The occurrence of polyenoic very long chain fatty acids with greater than 32 carbon atoms in molecular species of phosphatidylcholine in normal and peroxisome-deficient (Zellweger''s syndrome) brain.

The n-6 tetra- and pentaenoic fatty acids with carbon chain lengths greater than 32 found in normal brain are located predominantly in a separable species of phosphatidylcholine. A similar phospholipid is found in increased amounts in the brain of peroxisome-deficient (Zellweger's syndrome) patients, but the fatty acid composition differs in that penta- and hexaenoic derivatives predominate. Our data strongly suggest that the polyenoic very long chain fatty acids are confined to the sn-1 position of the glycerol moiety, while the sn-2 position is enriched in saturated, monounsaturated and polyunsaturated fatty acids with less than 24 carbon atoms. It is postulated that these unusual molecular species of phosphatidylcholine may play some, as yet undefined, role in brain physiology.     

Transmission electron microscopy of small numbers of sorted cells

A new method that allows the transmission electron microscopic examination of as few as 1 x 10(4) cells obtained by flow cytometric sorting is described. The approach involves "sandwiching" fixed cells in an agarose case by a microcentrifugation system consisting of small-diameter cell-centrifugation tubes and subsequent processing of the cells by conventional techniques. The advantages offered by this method are discussed.     

Neuropathy Target Esterase: Rates of Turnover In Vivo Following Covalent Inhibition with Phenyl Di-n-Pentylphosphinate

Phenyl di-n-pentylphosphinate is a reasonably stable easily synthesized inhibitor of neuropathy target esterase (NTE) with low anticholinesterase activity. Like phenylmethylsulphonyl fluoride it protects hens against neuropathic effects of compounds such as diisopropylphosphorofluoridate. At intervals up to 15 days after dosing hens (10 mg/kg s.c. to inhibit 90% NTE) assays were made of catalytically active and of phosphinylated NTE in autopsy tissue. The sum of these components was always within the range of catalytic activity in undosed controls. However, the half-life of reappearance of active NTE was 2.07 days +/- 0.13 (SD, n = 6) for brain and 3.62 days +/- 0.23 (SD, n = 6) for spinal cord--shorter than after dosing with phenylmethylsulphonyl fluoride. It is proposed that: (1) The physiological turnover mechanism cannot distinguish between catalytically active and di-n-pentylphosphinylated NTE although initiation of organophosphate-induced delayed polyneuropathy might involve recognition of aged di-alkyl-phosphorylated NTE as "foreign". (2) The short half-lives indicate a slow spontaneous dephosphinylation of inhibited NTE occurs in vivo as well as de novo synthesis. The difference in half-lives for brain and spinal cord NTE may be due to different rates of synthesis de novo or (more likely) to different rates of spontaneous reactivation of the inhibited NTE in the two tissues.     

Mechanisms of glucocorticoid function in human leukemic cells: analysis of receptor gene mutants of the activation-labile type using the covalent affinity ligand dexamethasone mesylate

In the cultured acute lymphoblastic leukemic (ALL) cell line, clones of sensitive cells are killed by receptor-occupying concentrations of glucocorticoids. In addition, several types of resistance have been identified. The types of resistance are r- (glucocorticoid binding site loss), ract/l (activation labile receptors) and r+ly- (defective lysis mechanism). The two types of receptor mutants have been examined for the presence and expression of the glucocorticoid receptor (GR) gene. Southern blot analysis, using a full-length cDNA probe for human GR, shows that the gene in both is grossly intact. Examination of the expression of the gene by Northern blots reveals the presence of normal, 7-kb message in both types of receptor mutants, though in amounts somewhat reduced from wild-type. This report focuses on the activation labile mutants. Since characterization of these mutants suggests that they can bind ligand but not retain it during activation, we hypothesized that they would respond normally to a ligand that could not be lost during activation. This seems to be the case. When the covalent affinity ligand dexamethasone mesylate, itself a partial glucocorticoid agonist/antagonist, is used, the ract/l cells are killed to an extent corresponding to that evoked by a sub-optimal concentration of the full agonist dexamethasone. We conclude: (1) that the ract/l receptors can function to kill cells if provided a ligand that they do not lose during activation; (2) that the partial agonist activity of dexamethasone mesylate for cell killing is not due to release of a small amount of free dexamethasone; (3) that the poor agonist activity of dexamethasone mesylate receptor complexes suggests that the role of steroid is strictly to participate in conversion of the receptor to its DNA binding form, after which presence of the steroid actually interferes with proper receptor action.     

Bioflavonoid interaction with rat uterine type II binding sites and cell growth inhibition

Competition analysis with a number of known bioflavonoids demonstrated that these compounds (luteolin, quercetin, pelargonin) compete for [3H]estradiol binding to cytosol and nuclear type II sites in rat uterine preparations. The inhibition of [3H]estradiol binding to type II sites was specific and these bioflavonoids did not interact with the rat uterine estrogen receptor. Since estradiol stimulation of nuclear type II sites in the rat uterus is highly correlated with cellular hypertrophy and hyperplasia, we assessed the effects of these compounds on the growth of MCF-7 human breast cancer cells in culture and on estradiol stimulation of uterine growth in the immature rat. The data demonstrated that addition of quercetin (5-10 micrograms/ml) to MCF-7 cell cultures resulted in a dose-dependent inhibition of cell growth (DNA/flask). This effect was reversible by removal of quercetin from the culture medium, or by the addition of 10 nM estradiol-17 beta to these cell cultures containing this bioflavonoid. Since estradiol-17 beta (10 nM) stimulated nuclear type II sites and proliferation of MCF-7 cells, we believe bioflavonoid inhibition of MCF-7 cell growth may be mediated through an interaction with nuclear type II sites. This hypothesis was confirmed by in vivo studies which demonstrated that injection of luteolin or quercetin blocked estradiol stimulation of nuclear type II sites in the immature rat uterus and this correlated with an inhibition of uterine growth (wet and dry weight). These studies suggest bioflavonoids, through an interaction with type II sites, may be involved in cell growth regulation.     

Calcitonin gene-related peptide in human fetal lung and in neonatal lung disease

The ontogeny of calcitonin gene-related peptide immunoreactivity (CGRP-IR) was evaluated immunohistochemically in 67 human fetal or newborn lungs previously analyzed for calcitonin immunoreactivity (CT-IR). CGRP-IR was present by 10 weeks of gestation in rare, solitary neuroendocrine (NE) cells of developing conducting airways in two of eight first-trimester lungs. During the second trimester, cells with CGRP-IR were found consistently (21/23 fetuses). However, the numbers of positively staining cells did not appear to increase in these fetuses or in third-trimester infants dying of non-pulmonary causes. The highest concentrations of CGRP-IR cells were seen in lungs of premature infants with advancing chronic lung disease associated with bronchopulmonary dysplasia (BPD). CGRP-IR was seen earlier in gestation and in greater numbers of NE cells than was calcitonin immunoreactivity (CT-IR) reported previously in these same fetal lungs (Lab Invest 52:52, 1985). Its presence paralleled that of CT-IR in postnatal chronic lung disease.     

Allometric shape change and heterochrony in the freeliving coral Trachyphyllia bilobata (Duncan)

Regression analysis has been used to study the relationship between age, size, shape, and surface area in two ancestral-descendant populations of the Neogene Caribbean coral Trachyphyllia bilobata. Analyses of the relationship between size and age show that the relationship is isometric and that little difference occurs between populations in mean corallite length or height and in their rates of growth. Onset of columella growth is significantly earlier, however, in the descendant population. Studies of the relationship between size and shape show that growth is allometric, with shape change occurring in both corallum elongation and pinching of the corallite wall during ontogeny. In the descendant population, pinching and elongation initiate earlier in the ontogeny of the coral. These results suggest that the evolutionary development of the meandroid form in freeliving corals has been accomplished by heterochrony, involving a complex set of disassociated peramorphic changes in ontogeny accompanied by paedomorphic changes in astogeny. Further analyses show that the observed heterochronic changes serve to decrease corallum surface area which may in turn enhance sediment removal and nutrition in unstable habitats.