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Biomedical subjects

S Tajima

Publications and source records attributed to S Tajima.

At least 19 recordsLinked to original sources
PubMed1996

Erythropoietin-independent erythrocyte production: signals through gp130 and c-kit dramatically promote erythropoiesis from human CD34+ cells.

Erythropoietin (EPO) is the primary humoral regulator of erythropoiesis and no other factor has previously been reported to support proliferation and terminal maturation of erythroid cells from hemopoietic stem cells. Here we show that stimulation of glycoprotein (gp130) by a combination of recombinant human soluble interleukin 6 receptor (sIL-6R) and IL-6 but not sIL-6R or IL-6 alone can support proliferation, differentiation, and terminal maturation of erythroid cells in the absence of EPO from purified human CD34+ cells in suspension culture containing stem cell factor (SCF). A number of erythroid bursts and mixed erythroid colonies also developed in methylcellulose culture under the same combination. The addition of anti-gp130 monoclonal antibodies but not anti-EPO antibody to the same culture completely abrogated the generation of erythroid cells. These results clearly demonstrate that mature erythroid cells can be emerged from hemopoietic progenitors without EPO in vitro. Together with the previous reports that human sera contain detectable levels of sIL-6R, IL-6, and SCF, current data suggest that gp130 signaling in association with c-kit activation may play a role in human erythropoiesis in vivo.

Antigens, CD↗
PubMed1996

Immunohistochemical studies of glycosaminoglycans in nanophthalmic sclera.

BACKGROUND: It has been observed that there may be some connection between scleral collagen changes, glycosaminoglycan (GAG) changes and nanophthalmic uveal effusion. In a previous paper we reported increased dermatan sulfate (DS) and chondroitin sulfate (CS) contents in nanophthalmos sclera using biochemical analysis. In the present work we used immunohistochemical methods to assess the GAG changes in nanophthalmos sclera. METHODS: Scleral specimens from three nanophthalmic patients and five controls were processed to immunostaining with peroxidase-antiperoxidase (PAP) methods. Monoclonal antibodies for chondroitin (Ch), chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S), DS and keratan sulfate (KS) were used. The specimens were observed under a light microscope. RESULTS: Nanophthalmos scleral samples exhibited strong KS staining compared with the control specimens. Both the nanophthalmos and the control samples were stained with anti-Ch, anti-C4S, anti-C6S and anti-DS antibodies to the same intensity. CONCLUSION: Our results suggest that an increased or abnormal distribution of KS may contribute to abnormalities in collagen fibers.

Adolescent↗
PubMed1996

Immunolocalizations of human gelatinase (type IV collagenase, MMP-9) and TIMP (tissue inhibitor of metalloproteinases) in normal epidermis and some epidermal tumors.

The matrix metalloproteinases (MMPs) MMP-2 and MMP-9 (gelatinases) have been suggested as serving an important role in cleaving the basement membrane structure. Tissue inhibitors of metalloproteinases TIMPs (particularly TIMP-1) are known to inhibit MMPs. Based on this background, we raised monoclonal antibodies against human gelatinase (MMP-9) and human recombinant TIMP (TIMP-1), and immunostained these two components in skin from patients with squamous cell carcinoma (SCC), Bowen's disease (BD) and keratoacanthoma (KA). MMP-9 showed positive staining mainly in the granular layer of normal epidermis. In some cases of SCC and BD, MMP-9 showed positive staining in the dysplastic lesions even in the basal layer. TIMP showed a thorough positivity in normal epidermis. Unstained regions with this antibody were observed in SCC and BD. These results suggest that an altered staining pattern for MMP-9 and TIMP may be closely related to the malignant transformation of SCC and BD.

Animals↗
PubMed1996

Melanogenesis, biosynthetic phenotype of fibronectin and collagen, and migrating activity in cloned B16 mouse melanoma cells.

Three clones, melanotic (M3), amelanotic (A4 and A7) cells were isolated from B16 F10 mouse melanoma cell line. These clones exhibited different biosynthetic activities of fibronectin and collagen: A4 clone showed relatively active synthesis of both collagen and fibronectin, and A7 clone exhibited most active fibronectin synthesis, whereas no significant synthesis of these molecules was observed in M3 clone. No significant difference in growth rate was observed in these three clones. Migrating activities measured by basement membrane matrix-coated dishes were greater in A7 clone than in A4 and M3 clones. Messenger RNA levels of collagen and fibronectin paralleled collagen and fibronectin synthesis in these clones whereas tyrosinase mRNA level was unaltered between melanotic (M3) and amelanotic (A4 and A7) cells. These results indicate that B16 melanoma cells have heterogeneous cell populations consisting of different biosynthetic and metastatic properties. These clones may provide good tools for studying the relationship between the phenotypes of melanogenesis biosynthesis of extracellular macromolecules and migrating activity of melanoma cells.

Animals↗
PubMed1996

A compound heterozygote for familial hypercholesterolaemia with a homozygous mother.

Homozygous familial hypercholesterolaemia (FH) is a rare disorder in which the patients develop severe hypercholesterolaemia and premature coronary atherosclerosis from childhood. Here we report a unique family with clustering of homozygous FH. The proband was a 25-year-old man, who showed marked hypercholesterolaemia, multiple xanthomas and severe coronary atherosclerosis. His mother also showed the typical characteristics of homozygous FH. Sequencing analysis of the low-density lipoprotein receptor gene revealed that he was a compound heterozygote, carrying two different point mutations. One was a novel mutation, FH Wakayama (Cys-->Ser at 317), derived from his mother, and the other was a recurrent mutation, FH Niigata (T-->C at 1845 + 2, 5' splice signal in intron 12), derived from his father. The proband we report seems to be a very rare case of an FH homozygote born from a homozygous mother.

Adult↗
PubMed1996

Solitary piloleiomyoma with an unusual clinical appearance.

We present a 66-year-old man with a solitary piloleiomyoma on the forehead. The lesion had unusual clinical features; it was a crusted and dome-shaped nodule, which exhibited rapid growth. Histology revealed the typical features of piloleiomyoma. The unusual clinical features may have been due to its unique location, in which the downward growth of the tumor was prevented by the underlying frontal bone.

Aged↗
PubMed1996

A case of acrogeria associated with late-onset focal dermal elastosis.

We report a case of acrogeria. Biochemical analysis of patient fibroblasts demonstrated unaltered type III collagen synthesis. An RNA hybridization assay demonstrated that the levels of type I and elastin mRNAs were normal. The patient had multiple yellow papules on the neck, axilla and inguinal area which clinically and histologically resembled late-onset focal dermal elastosis. Both diseases may have some similar abnormal metabolism of connective tissue.

Adolescent↗
PubMed1996

Systemic scleroderma and perforating granuloma annulare: differential diagnosis from calcinosis.

BACKGROUND: Systemic scleroderma is a disorder of unknown etiology with skin sclerosis. Its major histological features are swollen and homogenized collagen bundles. OBJECTIVE AND METHODS: We describe 2 patients with systemic sclerosis who have multiple umbilicated nodules indistinguishable from calcinosis cutis. RESULTS: Histological examinations including Von Kossa staining revealed features of perforating granuloma annulare, but not of calcinosis cutis. CONCLUSION: The association may not be fortuitous but both diseases may be etiologically related.

Adult↗
PubMed1995

Isolation of a novel cDNA that encodes a protein localized to the pre-acrosome region of spermatids.

We have identified a novel cDNA clone, named AZ1, obtained from a cDNA library of mRNA prepared from C3H10T1/2 cells that had been transiently exposed to 5-azacytidine, a potent demethylating reagent. The amount of transcript increased with 5-azacytidine treatment of C3H10T1/2 cells and the transcript was highly expressed in mouse testis. As the mutant mouse jsd/jsd, which has a defect in germ cell maturation, barely expressed the transcript, the message was expected to be expressed specifically in spermatocytes. The mRNA was detected at significant levels in the testes from mice aged 16 days after birth, suggesting that its expression started at the pachytene spermatocyte stage. The elucidated nucleotide sequence contained a 2841-nucleotide open reading frame, and the expected amino acid sequence had a molecular mass of 107,254 Da. Specific antibodies raised against the fusion protein including glutathione S-transferase revealed an approximately 130-kDa band of a translation product in testis and in cultured cells transfected with AZ1 cDNA in the expression vector on Western-blot analysis. The protein was localized to the pre-acrosome region of round and elongated spermatids. However, it was not detected at a more advanced stage of spermatids, i.e. just before their release from Sertoli cells. This protein may play an important role in spermatogenesis.

Acrosome↗
PubMed1995

A single point mutation in the splice donor site of the low-density-lipoprotein-receptor gene produces intron read-through, exon-skipped and cryptic-site-utilized transcripts.

Familial hypercholesterolemia is a genetic disorder caused by mutations of the low-density-lipoprotein (LDL) receptor gene. We characterized the structures of LDL receptor mRNA transcripts in the fibroblasts of a homozygous patient carrying a single base substitution (T-->C) at the 5' splice donor site of intron 12 of the LDL receptor gene. We identified three aberrant transcripts as a consequence of intron-12 read-through, exon-12 skipping and utilization of a cryptic splice donor site. Only a point mutation at the 5' splice donor site caused the production of three alternatively spliced products. None of these transcripts produced a functional LDL receptor protein in this patient.

Alternative Splicing↗
PubMed1995

Stimulation of cell proliferation and autoregulation of elastin expression by elastin peptide VPGVG in cultured chick vascular smooth muscle cells.

Synthetic elastin peptides, VPGVG or its polymer (VPGVG)n, enhanced the proliferation of smooth muscle cells 1.5-fold during 48 h treatment at the concentrations over 10(-6) M or 1.0 microgram/ml, respectively. Monomeric and polymeric VPGVG sequences reduced elastin synthesis and its mRNA level to one-third and one-half of control respectively under the conditions in which the proliferation of cells were enhanced, but did not change collagen synthesis as measured by bacterial collagenase digestion. The elastin-specific autoregulation by elastin fragments may reflect the feedback regulation of elastin expression which may play an essential role in elastin metabolism under the normal and diseased conditions.

Amino Acid Sequence↗
PubMed1995

Cell cycle-dependent regulation of elastin gene in cultured chick vascular smooth-muscle cells.

A study was made of the relationship between elastin expression and the proliferative state of chick vascular smooth-muscle cells. Confluent cells of primary culture brought to a quiescent state by the deprivation of serum for 72 h exhibited a 5-, 3.5- and 2-fold increase in elastin synthesis, elastin mRNA level and transcriptional activity of elastin gene respectively over those in the proliferative state. On re-addition of serum in serum-deprived culture, cells started to proliferate, and elastin synthesis, its mRNA level and transcription of the gene decreased to the level of a proliferative state within 24 h, indicating that elastin expression in smooth-muscle cells was controlled by their growth states at least in part at a transcriptional level. A comparable increase in elastin mRNA level was observed when the cell growth was arrested by suspension culture for 72 h. When the cells were synchronized at the G1/S phase with thymidine/hydroxyurea treatment, elastin expression at the G1/S phase was greater than that at the G2/M phase during cell cycling. Elastin mRNA level at the G0 phase brought about by serum-deprivation or suspension culture predominated over that at the G1/S phase during cell cycling. These results indicate that gene expression of elastin and cell cycle are tightly coupled, which is independent of the presence of serum or adhesive state, and that elastin expression could be a biochemical marker for the growth states of smooth-muscle cells.

Animals↗
PubMed1995

Overexpression of DNA methyltransferase in myoblast cells accelerates myotube formation.

We overexpressed mouse DNA methyltransferase in murine C2C12 myoblast cells and tested the isolated clones for their ability to differentiate. Significant numbers of the clones showed distinct myotubes 24 h after the isolated transformants had been induced to differentiate, whereas the parent C2C12 cells did not form myotubes at this time point. Transfection of the vacant vector or the plasmid containing the reverse-oriented DNA methyltransferase cDNA did not provide significant numbers of transformants with the accelerated differentiation phenotype, suggesting that the effect is caused by the expression of DNA methyltransferase. The expressions of skeletal muscle myosin and creatine kinase in clones that showed the accelerated differentiation-phenotype were also induced about 24 h earlier and at higher levels relative to the parent C2C12 or the control cells, indicating that the entire process of myogenesis had been accelerated. All the methyltransferase-transfected clones, regardless of their phenotypes, demonstrated about threefold higher DNA methyltransferase activity and higher methylation levels than those of the clones transfected with vector alone or the reverse-oriented plasmid. At the early stage of transfection of the sense-oriented plasmid, high de novo methylation activities were detected. We consider it likely that this high de novo methylation activity is the reason for the high methylation levels and the accelerated myotube formation of the clones transfected with the sense-oriented plasmid. In some transformants which showed the accelerated differentiation phenotype, MyoD1 was already fully expressed under the growth conditions while, in control cells, MyoD1 was expressed at low levels. This elevated level of MyoD1 transcription could account for the accelerated myotube formation observed in the transformants. The methylation state of the HpaII sites in exon 1 through exon 2 of the MyoD1 gene and the expression of the MyoD1 transcript are positively correlated.

Animals↗
PubMed1995

Nucleotide sequence and the molecular evolution of a new A2 gene in the DQ subregion of the bovine major histocompatibility complex.

cDNA clones encoding the bovine major histocompatibility complex (MHC) class II DQ alpha chain were isolated. One clone, MQ9, encoded a primary translated product of 255 amino acids, with a signal peptide of 23 amino acids and a mature polypeptide of 232 amino acids. A new A2 gene in the DQ subregion of the bovine genome was identified from a comparison of amino acid sequences encoded by class II A genes among several species and the construction of a phylogenetic tree. It was revealed that MQ9 is most closely related to the ovine DQA2 genes among sequences from various mammalian species. By contrast, the BoLA-DQA genes previously isolated are more closely related to ovine DQA1 than to the BoLA-DQA2 gene, and they represent BoLA-DQA1 genes. Thus, the presence of two BoLA A genes, which may be expressed and functional in the bovine, as well as in sheep was confirmed. A large number of amino acids unique to products of DQA2 genes of bovine and ovine origin were identified when the predicted amino acid sequences for both species were compared, and most of the DQA2-specific residues were located in the alpha 1 domain and were conserved with respect to products of DQA1 genes of ruminants. Thus, several characteristics of the bovine DQA genes were found to differ from those of human and rodent genes, despite similarities in gene structure and in nucleotide sequence.

Amino Acid Sequence↗
PubMed1995

Presence of elastin-related 45-kDa fragment in culture medium: specific cleavage product of tropoelastin in vascular smooth muscle cell culture.

Significant amount of 45-kDa polypeptide was found to be present in the cultured medium of chick aortic smooth muscle cells. The polypeptide as well as tropoelastin (65-kDa) reacted with monoclonal antibody for tropoelastin. Pulse-chase experiments revealed that the relative density of the 45-kDa polypeptide to tropoelastin increased with chase periods. Partially purified radioactive tropoelastin (65-kDa) was converted to a 45-kDa polypeptide fragment in the test tube. The processing from the 65- to the 45-kDa polypeptide in the test tube was inhibited by ethylenediaminetetraacetic acid but not by N-ethylmaleimide and aminophenylmethylsulfonyl fluoride. These results indicate that the 45-kDa fragment is a degradation product of tropoelastin and that processing is mediated by enzymatic cleavages with metal proteinase. Fully hydroxylated tropoelastin treated with ascorbic acid was more resistant to the enzymes than underhydroxylated tropoelastin with scorbutic condition, suggesting that the structural stability of tropoelastin is also involved in the processing rate.

Animals↗
PubMed1995

Enhanced tropoelastin-degrading activity during cell passages in cultured smooth muscle cells.

Tropoelastin expression in vascular smooth muscle cells during serial cell passages from the primary to the tertiary culture was studied. The level of tropoelastin was found to be greatly reduced as the number of cell passages increased. The translational activity and level of elastin mRNA were essentially unchanged throughout the cell passages. The reduction in tropoelastin expression was not due to the repetitive trypsin treatment nor to the prolyl hydroxylation level of the newly-synthesized elastin. A comparable decline in tropoelastin expression was also found with increasing cell division in the primary cultures plated at different cell densities. A pulse-chase experiment revealed that the newly-synthesized elastin in the tertiary culture degraded more rapidly than that in the primary culture. The culture medium harvested from the tertiary culture exhibited a higher tropoelastin-degrading activity than that from the primary culture in the test-tube. The degrading activity of the tertiary culture was inhibited by the addition of 1 mM ethylenediaminetetraacetic acid or 1 mM phenylmethylsulfonyl fluoride, but not by 1 mM N-ethylmaleimide. These results suggest that the reduction in tropoelastin expression during the cell passages from the primary to the tertiary culture is due to the enhanced tropoelastin-degrading activity of the tertiary culture. The transition to tropoelastin-degrading phenotype during cell passages may explain the biological mechanisms of smooth muscle cell migration from the media to the intima observed in the pathological conditions.

Animals↗