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Biomedical subjects

D C Quinlan

Publications and source records attributed to D C Quinlan.

18 recordsLinked to original sources
PubMed1994

Altered retinoblastoma protein expression and prognosis in early-stage non-small-cell lung carcinoma.

BACKGROUND: Altered retinoblastoma (RB [also known as RB1]) gene expression was initially found in a small cohort study to occur in five (22%) of 23 patients with primary stage I and II non-small-cell lung carcinomas (NSCLCs). Putative mutation of the p53 gene (also known as TP53) has also been found to occur frequently in stage I and II NSCLCs and to be associated with more aggressive disease and a poorer prognosis. PURPOSE: Our purpose was to determine the Rb protein status in the same cohort that had been previously studied for their p53 protein status and to document whether loss of Rb protein expression was also an important factor in overall survival. METHODS: One hundred one stage I or II NSCLC specimens were analyzed by immunohistochemical staining. These paraffin-embedded tumor sections were obtained from individual paraffin blocks prepared for each patient in the previous study. Patient survival status was obtained from hospital and tumor registry records. RESULTS: Altered Rb protein expression was found in 24 of 101 stage I and II NSCLCs. The median survival was 32 months for patients with Rb-positive (Rb+) tumors and 18 months for individuals in whom expression of Rb protein was absent or altered (Rb-) in tumor cells. Log-rank analysis of the differences in overall survival was statistically significant (P = .007). When these results were combined with the p53 status in the same tumor, the median survival was 12 months for those individuals who had theoretically the worst pattern (Rb-/p53+) and 46 months for those patients with theoretically the best pattern (Rb+/p53-) (P < .001). The Rb+ and Rb- groups in this cohort were well balanced with respect to the distribution of age, disease stage, histologic types, p53 status, and sex. Using a multivariate proportional hazards regression model, both altered Rb and p53 status were found to be significantly associated with poor prognosis (P = .005 and .012, respectively) in the overall cohort. CONCLUSION: Altered Rb protein expression is an independent prognostic marker for overall decreased survival in early-stage NSCLC as detected by absence of nuclear Rb protein staining. There appears to be a poorer prognosis when loss of Rb protein function and mutated p53 protein occur in the same tumor. IMPLICATIONS: If these findings can be confirmed in larger prospective studies, the results would suggest that both the Rb and p53 status should be utilized as independent prognostic factors in early-stage NSCLC.

Adenocarcinoma↗
PubMed1994

Potential application of p53 as an intermediate biomarker in Barrett's esophagus.

Diagnosis of the neoplastic progression in Barrett's esophagus using the histologic classification of dysplasia is frequently difficult. The tumor suppressor protein p53, when mutated, confers a promoter effect on cell growth. The purpose of this study was to evaluate the applicability of p53 as an intermediate biomarker of malignancy in Barrett's esophagus. Archival analysis of 100 biopsy specimens of Barrett's esophagus and 10 esophageal adenocarcinomas were compared with 35 chronic esophagitis biopsy specimens. Immunocytochemistry using an anti-p53 monoclonal antibody was performed and elevated immunoreactivity quantitated microscopically. Data were analyzed using a logistic regression model. Significant p53 immunoreactivity occurred as follows: chronic esophagitis (0%), Barrett's esophagus without dysplasia (10%), with low-grade dysplasia (60%), with high-grade dysplasia (100%), and adenocarcinoma (70%). All cases of Barrett's esophagus were significantly immunoreactive when compared with the chronic esophagitis cases (p = 0.001). There was an increase in p53 immunoreactivity as the histologic classification progressed toward adenocarcinoma (p = 0.001). Progression to high-grade dysplasia may be predicted based on p53 immunoreactivity. These findings suggest a role for p53 as an intermediate biomarker in Barrett's esophagus.

Adenocarcinoma↗
PubMed1992

Accumulation of p53 protein correlates with a poor prognosis in human lung cancer.

Mutations in the gene coding for the p53 tumor suppressor protein are common in a variety of human cancers. To assess the role of a putative mutated p53 protein in human lung cancer, a monoclonal antibody recognizing it was used in an immunoperoxidase detection system. A total of 114 cases of Stage I and II adenocarcinomas and squamous cell carcinomas were studied. The staining pattern was always intranuclear and heterogeneous. When the median or mean survival time was compared between cases, p53 accumulation had a statistically significant negative prognostic value. This was supported by a Kaplan-Meier survival plot of p53 producers and nonproducers. In 7 of 24 Stage II cases that were negative for p53 in the primary tumor, metastatic regional lymph nodes were p53-positive. These latter cases had greatly reduced survival times. Thus, p53 accumulation in primary tumors (and regional lymph nodes) may identify a subgroup of lung cancer patients with a prognosis of more aggressive disease.

Adenocarcinoma↗
PubMed1987

Dynamics of glycoprotein metabolism in familial polyposis coli-derived colonic epithelial tissue.

Familial polyposis coli is a disease state that offers an opportunity to obtain colon tissue in a predictable manner in terms of identifying putative premalignant markers. Our results support the hypothesis that a sequential series of alterations occur in the dynamics of glycoprotein metabolism as normal colonic mucosa progresses to colon adenocarcinoma. Specifically, both nonmalignant polyps and colon adenocarcinoma tissue expressed diminished glycosylation of proteins, whereas an elevated level of activity for membrane-associated glycosyltransferases does not appear until colon cancer itself develops.

Adenocarcinoma↗
PubMed1983

Cell surface and cytoskeletal interactions in a temperature-sensitive strain of SV40-transformed mouse fibroblasts.

In order to assess the role of cytoskeletal structure in modulating cell surface topography during cell transformation, cytoskeletal organization of 3T3 mouse cells transformed with a tsA mutant of simian virus 40 (SV40) was studied in detail by correlative light and electron microscopy. Detergent-extracted, critical-point dried whole cells observed in the electron microscope were seen to contain well-organized microfilament bundles (stress fibers) traversing the longitudinal axis of cells grown at the restrictive temperature (39 degrees C). When grown at the permissive temperature (32 degrees C), cells prepared in this manner were not observed to contain such structures. However, when semithin sections (0.5 micron) were viewed by transmission electron microscopy at 120 kV, short microfilament bundles were seen in 32 degrees C-grown cells. There was an alteration in the morphology of these structures at sites of attachment to the substratum (focal contacts), and they were shorter in length than microfilament bundles of 39 degrees C-grown cells. A difference was also observed between the two phenotypes in the layer of microfilaments associated with the dorsal cell surface. Since it is this layer that directly determines cell surface architecture, it is proposed that changes in microfilament bundle-generated surface tension are responsible for alterations of this layer, leading to an altered cell surface morphology. Tension may be modified by disturbances in focal contacts (or adjacent regions) or altered actin-associated protein(s).

Animals↗
PubMed1982

Sodium-gradient-stimulated transport of L-alanine by plasma-membrane vesicles isolated from liver parenchymal cells of fed and starved rats. Crucial role of the adrenal glucocorticoids.

The ability of liver efficiently to take up amino acids, particularly l-alanine, during starvation was studied in a cell-free system by isolating plasma-membrane vesicles in a transport-competent state from rat liver parenchymal cells. These membrane vesicles have the capacity to accumulate l-alanine against an apparent concentration gradient when exposed to an artificial and transient transmembrane Na(+) gradient (extravesicular Na(+) concentration greater than inside). The rate of accumulation of l-alanine is dependent on the plasma-membrane vesicle concentration, and the steady-state concentration attained is inversely related to the osmolarity of the medium. The Na(+)-mediated stimulation is not exhibited if the membrane vesicles are pre-equilibrated with NaCl, if K(+) or Li(+) are substituted for Na(+), or if SO(4) (2-) replaces Cl(-) as the counterion. The apparent active transport of l-alanine into the membrane vesicles appears to occur by an electrogenic mechanism: (1) the use of NaSCN significantly heightens the early concentrative phase of transport when compared with the effect of NaCl; (2) an enhanced active transport is also observed when a valinomycin-induced K(+) efflux occurs concomitant with Na(+) and l-alanine influx. Plasma-membrane vesicles isolated from liver parenchymal cells of a 24 h-starved rat exhibit an initial l-alanine transport rate that is 3-4 times that for membrane vesicles derived from a fed animal. The increased rate of l-alanine transport by plasma-membrane vesicles from starved animals can be obliterated by adrenalectomy and restored by administration of glucocorticoid. These results establish that stimulation of the gluconeogenic pathway by starvation involves a plasma-membrane-localized change affecting l-alanine transport which is regulated in part by the glucocorticoid hormones.

Adrenalectomy↗
PubMed1981

Hexose sugar transport activity in a mouse fibroblast cell line temperature sensitive for expression of the transformed phenotype.

A temperature-sensitive mouse fibroblast cell line was used to examine the relationship between hexose sugar uptake rates and the control of cell growth. The cell line (ts-H6-15) is a derivative of SV-3T3 cells, exhibiting a transformed phenotype at 32 degrees C and a normal phenotype at 39 degrees C. For cells actively growing at either temperature, a marked decrease in the rate of 3-O-methyl-D-glucose (3-O-MeG) transport is observed as cell population density increases. At cell population densities tested, 3-O-MeG transport rates (at a common assay temperature) were greater in H6-15 cells grown at 32 degrees C than at 39 degrees C, with the enhancement being maximal at the lowest cell densities. The effect of low serum-arrest on H6-15 cells revealed that cells growing at 39 degrees C arrest in G1, while cells at 32 degrees C stop more randomly throughout their cycle. Under conditions of low serum-arrest the rate of 3-O-MeG transport remained as high as in actively growing cells at both 32 degrees C and 39 degrees C. However, 2-deoxyglucose uptake rates were growth state-dependent at 39 degrees C, indicating perhaps metabolic as well as membrane-level control of sugar accumulation. These results further demonstrate that rates of hexose sugar transport by themselves are not always absolutely correlated with rates of cell proliferation and, thus, may be reliable predictors of cell growth potential.

Animals↗
PubMed1981

Morphology of temperature-sensitive strains of viral-transformed 3T3 mouse fibroblasts in vitro as determined by SEM and STEM.

The morphological characteristics of BALB/c 3T3 mouse fibroblasts transformed with two temperature-sensitive strains of simian virus 40 (SV-40) were studied. When grown at the permissive temperature (32 degrees C, transformed phenotype) strains A7B4b and A209B4a both closely resemble the morphology described for wild-type SV-40 cells. Scanning electron images show numerous knobs and filopodia as characteristic surface features in both subconfluent and confluent cultures. Scanning-transmission electron images reveal that the two strains are also similar in intracellular structure. When grown at the restrictive temperature (39 degrees C, untransformed phenotype) the two strains differ from each other. The A209B4a strain is more elongated in subconfluent cultures, and shows interactions of filopodia at cell-cell interfaces when confluent similar to those described for wild-type 3T3 cells. This strain also shows intracellular accumulation of polymorphic material thought to be secondary lysosomes. The A7B4b cells are flattened but generally not elongated when subconfluent, and do not show any obvious interactions at confluency. Intracellularly, this strain also differs from A209B4a.

Animals↗
PubMed1977

The regulation by fibroblast growth factor of early transport changes in quiescent 3T3 cells.

This study involves the use of fibroblast growth factor (FGF) as a substitute for exogenous serum to examine the early transport changes which occur when quiescent 3T3 cells re-initiate active growth. FGF, in nanogram amounts, together with insulin and dexamethasone, can induce mitogenesis and mitosis in 3T3 cells GO-arrested by holding in growth medium containing 0.8% calf serum. In terms of quiescent cell transport activity enhancement, FGF is 300,000-fold more effective than fresh serum, on a protein basis. In addition, very short exposure of serum-depleted cells to FGF indicates that a distinct temporal or time sequence exists in the transport system activation process. For example, uptake of alpha-aminoisobutyric acid (AIB) and uridine are stimulated very rapidly, whereas hypoxanthine uptake does not respond until much later. Closer analysis shows that AIB uptake is maximally enhanced within zero to two minutes after FGF addition to cells. Finally, the stimulatory effect of FGF on transport system activities is specific in terms of the proliferative state of the cells to which it is added, and in terms of the uptake systems which respond to it.

Adenosine↗
PubMed1976

Group translocation of the ribose moiety of inosine by vesicles of plasma membrane from T(3 cells transformed by Simian virus 40.

Plasma membrane vesicles are isolated from Simian virus 40-transformed Balb/c mouse 3T3 (SV-3T3) cells. These membrane vesicles contain no significant contamination by mitochondria, endoplasmic reticulum, or lysosomes as determined by marker enzyme analysis. The use of [U-14C] inosine as a transport substrate results in the accumulation of labeled ribose-1P as transport product by the plasma membrane vesicles. This suggests the action of purine nucleoside phosphorylase (the enzyme which mediates the phosphorolysis of inosine to ribose-1-P and hypoxanthine0 before, during, or after the transport step. Neither inosine nor significant amounts of hypoxanthine are found intravesicularly. The Km for inosine, the substrate in this reaction which leads to the accumulation of ribose-1-P by the plasma membrane vesicles, is 35 to 45 muM while the Vmax for ribose-1-P accumulation is 100 to 120 pmol/min/mg of plasma membrane protein...

Binding, Competitive↗
PubMed1976

The existance of a group translocation transport mechanism in animal cells: uptake of the ribose moiety of inosine.

After exposure to inosine, transport-competent plasma membrane vesicles isolated from SV-40-transformed Bal/c 3T3 cells accumulate intravesicular ribose 1-PO4 at a concentration 200-fold greater than the extravesicular concentration. An analysis of the purine nucleoside phosphorylase activity distribution in various subcellular fractions, relative to other enzyme activities, indicated the presence of plasma membrane-associated purine nucleoside phosphorylase activity. The plasma membrane vesicles appear relatively impermeable to hypoxanthine. However, hypoxanthine, which is a competitive inhibitor of the transport reaction, is the only compound tested capable of mediating efflux of already accumulated ribose 1-PO4. In addition, hypoxanthine does not result in the efflux of transported uridine which is accumulated in these membrane vesicles as uridine. Exogenous ribose 1-PO4 neither results in counterflow nor does it inhibit the original uptake reaction. The following transport reaction is proposed: uptake occurs by group translocation, mediated by membrane-localized purine nucleoside phosphorylase. The data are consistent with sites for inosine and hypoxanthine being on the outer membrane surface whereas the ribose 1-PO4 site is only on the inner surface.

Adenine↗
PubMed1976

Sodium-stimulated amino acid uptake into isolated membrane vesicles from Balb/c 3T3 cells transformed by simian virus 40.

Mediated uptake of amino acids by membrane vesicles isolated from Balb/c 3T3 cells transformed by simian virus 40 has been demonstrated. Initial rates of transport of radioactively labeled L-leucine and alpha-aminoisobutyric acid were enhanced by the addition of NaCl (100 mM) to the reaction mixture at the start of the uptake process. This enhancement included a prominent "overshoot" during initial uptake. Slight stimulation of alpha-aminoisobutyric acid uptake was seen with K+, but none with Li+. The mediated nature of the uptake event for L-leucine was shown by saturation kinetics and by inhibition with L-valine. The transport assay measured predominantly intravesicular amino acid uptake rather than binding, as shown by the variation of uptake in response to changes in extravesicular osmolarity. Electron microscopy confirmed the presence of closed vesicles. Thus, amino acid transport has been characterized in an in vitro membrane vesicles system which should prove useful for studies of growth control.

Amino Acids↗
PubMed1974

An altered rate of uridine transport in membrane vesicles isolated from growing and quiescent mouse 3T3 fibroblast cells.

Balb/c 3T3 and simian virus-40-transformed Balb/c 3T3 (SV-3T3) cells were examined for their ability to transport nucleosides. Confluent (quiescent) 3T3 cells transported uridine at a rate 3-4 fold lower than did subconfluent cells. Adenosine uptake was independent of cell population density. Both adenosine and uridine were transported at the same rate by confluent and sub-confluent SV-3T3 cells. A membrane vesicle population (plasma membrane and endoplasmic reticulum) was isolated from 3T3 and SV-3T3 cells by nitrogen cavitation. With membrane vesicles derived from SV-3T3 cells it was determined that uptake occurred by a mediated process. Uptake of adenosine and uridine by membrane vesicles isolated from 3T3 and SV-3T3 cells demonstrated the same pattern as found in whole cells; that is, membrane vesicles from confluent 3T3 cells transported uridine at a rate 3-fold lower than did membrane vesicles from subconfluent 3T3 cells. Much of the adenosine taken up was converted to inosine, hypoxanthine, and ribose 1-phosphate, whereas uridine transport resulted only in the accumulation of uridine. Results obtained with membrane vesicles indicate that the lowered rate of uridine transport by confluent 3T3 cells seems due to an alteration in the membrane itself or a component(s) thereof, rather than to changes in subsequent cellular metabolic processes.

Adenosine↗
PubMed1974

Partial purification of a membrane-associated deoxyribonucleic acid complex from Mycoplasma gallisepticum.

A Mycoplasma gallisepticum subcellular fraction (P2), which contains the deoxyribonucleic acid replication complex, can be isolated by differential centrifugation of freeze-thaw-lysed cells. The nascent deoxyribonucleic acid is released from P2 by Lubrol-WX, sodium dodecyl sulfate, Pronase, and deoxyribonuclease, but not by saponin, ribonuclease, phospholipase C, or high-frequency sonic treatment. Sonic treatment further fractionates the cell ghost and allows partial purification, on sucrose density gradients, of a deoxyribonucleic acid replication complex attached to the cells' polar membrane-bleb-infrableb structures.

Carbon Radioisotopes↗
PubMed1973

Deoxyribonucleic acid synthesis in synchronously growing Mycoplasma gallisepticum.

Early log-phase cells of Mycoplasma gallisepticum A5969 were synchronized by holding in Eagle minimal essential medium (MEM) for 2 h. When transferred out of MEM into tryptose medium, the cells exhibited synchronous growth. Deoxyribonucleic acid (DNA) synthesis proceeded continuously during this growth but stopped during the period of cell division. One round of DNA replication was observed per cell doubling, and a unique region of DNA was found to be permanently bound to the membrane.

Cell Division↗
PubMed1972

Membrane association of the deoxyribonucleic acid growing-point region in Mycoplasma gallisepticum.

Newly replicated deoxyribonucleic acid (DNA) in Mycoplasma gallisepticum A5969 is membrane associated. Cells pulse-labeled 1 to 3 min with (3)H-thymidine are lysed by a freeze-thaw procedure. After brief sonic treatment to shear the DNA, differential centrifugation gives a cell fraction (P2) that is enriched sevenfold for pulse-labeled DNA. P2 contains 80% of the total adenosine triphosphatase activity, 65% of the total cholesterol, and morphologically intact terminal bleb structures. Three to four minutes are needed to fully label the DNA growing-point region, whereas 7 to 8 min are required to "chase" 50% of the (3)H-labeled DNA. This pulse-chase removes 80 to 85% of the nascent DNA from the P2 fraction.

Adenosine Triphosphatases↗