PubMed Health⌕ Search

Biomedical subjects

J M Connolly

Publications and source records attributed to J M Connolly.

At least 73 records · Page 4Linked to original sources
PubMed1991

High-fiber diet reduces serum estrogen concentrations in premenopausal women.

The effect of dietary fiber on serum estrogen concentrations was studied in 62 premenopausal women. Their estimated daily dietary fiber intake was increased from an average of approximately 15 g to 30 g/d by supplementation with wheat, oat, or corn bran. This was done without any significant decrease in dietary fat consumption. After 2 mo on the high-fiber diets, the wheat-bran-supplemented group showed significant reductions in serum estrone (P less than 0.002) and estradiol (P less than 0.02) but no change in serum progesterone or sex-hormone-binding globulin concentrations. The serum estrogens were unaffected by dietary-fiber supplementation with oat or corn brans.

Adult↗
PubMed1990

Effects of fatty acids and inhibitors of eicosanoid synthesis on the growth of a human breast cancer cell line in culture.

Dietary lipids may influence breast cancer progression and prognosis. The MDA-MB-231 human breast cancer cell line was used to examine the direct effects of the various classes of free fatty acids (FAs) on growth in serum-free medium and the involvement of eicosanoid biosynthesis. Linoleic acid, an omega 6 FA, stimulated MDA-MB-231 cell growth with an optimal effect at a concentration of 0.75 microgram/ml, whereas oleic acid, an omega 9 FA, produced growth stimulation at 0.25 microgram/ml but was inhibitory at higher concentrations. Docosahexaenoic acid exhibited a dose-related inhibition of cell growth at concentrations ranging from 0.5 to 2.5 micrograms/ml; eicosapentaenoic acid, also an omega 3 FA, was less effective. Similar inhibitory effects occurred with saturated FAs. Indomethacin, which at high concentrations is an inhibitor of both the cyclooxygenase- and lipoxygenase-catalyzed pathways of eicosanoid synthesis, suppressed cell growth stimulation by an otherwise optimal dose of linoleic acid when present at 40 micrograms/ml. Experiments with piroxicam, nordihydroguaiaretic acid, and esculetin, other inhibitors of eicosanoid biosynthesis with varying selectivity for enzymes of the prostaglandin and leukotriene pathways, indicated that MDA-MB-231 cell growth was dependent on leukotriene rather than prostaglandin production.

Breast Neoplasms↗
PubMed1990

The polypeptide of immunoglobulin G influences its galactosylation in vivo.

To examine the nature of the factors influencing the galactosylation pattern of the heavy chain of murine immunoglobulin G (IgG), cell fusion was performed between a myeloma (P3x63Ag8) and a hybridoma (Sp2HL/Bu) cell line which secrete different IgGs possessing structurally distinct CH2-linked oligosaccharide moieties. The glycosylation patterns of the IgGs of the parental and fused cells were studied. Pronase digestion of the purified heavy chains and subsequent end labeling with fluorescein isothiocyanate produced fluoresceinated glycopeptides which were detected and purified by polyacrylamide gel electrophoresis. Structural information was obtained by enzymatic digestion, lectin affinity chromatography, and methylation analysis. IgGs from both parental lines possessed oligosaccharide units displaying microheterogeneity based upon a common symmetrical biantennary structure terminating in beta-GlcNAc. The structures of both IgGs, however, differed in the pattern of the mono- and digalactosylated components. Clones, selected following the fusion of the parental cells, were expanded; and the individual IgGs were purified. All clones produced homodimeric IgG1 and IgG2b as well as heterodimeric IgG possessing both the gamma 1 and gamma 2b heavy chains. Analysis of the carbohydrate moieties of the gamma 1 chain from the homodimeric and heterodimeric IgGs and of the gamma 2b chain from the heterodimeric molecule demonstrates that the polypeptide structure of the heavy chain influences the terminal galactosylation of the glycan unit at the conserved site of glycosylation of IgGs.

Acetylglucosaminidase↗
PubMed1990

Peptide ligand-induced conformation and surface expression of the Ld class I MHC molecule.

Newly synthesized major histocompatibility complex (MHC) class I molecules in the endoplasmic reticulum are thought to bind peptides of foreign and endogenous antigens. Several lines of evidence indicate that beta-2 microglobulin (beta 2m) and/or peptide ligand participate in the intracellular transport and surface expression of class I molecules, but the nature of their involvement is still unclear. Here we present evidence that culturing non-mutant cells (fibroblast, thymoma or mastocytoma) with a peptide ligand specific for the Ld class I molecule of the mouse leads to a dramatic (fourfold) and specific induction of Ld surface expression. Surprisingly, this peptide ligand-induced expression of Ld does not result in an increased intracellular association of Ld with beta 2m. These findings demonstrate that the previously reported decrease in surface expression of Ld results from its failure to be saturated with endogenous self-peptide ligands. This unique feature of Ld could also contribute to the fact that several virus-specific cytotoxic T cell responses have been found to be Ld-restricted.

Animals↗
PubMed1990

Production of epidermal growth factor and transforming growth factor-alpha by the androgen-responsive LNCaP human prostate cancer cell line.

Epidermal growth factor (EGF)-related polypeptides may be involved in the growth of human prostate cancer cells, and in the androgen stimulation of hormone-responsive prostatic carcinomas. We have shown that androgen-responsive LNCaP cells, like the autonomous DU 145 human prostate cancer cell line, synthesize and secrete EGF and related polypeptides, including immunoreactive transforming growth factor-alpha (TGF-alpha). As determined by radioimmunoassay, intracellular EGF levels were approximately 100 times those of TGF-alpha, but together these accounted for less than half of the total EGF-like polypeptides detected in a radioreceptor assay. Although LNCaP cell growth was stimulated by dihydrotestosterone (DHT), there was no evident effect on immunoreactive EGF levels in the medium after correction for cell number. Moreover, metabolic labeling experiments showed no effect of the androgen on EGF synthesis by LNCaP cells. Gel filtration chromatography of conditioned medium showed both high molecular weight species and the mature 6,000 dalton form of immunoreactive EGF. We conclude that although LNCaP prostate cancer cell growth is stimulated by DHT, it is unlikely that it is mediated directly via increased EGF synthesis by the tumor cells.

Androgens↗
PubMed1990

Dietary prevention of breast cancer.

A review of the epidemiological and experimental data suggests that dietary modification does have a place in breast cancer prevention. This needs to be considered at three levels: dietary recommendations for the general population, dietary intervention in the management of women at high risk for breast cancer, and the place for dietary therapy as a component of regimens intended to reduce risk of recurrence after primary treatment of breast cancer ('secondary prevention'). Based on present evidence, a dietary approach to breast cancer prevention should include weight control, when indicated, a reduction in dietary fat intake to approximately 20% of total fats, and an increase in fiber consumption to 25-30 g day-1. Further studies are indicated to determine the optimal qualitative composition of dietary fat consumed, and the nature of supplemental fiber.

Body Weight↗
PubMed1990

Recognition by CD8 on cytotoxic T lymphocytes is ablated by several substitutions in the class I alpha 3 domain: CD8 and the T-cell receptor recognize the same class I molecule.

The CD8 molecule on class I-reactive cytotoxic T lymphocytes (CTLs) is believed to function as a coreceptor along with the alpha beta T-cell receptor. Whereas the alpha beta T-cell receptor recognizes polymorphic residues in the alpha 1/alpha 2 domains of the class I molecule, the CD8 molecule is believed to recognize monomorphic class I residues. Our previous experiments suggested that residue 227 in the alpha 3 domain of major histocompatibility complex class I molecules contributes to the determinant recognized by CD8. By using a panel of site-directed mutants of H-2Dd, this observation has been extended herein. Our findings indicate that for recognition by CD8-dependent CTLs, residue 227 must be either glutamic acid or aspartic acid and cannot be either basic or uncharged. However, the recognition by CD8-independent CTLs is unaffected by any of the substitutions at position 227 of H-2Dd. Similarly, alterations of other charged residues at positions 222, 223, and 229 have an analogous effect to substitution at residue 227, whereas substitutions at residues 192 and 232 do not affect the reactivity of CD8-dependent or CD8-independent CTLs. In addition, mutant H-2Dd molecules that are not recognized by CD8-dependent CTLs are unable to stimulate a primary CTL response, yet they can stimulate a secondary CD8-independent H-2Dd-specific CTL response. These findings suggest that CD8 recognition is obligatory for the priming of class I-dependent CTL responses. Since endogenous class I molecules were expressed by all of the transfected cell lines, these findings provide direct genetic evidence that CD8 and the alpha beta T-cell receptor must interact with the same class I molecule.

Amino Acids↗
PubMed1989

Stimulation of growth of human breast cancer cell lines in culture by linoleic acid.

Linoleic acid, an omega-6 unsaturated fatty acid, stimulated growth of the MDA-MB-231 and MCF-7 human breast cancer cell lines in culture. Responses of the estrogen-independent MDA-MB-231 cells both in serum-free medium and with 1% fetal bovine serum added were positively correlated with linoleic acid concentration over the entire range examined (5-750 ng/ml). Growth stimulation of the estrogen-responsive MCF-7 cell line was maximal at a LA concentration of 500 ng/ml when cultured in 1% fetal bovine serum-containing medium with added estradiol. Linoleic acid had no mitogenic effect on three human cancer cell lines derived from sites other than breast, or on untransformed 3T3 cells.

Breast Neoplasms↗
PubMed1989

Locus-specific regulation of Kd, Dd, and Ld class I genes in the BALB/c S49 lymphoma sublines.

The S49 tumor sublines are variants isolated from a single parent BALB/c tumor which demonstrate locus-specific shut-off of their Kd, Dd, and/or Ld surface molecules. Four phenotypically different sublines were characterized: wild-type (Kd Dd Ld)+, a null phenotype (Kd Dd Ld)-, and two intermediates (Kd Dd)+ (Ld)- and (Kd)+ (Dd, Ld)-. Nonexpressed class I Ag were not induced with IFN-gamma even though the expressed antigens were found to be appropriately responsive. Southern blot analysis using probes specific for the 5', exonic or 3' portions of the Kd, Dd, and Ld genes indicated that these S49 sublines have no major chromosomal aberrancies. Northern blot analysis of RNA from each S49 subline using locus-specific oligonucleotide probes revealed message only for the expressed Ag. Thus, the locus-specific shut-off of class I expression in these tumors appears to be acting at the transcriptional level. RNase protection analysis confirmed this result and furthermore demonstrated that the repression is exquisitely specific for the Kd, Dd, and Ld genes as other "class I-like" messages were detected in each cell line. These findings are discussed in the context of the various cis- and trans-acting mechanisms that have been proposed to regulate class I expression.

Animals↗
PubMed1989

Secretion of epidermal growth factor and related polypeptides by the DU 145 human prostate cancer cell line.

Epidermal growth factor (EGF)-like proteins may be important in regulating the growth of human prostate cancer cells and in the development of hormone independence. In the present study, we demonstrated that the DU 145 human prostate cancer cell line secretes EGF-like polypeptides into serum-free culture medium. The production of both authentic EGF and transforming growth factor-alpha was demonstrated by specific radioimmunoassays. In addition to 6-7k monomers, immunoreactive higher molecular weight species were isolated by gel chromatography. The DU 145 cells also specifically bound human EGF, with both high- (Kd 1.8 x 10(-10) M) and low- (Kd 1.1 x 10(-9) M) affinity binding sites. Exogenous EGF stimulated 3H-thymidine incorporation when cells were plated at low density in serum-free culture medium, while at higher cell density neither cell division nor 3H-thymidine incorporation was significantly altered. We postulate that DU 145 cells may be autologously stimulated by endogenously produced EGF and related polypeptides, which, under normal culture conditions, precludes any further effect of exogenous EGF.

Cell Division↗
PubMed1988

The Lyt-2 molecule recognizes residues in the class I alpha 3 domain in allogeneic cytotoxic T cell responses.

The involvement of the different domains of the MHC class I molecule in CTL recognition was investigated. mAbs specific for the alpha 1/alpha 2 domains of H-2Ld interfered with both the primary and secondary generation and effector function of in vitro Ld-specific CTL. mAbs specific for the alpha 3 domain of H-2Ld interfered with the generation and function of primary in vitro Ld-specific CTL; however, there was no effect on the in vitro generation of secondary CTL and only partial inhibition of their function. In vivo treatment with graft-specific antibodies to both the alpha 3 domain and the alpha 1/alpha 2 domains together resulted in a dramatic enhancement of Ld- or Dd-disparate skin grafts, whereas the individual mAbs showed minimal effects. This suggested that the class I alpha 3 domain is recognized by alloreactive CTL. Several approaches were undertaken to examine whether recognition of the alpha 3 domain determinants is mediated by the Lyt-2 molecule. When mAbs specific for the alpha 3 domain of either H-2Ld or H-2Dd were used in vivo and in vitro, the resulting CTL population was not inhibited by antibody to the alpha 3 domain and was only partially inhibited by antibody to Lyt-2. We therefore observed a correlation between the effects of antibody to the class I alpha 3 domain of the target molecule and antibody to the Lyt-2 molecule on the CTL. To further test the relationship between CTL recognition of the alpha 3 domain and the involvement of Lyt-2, we used a cell expressing a mutation in the alpha 3 domain of the Dd molecule. The mutation resulted in a single amino acid substitution of glu to lys at residue 227 of the alpha 3 domain. Consistent with an earlier report, cells expressing the mutant Dd lys molecule were not lysed by CTL from a primary stimulation against the wild-type Dd glu molecule. However, this same cell line was killed by the Lyt-2-independent secondary Dd-specific CTL generated in the presence of antibody to the alpha 3 domain in vivo and in vitro. Furthermore, cells expressing the mutant Dd lys molecule failed to stimulate a primary response. In conclusion, several independent lines of evidence indicate that residues in the alpha 3 domain of the class I molecule are involved in recognition by the Lyt-2 molecule, and that Lyt-2-mediated recognition can be specifically blocked using mAb to determinants in the alpha 3 domain.

Animals↗
PubMed1988

Epidermal growth factor-like proteins in breast fluid and human milk.

Epidermal growth factor (EGF), and the transforming growth factor-alpha (TGF-alpha) family of proteins, which also bind to the EGF receptor, have been associated with human breast cancer. The total EGF-like proteins were determined by a radioreceptor assay, and TGF-alpha by radioimmunoassay, in human milk and breast fluid samples. The breast fluids were collected by nipple aspiration from healthy premenopausal women. Both the 24 milks and 18 breast fluids assayed contained EGF-like proteins, at concentrations ranging from 32-600 ng/ml (median, 140 ng/ml), and 62-654 ng/ml (median, 205 ng/ml) respectively. Immunoreactive TGF-alpha proteins were detected at higher levels in 21 breast fluids (range, 0-50.0; median 5.1 ng/ml) than in 24 milk samples (range, 0-8.4; median, 0.8 ng/ml).

Adult↗
PubMed1987

Quantitative differences in the effects of mouse and human epidermal growth factors on A431 human tumor cells.

Human and mouse epidermal growth factor (hEGF; mEGF) bind to the same two classes of receptor sites on cell membranes prepared from the human epidermoid carcinoma cell line A431. However, the affinities of mEGF for the low affinity receptors (K(d) 3.5 X 10-9 M) and high affinity receptors (K(d) 2.7 X 10-10 M) are lower than those of hEGF (K(d) 1.2 X 10-9 M and 1.9 X 10-10 M respectively). In consequence, artefactually high results are obtained when mEGF is used for radioreceptor assays of EGF-like proteins in human biological fluids. Both species of EGF stimulated A431 cell growth at low concentrations and inhibited at high concentrations; however, hEGF enhanced cell growth over a wider concentration range (up to 1.6 nM), and had a higher maximal stimulating effect (38% vs. 25%).

Animals↗
PubMed1986

Structure and expression of Fc gamma receptors on mouse suppressor T cell hybridomas.

Antigen-specific and idiotype-specific mouse suppressor T cell hybridomas were analyzed for the presence and specificity of Fc gamma receptors (Fc gamma R) by EA rosetting and by flow microfluorometry with the use of monoclonal antibodies. We found that four hybridomas expressed Fc gamma R specific for IgG1 and IgG2b, one of which became Fc gamma R- during prolonged culture. Four other hybridomas and the fusion parent, BW5147, consistently lacked Fc gamma R. The 125I-labeled Fc gamma R were isolated from surface radioiodinated hybridoma cells solubilized with 1% Nonidet P-40, were purified by using single or repetitive chromatography on mouse IgG-Sepharose columns, and were analyzed by SDS-PAGE. An 125I-labeled 56,000 to 61,000 Mr macromolecule was isolated from each of the Fc gamma R+ hybridomas, but from none of the Fc gamma R- hybridomas nor from BW5147 cells. This macromolecule rebound to insolubilized mouse IgG1, IgG2b, and human Fc fragments, but not to insolubilized mouse IgG2a, IgG3, or IgA or human F(ab')2 fragments, consistent with the specificity observed for Fc gamma R on intact hybridoma cells. The mouse suppressor T cell Fc gamma R differs in size and specificity from mouse B cell Fc gamma R. A 70,000 Mr protein expressed on all hybridomas and on BW5147 cells was radiolabeled and, despite preclearing with ovalbumin-Sepharose, bound to the mouse IgG-Sepharose columns, presumably due to mouse antibodies to gp-70. This macromolecule was completely and specifically removed by using goat antiserum to gp-70.

Animals↗
PubMed1981

Fc (IgG) receptor distributions in homogeneous and heterogeneous cell populations by flow microfluorometry.

A flow microfluorometric method has been developed for quantitating the numbers of Fc receptors on individual cells. The cells were equilibrated at 0 degrees C with radiolabeled, affinity-crosslinked rabbit IgG dimers, washed, and treated with fluorescent antibodies against rabbit IgG. The stained cells were analyzed for fluorescence emission by using a fluorescence-activated cell sorter and for bound dimer molecules by using a gamma counter. Standard curves relating fluorescence emission to numbers of dimer molecules bound to cells were used to determine Fc receptor distributions on P388D1 cells, human peripheral blood lymphocytes, and normal mouse spleen cells. Essentially all of the P388D1 cells bore Fc receptors, distributed in a skewed Gaussian profile having a peak at 2 X 10(5) receptors per cell. Human peripheral blood lymphocytes and mouse spleen cells contained positive and negative subpopulations. The percentage of positive cells in human lymphocytes from different donors ranged from 50 to 25; the receptor distributions of these cells were symmetrical and similar in all donors in shape and average receptor density (4.2 X 10(4) receptors per cell). Mouse spleen cells contained 55% positive cells with nonsymmetrical heterogeneous distributions of receptor densities. These cells peaked at 1 to 2 X 10(4) receptors per cell, but significant numbers of cells had receptor densities 10- to 20-fold greater.

Animals↗
PubMed1980

The cellular nature of concanavalin A-stimulated antibody-dependent cell-mediated cytotoxicity.

Enhanced antibody-dependent cytotoxic cell (ADCC) activity, generated in vitro by concanavalin A (Con A) stimulation of human peripheral blood lymphocytes, was analyzed to identify the nature of the killer (K) cell. Stimulated and unstimulated 2-day cultured cells were separated into sheep erythrocyte rosetting (E+) and non-rosetting (E-) subpopulations, and ADCC activity was determined by using the slopes of linear dose-response plots as a measure of K cell activity. Quantitative comparisons were made by examining activity on a per cell basis ("specific" activity) and total K cell activity recovered per subpopulation. This analysis revealed that although the E- cells were enriched for specific ADCC activity, the E+ subpopulation contained the major proportion of the total recovered activity and nearly all of the Con A-induced increase in K cell activity.

Antibody-Dependent Cell Cytotoxicity↗