Establishment of a DNA bank of human origin

High molecular weight DNA extracted from 50 unrelated parisian individuals constitutes a "DNA-thèque" which can be used at the population level. This procedure allows preparation from a variety of tissues such as liver, placenta and leucocytes. The "quality" of the DNA obtained (about 50 Kb, RNA and protein free) and the efficiency of the method (in terms of the yield of DNA obtained) are considered. Blood is the easiest material to obtain and permits the obtaining of a reasonable amount of human and primate DNA, particularly when familial investigation is planned for polymorphism studies. One of the objects of this article is to encourage others to make use of our bank, for studies in population and evolutionary genetics.     

Alpha-1-antitrypsin deficiency and juvenile liver disease Ultrastructural observations compared with light microscopy and routine liver tests

Sixteen children (aged between 1 month and 20 years) with alpha-1-antitrypsin deficiency (PiZ) were investigated by liver biopsy on one or more occasions. Eight patients had suffered from neonatal cholestasis, and two of them were investigated during the cholestatic period as well. The clinical status and liver function tests were compared with the light and electron microscopical findings. According to the light microscopical analyses at the latest investigation, the cholestatic and noncholestatic patients were classified as healthy, fibrotic or cirrhotic cases. All livers displayed periodic acid-Schiff positive, diastase-resistant globules in some but not all periportally located hepatocytes. By electron microscopy accumulation of retained secretory material was found in all PiZ patients. This accumulation was most conspicuous in the smooth endoplasmic reticulum of hepatocytes. alpha-1-antitrypsin deficiency seems to affect some, but not all hepatocytes. In the affected cells disappearance or hypotrophy of the Golgi complex could be observed. The intracellular transport of very low density lipoproteins (VLDL) was apparently not affected. The migration block in alpha-1-antitrypsin deficiency seems to occur before transportation to the Golgi complex. The extent of the involvement was not strictly age-dependent. There was no ultrastructural evidence of subclinical cholestasis as a possible triggering factor in the development of cirrhosis.     

Swelling and ion uptake in cat cerebrocortical slices:

Cat cerebrocortical slices incubating in medium containing normal K+ concentrations were exposed to a number of different transmitters. Norepinephrine, histamine and adenosine or 2-chloroadenosine caused increased swelling of the slices associated with an increased Na+ and Cl- content. These effects were seen only when both Cl- and HCO3- were present in the medium, and were inhibited by a number of anion transport inhibitors. These characteristics were identical to those of the HCO3(-)-dependent component of the swelling induced by high K+ levels in the medium. Other transmitters, namely 5-hydroxytryptamine, dopamine, and gamma-amino butyric acid, were ineffective. The effects of norepinephrine, histamine and 2-chloroadenosine were antagonised by propranolol and phentolamine, chlorpheniramine and diphenhydramine, and theophylline respectively. These antagonists also inhibited HCO3(-)-dependent, K+-stimulated swelling. The transmitters which induced swelling also stimulated the carbonic anhydrase activity of cerebrocortical slices. We conclude from these data that the HCO3(-)-dependent component of K+-stimulated swelling may be due to K+-stimulated release of transmitters. Furthermore, the fact that the transmitters which induce swelling have also been reported to be most effective in increasing cAMP content in both brain slices or cultured astrocytes is consistent with the swelling response being mediated via cAMP-induced changes and being predominantly localized to astrocytes.     

Plasmapheresis in the initial treatment of insulin-dependent diabetes mellitus in children.

Several factors indicate that autoimmune mechanisms may play a part in the aetiology of insulin-dependent diabetes mellitus. At the onset of the disease in 10 children (aged 11-16 years) plasmapheresis was performed four times over one to two weeks. Seventeen age-matched children with the same clinical features served as controls. The C-peptide concentrations at onset were the same in the two groups, but after one month the children treated with plasmapheresis had significantly higher values. This difference became even more pronounced after three, nine, and 18 months, both during fasting and at the maximum response to a standardised meal. The study group also had a significantly more stable metabolism, longer partial remission, and no higher insulin requirement. Of the 10 treated children islet-cell cytoplasmic antibodies were present in seven before plasmapheresis and in nine during treatment. The antibodies remained detectable in five and six out of nine patients at one and six months respectively after plasmapheresis. Although the mechanisms are obscure, plasmapheresis performed at the onset of insulin-dependent diabetes mellitus may help to preserve beta-cell function.     

Gene fusion vectors based on the gene for staphylococcal protein A

Two plasmid vectors, containing the gene coding for staphylococcal protein A and adapted for gene fusion, have been constructed. These vectors will allow fusion of any gene to the protein A gene, thus giving hybrid proteins which can be purified, in a one-step procedure, by IgG affinity chromatography. As an example of the practical use of such vectors, the protein A gene has been fused to the lacZ gene of Escherichia coli. E. coli strains containing such plasmids produce hybrid proteins with both IgG binding and β-galactosidase activities. The hybrid protein(s) can be immobilized on IgG-Sepharose by its protein A moiety with high efficiency without losing its enzymatic activity and they can be eluted from the column by competitive elution with pure protein A. The fused protein(s) also binds to IgG-coated microtiter wells which means that the in vivo product can be used as an enzyme conjugate in ELISA tests.     

The effect of proteoglycans of cartilage and over-sulphated polysaccharides on the development of calcium-hydroxy-apatite (CHA) crystal formation in vitro

A coulometric model system is described which facilitates the quantitative study of the kinetics of transformation of amorphous calcium phosphate (ACP) into calcium-hydroxy-apatite (CHA) crystals. Proteoglycans of high molecular weight and over-sulphated polysaccharides (Arteparon, dextran sulphate) delayed CHA crystal formation. The results have enabled us to characterize the structure activity relationship of inhibitors of CHA formation, and to postulate a general structural requirement for molecules with inhibitory effect. As working mechanism, binding of calcium ions by sulphate groups of polyanions was supposed, which might reversibly impair "the critical nuclei formation", and/or further deposition of calcium ions in the CHA crystals. The clinical, therapeutical significance of the determination of the threshold concentration of different compounds is discussed.