Essential fatty acid deficiency lowers the activity of the acetylated low density lipoprotein receptor of rat peritoneal macrophages.

We compared phospholipid fatty acid composition, cholesterol ester accumulation, and receptor-mediated binding, internalization, and degradation of acetylated low-density lipoprotein (acetyl-LDL) in cultured peritoneal macrophages from rats fed an essential fatty acid deficient or control diet. The deficient diet increased the 5,8,11-eicosatrienoic acid and decreased the omega 6 fatty acid content of macrophage phospholipid relative to control. The deficient diet did not affect macrophage uptake of [1-14C]oleate; however, it lowered the accumulation of intracellular labelled cholesteroyl oleate to 66% of the control. This effect was attributed to a diminution of the specific binding of acetyl-LDL, and not to acetyl-LDL internalization nor to degradation. The results demonstrate the sensitivity of the acetyl-LDL receptor to changes in its membrane environment, brought about through dietary means.     

Inhibition of human immunodeficiency virus replication in acutely infected CD4+ cells by CD8+ cells involves a noncytotoxic mechanism.

The mechanism by which CD8+ T cells from human immunodeficiency virus (HIV)-infected individuals suppress HIV replication in acutely infected CD4+ T cells was investigated. Cytotoxicity was not involved, as the antiviral activity of the CD8+ cells did not correlate with the ability to lyse HIV-infected or uninfected CD4+ T cells. In addition, the frequency of HIV-infected CD4+ cells increased during coculture with CD8+ T cells even in the absence of detectable levels of virus replication. Moreover, separation of the CD4+ and CD8+ cells by a 0.4-micron-pore-size filter delayed HIV replication, indicating a role, at least in part, for a soluble factor. However, cell contact was required for optimal antiviral activity. These results extend further the observation on the mechanism of antiviral HIV activity by CD8+ cells from infected individuals. They support the conclusion that CD8+ cells can play a major role in preventing development of disease in HIV-infected individuals.     

Several CD4 domains can play a role in human immunodeficiency virus infection in cells.

The human immunodefiency virus (HIV) uses the human CD4 glycoprotein as a receptor for infection of susceptible cells. Cells expressing a series of mutated forms of the CD4 gene have shown a variability in their ability to support replication of three HIV type 1 (HIV-1) and three HIV-2 strains. Moreover, when different stages of virus production were examined by a variety of assays, a consistent delay was observed in all cell lines containing CD4 mutants compared with those with intact full-length CD4. Cells expressing the CD4.415 mutant (modified at the serine 415 corresponding to a phosphorylation site of the cytoplasmic domain) showed only a minimal effect on virus replication. Cells expressing CD4.403 and CD4.401 mutants (lacking the whole cytoplasmic domain) manifested a moderate delay in production of virus progeny. The most substantial effect on HIV replication was observed in cells expressing a chimeric hybrid containing sequences corresponding to the first 177 residues of the N-terminal CD4 fused to CD8 sequences encoding the hinge, transmembrane, and cytoplasmic domains of the human CD8. Furthermore, in a cell-to-cell contact assay, fusion was absent when the CD4 proximal membrane domain was replaced by the CD8 counterpart. In addition, a strong correlation between the down-modulation of the surface CD4 and HIV expression was observed. These observations suggest that in addition to the known binding region, other domains of CD4 could play an important role in regulating HIV entry of cells.     

Effects of antiplatelet agents on platelet aggregation induced by platelet--activating factor (PAF) in human whole blood.

Impedance aggregometry was used to evaluate the potency of anti-platelet agents on Platelet Activating Factor (PAF)--induced platelet aggregation in citrated human whole blood. Drugs were tested for ability to inhibit maximum aggregation to PAF. Dose response curves were obtained and the concentration of drug producing 50% inhibition of maximum aggregation (ED50) determined. ED50's (microM) for specific PAF antagonists WEB 2086, Ro 19-3704, FR-900452, BN 52021, L-652,731, CV 3988, WEB 2118 and 48740 RP are: 0.39, 2.4, 4.7, 19.5, 21.0, 5.32, 161.0, 924.0, respectively. ED50's for non-specific PAF antagonists, diltiazem, propranolol, ketotifen, procaine HCL, and lidocaine HCL are: 38.0, 56.0, 250.0, 513.0 and 768.0, respectively. Ibuprofen was inactive at 2300 microM. Results are consistent with concept that there are specific receptors on platelets mediating PAF-induced aggregation in whole blood. Aggregation is inhibited potently by specific and competitive PAF receptor antagonists. Whole blood aggregometry may be a valid method for predicting in vivo activity of PAF antagonists.     

Cancer: improving early detection and prevention. A community practice randomised trial.

OBJECTIVE--To test the impact of physician education and facilitator assisted office system interventions on cancer early detection and preventive services. DESIGN--A randomised trial of two interventions alone and in combination. SETTING AND SUBJECTS--Physicians in 98 ambulatory care practices in the United States. INTERVENTIONS--The education intervention consisted of a day long physician meeting directed at improving knowledge, attitudes, and skills relevant to cancer prevention and early detection. The office system intervention consisted of assistance from a project facilitator in establishing routines for providing needed services. These routines included division of responsibilities for providing services among physicians and their staff and the use of medical record flow sheets. MAIN OUTCOME MEASURES--The proportions of patients provided the cancer prevention and early detection services indicated annually according to the US National Cancer Institute. RESULTS--Based on cross sectional patient surveys, the office system intervention was associated with an increase in mammography, the recommendation to do breast self examination, clinical breast examination, faecal occult blood testing, advice to quit smoking, and the recommendation to decrease dietary fat. Education was associated only with an increase in mammography. Record review for a patient cohort confirmed cross sectional survey findings regarding the office system for mammography and faecal occult blood testing. CONCLUSION--Community practices assisted by a facilitator in the development and implementation of an office system can substantially improve provision of cancer early detection and preventive services.     

Assay of inorganic sulfate in biologic fluids by nonsuppressed (single-column) ion chromatography

An assay using nonsuppressed (single-column) anion chromatography was developed to determine the concentration of inorganic sulfate in biologic fluids. A conventional HPLC system with an anion-exchange column and conductimetric detector interfaced with an automatic injector and integrator was used. The mobile phase for the chromatography of urine and serum samples is 4 mM potassium hydrogen phthalate, pH 4.5, and potassium iodide is used as the internal standard. For cerebrospinal fluid samples, the mobile phase is modified by addition of 10% of a 4 mM phthalic acid solution. Results of the HPLC assay were found to correlate well (r = 0.991 and 0.999) with those of two commonly used spectrophotometric methods for urine and serum inorganic sulfate determinations. However, the concentrations determined by ion chromatography were 2.5 to 10% lower, possibly due to less assay interference by other substances following chromatographic separation of sulfate. Anion chromatography using a single-column system is a convenient and relatively inexpensive method with sufficient sensitivity for the determination of inorganic sulfate concentrations in urine, serum, and cerebrospinal fluid.     

Neurohypophysial hormones of the 1-month-old bovine fetus: absence of vasotocin during mammal development

The neurohypophysial hormones of the 1-month-old bovine fetus have been identified by their positions in ion-exchange chromatography and their retention times in high-pressure reverse-phase partition chromatography. Arginine vasopressin and oxytocin have been recognized. The molar ratio vasopressin/oxytocin in neurohypophysis is about 6 in the 1-month-old fetus compared with 4 in the 3-month-old fetus, 2.7 in the 7-month-old fetus and 1 in the adult. Vasotocin is virtually absent even in the early fetus (less than 0.1% of arginine vasopressin). The occurrence of a vasotocin gene expressed in the fetus but silent in the adult appears unlikely.     

Evidence that TET protein functions as a multimer in the inner membrane of Escherichia coli.

The inner membrane TET (TetA) protein, which is involved in Tn10-mediated microbial tetracycline resistance, consists of two domains, alpha and beta, both of which are needed for tetracycline resistance and efflux (M.S. Curiale, L.M. McMurry, and S.B. Levy, J. Bacteriol. 157:211-217, 1984). Since tetracycline-sensitive mutants in one domain can partially complement sensitive mutants in the other domain and since some sensitive mutants show dominance over the wild type, a multimeric structure for TET in the membrane had been suggested. We have studied this possibility by using tetA-phoA gene fusions. We fused all but the last 40 base pairs of the tetA gene with the carboxy terminus of the phoA gene for alkaline phosphatase (PhoA), whose activity requires its dimerization in the periplasm. The tetA-phoA fusion protein was under control of the tetracycline-inducible regulatory system for the tetA gene. Induction led to the synthesis of a 78,000-dalton inner membrane protein. Tetracycline resistance was expressed at reduced levels, consistent with the terminal beta domain deletion. Alkaline phosphatase activity was also present, but at low levels, suggesting that some, but not all, of the fusion proteins had their carboxy-terminal ends in the periplasm. When wild-type or mutant TET proteins were present in the same cell with the fusion protein, the tetracycline resistance level was affected (raised or lowered); however, phosphatase activity was reduced only when TET proteins with intact or near-intact beta domains were present. These findings suggest that TET functions as a multimer and that intact beta domains, on TET molecules in the heterologous multimer, either allow fewer PhoA moieties to project into the periplasm or sterically hinder PhoA moieties from dimerizing.     

Survival of Escherichia coli with and without ColE1::Tn5 after aerosol dispersal in a laboratory and a farm environment.

The survival of a laboratory strain and a naturally occurring fecal strain of Escherichia coli, with and without a Tn5-containing derivative of ColE1, was examined after aerosol dispersal in a laboratory office and a barn under ambient temperature and humidity conditions. Following the release of paired strains, air and diverse types of surfaces were assayed for the test organisms. In both environments, the number of airborne bacteria declined rapidly within the first 2 h. Longer survival was found on surfaces and varied with surface type: recovery was greatest from wood products. Organisms persisted for 1 day in the office and for up to 20 days in the barn. Survival of the fecal strain was better than that of the laboratory strain in both test environments. In general, plasmid-bearing strains fared similarly to their plasmidless parents, but in several comparisons the ColE1::Tn5-containing strain showed enhanced survival. These studies have implications for the present and proposed release of genetically engineered organisms with and without plasmid vectors.