Transduction of human colony-stimulating factor-1 (CSF-1) receptor into interleukin-3-dependent mouse myeloid cells induces both CSF-1-dependent and factor-independent growth.

A retroviral vector encoding the receptor for human colony-stimulating factor-1 (CSF-1) was introduced into murine myeloid FDC-P1 cells which require interleukin-3 (IL-3) for their proliferation and survival in culture. Cells expressing the CSF-1 receptor (CSF-1R), selected by fluorescence-activated cell sorting in the continued presence of murine IL-3, formed colonies in semisolid medium and were able to proliferate continuously in liquid cultures containing human recombinant CSF-1. Thus, although they do not synthesize endogenous murine CSF-1R, FDC-P1 cells express the downstream components of the CSF-1 mitogenic pathway necessary for its signal-response coupling. After receptor transduction, slowly proliferating factor-independent variants that produced neither CSF-1 nor growth factors able to support the proliferation of parental FDC-P1 cells also arose. When the human CSF-1R was expressed in FDC-P1 cells under the control of an inducible metallothionein promoter, the frequencies of both CSF-1-responsive and factor-independent variants increased after heavy-metal treatment. In addition, a monoclonal antibody to human CSF-1R arrested colony formation by both the CSF-1-dependent and factor-independent cells but did not affect their growth in response to IL-3. Therefore, the induction of both the CSF-1-dependent and factor-independent phenotypes depended on expression of the transduced human CSF-1R.     

Tandem linkage of human CSF-1 receptor (c-fms) and PDGF receptor genes

A 5' untranslated exon of the human CSF-1 receptor gene (c-fms) is separated by a 26 kb intron from the 32 kb receptor coding sequences. Nucleotide sequence analysis of cloned genomic DNA revealed that the 3' end of the PDGF receptor gene is located less than 0.5 kb upstream from this exon. Similarities in chromosomal localization, organization, and encoded amino acid sequences suggest that the genes encoding the CSF-1 and PDGF receptors arose through duplication. The as yet unidentified c-fms promoter/enhancer sequences may be confined to the nucleotides separating the two genes or could potentially lie within the PDGF receptor gene itself.     

Genomic organization, chromosomal localization, and independent expression of human cyclin D genes.

Murine cDNA clones for three cyclin D genes that are normally expressed during the G1 phase of the cell cycle were used to clone the cognate human genes. Bacteriophage and cosmid clones encompassing five independent genomic loci were partially sequenced and chromosomally assigned by an analysis of somatic cell hybrids containing different human chromosomes and by fluorescence in situ hybridization to metaphase spreads from normal peripheral blood lymphocytes. The human cyclin D1 gene (approved gene symbol, CCND1) was assigned to chromosome band 11q13, cyclin D2 (CCND2) to chromosome band 12p13, and cyclin D3 (CCND3) to chromosome band 6p21. Pseudogenes containing sequences related to cyclin D2 and cyclin D3 mapped to chromosome bands 11q13 and 6p21, respectively. Partial nucleotide sequence analysis of exons within each gene revealed that the authentic human cyclin D genes are more related to their mouse counterparts than to each other. These genes are ubiquitously transcribed in human tumor cell lines derived from different cell lineages, but are independently and, in many cases, redundantly expressed. The complex patterns of expression of individual cyclin D genes and their evolutionary conservation across species suggest that each family member may play a distinct role in cell cycle progression.     

Usefulness of linked DNA probes for prenatal diagnosis of cystic fibrosis: report of a case in a 1:4 risk pregnancy

Prenatal diagnosis of cystic fibrosis was performed with linked DNA probes in a couple with a 1:4 risk. The limits and the future of molecular prenatal diagnosis are discussed.     

A specific immunological probe for the major myelin proteolipid. Confirmation of a deletion in DM-20

Major myelin proteolipid (MMPL, also called PLP) and DM-20 are the two major intrinsic membrane proteins of CNS myelin. A specific immunological probe was obtained for MMPL by raising antibodies against the synthetic tridecapeptide 117-129 of MMPL. Antibodies against this peptide reacted with the MMPL but did not cross react with DM-20, while both proteolipids had been shown previously to be recognized by antibodies directed against the C-terminal hexapeptide of MMPL. This is in accordance with previous findings showing that DM-20 differs only from MMPL by a deletion of residues 100-140 (+/- few units). Furthermore, this site-specific immunological probe also recognizes MMPL in its native form in oligodendrocytes in primary glial cell cultures.     

Primary structure determination of five sialylated oligosaccharides derived from bronchial mucus glycoproteins of patients suffering from cystic fibrosis. The occurrence of the NeuAc alpha(2----3)Gal beta(1----4)[Fuc alpha(1----3)] GlcNAc beta(1----.) structural element revealed by 500-MHz 1H NMR spectroscopy

The structure of sialylated carbohydrate units of bronchial mucins obtained from cystic fibrosis patients was investigated by 500-MHz 1H NMR spectroscopy in conjunction with sugar analysis. After subjecting the mucins to alkaline borohydride degradation, sialylated oligosaccharide-alditols were isolated by anion-exchange chromatography and fractionated by high performance liquid chromatography. Five compounds could be obtained in a rather pure state; their structures were established as the following: A-1, NeuAc alpha(2----3)Gal beta(1----4) [Fuc alpha(1----3)]GlcNAc beta(1----3)Gal-NAc-ol; A-2, NeuAc alpha(2----3)Gal beta(1----4)GlcNAc beta(1----6)-[GlcNAc beta (1----3)]GalNAc-o1; A-3, NeuAc alpha(2----3)Gal beta-(1----4)[Fuc alpha(1----3)]GlcNAc beta(1----3)Gal beta(1----3) GalNAc-o1; A-4, NeuAc alpha(2----3)Gal beta(1----4)[Fuc alpha(1----3)]Glc-NAc NAc beta(1----6)[GlcNAc beta(1----3)]GalNAc-o1; A-6,NeuAc alpha-(2----3) Gal beta(1----4)[Fuc alpha(1----3)]GlcNAc beta(1----6)[Gal beta-(1----4) GlcNAc beta(1----3)]GalNAc-o1. The simultaneous presence of sialic acid in alpha(2----3)-linkage to Gal and fucose in alpha(1----3)-linkage to GlcNAc of the same N-acetyllactosamine unit could be adequately proved by high resolution 1H NMR spectroscopy. This sequence constitutes a novel structural element for mucins.     

Cortisol response in heifers to artificial insemination, natural mating, and no mating at estrus

Eight mixed-breed dairy heifers were used in a random block design to study the stress-producing effect of various types of mating on heifers. Treatments consisted of artificial insemination, natural mating, or not mating at estrus. Stress was evaluated by measuring blood plasma cortisol. Blood samples were collected by indwelling jugular catheters at 60 and 30 min before treatment; immediately before treatment (0 min); and 5, 15, 30, 60, 120 and 180 min after treatment. Means for cortisol were not significant for treatment or treatment across time. Mean cortisol response to the artificial insemination, natural mating and no mating treatments were 5.36, 8.05 and 5.56 ng/ml, respectively. These results suggest that the use of artificial insemination does not impose and added stress at estrus.