Bilateral centripetal lipid keratopathy with diffuse anterior scleritis.
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Cataract↗
Biomedical subjects
M Morisawa
Publications and source records attributed to M Morisawa.
Brachyury (T) gene expression and notochord development in Oikopleura longicauda (Appendicularia, Urochordata).
Appendicularia (Larvacea) is a subgroup of Urochordata (Tunicata) comprised of holoplanktonic organisms that retain their tailed architecture throughout their life history, while other tunicates, including ascidians and doliolids, resorb the tail after metamorphosis. In order to investigate the characteristics of the appendicularian unresorbed notochord, we isolated a partial genomic clone and a full-length cDNA sequence homologous to the mouse Brachyury (T) gene from the appendicularian Oikopleura longicauda. Brachyury is known to be predominantly expressed in the notochord cells and plays an important role in their differentiation in other chordates. While phylogenetic analysis robustly supports the orthology of the isolated Brachyury gene, the exon-intron organization found in the genomic clone was distinct from that well-conserved among other T-box genes. In addition to a detailed observation of notochord development in living specimens, whole-mount double in situ hybridization was carried out using a Brachyury probe along with a muscle actin probe. The Brachyury transcripts were found in the notochord of the tailbud embryos and persisted into later stages. The present study highlights characteristics of notochord development in the appendicularian. Furthermore, these results provide basic knowledge for comprehensive understanding of the cellular- and molecular-based mechanisms needed to build the characteristic cytoarchitecture of notochord that varies among tunicate species.
Amino Acid Sequence↗
Immunochemical assay of hemoglobin with N(epsilon)-(carboxymethyl)lysine at lysine 66 of the beta chain.
BACKGROUND: N(epsilon)-(Carboxymethyl)lysine (CML), a well-characterized and major advanced glycation end product structure, is produced via a Maillard reaction by nonenzymatic glycation and/or oxidation. Although few of the carboxymethylation sites of lysine residues on proteins have been identified, it is known that the possible lysine glycation site in hemoglobin (Hb) is Lys-66 on the beta chain. We aimed to develop an assay for the Hb with a CML (CML-Hb) site specific to Lys-66 on the Hb beta chain and to determine whether the lysine residue at that site is carboxymethylated. METHODS: Ala-His-Gly-Lys-Lys(CM)-Val-Leu-Gly-Ala-Phe-Ser-Cys, the peptide derived from the beta chain of human Hb, was synthesized as an immunogen, and a monoclonal antibody against the peptide was prepared. A latex immunoassay method was established using the antibody on an automatic analyzer. In this study, 20 samples from healthy subjects and 80 samples from nondiabetic patients undergoing hemodialysis (HD) were analyzed. RESULTS: The latex immunoassay method using the antibody correlated significantly with the ELISA method using the antibody (r = 0.95; P <0.001). Between healthy subjects (n = 20) and nondiabetic HD patients (n = 80), a significant difference was seen in circulating CML-Hb (525 +/- 76 vs 778 +/- 137 pmol CML/mg of Hb; P <0.0001). CONCLUSION: The latex method for the CML-Hb site specific to Lys-66 on the beta chain can measure large numbers of samples on an automatic analyzer.
Acetaldehyde↗
Muscle actin genes and muscle cells in the appendicularian, Oikopleura longicauda: phylogenetic relationships among muscle tissues in the urochordates.
Appendicularians (larvaceans) are planktonic tunicates. They possess a tail throughout their life, which marks a distinct difference between appendicularians and the other tunicate groups, including salps, doliolids, pyrosomes, and ascidians. We isolated cDNA clones encoding muscle-type and cytoplasmic-type actin isoforms from the appendicularian, Oikopleura longicauda. The types of the actin isoforms were confirmed by amino acid identities in the diagnostic residues compared to those of the other chordate muscle and cytoplasmic actins. Interestingly the appendicularian muscle actin isoform sequence has an intermediate feature between the ascidian tail (larval) muscle actin isoform and the body-wall (adult) muscle actin isoform. Analysis of a genomic clone from a muscle actin gene revealed that it contains only one intron in the coding region. It is located at a position that does not correspond to those of any introns reported in other deuterostome actin genes. Whole-mount in situ hybridization shows that the muscle actin transcript is detected specifically in the tail muscle cells of late tailbud-stage embryos. A signal is also found transiently in the region where the heart will form. In adults, the gene is expressed in tail muscle cells but not in the heart. Together with results of cytochemical and histochemical studies demonstrating the structure of muscle tissue, the present study highlights characteristics of appendicularian muscle organization, which are compared with those of the larval and adult muscle of ascidians.
Actins↗
Membrane hyperpolarization removes inactivation of Ca2+ channels, leading to Ca2+ influx and subsequent initiation of sperm motility in the common carp.
Change of osmolality surrounding spawned sperm from isotonic to hypotonic causes the initiation of sperm motility in the common carp. Here we show that membrane-permeable cAMP does not initiate motility of carp sperm that is quiescent in isotonic solution, and that motility of the demembranated sperm can be reactivated without cAMP. Furthermore, the cAMP level does not change during the initiation of sperm motility, and inhibitors of protein kinase do not affect sperm motility, suggesting that no cAMP-dependent system is necessary for the regulation of sperm motility. Sperm motility could not be initiated in Ca(2+)-free hypoosmotic solutions, and significant increase in the intracellular Ca(2+) level was observed by a Ca-sensitive fluorescence dye during hypoosmolality-induced active motion period. The demembranated sperm cells were fully reactivated in the solutions containing 10(-7) to 10(-5) M Ca(2+). Ca(2+) channel blockers such as verapamil and omega-conotoxin reversibly inhibited the initiation of sperm motility, suggesting that Ca(2+) influx is the prerequisite for the initiation of carp sperm motility. Motility of intact sperm was completely blocked; however, that of the demembranated sperm was not inhibited by the calmodulin inhibitor W7, suggesting that the calmodulin bound close to the plasma membrane participated in the initiation of sperm motility. Flow cytometric membrane potential measurements and spectrophotometric measurements by using fluorescence dyes showed transient membrane hyperpolarization on hypoosmolality-induced motility. This article discusses the role of membrane hyperpolarization on removal of inactivation of Ca(2+) channels, leading to Ca(2+) influx at the initiation of carp sperm motility.
Animals↗
Cyclic AMP- and calmodulin-dependent phosphorylation of 21 and 26 kda proteins in axoneme is a prerequisite for SAAF-induced motile activation in ascidian spermatozoa.
Sperm activating and -attracting factor (SAAF), derived from the egg of the ascidian Ciona, activates sperm motility through adenosine 3':5'-cyclic monophosphate (cAMP)-synthesis. A demembranated preparation of intact immotile sperm without SAAF was shown to require cAMP for reactivation. However, a demembranated preparation of intact motile sperm treated with SAAF did not require cAMP for reactivation, suggesting that cAMP is a prerequisite factor for SAAF-dependent activation of sperm motility. Furthermore, a cAMP-dependent protein kinase (PKA) inhibitor, H-89, was found to inhibit sperm motility. During in vivo or in vitro activation of sperm motility by SAAF or cAMP, a 26 kDa axonemal protein and 21 kDa dynein light chain were phosphorylated, respectively, suggesting the involvement of PKA-dependent phosphorylation of these proteins in sperm activation. The calmodulin antagonist, W-7, and an inhibitor of calmodulin-dependent myosin light chain kinase, ML-7, also inhibited the activation of sperm motility. Inhibition was reversed by the addition of phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine. Demembranated preparations of immotile sperm in the presence of W-7 or ML-7 were reactivated by cAMP, suggesting that calmodulin participated in sperm activation and that cAMP synthesis was followed by activation of a calmodulin-dependent mechanism.
Animals↗
Phosphorylation of axonemal 21 kDa and 26 kDa proteins modulates activation of sperm motility in the ascidian, Ciona intestinalis.
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Animals↗
Membrane hyperpolarization by sperm-activating and -attracting factor increases cAMP level and activates sperm motility in the ascidian Ciona intestinalis.
In the ascidian Ciona intestinalis (and C. savignyi), sperm-activating and -attracting factor (SAAF) is released from the egg at fertilization and stimulates both Ca(2+) influx and a transient increase in cAMP level of the sperm, leading to the activation of sperm motility (M. Yoshida et al., 1994, Dev. Growth Differ. 36, 589-595). In this paper we show in C. intestinalis that valinomycin, a potassium-selective ionophore, as well as SAAF, activated sperm motility, and this activation was suppressed by extracellular high K(+). Membrane potential measurements showed that both SAAF and valinomycin increase K(+) permeability of sperm and induce membrane hyperpolarization, the amplitude of which depends on the external K(+) concentration. The membrane potential and intracellular K(+) concentration of Ciona sperm without SAAF were estimated to be about -50 mV and 560 +/- 40 mM, respectively. After treatment with SAAF or valinomycin the membrane potential became almost equal to the equilibrium potential of K(+) (-100 mV), and the cAMP level increased in artificial seawater. A potent voltage-dependent K(+) channel blocker, MCD peptide, at the concentration of 10 microM blocked SAAF-induced hyperpolarization of the cells, increase in cAMP, and sperm motility. These results suggest that membrane hyperpolarization produced by the opening of K(+) channels elevates cAMP synthesis and leads to the activation of sperm motility in Ciona.
Animals↗
Loss of test cells leads to the formation of new tunic surface cells and abnormal metamorphosis in larvae of Ciona intestinalis (Chordata, ascidiacea).
The larvae of the ascidian Ciona intestinalis from which the chorion with the test cells and follicle cells were removed developed normally without the test cells until the early tailbud stage. A number of round-shaped cells morphologically similar to the test cells but with different lectin affinities and autofluorescence, then appeared on the neck region of the demembranated embryos. The new cells had three different types: round, particulate, and granular, and these cells increased in number after the late tailbud stage. The morphology of the adhesive papillae, tunic layers and epidermis of the demembranated larvae was similar to that of control larvae; however, the affinity to lectins was different in the swimming period. Control larvae attached to the substratum after the swimming period, resorbed the tail completely and underwent rotation of the visceral organs. Conversely, rotation occurred before completion of tail resorption in the demembranated larvae. Furthermore, the metamorphic events progressed more slowly in the demembranated larvae. These results suggest that the test cells play important roles in normal development and morphogenesis of ascidian larvae.
Animals↗
Purification and characterization of prolyl endopeptidase from the Pacific herring, Clupea pallasi, and its role in the activation of sperm motility.
Protease activities with specificity toward synthetic substrates, Suc-Gly-Pro-Leu-Gly-Pro-MCA for prolyl endopeptidase or collagenase-like peptidase, and Suc-Ala-Ala-Pro-Phe-MCA for chymotrypsin were identified in the detergent-soluble fraction of herring spermatozoa. The enzyme activities increased in the presence of herring sperm-activating protein (HSAP). Among them a prolyl endopeptidase [EC. 3. 4. 21. 26] was purified to near homogeneity from herring testis. The molecular mass of the enzyme was 79 kDa and the properties of the enzyme were quite similar to prolyl endopeptidase from other tissues or cells. Both the enzyme activation and the sperm motility activation by HSAP were inhibited by benzyloxycarbonyl-L-thioproline-thioprolinal, a specific inhibitor for prolyl endopeptidase. Furthermore, the motility activation by HSAP was inhibited by substrates of the prolyl endopeptidase. Western blotting with mouse anti-prolyl endopeptidase serum revealed the presence of 79 kDa prolyl endopeptidase in the tail fraction of herring sperm. These results suggest that prolyl endopeptidase exists on the surface of the sperm tail and interacts with the HSAP.
Amino Acid Sequence↗
Sperm-activating proteins obtained from the herring eggs are homologous to trypsin inhibitors and synthesized in follicle cells.
The activation of sperm motility by the egg is an ubiquitous phenomenon in the animal kingdom, but the molecules by which the egg activates sperm motility have been clarified in only a few invertebrate species. In the Pacific herring, Clupea pallasii, mature unfertilized eggs release the sperm-activating proteins which are prerequisite to successful fertilization. Complementary DNA clones encoding herring sperm-activating proteins were isolated from a herring ovarian complementary DNA library and amino acid sequences were deduced. The herring sperm-activating protein(s) is a secretory product(s) with a strong homology to Kazal-type trypsin inhibitors, such as mammalian acrosin inhibitors. The sperm-activating proteins were globally distributed in the outermost layer of the egg chorion and its gene was found to be expressed in the follicle cells which surround developing oocytes. These results suggest that in the Pacific herring, trypsin inhibitor-like proteins are synthesized in the follicle cells, secreted, accumulated in the egg chorion during oocyte development, and released into the milieu at spawning to activate the motility of spermatozoa at the time of gamete interaction.
Amino Acid Sequence↗
Role of two series of Ca2+ oscillations in activation of ascidian eggs.
Changes in [Ca2+]i are an essential factor regulating egg activation. Matured ascidian eggs are arrested at metaphase I, and two series of [Ca2+]i transients have been observed after fertilization: Ca2+ waves just after fertilization (Series I) and [Ca2+]i oscillation between the first and second polar body extrusion (Series II). We investigated mechanisms involved in the elevation of [Ca2+]i and the role of the [Ca2+]i transients during egg activation in Ciona savignyi. The monoclonal antibody 18A10 against IP3 receptor type 1, which inhibits IP3-induced Ca2+ release in hamster and mouse eggs, did not show substantial inhibitory effects on series I or egg deformation, whereas Series II and the first cell division were inhibited by the antibody. Ruthenium red, an inhibitor of ryanodine receptor-mediated Ca2+ release, had no apparent effect of [Ca2+]i transients and other events related to the egg activation. Microinjection of IP3 into unfertilized eggs induced [Ca2+]i transients similar to those seen in Series I, whereas injection of cyclic ADP ribose, an agonist of ryanodine receptors, rarely induced [Ca2+]i transient. Adenophostin B, a potent nonmetabolizable agonist of IP3 receptors, induced [Ca2+]i oscillations which continued after first polar body extrusion, without separation to two series, and led to extrusion of first and second polar bodies. These results suggest that Series II is driven by the mouse type 1-like IP3 receptor while Series I seems to be mediated by another type of IP3 receptor. Injection of IP3 only induced the first polar body extrusion and the egg was arrested at metaphase II even when a higher amount of IP3 was injected. On the other hand, reinjection of IP3 after the first polar body extrusion led to emission of the second polar body. Thus, Series I and II of [Ca2+]i transients are likely to be required for metaphase-anaphase transition in meiosis.
Adenosine↗
Proteasomes regulate the motility of salmonid fish sperm through modulation of cAMP-dependent phosphorylation of an outer arm dynein light chain.
Proteasomes are involved in ATP-dependent regulation of sperm motility in salmonid fish. We have demonstrated here by immunoelectron microscopy that proteasomes are located at the structure of the chum salmon sperm flagellum that attaches at the base of the outer arm dynein and extends toward the plasma membrane. Furthermore, substrates and inhibitors of proteasome inhibit the cAMP-dependent phosphorylation of a 22 kDa axonemal protein in chum salmon sperm. The 22 kDa phosphoprotein was solubilized by treatment of the axoneme with a high salt solution and subsequent sucrose density gradient centrifugation of the extract revealed that it cosedimented with 19 S outer arm dynein, indicating that it is a dynein light chain. These results suggest that proteasomes modulate the activity of outer arm dynein by regulating cAMP-dependent phosphorylation of the 22 kDa dynein light chain.
Animals↗
Aminopeptidase-like protease released from oocytes affects oocyte surfaces and suppresses the acrosome reaction in establishment of polyspermy block in oocytes of the mussel Mytilus edulis.
Suppression of the acrosome reaction of sperm on fertilized oocytes inhibits sperm-oocyte binding and is considered one of the three mechanisms responsible for polyspermy block in oocytes of the mussel Mytilus edulis (Togo et al., 1995). When unfertilized oocytes were inseminated in the presence of aminopeptidase inhibitors and the fertilized oocytes were inseminated again, neither the acrosome reaction nor sperm binding to fertilized oocytes were suppressed, suggesting that aminopeptidase-like protease participates in suppression of the acrosome reaction. The supernatant solution obtained after centrifuging a suspension of fertilized oocytes hydrolyzed aminopeptidase substrates, and these activities were inhibited strongly or effectively by aminopeptidase inhibitors. When unfertilized oocytes were incubated with this supernatant solution and inseminated, both the number of sperm bound and the acrosome reaction rate decreased, and these effects were reversed by conducting this treatment in the presence of aminopeptidase inhibitors. These results suggest that aminopeptidase-like protease released from the oocyte at fertilization affects the oocyte surface to suppress the acrosome reaction and consequently inhibits sperm-oocyte binding.
Acrosome↗
Degradation of vitellogenins by 170 kDa trypsin-like protease in the plasma of the tilapia, Oreochromis niloticus.
Proteolytic degradation of plasma vitellogenins during purification procedure has been noted in several teleost fishes. We have characterized here a trypsin-like serine protease in the plasma of the tilapia, Oreochromis niloticus, which degrades vitellogenins. The molecular mass of the protease was estimated as 230 kDa by gel filtration and as 170 kDa both by nondenaturing and by SDS-polyacrylamide gel electrophoresis. The protease efficiently hydrolyzed the synthetic peptide substrates for trypsin-like proteases but not the substrates for chymotrypsin-like proteases nor aminopeptidases. Hydrolysis of the peptide substrates was strongly inhibited by leupeptin, aprotinin and N-tosyl-L-lysine chloromethyl ketone and to certain extent by chymostatin, 3,4-dichloroisocoumarin, phenylmethanesulfonyl fluoride, and soybean trypsin inhibitor. Leupeptin and aprotinin also inhibited the degradation of a vitellogenin in the plasma. Although the physiological functions of the 170 kDa protease in vivo have not been elucidated, the results on exzymatic properties of this protease will be useful for the isolation and characterization of vitellogenin not only in tilapia but also in other organisms.
Amino Acid Sequence↗
Test cell migration and tunic formation during post-hatching development of the larva of the ascidian, Ciona intestinalis.
Morphological changes in the tunic layers and migration of the test cells during swimming period in the larva of the ascidian, Ciona intestinalis, were observed by light and electron microscopy. The swimming period was divided into three stages. In stage 1, further formation of juvenile tunic layer started only in the larval trunk and neck region. In stage 2, the layer became swollen in the ventral and dorsal sides of the neck region and in stage 3, the swelling expanded backward. Concomitantly with these changes, the outermost larval tunic layer (outer cuticular layer), which had been formed before hatching, also swelled in the neck region in stage 2 and formed two humps in stage 3, although the layer did not change in the tail region during the swimming period. Test cells that were present over the entire larval tunic layer in stage 1 began to move from the surface of the fin toward that of the side of the body in stage 2, and finally gathered to form six bands running radially from the anterior end to the posterior end of the trunk region and aligned along the lateral sides of body in the tail region in stage 3. In electron microscopic observations, pseudopodia protruding from the test cells invaded the larval tunic, following which they extended proximate to the juvenile tunic in the trunk region. In the tail region, which had no juvenile tunic layer as that described, the pseudopodia invaded and remained adjacent to the surface of the epidermis or the sensory cilia protruded from the epidermis. Metamorphosis of the larvae, further tunic formation, degradation of adhesive papilla, attachment of larva to the substratum and tail resorption commenced after these morphological changes occurred. The possible role of the test cells in metamorphosis is discussed.
Animals↗
Multiregional introgressions inferred from the mitochondrial DNA phylogeny of a hybridizing species complex of gobiid fishes, genus Tridentiger.
Partial sequences of the cytochrome b gene (402 bp) in mtDNA were determined for brackishwater gobiid fishes, genus Tridentiger, collected from geographically distant locations in the Japanese Archipelago, and their interspecific and geographic variations were analyzed and compared. Contrary to the results of a previous allozyme analysis which revealed the existence of considerable genetic divergence (Nei's genetic distance > 0.5) between T. obscurus and T. brevispinis, the mtDNA haplotypes (mitotypes) of these two species were very similar and could not be distinguished by any of the neighbor-joining, maximum-likelihood or parsimony analyses. Hybrid individuals between the two species were also found, with several mitotypes being shared by both species and their hybrids. The phylogenetic relationships of mitotypes were divided into three subgroups, the geographical distributions of the latter being allied to geographical features of the Archipelago. These results suggested the occurrence of multiregional introgression between the two species, with mitotypes transferring from one species to the other.
Animals↗
[Current status and problems of home parenteral nutrition for terminal cancer patients from the viewpoint of indication].
Since June 1993, we have been performing home parenteral nutrition (HPN) for end-stage cancer patients. We studied how many patients could be treated with HPN, why they were not able to be treated with HPN in the terminal stage cancer patients, 158 cases, who admitted and died in our ward from June 1993 to April 1997. Eighty-six patients (54.4%) were considered to not be indicated for HPN, due to general weakness (28 cases), dyspnea (17 cases), need for medical care other than HPN (34 cases), and inability to understand this treatment (1 case). Fifty-six patients (35.4%) were considered to have been able to be treated with HPN and could have had returned home. Fifteen patients (26.8%) were given HPN. Four cases wanted to stay in the hospital after they knew the truth and the prognosis of their disease. Another four patients became weak while preparing for HPN and could not go home. Eighty percent of patients could not to be treated with HPN because of the factors of their family. Eight patients had no family to cared for them. Twelve patients were refused to the truth of their disease. The families of 13 cases refused to accept and care for them in their home. We may conclude that co-operation among patients, their family and the medical staff is the most important factor in providing HPN for end-stage cancer patients.
Adult↗