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Biomedical subjects

S S Spicer

Publications and source records attributed to S S Spicer.

At least 19 recordsLinked to original sources

Ultrastructural localization and semiquantitative analysis of glycoconjugates in the tectorial membrane.

The tectorial membrane of the gerbil cochlea was analyzed with lectin-gold cytochemical methods for demonstrating and characterizing glycoconjugates (GCs) in situ. Binding of lectins from Limax flavus (LFA), Lens culinaris (LCA), Datura stramonium (DSA), Ricinus communis (RCA I), Ulex europeus (UEA I) and Phaseolus vulgaris (PHA L) was assayed semiquantitatively on ultrathin sections. Binding occurred throughout the tectorial membrane with all lectins except UEA I but the labelling density with a given lectin differed among substructures. The cover net disclosed the highest level of GC with four lectins whereas the fibrous layer revealed the lowest level. DSA, LCA and PHA L demonstrated considerable similarity between the cover net and the marginal band in content of GC with N-linked oligosaccharide. The cover net differed from the marginal band, however, in containing more RCA I reactive GC with terminal lactosamine. Hensen's stripe, with which inner hair cell stereocilia are thought to interact, differed from other substructures in containing the highest level of PHA L-reactive traintennate N-linked chains and except for the basal layer the lowest concentration of GC with terminal lactosamine. Fucosylated GC detectable with UEA I-gold was present at low levels in all substructures except the cover net and marginal band. Distribution of GCs in the fibrous layer and less consistently in the cover net differed between limbal and middle zones. The differences observed here in the carbohydrate composition among substructures in the tectorial membrane support and extend previous cytochemical observations and imply a role for different classes of GCs in determining the biophysical and physiological properties of the tectorial membrane.

Animals

Creatine kinase in epithelium of the inner ear.

Epithelium of the inner ear in the gerbil and mouse was examined immunocytochemically for presence of creatine kinase (CK). Marginal cells of the cochlear stria vascularis and dark cells and transitional cells of the vestibular system were found to contain an abundance of the MM isozyme (MM-CK). CK in these cells concurs with that which is coupled to Na,K-ATPase in other cells and is considered to supply ATP for the Na,K-ATPase that mediates the high KCl of endolymph. Inner hair cells revealed content of the BB isozyme and in this respect resembled the energy-transducing photoreceptor cells in retina. In addition, outer phalangeal (Deiters') cells stained for both MM- and BB-CK whereas inner phalangeal cells evidenced content of only the BB isozyme. Immunolocalization of CK appeared similar in mouse and gerbil inner ear. Specificity of the staining was affirmed by observations in agreement with those reported for CK in various cell types and by staining with antisera from more than one source.

Animals

Histochemical analysis of glycoconjugates in gelatinous membranes of the gerbil's inner ear.

The gelatinous membranes of the gerbil inner ear were analyzed histochemically for glycoconjugates with a battery of twenty horseradish peroxidase-conjugated lectins. Glycoconjugates with mannose (Man) and/or glucose (Glc), galactose (Gal), fucose (Fuc), N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc) and N-acetylneuraminic acid (NeuAc) were detected in the tectorial and otolithic membranes and cupula. Differences in lectin reactivity were observed between tectorial and vestibular membranes and also among zones and between the medial and lateral regions of the middle zone of the tectorial membrane. The distribution of staining differed markedly for several lectins that bind specifically to GalNAc or to GlcNAc but vary in affinity for oligosaccharides containing these sugars in different sequences or linkages. The findings suggest presence of the terminal disaccharides GalNAc alpha 1,3Gal in tectorial membrane and Gal beta 1,3GalNAc in vestibular membranes. Lectin binding profiles provided evidence that the limbal zone's fibrous and attachment layers contain mainly O-glycosidically linked oligosaccharides whereas the middle zone's medial fibrous layer contains both O- and N-linked chains. The remaining regions of the tectorial membrane contain mainly N-linked oligosaccharides with bisected biantennary type chains predominating. Additionally, the marginal band and the middle zone's basal layer contain abundant N-linked oligosaccharides with a triantennary structure.

Animals

Differentiation of inner ear fibrocytes according to their ion transport related activity.

Fibrocytes in the lateral wall and limbus of the gerbil cochlea evidenced a capacity for ion transport activity by immunostaining for transport mediating enzymes including Na,K-ATPase, carbonic anhydrase (CA) and creatine kinase (CK). Fibrocytes of the spiral ligament unlike those in the suprastrial region and limbus decreased in abundance from base to apex. Spiral ligament fibrocytes at a given position along the cochlea varied in content of transport related enzymes, and on the basis of immunostaining, location and orientation, were classified into four types. Type I fibrocytes under the stria vascularis stained for CA isozymes II and III and CK isozyme BB. Type II fibrocytes under the outer sulcus and spiral prominence epithelium were found to contain only Na,K-ATPase. Type III fibrocytes lying adjacent to bone in the inferior region of the spiral ligament contained CA II and III and CK isozymes BB and MM. Type IV fibrocytes located more superficially in the inferior part of the spiral ligament stained variably for all the enzymes. Superficial fibrocytes in the suprastrial area disclosed Na,K-ATPase whereas the underlying fibrocytes stained for CA and CK. Limbal fibrocytes reacted with antisera to all the enzymes except CA III. Most fibrocytes in stromal plates beneath the vestibular system's neurosensory epithelium contained Na,K-ATPase and CA II but not CA III. These findings point to cooperativity in fluid and ion transport between epithelial cells and neighboring fibrocytes and demonstrate functional diversity of fibrocytes of the inner ear providing a basis for classifying those in the spiral ligament.

Animals

Distribution of glycoconjugates in ion transport cells of gerbil inner ear.

Ion transport cells in gerbil inner ear were differentiated histochemically by staining glycoconjugates (GCs) with a battery of horseradish peroxidase-conjugated lectins. Strong staining with PSA and LCA showed a high content of N-linked oligosaccharides in transport cell GCs. Reactivity with PHA-L and PHA-E identified GC with triantennary and with bisected biantennary N-linked oligosaccharides, respectively, in these cells. High affinity for DSA and PWM demonstrated abundant N-acetyl lactosamine in N-linked side chains. Ion transporting epithelial cells reacting with lectins specific for N-linked oligosaccharides included strial marginal cells and outer sulcus cells of the cochlea and dark cells, transitional cells, and planum semilunatum cells of the vestibular system. In general, all of the inner ear transport epithelial cells revealed a similar lectin binding profile, with the one exception that SBA reacted strongly with ion transporting cells in the vestibular system but only weakly with those in the cochlea. Fibrocytes specialized for ion transport located in distinct areas in the suprastrial and inferior regions of the spiral ligament also stained with lectins that demonstrate N-glycosylation. However, transport fibrocytes differed from transport epithelial cells in two ways. First, they reacted e with HPA, DBA, VVA, and SJA specific for O-linkages and second, they failed to react with UEA I. The staining pattern for N-glycosylated GC resembled that for Na+, K(+)-ATPase in inner ear, suggesting a relationship between these constituents.

Animals

Comparative distribution of carbonic anhydrase isozymes III and II in rodent tissues.

Carbonic anhydrase (CA) III was demonstrated immunocytochemically in epithelium in some regions of salivary gland ducts, colon, bronchi, and male genital tract and in adipocytes, in addition to skeletal muscle and liver where the isozyme was previously localized. Basal cells beneath the submandibular gland's excretory ducts in guinea pig stained for CA III. Carbonic anhydrase III occurred alone in some and with CA II in other sites but was often absent from CA-II-containing types of cells. This was exemplified by CA III's abundance in CA-II-positive proximal colon and its sparsity in the CA-II-rich distal colon of the mouse. Striated ducts in guinea pig, but not mouse salivary glands, stained darker for CA and appeared accordingly to function more actively in ion transport compared with excretory ducts. Carbonic anhydrase content varied among genera in liver and pancreas and between mouse species and strains in salivary glands and kidney. Newly observed murine sites of CA II activity included Auerbach's plexus and a population of leukocytes infiltrating the lamina propria in small intestine, and several types of cells in the male genital tract. In immunoblot tests, antisera to CA III showed no cross reactivity with antisera to CA II, but those to CA II disclosed weak cross reactivity with CA III.

Adipose Tissue

Giant mitochondria distinct from enlarged mitochondria in secretory and ciliated cells of gerbil trachea and bronchioles.

Numerous mitochondria ranging from slightly larger than normal to several micrometers in diameter (giant) were found in about one-half the serous secretory cells in the surface epithelium of the normal gerbil trachea and proximal bronchi. Tracheal serous cells of mice also were found to contain numerous giant mitochondria. Clara cells of gerbil bronchioles contained abundant giant mitochondria in addition to normal tubular mitochondria and the second population of enlarged spherical mitochondria that have been described in Clara cells of several genera. In contrast, mouse Clara cells revealed the normal tubular and the enlarged spherical mitochondria but no giant mitochondria. A survey of a number of cell types in gerbils failed to disclose hypertrophied mitochondria outside tracheobronchial surface epithelium and bronchioles. The mitochondrial enlargement resulted from an increase of matrix but not cristae. The expansion of matrix displaced the relatively sparse cristae into small collections compressed against the outer membrane. The prevalence of giant mitochondria and of granular endoplasmic reticulum is similar among cells, and these two organelles are codistributed within cells. The megamitochondria and granular reticulum occupy a central stratum, whereas normal mitochondria occur in the apical and basal regions. The giant mitochondria are considered related to a normal biologic activity that is characteristic of respiratory tract epithelium of mice and gerbils selectively and is more prominent in secretory cells than in ciliated cells.

Animals

Glycoconjugate unique to migrating primordial germ cells differs with genera.

Previous cytochemical studies showing that rat primordial germ cells (PGCs) possess a unique surface glycoconjugate containing terminal alpha-N-acetylgalactosamine were extended in this study to determine whether a similar distinctive glycoconjugate coats the surface of PGCs in the mouse. The results showed that mouse PGCs fail to react with peroxidase-conjugated lectins specific for localizing glycoconjugate with terminal N-acetylgalactosamine. All available lectin conjugates with affinity for other terminal sugars or internal sugar linkages also failed to stain mouse PGCs except for the conjugates that bind to alpha-fucose. One fucose-specific lectin conjugate stained only PGCs in the early mouse embryo but stained additional sites in more mature embryos and lost reactivity with PGCs after gestational day 14. Another fucose-specific conjugate stained PGCs until day 15, but with less selectivity, and a third such conjugate bound to several sites, but not to PGCs. The results suggest that the developmental mechanisms mediating cellular interaction, migration, and differentiation may be similar in different genera, but the specific structure of the cell surface glycoconjugate involved in these mechanisms differs.

Animals

Distribution of glycoconjugates in the optic vesicle and optic cup.

Lectin histochemical methods and immunohistochemical techniques have been utilized to investigate and partially characterize glycoconjugates in the developing eye. Peanut-lectin-binding sites associated with radial glial cells were found in the diencephalon. In the optic primordia, binding sites associated with radial glia were masked by terminal sialic acid, and only reacted with peanut lectin when pretreated with sialidase. This finding indicates that glycoconjugates associated with diencephalic radial glia contain terminal galactose-beta-(1----3)N-acetyl galactosamine, but glycoconjugates associated with radial glia in the optic primordia contain sialic acid----galactose-beta(1----3)N-acetyl galactosamine. The selective distribution of galactose, N-acetyl galactosamine and fucose associated with radial glial cells has also been demonstrated. We postulate that these distributions mediate the shaping of the developing eye.

Animals

Immunolocalization of Na+,K(+)-ATPase and carbonic anhydrase in the gerbil's vestibular system.

The distribution of two ion transport enzymes in the vestibular system was investigated immunocytochemically. Immunostaining demonstrated abundant Na+,K(+)-ATPase in the basolateral plasmalemma of all dark cells and of cuboidal (transitional) cells bordering maculae and planum semilunatum cells bordering cristae. Na+,K(+)-ATPase was also present in nerve terminals impinging on vestibular hair cells and around nerve fibers and ganglion cells. Na+,K(+)-ATPase containing cells with fine intertwining processes were found within the perilymphatic stroma beneath maculae and cristae. These cells and interspersed nerves form a distinct, highly cellular plate that lies under neurosensory epithelium selectively. The catalytic alpha subunit of Na+,K(+)-ATPase in vestibular epithelia differs antigenically from the alpha subunit in nerves and from the alpha subunit in salivary gland and renal epithelium. Carbonic anhydrase (CA) isozyme II was localized in the apex of all supporting cells in neurosensory epithelia. In contrast, CA II immunostaining varied in vestibular dark cells showing heterogeneity in ion transport activity among these cells. Immunostaining evidenced CA II also in perilymphatic stromal cells which were presumably fibroblastic in nature and which correspond in location with the Na+,K(+)-ATPase positive cells under the vestibular neurosensory epithelium.

Animals

Lectin binding beneath the epithelium and in smooth muscle cells in the developing bronchial tree.

Components of the subepithelial stratum in developing rat lung reacted transiently with Maclura pomifera agglutinin (MPA) and Aleuria aurantia agglutinin (OFA) conjugated to horseradish peroxidase. These lectins possess selective affinity for and serve to localize glycoconjugates (GCs) with terminal Gal/GalNAc and Fuc, respectively. Staining was strongest with both lectins in the proximal bronchial tree and decreased peripherally to growing buds where it was absent. MPA staining of subepithelial structures decreased from the pseudoglandular through the canalicular period and disappeared by the terminal sac stage. Disappearance of this subepithelial reactivity coincided with appearance of apical MPA-positive glycoconjugate in the canalicular period. OFA stained selectively a layer of flattened cells and a thin extracellular stratum under the epithelium of proximal bronchi in the canalicular period. This lectin affinity extended farther peripherally in the pseudoglandular interval and diminished thereafter. The layer of OFA-positive cells underlying the epithelium was identified immunohistochemically as immature smooth muscle. These muscle cells gained contractile protein while losing surface lectin reactivity during fetal development. The high iron diamine method localized sulfated GC in basement membrane of proximal respiratory passages in the fetal lung. The results attest to the involvement of specific GCs in mediating epithelial-mesenchymal cell interaction during critical stages of bronchial morphogenesis.

Animals

Immunohistochemical localization of sodium-potassium-stimulated adenosine triphosphatase and carbonic anhydrase in human colon and colonic neoplasms.

Sodium-potassium-stimulated adenosine triphosphatase and carbonic anhydrase isozymes I and II were localized immunocytochemically in adenomas, adenocarcinomas, and normal epithelium of human colon harboring non-neoplastic lesions. Non-neoplastic control colon showed carbonic anhydrase I and II in the cytoplasm of the columnar cells lining the upper half of the crypts. Antiserum to sodium-potassium-stimulated adenosine triphosphatase bound to the basolateral but not the apical plasmalemma of columnar epithelial cells. Staining was most intense in the superficial cells, which also contained carbonic anhydrase, but was also evident to a lesser degree in cells deep in the crypts. Adenomas and adenocarcinomas failed to stain for content of carbonic anhydrase but retained basolateral sodium-potassium adenosine triphosphatase positivity. The staining characteristics of colonic neoplasms for the two enzymes involved in the transport function of colonic epithelium thus resembled those of the less mature cells lining the base of normal crypts.

Adenocarcinoma

Glycoconjugate with terminal galactose. A selective property of macrophages in developing rat lung.

Pulmonary macrophages in pre- and postnatal rats were examined histochemically with a battery of peroxidase labeled lectins. Among them, Griffonia simplicifolia agglutinin I-B4 (GSA I-B4) which binds specifically to terminal alpha-galactose showed selective affinity in lung for the monocyte-macrophage line. These cells were demonstrable with GSA I-B4 from the 14th day of gestation through the adult. Extension to the ultrastructural level showed strong selective binding of this lectin to the surface of the plasmalemma and inner face of membranes limiting phagosomes in macrophages. At day 14 of gestation, monocyte-like cells positive with GSA I-B4 were scattered in various organs including lung. The lectin reactive cells in lung increased in number and size with development, infiltrating the interstitium through day 20 of gestation and then also entering the alveolar space. These findings suggest that GSA I-B4 recognizes a surface glycoconjugate characteristic of the pulmonary monocyte-macrophage line. Such selective lectin affinity offers a marker for detecting the pulmonary macrophages and examining their kinetics by light and electron microscopy.

Aging

Comparison of glycoconjugates at the surface of developing type II pneumocytes and Clara cells.

The apical surface coat of type II pneumocytes and Clara cells in pre- and post-natal rat lung was examined with lectin histochemical methods. Lectins from Helix pomatia (HPA), peanut (PNA) and Maclura pomifera (MPA) were conjugated with horseradish peroxidase and used to stain paraffin sections of fixed lung with or without certain pre-treatments. HPA and MPA were observed to react with almost all type II pneumocytes at postnatal day 1. Type II pneumocytes that stained with a sialidase-PNA sequence increased from a few positive cells at postnatal day 5 to many in the adult. It has been reported that the surface coat of type II pneumocytes closely resembles that of Clara cells in its staining with histochemical methods employing cationic dyes or lectins including MPA and PNA. However, staining with HPA, especially after periodic acid oxidation, revealed many type II pneumocytes with strong reactivity but showed only a few Clara cells that were faintly positive. HPA also stained alveolar macrophages. The HPA affinity of macrophages, however, was labile to oxidation with periodic acid or galactose oxidase unlike that of type II pneumocytes. This difference suggests that HPA recognizes more than one type of sugar structure.

Age Factors

Transcutaneous oxygen tension measurements during hemorrhagic hypoperfusion using Trendelenburg and the pneumatic antishock garment.

Transcutaneous oxygen tension (PtCO2) was observed during hemorrhagic hypoperfusion using four therapeutic modalities: pneumatic antishock garment (PASG), 20 degrees Trendelenburg positioning, combined PASG-Trendelenburg, and whole blood infusion. Anesthetized mongrel dogs were mechanically ventilated. A heated transcutaneous oxygen sensor was applied to the skin overlying the sternum. Animals were bled over 10 min of 25% of their calculated blood volume. A therapeutic intervention was applied at the onset of hemorrhage (PASG, Trendelenburg, PASG-Trendelenburg or control). All animals were observed for 20 min, then during a 10-min shed blood reinfusion period, and for 20 min thereafter. PtCO2 was measured continuously and the following were measured serially: cardiac output, mean arterial pressure (MAP), mixed venous oxygen tension (MvO2), and arterial oxygen tension (PaO2). Cardiac index (CI) and the oxygen extraction ratio were calculated. PtCO2 decreased immediately after hemorrhage in all animals. Control values remained consistently below values for active interventions during this time. All groups regained baseline levels of PtCO2 after reinfusion of shed blood volume. PaO2 remained nearly constant during all experiments. MAP and CI fell in all groups following hemorrhage but did so less precipitously in the PASG group. The PASG and PASG-Trendelenburg groups showed the greatest increase in CI during reinfusion. These results suggest that when PtCO2 is relied upon as an indicator of adequacy of resuscitation during moderate hemorrhagic shock, that cutaneous perfusion may be improved by the PASG or Trendelenburg position, and that perfusion is most effectively restored by blood infusion.

Animals

Mice carrying a CAR-2 null allele lack carbonic anhydrase II immunohistochemically and show vascular calcification.

Mutant mice reported to lack carbonic anhydrase isoenzyme II (CA II) have been examined here for immunocytochemical evidence of CA II and for histopathologic change. All histologic sites that immunostain for CA II in a wide range of organs in normal mice failed to show such immunoreactivity in the homozygous mutants. The CA II-deficient mice differed from controls in evidencing an age dependent medial calcification of small arteries in a number of organs. The male genital tract revealed the most extensive arterial calcinosis and males were possibly more affected in general than females. One or another Car-2n/Car-2n mouse showed changes additionally in uterus, small bowel, lymph nodes, or renal pelvis.

Aging

Heterogeneity of macrophages evidenced by variability in their glycoconjugates.

Staining rat tissues with a battery of 15 lectin-horseradish peroxidase conjugates showed that macrophages contain glycoconjugates possessing terminal alpha, beta-galactose, N-acetylgalactosamine, fucose, and N-acetylneuraminic acid, plus two terminal disaccharides. Dissimilar binding of lectins by different phagocyte populations in the same or different organs evidenced variability in glycoconjugates according to the location of the macrophages. With a group of four lectins, macrophages stained most intensely in lung, next strongest in splenic red pulp and lymph node sinuses, and weakest in skin and liver. Two populations of macrophages were newly recognized in spleen on the basis of content of fucose-rich glycoconjugate. These included a necklace-like band of macrophages at the border between marginal zone and germinal center and distinctive macrophages dispersed throughout the marginal zone. In lymph nodes, phagocytes stained strongly in the germinal centers and weakly in sinuses for glycoconjugate with N-linked oligosaccharides and conversely for glycoconjugate with terminal beta-galactose. Variable lectin binding indicated heterogeneity of thymic macrophages. Lectin cytochemistry offers increased sensitivity for detecting macrophages in tissue sections, provides selective staining that shows the prevalence and distribution of the phagocytes and differentiates macrophages into separate subtypes.

Animals