A panel of monoclonal antibodies to the 69 kDa glycosyl phosphatidylinositol anchored lymphocyte differentiation antigen ecto-5'-nucleotidase (ecto-5'-NT, CD73) was produced using highly purified human placental 5'-NT as immunogen. Antibodies 1E9.28.1 and 7G2.2.11 inhibit soluble placental 5'-NT activity and recognize lymphocyte CD73 in indirect immunofluorescence and immunoprecipitation assays. In addition, 1E9.28.1 induces vigorous T cell proliferation in the presence of submitogenic doses of phorbol myristate and F(ab')2 goat anti-mouse Ig. Both antibodies can be used to purify the three major forms of placental 5'-NT by affinity chromatography. By two-color immunofluorescence, CD73 was found to be expressed on 19 +/- 5% of CD3+, 11 +/- 4% of CD4+, 51 +/- 14% of CD8+, 25 +/- 8% of CD28+, 15 +/- 5% of CD29+, 27 +/- 7% of CD45RA+, and 70 +/- 6% of CD19+ lymphocytes. Within T cells, CD73 expression is restricted to the CD28+ subset. Thus, CD73 is found on subsets of both T and B lymphocytes, with the highest expression on B cells and CD8+ T cells. In sections of hyperplastic tonsil, CD73 expression is restricted to the small lymphocytes of the follicular mantle zone, a small subset of extrafollicular lymphocytes situated within the epithelium of the tonsillar crypt, and to follicular dendritic cells within the lower part of the "light-zone." CD73 is also detected on subsets of endothelial cells of capillaries and venules and the basal layer of non-keratinizing squamous epithelium and transitional cell type mucosa of many tissues. Given the tissue distribution of CD73, along with its glycosyl phosphatidylinositol membrane anchoring and the observation that some CD73 antibodies are mitogenic, we propose that this interesting antigen may play a role in cell activation, lymphocyte homing, and/or cell adhesion.

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The urine and plasma of epileptic patients receiving therapeutic doses of valproic acid (2-propylpentanoic acid) was found to contain five doubly unsaturated metabolites of valproic acid, which were identified äs their trimethylsilyl derivatives by GC/MS. A series of reference substances was synthesized but only two of them were identical with native metabolites: 2(2-propenyl)-4-pentenoic acid (= 4.4'-diene) and E-2propyl-2.4-pentadienoic acid (E-2.4-diene). The mass-spectra of the five native metabolites are given. Preliminary quantitative data obtained from four groups of patients indicate increased formation of doubly unsaturated metabolites when valproic acid-induced side-effects are present, and in cases of fatal hepatic failure. The 4.4'-diene has hitherto been found only in fatal cases with hepatic injury. Quantitative data are presented äs % of the sum of valproic acid plus all its detected metabolites. Fünf zweifach ungesättigte Metabolite der Valproinsäure in Urin und Plasma von epileptischen Patienten unter Behandlung mit Valproinsäure Zusammenfassung: In Urin und Plasma von epileptischen Patienten unter Behandlung mit Valproinsäure (2-Propylpentansäure) konnten insgesamt fünf zweifach ungesättigte Metabolite der Valproinsäure als Trimethylsilyl-Derivate durch GC/MS identifiziert werden. Eine Reihe von Referenzsubstanzen wurde synthetisiert, aber nur zwei Von ihnen waren mit nativen Metaboliten identisch: 2(2-PropenyI)-4-pentensäure (= 4.4'-dien) und E-2-Propyl-2.4-pentadiensäure (= E-2.4-dien). Die Massenspektren der fünf nativen Metabolite werden angegeben. Vorläufige quantitative Ergebnisse aus vier Gruppen von Patienten zeigen eine erhöhte Bildung von zweifach ungesättigten Metaboliten bei Valproinsäure-induzierten Nebenwirkungen und bei letalen Leberschäden. Das 4.4'-Dien konnte bisher nur bei Todesfällen mit Leberintoxikation nachgewiesen werden. Die quantitativen Werte werden in % der Summe aus Valproinsäure und allen nachgewiesenen Metaboliten angegeben.

Water proton nuclear relaxation measurements are used to detect and characterize four distinct intermediate states for Gd3+ bound to CaZ+ sites of sarcoplasmic reticulum Ca *+-ATPase in complexes with ATP analogues. In the absence of nucleotides, Gd3+ binds to two occluded Cazf transport sites on CaZ+-ATPase which have a low accessibility to solvent water. In the presence of the nonhydrolyzable ATP analogue, Co(NH,),AMPPCP, a new state for bound Gd3+ (still occluded and with fewer waters of hydration) is observed. In the presence of Co(NH,),ATP or ATP, two additional states for bound Gd3+ are detected in the NMR studies. The first of these probably represents an intermediate state for bound Gd3+ during ATP hydrolysis. The latter is the most occluded Gd3+ site yet observed in these studies and is probably analogous to the highly occluded E,-P state observed with CrATP [( 1987) Biochim. Biophys. Acta 898,313-3221.