Pregnancy hormone concentrations in marijuana users

The maternal serum concentrations of human chorionic gonadotropin, pregnancy-specific beta-l-glycoprotein, placental lactogen, progesterone, 17-hydroxyprogesterone, estradiol and estriol were measured in 13 women who smoked marijuana regularly throughout pregnancy. Cannabinoid use in these women was confirmed by RIA measurements of their sum Δ ⁹- tetrahydrocannabinol (THC) concentrations. These THC using women were matched within $\text{2}\text{1}\text{2}$ weeks of gestational age with 13 pregnant non-THC using controls drawn from the same population. Placental protein and steroid hormone concentrations were within established normal ranges for gestational age and there were no significant differences between the groups in the concentrations of any of the protein and steroids measured. In addition, no significant differences between THC users were found following linear regression analysis of placental hormone concentrations as a function of gestational age. Thus, this study suggests that marijuana use during pregnancy does not significantly alter the circulating maternal concentrations of trophoblastic protein hormones or major fetoplacental steroid hormones.     

The beta-major and beta-minor globin genes in murine erythroleukemia cells replicate during the same early interval of the S phase

The time of replication in S phase was determined for the 7.3 kb EcoRI segment containing the beta-major globin gene and the 14 kb EcoRI segment containing the beta-minor globin gene in a murine Friend erythroleukemia virus transformed (MEL) cell line. Cells were obtained from different intervals of S phase by centrifugal elutriation to avoid artifacts of chemical synchronization. Newly synthesized DNA from different parts of S phase were obtained by isolation of 5-bromouracil (BU) labelled DNA from these cells. The BU-DNA synthesized during four different intervals of S was transferred to diazobenzyloxymethyl (DBM) paper and hybridized with a beta-globin cDNA probe. Quantitation showed that both beta-globin segments were replicated in the first quarter of the S phase with no significant difference in their time of replication in this MEL cell line.     

T cell responses specific for subregions of allogeneic MHC molecules.

The repertoire of C3H (H-2k) CD4+ T cells for I-Ab allopolymorphisms was analyzed by studying the responses of unprimed populations of T cells and of I-Ab-specific T cell clones for recombinant MHC molecules containing combinations of polymorphic subregions of the alpha- and beta-chains from the I-Ab and I-Ak molecules. In this system, polymorphisms in the predicted MHC alpha-helices were more potent than polymorphisms in the beta-strands in stimulating unprimed alloreactive T cells. Similarly, 75% of I-Ab-specific T cell clones responded to recombinants containing b polymorphisms in both the alpha- and beta-chains helices and tolerated the substitution of k polymorphisms in the beta-pleated sheet. Furthermore, 20% of the clones responded to a molecule containing allogeneic b residues in just the beta-chain helix. The results demonstrate that the T cell response to allogeneic MHC molecules consists largely of sets of T cells with overlapping specificities for subregions of the MHC molecule. In addition, they highlight the importance of the alpha-helices in these responses and a diminished role for polymorphisms in the beta-strands when, as in the present case, MHC structure and conformation is tolerant of beta-sheet substitutions. These results sharply contrast with observations made in the analysis of Ag-specific T cells and lead to the suggestion that a subset of alloreactive T cells are not peptide specific and can directly recognize MHC polymorphisms.     

Three-dimensional structure at 5 Å resolution of cytosolic aspartate transaminase from chicken heart

An X-ray study of orthorhombic crystals of cytosolic aspartate transaminase from chicken heart has been carried out at 5 Å resolution. The crystals belong to space group P2₁2₁2₁, with unit cell dimensions $\text{a = 62.7}\text{A}\text{?}\text{, b = 118.1}\text{A}\text{?}\text{, c = 124.5}\text{A}\text{?}$. The electron density map has been calculated on the basis of five heavy-atom derivatives. The model of the molecule derived from this map revealed clearly two subunits of similar structure related by a non-crystallographic dyad. The secondary structure of the protein comprises nine helical segments per subunit.The enzyme has been shown to be catalytically active in the crystal form. Removal of the coenzyme from the crystals made it possible to derive from the difference Fourier map the position of the active site in the enzyme molecule.Significant conformational changes have been observed which accompany the interconversion of intermediates of the enzymic reaction.     

B and T lymphocyte attenuator-mediated signal transduction provides a potent inhibitory signal to primary human CD4 T cells that can be initiated by multiple phosphotyrosine motifs

The B and T lymphocyte attenuator (BTLA) is a recently identified member of the CD28 family of cell receptors. Initial reports demonstrated that mice deficient in BTLA expression were more susceptible to experimental autoimmune encephalomyelitis, indicating that BTLA was likely to function as a negative regulator of T cell activation. However, cross-linking of BTLA only resulted in a 2-fold reduction of IL-2 production, questioning the potency with which BTLA engagement blocks T cell activation. We established a model in which BTLA signaling could be studied in primary human CD4 T cells. We observed that cross-linking of a chimeric receptor consisting of the murine CD28 extracellular domain and human BTLA cytoplasmic tail potently inhibits IL-2 production and completely suppresses T cell expansion. Mutation of any BTLA tyrosine motifs had no effect on the ability of BTLA to block T cell activation. Only mutation of all four tyrosines rendered the BTLA cytoplasmic tail nonfunctional. We performed structure-function studies to determine which factors recruited to the BTLA cytoplasmic tail correlated with BTLA function. Using pervanadate as a means to phosphorylate the BTLA cytoplasmic tail, we observed both Src homology protein (SHP)-1 and SHP-2 recruitment. However, upon receptor engagement, we observed only SHP-1 recruitment, and mutations that abrogated SHP-1 recruitment did not impair BTLA function. These studies question whether SHP-1 or SHP-2 have any role in BTLA function and caution against the use of pervanadate as means to initiate signal transduction cascades in primary cells.     

SecA2 functions in the secretion of superoxide dismutase A and in the virulence of Mycobacterium tuberculosis

Tuberculosis remains a severe worldwide health threat. A thorough understanding of Mycobacterium tuberculosis pathogenesis will facilitate the development of new treatments for tuberculosis. Numerous bacterial pathogens possess specialized protein secretion systems that are dedicated to the export of virulence factors. Mycobacterium tuberculosis is part of a developing group of pathogenic bacteria that share the uncommon property of possessing two secA genes (secA1 and secA2). In mycobacteria, SecA1 is the essential 'housekeeping' SecA protein whereas SecA2 is an accessory secretion factor. Here we demonstrate that SecA2 contributes to the pathogenesis of M. tuberculosis. A deletion of the secA2 gene in M. tuberculosis attenuates the virulence of the organism in mice. By comparing the profile of proteins secreted by wild-type M. tuberculosis and the DeltasecA2 mutant, we identified superoxide dismutase A (SodA) as a protein dependent on SecA2 for secretion. SodA lacks a classical signal sequence for protein export. Our data suggests that SecA2-dependent export is a new type of secretion pathway that is part of a virulence mechanism of M. tuberculosis to elude the oxidative attack of macrophages.     

Activation of chloride secretion in cystic fibrosis cells and tissues by the substituted imidazole SRI 2931

Recent interest in nucleotides and related agents as part of clinical trials in cystic fibrosis (CF) therapy have elicited efforts to identify novel compounds capable of activating transepithelial chloride (Cl(-)) transport in CF cells and tissues. From a library of nucleosides, bases, and other substituted heterocycles, 341 compounds were screened for their ability to activate anion transport in CF cells grown on permeable supports. One compound, SRI 2931, was found to confer prolonged and potent activity when administered to the apical surfaces of CF pancreatic epithelial cells, primary CF nasal epithelial cells, non-CF human colonic epithelial cells, and intact tissue taken from mouse models for CF. Concentrations of SRI 2931 (20 microM), which activated Cl(-) transport, had minimal effect on cell proliferation. SRI 2931 was not calcium (Ca(2+)) or cAMP dependent, suggesting important differences from conventional chloride secretagogues. The compound selectively released ATP from the apical, but not basolateral, surfaces of CF cells grown on permeable supports. The magnitude, longevity, and mechanism of action of the response provide a tool for dissecting pathways of epithelial ATP extracellular signaling and Cl(-) permeability.