• BMB Reports
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• v.38 no.6
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• pp.717-724
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• 2005

The nontoxic B subunit of cholera toxin (CTB) can significantly increase the ability of proteins to induce immunological tolerance after oral administration, when it was conjugated to various proteins. Recombinant CTB offers great potential for treatment of autoimmune disease. Here we firstly investigated the feasibility of silkworm baculovirus expression vector system for the cost-effective production of CTB under the control of a strong polyhedrin promoter. Higher expression was achieved via introducing the partial non-coding and coding sequences (ATAAAT and ATGCCGAAT) of polyhedrin to the 5' end of the native CTB gene, with the maximal accumulation being approximately 54.4 mg/L of hemolymph. The silkworm bioreactor produced this protein vaccine as the glycoslated pentameric form, which retained the GM1-ganglioside binding affinity and the native antigenicity of CTB. Further studies revealed that mixing with silkworm-derived CTB increases the tolerogenic potential of insulin. In the nonconjugated form, an insulin : CTB ratio of 100 : 1 was optimal for the prominent reduction in pancreatic islet inflammation. The data presented here demonstrate that the silkworm bioreactor is an ideal production and delivery system for an oral protein vaccine designed to develop immunological tolerance against autoimmune diabetes and CTB functions as an effective mucosal adjuvant for oral tolerance induction.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.4
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• pp.241-244
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• 2011

A lectin from the hemolymph of purse crab, Philyra pisum, was found to have anti-proliferative activity on human lung cancer cells by our laboratory. In this study, P. pisum lectin (PPL) was molecularly characterized including molecular mass, amino acid sequences, amino acid composition, and the effects of metal ions, temperature, and pH on the activity. We found that PPL showed mitogenic activity on human lymphocytes and BALB/c mouse splenocytes. The mitogenic activity (maximum stimulation index, $SI=9.57{\pm}0.59$) of PPL on human lymphocytes was higher than that of a standard well-known plant mitogen, concanavalin A (maximum $SI=8.80{\pm}0.59$). The mitogenic activity mediated by PPL is required for optimum dosing, and higher or lower concentrations caused decreases in mitogenic response. PPL also induced mitogenic activity on mouse splenocytes, however, the maximum SI ($1.77{\pm}0.09$) on mouse splenocytes of PPL was lower than that ($2.14{\pm}0.15$) of concanavalin A. In conclusion, PPL is a metal ion-dependent monomer lectin with mitogenic activity, and could be used as a lymphocyte or splenocyte stimulator.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.14 no.8
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• pp.4106-4110
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• 2013

Apolipophorin-III (apoLp-III) was isolated and purified from the last larval hemolymph of Galleria mellonella by the KBr gradient ultracentrifugation and gel chromatography (Sephadex G-100). In this paper, we examined that apoLp-III is taken up into the last larval fat bodies in Galleria mellonella. The last larval fat body tissues were incubated at room temperature for 30 min with fluorescein isothiocyanate (FITC)-labeled apoLp-III (FITC-apoLp-III). Fluorescein microscopy and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) revealed that the last larval fat body tissues internalize FITC-apoLp-III. The results show that the apoLp-III is taken up by the last larval fat body.

• Journal of Sericultural and Entomological Science
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• v.31 no.2
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• pp.72-81
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• 1989

Antheraea yamamai vitellin was purified from matured eggs by polyacryamide gel electrophoresis, also stage dependent appearance, immunological comparison and relative content of the protein were investigated. 1. Vitellogenin, the precursor of vitellin, was first detected in the larval hemolymph at the late spinning stage by polyacrylamide gel electrophoresis and immunoelectrophoresis. 2. The electrophoretic mobility of the vitellin was identical with that of Bombyx mori and of Bombyx mandarina. However, the specific antiserum against A. uamamai vitellin did not react with either that of Bombyx mori or Bombyx mandarina in immumo-diffusion test. 3. Relative content of A. yamamai vitellin to the total soluble egg protein was 46.0 percent and did not change till eight days after oviposition. But the content started to decline from ten days after oviposition and was negligible in the five or serventeen month old eggs.

• International Journal of Industrial Entomology and Biomaterials
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• v.17 no.2
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• pp.217-221
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• 2008

Dung beetle larvae at the $3^{rd}$ instar were injected with lipopolysaccaride and inducible proteins were examined within a pI level of 3-10 and a size level by proteomics, including 1-D SDS PAGE analysis and antibacterial assay. The immune infected larvae extracts provided seven protein bands in one-dimensional electrophoresis and its antibacterial activity also checked. Hemolymph protein from immune infected larvae of the dung beetle were separated by twodimensional gel electrophoresis and compared with those from native larvae. In 2-D gel electrophoresis, we detected 63 immune infected unique and 32 up-regulated proteins, and 36 proteins that were down-regulated or not present in treated gel. Ten protein spots from unique proteins and those presented as different level of abundance in infected and native larvae were specially expressed. These differentially expressed proteins were proposed to be involved in the defense mechanism against microorganism.

• Korean journal of applied entomology
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• v.34 no.1
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• pp.20-32
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• 1995

Adult diapause in insects is characterized by suppression of reproductive development. It is induced by environmental cues such as photoperiod, temperature, food availability, and other conditions Diapause-inducing environment is recognized and analyzed by the brain of the insects. The interpreted information is conveyed via endocrine system to target tissues such as ovaries, fat body, and other tissues. From this signal hierarchy of a brain-endocrine-target tissue axis, several factors are involved to express a diapause trait in a quantitative mode, even though the insects show a binomial phenotye between being in diapause or not. Recent works estimated that the number of the factors is relatively small by a series of crossing trials between high and low diapause lines. Heritability of the diapause is quite high (ca. 70%) in some species. Epistasis, sex-linkage, pleiotropism, and other nongenetic components also affect diapause inheritance. Most physiological studies have been focused on control mechanisms of the juvenile hormone (JH) synthesis in corpora allata (CA) because JH level in hemolymph of teneral adults is critical to decide a later developmental mode. Allatostatin, an antagonizer of JH synthesis, has been believed to be a potent brain message to CA for adult diapause induction.

• Journal of Aquaculture
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• v.20 no.4
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• pp.205-211
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• 2007

Stress-inducible proteins may function in part as molecular chaperones, protecting cells from damage due to various stresses and helping to maintain homeostasis. We examined the mRNA expression patterns of a 68-kDa heat shock protein (HSP68) and 78-kDa glucose-regulated protein (GRP78) in relation to physiological changes in Pacific oyster Crassostrea gigas under osmotic stress. Expression of HSP68 and GRP78 mRNA in the gill significantly increased until 48 h in a hypersaline environment (HRE) and 72 h in a hyposaline environment (HOE), and then decreased. Osmolality and the concentrations of $Na^+$, $Cl^-$, and $Ca^{2+}$ in the hemolymph of HRE oysters significantly increased until 72 h (the highest value) and then gradually decreased; in HOE oysters, these values significantly decreased until 72 h (the lowest value), and then increased. These results suggest that osmolality and $Na^+$, $Cl^-$, and $Ca^{2+}$ concentrations were stabilized by HSP68 and GRP78, and indicate that these two stress-induced proteins play an important role in regulating the metabolism and protecting the cells of the Pacific oysters exposed to salinity changes.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.466-471
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• 2005

An Escherichia coli strain with high phytase activity was screened from pig excreta. The phytase gene, appA, was amplified by PCR technique. To obtain large amounts of appA phytase, the appA gene was subcloned into the baculovirus transfer vector pVL1393 under the control of the Polyhedrin promoter. The recombinant baculovirus harboring the appA gene was obtained after co-transfection and screening. The early $5^{th}$ instar larvae of silkworm were infected with the recombinant virus. Using this system, the appA phytase was overproduced up to 7,710 U per ml hemolymph. SDS-PAGE analysis revealed the baculovirus-derived appA phytase to be approximately 47 kDa in size. The optimal temperature and pH of the expressed phytase were $60^{\circ}C$ and pH 4.5, respectively. The enzymatic activity was increased by the presence of 1 mM $Ca^{2+}$, 1 mM $Mn^{2+}$, or $0.02\%$ Triton X-100.

• Journal of Aquaculture
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• v.16 no.3
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• pp.196-201
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• 2003

Sessile organisms such as the oyster Crassostrea gigas have been given much attention as a potential biomonitoring indicator to assess the impact of toxicants on aquatic organism. In this study, we exposed cells isolated from gill of oyster (Crassostrea gigas) to hydrogen peroxide in vitro. In addition oysters were in vivo exposed to pyrene and benzo(a)pyrene at various concentrations for 2 weeks. Comet assay was used to detect DNA single strand breaks and to investigate the application of this technique as a tool for aquatic biomonitoring. Hydrogen peroxide increased DNA single strand break with increasing concentration after 30 minutes exposure in vitro. Pyrene and benzo(a)pyrene caused DNA damage only at very high concentration (100 $\mu\textrm{g}$/L or 1000 $\mu\textrm{g}$/L) at two week exposure in vivo. DNA damage was relatively higher at hemocyte than at gill. It suggested that metabolized PAHs are transferred to hemolymph from digestive gland which have a relatively high enzyme activity, and attacked the DNA of hemocyte, while gill accumulated PAHs without degrading them to their metabolites due to low enzyme activity at gill. Both in vitro and in vivo exposure experiments showed that the comet assay is an effective tool on screening whether the organism are exposed to genotoxic contaminants.

• The Korean Journal of Zoology
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• v.33 no.3
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• pp.346-353
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• 1990

Two storage proteins, storage protein-1 (SPl) and storage protein -2 (SP2) were found in hemolymph and fat body during the development of Lymantria dispar L. SP1 has a molecular weight of 440, 000 and consists of six identical subunits (MW = 72, 000). The pI value of SP1 was 6.2. SP1 shows a similar high concentration during the late larval stage in both male and female. However, SP1 represents a quite different pattern during pupal stage between male and female. SP1 gradually decreases in male but increases in female. SP1 is immunologically identical to yolk protein. Also, SP1 of L. dispar shows immunologically partial reactions with storage proteins of Hyphantria cunea and Galleria mellonella.