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S Temple

Publications and source records attributed to S Temple.

10 recordsLinked to original sources
PubMed1992

Rat embryonic septal neurons survive and express cholinergic properties in isolation and without nerve growth factor.

We studied survival and expression of cholinergic properties in embryonic septal neurons grown in very low density microcultures (1-7 cells per Terasaki well). Even in cultures containing only a single neuron, at least 10% of plated neurons survived for 2 weeks or more in medium containing fetal calf serum or an acid-stable fraction (55,000 Da) of horse serum. Of these surviving neurons, 30-40% stained positively for acetylcholinesterase (AChE) or nerve growth factor (NGF) receptor, even though the culture medium lacked detectable levels of NGF, brain-derived neurotrophic factor, and fibroblast growth factor. Addition of NGF or an antibody against NGF had no effect on either neuronal survival or the percentage of neurons staining positively for AChE or NGF receptor after 18-20 days in vitro. There was no cell division in medium containing the serum fraction, but when 10% fetal calf serum was present cell division occurred in some of the cultures, and in half of these cases at least one of the clonal progeny became AChE-positive. These results demonstrate that some embryonic septal cells can survive at least 2 weeks and develop cholinergic neuronal properties in the absence of other cells or NGF.

Acetylcholinesterase↗
PubMed1989

Division and differentiation of isolated CNS blast cells in microculture.

The mechanism of transformation of the overtly similar cells of the neural plate into the numerous and diverse cell types of the mature vertebrate central nervous system (CNS) can better be understood by studying the clonal development of isolated CNS precursor cells. Here I describe a culture system in which blast cells (cells capable of division) isolated from embryonic day 13.5-14.5 rat forebrain can divide and differentiate into a variety of clonal types. Most clones contain only neurons or glia; 22% contain both neurons and non-neuronal cells. For the division of blast cells, live conditioning cells need to be present indicating that environmental signals influence proliferation. Heterogeneous clones develop in homogeneous culture conditions, so factors intrinsic to the blast cells are probably important in determining the number and type of clonal progeny.

Animals↗
PubMed1988

Pulmonary toxicity of trichloroethylene: induction of changes in surfactant phospholipids and phospholipase A2 activity in the mouse lung.

Trichloroethylene (TCE) is a common organic solvent in use as a dry cleaning agent as well as an inhalant anesthetic. Nevertheless the effects of this material on the pulmonary surfactant which prevents alveolar collapse at maximal expiration is not known. Therefore, we have examined the effect of TCE on the intra- and extracellular surfactant pools and the activity of phospholipase A2, an enzyme which controls the remodeling of phosphatidylcholine to dipalmitoylphosphatidylcholine, the primary constituent of the pulmonary surfactant. Male CD-1 mice were treated ip with 2500 or 3000 mg/kg TCE. Twenty-four hours later mice were anesthetized and the lungs lavaged. Mice were then killed, the lungs perfused and excised, and subcellular fractions including lamellar bodies prepared. Some lungs were prepared for ultrastructural examination. Phospholipase A2 was assayed in all subcellular fractions. Phospholipid was assayed in the lavage (extracellular surfactant) and the lamellar bodies (intracellular surfactant). TCE (2500 mg/kg) caused selective exfoliation of Clara cells. However, only the dose of 3000 mg/kg TCE produced a significant decrease in the intracellular surfactant phospholipid. Minimal changes occurred in the phospholipid profiles. Phospholipase A2 specific activity was significantly decreased at both dosages within the lung microsomal fraction. In addition after treatment with 3000 mg/kg TCE the enzyme activity in the lamellar body fraction was significantly increased. These data suggest that inhalation of TCE may damage the enzymes which are responsible for synthesizing the pulmonary surfactant resulting in lower amounts of surfactant being stored and available for secretion into the alveolus.

Animals↗
PubMed1988

Characterization and comparison of the role of beta-agonists on in vivo and in vitro surfactant-related phospholipid synthesis and secretion by fetal rabbit lung and isolated type II alveolar cells.

The role of beta-adrenergic stimulation in surfactant synthesis and secretion was investigated in the fetal lung. Fetuses were treated with isoxsuprine or saline on gestational day 24 by ip injection. Three days later the fetal lungs were lavaged and intracellular surfactant was isolated on a sucrose gradient. Concurrently undifferentiated type II alveolar cells were isolated from 24-day fetal rabbit lung and grown in vitro. In the in vivo portion of the study, examination of surfactant pool sizes revealed that only saline treatment produced a significant elevation in tissue-stored or secreted surfactant compared to untreated controls. Isoxsuprine appeared to inhibit the saline-induced increase. In the case of the intracellular surfactant, the phosphatidylcholine content per gram of lung was significantly increased after saline treatment. In vitro response of isolated type II alveolar cells to isoxsuprine was dependent on prior incubation of the cells for 24 h with conditioned medium. Isoxsuprine stimulated a dose-dependent decrease in the intracellular stores of radioactively labeled DSPC after 24 h of exposure to the drug. A corresponding increase in labeled DSPC in the culture medium was observed. Forth-eight hours after exposure to the drug, those cells that had secreted the highest level of DSPC displayed the highest levels of renewed synthesis of DSPC. This study indicates that the immature fetal lung can be induced to synthesize surfactant-related phospholipid by the stress of laparotomy and/or drug administration. Short-term exposure to beta-agonists is insufficient to stimulate secretion of surfactant stores. In contrast, isolated type II alveolar cells exposed to isoxsuprine respond by secreting DSPC.

Adrenergic beta-Agonists↗
PubMed1987

Profile of phospholipase A2 activity in subcellular fractions and lamellar bodies of developing, neonatal and adult rabbit lung. Correlation with intracellular levels of disaturated phosphatidylcholine.

Phospholipase A2 activity was determined in subcellular fractions and lamellar bodies of fetal, neonatal and adult rabbit lungs. Specific activity in most fractions decreased from the 24th to the 28th day of gestation. All fractions except the mitochondrial and the nuclear fractions exhibited a sharp increase in activity in the newborn lung. Specific activity in the adult lung generally declined in comparison to neonatal values. During gestation total enzyme activity per gram of lung was concentrated in the cytosolic fraction. With the exception of the lamellar body fraction, the total content of phospholipase A2 activity increased dramatically in all fractions from the neonatal lung. The lamellar body fractions displayed both low specific activity and low total enzyme activity during gestation. Specific activity increased dramatically in the neonatal and adult lung but still accounted for only a small fraction of the activity in comparison to the other subcellular fractions. The subcellular content of disaturated phosphatidylcholine (PC) appeared to correlate well with the activity of phospholipase A2 in the neonatal mitochondrial, microsomal and cytosolic fractions. Since decreasing prenatal enzyme levels are associated with increasing disaturated PC content, the alkaline and calcium-dependent phospholipase A2 may not be directly involved in disaturated PC synthesis in the fetus. However, postnatally, the correlation between the pattern of production of disaturated PC and the activity of the phospholipase A2 indicates a role for this enzyme in surfactant-related disaturated PC synthesis.

Animals↗
PubMed1987

Characterization and distribution of a phospholipase A2 activity from adult rabbit lung.

A phospholipase A2 activity was characterized in adult rabbit lung. This activity was calcium- and deoxycholate-dependent and displayed an alkaline pH optimum. Km and Vmax were 0.176 mM and 256.8 pmoles/min./mg protein respectively. The microsomal fraction displayed the highest enzymatic specific activity; the lowest activity was present in the cytosol. Yet this latter fraction accounted for the majority of the total activity. Although the specific activity was high within the lamellar body fraction this compartment contained only approximately 2% of the total activity. Phospholipase A2 activity was inhibited by bromophenacyl bromide, chlorpromazine and mepacrine in decreasing order of effectiveness. Treatment of the microsomes with increasing concentrations of NaC1 indicated that the lung phospholipase A2 activity was relatively loosely bound to the microsomal membranes and was maximally removed with salt at a concentration only slightly higher than physiological. Addition of calmodulin to the enzyme assay did not significantly alter hydrolysis of labelled phosphatidylcholine.

Animals↗
PubMed1986

Clonal analysis of oligodendrocyte development in culture: evidence for a developmental clock that counts cell divisions.

The clonal development of oligodendrocytes was studied by culturing individual oligodendrocyte--type-2 astrocyte (O-2A) progenitor cells on monolayers of type-1 astrocytes, which stimulate O-2A progenitor cells to divide. Oligodendrocytes developed by a proliferative lineage in which clonal progeny differentiated together after a number of cell divisions. Most O-2A progenitor cells had similar cell cycle times (1-2 days), but their proliferative capacity varied greatly: some divided only once while others divided up to eight times before differentiating. sister cells behaved similarly when recultured separately on astrocyte monolayers. These findings are consistent with the cell-division-counting hypothesis previously proposed to explain the timing of oligodendrocyte differentiation. They also unambiguously establish the phenotype of O-2A progenitor cells in vitro and demonstrate that these cells respond directly to growth factors produced by type-1 astrocyte monolayers.

Animals↗
PubMedDate not supplied

Differentiation of a bipotential glial progenitor cell in a single cell microculture.

Although it is known that most cells of the vertebrate central nervous system (CNS) are derived from the neuroepithelial cells of the neural tube, the factors determining whether an individual neuroepithelial cell develops into a particular type of neurone or glial cell remain unknown. A promising model for studying this problem is the bipotential glial progenitor cell in the developing rat optic nerve; this cell differentiates into a particular type of astrocyte (a type-2 astrocyte) if cultured in 10% fetal calf serum (FCS) and into an oligodendrocyte if cultured in serum-free medium. As the oligodendrocyte-type-2 astrocyte (0-2A) progenitor cell can differentiate along either glial pathway in neurone-free cultures, living axons clearly are not required for its differentiation, at least in vitro. However, the studies on 0-2A progenitor cells were carried out in bulk cultures of optic nerve, and so it was possible that other cell-cell interactions were required for differentiation in culture. We show here that 0-2A progenitor cells can differentiate into type-2 astrocytes or oligodendrocytes when grown as isolated cells in microculture, indicating that differentiation along either glial pathway in vitro does not require signals from other CNS cells, apart from the signals provided by components of the culture medium. We also show that single 0-2A progenitor cells can differentiate along either pathway without dividing, supporting our previous studies using 3H-thymidine and suggesting that DNA replication is not required for these cells to choose between the two differentiation programmes.

Animals↗