34 research outputs found
Identification of molecular candidates which regulate calcium-dependent CD8+ T-cell cytotoxicity
Cytotoxic CD8+ T lymphocytes (CTL) eliminate infected cells or transformed tumor cells by releasing perforincontaining cytotoxic granules at the immunological synapse. The secretion of such granules depends on Ca2+-
influx through store operated Ca2+ channels, formed by STIM (stromal interaction molecule)-activated Orai
proteins. Whereas molecular mechanisms of the secretion machinery are well understood, much less is known
about the molecular machinery that regulates the efficiency of Ca2+-dependent target cell killing. CTL killing
efficiency is of high interest considering the number of studies on CD8+ T lymphocytes modified for clinical use.
Here, we isolated total RNA from primary human cells: natural killer (NK) cells, non-stimulated CD8+ T-cells,
and from Staphylococcus aureus enterotoxin A (SEA) stimulated CD8+ T-cells (SEA-CTL) and conducted whole
genome expression profiling by microarray experiments. Based on differential expression analysis of the transcriptome data and analysis of master regulator genes, we identified 31 candidates which potentially regulate
Ca2+-homeostasis in CTL. To investigate a putative function of these candidates in CTL cytotoxicity, we transfected either SEA-stimulated CTL (SEA-CTL) or antigen specific CD8+ T-cell clones (CTL-MART-1) with siRNAs
specific against the identified candidates and analyzed the killing capacity using a real-time killing assay. In
addition, we complemented the analysis by studying the effect of inhibitory substances acting on the candidate
proteins if available. Finally, to unmask their involvement in Ca2+ dependent cytotoxicity, candidates were also
analyzed under Ca2+-limiting conditions. Overall, we identified four hits, CCR5 (C-C chemokine receptor type
five), KCNN4 (potassium calcium-activated channel subfamily N), RCAN3 (regulator of calcineurin) and BCL (Bcell lymphoma) 2 which clearly affect the efficiency of Ca2+ dependent cytotoxicity in CTL-MART-1 cells, CCR5,
BCL2, and KCNN4 in a positive manner, and RCAN3 in a negative way
ChemInform Abstract: Discovery of Olodaterol (V), a Novel Inhaled β2-Adrenoceptor Agonist with a 24 h Bronchodilatory Efficacy.
Synthesis of 2-Hydroxy-6-{[(16R)-Ăź-D-mannopyranosyloxy]heptadecyl}benzoic Acid, a Fungal Metabolite with GABA<sub>A</sub> Ion Channel Receptor Inhibiting Properties
An expeditious total synthesis of the physiologically active fungal metabolite 1 is described. The stereoselective formation of its Ăź-D-mannopyranosidic linkage is achieved in two steps upon reaction of the hexopyranos-2-ulosyl bromide 15 with the glycosyl acceptor 13, followed by reduction of the resulting Ăź-D-glycos-2-uloside 16. Alcohol 13 was efficiently prepared via a Suzuki reaction of the aryltriflate 11 with the 9-alkyl-9-BBN derivative 10
A practical synthesis of β-d-mannopyranosides
An indirect yet highly efficient protocol for the β-d-mannosylation of sterically hindered alcohols is reported. Trichloroacetimidate 5 is used a key building block which is converted into the desired mannosides 9 via triflates 8 by an ultrasound promoted β-d-gluco → β-d-manno inversion process
Total Synthesis of Caloporoside
The first total synthesis of the fungal metabolite caloporoside 1, a strong and selective inhibitor of phospholipase C, is described. Both sugar units of its complex disaccharidic segment were obtained from 3,4,6-tri-O-benzyl-d-glucopyranose 14 as a common building block, with d-gluco → d-manno inversions as the key strategic elements. This particular substitution reaction occurred readily on the acyclic segment (27 → 28), whereas ultrasonication was required to override adverse stereoelectronic effects upon formation of β-d-mannopyranoside unit 34. The (16R)-hydroxyheptadecylsalicylic acid part of 1 was efficiently prepared by a palladium-catalyzed Suzuki cross coupling reaction of aryltriflate 7 with the 9-alkyl-9-BBN derivative formed from alkene 6 and 9-H-9-BBN