Search PubMed⌕ Search

Biomedical subjects

S C Brown

Publications and source records attributed to S C Brown.

At least 55 records · Page 3Linked to original sources
PubMed1997

DNA base composition of Allium genomes with different chromosome numbers.

The present report examines whether the presence of an additional chromosome can be detected as modifying the nuclear DNA amount and base composition of the cell, determined here by flow cytometry of interphasic nuclei, using four monosomic additions (chromosomes 3C, 4C, 7C and 8C transmitted from Allium cepa to Allium fistulosum L.). A. cepa differs significantly from A. fistulosum in genome size (2C DNA = 33.2 pg in A. cepa and 23.3 pg in A. fistulosum) as well as in GC content (38.7% and 39.8%, respectively). The presence of an extra chromosome of A. cepa obviously increases the nuclear DNA amount above the A. fistulosum value but also alters the apparent mean GC content. By comparing the monosomic additions and the parental background, the DNA amount and base composition of each of the four single chromosomes were calculated to quantify the GC content per chromosome and therefore to deduce their initial contribution to the A. cepa genome. Taken individually, some chromosomes are atypical in terms of GC content: the single chromosome 3C is AT-rich, having only about only 25% GC. However, the three other chromosomes examined are typical of the A. cepa genome in base composition. Indeed, this biological panel gives access to chromosomal features via a cytometric assay of nuclei. It should facilitate quantification of GC-rich repetitive sequences forming heterochromatic domains located mainly at the telomeres in the monocotyledonous A. cepa genome.

Allium↗
PubMed1997

NMR structure of a protein kinase C-gamma phorbol-binding domain and study of protein-lipid micelle interactions.

Classical protein kinase C (PKC) family members are activated by the binding of various ligands to one of several cysteine-rich domains of the enzyme. The natural agonist, diacylglycerol (DAG), and the natural product superagonist, phorbol dibutyrate (PDB), activate the enzyme to produce wide-ranging physiological effects. The second cysteine-rich (Cys2) domain of rat brain PKC-gamma was expressed and labeled with 15N and 13C, and the solution structure was determined to high resolution using multidimensional heteronuclear NMR methods. The phorbol binding site was identified by titrating this domain with phorbol-12,13-dibutyrate (PDB) in the presence of organic cosolvents. Titrations of this domain with lipid micelles, in the absence and presence of phorbols, indicate selective broadening of some resonances. The observed behavior indicates conformational exchange between bound and free states upon protein-micelle interaction. The data also suggest that half of the domain, including the phorbol site and one of the zinc sites, is capable of inserting into membranes.

Amino Acid Sequence↗
PubMed1997

Utrophin-dystrophin-deficient mice as a model for Duchenne muscular dystrophy.

The absence of dystrophin at the muscle membrane leads to Duchenne muscular dystrophy (DMD), a severe muscle-wasting disease that is inevitably fatal in early adulthood. In contrast, dystrophin-deficient mdx mice appear physically normal despite their underlying muscle pathology. We describe mice deficient for both dystrophin and the dystrophin-related protein utrophin. These mice show many signs typical of DMD in humans: they show severe progressive muscular dystrophy that results in premature death, they have ultrastructural neuromuscular and myotendinous junction abnormalities, and they aberrantly coexpress myosin heavy chain isoforms within a fiber. The data suggest that utrophin and dystrophin have complementing roles in normal functional or developmental pathways in muscle. Detailed study of these mice should provide novel insights into the pathogenesis of DMD and provide an improved model for rapid evaluation of gene therapy strategies.

Animals↗
PubMed1997

Genetic correction of dystrophin deficiency and skeletal muscle remodeling in adult MDX mouse via transplantation of retroviral producer cells.

Duchenne muscular dystrophy (DMD) is an X-linked, lethal disease caused by mutations of the dystrophin gene. No effective therapy is available, but dystrophin gene transfer to skeletal muscle has been proposed as a treatment for DMD. We have developed a strategy for efficient in vivo gene transfer of dystrophin cDNA into regenerating skeletal muscle. Retroviral producer cells, which release a vector carrying the therapeutically active dystrophin minigene, were mitotically inactivated and transplanted in adult nude/mdx mice. Transplantation of 3 x 10(6) producer cells in a single site of the tibialis anterior muscle resulted in the transduction of between 5.5 and 18% total muscle fibers. The same procedure proved also feasible in immunocompetent mdx mice under short-term pharmacological immunosuppression. Minidystrophin expression was stable for up to 6 mo and led to alpha-sarcoglycan reexpression. Muscle stem cells could be transduced in vivo using this procedure. Transduced dystrophic skeletal muscle showed evidence of active remodeling reminiscent of the genetic normalization process which takes place in female DMD carriers. Overall, these results demonstrate that retroviral-mediated dystrophin gene transfer via transplantation of producer cells is a valid approach towards the long-term goal of gene therapy of DMD.

Animals↗
PubMed1997

Chest pain and cocaine use in 18 to 40 year-old persons: a retrospective study.

This study examined the incidence of chest pain and cocaine use in 18 to 40 year olds who were seen in a public inner city emergency department. The medical records of a convenience sample of 386 subjects were reviewed during a 6 month period. Results revealed a significant differences in physiological, diagnostic, and patient history data associated with chest pain and cocaine use. These data that highlight the clinical features of cocaine users can be used as a profile for the management of cocaine-related cardiovascular problems.

Adolescent↗
PubMed1996

The carboxy-terminal region of human interleukin-5 is essential for maintenance of tertiary structure but not for dimerization.

The C-terminal region of interleukin-5 has previously been suggested to be important for biological activity [Mackenzie et al., (1991), Mol. Immunol. 28, 155-158; Kodama et al. (1991), Biochem. Biophys. Res. Commun. 178, 514-519]. We have investigated this region by making a series of truncation mutants. The proteins were expressed in Escherichia coli, purified from inclusion bodies, and were able to refold with the disulfide homodimeric topology typical of interleukin-5. Analysis of the truncated carboxy-terminal proteins in an interleukin-5-dependent proliferation assay on TF-1 cells showed a rapid loss of activity as the C-terminal was shortened by more than two amino acids. This loss of biological activity correlated with a drop in binding affinity to both the alpha chain of the receptor and the high-affinity complex consisting of the alpha and beta subunits. Analysis of the proteins by 1H-NMR showed that the truncated mutants have higher exchange rates with solvent, indicating a less rigid structure. The carboxy-terminal region is therefore necessary to maintain the stability of the four-helix bundle and to orient correctly the important residues of the fourth helix. Inspection of the structure determined by X-ray crystallography shows that Trp-110 acts as the major residue in anchoring the fourth helix.

Amino Acid Sequence↗
PubMed1995

Flow sorting of the Y sex chromosome in the dioecious plant Melandrium album.

The preparation of stable chromosome suspensions and flow cytometric sorting of both the Y sex chromosome of the white campion, Melandrium album, and the deleted Y chromosome of an asexual mutant, 5K63, is described. The principle has been to maintain transformed roots in vitro, synchronise and block mitosis, reduce cells to protoplasts, and lyse these to release chromosomes. Such in vitro material, unlike many cell suspensions, showed a stable karyotype. Factors critical to producing high-quality chromosome suspensions from protoplasts include osmolality of isolation solutions and choice of spindle toxin and of lysis buffer. Agrobacterium rhizogenes transformed young growing root cultures were synchronised at G1/S with 50 microM aphidicolin for 24 h and released to a mitotic block with 30 microM oryzalin for 11 h. Protoplast preparations from such tissue routinely had metaphase indices reaching 15%. Suspensions of intact metaphase chromosomes, with few chromatids, were obtained by lysing swollen mitotic protoplasts in a citric acid/disodium phosphate buffer. Except for the presence of clumps of autosomal chromosomes near the X and Y chromosome zones, monoparametric histograms of fluorescence intensities of suspensions stained with 4',6-diamino-2-phenylindole showed profiles similar to theoretical flow karyotypes. Two types of Y chromosomes, one full-length and one partially deleted (from the asexual mutant), could be sorted at 90% purity (21-fold enrichment of Y). These results are discussed in the context of sex determination and differentiation in higher plants.

Cell Line, Transformed↗
PubMed1995

Artificial antisense RNA regulation of YBR1012 (YBR136w), an essential gene from Saccharomyces cerevisiae which is important for progression through G1/S.

YBR1012 (YBR136w) is an essential gene from Saccharomyces cerevisiae identified during the systematic sequencing of part of the right arm of chromosome II. We previously constructed a conditional allele of YBR1012 based on antisense RNA, by inserting a small fragment of this gene downstream from the inducible UASGAL10-CYC1 promoter. Several other antisense RNA constructions have since been made and their activity tested. The response of the system appears to be very delicate, as the presence or absence of 13 nucleotides of polylinker in the 300 nucleotide antisense transcript can dramatically modify its effectiveness. The most effective antisense RNA construction was used in flow cytometry studies to investigate the role of ybr1012p. The results show that during the antisense RNA block some 80% of the cells are arrested with their DNA unreplicated, suggesting that Ybr1012p is needed for progression through G1 or early S phase.

Cell Cycle↗
PubMed1995

Modulation of membrane fluidity and lipidic metabolism in transformed rat fibroblasts induced by the sesquiterpenic hormone farnesylacetone.

Farnesylacetone is a natural terpene extracted from androgenic glands of the crustacean Carcinus maenas and is capable of inhibiting proliferation, notably in transformed mammalian cells. Flow cytometry with three lipophilic probes, diphenylhexatriene, trimethylammonium-diphenylhexatriene, and Nile red, has revealed modifications of the lipidic metabolism in transformed FR3T3-mTT4 rat fibroblasts treated by farnesylacetone, including changes in membrane fluidity. Farnesylacetone strongly increased the number of neutral lipidic droplets in the cytoplasm. Moreover, after prolonged terpene treatment, the membrane fraction of cells contained a substantial level of triglycerides. Farnesylacetone provoked an immediate but transitory increase in membrane fluidity of the cell membrane. The change in measured lipid fluidity appears to be due to these triglycerides rather than to the phospholipids.

Animals↗
PubMed1995

Glomerular filtration rate measurement: a neglected test in urological practice.

OBJECTIVES: To assess the accuracy and reproducibility of methods used to measure glomerular filtration rate (GFR) in clinical practice. PATIENTS AND METHODS: Simultaneous clearances of iohexol, 99mTc-diethylene-triamine-penta-acetic acid (DTPA), 24 h renal creatinine clearance and creatinine clearance estimated from a serum sample only, were obtained in 31 patients. Accuracy was calculated relative to iohexol clearance. The reproducibility of each method was determined by repeat measurements in the same individuals: three 99mTc-DTPA clearances in 21 patients; three 24 h creatinine clearance estimations in 12 patients; and three serum creatinine clearance estimations in 21 patients. RESULTS: The mean differences between 99mTc-DTPA clearance, 24 h renal creatinine clearance and estimated creatinine clearance compared with the reference method were 0.2 mL/min, 21.6 mL/min and 0.6 mL/min, respectively. 99mTc-DTPA clearance had fairly tight 95% limits of agreement (12.2 mL/min) compared with 35.4 mL/min for 24 h creatinine clearance and 25.8 mL/min for estimated creatinine clearance. The reproducibility for each method was 5.4%, 24.3% and 6.1%, respectively. CONCLUSION: Single-injection 99mTc-DTPA clearance provides an accurate and reproducible method of GFR measurement that is suitable for objective monitoring of renal function. Twenty-four hour creatinine clearance is neither sufficiently accurate nor reproducible for this purpose. Creatinine clearance estimation from a serum sample without urine collection may be preferable to the more traditional 24 h collection method.

Creatinine↗
PubMed1995

Spatial and temporal distribution of [Ca2+]i in normal human myotubes. A fura-2 imaging study.

The spatio-temporal distribution of intracellular, free calcium ions, [Ca2+]i, induced in human myotubes by electrical stimulation typically showed a relatively large increase of [Ca2+]i in the vicinity of the plasmalemma. The similarity of this distribution, with that observed after the application of caffeine, and the lack of any effect of lanthanum, strongly suggest that the main source of Ca2+ participating in the electrically induced transient is the sarcoplasmic reticulum. Aneurally cultured human myotubes therefore display a 'skeletal muscle type' coupling between membrane depolarization and calcium release. However, the relatively slow time course of the electrically induced transients compared to rat and mouse myotubes, together with the inability of Ca2+ released from the sarcoplasmic reticulum to activate the contractile machinery, implies that aneurally cultured human myotubes achieve only a limited degree of differentiation. The relevance this may have to an apparent delay between the electrically induced rise in intranuclear Ca2+ relative to cytosolic Ca2+ remains to be determined but, at this stage of differentiation, there appears to be some form of barrier to free diffusion between the two cellular compartments.

Caffeine↗
PubMed1994

NMR solution structure of a peptide nucleic acid complexed with RNA.

Peptide nucleic acids (PNA) incorporating nucleic acid bases into an achiral polyamide backbone bind to DNA in a sequence-dependent manner. The structure of a PNA-ribonucleic acid (RNA) complex was determined with nuclear magnetic resonance methods. A hexameric PNA formed a 1:1 complex with a complementary RNA that is an antiparallel, right-handed double helix with Watson-Crick base pairing similar to the "A" form structure of RNA duplexes. The achiral PNA backbone assumed a distinct conformation upon binding that differed from previously proposed models and provides a basis for further structure-based design of antisense agents.

Magnetic Resonance Spectroscopy↗
PubMed1994

Muscle at birth in mice selected for large and small body size.

Selection for divergent body weight at 6 wk of age in the Q strain mouse has produced large (QL) and small (QS) mice which differ 2-fold in their adult body weight. The purpose of this investigation was to identify some of the cellular mechanisms which underlie the early divergence in size between the 2 lines. At birth, QL mice (for similar litter sizes) were 28% heavier and 6% longer than QS mice. This was reflected by measurements of longitudinal bone length which were greater in QL (tibia 6.2%, humerus 4.2%) compared with QS mice. Fibre number was found to be 18 and 17% greater in the biceps brachii and soleus muscles respectively of the QL mice. It was concluded that this was not a consequence of any alteration in the ratio of developing secondary to primary myofibres in either muscle. Fibre cross-sectional areas were only significantly different between the QL and QS for the soleus muscle, which might be explained by the relatively greater divergence in the length of its supporting bone (tibia) between the QL and QS compared with the humerus. Estimates of nuclear number showed that there were significantly more nuclei in biceps brachii muscle of QL than in the QS mice which could be attributed to the difference in fibre number, although no such differences were found for the soleus muscle. There was no apparent alteration in the proportion of nuclei found within the fibres of the biceps muscle. Overall the results indicate that selection in this situation has acted through the normal cellular processes of growth.

Animals↗