Biomedical subjects

F H White

Publications and source records attributed to F H White.

At least 55 records · Page 3Linked to original sources

PubMed1984

Quantitative ultrastructure of cytolytic lymphocytes mediating allograft rejection in the mouse. I. Cellular alterations in T lymphocytes during specific target cell lysis.

A quantitative ultrastructural analysis of cytolytic T lymphocytes (CTL) is presented which allows both the distinction of these cells from normal T lymphocytes and permits the demonstration of ultrastructural alterations of putative CTL following interaction with target cells (TC). Alloreactive CTL were generated in C57BL/10 mice receiving intraperitoneal fibroblastic allografts and target-binding splenic lymphocytes (TBSL) were concentrated by specific immunoadsorption on fibroblast monolayers. TBSL were subjected to ultrastructural quantification either at the onset of TC interaction or following 30 or 60 min incubation at 37 degrees C. By means of simple stereological relationships it was shown that, in comparison with normal, non-cytolytic splenic T lymphocytes, TBSL were slightly larger cells, displaying around 60% more cytoplasm, a similarly-sized nucleus and approximately triple the volume of Golgi apparatus. During the first 30 min of interaction with TC, the target binding surface of the TBSL plasma membrane decreased in area. This change was accompanied by a polarization of the TBSL towards the target. Incubation of lymphocytes with TC for a further 30 min resulted in a general polarization of lymphocytic cellular constituents away from the TC. These results were only attainable by objective quantitative analysis and are discussed in relation to possible mechanisms of CTL-mediated lysis.

Animals↗

PubMed1984

Effects of structure-forming solutes on chicken egg white lysozyme after reductive cleavage of disulfide bonds.

As an approach to learning more about structure-forming processes in chicken egg white lysozyme, the reduced protein was studied in the presence of methanol, 2-chloroethanol, trifluoroethanol, and sodium dodecyl sulfate, at various concentrations in aqueous solution. The mixtures were subjected to gel filtration chromatography, with circular dichroism (CD) examination of the resulting polymers and monomers. Changes in CD behavior, indicating development of ordered structure, were seen to be associated only with polymer formation. Contributing to these changes were increases in beta structure as well as alpha-helix. Since the solutes studied are known as helix formers, it is suggested that studies on many proteins, carried out in the presence of these and other structure-modifying solutes, may have been complicated by formation of beta structure as well as by polymerization. The reduced monomer formed no structure in the hydrophobic environment afforded by these solutes, but only randomized further. Thus, hydrophobic interactions may be of no significance to lysozyme chain folding in its early stages. As an alternative, the long range interactions stabilized by native tertiary cross links may be critical for the transition to native secondary structure.

Animals↗

PubMed1984

Evidence for interaction of substrate analogs with chicken eggwhite lysozyme after exhaustive reduction of disulfide bonds.

In a study of factors that influence the remaining secondary structure of reduced chicken eggwhite lysozyme, N-acetyl-D-glucosamine (NAG) and N,N'-diacetylchitobiose (di-NAG) were found to alter the circular dichroic (CD) spectrum of the reduced protein and its carboxymethyl derivative (Cml). Thus, negative ellipticities in the far u.v. were greater in the presence of the analogs, with NAG being the more effective. For Cml, curve fitting analysis of the CD data indicated an increased helical content in the presence of NAG by an average of 3% of the chain length, while beta-structure decreased by an equivalent amount. Other compounds structurally related to NAG produced no similar effects on the CD spectrum of Cml, nor were comparable effects of NAG in evidence on the Cm reduced derivatives of ribonuclease, chymotrypsin, wheat germ agglutinin, or alpha-lactalbumin. The effect therefore appears specific between NAG and Cml. Conversion of the tryptophan residue at Position 62 of Cml to the oxindolealanyl derivative prevented these effects of NAG, and this residue may therefore participate in the interaction. During a 4-day incubation at room temperature, the analog preserved the CD spectrum of Cml as well as its concentration. This effect was nearly specific when compared with other Cm reduced proteins and with other carbohydrates. Only one, N-acetyl mannosamine, was effective in preserving the concentration of Cml, but not the CD spectrum. Since D-glucosamine was entirely without effect on either the CD spectrum of Cml or on its change during incubation, the acetyl group appears essential for the NAG-Cml interaction. The specificity between NAG and Cml is tentatively accounted for in terms of interactions with the primary structure, rather than with the remaining secondary structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylglucosamine↗

PubMed1984

A qualitative ultrastructural study of the intercellular spaces between epithelial cells treated in vivo with DMBA.

One of the features of epithelial dysplasia at the histological level is known as "loss of cellular adherence" in which adjacent epithelial cells appear more widely separated from each other than in normal tissues. In this study we examine the effects of the carcinogen DMBA on the epithelium of the hamster cheek-pouch with particular emphasis on the dimensions of the intercellular spaces. DMBA-induced lesions were processed for electron microscopy and assigned to hyperplasia, dysplasia and carcinoma groups, using defined criteria on toluidine blue-stained 1 micron Araldite sections. Untreated pouches were used as a control. At the light-microscopical level, intercellular spaces in hyperplastic epithelium appeared similar to those present in untreated tissue but increased progressively in dysplastic and carcinomatous lesions. Spaces were generally wider between basal and spinous cells than between granular cells, although wide variations were observed between tissue blocks demonstrating similar histological features and also within adjacent areas of the same block. At the ultrastructural level, untreated and hyperplastic tissue showed only occasional focal separations of adjacent plasma membranes; these spaces were more frequent between cells of lower strata. In sections from dysplasias and carcinomas, spaces were always extensive and were occupied by numerous villous or foliate membrane-bound cytoplasmic extensions. These were often attached to each other by desmosomes of apparently normal morphology but of a lower frequency than in untreated epithelium. The increased epithelial separation as indicated by the increased intercellular spaces during chemical carcinogenesis may be a result of any or all of the following factors: desmosomal disruption or their failure to develop; the production of cell-surface molecules which are less adhesive; inflammatory oedema and direct alterations on intercellular junctions and cell-surface components by infiltrating inflammatory cells.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1984

Alterations in the volume of the intercellular space between epithelial cells of the hamster cheek-pouch: quantitative studies of normal and carcinogen-treated tissues.

The present report investigated the extent of the epithelial dysplastic feature known as "loss of cellular adherence" at the ultrastructural level by quantifying the volume of the intercellular space during hamster cheek-pouch carcinogenesis. Following topical application of DMBA to cheek-pouches, lesions were classified as hyperplasia, dysplasia and carcinoma, with untreated pouches serving as a control. Stereological point counting procedures were used to determine the volume density of intercellular space in defined basal, spinous and granular layers for each group. In general, progressive increases in volume density were detected within each stratum during carcinogenesis. These results indicate that increasing separation of epithelial cells occurs during carcinogenesis, although it is not yet known whether this results from loss of cohesion between specialised (i.e., desmosomal) or nonspecialised membrane areas. In addition, a simple indicator of pathological alteration, the Pathological Alteration Ratio (PAR), is described and was used to evaluate existing published data for intercellular spaces in various oral mucosal conditions. Values of the PAR were found to be substantially higher in carcinogen-treated epithelia than in reports describing changes in wound healing, lichen planus and leukoplakia simplex. These objective techniques are of value for investigating the pathogenesis of diseased epithelium and may find applications in the diagnosis of oral premalignant lesions.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1984

Desmosomes in hamster cheek pouch epithelium: their quantitative characterization during epithelial differentiation.

Desmosomes in stratified squamous epithelia appear to exhibit quantitative alterations during differentiation. In this work we use stereological and other morphometric methods to quantify these structures in epithelial cells from defined basal, spinous and granular strata. Hamster cheek pouch mucosa from five animals was processed for electron microscopy using strictly standardized techniques and a stratified random sampling procedure was used to obtain micrographs of cells from basal, spinous and granular layers. Stereological intersection counting techniques were used to determine for each layer the relative surface area of plasma membrane occupied by desmosomes (Ss), the number of desmosomes per unit surface area of plasma membrane (Ns), the mean individual desmosomal diameter (delta) and the mean individual desmosomal surface area (s). In addition, estimates of nuclear volume were obtained by direct measurement of nuclear profiles and volume-to-surface ratios were obtained by a combination of point and intersection counting, which enabled estimates for the volume (Vcell) and plasma membrane surface area (SPM) of the 'average' cell within each stratum to be acquired. Using this information, it was then possible to calculate both the total surface area (S) and the number (N) of desmosomes on the plasma membranes of average cells. The parameters Ss and Ns showed progressive increases between basal and granular layers, whereas values for delta and s were lower in granular cells when compared with basal and spinous cells. The parameters Vcell, SPM, S and N all increased progressively and significantly during differentiation. Between basal and granular layers, the mean cell volume and surface area had each increased approximately threefold, whereas the surface area and number of desmosomes on the average cell plasma membrane had increased approximately seven- and eleven-fold, respectively. Granular cells thus possess more numerous desmosomes, which occupy a greater proportion of the plasma membrane area but which are individually smaller, when compared with basal and spinous layers.

Animals↗

PubMed1984

Ultrastructural morphometric of gap junctions during differentiation of stratified squamous epithelium.

The presence of gap junctions in stratified epithelia has now been extensively documented, but there have been few attempts to quantify them. In the present report, samples of hamster cheek pouch mucosa were processed for electron microscopy and electron micrographs from defined basal, spinous and granular layers were obtained. Using a combination of direct measurement and stereological intersection counting techniques, the relative surface areas of peripheral gap junctions (i.e. those in direct contact with the epithelial plasma membrane) and annular gap junctions (i.e. those present as complete, approximately circular profiles within the epithelial cell cytoplasm) were determined. Following estimation of the plasma membrane surface area of 'average' epithelial cells from each of the defined strata, relative values were transformed into absolute data. Data from peripheral and annular junctions were pooled to provide an estimate of total gap junctions area. Relative surface area estimates were similar for peripheral, annular and total gap junctions, in that values were invariably highest in the spinous layer and lowest in the granular layer. Absolute data indicate that there is more than a threefold increase in the area of membrane differentiated into gap junctions in the average spinous cell when compared with the average basal cell. Values for total gap-junctional areas in the average granular cell are reduced somewhat with respect to the average spinous cell and this is effected by a decrease in the area of peripheral gap junctions. We conclude that there is synthesis of gap junctions between basal and spinous cells, which is followed by evidence of degradation between spinous and granular cells. The magnitude of the estimates of area is comparable to those obtained from other stratified and non-stratified epithelia and it would thus appear that gap junctions may play a significant role in cellular control processes in all viable epithelial strata.

Animals↗

PubMed1984

Locus of control and oral health.

People with an "internal" locus of control believe that ability and effort are important factors in affecting what happens to them, whereas those with an "external" locus of control believe that this is more a matter of luck or fate. Patients whose oral health was rated as being "good" scored in a more internal fashion on a locus of control questionnaire than patients whose oral health was "moderate" or "poor," suggesting that locus of control beliefs are important in the design of preventive programs on a personalized basis.

Adolescent↗

PubMed1984

Some aspects of desmosomal morphology during differentiation of hamster cheek pouch epithelium.

Desmosomes function as intercellular attachment devices. In keratinised epithelia, squames are shed from the surface and these are replaced by less differentiated cells of the basal or parabasal layers, which migrate progressively towards the surface. In order to perform these migratory activities, cellular movement must occur, with the implication that cells must be able to make and break their desmosomal attachments. The present study investigates desmosomal ultrastructure with the objective of suggesting mechanisms by which desmosomes are altered during differentiation. The majority of desmosomes in basal, spinous and granular layers were similar in morphology, with differences being quantitative rather than qualitative. Desmosomes appeared to be more frequent and of smaller dimensions in granular cells when compared with those present on cells from lower strata. Upon keratinisation, desmosomal cytoplasmic structural specialisations disappeared and the intercellular contact layer became more prominent. A number of features which might represent desmosomal formation or synthesis were observed in nucleated cellular layers. These included an apparent budding or fusing of attachment plaques from existing desmosomes. Asymmetrical attachment plaques were prevalent with a markedly reduced number of tonofilaments associated with one plaque or an attachment plaque which was generally or locally reduced in electron density. Other appearances suggested that de novo desmosomal formation occurred as a result of the production of either single or paired attachment plaques, often containing fine filaments extending for a short distance into the cytoplasm. In view of the increased frequency of desmosomes in granular cells, the majority of these desmosomal abnormalities may represent synthetic rather than degradative activities.

Animals↗

PubMed1984

Morphometric studies of the epidermal-dermal junction in the rat ear: some effects of experimental friction on epidermis and anchoring fibrils.

The aim of this investigation was to determine whether, following the application of experimental friction to the skin, qualitative an quantitative differences could be induced in the epidermis and in anchoring fibrils. The skin of rat ears was subjected to repeated mild fractional stimulation with a rotating bristle brush moistened with saline thrice weekly for three weeks, untreated ears being used as controls. A stratified random sampling design was followed for the morphometric analysis of interfollicular epidermis. With the light microscope, the lengths of the granular cell/keratin (BGK) and epithelial-connective tissue (BEC) junctions were measured using an image analyser, as was the epithelial thickness (T). The ratio BEC/BGK provides an index of the degree of irregularity of the epithelial-connective tissue junction. Using electron microscopy, the numbers of anchoring fibrils per unit length of lamina densa (NB) were determined using stereological intersection counting. There are no significant differences in BEC/BGK or T between control and experimental groups, but anchoring fibril frequency NB increased from 10.29 micrometers in control epidermis to 18.9 micrometers in friction-treated epidermis. It is concluded that a mild frictional stimulus which fails to produce significant alterations in epidermal thickness can produce a marked increase in anchoring fibril frequency. This response may reflect a functional alteration associated with increased mechanical loading, in which epidermal-dermal adherence is augmented by either synthesis of anchoring fibrils or their insertion into the lamina densa.

Animals↗

PubMed1983

Inter-animal variation and its influence on the overall precision of morphometric estimates based on nested sampling designs.

This paper provides additional experimental evidence that biological variation between individuals is likely to be the major factor influencing the overall precision and efficiency of nested sampling schemes for morphometric analysis of thin sections. Four distinct experimental systems (two based on nervous tissue and two on epithelia) have been investigated. Morphometric estimates were obtained from measurements made on micrographs generated by sampling tissues at several levels. Sources of sampling variation were isolated so that the contributions to overall variance made by inter-animal differences could be evaluated. In each case, biological variation was the cardinal component of total observed variance between animals. Relative contributions ranged from 53% to 78%. Examining more animals would be the most efficient way of reducing the variance of the group mean in these sampling designs.

Animals↗

PubMed1983

Stereological studies of differentiation in hamster cheek pouch epithelium: variations in the volume and frequency of mitochondria.

The present report describes quantitative studies of mitochondria from stratified squamous epithelial cells from hamster cheek pouch mucosa in differentiating compartments. Mucosa was processed for electron microscopy and, using strict sampling procedures, electron micrographs of epithelium from defined basal, spinous and granular strata were acquired. Using stereological and other morphometric methods, the volume density, numerical density and volume-to-surface ratio of mitochondria were estimated in each cellular layer, and these data were transformed into absolute values characterising the average cell in these layers. The results suggest that both volumetric and numerical densities of mitochondria decreased progressively during epithelial differentiation; conversely, the total volume and number of mitochondria present in granular cells were consistently higher than in basal cells, as was the volume of individual mitochondria. These quantitative changes may reflect functional alterations in the metabolism of epithelial cells during differentiation and provide a base line for further work on experimental epithelial biology and pathology.

Animals↗

PubMed1983

Volumetric alterations in tonofibrils during epithelial differentiation in hamster cheek pouch mucosa.

The present report describes the use of stereological procedures to evaluate the volume of tonofibrils present in defined epithelial strata undergoing differentiation. Hamster cheek pouch epithelium was processed for electron microscopy and, following strict sampling procedures, micrographs were obtained from basal, spinous and granular layers. Using stereological point counting techniques, the volume density of tonofibrils present in unit volume of epithelial cytoplasm was determined for each layer. From previously determined estimates of cytoplasmic volume in this tissue, it was possible to calculate the absolute volume of tonofibrils present in the 'average' epithelial cell of each layer. The results describe essentially similar volume density data for each of the layers investigated. However, estimates of tonofibril volume in the 'average' cell increase more than threefold between basal and granular layers. It is concluded that as the epithelial cell differentiates through spinous and granular strata, synthesis of tonofilaments may occur.

Animals↗

PubMed1982

Studies on the relationship of disulfide bonds to the formation and maintenance of secondary structure in chicken egg white lysozyme.

The dependence of secondary structure on SS content was examined to gain further insight into the role of SS bond formation in chain folding. Lysozyme was reduced to various levels, and the products were studied with respect to remaining native structure. Reductions were carried out in the absence of denaturant. Thus, the observed changes were not attributable to a general denaturing effect, but more specifically to cleavage of SS bonds. The products of reduction were found by gel filtration chromatography to contain both polymeric and monomeric forms. By "fingerprinting", the monomers gave no indication of having undergone SS interchange, and specific cleavage of native SS bonds was indicated by a stepwise disappearance of SS-containing peptides with increasing reduction level. Ion-exchange chromatography of the carboxymethylated monomers gave the three possible reduction intermediates, containing two, four, and six carboxymethylcysteine residues per mole, in addition to the fully reduced form. The intermediates exhibited circular dichroic (CD) characteristics that were clearly nonnative, indicating alteration of the secondary structure throughout all stages of reduction. Concomitantly, there were extensive losses of enzymatic activity at intermediate levels, while no activity was found for the fully reduced form. Partially and fully reduced lysozymes were also examined for their abilities to resume native secondary structure in the absence of SS formation, as would be evidenced by a reversion to native CD characteristics. Despite literature indications of native structure in the reduced protein as observed from examination of CD behavior, none was found in the present effort, under a variety of experimental conditions. These observations are consistent with an essential role of SS bonds in forming and maintaining a major portion of the native secondary structure in lysozyme.

Animals↗

PubMed1982

Towards economy of effort in quantitative ultrastructural pathology: efficient sampling schemes for studying experimental carcinogenesis.

Hamster cheek pouch mucosa was painted with DMBA in order to compare samples of epithelium at different pathological stages of carcinogenesis. Stereological methods were applied to ultrathin sections of tissue to estimate one ultrastructural index of change, the surface ratio of lamina densa compared with the overlying plasma membrane of cells in stratum basale. Analysis of variance techniques were then employed to isolate and quantify the contributions which different levels of sampling (animals, tissue blocks, microscopic fields) made to the total observed variation in this surface ratio. Economical sampling schemes for future use were calculated from the sampling variances by taking into account the relative costs at each sampling level. Though illustrated by means of the hamster cheek pouch-DMBA model, our results are pertinent to many other experimental models for quantitative histopathology.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1982

Cellular and nuclear volumetric alterations during differentiation of normal hamster cheek pouch epithelium.

The aim of this investigation was to determine whether volumetric changes occurred in differentiating layers of stratified squamous epithelium. Samples of cheek pouch mucosa from 5 hamsters were obtained, prepared for electron microscopy using carefully controlled methods, and electron micrographs obtained from defined basal, spinous and granular layers of the epithelium. Stereological point counting procedures were used to determine the ratio of nucleus to cytoplasm for each of the defined cell layers. From direct measurement of nuclear profiles, major and minor axes were transformed to diameters of circles of equivalent area and it was thus possible to obtain an estimate of nuclear volume. Using the previously determined nuclear-cytoplasmic ratio, the volume of cytoplasm and hence cell volume, could be estimated for the cell layers. Between basal and granular layers, nuclear-cytoplasmic ratios decreased from 0.42 to 0.08, whereas cytoplasmic and cellular volumes increased progressively from 248 to 1052 microns 3 and from 352 to 1144 microns 3, respectively. Nuclear axial ratios were highest in the granular layers. These methods can be used in a variety of comparative ultrastructural studies of epithelia, and will also prove valuable in generating additional biological information from more conventionally presented stereological data.

Animals↗

PubMed1982

Stereological methods for quantifying cell surface specializations in epithelia, including a concept for counting desmosomes and hemidesmosomes.

We present stereological procedures for morphometric evaluation of intercellular junctions appearing in ultrathin sections of epithelia. A model representing desmosomes and hemidesmosomes as flat, circular discs differentiated within the plasma membrane of epithelial cells forms a basis for estimating number per unit volume of tissue or per unit area of cell surface. Methods for estimating relative surfaces are also provided. Consideration is given to problems of tissue sampling, varying feature size and finite section thickness. The procedures are improvements over earlier estimates of number per unit area of section or per unit length of membrane. Worked examples of calculations are included and methods are illustrated further by reference to quantitative studies of hamster cheek pouch epithelium during experimental carcinogenesis.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1982

Quantitative light and electron microscopical studies of the epithelial-connective tissue junction in intraoral mucosae.

The present study was designed to clarify the morphological nature of epithelial-connective tissue attachment by quantitative comparison of palatal and buccal mucosae. Tissue samples were obtained from ferrets and, following a strict sampling regime, sections were obtained for quantitative light and electron microscopy. Electron micrographs from the epithelial-connective tissue junction were subjected to analysis by serological intersection counting. Quantitative results show that palatal epithelium has a comparatively convoluted interface, and is twice as thick as buccal epithelium. Stereological data are presented for relative surface area of basal plasma membrane occupied by hemidesmosomes, mean hemidesmosomal diameter and numerical density of hemidesmosomes per unit area of basal plasma membrane. Larger hemidesmosomes are found in the palatal epithelium. Results suggest that a prominent rete-ridge pattern, as seen in the palatal epithelium, along with a greater specialized surface for adhesion, may be responsible for providing epithelial-connective tissue stability in regions subjected to high mechanical stress.

Animals↗