Studies on an antineoplastic fraction from human urine. Characterization of the major protein in this fraction.

A fraction has been isolated from human urine which exhibits antiproliferative activity against human tumour cell lines without affecting the growth of several normal diploid cell lines or tumour cells of mouse or hamster origin. The major protein present in this fraction has been characterized and tentatively designated antineoplastic urinary protein (ANUP). An S020,W value of 3.69 S was obtained by sedimentation velocity analysis, and a subunit molecular mass of 16 300 Da was obtained by sedimentation equilibrium and by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Centrifugation data also indicated that the protein self-associates. The amino acid analysis of ANUP was consistent with its low pI (4.2) as determined by chromatofocusing analysis. Furthermore, the amino acid composition exhibited some features similar to collagen, as shown by high levels of proline and glycine, the absence of cysteine, and the presence of low levels of hydroxyproline.     

白蜡虫泌蜡期天敌群落的研究

选择有代表性的１０个白蜡主产地,研究了各地白蜡虫天敌群落的物种组成和群落结构,并进行了聚类分析和排序。结果表明,白蜡白泌蜡期天敌群落共有捕食和寄生性天敌１８种,各群落的优势种和物种变化较小,群落性程度较高,相似性系数最高达９５．１２％,最低为７３．８６％。影响群落相似性的主要因子为产区,地理位置,捕食性瓢虫的种类和数量。     

Studies on the mechanisms of action of BCG

Summary Bacillus Calmette-Guérin was given intravenously (i.v.) or subcutaneously (s.c.), 14 days before antigenic or mitogenic stimulation.When given i.v., BCG increased plaque-forming cell production against thymus-dependent antigens (sheep red blood cells and a hapten coupled to hemocyanin) and against a thymus-independent antigen (the same hapten coupled to polymerized flagellin). BCG given s.c. was generally ineffective in potentiating humoral responses.In contrast, a marked depression of spleen cell reactivity in vitro to a T-cell mitogen, concanavalin A (Con A) but an increased response to a B-cell mitogen, dextran sulfate, were observed after BCG i.v., and to a lesser extent after BCG s.c. This inhibitory effect was reversed after removal of cells adherent to nylon or plastic. Normal spleen cell reactivity to Con A was strongly depressed when the cells were cultivated in presence of unfractionated or plastic-adherent BCG-treated spleen cells, suggesting that BCG induces suppressor cells which inhibit T-cell responses in vitro. Similar impairment of T-cell functions in vivo was not observed since BCG given i.v. significantly increased the intensity of delayed-type hypersensitivity to picryl chloride.BCG also induced macrophage activation, measured by macrophage tumoricidal activity in vitro, more strongly and more rapidly when given i.v. than s.c.Modifications of the distribution of lymphocytes from the circulation to the organs were observed. An early increase of lymphocyte migration to the spleen and a long-lasting depression of cell migration to mesenteric lymph nodes and bone marrow were seen after BCG given i.v. When given s.c., BCG induced a progressive increase of cell migration to the draining lymph nodes concomitant with a depression of migration to mesenteric nodes.These results demonstrate the complexity of the mode of action of BCG, but many of its effects on immune responses may result from a primary action on macrophages.     

Studies on the compartmentation of lipid in adipose cells. I. Subcellular distribution, composition, and transport of newly synthesized lipid: liposomes

The subcellular distribution and composition of endogenously synthesized lipid in isolated white adipose cells were studied to determine the nature and extent of lipid compartmentation. After brief incubation of cells with labeled glucose, acetate, or palmitic acid, over 90% of newly synthesized triglyceride was localized in the bulk-lipid phase, indicating rapid intracellular transport and storage. From 13 to 20% of the newly formed lipid was diglyceride, and over 95% of it was localized in the central lipid-storage vacuole rather than in organelle systems concerned with esterification, thus indicating intracellular segregation of newly synthesized partial glycerides. Most of the newly synthesized phosphatides partitioned with membranous organelles. Synthesis of cholesterol or cholesteryl ester was negligible. After brief incubation of cells with labeled glucose, the relative specific activity of organelle triglyceride was mitochondria > microsomes > liposomes > soluble supernatant > bulk lipid. In pulse-chase studies the specific activity of organelle triglyceride decreased and that of the bulk fraction increased reflecting intracellular lipid transport. The data suggest that a significant proportion of newly formed lipid is transferred from mitochondrial membranes into the storage vacuole by direct lipid-lipid interaction. Liposomes, which consist of small enclosed lipid droplets resembling chylomicrons, contained triglycerides of specific activity similar to microsomal triglyceride. While the evidence that liposome triglyceride may be microsomal in origin is indirect, the results do indicate that the liposome fraction represents a phase in the transport and(or) storage of new glyceride. At least two forms of compartmentation of newly synthesized lipids occurred. The first, termed "structural," refers to localization of lipids to organelle fractions. The second type of compartmentation, termed "chemical," concerns the intracellular segregation of a specific lipid class. The accumulation and segregation of newly synthesized diglyceride in the bulk storage pool are examples of the latter form of compartmentation.     

Studies on phospholipase C from Pseudomonas aureofaciens. I. Purification and some properties of phospholipase C.

Phospholipase C (phosphatidylcholine cholinephosphohydrolase, EC 3.1.4.3) from Pseudomonas aureofaciens was purified 3600-fold from the culture filtrate with a recovery of 1.6%. Purification was performed with the useof (NH4)2SO4 precipitation, Sephadex G-100 gel filtration and by ion-exchange chromatography on DEAE-Sephadex A-50 and CM-Sephadex C-50. The purified enzyme appeared to be homogeneous as revealed by polyacrylamide disc gel electrophoresis at pH 9.3. The molecular weight was estimated to be 35 000 by gel filtration on Sephadex G-75. Under our experimental conditions, phosphatidylethanolamine was more rapidly hydrolysed than phosphatidylcholine. Lyso forms of these two phosphatides were poor substrates. Phosphatidylserine, phosphatidylglycerol, phosphatidylinositol, cardiolipin and sphingomyelin were not hydrolysed. The enzyme activity with phosphatidylcholine as substrate was slightly stimulated by Ca2+, Mg2+, and Mn2+. However, these cations inhibited the activity with phosphatidylethanolamine as substrate. An anionic detergent, sodium deoxycholate, slightly enhanced the activity when phosphatidylcholine and phosphatidylethanolamine were used as substrates. A cationic detergent, cetyltrimethylammonium bromide, inhibited enzyme activity. EDTA and o-henanthroline inhibited the activity of the enzyme to a marked degree.     

Fluid lipid fraction in rod outer segment membrane.

Rod outer segment membrane is analyzed using the spin resonance label technique by means of two probes. The solubility of the first label,2,2,6,6-tetramethylpiperidin-1-oxyl, is correlated with the membrane fluidity which is measured using a stearic acid spin probe. The two values are compared to the solubility-fluidity relationship which characterizes a model system in which all lipids are in a fluid state. The analysis leads to the conclusion that only two thirds of the membrane lipids are fluid. This conclusion is reinforced by the observation that partial lipid removal leaves rigid lipids associated with the rhodopsin molecules.     

Studies of pesticide effects on Chlorella metabolism I. Effect of herbicides on complex lipid biosynthesis

Various herbicides were tested for possible effects on biosynthesisof complex lipids in Chlorella using ¹⁴C-U-acetate as a precursor.Among the herbicides tested, Hill reaction inhibitors causedinhibition of ¹⁴C-incorporation by 60 to 80 per cent at a concentrationof 5 mg/liter, and noticeable alteration in the labeling patternof glyco- and phospholipids. Proportions of mono- and digalactolipidsmarkedly decreased while those of phosphatidyl ethanolamineand phosphatidyl glycerol increased. In terms of net synthesis,phosphatidyl ethanolamine was affected, if at all, only slightlyby Hill reaction inhibitors, while the other complex lipidsdecreased to various degrees. (Received February 9, 1973; )