Association of the sea urchin EGF-related peptide, EGIP-D, with fasciclin I-related ECM proteins from the sea urchin Anthocidaris crassispina.

Exogastrula-inducing peptides (EGIP) of the sea urchin Anthocidaris crassispina are endogenous peptides related to epidermal growth factor (EGF), which induce exogastrulation in the embryo. Recently, a protein(s) from sea urchin embryos that binds to one of the EGIP, EGIP-D (EGIP-D-binding protein, EBP) was purified. The isolation and characterization of the cDNA clones for two EBP proteins (EBP-alpha and EBP-beta) is reported. The two EBP proteins were highly similar in structure to each other; both possessed putative cell-binding sites and two repeated sequences characteristically seen in the insect neuronal cell adhesion protein, fasciclin I. The EBP showed similarity with other sea urchin proteins HLC-32, Bep1, and Bep4. It has been confirmed that bacterially expressed EBP proteins associate with EGIP-D as does native EBP, suggesting the interaction between EGF-related proteins and fasciclin I-related proteins. An EBP transcript of 1.4 kb was strongly expressed in immature ovaries but not in immature testes. A somewhat lower level of the transcript existed in unfertilized eggs and the amount gradually declined to an almost undetectable level by the pluteus stage. The EBP proteins were present throughout embryonic development at nearly constant levels. Although most of the proteins were distributed rather evenly in the cytoplasm, a small portion was detected on the apical surface of blastomeres and ectodermal cells, showing that EBP are components of the hyaline layer.

Expression of a src-type protein tyrosine kinase gene, AcSrc1, in the sea urchin embryo.

By screening a cDNA library and 3'-rapid amplification of cDNA ends, the cDNA for a non-receptor type protein tyrosine kinase from the sea urchin Anthocidaris crassispina was analyzed. The deduced protein (AcSrc1) with the highest identity of about 60% to mammalian Src family kinases shows the characteristic features of the Src family. AcSrc1 mRNA is maternally expressed in unfertilized eggs, while zygotic expression is first detected in blastulae and continues through the pluteus stage. Zygotic mRNA expression, visualized by in situ hybridization, is detected specifically in archenteron at the gastrula stage, while it is restricted in plutei to the midgut and hindgut, suggesting specific roles for AcSrcl in the formation and/or functions of the digestive tract. Meanwhile, western blot analysis has shown that the AcSrc1 protein is constantly expressed throughout embryogenesis. By immunostaining, it was found that the protein (distributed evenly in the cytoplasm of unfertilized eggs) is translocated to the membrane after fertilization. All through the following development, AcSrcl was localized to the peripheries of different embryonic cells, although at a relatively low level of localization at the boundaries between adjacent cells.

[Corneal curvature of myopia].

We classified myopia by total refractive error, measured the corneal curvature, and analyzed the relationship between the diopter value and corneal curvature in growing teen-agers and in adults in their twenties, whose myopic progress is presumed to have stopped. We adopted as the harmonic average the central 3.0 mm of the cornea. We also examined peripheral corneal curvature in upper side, lower side, temporal side and nasal side at about 4.5 mm and 6.5 mm diameter from the center. There was very little difference in the harmonic average of corneal curvature from -1D to -6D. Similarly, there was very little difference in the peripheral corneal curvature. The corneal curvature was larger from central to periphery in all directions. The upper side was larger than lower side and the nasal side was larger than the temporal side. We concluded that the refractive power did not depend on the corneal refractive power in mild to moderate myopic patients.

Efficacy of ivermectin against Strongyloides stercoralis in humans.

Okinawa Prefecture is an endemic area of Strongyloides stercoralis infection. Since treatment of this infection remains unsatisfactory, we evaluated the efficacy of ivermectin. Twenty-three patients were treated with a single oral dose of ivermectin (mean +/- SD, 105.5 +/- 20.8 mcg/kg of body weight), followed by a second dose two weeks later. The rate of cure was 85.7% at 2 weeks after the first treatment, and 90.5% at 2 weeks after the second treatment. Side effects occurred in 2 patients (8.7%), but they were mild and transient. The results indicate that ivermectin might be useful and relatively safe for the therapy of Strongyloides stercoralis infection as an alternative to thiabendazole or mebendazole.

[Preventive effect of Clostridium butyricum M588 against the proliferation of Clostridium difficile during antimicrobial therapy].

Fecal flora of the patients without remarkable gastrointestinal diseases were studied. All patients were treated with antimicrobial drugs with or without Miya-BM (the preparation of Clostridium butyricum M588). The stools were examined before and after (during) antimicrobial treatment. Fecal flora of 69 patients before medication was almost the same with that of healthy adults as has been reported by Mitsuoka et al. After giving antimicrobials, most bacterial genus except Enterococcus and Yeasts in the stool decreased their detection rate and their population. This floral change was not much influenced by co-administration of Miya-BM. The detection rate of Clostridium difficile and/or the toxin A from the stool were markedly increased after giving antimicrobials. When Miya-BM was co-administered with antimicrobials, however, the detection of C. difficile and/or toxin A was very rare. C. butyricum M588 was recovered from 7 cases out of 10 patients treated with antimicrobials and Miya-BM. Non-spore form of C. butyricum was dominant in the feces of 3 cases, and spore form was dominant in the other 4. This result showed that administered C. butyricum M588 germinated in intestinal tract.

[Inhibition of enteropathogens by Clostridium butyricum MIYAIRI 588].

The inhibitory effect of Clostridium butyricum MIYAIRI 588 against various enteropathogens was investigated in mixed cultures. It was observed that C. butyricum M588 inhibited the growth of Vibrio cholerae O1, V. cholerae non-O1, Aeromonas hydrophila, and Shigella flexneri. Considering that the interaction between C. butyricum and Shigella is especially important because of their proliferation site in the lower intestine, further examinations were carried out on Shigella in particular. Results were as follows: 1) In BHI broth culture of Shigella, the pH of culture fluid went down to 5.2, but the growth of Shigella was not inhibited. 2) In the mixed culture of Shigella and C. butyricum, the growth of Shigella was inhibited, nevertheless the pH of the culture fluid was 5.6. 3) In the mixed culture with phosphate buffered BHI maintaining the pH higher than 6.0, the growth of Shigella was inhibited. 4) In case of pure culture of C. butyricum in BHI broth, the pH of culture fluid indicated 5.5, and Shigella failed to grow in the cell free culture supernatant. 5) The growth of Shigella was not inhibited in the culture supernatant when the pH was adjusted at 7.2. These results suggested that the inhibition of Shigella in the mixed culture with C. butyricum was not due to a single factor such as pH or fatty acid etc. but due to multifactors including live cells of C. butyricum.

Effects of short chain fatty acids on the production of heat-labile enterotoxin from enterotoxigenic Escherichia coli.

Each addition of some short chain fatty acids (SCFAs) into casamino acids-yeast extract culture media at a concentration of 2 mg/ml reduced the production of heat-labile enterotoxin (LT) from enterotoxigenic Escherichia coli in proportion to the elongation of carbon chain from C-2 to C-7. The LT-production was inversely recovered by the addition of longer chain fatty acids. The reduction of LT-production by SCFAs seems to depend on the disturbance of the biosynthesis of LT itself, since LT was not detected in the cells treated with n-heptyric acid at 2 mg/ml, which abolished the LT-production.