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R Seki

Publications and source records attributed to R Seki.

At least 37 records · Page 2Linked to original sources
PubMed1994

V delta 5+ T cells of BALB/c mice recognize the murine heat shock protein 60 target cell specificity.

The purpose of this paper was to study the gamma delta T-cell receptor repertoire and target specificity following stimulation of peripheral T cells of BALB/c mice with autologous or bacterial ligands. The expression of V gamma and V delta chain families in T cells that had been expanded by stimulation in syngeneic mixed lymphocyte culture or with purified protein derivative (PPD) was determined by the semiquantitative DNA polymerase chain reaction (PCR) method. Responder T cells to either of these stimuli strongly expressed both V delta 5 and V delta 6 genes. However, addition of the ML 30 anti-murine heat shock protein (hsp) 60 monoclonal antibody (mAb) to the cell culture selectively inhibited only the expansion of V delta 5 T cells. A V delta 5 T-cell hybridoma (KMT-5), which recognized syngeneic splenic and fibrosarcoma Meth A cells but not allogeneic cells, was produced by cell fusion from autoreactive blast cells. Incubation of the KMT-5 hybridoma in the presence of ML 30 antibody blocked the stimulation of interleukin-2 (IL-2) secretion by syngeneic target cells. It was also found that the DNA of KMT-5 hybridoma and of the autoreactive gamma delta T cells contained the BALB/c invariant delta (BID) chain sequence. It is concluded from these results that BALB/c peripheral V delta 5 T cells recognize an autologous hsp 60 target specificity in a V delta-gene restricted manner. We also propose that T cells of this V gene family may be involved in the immune surveillance of certain tumours and intracellular infections.

Animals↗
PubMed1993

Mechanism of chromosomal integration of transgenes in microinjected mouse eggs: sequence analysis of genome-transgene and transgene-transgene junctions at two loci.

Production of transgenic animals is a key technique in modern biology, but the process of chromosomal integration of transgenes in microinjected eggs is still not fully understood. To gain information on the mechanisms involved in this process, we have cloned two transgene loci and their corresponding pre-integration sites and compared the junction sequences with the parental nucleotide (nt) sequences. No extensive DNA rearrangements were detected at these loci: only simple deletions (caused by the integration of the transgene concatemers) were present in the host genome. Analysis of three transgene-transgene junctions within the concatemers showed that 'nibbling' of ends (up to 3 nt) had occurred at some ends prior to joining. At all four genome-transgene junctions, short homologies of 1 to 3 nt were found, and at least three of these junctions were associated with the consensus sequence for topoisomerase-I cleavage sites. Moreover, three of the four integration junctions occurred in the terminal regions of the injected sequence, at positions only a few nt away from the ends. These results suggest that linear, but not circular, concatemers were preferentially integrated at their ends utilizing short homologies to the host genome.

Animals↗
PubMed1993

Extensive N nucleotide addition in junctional region of T cell receptor V gamma 5 genes rearranged in fetal liver-derived thymocytes in radiation chimera mice.

The V gamma 5 chain of T cell receptor gamma delta is preferentially expressed by murine fetal thymocytes. The fetal V gamma 5 gene is known to be homogeneous and lacks N nucleotide addition. We previously reported that V gamma 5+ cells were detected among donor-derived thymocytes from irradiated C3H/He mice soon after reconstitution with AKR/J fetal liver (FL) cells, but that only a few V gamma 5+ were detected among donor-derived thymocytes from irradiated C3H/He mice reconstituted with adult bone marrow (ABM) cells. The results suggest that preferential use of V gamma 5 is determined at the level of T cell precursor generation. In the present report, we analyzed whether the junctional region of FL-derived V gamma 5 genes in the FL chimeras is of fetal type. Sequencing analysis showed that V gamma 5 genes from FL chimeras contained extensive N nucleotide addition and were diverse both in nucleotide sequences and deduced amino acid sequences. V gamma 5 genes from donor-derived thymocytes of ABM chimeras, which by polymerase chain reaction were revealed to be less frequent than those of FL chimeras, also showed extensive N addition. Furthermore, the terminal deoxynucleotidyl transferase (TdT) gene, which encodes an enzyme that adds N nucleotides into the junctional region, was expressed at high level in the donor-derived thymocytes of FL chimeras and normal 8-week-old AKR/J thymocytes but at low level in day 17 fetal AKR/J thymocytes. Our results suggest that the preferential rearrangement of the V gamma 5 gene is determined at the level of T cell precursor generation but the homogeneous V gamma 5 sequence of fetal type is generated only in the fetal thymic microenvironment where TdT gene expression is low or absent. The nomenclature of murine TcR gamma chains is according to Reilly et al. (Nature 1986. 321: 878). The relationship between the different nomenclature systems is summarized in Takagaki et al. (J. Immunol. 1989. 141: 2112).

Animals↗
PubMed1991

Inherited type of allelic methylation variations in a mouse chromosome region where an integrated transgene shows methylation imprinting.

It is still unclear whether or not parent-of-origin-dependent differential methylation observed in some transgenes reflects genomic imprinting of endogenous genes. We have characterized a transgene locus showing such methylation imprinting together with the corresponding native chromosome region. We show that only part of the transgene is affected by methylation imprinting and the methylation pattern is established before early prophase I during spermatogenesis. Interestingly, the native genomic region, which is mapped to the proximal chromosome 11, shows no evidence of methylation imprinting but displays heritable, strain-specific type of allelic methylation differences. The results demonstrate that transgenes do not necessarily reflect the methylation status of either the surrounding or corresponding chromosome region. In addition, inherited type of allelic methylation variations previously described in human may be widespread in mammals.

Animals↗
PubMed1991

Exonic point mutations in NADH-cytochrome B5 reductase genes of homozygotes for hereditary methemoglobinemia, types I and III: putative mechanisms of tissue-dependent enzyme deficiency.

We analyzed the NADH-cytochrome b5 reductase gene of hereditary methemoglobinemia type I and type III, by using PCR-related techniques. The mutation in type I is a guanine-to-adenine substitution in codon 57 of exon 3 of the NADH-cytochrome b5 reductase gene, and the sense of this codon is changed from arginine to glutamine. In type III the mutation is a thymine-to-cytosine transition in codon 148 of exon 5, causing leucine-to-proline replacement in type III. The former mutation abolishes the MspI recognition site. Homozygosity for the former mutation in a patient with type I was confirmed by restriction analysis of PCR-amplified fragments and by dot blot hybridization of amplified products with allele-specific oligonucleotide probes. The latter mutation generates a recognition site for MspI. Amplification of exon 5 by PCR followed by digestion with MspI revealed homozygosity for this mutation in patients with type-III. Putative mechanisms of tissue-dependent enzyme defects in hereditary methemoglobinemia are discussed.

Adult↗
PubMed1988

Scanning electron microscopic observations on vascular casts of ciliary processes in normal and topical epinephrine-treated rats.

Existence of neurohumoral control of the ciliary process blood flow was investigated in normal adult Wistar albino rats, utilizing the vascular casting method with a scanning electron microscope, comparing the ciliary process vessels in epinephrine-treated eyes to those in control eyes. The ciliary process microvasculature consisted of marginal capillaries, intraprocess capillaries and collecting veins. The supplying arterioles of the ciliary processes were derived from the iridociliary arterial circle. The arterioles were constricted or annularly impressed just before connecting with the marginal capillaries in the controls. The epinephrine instillation resulted in significant constriction of all the ciliary process vessels. Especially the ciliary process arterioles were narrowed in their full length in contrast to the focal constrictions in the controls. The diameter of the constricted arterioles in the epinephrine-treated eyes was the same as the focal constrictions in the controls. These results suggested that sphincter muscles existed in the wall of the ciliary process arterioles of the rat and that neurohumoral blood flow control was related to the constriction of the ciliary process arterioles resulting from the epinephrine administration.

Administration, Topical↗