Molecular cytogenetic analysis of the highly repetitive DNA in the genome of Apodemus argenteus, with comments on the phylogenetic relationships in the genus Apodemus.

The DNA of Apodemus argenteus was digested with DraI, and the resultant DraI fragment of highly repetitive DNA was isolated and analyzed by DNA filter hybridization, cloning, sequencing, and fluorescence in situ hybridization (FISH). Southern blot hybridization and nucleotide sequencing revealed that most of the DraI fragment consisted of a 230-bp repeating unit and contained no sex-chromosome-specific nucleotide sequences. The DraI fragment included the CENP-B box-like sequence, with a strong homology to the human CENP-B box sequence.

Disease model: human aging.

Very little is known about the molecular mechanisms of human aging. This, at least in part, derives from a paucity of appropriate animal models of aging. Until recently, the senescence-accelerated mouse was the only mammalian model of aging.

Aging mechanisms.

Aging (senescence) has long been a difficult issue to be experimentally analyzed because of stochastic processes, which contrast with the programmed events during early development. However, we have recently started to learn the molecular mechanisms that control aging. Studies of the mutant mouse, klotho, showing premature aging, raise a possibility that mammals have an "anti-aging hormone.

In vivo klotho gene delivery protects against endothelial dysfunction in multiple risk factor syndrome.

The klotho gene, originally identified by insertional mutagenesis in mice, suppresses multiple aging phenotypes (e.g., arteriosclerosis, pulmonary emphysema, osteoporosis, infertility, and short life span).

Decreased insulin production and increased insulin sensitivity in the klotho mutant mouse, a novel animal model for human aging.

We have recently identified a novel gene, klotho (kl), which may suppress several aging phenotypes. A defect of kl gene expression in the mouse results in a syndrome resembling human aging, such as arteriosclerosis, skin atrophy, osteoporosis, and pulmonary emphysema. To determine whether mouse homozygotes for the kl mutation (kl/kl) show abnormal glucose metabolism, an oral glucose tolerance test (OGTT) was performed at 6 to 8 weeks of age.

Targeted disruption of Na+/Ca2+ exchanger gene leads to cardiomyocyte apoptosis and defects in heartbeat.

Ca(2+), which enters cardiac myocytes through voltage-dependent Ca(2+) channels during excitation, is extruded from myocytes primarily by the Na(+)/Ca(2+) exchanger (NCX1) during relaxation. The increase in intracellular Ca(2+) concentration in myocytes by digitalis treatment and after ischemia/reperfusion is also thought to result from the reverse mode of the Na(+)/Ca(2+) exchange mechanism. However, the precise roles of the NCX1 are still unclear because of the lack of its specific inhibitors.

Improvement of multiple pathophysiological phenotypes of klotho (kl/kl) mice by adenovirus-mediated expression of the klotho gene.

Background: We have established a novel mouse mutant, klotho (kl), by insertional mutation of a transgene and identified the structural gene. The mouse homozygous for the mutation exhibits multiple pathological conditions resembling age-related disorders in humans and can be regarded as a model of human premature aging syndromes. However, the pathophysiological role of Klotho protein has not been clarified.

Isolation and characterization of Na+/Ca2+ exchanger gene and splicing isoforms in mice.

The Na+/Ca2+ exchanger gene NCX1 is ubiquitously expressed in mammalian tissues, and encodes several isoforms through alternative RNA splicing. In this report, we describe the gene structure that gives rise to the multiple isoforms, and the tissue-specific expression of these isoforms in mice. The mouse NCX1 gene contains a cluster of six exons (A, B, C, D, E, and F) which encode a variable region in the large intracellular loop of the protein, as previously reported in rabbits and humans.

Impairment of B lymphopoiesis in precocious aging (klotho) mice.

Inactivation of the klotho gene in mice results in multiple disorders that resemble human aging after 3 weeks of age. Because hematopoiesis, especially B lymphopoiesis, is affected in humans and mice by aging, we analyzed the hematopoietic state in homozygous klotho (kl/kl) mice. The kl/kl mice showed thymic atrophy and a reduced number of splenocytes.

Cytogenetic analysis of the tamaraw (Bubalus mindorensis): a comparison of R-banded karyotype and chromosomal distribution of centromeric satellite DNAs, telomeric sequence, and 18S-28S rRNA genes with domestic water buffaloes.

The karyotype of the tamaraw (Bubalus mindorensis, 2n = 46) was investigated by RBG-banding technique and compared with those of the river and the swamp cytotypes of domestic water buffalo (B. bubalis). The tamaraw karyotype consisted of 6 submetacentric and 16 acrocentric autosome pairs (NAA = 56), and X and Y chromosomes.

Increases in intracellular calcium of arterial smooth muscle cells in transgenic mice overexpressing Na+/H+ exchanger.

We produced transgenic mice overexpressing Na+/ H+ exchanger as a model of salt-sensitive hypertension and reported that dietary salt loading elevates blood pressure in these transgenic mice. We speculate that this blood pressure elevation may be attributed to the elevation of intraarterial smooth muscle Ca2+ concentration through Na+/Ca2+ exchange. To test this hypothesis, we measured the isometric tension of aortic rings and intracellular free calcium ([Ca2+]i) of cultured smooth muscle cells.

Establishment of the anti-Klotho monoclonal antibodies and detection of Klotho protein in kidneys.

A novel gene, klotho (kl), which is involved in the development of a syndrome resembling human aging in mice, was recently identified. The kl gene encodes a single-pass membrane protein whose extracellular domain carries homology to beta-glucosidases. There also exists a splice variant of kl mRNA which encodes a putative secreted protein in both human and mouse.

Disruption of the klotho gene causes pulmonary emphysema in mice. Defect in maintenance of pulmonary integrity during postnatal life.

Homozygous mutant klotho (KL(-/-)) mice exhibit multiple phenotypes resembling human aging. In the present study, we focused on examining the pathology of the lungs of klotho mice and found that it closely resembled pulmonary emphysema in humans both histologically and functionally. Histology of the lung of KL(-/-) mice was indistinguishable from those of wild-type littermates up to 2 wk of age.

Dietary salt loading increases nitric oxide synthesis in transgenic mice overexpressing sodium-proton exchanger.

We studied the role of nitric oxide (NO) synthesis in amelioration of blood pressure elevation during dietary salt loading in transgenic mice overexpressing sodium proton exchanger. Systolic blood pressure rose after starting salt loading only in the high-salt group of transgenic mice. However, this elevation of blood pressure was not continued.

Independent impairment of osteoblast and osteoclast differentiation in klotho mouse exhibiting low-turnover osteopenia.

We recently identified a new gene, klotho, which is involved in the suppression of multiple aging phenotypes. The mouse homozygous for a disruption of the klotho locus (kl/kl) exhibited multiple pathological conditions resembling human aging. Histomorphometric analysis revealed low-turnover osteopenia in kl/kl mice.

BTEB2, a Krüppel-like transcription factor, regulates expression of the SMemb/Nonmuscle myosin heavy chain B (SMemb/NMHC-B) gene.

We have recently characterized the promoter region of the rabbit embryonic smooth muscle myosin heavy chain (SMemb/NMHC-B) gene and identified the 15-bp sequence, designated SE1, located at -105 from the transcriptional start site as an important regulatory element for its transcriptional activity in a smooth muscle cell (SMC) line. In this study, we attempted to isolate cDNA clones encoding for the transcription factors that control the expression of the SMemb gene through binding to this cis-regulatory element. We screened a lambdagt11 cDNA library prepared from C2/2 cells, a rabbit-derived SMC line, by using a radiolabeled concatenated oligonucleotide containing SE1 as a probe.

Molecular cloning of rat klotho cDNA: markedly decreased expression of klotho by acute inflammatory stress.

We have recently identified a novel gene, termed klotho, that is involved in the suppression of several aging phenotypes. The gene encodes a membrane protein that shares sequence similarity with the beta-glucosidases of bacteria and plants. In this study, we isolated rat klotho cDNA and examined its tissue distribution in rats.

Downregulation of the Klotho gene in the kidney under sustained circulatory stress in rats.

We recently reported the isolation of the klotho gene, which in predominantly expressed in the kidney and involved in human aging phenotypes. In our previous studies, we demonstrated that the Klotho protein or its metabolites may possibly function as humoral factor(s) and protect against endothelial dysfunction because acetylcholine-mediated NO production in arteries was impaired in heterozygous klotho deficient mice (kl/+). However, the pathophysiological significance of the Klotho protein has not been clarified yet.

Klotho protein protects against endothelial dysfunction.

Arteriosclerosis caused by aging is recognized to be a crucial risk factor of cardiovascular disease. We recently established klotho mouse which causes age-related disorders including arteriosclerosis. However, no information on endothelial function of klotho mouse or the physiological role of klotho protein as a circulating factor is available.

UTF1, a novel transcriptional coactivator expressed in pluripotent embryonic stem cells and extra-embryonic cells.

We have obtained a novel transcriptional cofactor, termed undifferentiated embryonic cell transcription factor 1 (UTF1), from F9 embryonic carcinoma (EC) cells. This protein is expressed in EC and embryonic stem cells, as well as in germ line tissues, but could not be detected in any of the other adult mouse tissues tested. Furthermore, when EC cells are induced to differentiate, UTF1 expression is rapidly extinguished.

Cytogenic studies of Hynobiidae (Urodela). XIV. Analysis of the chromosome of a Chinese salamander, Batrachuperus pinchonii (David).

The chromosome number of a Chinese salamander, Batrachuperus pinchonii, was re-examined Adults and embryonic specimens had a diploid number of 66, with 33 bivalents during meiosis, in contrast to previous reported results. Furthermore, when C-banding analysis was performed with embryos, chromosomes with banding patterns homoeologous to those of Salamandrella keyserlingii and Hynobius species were found. It appears, therefore, that Batrachuperus, Salamandrella and Hynobius might be derived from a common ancestral species in eastern Asia.

Structure of the mouse klotho gene and its two transcripts encoding membrane and secreted protein.

We previously established a novel mouse model for human aging and identified the genetic foundation responsible for it. A defect in expression of a novel gene, termed klotho (kl), leads to a syndrome resembling human aging in mice. The kl gene encodes a single-pass membrane protein whose extracellular domain carries homology to beta-glucosidases.

Identification of the human klotho gene and its two transcripts encoding membrane and secreted klotho protein.

Inactivation of the klotho (kl) gene in mice results in multiple disorders that resemble human aging. The mouse kl gene encodes a novel single-pass membrane protein with homology to beta-glucosidases. In this study, we have isolated a human homologue of the kl gene and determined its gene structure.

Mutation of the mouse klotho gene leads to a syndrome resembling ageing.

A new gene, termed klotho, has been identified that is involved in the suppression of several ageing phenotypes. A defect in klotho gene expression in the mouse results in a syndrome that resembles human ageing, including a short lifespan, infertility, arteriosclerosis, skin atrophy, osteoporosis and emphysema. The gene encodes a membrane protein that shares sequence similarity with the beta-glucosidase enzymes.

Isolation of the embryonic form of smooth muscle myosin heavy chain (SMemb/NMHC-B) gene and characterization of its 5'-flanking region.

To examine the molecular mechanisms that regulate the expression of the SMemb/NMHC-B gene, a nonmuscle myosin heavy chain isoform predominantly expressed in fetal aorta, we have isolated and characterized the 5'-flanking region of the rabbit SMemb/NMHC-B gene. Transient transfection experiments demonstrated that 105 base pairs of 5'-flanking sequence was necessary to direct high level transcription in C2/2 cells, vascular smooth muscle cells derived from rabbit aorta. An essential cis-regulatory element was localized between -100 and -91 base pairs from the transcription start site based on the results that replacement mutagenesis within this region significantly reduced promoter activity.