Enzyme activity profiles in mouse teratocarcinomas. A quantitative ultramicroscale analysis.

Nine tumour lines with different developmental capacities were derived from spontaneous as well as from one induced teratocarcinoma: three teratocarcinoma-derived rhabdomyosarcomas TDR 602, TDR 694, and TDR 114; two teratocarcinoma-derived neuroblastomas TDN 2151 and TDN 2283; two teratocarcinoma-derived endodermal tumours TDE 274 and TDE 113; one multipotential teratocarcinoma OTT 2289, and one undifferentiated teratocarcinoma OTT 2158. Quantitative analyses of ten catabolic enzymes, i.e. alkaline and acid phosphatase, alpha- and beta-galactosidase, alpha- and beta-glucosidase, alpha-mannosidase, alpha-fucosidase, beta-glucuronidase, and hexosaminidase were carried out at the 20-cell level, and specific enzyme activity profiles were established for each of the tumour lines studied. These profiles may be used for the biochemical identification of a tumour type at the single cell level in addition to morphological and biological criteria

Mice Deficient for the Myelin-Associated Glycoprotein Show Subtle Abnormalities in Myelin

Using homologous recombination in embryonic stem cells, we have generated mice with a null mutation in the gene encoding the myelin-associated glycoprotein (MAG), a recognition molecule implicated in myelin formation. MAG-deficient mice appeared normal in motor coordination and spatial learning tasks. Normal myelin structure and nerve conduction in the PNS, with N-CAM overexpression at sites normally expressing MAG, suggested compensatory mechanisms. In the CNS, the onset of myelination was delayed, and subtle morphological abnormalities were detected in that the content of oligodendrocyte cytoplasm at the inner aspect of most myelin sheaths was reduced and that some axons were surrounded by two or more myelin sheaths. These observations suggest that MAG participates in the formation of the periaxonal cytoplasmic collar of oligodendrocytes and in the recognition between oligodendrocyte processes and axons

Trace fear conditioning involves hippocampal α(5) GABA(A) receptors

The heterogeneity of γ-aminobutyric acid type A (GABA(A)) receptors contributes to the diversity of neuronal inhibition in the regulation of information processing. Although most GABA(A) receptors are located synaptically, the small population of α(5)GABA(A) receptors is largely expressed extrasynaptically. To clarify the role of the α(5)GABA(A) receptors in the control of behavior, a histidine-to-arginine point mutation was introduced in position 105 of the murine α(5) subunit gene, which rendered the α(5)GABA(A) receptors diazepam-insensitive. Apart from an incomplete muscle relaxing effect, neither the sedative, anticonvulsant, nor anxiolytic-like activity of diazepam was impaired in α(5)(H105R) mice. However, in hippocampal pyramidal cells, the point mutation resulted in a selective reduction of α(5)GABA(A) receptors, which altered the drug-independent behavior. In line with the role of the hippocampus in certain forms of associative learning, trace fear conditioning, but not delay conditioning or contextual conditioning, was facilitated in the mutant mice. Trace fear conditioning differs from delay conditioning in that the conditioned and unconditioned stimulus are separated by a time interval. Thus, the largely extrasynaptic α(5)GABA(A) receptors in hippocampal pyramidal cells are implicated as control elements of the temporal association of threat cues in trace fear conditioning

In vitro generation of mature, naive antigen-specific CD8⁺ T cells with a single T-cell receptor by agonist selection

Peripheral blood T cells transduced with a tumor-specific T-cell receptor (TCR) face problems of auto-reactivity and lack of efficacy caused by cross-pairing of exogenous and endogenous TCR chains, as well as short term in vivo survival due to activation and growth factor-induced differentiation. We here studied an alternative strategy for the efficient generation of naive CD8(+) T cells with a single TCR. TCR-transduced human postnatal thymus-derived and adult mobilized blood-derived hematopoietic progenitor cells (HPCs) were differentiated to CD4(+)CD8(+) double-positive T cells using OP9-Delta-like 1 (OP9-DL1) cultures. Addition of the agonist peptide induced double positive cells to cross-present the peptide, leading, in the absence of co-stimulation, to cell cycle arrest and differentiation into mature CD8(+) T cells. comprehensive phenotypic, molecular and functional analysis revealed the generation of naive and resting CD8(+) T cells through a process similar to thymic positive selection. These mature T cells show a near complete inhibition of endogenous TCRA and TCRB rearrangements and express high levels of the introduced multimerreactive TCR. Upon activation, specific cytokine production and efficient killing of tumor cells were induced. Using this strategy, large numbers of high-avidity tumor-specific naive T cells can be generated from readily available HPCs without TCR chain cross-pairing