PubMed Health⌕ Search

Biomedical subjects

R R Cowden

Publications and source records attributed to R R Cowden.

At least 37 records · Page 2Linked to original sources
PubMed1977

Ultrastructure and histochemistry of the supportive structures associated with the radula of the slug, Limax maximus.

The odontophore and connective tissue-filled portion of the radular sac (called the "collostyle") of the slug, Limax maximus, were examined by light and electron microscopy. While both of these structures grossly resemble vertebrate cartilage, neither is composed of a type of tissue with the microscopic appearance and histochemical properties of cartilage. The roughly U-shaped odontophore possesses a thin capsule composed of connective tissue. The parenchyma of the odontophore consists of modified muscle cells which are organized into irregular groups by incomplete trabeculae composed of conventional muscle cells. The odontophoral cells are variable in size; they contain glycogen-filled "cores" as well as bundles of peripherally located filaments resembling myofilaments; and they are innervated like muscle cells. The nuclei of the cells are located eccentrically in the glycogen-filled portions of the cells and typically contain prominent nucleoli. The nuclei are surrounded by multiple small Golgi complexes and pleomorphic dense bodies resembling lysosomes. The extracellular matrix of the odontophore is very sparse and contains glycogen and fibrillar material but no histochemically demonstrable acidic mucosubstances. The collostyle consists of a gelatinous type of tissue somewhat like vertebrate mucoid connective tissue. The abundant extracellular matrix contains cross-banded filaments, a flocculent material disposed in wavy indefinite strands, and small electron-dense particles. The matrix contains histochemically demonstrable neutral and weakly acidic mucosubstances. The cell population of the collostyle includes solitary muscle cells and fibrocytes containing large quantities of glycogen.

Animals↗
PubMed1976

Cytochemical evaluation of the Guard procedure a regressive staining method for demonstrating chromosomal basic proteins. I. Effects of fixation, blocking reactions, selective extractions, and polyacid "differentiation".

Appropriately fixed preparations stained by a modification of the Guard (1959) reaction for "sex chromatin" display selective staining of interphase chromatin and mitotic or meiotic chromosomes. This is a regressive staining method which seems to depend on the selective displacement of an acidic dye from less basic structures, and retention of the dye at more basic sites. The results obtained with the reaction can be controlled by the length of time that the preparations are "differentiated" in solutions containing phosphomolybdic and phosphotungstic acids (polyacids). After three- or four-hour exposures to polyacid solutions, all chromatin is stained. However, with longer differentiation, "condensed" chromatin can be stained preferentially. Of a number of fixatives investigated, only 10% formalin, ethanol-acetic acid (3:1), and Bouin's solution proved useful. Others resulted in diminished specificity or a total loss of selectivity. The most intense staining was obtained after formalin fixation. Less intense dyebinding was observed after fixation in 3:1 - probably due to extraction of some histone fractions-and the least amount of dye was bound in Bouin's-fixed chromatin - probably due to blockage of arginine residues by picric acid. The reaction was not affected by enzymatic removal of nucleic acids or the extraction of lipids. It was diminished by treatment with trypsin or weak acetylation, and it was completely prevented by strong acetylation, deamination, or extraction of basic proteins with HCl. The results presented suggest that the modified Guard (1959) procedure selectively demonstrates basic nucleoproteins. Further, by the use of regressive differentiation in polyacid solutions, the retention of dye in more condensed chromatin can be favored.

Animals↗
PubMed1976

Cytochemical characterization of the modified Guard procedure a regressive staining method for demonstrating chromosomal basic proteins. II. Substitution of dyes for biebrich scarlet.

A number of acidic dyes, including various fluorochromes, were substituted for biebrich scarlet in the modified Guard (1959) procedure, a regressive staining method which appears to demonstrate basic chromosomal proteins. These substitutions were made to test the possibility that dyes other than biebrich scarlet might provide advantages in sensitivity and/or contrast, or that more control could be exerted over the "differentiation" step in which solutions of phosphotungstic and phosphomolybdic acids (polyacids) are used. Of the dyes tested in this investigation, six were found to be especially suitable: procion brilliant blue, procion yellow, geranine G, brilliant sulfoflavine, eosin Y, and eosin B. While procion brilliant blue could be used as an absorption dye only, the other dyes were used more profitably as fluorochromes. The various dyes displayed considerable variability in the ease with which they could be displaced from substrates with polyacid solutions during the differentiation step. Procion brilliant blue, procion yellow, and geranine G were displaced gradually and thus resembled biebrich scarlet. In contrast, eosin B, eosin Y, and brilliant sulfoflavine were displaced more easily from all but the most highly condensed chromatin in substrates. Brilliant sulfoflavine yeilded exceptionally bright and nearly selective fluorescence of consensed chromosomes in division, "X" chromosomes of grasshopper spermatocytes, and sperm heads. Weak, but selective fluorescence was observed when monazo sulfonated dyes, including ponta chrome violet SW, eriochrome black, diamond red, and ponta chrome blue black, were substituted in the modified Guard procedure. Similar results were obtained with solochrome cyanin R. As expected, these dyes seemed to function as weakly acidic dyes.

Animals↗
PubMed1976

Some quantitative aspects of Acridine Orange fluorescence in unfixed, sucrose-isolated mammalian nuclei.

Nuclei were isolated from mouse liver and central nervous system (CNS). These nuclei were fluorochromed without fixation in a 0.25 M sucrose medium containing 5-5 X 10(-5) M Acridin Orange and measured with an incident microfluorometric system. In the case of mouse CNS nuclei, the major and minor axes of the nuclei were measured with a filar micrometer. Three modal values were obtained from the hepatocyte suspension corresponding to 2C, 4C, and 8C nuclei, respectively. While the CNS nuclei displayed substantial variability in size, the Acridine Orange emission values at 530 nm were nearly constant. The data suggest that under these conditions, Acridine Orange fuorescence reflects DNA content. Further, the 530 nm fluorescence emission is not affected by chromatin condensation or proteins complexed with DNA.

Acridines↗
PubMed1976

A comparison of two methods of preparing cells or nuclei for high-resolution microspectrophotometry.

Suspensions of isolated mouse thymus cells were subjected to two preparative methods: either they were dropped through several mm of 9:1 v/v ethanol-acetic acid fixative, allowed to stand for 1hr and then processed for staining; or they were fixed, passed through a graded ethanol series to 70% ethanol, centrifuged on to slides in a modified Shandon cytocentrifuge and then carried wet into the staining procedure. All preparations were stained by the Feulgen reaction and evaluated by high-resolution microspectrophotometry. While the two preparative procedures yielded similar results, there appeared to be less variability in the data obtained from the centrifuged cell populations.

Animals↗
PubMed1976

Fluorescence histochemistry of the surface epithelium and stroma in the rhesus monkey ovary.

The surface epithelium and stromal elements in the ovary of the rhesus monkey were studied with a variety of fluorescence-histochemical procedures to determine the relative distribution of specific chemical moieties. The surface (germinal) epithelium displayed high levels of cytoplasmic RNA and specifically intense fluorescence with methods that demonstrate basic, SH and SS groups of protein. The underlying basement membrane demonstrated an intense reaction when fluorochromed with hydroxynaphthoic acid hydrazide directed toward side-chain carboxyl groups of protein. The fluorescence pattern displayed by the underlying connective tissue was heterogeneous and related to its anatomical localization. It is suggested that this heterogeneity reflects specific, but as yet undefined, chemical differences in the fibrous elements of the ovary.

Animals↗
PubMed1975

A comparison of four quantitative cytochemical methods directed toward demonstration of DNA.

Populations of nuclei isolated from mouse brain tissue were stained by the following cytochemical methods considered stoichiometric for DNA: (1) the Feulgen reaction; (2) gallocyanin-chromalum after RNase; (3) pH 4.0 methylene blue after RNase; and (4) methyl green used in the presence of 2M magnesium chloride. Replicate preparations to be stained with gallocyanin-chromalum, methylene blue, and methyl green were acetylated prior to staining. All of these groups were examined by high-resolution scanning microspectrophotometry. The results indicated that of the methods examined, the Feulgen reaction, gallocyanin-chromalum used without prior acetylation, and methylene blue used with prior acetylation were the most useful in revealing differences attributable to variability in chromatin organization. The greatest variability in total extinction measurements was observed in acetylated, methylene blue-stained nuclei, while the least variability was observed in nuclei stained with methyl green in the presence of 2 M magnesium chloride. Acetylation produced different effects on dye-binding in different groups. It greatly increased binding in nuclei stained with methylene blue; it reduced binding in the methyl green-2 M magnesium chloride series.

Acetylation↗
PubMed1975

A fluorescent water soluable carbodiimide-2-hydroxy-3-naphthoic acid hydrozide reaction for the demonstration of carboxyl groups in proteins and mucosubstances.

The carbodiimide-2-hydroxy-3-haphthoic acid hydrazide reaction as developed by Geyer (1964) was used without subsequent diazonium coupling as a fluorescent method for the demonstration of carboxyl groups in both proteins and mucosubstances. The topological distribution of the fluorophore was similar to that reported by Geyer. Quantitative microfluorometric studies on cartilage sections revealed differences in detail between emissions in cartilage matrix mucoprotein as compared to the dense connective tissue associated with the perichondrium which consists principally of protein. It would also appear that the primary fluorescent emission of unstained preparations at 450 mm should be useful in microfluorometric determinations of proteins.

Animals↗
PubMed1975

Feulgen microspectrophotometric analysis of deoxyribonucleoprotein organization in larval and adult freshwater sponge nuclei.

Microspectrophotometric measurements of nuclei of adult choanocytes and archeocytes and larval archeocytes and flagellated ectodermal epithelia from stages III and IV were undertaken on tissues from the freshwater sponge, Eunapius fragilis (Leidy). The condensed nuclei of differentiated choanocytes and state IV flagellated epithelial cells presented integrated extinction values about 64% of those obtained for either adult or larval archeocytes. This suggests that such measurements represent sensitive indicators of deoxyribonucleoprotein-complex organization; thus, nuclear differentiation. Further, no large population of G2 (4C) nuclei was found among these populations as has been reported in two classes of coelenterates.

Animals↗
PubMed1975

Microspectrophotometric estimates of non-histone proteins in cell nuclei displaying differing degrees of chromatin condensation.

Nuclei isolated from mouse thymus, kidney, and liver were fixed in ethanol-acetic acetic acid; treated with dilute acid to extract histones; stained by three protein end-group procedures; and measured by scanning, integrating microspectrophotometry. Measurements were also made of nuclei isolated from the same organs and stained by the Feulgen procedure for DNA. Protein end-group procedures included pH 2.8 Biebrich scarlet (for basic groups), mercury orange (for sulfhydryl groups), and mercury orange after thioglycolate reduction (for the sum of sulfhydryl and disulfide groups). With the exception of the comparison between Feulgen-stained 2C liver and kidney nuclei, the integrated extinction values obtained for nuclei of a given organ differed significantly from the measurements of nuclei from other organs, regardless of the staining procedure. Furthermore, the integrated extinction values for 2C nuclei were highest in larger, more vesicular nuclei (from liver and kidney) and lowest in condensed thymocyte nuclei, except in the case of measurements of the disulfide content of the nuclei. In this instance, the values of integrated extinction were highest in condensed thymocyte nuclei, intermediate in kidney nuclei, and lowest in 2C liver nuclei. When 2C, 4C, and 8C liver nuclei were compared, the integrated extinction values of 4C nuclei were found to be approximately twice those of 2C nuclei whose disulfide and Feulgen values were, respectively, higher and lower than expected. The greater disulfide values and reduced Feulgen values obtained in thymocyte and 8C liver nuclei might be related to a greater degree of chromatin condensation in these nuclei, and therefore, to a reduction or selective restriction of their RNA transcriptional capacities.

Animals↗
PubMed1975

The histochemistry and structure of tentacle cartilage tissues in the marine polychaete, Sabella melanostigma.

The cartilages (or "chondroid" tissue) in tentacles of the polychaete annelid, Sabella melanostigma, have been examined by electron microscopy and a series of histochemical techniques for the demonstration of mucopolysaccharides and protein end-groups. The ultrastructural studies indicated that the cartilages possess an investing layer of dense connective tissue which differs significanly from the matrix material secreted between the chondrocytes. The cartilage matrix was positive for acidic mucins with levels of sulfation above those of mammalian chondroitins A and C. This matrix as well as the investing connective tissue were intensely PAS-positive. Sabella cartilage was also stained intensely by methods for demonstrating tryptophan, tyrosine, side-chain carboxyl groups, disulfide groups, and amino groups. It was not stained by the procedure for sulfhydryl groups. Some evolutionary aspects of cartilage and chondroid tissues were discussed.

Amino Acids↗
PubMed1975

Fluorescent demonstration of basement membranes with hydroxy-naphthoic acid hydrazide.

The hydroxy-naphthoic acid hydrazide (HNAH) procedure can be employed not only for selective fluorochroming of side chain carboxyl groups of proteins, but also for demonstration of certain exoplasmic structures---particularly basement membranes. The demonstration of basement membranes by the HNAH procedure is likely related to the relatively high levels of available side chain carboxyl groups in the noncollagen glycoproteins of basement membranes.

Animals↗
PubMed1975

Evaluation of two protein end-group reactions as potential fluorescent cytochemical methods.

Histochemical preparations stained by a variant of the Morel-Sisley reaction for protein tyrosine were found to produce a red fluorescence when excited by broadband blue light which is topologically identical to the distribution of chromophore when viewed by absorption (equal transmission) microscopy. The fluorescence mode of viewing preparations stained by this method gave greater sensitivity and contrast than the absorption mode. The p-dimethylaminobenzaldehyde (DMAB)-nitrate method for protein tryptophan did not result in a useful fluorescent end-group method. Preparations stained by this method displayed a pattern of generalized fluorescence of all structures except those which react in the final step of the p-DMAB-nitrite reaction. The specificity of the intermediate reaction product has yet to be established.

Amphibians↗
PubMed1975

Fluorescent cytochemistry of calid and algid normal and Lucké tumor-bearing kidneys.

Fluorescent cytochemical techniques, chiefly end-group methods, have been employed in the characterization of cells in calid and algid normal and LUCKE tumor-bearing kidneys of the leopard frog, Rana pipiens. Cytoplasmic RNA and glycogen distribution reflect the known patterns of general metabolic activity of the respective cells in question. Protein sulfhydryl distribution is similar in normal distal tubules and tumors while both differ significantly from proximal tubules. Viral inclusions in nuclei of algid tumor cells contain demonstrable DNA, protein sulfhydryls and disulfides, arginine and lysine-rich basic proteins, also both C-terminal and side chain carboxyl groups. Significant amounts of DNA are demonstrable in the cytoplasm of algid tumors. Fluorescence cytochemical methods directed toward end-groups or specific classes of macromolecules offer a particularly favorable option, because of increased sensitivity and higher levels of contrast, for comparative cytological investigations of the LUCKE tumor and of similar systems.

Adenocarcinoma↗