We separated an aqueous extract of Morus alba stem bark by gel filtration chromatography, ammonium sulfate fractionation, and chromatography on Superdex 200 10/300GL and Mono Q 5/50GL columns. The resultant water-soluble fraction significantly increased total lymphocyte proliferation up to 3-fold (at 10μg/ml) and stimulated the proliferation of isolated B cells (at 5, 10, and 20μg/ml), similar to the positive control (lipopolysaccharide). This fraction, with an apparent high molecular weight of ≥ 200k, consisted of 55.3% proteins and 9.6% sugars. The bioactive fraction treated with trypsin and pre-incubated either alone or with 2-mercaptoethanol and glycopeptitase F significantly reduced lymphocyte proliferation. These results suggest that a water-soluble glycoprotein-like complex of M. alba stem bark has therapeutic potential to boost immunity.

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Raw silk of different sizes (10, 27, 42, and 100d) was used to produce 200d of twisted yarn with twist numbers of 100, 200, 500, and 1,000T/m, and the untwist percentage, bending rigidity, and hysteresis of non-degummed and degummed yarn were examined. The results show that the physical properties of the produced silk yarn are highly dependent on the twist number and that the structure of twisted yarn is highly dependent on the raw silk size. Furthermore, yarn produced from thick raw silk has superior bending characteristics.

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In an insect cell line BmN4 derived from Bombyx mori, foreign genes were efficiently integrated in chromosomal DNA by the piggyBac transposition system composed of the donor plasmid harboring the piggyBac element and the helper plasmid expressing the transposase gene. An analysis of transformed cells, however, revealed that the presence of mRNA and activity of the transposase for more than 50 days after the co-transfection of the plasmids, suggesting the re-mobilization of the integrated piggyBac element could be caused by a continuous production of active transposase. To shorten the period of transposase production, four transposase mRNA constructs with different untranslated region (UTR) sequences were synthesized in vitro and used as alternatives for the helper plasmid. One mRNA construct, whose 5’ and 3’ UTR sequences except for the polyA tail were similar to those of the mature transcript expressed from the helper plasmid, was effective in the transposition of the piggyBac element from the co-transfected donor plasmid into the genome and was comparable to the helper plasmid in the efficiency of its transformation. In addition, the transfected mRNA disappeared earlier than the transcript from the helper plasmid and the transposase activity became undetectable after around 21 days of post-transfection during the antibiotic selection. The results clearly indicated that, as a source of transposase for the piggyBac-mediated transformation of BmN4 cells, and possibly other cultured cells and living organisms as well, the in vitro synthesized mRNA with the appropriate 5’ and 3’ UTR sequences is superior in the stability of the integrated piggyBac element to the conventional helper plasmid.

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Two types of silk fibroin-based scaffolds, with or without hydroxyapatite, were examined in a context for guided bone regeneration with potential application for dental surgery. Scaffolds were implanted to artificial bone defects on rabbit femur, which provides similar structure as alveolar bone and healing procedures were observed histologically. Both of the scaffolds were effective to prevent the infiltration of the other types of tissue such as fibrous connective tissue or cartilage, which happens in some cases without implantations and causes incomplete regeneration with subsided surface. In vivo degradation aspect of the scaffolds was significantly different from each other. The degradation was accelerated in the presence of hydroxyapatite, resulting in fragmentation of residual scaffold after eight weeks, where the scaffolds without hydroxyapatite remained uninterrupted. This difference suggests a possibility for tailor-made preparation of scaffolds with different biodegradability as well as controlled release of bioactive substrates.

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We have characterized a novel gene, CLEM 20, from the flesh fly Sarcophaga peregrina, which shows significant homology to the C-type lectin family. CLEM 20 mRNA was highly transcribed from the second day after eclosion only in the C-type lectin-producing tissue (CLPT) located at the tip of mouthparts. In the CLPT, CLEM 20 mRNA tended to be detected in several cells neighboring the muscle. CLEM 20 protein localized to a specific site in the cytoplasm of several cells in the CLPT. We had already reported that CLEM 36, another C-type lectin of this fly, was expressed in the CLPT and secreted into saliva (Yamamoto-Kihara and Kotani, 2004). CLEM 36 mRNA tended to be detected in several CLPT cells neighboring the opening site of the food canal, and CLEM 36 protein was synthesized abundantly in the cytoplasm and gathered in a specific part of the cytoplasm in the CLPT. Localization of CLEM 20 mRNA and protein in the CLPT seemed to be different from that of CLEM 36 mRNA and protein. Our results indicate that CLEM 20 is a distinct C-type lectin from CLEM 36 and that the localizable difference in the CLPT between CLEM 20 and 36 protein may reflect differences in the secretory process into saliva, such as the secretory rate or the amount of the produced protein.

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