Differential gene expression profiling in blood from patients with digestive system cancers

To develop a non-invasive and sensitive diagnostic test for cancer using peripheral blood, we evaluated gene expression profiling of blood obtained from patients with cancer of the digestive system and normal subjects. The expression profiles of blood-derived total RNA obtained from 39 cancer patients (11 colon cancer, 14 gastric cancer, and 14 pancreatic cancer) was clearly different from those obtained from 15 normal subjects. By comparing the gene expression profiles of cancer patients and normal subjects, 25 cancer-differentiating genes (p < 5.0 × 10⁻⁶ and fold differences >3) were identified and an “expression index” deduced from the expression values of these genes differentiated the validation cohort (11 colon cancer, 8 gastric cancer, 18 pancreatic cancer, and 15 normal subjects) into cancer patients and normal subjects with 100% (37/37) and 87% (13/15) accuracy, respectively. Although, the expression profiles were not clearly different between the cancer patients, some characteristic genes were identified according to the stage and species of the cancer. Interestingly, many immune-related genes such as antigen presenting, cell cycle accelerating, and apoptosis- and stress-inducing genes were up-regulated in cancer patients, reflecting the active turnover of immune regulatory cells in cancer patients. These results showed the potential relevance of peripheral blood gene expression profiling for the development of new diagnostic examination tools for cancer patients.     

Variation in Bitterness Potency When Introducing Gly-Gly Residue into Bitter Peptides

We previously reported that Gly-Gly-Arg-Pro and Arg-Pro-Gly-Gly, the derivatives of a bitter peptide Arg-Pro, had no bitterness although Gly-Arg-Pro and Arg-Pro-Gly had a bitter taste at the same level as that of Arg-Pro. To elucidate the mechanism of elimination of bitterness by the introduction of the Gly-Gly residues, we synthesized Gly-Gly derivatives of other bitter peptides such as Phe-Phe, Val-Val-Val, and Arg-Pro-Phe-Phe, and examined the effectiveness of Gly-Gly residues in eliminating bitterness. We suggest that, for Arg-Pro and Val-Val-Val, the Gly-Gly residue might prevent a hydrophobic group from binding to a taste receptor.     

Luciferase cDNA from Japanese firefly,Luciola cruciata: Cloning,structure and expression in Escherichia coli

We have cloned the cDNA for luciferase from lantern poly(A)⁺ RNA of a Japanese firefly, Luciola cruciata (Genji botaru in Japanese). This cDNA directed the synthesis of enzymatically active luciferase under the control of the lac promoter in Escherichia coli. The amino acid sequence predicted from the cDNA sequence shows that Genji firefly luciferase consists of 548 amino acids and has a molecular weight of 60,024. Considerable sequence homology was found upon the comparison of the Genji and North American firefly luciferases.     

The human platelet as a reproducible and sensitive cell for the detection and assay of platelet-activating factor.

A new procedure for the preparation of human platelets consistently sensitive to platelet-activating factor (PAF) in the low nanomolar range has been developed. Key to the success of this approach was the addition of adenosine during the isolation phase, providing an excellent recovery of stable cells, and the inclusion of ADP in the aggregation assay, providing increased sensitivity to PAF. Examination of the binding profile of tritium-labeled PAF to these platelets in the presence or absence of ADP revealed significant difference in the Kd values but not in the number of specific binding sites. Other reagents having an influence on the reactivity and stability of the human platelets, as regards its interaction with PAF, are described.     

Vinculin and Rab5 Complex Is Requited for Uptake of Staphyrococcus aureus and Interleukin-6 Expression

Vinculin, a 116-kDa membrane cytoskeletal protein, is an important molecule for cell adhesion; however, little is known about its other cellular functions. Here, we demonstrated that vinculin binds to Rab5 and is required for Staphylococcus aureus (S. aureus) uptake in cells. Viunculin directly bound to Rab5 and enhanced the activation of S. aureus uptake. Over-expression of active vinculin mutants enhanced S. aureus uptake, whereas over-expression of an inactive vinculin mutant decreased S. aureus uptake. Vinculin bound to Rab5 at the N-terminal region (1-258) of vinculin. Vinculin and Rab5 were involved in the S. aureus-induced phosphorylation of MAP kinases (p38, Erk, and JNK) and IL-6 expression. Finally, vinculin and Rab5 knockdown reduced infection of S. aureus, phosphorylation of MAPKs and IL-6 expression in murine lungs. Our results suggest that vinculin binds to Rab5 and that these two molecules cooperatively enhance bacterial infection and the inflammatory response.     

Oligodendrocyte Plasticity with an Intact Cell Body In Vitro

Demyelination is generally regarded as a consequence of oligodendrocytic cell death. Oligodendrocyte processes that form myelin sheaths may, however, degenerate and regenerate independently of the cell body, in which case cell death does not necessarily occur. We provide here the first evidence of retraction and regeneration of oligodendrocyte processes with no cell death in vitro, using time-lapse imaging. When processes were severed mechanically in vitro, the cells did not undergo cell death and the processes regenerated in 36 h. In a separate experiment, moderate N-methyl-D-aspartate (NMDA) stimuli caused process retraction without apparent cell death, and the processes regained their elaborate morphology after NMDA was removed from the culture medium. These results strongly suggest that demyelination and remyelination can take place without concomitant cell death, at least in vitro. Process regeneration may therefore become a target for future therapy of demyelinating disorders.