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Biomedical subjects

M Pierce

Publications and source records attributed to M Pierce.

At least 37 records · Page 2Linked to original sources
PubMed1986

Endocytosis of mannose-6-phosphate binding sites by mouse T-lymphoma cells.

The endocytosis and intracellular transport of mannose-6-phosphate conjugated to bovine serum albumin (Man-6-P:BSA) by mouse T-lymphoma cells were investigated in detail using several methods of analysis, both morphological and biochemical. Man-6-P:BSA was labeled with fluorescein or 125I and used to locate both surface and intracellular Man-6-P binding sites by light or electron microscopy, respectively. Incubation of cells with either fluorescent- or 125I-labeled Man-6-P:BSA at 0 degree C revealed a uniform distribution of the Man-6-P binding sites over the cell surface. Competition experiments indicate that the Man-6-P:BSA binding sites on the cell surface are the same receptors that can recognize lysosomal hydrolases. After as little as 1 min incubation at 37 degrees C, endocytosis of Man-6-P binding sites was clearly observed to occur through regions of the plasma membrane and via vesicles that also bound anticlathrin antibody. After a 5-15-min incubation of cells at 37 degrees C, the internalized ligand was detected first in the cis region of the Golgi apparatus and then in the Golgi stacks using both autoradiography and immunocytochemistry to visualize the ligand. The appearance of Man-6-P:BSA in the Golgi region after 15-30 min was confirmed by subcellular fractionation, which demonstrated an accumulation of Man-6-P:BSA in light membrane fractions that corresponded with the Golgi fractions. After a 30-min incubation at 37 degrees C, the internalized Man-6-P binding sites were localized primarily in lysosomal structures whose membrane but not lumen co-stained for acid phosphatase. These results demonstrate a temporal participation of clathrin-containing coated vesicles during the initial endocytosis of Man-6-P binding sites and that one step in the Man-6-P:BSA transport pathway between plasma membrane and the lysosomal structure can involve a transit through the Golgi stacks.

Animals↗
PubMed1985

Separation of metabolically radiolabeled O-methylmannitols and O-methylfucitol by reverse-phase high-performance liquid chromatography: applications to animal cell oligosaccharide structural analysis.

Reverse-phase high-performance liquid chromatography was utilized to separate efficiently and rapidly a standard mixture of various radiolabeled O-methylated mannitols and O-methylfucitol commonly encountered when vertebrate asparagine-linked oligosaccharides are subjected to permethylation, hydrolysis, and reduction with NaBH4. The following reduced, radioactive O-methylhexitols were resolved: 2,4-, 3,4-, and 3,6-di-O-methylmannitols; 3,4,6-tri-O-methylmannitol, 2,3,4-tri-O-methylfucitol, and 2,3,4,6-tetra-O-methylmannitol. To demonstrate the utility of this separation method in the analysis of metabolically radiolabeled asparagine-linked oligosaccharides, mouse lymphoma BW 5147 cells were metabolically radiolabeled with [2-3H]mannose and their glycopeptides prepared by Pronase digestion and fractionated by serial chromatography on immobilized lectins. Each fraction was subjected to methylation and hydrolysis, the released monosaccharides were reduced, and the radioactive O-methylhexitols were separated by reverse-phase HPLC. The relative amounts of the O-methylhexitols in each glycopeptide fraction analyzed were similar to those values determined by a combination of other separation systems.

Animals↗
PubMed1985

The experience with definitive irradiation of clinically limited squamous cell cancer of the trachea.

Squamous cell cancer of the trachea is an uncommon malignancy infrequently cured by either surgery or irradiation. Failure to control the primary tumor has been the most common cause for death. Our experience in three patients with small squamous cell cancer of the trachea treated by definitive irradiation is encouraging. The primary tumor was controlled in each patient. Histologic confirmation of tumor sterilization was observed in two. One patient remains alive and well at 54 months; one patient developed an epidural metastases at 16 months and subsequently died; and the third patient developed a separate primary lung cancer at 48 months. Two patients developed significant radiation complications.

Adult↗
PubMed1985

Significant attitude changes among residents associated with a pediatric nurse practitioner.

In the study reported here, significant changes in attitudes occurred over a three-year period among pediatric residents who had close association with a pediatric nurse practitioner (PNP) in their residency program. Residents entering the training program had an overall uncertain attitude toward PNPs but did respond favorably to the PNP's participation in the care of well children, patient education, and follow-up care. The greatest number of attitude changes toward the PNP took place after the first year of interaction between the residents and the PNP. After the first year, the residents favored the PNP's participation in the care of minor illnesses and stable chronic illnesses and in management of telephone calls from parents with concerns. Second-year residents also felt that the PNP was accepted by patients and physicians, that the PNP contributed to the reduction of health care costs, and that the PNP should have an expanded role. Upper-level residents continued to express favorable attitudes. The findings support the authors' hypothesis that favorable attitudes on the part of physicians toward PNPs can be cultivated during residency training by exposing residents to a PNP role model. In addition, the findings suggest that subsequent years of exposure will reinforce these favorable opinions if favorable opinions are established early in residency training.

Attitude of Health Personnel↗
PubMed1984

The effects of high-dose corticosteroids in patients with septic shock. A prospective, controlled study.

To determine whether corticosteroids are efficacious in severe septic shock, we conducted a prospective study of 59 patients randomly assigned to a methylprednisolone, dexamethasone, or control group. Patients were treated 17.5 +/- 5.4 hours (mean +/- S.E.M.) after the onset of shock, and 55 patients required vasopressor agents. Early in the hospital course, reversal of shock was more likely in patients who received corticosteroids than in those who did not. Four (19 per cent) of 21 methylprednisolone-treated, 7 (32 per cent) of 22 dexamethasone-treated, and none of 16 control patients had reversal of shock 24 hours after drug administration (corticosteroid groups vs. control group, P less than 0.05). Patients treated with corticosteroids within four hours after the onset of shock had a higher incidence of shock reversal (P less than 0.05). At 133 hours after drug administration, 17 (40 per cent) of 43 corticosteroid-treated patients had died, and 11 (69 per cent) of 16 control patients had died (P less than 0.05). However, these differences in reversal of shock and survival disappeared later in the course. Overall, 16 (76 per cent) of 21 patients receiving methylprednisolone, 17 (77 per cent) of 22 patients receiving dexamethasone, and 11 (69 per cent) of 16 controls in the hospital died. We conclude that corticosteroids do not improve the overall survival of patients with severe, late septic shock but may be helpful early in the course and in certain subgroups of patients.

Adolescent↗
PubMed1984

A ganglioside antigen on the rat pancreatic B cell surface identified by monoclonal antibody R2D6.

In an attempt to identify B cell specific antigens, we have generated a mouse monoclonal antibody, R2D6, which is directed against plasma membranes of rat pancreatic B cells but against no other pancreatic cells. R2D6 crossreacted with mouse and guinea pig B cells, but not with human or dog. The B cell specificity of R2D6 was utilized in fluorescence-activated cell sorting to prepare highly enriched separate populations of viable pancreatic islet B cells and A cells. R2D6 also recognized adrenal chromaffin cells, secretory cells in the anterior pituitary, and the myenteric plexus of the gastrointestinal tract. Trypsin, chymotrypsin, papain, ficin, and pronase had no effect on R2D6-binding to dissociated rat islet cells. However, neuraminidase treatment of intact cells reduced R2D6-binding by 75%. The antigen recognized by R2D6, Ag(R2D6), could be quantitatively extracted from rat islets by dichloromethane/methanol (2:1) and, after drying, was soluble in methanol alone as well as in phosphate-buffered saline. When the dichloromethane/methanol extract (DME) was bound to polyvinylchloride microtiter plates, antigenic activity was retained and remained insensitive to pronase. In this solvent-extracted form, antigenic activity was totally destroyed by neuraminidase. Therefore, sialic acid is either an integral part of, or is related sterically to the binding site (epitope) for R2D6. In high performance thin-layer chromatographs of the DME, developed in 60:40:9 chloroform/methanol/2.5 N ammonia, Ag(R2D6) migrated with a relative mobility (Rf) of 0.54 +/- 0.07 (n = 3), which was a position nearly coincident with the purified brain ganglioside, GD1a. The antigen bound to DEAE-Sephacel, was not inactivated by mild treatment with base (which hydrolyzes phospholipids) and eluted in ganglioside fractions upon C18 Sep-Pak and upon silicic acid chromatography. Hence, the solubility characteristics, enzyme sensitivities, and behavior of Ag(R2D6) in four chromatography systems are consistent with its identification as a ganglioside.

Animals↗
PubMed1983

Cell-cell recognition in yeast. Characterization of the sexual agglutination factors from Saccharomyces kluyveri.

The cell surface molecules responsible for sexual agglutination between haploid cells of opposite mating type from Saccharomyces kluyveri have been purified and characterized. The 17-factor, released from 17-cells by beta-glucanase digestion (Zymolyase), is a glycoprotein of 6 X 10(4) Da. Its binding activity is heat- and protease-labile, but it is stable to reducing agents and exo-alpha-mannosidase digestion. The 16-factor, released from 16-cells by Zymolyase digestion, has a molecular weight of 5 X 10(5) and is over 95% carbohydrate. An active binding fragment can be released from 16-factor, from the factor purified from a mutant of 16-cells (16(mnn1)-factor), and from the surfaces of the cells themselves by dithiothreitol treatment. The 16(mnn1)-binding fragment has a molecular weight of 2 X 10(4) and is 30% carbohydrate. Its binding activity is stable to heat and some proteases, but it is labile to pronase, carboxypeptidases A and Y, alpha-mannosidases, and mild periodate treatment. 125I-16(mnn1)-binding fragment adheres specifically to 17-cells but does not bind to 16-cells or cells of other yeast strains. The binding of the labeled fragment to 17-cells is characterized by a KA of 10(8) M-1, and 5 X 10(5) binding sites are present per cell. The purified intact factors are monovalent and appear to interact in a lock and key fashion to cause the specific agglutination of S. kluyveri 16- and 17-cells.

Amino Acids↗
PubMed1982

Quantification of ganglioside GM1 synthetase activity on intact chick neural retinal cells.

Neural retinal cells from 9-d-old chick embryos were assayed for uridine diphosphate (UDP)-galactose:ganglioside GM2 galactosyltransferase, or GM1 synthetase, activity using the oligosaccharide fragment of GM2, oligo-GM2, oligo-GM2, as the exogenous acceptor. The results demonstrated that this enzyme activity was present on the external surfaces of intact cells. Little difference between the specific activities of cell surface GM1 synthetase could be detected when cells derived from dorsal and ventral segments of the neural retina were compared. These results suggested that this cell-surface enzyme was not present in a concentration gradient along the dorsoventral axis of the neural retina.

Animals↗
PubMed1979

Galactosyltransferase from commerical preparations of fetuin.

A galactosyltransferase that transfers galactose from UDPgalactose to asialoagalacto fetuin or N-acetylglucosamine was partly purified from two commerical preparations of fetuin and its kinetic properties were characterized. Several other preparations of fetuin were also found to contain galactosyltransferase activity.

Animals↗
PubMed1978

32P-labeling patterns in rat pancreatic islets: tissue source of the radiophosphate released after glucose stimulation.

Immediately after stimulation with glucose in vitro, isolated rat pancreatic islets prelabeled with [32P]orthophosphate release a pulse of [32P]orthophosphate into the media (the "phosphate flush"). Islets have been rapidly frozen before, during, and after this pulse to assess the concurrent changes in the distribution of tissue radioactivity. Under the present experimental conditions, approximately 90% of the islet radioactivity was soluble in perchloric acid (PCA-soluble) immediately before stimulation and slightly more than half of that was present as [32P]orthophosphate. The tissue pool of [32P]orthophosphate declined 55% and 62% after 7 and 14 min of stimulation which, respectively, incorporated the peak and the end of the heightened efflux of radioactivity. The net decrementa in tissue orthophosphate could account for all of the radioactivity which was released during the "phosphate flush." During the 14-min period of stimulation, labeled ATP and GTP (which had accounted for 13% and 4% of total PCA-soluble radioactivity before stimulation) increased 51% and 35%, respectively, and labeled ADP and AMP (which had accounted for 5.4% and 1.5% of PCA-soluble counts) fell 36% and 77%, respectively. Certain other PCA-soluble components, such as phosphorylcholine and phosphorylethanolamine, and total PCA-insoluble radioactivity were not demonstrably altered. The findings indicate that the "phosphate flush" originates from a labile pool of tissue orthophosphate. It remains to be established whether the simultaneous changes in the turnover of selected nucleotides are coupled to the translocation of orthophosphate or are mediated separately.

Animals↗
PubMed1977

Biochemical investigations of retinotectal specificity.

An in vitro assay for retinotectal specificity has been described. The results show that, in the chick embryo, cells dissociated from the dorsal retina preferentially adhered to ventral tectal surfaces while cells from the ventral retina preferentially adhered to dorsal tectal surfaces. These adhesive preferences thus mimic the retinotectal specificity observed in vivo. The assay has been extended for use with plasma membrane preparations from retinal cells. Experiments in which retinal cells or tectal surfaces were treated with purified proteases and glycosidases have partially characterized the moieties responsible for the observed specificities. These results are consistent with a double gradient in the dorsal-ventral axis of complementary proteins and carbohydrates. The carbohydrate moiety would be expected to terminate in an acetylated hexosamine and to be more concentrated dorsally in both retina and tectum. A protein that is complementary to the hexosamine terminus would be localized in the ventral part of retina and tectum.

Animals↗
PubMed1976

Mutations masking the lambda cin-1 mutation.

A lambda cnc mutation masks the phenotype of the lambda cin-1 mutation. Eleven cnc mutations have been found. All appear to be identical and all map at a site just to the right of the cin-1 site. It is probable that cin-1 and cnc affect adjacent, or almost adjacent nucleotide pairs. Neither the cin-1 nor the cnc mutation involves large additions or deletions of genetic material.

Chromosome Mapping↗
PubMed1976

[32P] Orthophosphate efflux from pancreatic islets: graded response to glucose stimulation.

The dose-response relationships of the glucose-induced rapid transient efflux of [32P] orthophosphate from prelabeled pancreatic islets ("phosphate flush") have been investigated. Threshold levels for eliciting a "phosphate flush" were between 0.5 and 1.0 mg/ml glucose, the apparent "Km" for this event was 1.0-1.5 mg per ml and the apparent "Vmax" was reached at ambient glucose concentrations between 1.5 and 2.0 mg per ml. This dose-response curve is somewhat shifted to the left in comparison with previously published data for glucose-induced insulin release. Thus, the "phosphate flush" appears to display a more narrow dose-response curve to ambient glucose concentrations than the actual release of insulin. It is proposed that this may constitute further evidence that the "phosphate flush" reflects an early step in stimulus-secretion coupling and that there may be some loss of sensitivity as the glucose signal is transmitted from the site of recognition to the final site of hormone release.

Animals↗