Antigenic study of the differentiation of the human kidney using monoclonal antibodies

The production of monoclonal antibodies against human embryonic renal cells allowed to display on the adult human kidney some antigens typical of certain structures or tissues: the proximal convoluted tubule for EG 9-11 and EG 19-6 monoclonal antibodies, the glomerular basement membrane for EG 14-1, the urothelium for EE 24-6, the connective tissue for EK 8-1 and EK 17-1 and probably the capsular and tubular basement membranes for EK 8-1. Simultaneously, we could follow the spatial and temporal repartition of the antigens during the renal development. One of them (EI 16-1) seemed to disappear in the adult and might correspond to a foetal type-antigen.     

Physical Mapping of 49 Microsatellite Markers on Chromosome 19 and Correlation with the Genetic Linkage Map

We have regionally localized 49 microsatellite markers developed by Généthon using a panel of previously characterized somatic cell hybrids that retain fragments from chromosome 19. The tight correlation observed between the physical and the genetic orders of the microsatellites provide cytogenetic anchorages to the genetic map data. We propose a position for the centromere just above D19S415, from the study of two hybrids, each of which retains one of the two derivatives of a balanced translocation t(1;19)(q11;q11). Microsatellites, which can be identified by a standard PCR protocol, are useful tools for the localization of disease genes and for the establishment of YAC or cosmid contigs. These markers can also judiciously be used for the characterization of new hybrid cell line panels. We report such a characterization of 11 clones, 8 of which were obtained by irradiation-fusion. Using the whole hybrid panel, we were able to define the order of 12 pairs of genetically colocalized microsatellites. As examples of gene mapping by the combined use of microsatellites and hybrid cell lines, we regionally assigned the PVS locus between the 19q13.2 markers D19S417 and D19S423 and confirmed the locations of fucosyltransferase loci FUT1, FUT2, and FUT5.     

Five specificity patterns of (1----3)-alpha-L-fucosyltransferase activity defined by use of synthetic oligosaccharide acceptors. Differential expression of the enzymes during human embryonic development and in adult tissues.

The use of synthetic trisaccharides as acceptors led to the definition of five main (1----3)-alpha-L-fucosyltransferase activity patterns in human adult tissues: (I). Myeliod cells, granulocytes, monocytes, and lymphoblasts, transfer an alpha-L-fucopyranosyl group to O-3 of a 2-acetamido-2-deoxy-D-glucosyl residue of H blood-group Type 2 oligosaccharide [alpha-L-Fucp-(1----2)-beta-D-Galp-(1----4)-beta-D-GlcpNAc----R] with Mn2+ as activator. (II) Brain has the same acceptor specificity pattern as myeloid cells, but can also use Co2+ as activator. (III) Plasma and liver transfer an alpha-L-furopyranosyl group to H blood-group Type 2 and to sialyl-N-acetyllactosamine [alpha-NeuAc-(2----3)-beta-D-Galp-(1----4)-beta-D-GlcpNAc----R]. (IV) Intestine, gall bladder, kidney, and milk have the same activity as (III), but also transfer an alpha-L-fucopyranosyl group to O-4 of a 2-acetamido-2-deoxy-D-glucose residue of H blood-group Type 1 [alpha-L-Fucp-(1----2)-beta-D-Galp-(1----3)-beta-D-GlcpNAc----R] and sialyl Type 1 [alpha-NeuAc-(1----3)-beta-D-Galp-(1----3)-beta-D-GlcpNAc----R]. (V) Stomach mucosa is not able to use sialyl-N-acetyllactosamine, but can transfer an alpha-L-fucopyranosyl group to the other Type 1 and Type 2 acceptors. Unlike in adult tissue, a single myeloid-like pattern of (1----3)-alpha-L-fucosyltransferase activity was found at early stages of development in all tissues tested. This embryonic enzyme is later progressively replaced by enzymes or mixtures of enzymes having the corresponding adult patterns of enzyme expression. All lymphoblastoid cell lines and half of the tumor epithelial cell lines tested expressed the myeloid-like pattern of enzyme found in normal embryonic tissues. The remaining tumor epithelial cell lines expressed different forms of (1----3/4)-alpha-L-fucosyltransferase acceptor specificity patterns.     

Isolation and characterization of human Lp-B lipoprotein containing apolipoprotein B as the sole apolipoprotein

A sequential immunoaffinity chromatography procedure was developed to isolate from whole normolipidemic human plasma a subpopulation of apoB containing particles (Lp-B) which is virtually free of non apoB protein. The absence of non apoB protein in Lp-B was assessed by enzyme immunoassay against apolipoproteins A-I, A-II, A-IV, E, C-III and (a). Electron microscopy and fractionation of the isolated particles by gel filtration demonstrated that these particles were heterogeneous in size. However, most of them had diameters between 18 and 26 nm. These particles were found to be rich in cholesterol (molar ratio cholesterol/apoB = 2246 +/- 995) poor in triacylglycerol (molar ratio triacylglycerol/apoB = 555 +/- 518) and had a phospholipids/apoB molar ratio of 713 +/- 348. Most of the cholesterol was esterified (66% +/- 5%). Lp-B particles bound to the apoB, E receptor of HeLa cells with a lower affinity than LDL prepared by ultracentrifugation (1.030 kg/l less than d less than 1.053 kg/l). (KD = 18.9 vs 10.5 nmol/l).