The effect of serum on lipid synthesis and on the expression of oligodendrocyte marker-enzymes in oligodendrocyte-enriched glial cells in culture

Glial cells were isolated from the cerebra of 7-day old rats and the effect of serum on the development of these cells in culture was studied. The activities of the oligodendrocyte marker-enzymes, 2′3′-cyclic nucleotide 3′-phosphodiesterase and glycerol 3-phosphate dehydrogenase and the synthesis of the myelin-associated sulpholipid, sulphatide, were used to monitor the differentiation of these cells in vitro. The results indicate that serum: (i) represses lipogenesis, cholesterogenesis and sulphatide synthesis, (ii) lowers the expression of 2′3′-cyclic nucleotide 3′ phosphodiesterase and glycerol 3-phosphate dehydrogenase but not of lactate dehydrogenase and (iii) thus impairs the differentiation of oligodendrocytes.     

A flexible method to align large numbers of biological sequences

Summary A method for the alignment of two or more biological sequences is described. The method is a direct extension of the method of Taylor (1987) incorporating a consensus sequence approach and allows considerable freedom in the control of the clustering of the sequences. At one extreme this is equivalent to the earlier method (Taylor 1987), whereas at the other, the clustering approaches the binary method of Feng and Doolittle (1987). Such freedom allows the program to be adapted to particular problems, which has the important advantage of resulting in considerable savings in computer time, allowing very large problems to be tackled. Besides a detailed analysis of the alignment of the cytochrome c superfamily, the clustering and alignment of the PIR sequence data bank (3500 sequences approx.) is described.     

Relation of changes in amount and type of dietary fat to fecapentaenes in premenopausal women

Correlation studies suggest that fecal mutagenicity is increased in groups eating high-fat diets, the same groups who are often found to have high colorectal cancer incidence and mortality. The fecapentaenes are the best characterized class of fecal mutagens, but the relationship of dietary fat intake to the excretion of these potent genotoxins is unknown. We studied the effect of changes in amount and type of dietary fat on fecapentaene levels in 31 premenopausal women 20-40 years of age who participated in a controlled feeding study. After a pre-diet free-living period lasting 1 menstrual cycle, women were placed on a high-fat (40% energy from fat) diet for 4 menstrual cycles and then switched to a low-fat (20% energy from fat) diet for an additional 4 menstrual cycles. One-half the subjects were maintained throughout the study at a ratio of polyunsaturated-to-saturated fatty acids (P/S ratio) of 1.0, the other half at 0.3; body weight was constant. All meals during the controlled diet periods were prepared at the Human Study Facility of the Beltsville Human Nutrition Research Center. Fecapentaene and fecapentaene precursor levels were measured in acetone extracts from 3-day pooled stool samples collected during the study. No differences in fecapentaene or precursor levels were observed between the high- and low-fat diets at either P/S ratio. Fecapentaene and precursor levels were higher while on controlled diets than during the pre-diet free-living period, and levels declined again in the post-diet free-living period. We conclude that dietary fat has no significant effect on fecapentaene or precursor levels in acetone extracts of stool in premenopausal women. The effect of other dietary or non-dietary factors on fecapentaenes remains unknown.     

Carbon and nitrogen partitioning in young nodulated pea (wild type and nitrate reductase-deficient mutant) plants exposed to NH4NO3

Carbon and nitrogen partitioning was examined in a wild-type and a nitrate reductase-deficient mutant (A317) of Pisum sativum L. (ev. Juneau), effectively inoculated with two strains of Rhizobium leguminosarum (128C23 and 128C54) and grown hydroponically in medium without nitrogen for 21 days, followed by a further 7 days in medium without and with 5 mM NH₄NO₃. In wild-type symbioses the application of NH₄NO₃ significantly reduced nodule growth, nitrogenase (EC 1.7.99.2) activity, nodule carbohydrates (soluble sugars and starch) and allocation of [¹⁴C]-labelled (NO₃⁻, NH₄⁺, amino acids) in roots. In nodules, there was a decline in amino acids together with an increase in inorganic nitrogen concentration. In contrast, symbioses involving A317 exhibited no change in nitrogenase activity or nodule carbohydrates, and the concentrations of all nitrogenous solutes measured (including asparagine) in roots and nodules were enhanced. Photosynthate allocation to the nodule was reduced in the 128C23 symbiosis. Nitrite accumulation was not detected in any case. These data cannot be wholly explained by either the carbohydrate deprivation hypothesis or the nitrite hypothesis for the inhibition of symbiotic nitrogen fixation by combined nitrogen. Our result with A317 also provided evidence against the hypothesis that NO₃⁻ and NH₄⁺ or its assimilation products exert a direct effect on nitrogenase activity. It is concluded that more than one legume host and Rhizobium strain must be studied before generalizations about Rhizobium /legume interactions are made.     

Antibodies to pollen exines

A polyclonal antiserum and monoclonal antibodies have been prepared to purified pollen exines of Calocedrus decurrens Florin. The location of the antigen is in the exine, as shown by light-and electron-microscopic immunocytochemistry. The greatest reduction in antibody binding follows treatment of the exine with chemicals known to alter sporopollenin. These results provide evidence that sporopollenin is antigenic. Exines of ten species of gymnosperms and angiosperms also bound the polyclonal antiserum, indicating similarity of sporopollenin structure.     

Agonist-stimulated oscillations and cycling of intracellular free calcium in individual cultured muscle cells

Receptor regulation of [Ca2+]i was monitored in individual BC3H-1 muscle cells with intracellularly trapped fura-2 using digital imaging analysis techniques. Activation of alpha 1-adrenergic or H1-histaminergic receptors resulted in multiple bursts, or oscillations, of elevated [Ca2+]i with an average interval frequency of approximately 1.8 min-1. The duration of oscillatory behavior was generally more prolonged in response to phenylephrine than in response to histamine. Additionally, a larger fraction of the cells responded with [Ca2+]i oscillations to phenylephrine (approximately 90%) than to histamine (approximately 60%), although the majority of cells produced oscillations in response to both agonists. In most cells, the receptor-mediated [Ca2+]i oscillations continued for several minutes in the absence of extracellular Ca2+, although the amplitude of the individual peaks gradually decreased. The activation of [Ca2+]i oscillations by H1-receptors was more dependent upon extracellular Ca2+ than those elicited by alpha 1-receptors, reflecting the greater dependency of the histaminergic response on Ca2+ influx. Readdition of Ca2+ to the incubation buffer resulted in the resumption of the [Ca2+]i oscillations. These results indicate that considerable cycling of Ca2+ between the cytoplasm and the endoplasmic reticulum must occur. Receptor-mediated [Ca2+]i oscillations were much more prevalent in subconfluent cells than in confluent cells, possibly due to increased coupling of the cells at higher densities. The cells were capable of responding independently of one another, since sister cells displayed unique temporal responses immediately following cell division. Thus, the linkage of receptor occupancy to [Ca2+]i elevation is a functionally unique property for each individual cell and can be influenced by epigenetic factors.     

Differential inhibition of indirect immunofluorescence in rat liver sections incubated with antibodies against microsomal cytochromes P-450a, b, and c and epoxide hydrolase

Rat liver sections were incubated with antibodies (100-1000 micrograms IgG/ml) against microsomal cytochromes P-450a, P-450b, and P-450c, and epoxide hydrolase. Inhibition of indirect immunofluorescence, which progressed with higher concentrations of primary antibody, corresponded with antigen-enriched tissue in frozen liver sections from male and female rats. It was found in liver sections from phenobarbital-treated rats incubated with anti-P-450b and anti-epoxide hydrolase and from 3-methylcholanthrene-treated rats incubated with anti-P-450c. No inhibition was found in sections from untreated rats or rats receiving treatments that did not induce the specific antigen. No inhibition was found in sections incubated with anti-P-450a. Inhibition of immunofluorescence was abolished in frozen sections subjected to dehydration-rehydration protocols known to extract antigens, and was prevented by certain solvents and detergent-wash. Inhibition of immunofluorescence provides a unique method for confirming the antigen-rich regions of the liver lobules specific for microsomal expoxide hydrolase and the cytochrome P-450s.     

Cutaneous laser-Doppler flowmetry: influence of underlying muscle blood flow

To find whether the measurement of skin blood flow (SkBF) by laser-Doppler flowmetry (LDF) is influenced by blood flow to underlying skeletal muscle, five subjects performed mild forearm exercise to induce a metabolic hyperemia in muscle in both forearms. This exercise consisted of alternative opening and closing of both hands at a frequency of approximately 1/s for a duration of 3 min. This exercise was performed twice by each subject. Forearm blood flow (FBF) by plethysmography increased from 2.64 +/- 0.49 (rest) to 31.11 +/- 9.95 ml.100 ml-1.min-1 (immediately after exercise) (P less than 0.001). No statistically significant postexercise increase was observed in LDF measured on the dorsal (110 +/- 21 to 105 +/- 21 mV) or ventral surface (266 +/- 113 to 246 +/- 77 mV) of the forearm. LDF measured from the chest also showed no significant change, indicating that the exercise was too mild to have reflex effects on SkBF. Moreover, the slope of the logarithmic linear regression and the half-time for recovery during the postexercise period for FBF were not reflected in LDF measurements from any of the three sites. We conclude that LDF measured from the skin surface is not influenced by blood flow to underlying skeletal muscle.