Control of flatfish sperm motility by CO2 and carbonic anhydrase

Sperm motility in flatfishes shows unique characteristics. The flagellar movement either in vivo or in permeabilized models is arrested by the presence of 25-100 mM HCO3-, or by gentle perfusion with CO2 gas. To understand the molecular basis of this property, sperm Triton-soluble proteins and flagellar proteins from several species were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. An abundant 29-kDa protein was observed only in flatfish species. Partial amino acid sequences identified this protein as a carbonic anhydrase, an enzyme involved in the interconversion of CO2 and HCO3-. 6-ethoxyzolamide, a specific inhibitor of carbonic anhydrase inhibits sperm motility, especially at low pH. In the case of HCO3(-)-arrested sperm, the motility is restored by addition of 6-ethoxyzolamide. Taken together, these results suggest that a novel pH/HCO3(-)-dependent regulatory mechanism mediated by carbonic anhydrase is involved in the motility control in flatfish sperm.     

Dendritic cell immunoactivating receptor,a novel C-type lectin immunoreceptor,acts as an activating receptor through association with Fc receptor gamma chain

An increasing number of C-type lectin receptors are being discovered on dendritic cells, but their signaling abilities and underlying mechanisms require further definition. Among these, dendritic cell immunoreceptor (DCIR) induces negative signals through an inhibitory immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic tail. Here we identify a novel C-type lectin receptor, dendritic cell immunoactivating receptor (DCAR), whose extracellular lectin domain is highly homologous to that of DCIR. DCAR is expressed similarly in tissues to DCIR, but its short cytoplasmic portion lacks signaling motifs like ITIM. However, a positively charged arginine residue is present in the transmembrane region of the DCAR, which may explain its association with Fc receptor gamma chain and its stable expression on the cell surface. Furthermore, cross-linking of DCAR in the presence of gamma chain activates calcium mobilization and tyrosine phosphorylation of cellular proteins. These signals are mediated by the immunoreceptor tyrosine-based activating motif (ITAM) of the gamma chain. Thus, DCAR is closely related to DCIR, but it introduces activating signals into antigen-presenting cells through its physical and functional association with ITAM-bearing gamma chain. The identification of this activating immunoreceptor provides an example of signaling via a dendritic cell-expressed C-type lectin receptor.     

Effect of dietary short-chain fructooligosaccharides on the cecal microflora in gastrectomized rats

Total gastric resection is known to lead to changes in the microflora in the whole gastrointestinal tract. Dietary short-chain fructooligosaccharides (Sc-FOS) have been shown to also induce a change in the microflora in the large bowel by promoting an increase in the numbers of Bifidobacterium and Lactobacillus which have beneficial effects on the host. In the present study, 4-week-old male Sprague-Dawley rats received total gastrectomy or laparotomy, and each of these surgically treated groups was randomly divided into two experimental diet groups and given a 7.5% Sc-FOS diet or control diet. Enumeration and identification of the cecal bacteria was performed by using selective and non-selective media. In the gastrectomized rats, the total bacterial count, and the counts of Bacteroidaceae and Enterobacteriaceae were higher than those in the sham-operated rats. Sc-FOS promoted an increase in the numbers of Bifidobacterium and Lactobacillus, In the rats fed on the Sc-FOS diet, the predominant type of bacteria was Lactobacillus and in the rats fed on the control diet, it was Bacteroidaceae irrespective of gastrectomy. We confirmed that both gastrectomy and dietary Sc-FOS changed the composition of cecal microflora in the rats. Dietary Sc-FOS in the gastrectomized rats increased the proportions of Lactobacillus relative to other types of bacteria to levels similar to those seen in healthy normal rats, and decreased the proportion of Bacteroidaceae.     

Life cycle inventory analysis of co<Subscript>2</Subscript> emissions manufacturing commodity plastics in japan

The goal of this study was to calculate the average CO₂ emissions for manufacturing three commodity plastics, polyethylene (PE), polypropylene (PP), and polyvinyl chloride (PVC) in Japan. The CO₂ emissions were calculated from cradle to gate, excluding the calcination processes after use. As the results, the followings were observed: 1) The gross CO₂ emissions for the manufacture of plastics in Japan were 1.3, 1.4, and 1.7 kg-CO₂/kg-PE, PP, and PVC, respectively. These mainly reflected the difference of CO₂ emissions for the in-house electricity generation. 2) The CO₂ emissions for the electricity used for manufacturing PVC were higher than that used for PE and PP, because additional electricity was required for the electrolysis to produce chlorine. The gross electricity consumption for manufacturing PVC was 1.3 kWh/kg-PVC, and the other plastics consumed 0.5 kWh/kg-Products. In addition, the effects of energy saving were studied using a projected gas-diffusion electrode for the electrolysis of salt on the reduction of CO₂ emissions. It was estimated that the reduction in CO₂ emissions was 7% compared with the present PVC manufacturing processes.     

EST analysis of gene expression in testis of the ascidian Ciona intestinalis

To explore the gene expression underlying spermatogenesis, a large-scale analysis has been done on the cDNAs from testis of the ascidian, Ciona intestinalis. A set of 5,461 expressed sequence tags was analyzed and grouped into 2,806 independent clusters. Approximately 30% of the clusters showed significant sequence matches to the proteins reported in DDBJ/GenBank/EMBL database including a set of proteins closely related to the gene regulation during spermatogenesis, functional and morphological changes of spermatogenic cells during spermiogenesis, and physiological functions of sperm, as well as those with housekeeping functions commonly expressed in other cells. Some clones show similarities to the proteins present in vertebrate lymphocytes, suggesting a primitive immune system in ascidians. We have also found some genes that are known to participate in hormonal regulation of spermatogenesis in vertebrates. The large majority of the genes expressed in Ciona testis show no significant matches to known proteins and the further analysis of these genes may shed new light on the molecular mechanism of spermatogenesis and sperm functions.     

Dephosphorylation of Tctex2-related dynein light chain by type 2A protein phosphatase

Maspin is a 42kDa tumor suppressor protein that belongs to the serine protease inhibitor (serpin) family. It inhibits cell motility and invasion in vitro, and tumor growth and metastasis in nude mice; however, maspin's molecular mechanism of action has remained elusive. Maspin contains several tyrosine residues and we hypothesized that phosphorylation of maspin could play a role in its biological function. Our study reveals that maspin is phosphorylated on tyrosine moiety(ies) in normal mammary epithelial cells endogenously expressing maspin. In addition, transfection of the maspin gene, using either a stable or inducible system into maspin-deficient breast cancer cell lines, yields a protein product that is phosphorylated on tyrosine residue(s). Furthermore, recombinant maspin protein can be tyrosine-phosphorylated by the kinase domain from the epidermal growth factor receptor in vitro. These novel observations suggest that maspin, which deviates from the classical serpin, may be an important signal transduction molecule in its phosphorylated form.     

Evaluation of a mass screening program for lysinuric protein intolerance in the northern part of Japan

Lysinuric protein intolerance (LPI:MIM 222700) is an autosomal recessive disease characterized by defective transport of the dibasic amino acids. We recently reported a local cluster of LPI in the northern part of Japan (Koizumi et al., 2000). Mutational analysis of the LPI patients in this local cluster revealed they were exclusively homozygous for the R410X mutation. The effectiveness of early intervention with citrulline therapy (200 mg/kg per day) and protein restriction (1.5 g/kg per day) was confirmed in these patients. Mass screening was conducted in 4,568 newborn babies between 1999 and 2002, which was estimated to cover 100% of almost all newborns delivered in the screened area. Forty heterozygous newborns were found (0.88%), leading to an estimated incidence of LPI of 1:51,984. The number of people that required screening to detect one case was 51,984, and the cost for mass screening was 30 cents/person (a total of dollars 15,600). This is comparable to, or even less than, the cost of currently screened diseases in Japan. Therefore, we conclude that a mass screening program for LPI can be introduced effectively and economically into an area where an LPI cluster is located as the result of a founder mutation.     

Synthesis of chemotactic peptide analogs with a photoaffinity cross-linker as new probes for analysis of receptor-ligand interaction

Formylpeptide receptors are well-characterized receptors which participate in host defense responses of neutrophils. We designed and synthesized chemotactic peptide analog with p-benzoylphenylalanine (Bpa) and biotin to probe structural and mechanistic aspects of peptide-receptor interaction. These peptides possess biological activities which were dependent upon spacer residue length of and Bpa position. The covalent photoaffinity label was detected by Streptavidine-blot, which was inhibited by the parent peptide.     

Macrolide antibiotics inhibit prostaglandin E2 synthesis and mRNA expression of prostaglandin synthetic enzymes in human leukocytes

We investigated the action of macrolide antibiotics, which are considered to have anti-inflammatory activity, on lipopolysaccharide (LPS)-stimulated prostaglandin (PG) E2 synthesis and the expression of mRNAs for cytosolic phospholipase A2 (cPLA2), cyclooxygenase (COX)-1, and COX-2 in human leukocytes. The production of LPS-stimulated PGE2 was significantly increased in peripheral polymorphonuclear leukocytes (PMNLs) and in mononuclear leukocytes (MNLs). Amounts of mRNAs for COX-2 and cPLA2, but not for COX-1, were enhanced by LPS in PMNLs and MNLs. The LPS-enhanced PGE2 synthesis and the expression of cPLA2 and COX-2 mRNAs were inhibited by clarithromycin, azithromycin and dexamethasone in PMNLs and MNLs. The mRNA expression of COX-1 in PMNLs was decreased by clarithromycin and azithromycin. Macrolide antibiotics inhibited PGE2 synthesis in human leukocytes by suppressing cPLA2, COX-1, and COX-2 mRNA expression. These data indicate one mechanism of macrolide anti-inflammatory activity.