T Cell Factor-1 Controls the Lifetime of CD4+ CD8+ Thymocytes In Vivo and Distal T Cell Receptor α-Chain Rearrangement Required for NKT Cell Development

Natural killer T (NKT) cells are a component of innate and adaptive immune systems implicated in immune, autoimmune responses and in the control of obesity and cancer. NKT cells develop from common CD4+ CD8+ double positive (DP) thymocyte precursors after the rearrangement and expression of T cell receptor (TCR) Vα14-Jα18 gene. Temporal regulation and late appearance of Vα14-Jα18 rearrangement in immature DP thymocytes has been demonstrated. However, the precise control of lifetime of DP thymocytes in vivo that enables distal rearrangements remains incompletely defined. Here we demonstrate that T cell factor (TCF)-1, encoded by the Tcf7 gene, is critical for the extended lifetime of DP thymocytes. TCF-1-deficient DP thymocytes fail to undergo TCR Vα14-Jα18 rearrangement and produce significantly fewer NKT cells. Ectopic expression of Bcl-x_L permits Vα14-Jα18 rearrangement and rescues NKT cell development. We report that TCF-1 regulates expression of RORγt, which regulates DP thymocyte survival by controlling expression of Bcl-x_L. We posit that TCF-1 along with its cofactors controls the lifetime of DP thymocytes in vivo.     

Liver fatty acid-binding protein in two cases of human lipid storage

Summary FABPs in the various tissues play an important role in the intracellular fatty acid transport and metabolism. Reye's syndrome (RS) and multisystemic lipid storage (MLS) are human disorders characterized by a disturbance of lipid metabolism of unknown etiology. We investigated for the first time L-FABP in these two conditions. Affinity purified antibodies against chicken L-FABP were raised in rabbits, and found to cross-react specifically with partially purified human L-FABP. L-FABP content in liver samples of two patients with RS and MLS was investigated by immuno-histochemistry, SDS-PAGE and ELISA. L-FABP immuno-histochemistry showed increased reactivity in the liver of RS patient and normal reactivity in MLS liver. L-FABP increase in RS liver was confirmed by densitometry of SDS-PAGE and ELISA method. By these two methods the increase amounted to 180% and 199% (p < 0.02), respectively, as compared to controls. A possible role of L-FABP in the pathogenesis of RS is discussed.     

Affinity chromatography of Ankistrodesmus braunii nitrate reductase using blue dextran-sepharose (author's transl)

Blue Dextran has been coupled covalently to Sepharose-4B to purify the enzymatic complex NAD(P)H-nitrate reductase (EC 1.6.6.2) from the green alga Ankistrodesmus braunii by affinity chromatography. The optimum conditions for the accomplishment of the chromatographic process have been determined. The adsorption of nitrate reductase on Blue Dextran Sepharose is optimum when a phosphate buffer of low ionic strength and pH 6.5-7.0 is used. Once the enzyme has been bound to Blue Dextran Sepharose, it can be specifically eluted by addition of NADH and FAD to the washing buffer. However, none of the nucleotides added separately is able to promote the elution of the enzyme from the column. The elution can be also achieved, but not specifically, by increasing the ionic strength of the buffer with KCl. These results have made possible a procedure for the purification of A. braunii nitrate reductase which led to electrophoretic homogeneity, with an overall yield of 70% and a specific activity of 49 units/mg of protein.     

Ultrastructure of type-I salivary-gland acini in four species of ticks and the influence of hydration states on the type-I acini ofAmblyomma americanum

We describe the ultrastructure of type-I salivary-gland acini in two argasid and two ixodid species. The basic cell types in the agranular or type-I acini, and their associations, are very similar in argasids and ixodids; therefore, we propose an anatomical nomenclature for cells in the type-I acinus based on the adult ixodidsAmblyomma americanum andDermacentor variabilis, and the argasid adultArgas (Persicargas) arboreus and on nymphalOrnithodoros moubata. Four cell types were present in all specimens: one central lamellate cell, a variable number of peripheral lamellate cells, a variable number of peritubular cells depending on the species, and one circumlumenal cell. The lamellate cells had infolded basal plasma membranes that presented an amplified surface area to the hemolymph. These cells most likely secreted the fluid involved in water vapor uptake by ticks. ForAmblyomma americanum females, abundant K⁺-dependent, ouabain-sensitive Na⁺, K⁺-ATPase complexes were located on the infolded basal plasma membranes of the lamellate cells. Apical membranes of the lamellate cells, and plasma membranes of other cell types in the acinus had little or no evidence of Na⁺, K⁺-ATPase activity. Only the central lamellate cell extended from the hemolymph of the acinus to its lumen; peripheral cells did not contact the lumen. Except when the ticks were rehydrating, lipid inclusions were common features in the lamellate cells of the ixodids. Lipid inclusions were not seen in argasid type I acini; however, glycogen deposits were common. To determine if acinar cells respond to the changing hydration state of the tick, unfed femaleA. americanum were subjected to dehydration/rehydrating conditions. During rehydration, mitochondria in the lamellate cells changed from a matrix of medium electron-density and intermembrane space (orthodox configuration) to a matrix of greater density and larger intermembrane space (condensed configuration). The orthodox configuration was consistently observed in control and dehydrating ticks. The condensed configuration was the norm for mitochondria in lamellate cells of rehydrating ticks. Lipid inclusions were depleted in the rehydrating ticks compared to control or dehydrating ticks. Acini appeared to be reverting to the control or desiccated state when ticks were returned to low humidity, suggesting that these changes were cyclical. Nymphs ofO. moubata subjected to the same dehydration/rehydrating conditions showed no obvious ultrastructural changes.     

The role of humans in the diversification of a threatened island raptor

Background  

Anthropogenic habitat modifications have led to the extinction of many species and have favoured the expansion of others. Nonetheless, the possible role of humans as a diversifying force in vertebrate evolution has rarely been considered, especially for species with long generation times. We examine the influence that humans have had on the colonization and phenotypic and genetic differentiation of an insular population of a long-lived raptor species, the Egyptian vulture (Neophron percnopterus).     

Production of multimeric forms of CD4 through a sugar-based cross-linking strategy

We have developed a three-step cross-linking procedure that is specifically targeted at the carbohydrate on a protein and applied it to CD4 as a model system for studying the role of multivalent interactions in function. In the first step CD4 was oxidized with periodate, creating aldehydes that served as targets for the subsequent chemistry. Next the aldehydes were modified with cystamine, converting the reactive group into a thiol. Finally cross-linking through the thiol moiety was generated with the homobifunctional cross-linker bismaleimidohexane. With this procedure, approximately 60% of the CD4 was converted into higher molecular weight complexes that were soluble and retained function as assessed by glycoprotein gp120 binding activity. CD4 dimers and tetramers by mass were 4 and 15 times as active as CD4 monomer in blocking virus infection with HTLV-IIIB in an in vitro cellular assay. The cross-linking chemistry provides an efficient method for producing homomultimers of a glycoprotein.