Anti-senescence effect and molecular mechanism of the major royal jelly proteins on human embryonic lung fibroblast (HFL-I) cell line / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Royal jelly (RJ) from honeybee has been widely used as a health promotion supplement. The major royal jelly proteins (MRJPs) have been identified as the functional component of RJ. However, the question of whether MRJPs have anti-senescence activity for human cells remains. Human embryonic lung fibroblast (HFL-I) cells were cultured in media containing no MRJPs (A), MRJPs at 0.1 mg/ml (B), 0.2 mg/ml (C), or 0.3 mg/ml (D), or bovine serum albumin (BSA) at 0.2 mg/ml (E). The mean population doubling levels of cells in media B, C, D, and E were increased by 12.4%, 31.2%, 24.0%, and 10.4%, respectively, compared with that in medium A. The cells in medium C also exhibited the highest relative proliferation activity, the lowest senescence, and the longest telomeres. Moreover, MRJPs up-regulated the expression of superoxide dismutase-1 (SOD1) and down-regulated the expression of mammalian target of rapamycin (MTOR), catenin beta like-1 (CTNNB1), and tumor protein p53 (TP53). Raman spectra analysis showed that there were two unique bands related to DNA synthesis materials, amide carbonyl group vibrations and aromatic hydrogens. These results suggest that MRJPs possess anti-senescence activity for the HFL-I cell line, and provide new knowledge illustrating the molecular mechanism of MRJPs as anti-senescence factors.

IgE Binding Reactivity of Peptide Fragments of Bla g 4, a Major German Cockroach Allergen

Cockroaches have been recognized as a major cause of asthma. Bla g 4 is one of the most important German cockroach allergens. The aim of this study is to investigate IgE reactivity to the recombinant Bla g 4 (rBla g 4) in the sera of allergic patients and identify linear IgE binding epitope. For protein expression, full-length Bla g 4 (EF202172) was divided into 5 overlapping peptide fragments (E1: aa 1-100, E2: aa 34-77, E3: aa 74-117, E4: aa 114-156, and E5: aa 153-182). The full-length and 5 peptide fragments of Bla g 4 was generated by PCR and over-expressed in E. coli BL21 (DE3). The IgE binding reactivities of the full-length and peptide fragments were measured by ELISA using 32 serum samples of cockroach allergy. The sera of 8 patients (25%) reacted with rBla g 4. Four sera (100%) showed IgE-binding reactivity to full-length and peptide fragment 4, and 2 sera (50%) reacted with peptide fragment 2. One (20%) serum reacted with peptide fragment 3. The results of ELISA using overlapping recombinant fragments indicated that the epitope region was located at amino acid sequences 34-73 and 78-113, and major IgE epitope of Bla g 4 was located at amino acid sequences 118-152 of C-terminal. B-cell epitope analysis of German cockroach allergen Bla g 4 could contribute to the strategic development of more specific and potentially efficacious immunotherapy.

Intra- and interspecific variation of cuticular hydrocarbon composition in two Ectatomma species (Hymenoptera: Formicidae) based on Fourier transform infrared photoacoustic spectroscopy

We have been able to discriminate different castes and sexes of ants in the same colony by measuring cuticular hydrocarbon levels with Fourier transform infrared photoacoustic spectroscopy, compared by canonical discriminant function analysis. We have now applied this methodology to various colonies of two species of ants of the genus Ectatomma in the Brazilian Cerrado. There were clear interspecific differences in cuticular hydrocarbons of these ants, with a small intraspecific variation. The differences between colonies were greater in E. brunneum than in E. vizottoi. Genetic differences among the colonies and species were well estimated by Fourier transform infrared photoacoustic spectroscopy and statistical analyses.

Molecular cloning and expression of the C-terminus of spider flagelliform silk protein from Araneus ventricosus.

A cDNA coding for the C-terminus of spider flagelliform silk protein (AvFlag) was cloned from Araneus ventricosus. Analysis of the cDNA sequence shows that the C-terminus of AvFlag consists of 167 amino acids of a repetitive region and 87 amino acids of a C-terminal non-repetitive region. The peptide motifs found in spider flagelliform silk proteins, GPGGX and GGX,were conserved in the repetitive region of AvFlag. Phylogenetic analysis further confirmed that AvFlag belongs to the spider flagelliform silk proteins. The AvFlag cDNA was expressed as a 28 kDa polypeptide in baculovirus-infected insect cells. As a new expression approach for spider silk protein,the combination of polyhedrin and AvFlag creates a polyhedrin AvFlag fusion protein (61 kDa) that is produced as recombinant polyhedra; this provides a basis for the source of spider silk proteins for various applications.