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Biomedical subjects

K D Bauer

Publications and source records attributed to K D Bauer.

At least 55 records · Page 3Linked to original sources
PubMed1990

Flow cytometric evaluation of epithelial ovarian cancer.

We investigated the prognostic significance of deoxyribonucleic acid content and proliferative activity of tumor cell populations as measured by flow cytometry of the tumor specimens from 115 women with epithelial ovarian cancer. Deoxyribonucleic acid aneuploidy was found in 87 of 115 (76%) of these cancers with a mean deoxyribonucleic acid index of 1.6 and S-phase fraction of 14.7%. The S-phase fraction of the 28 (24%) diploid tumors was 7.0%. Deoxyribonucleic acid ploidy was significantly correlated with survival. S-phase fraction was significantly correlated with ploidy, residual tumor, histology, grade, ascites, time to recurrence, and survival. Diploidy versus aneuploidy were the best discriminating values for deoxyribonucleic acid index and an S-phase fraction of greater or less than 18% for that parameter. Multivariate analysis revealed stage, S-phase fraction, residual tumor, and grade to be independently associated with time to recurrence, and stage, age, S-phase fraction, and largest metastases were factors associated with survival. Deoxyribonucleic acid ploidy did not significantly improve either model. These results suggest that abnormalities of deoxyribonucleic acid content and the proliferative activity of tumor cell populations are reflective of their biologic activity.

Aneuploidy↗
PubMed1990

Quality control in the flow cytometric measurement of T-lymphocyte subsets: the multicenter AIDS cohort study experience. The Multicenter AIDS Cohort Study Group.

Since 1984, the Multicenter AIDS Cohort Study (MACS) has utilized four flow cytometry laboratories to measure T-lymphocyte subset levels semiannually in a large cohort of homosexual men. This report summarizes the steps taken in the MACS laboratories to attain comparability of lymphocyte subset determinations across the centers and over time. Identical flow cytometers, monoclonal antibodies, and analytic procedures have been used, and over a period of time, the procedure for sample preparation was also standardized. Interlaboratory proficiency testing utilizing identical specimens analyzed in the four laboratories was performed to evaluate the comparability of the data among the laboratories. Our results verify that such testing can identify technical bias in flow cytometric evaluations performed at different laboratories. Temporal laboratory consistency in flow cytometric measurements was evaluated using data from each site's HIV-seronegative homosexual reference group. Both sequential 95% confidence intervals (mean +/- 2 x SEM) and the within-person standard deviations of the immune measurements were considered. Significant variation in CD3, CD4, and CD8 lymphocyte subset percentages over time in the seronegative reference population was observed. Our observations indicate that the lymphocyte subset values of this seronegative group should be used to adjust those obtained on the seropositive study participants during a particular time period, thereby allowing improved discrimination of the effects of HIV on T cells in infected individuals. The data presented are of use for designing epidemiologic and intervention studies in HIV-1-infected individuals, especially for calculating sample sizes. The methods we have used to assess the quality of data in the MACS have general application to quality control programs in flow cytometry laboratories. In particular, comparison of sequential confidence intervals and within-person standard deviations for lymphocyte subset determinations on control populations are essential to a comprehensive proficiency testing program because they permit assessment of consistency within a laboratory over time.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Analysis of DNA ploidy and proliferative activity in relation to histology and N-myc amplification in neuroblastoma.

Diploid DNA content, advanced stage, unfavorable histology, and N-myc amplification are all associated with aggressive disease and poor prognosis in childhood neuroblastoma. DNA diploidy is associated with advanced stage and unfavorable histology, but the relationships among ploidy, N-myc amplification, and proliferative activity are not known. To determine if DNA diploidy is associated with N-myc amplification, we studied 29 neuroblastomas with flow cytometric analysis and Southern blot analysis. Clinical and histologic features were also evaluated. Sixty percent of the N-myc-amplified tumors were diploid, compared to 26% of the neuroblastomas, which lacked N-myc amplification (P = 0.11). In our analysis of proliferative activity and N-myc amplification, a higher mean percentage of cells in S phase was seen in the N-myc-amplified tumors (13.4%) than in the unamplified tumors (10%), but again the result was not statistically significant (P = 0.14). Significant associations were seen between unfavorable histology and DNA diploidy (P = 0.05), and between unfavorable histology and high proliferative activity (P = 0.007). Our data suggest that biologic factors other than N-myc amplification play a role in determining the aggressiveness of at least some diploid neuroblastomas.

Blotting, Southern↗
PubMed1989

Limitations in the measurement of c-myc oncoprotein and other nuclear antigens by flow cytometry.

Recent developments in cell fixation, quantitative immunocytochemistry, and flow cytometry allow for the quantification of a variety of oncoproteins and other proliferation-associated antigens in both fresh and archival pathology material. These studies provide evidence that the standard tissue deparaffinization/dissociation technique significantly reduces the amount of c-myc oncoprotein remaining for analysis. To examine the factor(s) responsible for this observation, individual variables of the deparaffinization/dissociation technique including type of fixative, pepsin concentration, pepsinization times, pH, and exposure to organic solvents were examined in HeLa-S3 cells. The cells were stained with monoclonal antibodies either to the c-myc oncoprotein or to p105, a prolifera-tion-associated nuclear antigen. Protein-levels were measured on the basis of anti-c-myc or anti-p105 immunofluorescence by flow cytometry and were found not be affected significantly by type of fixative, exposure to organic solvents, acid pH solution, or mechanical disruption. Levels of c-myc oncoprotein were reduced by over 50%, however, when cells were exposed to 0.5% pepsin, whereas p105 was more resilient with only an approximately 7% reduction following the same treatment. Thus, careful examination of aspects of the deparaffinization/dissociation technique appears to be a necessary prerequisite for quantification of specific nuclear proteins from dissociated tissue specimens.

Antibodies, Monoclonal↗
PubMed1989

Neoplasms of endometrial stroma: histopathologic and flow cytometric analysis with clinical correlation.

Ten neoplasms of endometrial stroma (one stromal nodule, four endolymphatic stromal myoses, and five stromal sarcomas) were compared using clinical data as well as histopathologic and flow cytometric parameters. None of the patients with stromal nodules or endolymphatic stromal myosis had extrauterine disease at presentation or tumors displaying a mitotic rate greater than 10/10 hpf (high-power fields), nuclear pleomorphism, atypical mitotic figures, DNA aneuploidy, or a high proliferative index (greater than 10% S phase cells). The stromal nodules were circumscribed and behaved in a benign fashion. The patients with endolymphatic stromal myosis had infiltrative tumors that behaved as low-grade cancers with good responses to therapy. Four of the five patients with stromal sarcoma had extrauterine disease at presentation as well as tumors characterized by a mitotic rate much greater than 10 mitoses/10 hpf, nuclear pleomorphism, atypical mitotic figures, DNA aneuploidy, and a high proliferative index. These four patients had aggressive disease with poor response to therapy. The fifth of the stromal sarcomas had a high mitotic rate, but lacked the other features linked with aggressive behavior; this patient has responded well to treatment. The mitotic count may not be the most useful criterion for predicting biologic behavior in endometrial stromal tumors since it does not always reflect an increased rate of cell turnover as demonstrated by the percentage of cells in the S phase. DNA analysis by flow cytometry yields a more accurate picture of tumor behavior.

Adult↗
PubMed1989

Development of antibodies to HIV-1 is associated with an increase in circulating CD3+CD4-CD8- lymphocytes.

This study investigated whether seroconversion with respect to human immunodeficiency virus, type 1 (HIV-1) was associated with an increase in lymphocytes expressing the CD3+CD4-CD8- phenotype. Proportions and absolute numbers of CD3+, CD4+, and CD8+ lymphocytes were determined prospectively over a 2.5-year period on 4954 homosexual and/or bisexual men participating in the Multicenter AIDS Cohort Study. Of the 4808 men whose serostatus at entry could be verified, 1745 were seropositive (SP) for antibodies to HIV-1 at entry into study, 2795 were uniformly seronegative (SN) for HIV-1 for 30 months, and 268 were seroconverters (SC) with respect to HIV-1 during this period. For each of six semiannual evaluations, proportions and numbers of CD3+CD4-CD8- lymphocytes (calculated as CD3- (CD4 + CD8] were both significantly greater in the SP group than in the SN group (P less than 0.001). Mean CD3+CD4-CD8- levels in the SC group were indistinguishable from those in the SN group before seroconversion, but by 3-9 months after seroconversion the SC group demonstrated absolute numbers of CD3+CD4-CD8- lymphocytes which were significantly increased (P less than 0.001) compared to the SN group using linear regression methods with adjustment for correlation of measurements within an individual over time. An additional significant increase occurred by 21-27 months after seroconversion (P = 0.006). These results are consistent with an association of HIV-1 seroconversion with an increase in circulating T lymphocytes expressing the CD3+CD4-CD8- phenotype (double negative T cells), a decrease in CD3-CD4-CD8+ natural killer cells, or both. An increase in double negative T cells could reflect a host defense mechanism against HIV-1 or effects of HIV-1 on T cell development.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1989

Cyst-derived cells do not exhibit accelerated growth or features of transformed cells in vitro.

Progressive renal enlargement is a prominent feature in autosomal dominant polycystic kidney disease (ADPKD), suggesting that the disease is due to hyperplasia and/or preneoplastic transformation of renal epithelial cells. In this study in vitro methods were developed to grow and propagate large numbers of cyst-derived epithelial cells from ADPKD kidneys and cortical epithelial cells from normal human kidneys (NK). In order to study their biologic features during early cell passages, cells were grown on Vitrogen (bovine dermal collagen)-FCS (fetal calf serum) coated dishes and fed a basic medium (DME:F12) supplemented with 10% FCS or a defined medium (Sens) containing insulin, transferrin, selenium, hydrocortisone, tri-iodothyronine and epidermal growth factor (EGF). Both ADPKD and NK cells grew as monolayers, were positive for keratin by immunohistochemistry and flow cytometry and had ultrastructural features of renal epithelial cells. Confluent NK and ADPKD monolayers formed domes. In contrast to NK cells, the growth and propagation of ADPKD cells were not supported by defined medium alone but required serum supplementation and ADPKD cells did not respond to growth factors (insulin, transferrin, EGF) that promoted the growth of NK cells. In serum supplemented media, the growth rate, cell doubling time and end cell number of ADPKD and NK cells were the same. Moreover, ADPKD cells did not exhibit any in vitro features of transformed cells: they were not immortal, they were sensitive to contact inhibition, they were anchorage dependent and they were not tumorigenic in nude mice. These findings do not support an increased rate of cell growth or cell transformation as causative factors in ADPKD.

Adult↗
PubMed1989

Tumor-promoting phorbol esters inhibit monocyte adherence to endothelial cells.

Monocyte adherence to endothelial cells (EC) is an important event in the development of a monocytic inflammatory response, yet the effects of inflammatory mediators on monocyte adherence to EC are not well described. We compared the effects of phorbol esters known to activate protein kinase C, including phorbol myristate acetate (PMA) and phorbol 12,13-dibutyrate (PDA), on monocyte adherence to bovine aortic EC. Human monocytes (purity 90 +/- 1% SE) were isolated by centrifugal elutriation to obtain monocytes not previously exposed to a surface. Kinetic studies revealed that 51Cr-labeled monocyte adherence to EC reached a plateau after a 45-min incubation. Concentrations of PMA between 10 and 1,000 ng/ml significantly decreased monocyte adherence to EC (26 +/- 10 and 35 +/- 8% decrease compared with control, respectively). Concentrations of PDA of 100 and 1,000 ng/ml had a similar inhibitory effect. In contrast, the chemotactic stimulus, zymosan-activated serum, significantly increased monocyte adherence (40 +/- 14% increase compared with control). Thus inflammatory stimuli have different effects on the adhesive interaction of monocytes to EC. This may provide a mechanism to selectively modulate monocyte egress from the circulation into extravascular inflammatory sites.

Animals↗
PubMed1989

Contrasting effects of inflammatory stimuli on neutrophil and monocyte adherence to endothelial cells.

Leukocyte adherence to endothelial cells (EC) is an important early event in inflammatory responses, which are often characterized by a predominance of either neutrophils (PMN) or monocytes. However, there is little information concerning the molecular events important in leukocyte adherence to EC. Intracellular activation of protein kinase C and the calcium-second messenger system leads to the stimulation of a number of important functions in PMN and monocytes. We compared the effects of members of these pathways on human PMN and monocyte adherence to cultured bovine aortic EC. We observed that phorbol myristate acetate, phorbol, 12,13-dibutyrate, L-alpha-1-oleoyl-2-acetoyl-sn-3-glycerol, and ionomycin each induced significant dose-dependent increases in PMN adherence to EC monolayers. In contrast, similar concentrations of each of these agents induced significant decreases in EC adherence of monocytes enriched by countercurrent centrifugal elutriation. Separate experiments determined that the differences in PMN and monocyte adherence to EC were not related to differences in oxidant production because 1) phorbol myristate acetate and L-alpha-1-oleoyl-2-acetoyl-sn-3-glycerol caused similar marked increases in both PMN and monocyte superoxide anion and hydrogen peroxide production and 2) ionomycin, which had opposing effects on PMN and monocyte adherence, had no effect on PMN and monocyte superoxide anion or hydrogen peroxide release. We conclude that activators of protein kinase C and the Ca-second messenger pathway have opposite effects on PMN and monocyte adherence to EC and that these effects are mediated by O2 radical-independent mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Endothelial cell subpopulations in vitro: cell volume, cell cycle, and radiosensitivity.

Vascular endothelial cells (EC) are important clinical targets of radiation and other forms of free radical/oxidant stresses. In this study, we found that the extent of endothelial damage may be determined by the different cytotoxic responses of EC subpopulations. The following characteristics of EC subpopulations were examined: 1) cell volume; 2) cell cycle position; and 3) cytotoxic indexes for both acute cell survival and proliferative capacity after irradiation (137Cs, gamma, 0-10 Gy). EC cultured from bovine aortas were separated by centrifugal elutriation into subpopulations of different cell volumes. Through flow cytometry, we found that cell volume was related to the cell cycle phase distribution. The smallest EC were distributed in G1 phase and the larger cells were distributed in either early S, middle S, or late S + G2M phases. Cell cycle phase at the time of irradiation was not associated with acute cell loss. However, distribution in the cell cycle did relate to cell survival based on proliferative capacity (P less than 0.01). The order of increasing radioresistance was cells in G1 (D0 = 110 cGy), early S (135 cGy), middle S (145 cGy), and late S + G2M phases (180 cGy). These findings 1) suggest an age-related response to radiation in a nonmalignant differentiated cell type and 2) demonstrate EC subpopulations in culture.

Animals↗
PubMed1989

Prognostic importance of DNA ploidy in medulloblastoma of childhood.

The deoxyribonucleic acid (DNA) content of 53 medulloblastomas was analyzed by means of flow cytometry and compared with the clinical and histological findings in the host patients. Analysis of DNA showed that about half of the tumors were diploid and the other half were aneuploid. More diploid tumors were found among patients of a young age, but the difference was without statistical significance. Cellular differentiation of the tumor did not correlate with DNA ploidy. No correlation was found between Chang's T staging system and the DNA ploidy, whereas the M staging correlated with the ploidy; diploid medulloblastomas had a greater tendency to metastasize than aneuploid medulloblastomas (p = 0.0003). Four-year survival was compared with the extent of resection and DNA ploidy. The patients with total resection and aneuploid medulloblastoma had a better prognosis than those with subtotal resection and diploid tumor (p = 0.001). There was only one survivor among eight patients with subtotally resected diploid medulloblastomas, while all of the seven patients with totally resected aneuploid medulloblastomas survived. Comparison of the G0/G1 phase fraction and S phase fraction in the surviving group and the deceased group offered no significant information.

Adolescent↗
PubMed1989

Quantification of the c-myc oncoprotein in human glioblastoma cells and tumor tissue.

The identification of a quantifiable oncoprotein marker in glial cells could lead to its use as an aid in the diagnosis, grading, and treatment of tumours of glial origin. In this study, monoclonal antibodies to the c-myc oncoprotein were used in conjunction with immunofluorescence microscopy, flow cytometry, and immunoblot analysis to quantitate and characterize the expression of this oncoprotein in neoplastic and benign cultured glial cells and brain-tumor tissue. Flow cytometric analysis revealed that the c-myc oncoprotein was highly expressed in neoplastic cell lines and in glioblastoma tumor specimens. In contrast, anti-c-myc oncoprotein staining was not present in a non-neoplastic glial cell line or in a benign brain tissue specimen. Immunoblot analysis revealed two distinct c-myc oncoprotein bands, having molecular weights of 64 and 75 kD. Densitometric determinations of the relative levels of the 64-kD protein were in good agreement with the determinations made by flow cytometry. Flow cytometry was also used to relate the quantity of the c-myc oncoprotein present in the cells to their cell cycle phase. In the malignant cultured cells, the protein underwent an approximate twofold increase as the cells progressed from G1/G0 to G2/M in the cell cycle. The present results suggest that the c-myc oncoprotein may prove to be a useful marker for the proliferation status and/or malignancy of glial cells.

Brain Neoplasms↗
PubMed1988

Flow cytometric DNA analysis of medulloblastoma. Prognostic implication of aneuploidy.

Paraffin-embedded surgical specimens from 26 infants and children with medulloblastomas treated between 1972 and 1981 were examined for DNA ploidy by flow cytometry (FCM). All patients received a standard treatment (a combination of maximum debulking of medulloblastoma and postoperative craniospinal irradiation with a posterior fossa boost of 5000 rad or more). They were studied to correlate the results of the findings of FCM DNA analysis with their final outcome, DNA ploidy, and extent of tumor resection. All seven patients with totally resected aneuploid medulloblastoma are alive, whereas only one of six patients with subtotally resected diploid medulloblastoma is alive (P = 0.0047). The current study suggests both DNA ploidy and extent of surgical resection are the most important determinant of patients' prognosis. Patients in selected group, particularly those with subtotally resected diploid tumor, are advised to undergo aggressive adjuvant chemotherapy.

Adolescent↗
PubMed1988

Genetic assessment of the strength of a cancer suppressor gene in hamster cells.

Tetraploid and near-tetraploid chemically transformed derivatives of the pseudodiploid hamster line BHK 21/clone 13 were prepared in four different ways and their ability to be suppressed for anchorage independence by fusion to the anchorage-dependent parental line was tested. In all cases the presence of a single normal genome, thought on genetic grounds to contain a single suppressor gene, was able to prevent the anchorage-independent growth of transformed lines whether they contained one or two complements of pseudodiploid chromosomes. Suppression of the single step in carcinogenesis that is registered by BHK cells as they transform to anchorage independence is thus unusually powerful and apparently independent of chromosome balance and of strict dosage effects.

Animals↗
PubMed1988

Application of multiparameter DNA content and immunofluorescence analysis in the investigation of human neoplasia.

In this discussion a number of IF strategies for possible application to the study of human neoplasia have been reviewed along with factors which can limit their use. Recently, advances in the application of FCM-based DNA content measurements have been shown to be very useful adjuncts in diagnostic pathology and oncology. The years ahead will allow exciting opportunities for gaining further insight into human neoplasia through the direct coupling of quantitative IF analyses with DNA measurements using multiparameter FCM.

Antibodies, Monoclonal↗
PubMed1988

Cell cycle progression in irradiated endothelial cells cultured from bovine aorta.

Logarithmically growing endothelial cells from bovine aortas were exposed to single doses of 0-10 Gy of 60Co gamma rays, and cell cycle phase distribution and progression were examined by flow cytometry and autoradiography. In some experiments, cells were synchronized in the cell cycle with hydroxyurea (1 mM). Cell number in sham-irradiated control cultures doubled in approximately 24 h. Estimated cycle stage times for control cells were 14.4 h for G1 phase, 7.2 h for S phase, and 2.4 h for G2 + M phase. Irradiated cells demonstrated a reduced distribution at the G1/S phase border at 4 h, and an increased distribution in G2 + M phase at 24 h postirradiation. Autoradiographs of irradiated cells after continuous [3H]thymidine labeling indicated a block in G1 phase or at the G1/S-phase border. The duration of the block was dose dependent (2-3 min/cGy). Progression of the endothelial cells through S phase after removal of the hydroxyurea block also was retarded by irradiation, as demonstrated by increased distribution in early S phase and decreased distribution in late S phase. These results indicate that progression of asynchronous cultured bovine aortic endothelial cells through the DNA synthetic cycle is susceptible to radiation inhibition at specific sites in the cycle, resulting in redistribution and partial synchronization of the population. Thus aortic endothelial cells, diploid cells from a normal tissue, resemble many immortal cell types that have been examined in this regard in vitro.

Animals↗