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Biomedical subjects

N Maeda

Publications and source records attributed to N Maeda.

At least 505 records · Page 28Linked to original sources
PubMed1988

Complex events in the evolution of the haptoglobin gene cluster in primates.

Southern blot analyses of genomic DNA show that new world monkeys have only one haptoglobin gene but that chimpanzees, gorillas, orangutans, and old world monkeys have three. Humans have two: haptoglobin (Hp) and haptoglobin-related (Hpr). These observations suggest that a triplication of the haptoglobin locus occurred after the divergence of the new world monkeys, followed by a deletion of one locus in humans. To investigate these events, we have cloned the haptoglobin gene cluster in chimpanzee. The organization of the Hp and Hpr genes in chimpanzees is the same as in humans, including a retrovirus-like sequence in the first intron of Hpr. The third gene, which we name Hpp for haptoglobin primate, is 16 kilobases downstream of Hpr. A second copy of the retrovirus-like sequence occurs between Hpr and Hpp. The nucleotide sequence of the chimpanzee Hpp gene suggests that it may code for a functional protein, but the chimpanzee Hpr gene has a single base deletion in exon 5 that causes a frameshift. Comparison of the human and chimpanzee sequences suggests that the human Hpr gene was generated by a homologous unequal crossover between ancestral Hpr and Hpp genes. The crossover point lies within a 1.3-kilobase region containing exon 5 and 500 nucleotides 3' to the genes, but the exact point is obscured by a subsequent gene conversion event.

Animals↗
PubMed1988

Erythrocyte aggregation as a determinant of blood flow: effect of pH, temperature and osmotic pressure.

The effect of pH, temperature and osmotic pressure on velocity of erythrocyte aggregation was quantitatively examined with a rheoscope combined with a video-camera, an image analyzer and a computer, (a) in an artificial medium containing fibrinogen and albumin and (b) in diluted autologous plasma. (1) With increasing pH of the medium, the velocity of erythrocyte aggregation increased. (2) The velocity of erythrocyte aggregation in the artificial medium increased as the temperature rose. However, in 70% autologous plasma the velocity was minimum at 15-18 degrees C, increasing both above and below this temperature (above 30 degrees C, the velocity saturated). (3) The velocity of erythrocyte aggregation decreased in hypotonic medium, while it increased in hypertonic medium (at osmotic pressures higher than 400 mOsm, the velocity decreased). The mechanism of erythrocyte aggregation is discussed with special reference to the morphological changes produced by pH, temperature and osmotic pressure, and the implications of the phenomena for oxygen transport to tissues in (patho)physiological situations are considered.

Adult↗
PubMed1988

An anionic charge barrier in the guinea pig cochlea.

A charge barrier has been found in the glomerular basement membrane of the kidney and plays an important role in the filtration of solutes. In the present study, we used electron microscopy to localize anionic sites of a similar charge barrier in the guinea pig cochlea. Polyethyleneimine (PEI) was used as a cationic marker to detect anionic sites. Our results showed a localization of PEI with regular interspaces, indicating the anionic sites to the charge in the capillary basement membrane of the stria vascularis and the spiral ligament, and in the basal lamina of Reissner's membrane and the spiral prominence. This charge barrier, as well as structural size barrier, may play an important role in the maintenance of normal inner ear functions.

Animals↗
PubMed1988

Establishment of hamster blastocyst-derived embryonic stem (ES) cells.

The establishment of four ES cell lines from the Syrian "golden" hamster (Mesocricetus auratus) is described. The cells can be maintained in the undifferentiated state when grown on primary mouse embryonic fibroblast feeder layers. In suspension culture they spontaneously differentiate into embryoid bodies of increasing complexity which contain a variety of tissues including embryonic ectoderm and myocardium. All four lines--one female and three male--are karyotypically normal with 44 chromosomes. Hamster is the second species from which ES cells have been established. As in mouse, the cells should be useful for developmental and transgenic studies.

Animals↗
PubMed1988

Establishment and characterization of a murine Gardner's osteosarcoma cell line (GOS/T): purification of bone-specific alkaline phosphatase (ALPase) and use of anti-ALPase as a diagnostic acid.

Gardner's murine osteosarcoma has been successfully cultured and maintained as a clonal cell line (GOS/T), capable of forming a tumor with neoplastic bone and osteoid tissue by inoculation of 1 x 10(7) cultured cells. Alkaline phosphatase (ALPase) level in the culture medium and the serum from the tumor-bearing mice increased significantly three times and 16 to 105 times, respectively. The bone-specific ALPase was purified by butanol extraction following gel-filtration through Sephadex G-200. The molecular weight of ALPase was determined as 420,000, which was three times 140,000, the molecular weight of a subunit. Immunofluorescence staining using anti-ALPase antiserum, which was made by inoculation of partially purified ALPase, revealed a positive reaction for GOS/T cell line and osteoblasts of the fetal mice. The data presented here suggest that the cell line of GOS/T can be regarded as an established cell line, and the immunological reaction with anti-ALPase antibody is a useful model system to study human osteosarcoma.

Alkaline Phosphatase↗
PubMed1988

Arterialized venous flap coverage for skin defects of the hand or foot.

This is a report on 15 patients whose skin defects of the hand or foot were covered using an arterialized venous flap, with only arterial blood flowing through the vein of the flap. Success appeared to be influenced by the donor site and size of the flap. When a small-size flap from the forearm was used, the success rate was almost 100 percent; however, there was a 50 percent failure rate when a large flap from the leg was used. This technique provides not only flap coverage for exposed bone and tendon, but also provides one-stage procedures for digits in need of revascularization and skin coverage.

Adult↗
PubMed1988

Targeted mutation of the Hprt gene in mouse embryonic stem cells.

The hypoxanthine-guanine phosphoribosyltransferase (Hprt) gene has been mutated in mouse blastocyst-derived embryonic stem cells by site-directed homologous recombination. Embryonic stem cells were electroporated in the presence of a targeting DNA fragment containing two specific features: (i) The targeting DNA contained a promoterless neomycin phosphotransferase (neo) gene that, when located within the endogenous Hprt locus, could be transcribed from the promoter of the target locus. (ii) The targeting fragment had two short regions of homology with the endogenous Hprt gene: one, 132 base pairs long and the other, 1.2 kilobase pairs long. Targeted cells in which the designed homologous recombination event occurred were isolated either by selection with G418 followed by 6-thioguanine or by selection with 6-thioguanine alone. Even though less than 2 kilobases of homology existed between the exogenous and target DNAs, an average of 2.6 embryonic stem cells were successfully targeted for every 10(5) colonies surviving electroporation. Six of the Hprt- cell lines showed homologous recombination. These six lines were further analyzed by nucleotide sequencing a fragment that spans one crossover point after amplification by the polymerase chain reaction. Four lines had the expected sequence, whereas two lines had small deletions abutting the 132-base-pair region of homology.

Amino Acid Sequence↗
PubMed1988

Structure and nucleotide sequence of the rat intestinal vitamin D-dependent calcium binding protein gene.

The vitamin D-dependent intestinal calcium binding protein (ICaBP, 9 kDa) is under transcriptional regulation by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], the hormonal active form of the vitamin. To study the mechanism of gene regulation by 1,25-(OH)2D3, we isolated the rat ICaBP gene by using a cDNA probe. Its nucleotide sequence revealed 3 exons separated by 2 introns within approximately 3 kilobases. The first exon represents only noncoding sequences, while the second and third encode the two calcium binding domains of the protein. The gene contains a 15-base-pair imperfect palindrome in the first intron that shows high homology to the estrogen-responsive element. This sequence may represent the vitamin D-responsive element involved in the regulation of the ICaBP gene. The second intron shows an 84-base-pair-long simple nucleotide repeat that implicates Z-DNA formation. Genomic Southern analysis shows that the rat gene is represented as a single copy.

Amino Acid Sequence↗
PubMed1988

Structure and regulation of the rat 1,25-dihydroxyvitamin D3 receptor.

Complementary DNA clones encoding the entire rat 1,25-dihydroxyvitamin D3 receptor were isolated, and the total 423-amino acid sequence was deduced. The amino acid sequence of the rat receptor is similar but not identical to the reported human receptor sequence. The cysteine-rich DNA-binding domain is completely conserved and the steroid-binding domain is greater than 93% conserved between rat and human. The cDNA was used as a probe in blot analysis of polyadenylylated RNA to show that the 1,25-dihydroxyvitamin D3 receptor mRNA is a single 4.4-kilobase mRNA that is expressed in intestine and kidney, slightly expressed in heart, and not detectable in liver and spleen. The receptor mRNA concentration is markedly increased during development of the rat intestine between day 14 and day 21, in accord with previous results obtained by measurement of receptor concentration by ligand binding or immunoblotting.

Amino Acid Sequence↗
PubMed1988

Isolation and expression of rat 1,25-dihydroxyvitamin D3 receptor cDNA.

The cDNA for the 1,25-dihydroxyvitamin D3 receptor has been isolated by screening a rat kidney lambda gt11 expression library with a mixture of three previously identified anti-receptor monoclonal antibodies. Protein produced from this cDNA reacts with each of the three monoclonal antibodies but does not react with an antibody specific for the porcine receptor or an irrelevant antibody. In addition, the expressed protein specifically binds 1,25-dihydroxyvitamin D3 but does not bind estradiol, progesterone, testosterone, cortisol, or aldosterone. The nucleotide sequence for this cDNA has been determined and used to deduce the amino acid sequence of the protein. This amino acid sequence contains a region that matches perfectly the partial amino acid sequence determined for the receptor isolated from porcine intestine. This amino acid sequence also closely matches the reported amino acid sequence of the putative DNA-binding finger of the avian 1,25-dihydroxyvitamin D3 receptor.

Amino Acid Sequence↗
PubMed1988

UGA suppressors in Saccharomyces cerevisiae: allelism, action spectra and map positions.

Sixty independent UGA suppressors of Saccharomyces cerevisiae have been studied. They are dominant and are divided into 16 groups (loci) by recombination. Suppressors representing these loci are divided into two classes by action spectra; four in class 1 (a broad action spectrum) and 12 in class 2 (a narrow action spectrum). Class 1 suppressors are less frequent in terms of not only total number but also number per locus than class 2 suppressors, indicating difference in either or both mutation frequency and selective pressure between suppressors of the two classes. Two of the class 1 suppressors, SUP152 and SUP161, do not recombine with SUP28 and SUP33, leucine-inserting UAA suppressors, respectively, indicating that they are mutations in genes coding for tRNA(Leu)UUA. Of the remaining two class 1 suppressors, SUP160 which causes lethality in the psi+ cytoplasm is mapped on chromosome XV very close to the centromere, and SUP165 on the right arm of chromosome XIV 44 cM distal to lys9. Of the class 2 suppressors, ten do not recombine with one or another of previously known UGA suppressors. The remaining two class 2 suppressors, SUP154 and SUP155, are mapped on the left and right arms of chromosome VII, respectively.

Alleles↗
PubMed1988

Molecular evolution of intergenic DNA in higher primates: pattern of DNA changes, molecular clock, and evolution of repetitive sequences.

A 3.1-kb intergenic DNA fragment located between the psi beta-globin and delta-globin genes in the beta-globin gene cluster was cloned from gorilla, orangutan, rhesus monkey, and spider monkey, and the nucleotide sequence of each fragment was determined. The phylogeny of these four sequences, together with two previously published allelic sequences from humans and one from chimpanzee, was constructed, and the accumulation of mutations in the region was analyzed. The sites of base substitutions are not evenly distributed within the region: two Alu repeats have accumulated 0.21 + 0.02 substitutions/site with 0.15 + 0.008 substitutions/site in the remainder of the fragment. The occurrence of substitutions at neighboring sites is more frequent than would be expected if they were independent. The observed excesses disappear when ancestral -CG- dinucleotide sites are excluded. The phylogenetic relationships of the sequences indicate that the human sequence shares a most recent coancestor with the chimpanzee sequence. The data also show that great apes have accumulated fewer mutations in this part of the genome than has the rhesus monkey. The relative rates of accumulation of 12 kinds of nucleotide substitution in the region during primate evolution are asymmetric in the DNA strands. From these rates of accumulation, the origin of a simple stretch of sequence near the 3' end of the 3.1-kb fragment was deduced to be a sequence comprising 50% T and 50% C on one strand. The two oppositely oriented Alu sequences in the 3.1-kb region were inserted at their present positions before the divergence of the New-World monkeys from other lineages. Our analysis shows that the nucleotide sequences of the two Alu repeats in spider monkey are unexpectedly similar both to each other and to the deduced ancestral sequence of Alu repeats. The data suggest that there has been some type of recombinational event between the spider monkey Alu repeats but that it was not a simple gene conversion.

Animals↗
PubMed1988

Purification and characterization of P400 protein, a glycoprotein characteristic of Purkinje cell, from mouse cerebellum.

P400 protein is a concanavalin A (Con A)-binding, 250-kilodalton glycoprotein characteristic of cerebellum. Extraction conditions for P400 protein were investigated, and complete solubilization of P400 protein from a submicrosomal fraction (P31 fraction) of mouse cerebellum was attained by the combination of 4% Zwittergent 3-14 and 4 M guanidinium chloride. The solubilized P400 protein was purified using Sepharose CL-4B and Con A-Sepharose chromatography. A monoclonal antibody (18A10) was prepared against P400 protein. Endo-beta-N-acetylglucosaminidase F digestion of P400 protein revealed that P400 protein has a small number of asparagine-linked oligosaccharide chains and that the epitope that is recognized by 18A10 monoclonal antibody is not on the asparagine-linked oligosaccharide portion. Tissue distribution of P400 protein was investigated by immunoblot analysis using 18A10 monoclonal antibody. P400 protein was abundant in the cerebellum, but a very small amount of P400 protein or related antigen was also detected in other parts of the nervous system and in nonneural tissues. Immunohistochemical studies indicated that P400 protein was distributed abundantly in the soma, the dendritic arborization, and the axon of the Purkinje cell. No immunoreaction was observed in the other types of cells.

Acetylglucosaminidase↗
PubMed1988

Characterization of microtubule-associated protein 2 from mouse brain and its localization in the cerebellar cortex.

Microtubule-associated protein (MAP) 2 was purified from the microtubule fraction of mouse brain by heat treatment and BioGel A-5m gel filtration. The purified preparation showed a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis using both a gradient gel (3.75-12.5%) and a low-percentage gel (5%), a finding indicating that MAP2B was absent under the conditions used. Amino acid analysis revealed that mouse MAP2 was an acidic protein with an isoelectric point (pI 4.5) and amino acid composition similar to those of porcine brain MAP2. Immunoblot analysis indicated that the antigens that reacted with MAP2 antiserum were present in large quantities in mouse brain. However, we also found a weak reaction in various tissues other than brain, and the major antigens involved were recognized to be common molecular species with the same molecular mass, 162 and 170 kilodaltons. Using antiserum against mouse brain MAP2, the developmental localization patterns of MAP2 in the mouse cerebellar cortex were studied by immunohistochemistry. MAP2 was mainly localized in the neuronal cells throughout development, with the expression in Purkinje cell dendrites being especially remarkable in the growth of arborization from postnatal day 3 to day 20. At the mature stage, the reaction was strong in the dendritic tree but very weak in the proximal dendrites and cell bodies.

Amino Acids↗
PubMed1988

1 alpha,25-Dihydroxyvitamin D3[1 alpha,25-(OH)2D3]-26,23-lactone inhibits 1,25-(OH)2D3-mediated fusion of mouse bone marrow mononuclear cells.

Vitamin D3 and its hormonally active metabolite 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] can be metabolized to a number of daughter metabolites, including 1 alpha,25-(OH)2D3-26,23-lactone; this latter compound has four diastereoisomers. The 23(S),25(R)-lactone (naturally occurring) and the 23(R),25(S)-1 alpha,25-(OH)2D3-26,23-lactone are both known to be able to inhibit bone resorption induced by 1 alpha,25-(OH)2D3 under in vivo or in vitro conditions. To understand the mechanism of the inhibitory action of these two isomers on bone resorption we investigated the effects of 1 alpha,25-(OH)2D3-26,23-lactone on unfractionated mouse bone marrow cells in vitro. The addition of 1 alpha,25-(OH)2D3 to these cultures dose-dependently stimulated the formation of multinucleated cells over a range of 10(-9) - 10(-7) M. The 23(S),25(S)- and 23(R),25(R)-1 alpha,25-(OH)2D3-26,23-lactones also increased the number of multinucleated cells, whereas the 23(S),25(R)- and 23(R),25(S)-1 alpha,25-(OH)2D3-26,23-lactones failed to do so. In addition, these latter two diastereomers inhibited the 1 alpha,25-(OH)2D3 stimulation of multinucleated cell formation, although the 23(S),25(S)- and 23(R),25(R)-1 alpha,25-(OH)2D3-26,23-lactones and 24R,25-(OH)2D3 did not. These multinucleated cells responded to calcitonin and contained tartrate-resistant acid phosphatase, both of which are characteristic of osteoclasts. The present data suggest that inhibition of multinucleated cell formation is the mechanism by which the 23(S),25(R)- or 23(R),25(S)-1 alpha,25-(OH)2D3-26,23-lactone inhibits bone resorption induced by 1 alpha,25-(OH)2D3.

Acid Phosphatase↗