Estimation of the size of freshwater ciliate populations by a sub-sampling technique

A technique is described for the estimation of the size of ciliate populations based on replicated counts. Sample drops were taken by automatic pipettes of different volume from activated sludge and from a small oligotrophic stream. Specific aims were: 1) the estimation of the number of species; 2) the estimation of the number of individual in each species; 3) the selection of a suitable sub-sample size and number of replicates. For each volume the cumulative increase in species taken in successive sub-samples was determined. For each species the minimum permissible sub-sample volume and the number of replicates required for an allowable error of 10% and 25% were determined. Lastly, the relationship between the mean number of individuals counted for each species and the relative coefficient of variation was visualized.     

Reactions of sugar nitro-alkenes with acetoacetic esters

3,4,5,6,7-Penta-O-acetyl-1,2-dideoxy-1-nitro-d-gluco-hept-1-enitol reacts with methyl acetoacetate in an unusual Michael reaction, giving the normal adduct (6), and a bicyclic derivative (9) that arises from quasi-dimerization of the former when a high concentration of the base is used. Acetylation of compound 9 gives the hydroxylamine O-acetate (10).From the reactions of 3,4,5,6,7-penta-O-acetyl-1,2-dideoxy-1-nitro-d-galacto-hept-1-enitol with ethyl and tert-butyl acetoacetate, the normal adducts (7 and 8) were isolated. The structures of compounds 6 to 10 were established on the basis of their spectral properties (u.v., i.r., mass, and ¹H- and ¹³C-n.m.r.)     

Amplification of lymph node cell tube leukocyte adherence inhibition (LAI) reactivity by leukocyte adherence inhibition factor (LAIF)

This investigation examines the immunologic basis for specific antigen-induced tube leukocyte adherence inhibition (LAI) reactivity of draining lymph node cells (LNC) from dogs with canine transmissible venereal sarcoma (CTVS). CTVS regressor LNC, macrophage-depleted LNC, and enriched T lymphocyte fractions, but not enriched B lymphocyte fractions, were specifically reactive to CTVS antigen extract in direct tube LAI. In addition, regressor LNC amplified tube LAI responses by generating supernatants with leukocyte adherence inhibition factor (LAIF) activity for normal dog indicator LNC and enriched peripheral blood mononuclear cells (PBMC) in an indirect tube LAI assay. However, macrophage-depleted LNC and enriched T lymphocyte fractions failed to generate supernatants with LAIF activity, suggesting that macrophage accessory cells play a central role in the amplification of tube LAI. Interestingly, CTVS regressor peripheral blood leukocytes (PBL) and PBMC, which were specifically reactive in direct tube LAI, also failed to generate supernatants with LAIF activity. These findings demonstrate a distinction between LAIF-mediated amplification and direct tube LAI reactivity, and suggest that leukocyte populations with differing cellular proportions and from different immunologic compartments may participate in tube LAI via different mechanisms.     

Differential effects of positive and negative proliferative stimuli on murine cytolytic and helper T-cell clones

Studies were performed to determine whether substances could be identified which exhibited differential regulatory effects--either positive or negative--on the growth of murine alloreactive cytolytic (Tc) and helper (Th) cloned T-cell lines. The following lines of evidence suggested that Tc and Th proliferate in response to the same growth factor (GF). (1) When GF-containing fluids from cultures of phorbol myristic acetate (PMA)-activated EL4 thymoma were fractionated by a variety of biochemical techniques. Tc and Th eluted together. (2) Absorption of GF-containing supernatants with either cloned Tc or cloned Th depleted GF activity for each to a similar extent, and GF eluted from either Tc or Th to which it had adsorbed supported the proliferation of Tc and Th equally well. (3) Lectin-depleted supernatants from cultures of concanavalin A (Con A)-activated Th stimulated the proliferation of Th as well as Tc. (4) Recombinant human interleukin (IL-2) supported the growth of Tc and Th with equal efficiency. On the other hand, the following observations indicated that Tc and Th differed in their responses to inhibitors of GF-driven proliferation. (1) Con A at greater than or equal to 0.3 micrograms/ml inhibited the GF-driven proliferation of each of three Th lines but not either of two Tc lines. To the contrary, Con A enhanced GF-dependent proliferation of Tc. (2) Like Con A, allogeneic splenocytes selectively depressed GF-driven proliferation of Th but not Tc. (3) A substance generated during the acid elution of GF from cells, possibly a modified fetal calf serum component, greatly reduced the GF-driven proliferation of Tc but not Th. These results suggest that differential control of the proliferation of Tc and Th in cellular immune responses may be achieved via negative regulatory signals and raise the possibility that substances which can selectively depress the proliferation of specific T-cell subsets might be found which would be of therapeutic value.     

Phospholipid Biosynthesis in Myelin: Presence of CTP: Phosphoethanolamine Cytidylyltransferase in Purified Myelin of Rat Brain

Highly purified myelin from rat brain was previously shown to contain the ethanolaminephosphotransferase which completes the synthesis of phosphatidyl ethanolamine. We have now obtained evidence for the presence in myelin of CTP:phosphoethanolamine cytidylyltransferase, the enzyme catalyzing formation of CDP-ethanolamine. Myelin was isolated by two different procedures, one based on the Norton-Poduslo method and the other involving repetitive gradients with osmotic shocking deferred to the end. The fact that activity remained constant through all but the earliest steps suggested that the enzyme is intrinsic to myelin. Comparison of subcellular fractions revealed that approximately half the total activity was in the supernatant, the remainder being distributed among the particulate fractions. Relative specific activity of myelin was 27-31% that of microsomes, thus eliminating the possibility of appreciable contamination by the latter. The possibility of adsorption of the soluble enzyme by myelin was rendered unlikely by retention of activity after washing the myelin with buffered sodium chloride or sodium taurocholate. Furthermore, relative specific activity of the cytidylyltransferase was 10-fold higher than that of lactate dehydrogenase (a cytosolic marker) in myelin. The apparent Km for CTP was approximately the same for myelin and microsomes, but that for phosphoethanolamine was significantly higher for myelin.     

The phytate and mineral content of some cereals,cereal products,legumes, legume products,snack bars,and nuts available in New Zealand

Analyses for phytate by an indirect precipitation method and for the minerals calcium (Ca), zinc (Zn), iron (Fe), copper (Cu), and manganese (Mn) by atomic absorption spectrophotometry were carried out on 100 foods available in New Zealand. Foods with 1% phytate (dry weight basis) included untoasted muesli, rolled oats, wheat germ, wheat bran, soybean, and some soy products. Most breads contained between 0.35 and 0.60% phytate; legumes on average had 0.62% phytate, as did snack bars. There was a wide variation in Ca and Zn contents: There was a tenfold variation in Ca content among the legume products, whereas there was a seventyfold variation in Zn content among the cereals. The phytate: Zn molar ratio, which is presumed to indicate the biovailability of Zn, was above 20∶1 for two-thirds of the cereals and almost all of the snack bars; it was above 15∶1 for one-third of the breads, almost all of the legumes, and half of the legume products. These high phytate: Zn molar ratios, as well as some Ca: phytate molar ratios above 6∶1, indicate that there might be a reduced biovailability of Zn in many of the foods analyzed in this study.     

Dye coupling in the organ of Corti

Summary Dye-coupling in an in vitro preparation of the supporting cells of the guinea-pig organ of Corti was evaluated by use of the fluorescent dyes, Lucifer Yellow, fluorescein and 6 carboxyfluorescein. Despite the presence of good electrical coupling in Hensen cells (coupling ratios >0.6) the spread of Lucifer yellow was inconsistent. Hensen cells are very susceptible to photoinactivation, i.e., cell injury upon illumination of intracellular dye; and this in conjunction with Lucifer Yellow's charge and K+-induced precipitability may account for its variability of spread. Fluorescein and 6 carboxyfluorescein, on the other hand, spread more readily and to a greater extent than Lucifer Yellow, often spreading to cell types other than those of Hensen. Dye spread is rapid, occurring within a few minutes. These results suggest that molecules of metabolic importance also may be shared by the supporting cells of the organ of Corti.     

Characterization of chlorophyll fluorescence quenching in chloroplasts by fluorescence spectroscopy at 77 K II. ATP-dependent quenching

In intact, uncoupled type B chloroplasts from spinach, added ATP causes a slow light-induced decline ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{≈ 3}\text{min}$) of chlorophyll a fluorescence at room temperature. Fluorescence spectra were recorded after fast cooling to 77 K and normalized with fluorescein as an internal standard. Related to the fluorescence quenching at room temperature, an increase in Photosystem (PS) I fluorescence (F₇₃₅) and a decrease in PS II fluorescence (F₆₉₅) were observed in the low-temperature spectra. The change in the $\text{F}{}_{735}\text{F}{}_{695}$ ratio was abolished by the presence of methyl viologen. Fluorescence induction at 77 K of chloroplasts frozen in the quenched state showed lowered variable (F_v) and initial (F₀) fluorescence at 690 nm and an increase in F₀ at 735 nm. The results are interpreted as indicating an ATP-dependent change of the initial distribution of excitation energy in favor of PS I, which is controlled by the redox state of the electron-transport chain and, according to current theories, is caused by phosphorylation of the light-harvesting complex.     

Isolation of a brain peptide identical to the intestinal PHI (peptide HI)

The isolation of a brain peptide identical to the intestinal peptide PHI (peptide HI) is described. The peptide was isolated from porcine brain extract using a chemical assay method based on its C-terminal isoleucine amide structure. The complete amino acid sequence of the peptide was found to be: His-Ala-Asp-Gly-Val-Phe-Thr-Ser-Asp-Phe-Ser-Arg-Leu-Leu-Gly-Gln-Leu-Ser-Ala- Lys-Lys-Tyr-Leu-Glu-Ser-Leu-Ile-NH2. This sequence is identical to the intestinal peptide thus demonstrating PHI to be a brain-gut peptide. The role of PHI in the central nervous system as a neurotransmitter or neuromodulator is discussed.     

Monoclonal antibody-directed immunopurification and identification of cytochromes P-450

A 28 amino acid peptide with diuretic and natriuretic activity has been purified from rat atrial muscle. The primary structure of this atrial peptide is H-Ser-Leu-Arg-Arg-Ser-Ser-Cys-Phe-Gly-Gly-Arg-Ile-Asp-Arg-Ile-Gly- (sequence in text) Ala-Gln-Ser-Gly-Leu-Gly-Cys-Asn-Ser-Phe-(Arg)-Tyr-OH. The biological activity of this peptide is identical to that of atrial natriuretic factor and cardionatrin I isolated from rat atria.