Advanced Materials Research Vols. 175-176

Abstract: Acetylcholinesterase (AChE), which contains two subfamilies, ace1 and ace2 in insects, was identified to be the target of organophosphorous and carbamate insecticides. To research the sequences and tissues expressions of two aces, full length cDNAs encoding two ace genes were cloned, designated as Bmm-ace1 and Bmm-ace2 from larvae of the Bombyx mandarina. The amino acid sequence of Bmm-ace1 shared 99.71 % homology with its homolog, Bm-ace1, in silkworm, Bombyx mori, with two mutations (G664S and S307P), and the amino acid sequence of Bmm-ace2 shared 99.37 % homology with Bm-ace2, in B. mori , with four mutations (M18I, N233S, I310V and G621S). Tissue expression analysis showed that ace1 gene expressed only in the brains and fat bodies of B. mandarina, while ace2 genes expressed in all the tissues tested. ace1 and ace2 expressed highly in brains and fat bodies. The present results are significant to the study of resistance evolution of Lepidorptera as well as the understanding of the mechanism of pesticide resistance of insects.

Abstract: In order to explore the roles of Bombyx mori glutathione S-transferase gene (BmGST) in detoxification and resistance to insecticides, we used real-time PCR method to detect the transcription levels of five BmGST genes in different tissues of the 5th instar larvae feeding on sodium fluoride treated mulberry leaves. The detection results were normalized by using 3 internal reference genes. The results indicated that the transcription levels of BmGSTs were different in various tissues. Transcription of BmGST genes could be induced by NaF, The normalized data with the above 3 internal reference genes indicated that, it is very important to choose adequate internal reference genes so as to ensure the reliability of the detection results.

Abstract: The present study was undertaken to clarify the change of induction of CYP4M5 and CYP4M9 expression level by rutin. In this study, we used dual-spike-in qPCR to examine expression profiles of the silkworm Bombyx mori CYP4M5 and CYP4M9 genes in the larval midgut and fatbody after exposure to rutin. In organization-course study, rutin at middle concentration (5×10-2ng/µL) caused significant upregulation of CYP4M5 and CYP4M9 genes at early time point (2h) in fatbody, higher concentration (5×10-1ng/µL) did secondly, while lower concentration (5×10-3ng/µL) caused little change. In the midgut of silkworm, rutin in all concentrations didn't affect the expression of the two genes. These findings showed that induction of CYP4M5 and CYP4M9 expression level by rutin are concentration depended and tissue-specific.Collectively, CYP4M5 and CYP4M9 genes may have some relationships with metabolism of rutin in silkworm.

Abstract: The Bombyx mori serves as model organism among the Lepidoptera insects. In the post-genomic era, in order to study gene function, the profiling of mRNA transcription has become a popular research field. Real-time quantitative RT-PCR (qRT-PCR) has become established as the sensitive method for detecting the expression level of low abundance mRNA, and it usually chooses one or several reference genes to standardize the expression level of target gene. Since the changes in amplification of reference gene can reflect the changes of RNA production, quality or cDNA synthesis efficiency. So choosing an appropriate reference gene can reduce the differences between tested samples. Based on the comparison of Standardization of three frequently-used reference genes (GAPDH, Actin-3, 28srRNA), and decide which is the best way to study gene expression level in silkworm, Bombyx mori.

Abstract: Mulberry (Molus alba L. and other plants of the genus Morus) has been cultivated in many Asian countries such as China, Korea, Japan and Thailand, and their leaves are used to feed silkworms (Bombyx mori L.). Additionally, the leaves are rich in alkaloid components including 1-deoxynojirimycin (1-DNJ) which is known as one of the most potent glucosidase inhibitors. Dietary mulberry 1-DNJ has been hypothesized to be beneficial for suppression of an abnormally high blood glucose level. In this study, we investigated the high 1-DNJ concentration cultivars from among over 500 various cultivars growing in the field at Experimental Farm of Shinshu University, to utilize the mulberry biomass in addition to feed silkworms. We extracted 1-DNJ from thirty-five varieties of mulberry leaves, and we determined the amount of 1-DNJ. As a result, some cultivars were found to contain a high DNJ concentration compared with Ichinose which is a standard cultivar.

Abstract: The transparent water-insoluble silk fibroin(SF) films were casted from the mixture solution of silk fibroin and xylitol/mannitol. The structure, surface morphology, solubility, mechanical properties and light transmittance of the blend films were measured. FT-IR, X-ray diffraction retuslts indicated that the films were mainly composed of Silk I structure. SEM showed the blend films with xylitol were miscible, whereas the blend films with mannitol had phase-separated structure. There were lots of nanopores in the blend films in the wet state. The insoluble SF /xylitol films had excellent mechanical properties while the SF / mannitol films were brittle. The mechanical property of SF/alcohol blend films were consistent with the human cornea in wet state.When the contents of xylitol were 10% and 20%, the blend films had high light transmittance which were similar to human cornea. In summary, the SF /xylitol film containing 10% xylitol provides a great potential to act as repairing materials for cornea.

Abstract: An attempt to change the structure of silk fibers and their properties for the biological application was studied by utilizing gamma radiation in various Co60 intensities (0 kGy, 30 kGy, 50 kGy, 100 kGy, 200 kGy, 500 kGy, 1000 kGy, 2000 kGy, 3000 kGy). With the increase of the gamma radiation intensity, SEM result shows that cracks and fragments were formed between microfibrils of the irradiated fiber significantly. Simultaneously SDS-PAGE results give the evidence that the molecular weight of the fibroin diminished. Furthermore, the breaking strength and elongation of irradiated fibers decreased gradually with the increasing Co60 intensity. Although no significant changes of the molecular conformations were found by FTIR and X-ray diffraction, the effects on molecular interactions of the silk fibroin, such as peptide bonding, hydrogen bond and intermolecular bonding force, were obviously observed and enhanced gradually with the increase of gamma radiation intensity.

Abstract: Electrospining technique is one of the hotest topics all over the world. The main form of the electrospun products is generally nonwoven fiber web with the poor mechanical properties. Consecutive PA6/MWNTs nanofiber filaments were successfully fabricated by an improved electrospining method. Peregal O was used as the bath to post-draw the as-spun filaments and the effects of draw ratio on their structures and properties were studied. The results show that with the increase of the draw ratio, the diameters of filament and fiber decrease; while the degree of orientation arrangement and crystallinity of the fibers is obviously improved and the breaking strength and initial modulus of the filaments increase. As the maximum draw ratio reaches 1.7, the breaking strength and initial modulus of the filament are 2.64 times and 4.2 times as compared to those of the control sample respectively.

Abstract: Silkworm silk has been recognized as a satisfactory biomaterial for long time due to its exceptional biocompatibility, biodegradability, mechanical properties etc. For example, silk fibers in the form of sutures have been used for centuries. The aim of this study is to discuss the potential usage of silk as the novel biomedical devices, such as blood vessels. In this study, cuit silks prepared from degummed raw silks were twisted as threads with four different yarn linear densities. A specific braiding machine was used to weave those threads into a tube. Subsequently two different groups of silk tubes were prepared. One was treated by ethanol and the other without. Thickness, porosity, mass per unit area of two groups of braided tubes were measured. Its mechanical properties were also studied. The influence of ethanol treatment and various yarn linear densities on its structural and mechanical properties was also studied. Results indicated that structural and mechanical properties of the tubes were significatly changed by the yarn linear densities and ethanol treatment. Conclusively, braided silk tube could be a potential blood vessel tissue engineering scaffold.

Abstract: To improve the physicochemical properties of calcium phosphate cement (CPC), silk fibroin (SF) in the different forms were added into CPC. The structure of the composites was studied by X-ray diffraction. The setting time was investigated by ISO Cement Standard Consistency Instrument. Scanning Electron Microscope was used to observe the surface morphology. Mechanical properties of samples were tested by Instron Universal Testing Machine. The results indicated that acicular crystal of hydroxyapatite (HA) was formed in the hardening body of both CPC with SF and the pure CPC. Addition of SF had no significant effect on the structure of SF/CPC composite. The setting time of CPC with SF was significantly shorter than that of the pure CPC (30.3 mins). The setting time of CPC by adding silk fibroin powder I (SFP) and silk fibroin fiber (SFF) was greatly shortened, which was only 11.7 minutes. The setting time of CPC with SFP decreased approximately by 1/3, while the setting time of the CPC with SFF decreased nearly by 1/2. With the adding of SF, the compressive strength of CPC increased significantly. There was a distinct increase in the work-of-compressive of CPC with the adding of SFF.