PubMed Health⌕ Search

Biomedical subjects

M E Brown

Publications and source records attributed to M E Brown.

At least 19 recordsLinked to original sources
PubMed1996

Analysis of basic fibroblast growth factor in rats with inherited retinal degeneration.

In RCS rats, photoreceptors degenerate between postnatal days 20 and 60, secondary to a genetic defect expressed in the neonatal retinal pigmented epithelium (RPE). Previous work has shown delay of the photoreceptor degeneration in this model by intraocular injection of basic fibroblast growth factor (bFGF). Evidence is presented here, from bFGF immunostaining and Northern analysis of bFGF mRNA, for reduced bFGF expression in uncultured RPE of dystrophic RCS pups. It is also shown that in the mutant eyes angiogenesis in the underlying choroid, which normally occurs between postnatal days 7 and 10, is markedly delayed, with irregular distribution of vessels, consistent with a reduction in this known angiogenesis factor. Mutational analysis of the bFGF transcript and gene by denaturing gradient gel electrophoresis and Southern analysis did not, however, reveal abnormalities in the coding sequence of this gene in RCS rats.

Animals↗
PubMed1996

The secretion of active recombinant human gastric lipase by Saccharomyces cerevisiae.

The expression of the complete human gastric lipase (HGL) gene in Saceharomyces cerevisiae grown in defined medium resulted in the secretion of active recombinant HGL (rec.HGL) to levels of up to approximately 11 mg/liter. Of the total measurable HGL activity, 90% was detected by assaying intact cells, suggesting that the majority of rec.HGL produced was secreted but stayed attached to the cell wall. The remaining 10% was present in the growth medium and from this source active rec.HGL was purified 300-fold by a combination of hydrophobic interaction and ion-exchange chromatography. Rec.HGL migrated on reduced SDS-PAGE as three bands with estimated molecular masses of 47,45, and 43 kDa. All three forms cross-reacted with an antibody raised to natural HGL and their treatment with Endo H showed them to be N-linked glycosylation variants of a single polypeptide. The 47-kDa species was isolated using lentil lectin Sepharose 4B and shown to possess a specific activity comparable to that of the natural enzyme. Rec.HGL had an acid pH activity optimum using either tributyrin or olive oil as substrate and did not lose activity if incubated in the presence of pepsin at pH 2.0. These results demonstrate that HGL secreted by Saccharomyces cerevisiae retained those properties of the natural enzyme required for its use in the treatment of pancreatic insufficiency.

Chromatography, Agarose↗
PubMed1996

Protein-bound and unbound forms of plasma cortisol in piglets after castration at seven or 14 days of age.

Male piglets from seven litters were used to evaluate the long-term effects of castration and age at castration on the concentrations of cortisol and its distribution in peripheral plasma. The piglets were allotted to one of three groups of 12 animals according to whether they had been castrated at seven or 14 days old or left intact. The animals were weighed and blood samples were collected from the anterior vena cava at three, seven, 10, 14, 17, 21, 28, 35 and 42 days of age. The plasma concentrations of albumin, total protein, cortisol, and the percentage distribution of cortisol among corticosteroid binding globulin-bound (CBG-bound), albumin-bound and unbound forms, were determined. Castration at seven or 14 days of age had no effect on any of the parameters measured. In all the piglets, the percentage of unbound cortisol decreased (P < 0.001) from 24.5 to 11.6 per cent over the period of sampling. The percentage of CBG-bound cortisol increased (P < 0.001) from 19 per cent on day 17 to 47 per cent on day 21, and albumin-bound cortisol declined from 61 to 37 per cent over the same period. These results agree with previous findings of age-related changes in the percentage distribution of cortisol in the piglet and demonstrate that these patterns are unaffected by castration.

Age Factors↗
PubMed1995

The fragment R collision: W. M. Keck telescope observations of SL9.

The W. M. Keck telescope was used to observe the impact of comet Shoemaker-Levy 9 (SL9) fragment R at a wavelength of 2.3 micrometers on 21 July 1994. The data showed three outbursts. The first flash lasted about 40 seconds and was followed 1 minute after its peak by a second flash that lasted about 3 minutes. A third, longer lasting flare began 6 minutes after the first flash and lasted for 10 minutes. At its maximum brightness, the flare outshone Jupiter. The two short flashes are probably associated with the initial meteor trail and the subsequent fireball, respectively. The bright flare occurred when the impact site rotated into view. These data show that the explosion ejected material at least 1300 kilometers above the visible cloud tops. The luminosity of the impact site during the long bright flare was probably maintained by the release of gravitational potential energy, as this material fell back onto the lower atmosphere.

Atmosphere↗
PubMed1995

Cell cycle-dependent regulation of p185neu: a relationship between disruption of this regulation and transformation.

Structure and function of p185neu receptor tyrosine kinase were found to be regulated in a cell cycle-dependent manner. In M phase, p185neu is hyperphosphorylated at serine and/or threonine residues. The phosphotyrosine [Tyr(P)] content of p185neu is at its highest level in G0/G1 phase, decreases through S and G2 phases, and reaches its lowest level in M phase. Phospholipase C-gamma (PLC-gamma) and GTPase-activating protein (GAP), substrates of p185neu, also have a similar profile of Tyr(P) content during the cell cycle. These results, along with in vitro immune complex kinase assays, suggest that the tyrosine kinase activity of p185neu is least active in M phase. Interestingly, the mutation-activated neu oncogene (neu*)-encoded protein product, p185neu* escaped from cell cycle regulation. Taken together, we demonstrate in this report that the structure and function of p185neu are regulated in a cell cycle-dependent manner, yet p185neu* escapes from this regulation and remains active through the cell cycle. Disruption of this cell cycle regulation may define a mechanism for p185neu*-mediated cellular transformation.

3T3 Cells↗
PubMed1995

Acquired cor triatriatum after orthotopic cardiac transplantation.

Orthotopic cardiac transplantation was performed in a 42-year-old woman with idiopathic cardiomyopathy. Postoperative right ventricular failure developed and a transesophageal echocardiogram demonstrated acquired cor triatriatum with marked obstruction to mitral valve inflow and severe right ventricular dilatation. At reexploration, redundant donor atrial tissue was excised correcting the cor triatriatum. She was alive and well with normal hemodynamic parameters 12 months after transplantation.

Adult↗
PubMed1995

An improved left ventricular cannula for chronic dynamic blood pump support.

Ventricular unloading with dynamic blood pumps can be markedly affected by the geometry of the cannula tip within the ventricular chamber. Due to the ability of these pumps to develop significant negative inflow pressure, existing cannula tips designed for passively filling blood pumps can be predisposed to inflow occlusion by intraventricular anatomic structures. A novel "trumpet" mouth cannula was constructed to overcome this limiting problem. This design was based on two criteria: to provide additional stenting to the ventricular apex, and to assure placement of the tip opening relative to the endocardial surface. This prototype cannula was evaluated in vivo against conventional caged, blunt, and tapered designs to assess anatomic and hemodynamic interaction within the ventricular apex. Post mortem examination revealed the inflow tract to be devoid of myocardial obstruction in all cases. These initial studies indicate that a trumpet mouth cannula can provide satisfactory hemodynamic performance required by dynamic blood pumps.

Animals↗
PubMed1995

Cytotoxic T lymphocyte response to hepatitis C virus-derived peptides containing the HLA A2.1 binding motif.

The HLA class I-restricted cytotoxic T lymphocyte (CTL) response is a major defense mechanism in viral infections. It has been suggested that the CTL response may contribute to viral clearance and liver cell injury during hepatitis C virus (HCV) infection. To test this hypothesis requires an understanding of the characteristics of HCV-specific cytotoxic effector cells and identification of the target antigens to which they respond. To begin this process we stimulated peripheral blood mononuclear cells (PBMC) from a group of HLA-A2 positive patients with chronic hepatitis C with a panel of 130 HCV-derived peptides containing the HLA-A2 binding motif. Effector cells were tested for their capacity to lyse HLA-A2-matched target cells that were either sensitized with peptide or infected with a vaccinia virus construct containing HCV sequences. Using this approach we have identified nine immunogenic peptides in HCV, three of which are derived from the putative core protein, three from the nonstructural (NS) 3 domain, two from NS4 and one from NS5. Selected responses were shown to be HLA-A2 restricted, mediated by CD8+ T cells and to recognize endogenously synthesized viral antigen. Unexpectedly, peptide-specific CTL responses could also be induced in sero-negative individuals, suggesting in vitro activation of naive CTL precursors. The precursor frequency of peptide-specific CTL was 10 to 100-fold higher in infected patients compared to uninfected controls, and the responses were greatly diminished by removal of CD45 RO+ (memory) T cells. Further quantitative studies are clearly required to establish whether a correlation exists between the HCV-specific CTL response and the clinical course of this disease. Definition of the molecular targets of the human CTL response to HCV creates this opportunity, and may also contribute to the development of a T cell-based HCV vaccine.

Amino Acid Sequence↗
PubMed1994

Selecting and designing cell lines for improved physiological characteristics.

We have developed several approaches to create cell lines with improved characteristics in cell culture. In some cases it has been possible to isolate natural variants with useful properties. Cholesterol independent variants of the mouse NSO myeloma cell line were isolated by cloning in a selective medium. A glutamine independent variant of a hyridoma was isolated by continuous (chemostat) culture under glutamine limited conditions in the presence of glutamate. Choline independent cells were isolated from a choline limited chemostat. In an alternative approach to modifying cell behaviour, we have used recombinant DNA techniques to introduce the glutamine synthetase (GS) gene to a hybridoma. This resulted in glutamine independence and increased productivity.

Animals↗
PubMed1993

Spontaneously arising immortal cell line of rat retinal pigmented epithelial cells.

A continuous cell line of rat retinal pigment epithelium (RPE), named BPEI-1, has been established and characterized. Sheets of pure RPE cells, uncontaminated by choroidal or neural retinal cell types, were isolated from eyes of 7-day-old Long Evans rats and established in primary culture. The primary RPE cells became extensively spread and grew slowly for approximately 1 month, at which time a colony of small rapidly dividing cells spontaneously appeared. Following trypsinization, most of the typical primary RPE cells did not survive and were quickly outnumbered by the smaller cells, which gave rise to a cell line that was grown continuously for several hundred generations. When growing at the maximal rate in media containing 20% FBS (doubling time 18 h), the cells were fibroblastic and nearly devoid of pigment, but were capable of morphologic transition back to a pigmented, epithelioid form when cultured under low serum conditions. Evidence that these cells originated from RPE included specific immunolabeling with antibodies to cellular retinaldehyde binding protein and cytokeratin, negative GFAP immunoreactivity, and demonstration of avid phagocytosis of isolated rod outer segments by these cells. Partial characterization of choroidal cells eliminated the latter cells as possible contaminants which could have given rise to the cell line. The BPEI-1 cell line, and other rat RPE cell lines currently being developed from pigmented normal (LE, RCS rdy+p+) and retinal dystrophic (RCS p+) rats should facilitate biochemical and molecular biological approaches to study of RPE cell function in health and disease.

Animals↗
PubMed1993

Clinical cardiopulmonary bypass without systemic anticoagulation.

Heparin-coated perfusion systems have been used to perform cardiopulmonary bypass with decreased systemic heparin dosages. We report the case of a 60-year-old man with end-stage liver disease and severe aortic stenosis who underwent an aortic valve replacement without systemic anticoagulation, employing a heparin-coated perfusion system. In patients with a contraindication to systemic anticoagulation, cardiopulmonary bypass with a heparin-coated system without systemic anticoagulation may be a viable alternative.

Aortic Valve↗
PubMed1993

In vitro propagation of human ocular surface epithelial cells for transplantation.

PURPOSE: To examine the possibility that ocular surface epithelial cells might be grown in culture for use as grafts. METHODS: The proliferative capacity of epithelial cells cultured from the conjunctiva, limbus, and central cornea of normal human eyes was compared. Single cells disaggregated from approximately 1 mm2 biopsy specimens were serially cocultured with lethally irradiated mouse 3T3 fibroblasts. To study the cells' ability to reform a stratified epithelium, confluent limbal cultures were released as an intact cell sheet with the enzyme Dispase and transplanted to a dermal connective tissue bed in nude mice. Attachment and differentiation properties of the reconstituted epithelium were examined immunohistochemically. RESULTS: Central corneal epithelial cells could not be propagated; they senesced in first or second passage. In contrast, limbal epithelial cells exhibited a substantial (i.e., mean of 23 population doublings) and conjunctival cells a moderate (i.e., mean of 11 population doublings) proliferative capacity. Within 4 days of transplantation to the nude mouse dermis, cultured limbal epithelial cells formed an epithelium 5-6 cell layers thick. The epithelium adhered firmly to the graft bed, and deposition of the basement membrane and anchoring fibril protein collagens IV and VII and laminin was detectable immunohistochemically. The transplanted epithelium displayed limbuslike compartmental expression of keratins K3, K13, and K19, and of the enzyme enolase. CONCLUSIONS: These results support the concept that corneal epithelial stem cells are located in the limbus and indicate that cultured autologous limbal cells may function as grafts to permanently restore the corneal epithelium after severe ocular surface injury.

Adult↗
PubMed1993

Tumors produced in rats with a single dosage of 1,2-dimethylhydrazine.

To evaluate possible nephroblastoma induction in young Sprague-Dawley male rats by 1,2-dimethylhydrazine (DMH), agents including inhibitors and stimulators of the carcinogenesis were tested concurrently in 2 experiments. In series A, rats, 27 days of age, were fed the following as supplements in a basal diet at the final wt% given: hydralazine (0.035%), disulfiram (250 ppm), ferrous sulfate heptahydrate (0.55%; 0.11 g% Fe), isotretinoin (240 ppm), dehydroepiandrosterone (0.30%) in addition to selenium (2 ppm; drinker). At day 15, DMH was injected s.c. at 108 mg base/kg; duration on the diets: 51 weeks. Series B comprised 33 day-old males which were partially hepatectomized (control and indomethacin at 10 mg/l by drinker) or bilaterally gonadectomized for comparison vs sham-operated, and intact groups on s.c. injection of estradiol benzoate (15 micrograms/kg), progesterone (30 mg/kg) and diallyl sulfide (100 mg/kg), the respective controls receiving the peanut oil vehicle. Treatments were begun 8 days post-operative and 17 days later, the single dosage of DMH as in the above was injected. The oil solutions were administered at the specified weekly levels for a total of 52 injections, 2 doses being introduced per week for the 1st 3 weeks. Colon adenocarcinomas comprised the main tumors and occurred in about 15-50% of the rats with total frequencies in the respective control ranges except for decrements with the disulfiram- and iron-fed groups. Renal changes were more involved with series B and nephroblastomas of the left kidney occurred in one animal each of the estradiol benzoate- and diallyl sulfide-injected groups. Of interest, bilateral nephroblastomas were present in one of the saline-injected controls which was gonadectomized. Under the conditions explored, concurrent treatment with DMH inhibitors or synergists had a minimal effect on nephroblastoma induction.

1,2-Dimethylhydrazine↗
PubMed1991

Human nasal glandular secretion of novel antioxidant activity: cholinergic control.

Exposure of the respiratory mucosa to oxygen-enriched air contributes to the generation of the lung damage in both adult respiratory distress syndrome and bronchopulmonary dysplasia. Recent work has identified the nasal submucosal gland as the source of diverse molecules important in mucous membrane host defense. We searched for the presence of antioxidant activity in nasal glandular secretions, the absence of which could possibly predispose to oxygen-induced injury. Employing a low molecular weight preparation of nasal secretions (a pooled concentrate passed over a 10,000-dalton molecular sieve), antioxidant activity capable of inhibiting both horseradish peroxidase and Fenton reagent reactions was discovered. The following lines of evidence suggest that submucosal glands are the source of this activity. (1) Antioxidant activity present in resting, baseline nasal washings is significantly increased after cholinergic stimulation either in response to topical methacholine or induced by a gustatory reflex. (2) Application of atropine reduced the antioxidant activity to baseline levels after either of the cholinergic stimuli. (3) Levels of antioxidant activity correlated very closely with the secretion of lactoferrin, a recognized product secreted solely from the serous cell of the submucosal gland. The antioxidant activity is due to novel, previously unrecognized molecules. This activity is found in nasal secretions containing molecules less than 10,000 daltons, is unaffected by N-ethyl maleimide (which inactivates glutathione, another low molecular weight antioxidant), is not associated with the capacity to reduce cytochrome c (as seen with ascorbic acid), and resides in the water soluble pool of secretions (in contrast to vitamin E, another putative antioxidant).(ABSTRACT TRUNCATED AT 250 WORDS)

Ascorbic Acid↗