The influence of postnatal nutritional deprivation on the phospholipid content of developing rat lung.

It has been previously reported that fasting may result in decreased lung surfactant production. In order to investigate this relationship and the role of nutrition in lung phospholipid synthesis, 21-day-old rats were exposed for 60 h to one of five dietary regimens: standard rat chow (controls), fasting, pure glucose, pure fat, or pure protein. After the period of fasting there was a 33% decrease in lung protein content, but there was no change in DNA content. Exposure to any of the experimental diets resulted in a decrease in tissue total phospholipid and phosphatidylcholine content per lung, but not per unit lung protein. Similarly lung lavage phospholipid and phosphatidylcholine content was decreased by 25% after fasting when expressed per lung or per unit DNA, but not per unit protein. Pulmonary cholinephosphotransferase (EC 2.7.8.2) activity was decreased in the fasted animals and those fed the protein diet, but not in the glucose or fat-fed animals. The activities of acetyl-CoA carboxylase (EC 6.4.1.2) and microsomal fatty acid elongation were decreased in all the experimental groups except for the glucose-fed group. It is concluded that fasting results in a decrease in lung cell size but not in lung cell number. Total phospholipid and phosphatidylcholine content in lung tissue and lung lavage is decreased per cell but not per unit cell mass.     

Purinoceptor agonists stimulate phosphatidylcholine secretion in primary cultures of adult rat type II pneumocytes

We examined the effect of purinoceptor agonists on phosphatidylcholine secretion in primary cultures of type II pneumocytes from adult rats. Surfactant is a major product of the type II cell and phosphatidylcholine is its principal component. Adenosine, AMP, ADP and ATP stimulated phosphatidylcholine secretion in a concentration-dependent manner. At the optimum concentration (1 mM), adenosine and AMP stimulated phosphatidylcholine secretion more than 2-fold, while ATP stimulated 5-fold and ADP almost 7-fold. Because of the magnitude of the response it is tempting to speculate that secretion of surfactant may be under purinoceptor regulation. None of these agents influenced cellular phosphatidylcholine synthesis or lactate dehydrogenase release into the medium, so the effects were primarily on secretion and were not secondary to effects on synthesis or cell damage. Non-metabolizable analogs of adenosine, 5'-N-ethyl-carboxyamidoadensoine (NECA) and L-N6-phenylisopropyladenosine (L-PIA), stimulated secretion to the same extent as adenosine and the effect of NECA was antagonized by 8-phenyltheophylline, suggesting a P1 purinoceptor-mediated mechanism. The stimulatory effect of ATP was diminished by alpha, beta-methylene ATP but only slightly by 8-phenyltheophylline, suggesting that, although part of the ATP effect could be explained by catabolism to adenosine, the P2 purinoceptor may also be involved in regulation of surfactant secretion.     

Epstein-Barr virus BZLF1 trans-activator specifically binds to a consensus AP-1 site and is related to c-fos.

Two regions of the Epstein-Barr virus BZLF1 trans-activator protein have sequence similarity to the c-fos protein. Part of the similarity corresponds to the region of c-fos which is similar to the DNA binding domain of c-jun and GCN-4. The structure of the exon which contains this region in c-fos and BZLF1 is also highly conserved between the two genes. Complete BZLF1 protein and a C terminal fragment were prepared either as purified fusion proteins or by in vitro translation from a BZLF1 cDNA. Gel retardation and DNase footprinting assays using these proteins show that BZLF1 is a sequence specific DNA binding protein capable of binding to a target sequence which contains a consensus AP-1 site.     

Molecular cloning of a cDNA for human pregnancy-specific beta 1-glycoprotein:homology with human carcinoembryonic antigen and related proteins

Human pregnancy-specific beta 1-glycoprotein (SP1) plays an essential role in normal pregnancy. It is also a well-characterized oncodevelopmental antigen, expressed aberrantly by all trophoblastic tumors and some other malignant cell types. Here we report the identification of a human placental cDNA encoding the SP1 polypeptide sequence. The coding sequence shows 95% identity at the nucleotide level with a distinct, recently published SP1 cDNA sequence (PSG16). Unexpectedly, the sequence is also highly homologous to the published sequence of human carcinoembryonic antigen (CEA). SP1, CEA and CEA-related nonspecific cross-reacting species thus belong to a group of closely related though antigenically diverse tumor-associated glycoproteins. Comparison of the deduced amino acid sequence of the SP1 cDNA with that of CEA provides insight into the modular nature of these related proteins. This may have implications for the genomic organization and evolution of the CEA gene family.     

Immunization with a vaccinia virus recombinant expressing herpes simplex virus type 1 glycoprotein D: long-term protection and effect of revaccination.

Previously we showed that mice immunized with a vaccinia virus vector expressing the herpes simplex virus type 1 (HSV-1) glycoprotein D (gD) gene (vaccinia/gD) were protected against both lethal and latent infections with HSV-1 for at least 6 weeks after immunization (K. J. Cremer, M. Mackett, C. Wohlenberg, A. L. Notkins, and B. Moss, Science 228:737-740, 1985). In the experiments described here, we examined long-term immunity to HSV following vaccinia/gD vaccination, the effect of revaccination with vaccinia/gD, and the impact of previous immunity to vaccinia virus on immunization with the gD recombinant. Mice immunized with vaccinia/gD showed 100, 100, and 80% protection against lethal infection with HSV-1 at 18, 44, and 60 weeks postimmunization, respectively. Protection against latent trigeminal ganglionic infection was 70, 50, and 31% at 6, 41, and 60 weeks postvaccination, respectively. To study the effect of reimmunization on antibody levels, mice vaccinated with vaccinia/gD were given a second immunization (booster dose) 3 months after the first. These mice developed a 10-fold increase in neutralizing-antibody titer (221 to 2,934) and demonstrated a significant increase in protection against lethal HSV-1 challenge compared with animals that received only one dose of vaccinia/gD. To determine whether preexisting immunity to vaccinia virus inhibited the response to vaccination with vaccinia/gD virus, mice were immunized with a recombinant vaccinia virus vector expressing antigens from either influenza A or hepatitis B virus and were then immunized (2 to 3 months later) with vaccinia/gD. These mice showed reduced titers of neutralizing antibody to HSV-1 and decreased protection against both lethal and latent infections with HSV-1 compared with animals vaccinated only with vaccinia/gD. We conclude that vaccination with vaccinia/gD produces immunity against HSV-1 that lasts over 1 year and that this immunity can be increased by a booster but that prior immunization with a vaccinia recombinant virus expressing a non-HSV gene reduces the levels of neutralizing antibody and protective immunity against HSV-1 challenge.     

Spatial and temporal organization of calcium signalling in hepatocytes.

Treatment of hepatocytes with agonists which act via the second messenger inositol 1,4,5-trisphosphate (Ins(1,4,5)P3), results in increases of cytosolic free Ca2+ [( Ca2+]i) which are manifest as a series of discrete [Ca2+]i transients or oscillations. With increasing agonist dose [Ca2+]i oscillation frequency increases and the initial latent period decreases, but the amplitude of the [Ca2+]i oscillations remains constant. Studies of these [Ca2+]i oscillations at the subcellular level have indicated that the [Ca2+]i changes do not occur synchronously throughout the cell, but initiate at a specific subcellular domain, adjacent to a region of the plasma membrane, and then propagate through the cell as a [Ca2+]i wave. For a given ceil, the locus of [Ca2+]i wave initiation is constant for every oscillation in a series and is also identical when the cell is sequentially stimulated with different agonists or when the phospholipase C-linked G protein is activated directly using AIF4-. The kinetics of the [Ca2+]i waves indicate that a Ca(2+)-activated mechanism is involved in propagating the oscillatory [Ca2+]i increases throughout the cell, and the data appear to be most consistent with a process of Ca(2+)-induced Ca2+ release. It is proposed that the ability to propagate [Ca2+]i oscillations into regions of the cell distal to the region in which the signal transduction apparatus is localized could serve an important function in allowing all parts of the cell to respond to the stimulus.     

Oxygenated cholesterols synergistically immobilize acyl chains and enhance protein helical structure in human erythrocyte membranes

Fourier transform infrared spectroscopy revealed that insertion of 20 alpha-hydroxycholesterol into human erythrocyte membranes (10% of total membrane sterol) immobilized the lipid acyl chains to a degree equivalent to enriching total membrane cholesterol by 50% (Rooney, M.W., Lange, Y. and Kauffman, J.W. (1984) J. Biol. Chem. 259, 8281-8285). Raman spectroscopy showed that the amount of acyl chain rotamers was not significantly altered by the presence of 20 alpha-hydroxycholesterol, indicating that acyl chain immobilization was limited to an inhibition of lateral motion. The presence of 20 alpha-hydroxycholesterol may synergistically enhance the acyl-chain-immobilizing behavior of membrane cholesterol. In addition, protein helical structure was not altered by 20 alpha-hydroxycholesterol. The insertion of 7 alpha-hydroxycholesterol into erythrocyte membranes resulted in an increase in protein helical structure which was comparable to that observed for erythrocyte membranes enriched with pure cholesterol by 50%. However, both acyl chain mobility and conformation were unchanged. These results suggest a synergistic behavior between oxysterols and cholesterol in modifying erythrocyte membrane packing.     

Interaction of fatty acids with lipid bilayers

We present a method by which it is possible to describe the binding of fatty acids to phospholipid bilayers. Binding constants for oleic acid and a number of fatty acids used as spectroscopic probes are deduced from electrophoresis measurements. There is a large shift in $\text{p}\text{K}$ value for the fatty acids on binding to the phospholipid bilayers, consistent with stronger binding of the uncharged form of the fatty acid. For dansylundecanoic acid, fluorescence titrations are consistent with the binding constants derived from the electrophoresis experiments. For 12-(9-anthroyloxy)stearic acid, fluorescence and electrophoresis data are inconsistent, and we attribute this to quenching of fluorescence at high molar ratios of 12-anthroylstearic acid to phospholipid in the bilayer.