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Biomedical subjects

L A Johnson

Publications and source records attributed to L A Johnson.

At least 145 records · Page 8Linked to original sources
PubMed1987

Gallbladder adenocarcinoma: the prognostic significance of histologic grade.

Gallbladder adenocarcinoma (GBA) postresection 5-year survival rates are less than 5%, but when histologically confined to the mucosa or submucosa, survival rates of 64% (5 years) and 44% (10 years) have been reported. Whether any other histologic features of GBA have prognostic significance is unknown. This investigation was conducted to determine if GBA histologic grade correlates with survival. Thirty patients with advanced stage GBA participating in Eastern Cooperative Oncology Group (ECOG) treatment protocol EST-2273 served as the study material. Using glandular tumor grade criteria recommended by others, a panel of 7 ECOG pathologists categorized the GBA as either predominantly low or high histologic grade. Each patient's GBA histologic section measured no less than 1.0 X 1.0 cm. Predominant grade was defined as being that grade present in greater than 75% of the histologic section. Patient survival times by grade were calculated from date of initiation of chemotherapy until date of death. The 13-week low grade GBA patient survival was significantly longer than the 7-week high grade GBA patient survival (p less than 0.01). Stratification of patients by either high or low predominant histologic grade is recommended in future GBA treatment studies.

Adenocarcinoma↗
PubMed1987

Flow sorting of X and Y chromosome-bearing spermatozoa into two populations.

The only established difference on which to base the separation of X and Y chromosome-bearing spermatozoa is chromosomal constitution. This difference is quantifiable both from chromosome morphology (karyotype) and from DNA content. Flow cytometric techniques were used to measure relative DNA content of the X and Y populations and to flow-sort spermatozoa from Chinchilla laniger. Epididymal spermatozoa were recovered in PBS, fixed in 80% ethanol, treated with papain and dithioerythritol, and stained for DNA with Hoechst 33342. Sperm nuclei were analyzed and sorted on an EPICS V flow cytometer/cell sorter, modified specifically for spermatozoa. Two clearly resolved peaks (coefficient of variation less than 1.5%) with approximately 7.5% difference in DNA content between X and Y chromosome-bearing spermatozoa were evident. Sperm nuclei were sorted from a portion of the X and Y peaks at a rate of 55 nuclei/sec for each population. Purities of individual X and Y populations averaged 95% as determined by reanalysis of the sorted populations. Successful sorting of Chinchilla X and Y chromosome-bearing spermatozoa into separate populations may aid in the identification of a biochemical marker that could be used to discriminate between the two sperm populations and lead to a practical procedure for sexing spermatozoa.

Animals↗
PubMed1987

Effect of liquid storage and cryopreservation of boar spermatozoa on acrosomal integrity and the penetration of zona-free hamster ova in vitro.

The effect of liquid storage and cryopreservation of boar spermatozoa on sperm motility, acrosomal integrity, and the penetration of zona-free hamster (ZFH) ova was examined. The sperm penetration assay (SPA) provides valuable information on specific events of fertilization and is a potentially useful indicator of sperm fertility. Ejaculated semen from 4 boars was subjected to 3 treatments: fresh (FRE, no storage), liquid-stored (LIS, stored at 18 degrees C for 3 days), and frozen (FRO, frozen by pellet method and stored at -196 degrees C for 3 days). A highly motile sperm population was isolated by the swim-up procedure (1 hr). FRE and LIS were incubated an additional 3 hr at 39 degrees C in a Tris-buffered medium to elicit capacitation and the acrosome reaction. Sperm motility and acrosomal integrity were assessed before and after incubation. For the SPA, sperm and eggs were incubated at 39 degrees C for 3 hr in Hams F-10 medium. Each egg was assessed for sperm penetration, sperm binding, and stage of development. Percentages of sperm motility and sperm with a normal apical ridge (NAR) prior to incubation were 78 and 78 (FRE), 75 and 69 (LIS), and 28 and 50 (FRO). After incubation, percentages of motility, NAR, and acrosome-reacted sperm were 34, 10, and 73 (FRE); 43, 24, and 51 (LIS); and 18, 13, and 59 (FRO). A somewhat higher (P less than .05) percentage of ZFH ova was penetrated by FRE (45.8) than by LIS (42.0). Penetration of ZFH ova by FRO was markedly (P less than .05) reduced (30.2). Sperm penetration was not significantly correlated with motility or acrosomal integrity before or after incubation, regardless of treatment. These data suggest that the SPA can be used in conjunction with conventional measures of semen analysis in assessing the potential fertilizing capacity of boar sperm and that liquid storage is superior to frozen storage with respect to preserving sperm fertility.

Acrosome↗
PubMed1987

Flow cytometry of X and Y chromosome-bearing sperm for DNA using an improved preparation method and staining with Hoechst 33342.

A new and improved method of preparing mammalian spermatozoa for high resolution flow cytometric DNA analysis and flow sorting is described. Ejaculated or cryopreserved sperm were briefly sonicated to remove tails and then stained with Hoechst 33342. This simple procedure was found superior to more severe treatments of dimethylsulfoxide washes, fixation in 80% ethanol, and protease digestion of the sperm membranes and tails by papain. Flow cytometric DNA analyses of sperm samples subjected to varying sonication times indicated that X and Y chromosome-bearing sperm populations could be well resolved with as little as 15-sec sonication. In addition, a comparison of sonicated samples stained with four concentrations of bisbenzimide (Hoechst 33342) or 4',6-diamidino-2-phenylindole (DAPI) indicated that 2.5 or 5.0 micrograms/ml of Hoechst was sufficient to resolve the X and Y sperm populations. In order to quantitatively describe the flow cytometric data, several indices (sample quality, orientation and splitting) were developed.

Animals↗
PubMed1987

Incidence of chromosome aberrations in mammalian sperm stained with Hoechst 33342 and UV-laser irradiated during flow sorting.

The separation of two sperm populations is possible using the technique of flow sorting, provided that a significant difference exists in the DNA content of X- and Y-bearing sperm. In order to ascertain whether or not chromosome damage was induced in sorted sperm, chromosome preparations were made from isolated sperm that had been microinjected into hamster eggs. While egg chromosomes exhibited a low frequency of chromosome aberrations, ranging from 4 to 7%, a large proportion of sperm cells exhibited chromosome damage. Between 29% of unstained and unsorted sperm and 38% of stained and unsorted sperm exhibited some type of chromosomal abnormality and this proportion increased to 50% in sorted sperm. If only damaged sperm nuclei are considered, the two unsorted sperm groups had a mean of 0.6 breaks, 0.8 triradial exchanges, and 0.2 quadriradial exchanges per nucleus. However, sorted sperm, which were stained with a fluorochrome and exposed to UV-laser irradiation, exhibited a mean of 2.9 breaks, 2.6 triradial, and 1.9 quadriradial exchanges per nucleus in which damage occurred. These observations indicate that the treatments and manipulations to which sperm nuclei are subjected during flow sorting cause chromosomal aberrations, and that exposure of the cells to UV-laser irradiation contributes substantially to the chromosome damage observed.

Animals↗
PubMed1986

Transmission of chronic idiopathic vitritis in mice by inoculation of human vitreous containing leucocyte phagolysosomal bacteria-like bodies.

Vitreous humour from chronic idiopathic vitritis (CIV) patients containing 0.5-0.7 micron diameter bacteria-like bodies (BLB) in polymorphonuclear leucocytes was inoculated into the eyelids of 100 mice. 200 control mice received either eye-bank vitreous or saline. After 12 months, 53 mice that received CIV vitreous, but none of the controls, had clinical signs of ocular inflammation (p less than 0.05); 15 of the mice that received CIV vitreous and none of the controls had histological evidence of chronic deep ocular inflammation, including vitritis (p less than 0.05); 95 CIV-vitreous-inoculated and 38 control mice were dead (p less than 0.05); and 3/3 of the CIV-vitreous group, compared with 0/3 controls, that were killed for histological assessment had phagolysosomal BLB identical to those in the CIV-vitreous inocula. The findings indicate that the BLB are pathogenic for mice.

Animals↗
PubMed1986

Modification of a laser-based flow cytometer for high-resolution DNA analysis of mammalian spermatozoa.

Modification of a Coulter EPICS V orthogonal laser-based flow cytometer/cell sorter allows resolution of X and Y mammalian sperm populations based on DNA content. The modification consists of beveling the sample injection tube situated in the flow chamber, adding a second fluorescence detector directly forward along the laser beam axis, and routing the collected fluorescence through an optical fiber bundle to one of the existing photomultiplier tubes. The X and Y chromosome-bearing spermatozoa from the bull, boar, and ram can be resolved using this system.

Animals↗
PubMed1986

Carcinoids: the prognostic effect of primary site histologic type variations.

Carcinoids are histologically classified as insular (A), trabecular (B) glandular (C), undifferentiated (D) or mixed. These have prognostic significance, i.e. Group 1 (most favorable, A + C); 2 (favorable, A, B, A + B); 3 (relatively unfavorable, all non A + C or A + B mixed types); and 4 (unfavorable, C, D). Midgut primaries have a better prognosis than either foregut or hindgut/cloacal primaries. Carcinoids from 114 Eastern Cooperative Oncology Group patients were studied to determine if primary site prognostic differences result from histologic prognostic group occurrence rate differences across primary sites. By primary site the following rates were observed: Foregut: 1 (0%), 2 (79.2%), 3 (12.5%), 4 (8.3%); midgut: 1 (26.7%), 2 (58.7%), 3 (6.6%), 4 (8.0%); hindgut/cloaca: 1 (0%), 2 (42.9%), 3 (42.9%), 4 (14.2%); nongut: 1 (0%), 2 (75.0%), 3 (12.5%), 4 (12.5%), p less than 0.01. The results demonstrate that primary site prognostic differences are highly dependent upon histologic prognostic group occurrence rate variations across primary sites. In addition multivariate analysis of survivorship by both histologic type (p less than 0.05) and primary site (p less than 0.05) demonstrated that each variable has independent prognostic significance.

Aged↗
PubMed1986

Gallbladder adenocarcinoma: prognostic significance of tumor acid mucopolysaccharide content.

The gall bladder mucosa is composed of neutral mucopolysaccharide and protein radical containing secretory cells, protein radical containing migratory cells, and neutral and acid mucopolysaccharides plus sialic acid containing goblet cells. The prognostic significance of histologic or histochemical parameters in gall bladder adenocarcinoma (GBA) are unknown. To determine if histochemical acid mucopolysaccharide content in GBA has prognostic value, GBA histologic sections from 26 advanced stage disease patients participating in Eastern Cooperative Oncology Group (ECOG) treatment study EST-2273 were stained with alcian blue at pH. 1.0, assessed by a pathology panel for either high (greater than 50%) or low (less than 50%) acid mucopolysaccharide content, and correlated with patient survival. Initial panel unanimous concurrence rate on acid mucopolysaccharide content was 88.9%. Median survival times from the start of chemotherapy to date of death for high acid mucopolysaccharide content GBA was 14 weeks versus five weeks for the low content GBA (P less than 0.0001). The results indicate that high acid mucopolysaccharide content in GBA significantly improves prognosis. ECOG recommends stratification by acid mucopolysaccharide content in future GBA treatment investigations.

Adenocarcinoma↗
PubMed1986

Purification to homogeneity of the human skin chymotryptic proteinase "chymase".

The chymotrypic proteinase "chymase" has been purified to apparent homogeneity from human skin. Our procedure differs from previously published partial purifications in that it does not involve affinity chromatography, most of the steps are carried out in 2 M KCl which stabilizes the enzyme, detergent is used to protect the enzyme in low-ionic-strength media, and troublesome concentration steps are avoided by using very small columns of high-capacity exchangers. The high-salt skin extract is applied successively to columns of hydroxyapatite, copper chelate Sepharose, and Sephadex G-100 in 2 M KCl. After dialysis against a zwitterionic detergent, the enzyme is adsorbed onto a 0.4-ml column of CM-Sepharose. An alkaline wash removes the remaining contaminants from the highly cationic enzyme, which is then eluted with 1 M KCl in a final volume of 2 ml. Sodium dodecyl sulfate electrophoresis reveals a single diffuse band of Mr 30,000. Recoveries range from 20 to 40% with yields of 0.2 to 0.4 mg of enzyme from 200 g of skin. Specific activities vary from 600 to 1400 units/mg for the hydrolysis of acetyltyrosine ethyl ester.

Chromatography↗
PubMed1986

Protective immunity to Ascaris suum: analysis of swine peripheral blood cell subsets using monoclonal antibodies and flow cytometry.

Outbred domestic swine or SLA inbred miniature swine were exposed to Ascaris suum either naturally on contaminated lots or by inoculation with UV-irradiated attenuated eggs. Both inbred and outbred swine developed virtually complete protection to a challenge of 10 000 eggs after natural exposure, but inbred swine were less resistant than outbred swine after UV-egg exposure. Flow cytometric analysis of peripheral blood mononuclear cells from these animals, performed to determine changes in cell subsets including helper T-cells, cytotoxic/suppressor T-cells, macrophages, and cells expressing class II major histocompatibility antigens, showed that both outbred and inbred swine had similar responses after parasite exposure. The levels of helper T-cells and cytotoxic/suppressor T-cells did not change after parasite exposure, while there was an appreciable but transient increase in macrophages only in those swine naturally exposed to A. suum. Swine exposed to A. suum, both naturally and by inoculation with UV-eggs, showed an increase in the amount of class II antigens detectable per cell. In a second set of experiments, outbred swine were exposed to A. suum naturally or by repeated experimental inoculation with different doses of normal eggs, and protective immunity and changes in blood cell subsets were determined. The greatest change in blood cell subsets was found at 3 and 5 weeks after initial parasite exposure, when macrophages were elevated moderately in a group of pigs inoculated every other day with 1000 eggs and markedly in a group that was naturally exposed; class II antigen expression was also increased during this period. These increases preceded peak serum antibody responses, which were lower in the naturally-exposed group relative to the experimentally-inoculated group. Both groups had high levels of protective immunity. This suggests than natural exposure to A. suum may activate cells and enhance specific immune responses to give high levels of protection.

Animals↗
PubMed1986

Chronic idiopathic vitritis. Ultrastructural properties of bacteria-like bodies within vitreous leukocyte phagolysosomes.

In chronic idiopathic vitritis (CIV) corticosteroid treatment failures, vitrectomy is beneficial. Searching for vitreous microbial agents, 14 vitrectomy specimens from 11 corticosteroid-failing CIV patients were inoculated into numerous in vitro cultural systems; Gram's-, Giemsa-, periodic acid-Schiff- (PAS), and Dieterle-stained centrifuged sediment smears were studied with the light microscope; and the sediment was examined electron microscopically. None of the specimens demonstrated in vitro growth. However, by light microscopy the smears of ten specimens from 8 of the 11 patients demonstrated, in a background of predominantly mononuclear leukocytes, a few polymorphonuclear leukocytes with minute cytoplasmic Gram's variable coccal bodies. By electron microscopy those ten specimens showed morphologically similar 0.5-0.7-micron, thick-walled, coccal-shaped, bacteria-like bodies and 0.03-micron electron-dense spheric particles within polymorphonuclear leukocyte phagolysosomes. The results suggest that CIV vitreous, sterile by contemporary laboratory technics, commonly demonstrates these phagolysosomal bacteria-like bodies. Innovative attempts should be made to cultivate these bacteria-like bodies. Animal pathogenicity studies, using these vitreous specimens as inocula, have been conducted. The results of that investigation will be the subject of another report.

Adolescent↗
PubMed1986

Assessment of spermatozoal function using dual fluorescent staining and flow cytometric analyses.

Spermatozoa from bulls, boars, dogs, horses, mice, and men were examined using a fluorogenic stain consisting of the membrane-permeant substrate carboxyfluorescin diacetate (CFDA) and the relatively membrane-impermeant nuclear stain propidium iodide (PI). Three distinct populations of spermatozoa were discernible in samples from each species upon microscopic examination. Individual spermatozoa, presumed to be viable because of their motility, retained products of the fluorescein chromophore throughout the cell. A second population of spermatozoa in which the nuclei stained red with PI retained the green fluorescein fluorophore mainly in the acrosome. A third population, presumed to be degenerate spermatozoa, possessed only red fluorescent nuclei. These populations were quantified using dual parameter flow cytometry in 14 samples of cryopreserved bovine spermatozoa for which fertility and seminal quality data were available. Flow cytometric analyses were highly correlated with other seminal quality measurements. Sequential flow cytometric analyses provided the ability to rapidly quantitate changes in specific fluorescently stained populations. The ability to make rapid quantitative measurements should allow development of new and presumably more reliable information on the functional aspects of spermatozoa.

Acrosome↗
PubMed1985

Flow cytometric determination of the proportions of X- and Y-chromosome-bearing sperm in samples of purportedly separated bull sperm.

A rapid assay for determining the proportions of X- and Y-chromosome-bearing sperm in semen samples would benefit research aimed at sex ratio control through sperm separation. It also would be of value for quality control should a separation technique be developed. Flow cytometric methods capable of measuring sperm DNA content precisely enough to resolve and quantify the X and Y populations in many mammalian species have been developed. They are effective for fresh and cryopreserved sperm of most domestic animals. Results are reported of flow cytometric analyses of bull sperm samples from seven commercial and academic sources after processing with procedures purported to separate the X and Y populations. In no case was enrichment of either sperm population observed. Breeding trials carried out by the sources of two of the sets of samples showed these procedures were ineffective in altering the sex ratio.

Animals↗
PubMed1984

Primary liver cancer. An Eastern Cooperative Oncology Group Trial.

One hundred ninety-two patients with unresectable primary liver cancer studied by members of the Eastern Cooperative Oncology Group (ECOG) were evaluable in a prospectively randomized clinical trial. Patient discriminants such as performance status were carefully evaluated to assess their influence on prognosis and to evaluate the importance of patient status on response and survival. Patients who were totally bedridden or with signs of overt liver failure were not entered on study. The median survival time for all evaluable previously untreated patients was 17 weeks (19 weeks for North American and European, and 10 weeks for South African black patients). Among the South African patients, however, there was a significantly larger proportion with an initially poor performance status. Prognostic variables (performance status, jaundice, and reduced appetite) dominate any differences among the treatments studied. Among North American and European patients on intravenous (IV) 5-fluorouracil (5-FU) + Methyl-CCNU (MeCCNU) + Adriamycin (ADM, doxorubicin), the 19% response rate is offset by 63% with severe toxicity and a median survival time of only 17 weeks, making this treatment unacceptable clinically. The median survival time of North American and European patients treated with IV 5-FU +/- MeCCNU was 28 weeks in contrast to a median survival time of 12 weeks with ADM (P less than or equal to 0.01). EST 2273 was the ECOG study of patients with primary liver cancer. The results of the first part of the trial were published in 1978. This report updates those findings and reports the results of patients entered subsequently on the second part of that study after it was amended in 1979. With more than 300 evaluable patients in EST 2273, this duet of studies is the largest ever conducted in patients with primary liver cancer, and draws a new baseline from which to measure the disease and its response to treatment.

Adenoma, Bile Duct↗