‐1,2,3,4‐tetrahydroquinolin‐6‐yl)‐3‐oxo‐propenyl]benzoic acid (AGN193198) neither binds effectively to retinoic acid receptors (RARs) and retinoid X receptors (RXRs) nor transactivates in RAR‐ and RXR‐mediated reporter assays. Even so, AGN193198 is potent in inducing apoptosis in human prostate and breast carcinoma cells (Keedwell et al., Cancer Res 2004;64:3302–12). Here, we extend these findings to show that AGN193198 potently and rapidly induces apoptosis in bladder carcinoma cell lines. One micromolar of AGN193198 completely abolished the growth of the transitional cell carcinoma lines UM‐UC‐3 and J82, and the squamous cell carcinoma line SCaBER; the transitional cell papilloma line RT‐4 was slightly less sensitive to the growth inhibitory effect of AGN193198. Treated cells accumulated in the G2M phase of the cell cycle. This was accompanied by apoptosis, as revealed by staining cells for exposure of phosphatidylserine at their surface (binding of Annexin V) and FACS analysis of propidium iodide labeled cells. As reported for prostate cancer cells, AGN193198 provoked rapid activation of caspases‐3 (by 6 hr), ‐8 (by 16 hr) and ‐9 (by 6 hr) in bladder cancer cells. These findings suggest that AGN193198 and related compounds, whose mechanism of action does not appear to involve RARs and RXRs, may be useful in the treatment of bladder cancer. © 2005 Wiley‐Liss, Inc.
Three innovations in cost accounting
Authors: Vercio Alan , Williams Brant
Published: February 2005

Leading‐edge cost measurement does not necessarily mean the latest “alphabet soup” solution or yet another vendor's software solution. The foundation for useful and relevant cost measurement has a long and time‐tested history. © 2005 Wiley Periodicals, Inc.
The behavior of transition metal nitrido bonds towards protonation rationalized by means of localized bonding schemes and their weights
Authors: Bachler Vinzenz
Published: February 2005

A new computational scheme is applied to rationalize the different protonation behaviors of the nitrido complexes [L′MnV(N)(acac)]+, [LCrV(N)(acac)]+, and [LVV(N)(acac)]+. L and L′ represent the macrocycles 1,4,7‐triazacyclononane and its N‐methylated derivative, respectively, and acac is the bidentate monoanion pentane‐2,4‐dionate. The bonds of the complexes are partitioned into bonds to be investigated and bonds of lesser interest. The investigated bonds are the transition metal nitrido bonds MVN| (M = Mn, Cr, and V) and the bonds of lesser interest are located in the ligands. The ligand bonds are described by means of the strongly occupied natural bond orbitals. The electrons in the MVN| nitrido bonds, however, are treated more accurately. A full configuration interaction procedure is applied in the space spanned by the strongly occupied natural bond orbitals and their corresponding antibonding orbitals. Localized bonding schemes and their weights are obtained for the dπ‐pπ bonds of interest. This is achieved by representing the two‐center natural bond orbitals for a dπ‐pπ bond by the one‐center natural hybrid orbitals localized at the bond atoms. The obtained bonding schemes are close to orthogonal valence bond structures. Their weights indicate that the nitrido nitrogen in [LVV(N)(acac)]+ is more easily protonated than the nitrido nitrogens in [L′MnV(N)(acac)]+ and [LCrV(N)(acac)]+. This result is in good accord with experiment. © 2005 Wiley Periodicals, Inc. J Comput Chem 26: 532–551, 2005
Polymorphism of a long‐chain cycloparaffin (CH2)120
Authors: Ihn Kyo Jin , Tsuji Masaki , Kawaguchi Akiyoshi , Katayama Ken‐Ichi
Published: February 2005

The polymorphism of a long‐chain cycloparaffin (CH2)120 and chain packing in its crystals were discussed on the basis of some results obtained mainly by transmission electron microscopy. Monoclinic and orthorhombic single crystals of (CH2)120 were isothermally grown together from a dilute solution in p‐xylene. Lozenge‐shaped orthorhombic single crystals were more frequently observed than lath‐shaped monoclinic ones. The basal surfaces of orthorhombic and monoclinic single crystal platelets wore decorated with vapor‐deposited polyethylene [PE]. Orthorhombic single crystals of (CH2)120 with the (110) twin boundary and monoclinic ones with the (100) twin boundary were also observed.

Rod‐like edge‐on crystals of (CH2)120 were grown from a dilute p‐xylene solution onto the (001) surface of alkali halides. The crystal system of the (CH2)120 edge‐on crystals depended on the kind of substrate. The monoclinic crystal was grown on NaCl, the orthorhombic one on KBr and KC1. The monoclinic form of (CH2)120 edge‐on crystal was transformed to the orthorhombic one by annealing on NaCl. In both monoclinic and orthorhombic edge‐on crystals, the molecular plane determined by two zigzag stems in a molecule of (CH2)120 was parallel to the substrate surface and the molecular axis (crystallographic c‐axis) was perpendicular to the longer side of the rod‐like edge‐on crystals. The sub‐cell dimensions of the stem chains in both forms of (CH2)120 crystals were very similar to those of monoclinic and crthorhombic PE crystals, respectively.
Immunoelectron microscopical distribution of histones H2B and H3 and protamines in the course of mouse spermiogenesis
Authors: Biggiogera Marco , Muller Sylviane , Courtens Jean Luc , Fakan Stanislav , Romanini Maria Gabriella Manfredi
Published: February 2005

We have followed the fine structural distribution of two nucleosomal core histones, H2B and H3, and of protamines in the course of mouse spermiogenesis by means of specific antibodies and ultrastructural immunocytochemistry.

Our results demonstrate that the nuclear labeling density of histone H2B decreases during steps 6–8 and then increases again in step 9–10 spermatids, while the labeling for histone H3 is constant throughout this period. In step 12 spermatids, the anti‐H2B antibody labels mainly the central area of the nucleus. The first signs of protamine labeling are present in step 12 spermatids, where the gold grains can be found over the periphery of the nucleus. Later on, protamine labeling constantly increases and, by the end of spermiogenesis, the whole nucleus is labeled.

We suggest that the morphological and structural differences between the central area and the periphery of mouse spermatids are, at least partly, due to a difference in the protein moiety associated with DNA. The central area, which is peculiar to the mouse and has been previously considered as a focus of chromatin condensation, represents, however, the last nuclear region containing histones and consequently the last area where the substitution of histones by protamines takes place.
Technique of reflection electron microscopy
Authors: Hsu Tung
Published: February 2005

Details of the technique of reflection electron microscopy (REM) are described. Step by step instruction is given on how to do it on an ordinary electron microscope. Also given are some specimen preparation techniques and strategy of REM investigation.
Observation of atomic steps on single crystal surfaces by a commercial scanning electron microscope
Authors: Uchida Yuji , Weinberg Gisela , Lehmpfuhl Gunter
Published: February 2005

Atomic steps on (111) and (100) crystal surfaces of Pt were observed using a commercial scanning electron microscope (SEM) in secondary electron mode. By comparing the SEM images and those by reflection electron microscopy (REM), the observed contrast was confirmed to be that from atomic steps on crystal surfaces. The contrast mechanism is briefly discussed. One application of this imaging technique is also shown.
Observation of surfaces by reflection electron holography
Authors: Osakabe Nobuyuki
Published: February 2005

Reflection electron holography is described as a method to observe sub‐Å surface morphology. Phase shift of a Bragg‐reflected electron wave was measured by means of holographic interferometry using an electron microscope equipped with a field emission electron gun and an electron biprism. A short wavelength of high energy electrons is the essential key to the high vertical sensitivity of this method, since geometrical path differences produced by the surface topography are measured in units of wavelengths in interferometrical measuring. Phase shift at a monoatomic step and the displacement field around a dislocation emerging on the surface were observed.
Section staining for electron microscopy using tannic acid as a mordant: A simple method for visualization of glycogen and collagen
Authors: Afzelius BjöRn A.
Published: February 2005

The possibility that tannic acid and other classical mordants can be used in the electron microscopial section staining technique has been tested. A mordant can be defined as a chemical that combines with both a certain specific tissue component and the staining substance and thereby permits a staining reaction that otherwise will not be obtained. The following features were found to characterize section staining of tannic acid mordanted sections. Tannic acid apparently blocks those sites that normally would be contrasted by uranyl acetate or some other staining compounds. Ribosomes remain unstained. Glycogen particles, on the other hand, were stianed, whereas they are not in non‐mordanted sections. In fact, glycogen was the only cytoplasmic component to be contrasted by the uranyl acetate, and collagen the only extracellular component. Several different section staining solutions gave the same staining patterns of examined cells and tissues. Specificity of the reaction thus seems to depend on the mordant rather than on the heavy atom section stain. Some other tested mordants, which have also been used in the light microscopical technique, did not give any useful new information.
The ultrastructure of mammalian pinealocytes: A systematic investigation
Authors: Bhatnagar Kunwar P.
Published: February 2005

Pinealocytes are not only the principal cellular components of the pineal gland, but they are also the principal synthetic machinery of this enigmatical gland with highly diverse and often questionable empyreal roles assigned to it. Ultrastructural descriptions of pinealocytes belonging to some 70 species of mammals (a mere 2% or less of the over 4,200 mammalian species) have been summarized from the available literature with new observations on 12 species of chiropterans. Space limitation precluded any treatment of the supporting glia, neural elements, and the perivascular spaces. A detailed table lists nearly all mammalian species whose pineal ultrastructure has been investigated. Blanks in this table point to the necessity of studies on those particular groups. A tabular listing of unusual structures reported within the pinealocyte cytoplasm points out the impending experimental work on these species. Such studies using the latest techniques might provide clearer insights into the functional role of the pineal gland as an important and integral component of the neuroendocrine axis. Whereas sufficient structural information now exists on cytoplasmic organelles such as synaptic ribbons and spherules, annulate lamellae, subsurface cisterns, and the several types of synaptic arrangements seen in relation to the pinealocyte soma and its processes, the functional role of these structures in pineal synthetic processes remains to be elucidated.
Morphofunctional aspects of the mammalian pineal gland
Authors: Karasek Michal , Reiter Russel J.
Published: February 2005

The primary aim of this review is to present the current state of knowledge of the ultrastructure of the mammalian pineal gland, with emphasis on its functional aspects. Basic ultrastructural features of the mammalian pinealocytes are presented with special attention paid to ultrastructural aspects of pineal secretion.
The embryonic pineal body as a multipotent organ
Authors: Watanabe Kenji , Araki Masasuke , Iwasaki Hiromichi
Published: February 2005

The repertoire of differentiating potency of mammalian and avian pineal cells has been examined utilizing cell culture technique. Skeletal muscle fibers are differentiated from pineal cells of the rat under the usual culture condition and from those of quail under hypertonic conditions. Myogenesis of pineal cells may be explained from the ontogeny of the pineal body. Anlagen of a pineal body are situated in bilateral cephalic neural folds, which also supply multipotent neural crest cells. In some conditions, almost all quail pineal cells are able to differentiate into pigmented epithelial cells and/or lens cells. Opsin containing cells found in culture of rat pineal cells may be in a similar category reflecting the “third eye”: the phylogenetic ancestor of the pineal body of avian and mammalian species. Neuron‐like cells have also been reported and neuronal morphology has been intensified under the effect of testicular hyaluronidase. The cytodifferentiation described above is suggested to be different expressions of a single type of progenitor cells in the pineal body. In relation to multipotentiality of pineal cells, the original differentiating state of pineal cells is interesting; it has been found that tyrosinase is expressed from the beginning of pineal formation and that its expression is stage‐specific (during embryonic period) and site‐specific (predominance in the dorsal half of the pineal body and in the apical cytoplasm of the pineal cell). In the 8 day quail embryo used for culture studies, three differentiating states as to tyrosinase are noticed. However, the distinction may be apparent, as even the cells negative in tyrosinase in this stage are still ready to express tyrosinase in the suitable culture condition. Pineal cells are flexible and surprisingly susceptible to environmental factors and useful for the study of cell differentiation in general. © 1992 Wiley‐Liss, Inc.
The microcomputer and image analysis in diagnostic pathology
Authors: Jarvis L. R.
Published: February 2005

This paper presents a snapshot view of the influence and direction of microcomputer technology for image analysis techniques in diagnostic pathology.

Microcomputers have had considerable impact in bringing image analysis to wider application. Semi‐automated tracing techniques are a simple means of providing objective data and assist in a wide range of diagnostic problems. From the common theme of reducing subjectivity in diagnostic assessment, an extensive body of research has accrued. Some studies have addressed the need for quality control for reliable, routine application.

Video digitizer cards bring digital image analysis within the reach of laboratory budgets, providing powerful tools for investigation of a wide range of cellular and tissue features. The use of staining procedures compatible with quantitative evaluation has become equally important. As well as assisting scene segmentation, cytochemical and immunochemical staining techniques relate the data to biological processes.

With the present state of the art, practical use of microcomputer based image analysis is impaired by limitations of information extraction and specimen throughput. Recent advances in colour video imaging provide an extra dimension in the analysis of multi‐spectral stains. Improvements will also be felt with predictable increase in speed of microprocessors, and with single chip devices which deliver video rate processing. If the full potential of this hardware is realized, high‐speed, routine analysis becomes feasible. In addition, a microcomputer imaging system can play host to companion functions, such as image archiving and transmission.

With this outlook, the use of microcomputers for image analysis in diagnostic pathology is certain to increase. However, it is the software in both design and concept which ultimately governs the performance which can be achieved. Progress may be made by structured software techniques, by application of mathematical principles, or by use of expert systems for data or image interpretation. © 1992 Wiley‐Liss, Inc.
Preservation and visualization of the sea urchin embryo blastocoelic extracellular matrix
Authors: Cherr Gary N. , Baldwin John D. , Summers Robert G. , Morrill John B.
Published: February 2005

Several methods were utilized to visualize the structure and orientation of the blastocoelic extracellular matrix (ECM) in Strongylocentrotus purpuratus embryos at the mesenchyme blastula stage. Rapid freezing in liquid propane cooled to LN2 temperatures followed by freeze substitution was used to preserve the ECM without shrinkage due to dehydration. Scanning, transmission, and light microscopy were employed to elucidate the ECMs' structure. The blastocoelic ECM consisted of parallel fibrillar sheets that were interconnected by finer filaments and oriented along the animal‐vegetal axis. The ECM completely filled the blastocoelic cavity as viewed by scanning electron microscopy. The basal lamina could be distinguished from the blastocoelic ECM as a thin coat on the plasma membrane of epithelial cells; the ECM was in contact with this coat. In contrast, the blastocoelic ECM attached directly to the plasma membrane of primary mesenchyme cells (PMC) which did not possess a basal lamina. The blastocoelic ECM was isolated as an intact “bag” and probed in a hydrated state with Con A and alcian blue. Confocal microscopy confirmed that the entire blastocoel was filled with a fibrillar ECM. These approaches offer advantages for future studies of the ECMs of sea urchin embryos and their roles in gastrulation. © 1992 Wiley‐Liss, Inc.
Ultrastructural imaging of freeze‐fractured plant cells in the scanning electron microscope
Authors: Barnes Susan H.
Published: February 2005

The application of the freeze‐fracture and cytoplasmic maceration technique in ultrastructural studies of plant cells is described. A major advantage of the technique is, that by extracting mobile cytoplasmic components from the freeze‐fractured cells, surface relief is introduced and three‐dimensional information is obtainable. The details of specimen preparation are described and the results obtained are reviewed. The use of chitosan embedding for very small or fragile specimens is described. © 1992 Wiley‐Liss, Inc.
Freeze‐substitution of chemically stabilized samples for biological field emission scanning electron microscopy
Authors: Peters Klaus‐Ruediger , Pohl Roderike
Published: February 2005

The high resolution imaging capabilities of modern field emission scanning electron microscopes require adequately improved tissue preparation procedures to prevent the collapse of macromolecular structures and the extraction of molecules. A routine cryo‐stabilization technique is described which utilizes chemical crosslinking and cryo‐dehydration for mechanical and chemical stabilization of protein and lipid structures and increase of electrical conductivity of the sample. Thiocarbohydrazide (TCH) serves as a general mordant for osmium tetroxide crosslinking. However, extensive washing after all impregnation steps is necessary to dissolve unspecific osmium black precipitations at the sample surface. Collagen I aggregates showed increased stability against collapse after TCH osmification alone, whereas pulmonary surfactant liposomes require additional freeze‐substitution in methanol and Freon 113 for stabilization during critical point drying. Environmental scanning electron microscopy (at water vapor pressures of 5–10 torr within the specimen chamber) was used to control, in the wet phase, the stabilization procedure at the level of chemical crosslinkage. It could be confirmed that tannic acid, often used to stabilize lipids, eads to artificial rearrangement of bilayered liposomes into compact presumable multilayered bodies, whereas the TCH osmification preserved liposome structures and their aggregates. The increase of electrical conductivity of sliced tissue was demonstrated on kidney. Support technologies for the cryo‐stabilization procedures are described in detail, as well as simple routines for first stabilization trials with new samples. On pulmonary tissue, the excellent preservation of alveolar shape and fine structures of intermediate forms of surfactant are described. © 1992 Wiley‐Liss, Inc.
Suspected chemoreceptors in coelenterates and ctenophores
Authors: Kass‐Simon G. , Hufnagel L. A.
Published: February 2005

Chemoreceptors in coelenterates and ctenophores have not been identified with certainty. Among prospective chemoreceptive cells are the sensory nerve cells, the cnidocyst bearing cnidocytes, and the epitheliomuscular cells that are likely to be involved in feeding or aggression. Both behaviors are mediated by coordinated chemical and mechanical reception. This is reflected in the close apposition of putative chemo‐ and mechanoreceptors. Among the structures that have been designated as likely chemo‐ and/or mechanoreceptors are stereocilia, kinocilia, and/or microvilli which are universally present on all the putative chemoreceptor complexes, while gland cells and mucous secretions are prevalent. Evidence that the actin‐containing stereocilia are chemically modulated mechanoreceptors is presented for several forms. © 1992 Wiley‐Liss, Inc.
Fine structure of olfactory sensilla in myriapods and arachnids
Authors: Tichy Harald , Barth Friedrich G.
Published: February 2005

Structural features of various types of olfactory sensilla are reviewed. (1). Sensilla basiconica which differ in form and size are found on the antennae of centipedes and millipedes. Their walls show longitudinal slits or grooves that either open into the sensillum lumen or do not penetrate the cuticle. In other such sensilla the outer surface is pierced by pores and the inner surface grooved and pocketed. These sensilla are innervated by one to six sensory cells. Their unbranched outer dendritic segments extend to the tip of the sensillum. The sensory cells are surrounded by two or three sheath cells which terminate at the sensillum base or form a continuous tube around the entire length of the outer dendritic segments. (2) Temporal organs of centipides are located between the insertion of the antenna and the ocelli. These sensilla consist of a shallow cuticular ring with a central sensory plate made up by a layer of unperforated cuticle or a capsule with a mushroom‐shaped structure inside formed by fibrous‐looking cuticle. A dozen sensory cells with unbranched outer dendritic segments innervate each sensillum. They extend toward the sensory cuticle and pass just below it. Numerous sheath cell processes run parallel to the outer dendritic segments up to the sensory cuticle. (3) Thread‐like flagella of Pauropoda are found on the antennae. They possess a flexible unperforated cuticular wall. These sensilla contain nine sensory cells surrounded by several sheath cells which form a continuous cytoplasmic tube around the outer dendritic segments. (4) Single‐walled sensilla with numerous plugged pores penetrating the cuticular wall occur on the tarsus of the first leg in ticks. Each sensillum is innervated by 4–15 sensory cells. Three sheath cells terminate in the base of the sensillum. (5) Double‐walled sensilla with spoke canals are found on the first tarsus of ticks. Their shaft is longitudinally grooved. Pore canals lead inward from the bottom of the grooves and open into vase‐shaped chambers. From its base these canals extend into the lumen of the sensillum which contains unbranched outer dendritic segments of 1–2 sensory cells. (6) Single‐walled sensilla with pore openings occur on the distal tarsal segments of the first leg of whip spiders. These sensilla are innervated by 40–45 sensory cells. Their unbranched outer dendritic segments fill the shaft lumen and extend partly into the wall pores. Microvillus‐shaped sheath cell processes line the inner surface of the cuticular wall. (7) Tarsal organs are located dorsally on the tarsus of all legs and pedipalps of spiders. These sensory organs consist of a cuticular capsule with a dome‐shaped projection inside. It is situated on the proximal sidewall of the capsule and possesses 7 pore canals that enclose the dendritic tips of 2–3 sensory cells, giving a total of 20 sensory cells. Each group of dendrites terminating in an individual pore canal is encased by 2 sheath cells. © 1992 Wiley‐Liss, Inc.
Hapten‐tagged plasma proteins as immunocytochemical probes for the study of vascular permeability
Authors: Ghitescu Lucian , Bendayan Moïse
Published: February 2005

Bovine serum albumin and transferrin were covalently coupled with fluorescein isothiocyanate and digoxigenin, respectively, and intravenously co‐injected in equal amounts in mouse. The derivation of the two proteins induces minor alterations of their physicochemical properties as well as of their physiological functions. The two tracers were revealed within vascular and extravascular compartments of diaphragm by quantitative postembedding immunocytochemistry, using antibodies against each of the haptens in conjunction with the protein AG‐gold complexes. The influence of different fixatives and embedding protocols on the immunodetectability of the hapten‐tagged proteins was assessed. Both resist reasonably well to osmication and embedding in Epon. None of the haptens reacted with the heterologous antibody. At 30 minutes after injection, the tracers were detected in blood plasma, interstitium, and endothelial plasmalemmal vesicles. The presence of both proteins within the interendothelial clefts was inconspicuous. The ratios between the labeling densities found over endothelium, interstitial space, and vascular lumen were similar for both tracers. This suggests that the endothelium of mouse diaphragm capillaries might exhibit comparable permeabilities towards serum albumin and transferrin which are similar in size and charge. The study shows that hapten‐tagged polypeptides are close to the corresponding native macromolecules, and represent interesting tools for the morphological study of dynamic processes such as transcytosis. © 1992 Wiley‐Liss, Inc.
Phylogeny of the vomeronasal system and of receptor cell types in the olfactory and vomeronasal epithelia of vertebrates
Authors: Eisthen Heather L.
Published: February 2005

In this paper, the evolutionary origin of the vomeronasal system as a discrete sensory system separate from olfaction is examined. The presence of a discrete vomeronasal system appears to be a derived character in tetrapods, and its presence in larval amphibians indicates that the system did not arise as a terrestrial adaptation. The vomeronasal system has been lost independently in several taxa, including crocodilians, some bats, cetaceans, and some primates. The presence of microvillar receptor cells in the vomeronasal epithelium appears to be the ancestral condition for tetrapods, and alternative hypotheses concerning the ancestral condition for receptor cell types in the vertebrate olfactory epithelium are discussed. Finally, the possibility that the vomeronasal system is present in some fishes in a form that has not been recognized is discussed in relation to the phylogenetic distribution of receptor cell types in vertebrates. © 1992 Wiley‐Liss, Inc.
Ciliated and microvillar receptor cells degenerate and then differentiate in the olfactory epithelium of rainbow trout following olfactory nerve section
Authors: Zielinski Barbara S. , Hara Toshiaki J.
Published: February 2005

We used scanning (SEM) and transmission (TEM) electron microscopy to examine ultrastructural changes in the olfactory epithelium (OE) of rainbow trout following unilateral olfactory nerve section. Both ciliated receptor cells (CRC) and microvillar receptor cells (MRC) degenerated and subsequently differentiated from unidentified precursor cells. The following changes took place in fish that were held at 10°C at the stated period following olfactory nerve section: on day 7, MRC and CRC contained intracellular vacuoles; on day 12, the olfactory knobs appeared disrupted; by day 26, olfactory receptor cells were absent from the OE; on day 42, there were receptor cell bodies and a few CRC with short cilia at the apical surface; and opn day 55, a small number of both CRC and MRC had differentiated. By day 76, both CRC and MRC repopulated the OE. Degenerative changes in the cytoplasm of the sustentacular cells (SC) and ciliated nonsensory cells (CNC) were observed in the first 26 days following olfactory nerve section, but these cells remained intact throughout the experiment. The degeneration and subsequent differentiation of CRC and MRC supports and extends previous observations that both cells types are olfactory receptor neurons with axons that extend along the olfactory nerve to the olfactory bulb. © 1992 Wiley‐Liss, Inc.
Morphology of olfactory epithelium in humans and other vertebrates
Authors: Morrison Edward E. , Costanzo Richard M.
Published: February 2005

Human olfactory epithelium is similar in organization and cell morphology to that of most vertebrate species. The epithelium has a pseudostratified columnar organization and consists of olfactory neurons, supporting and basal cells. Near the mucosal surface there are also microvillar cells. These cells have neuron‐like features and may be chemoreceptors. Human olfactory epithelium is not a uniform sensory sheet. Patches of non‐sensory tissue often appear in what was thought to be a purely olfactory region. The significance of these patches has not been determined, but they could reflect exposure to environment agents or changes that occur during the normal aging process.

In order to better understand the human olfactory system, further knowledge of the normal structure is necessary. This review addresses the morphology of the human olfactory epithelium and the remarkable plasticity of the vertebrate olfactory system. © 1992 Wiley‐Liss, Inc.