A Fully-Human Antibody Specifically Targeting a Membrane-Bound Fragment of CADM1 Potentiates the T Cell-Mediated Death of Human Small-Cell Lung Cancer Cells

Small-cell lung cancer (SCLC) is the most aggressive form of lung cancer and the leading cause of global cancer-related mortality. Despite the earlier identification of membrane-proximal cleavage of cell adhesion molecule 1 (CADM1) in cancers, the role of the membrane-bound fragment of CAMD1 (MF-CADM1) is yet to be clearly identified. In this study, we first isolated MF-CADM1-specific fully human single-chain variable fragments (scFvs) from the human synthetic scFv antibody library using the phage display technology. Following the selected scFv conversion to human immunoglobulin G1 (IgG1) scFv-Fc antibodies (K103.1–4), multiple characterization studies, including antibody cross-species reactivity, purity, production yield, and binding affinity, were verified. Finally, via intensive in vitro efficacy and toxicity evaluation studies, we identified K103.3 as a lead antibody that potently promotes the death of human SCLC cell lines, including NCI-H69, NCI-H146, and NCI-H187, by activated Jurkat T cells without severe endothelial toxicity. Taken together, these findings suggest that antibody-based targeting of MF-CADM1 may be an effective strategy to potentiate T cell-mediated SCLC death, and MF-CADM1 may be a novel potential therapeutic target in SCLC for antibody therapy.     

Digestibility and physicochemical properties of granular sweet potato starch as affected by annealing

The effects of annealing on the digestibility, morphology, and physicochemical characteristics of four types of granular sweet potato starches [Yulmi (YM), Yeonwhangmi (YHM), sweet potato starch from Samyang Genex (SSPS), and commercial sweet potato starch (CSPS)] were investigated. Annealing was performed at 55°C and 90% moisture content for 72 h. Morphology, the branched chain distribution of amylopectin, and the X-ray diffraction pattern remained unchanged during the annealing process. The slowly digestible starch content in annealed YM, YHM, and SSPS starches increased, but did not change in annealed CSPS. The gelatinization temperatures increased, but the gelatinization temperature range decreased with annealing. The swelling factor and amylose leaching decreased, while the close packing concentration increased. Rapid Visco Analyser analysis revealed that annealed starches possessed thermal stability and higher pasting temperatures. It is suggested that the enhanced packing arrangement formed during annealing impacts the digestibility and physicochemical properties of sweet potato starches.     

Changes in S-allyl cysteine contents and physicochemical properties of black garlic during heat treatment

This study aimed to investigate the effects of temperature on the black garlic manufacturing process. The moisture content, pH, browning intensity, S-allyl cysteine (SAC) content and antioxidant activity, including DPPH radical scavenging activity and reducing power, were determined. The moisture content of garlic gradually decreased throughout the heating process. The rate of moisture removal was higher at high temperatures compared with low temperatures. The pH also decreased more significantly in garlic heated at high temperatures. The browning intensity increased with increasing temperature. The SAC contents of black garlic were significantly different according to heating temperature; the garlic samples heated at a low temperature had a higher SAC contents. Antioxidant activity, as determined by the DPPH radical scavenging activity and reducing power, increased when the garlic was exposed to higher temperatures.     

Growth characteristics and development of a predictive model for Bacillus cereus in fresh wet noodles with added ethanol and thiamine

Response surface methodology was used to determine growth characteristics and to develop a predictive model to describe specific growth rates of Bacillus cereus in wet noodles containing a combination of ethanol (0 to 2% [vol/wt]) and vitamin B(1) (0 to 2 g/liter). B. cereus F4810/72, which produces an emetic toxin, was used in this study. The noodles containing B. cereus were incubated at 10°C. The growth curves were fitted to the modified Gompertz equation using nonlinear regression, and the growth rate values from the curves were used to establish the predictive model using a response surface methodology quadratic polynomial equation as a function of concentrations of ethanol and vitamin B(1). The model was shown to fit the data very well (r(2) = 0.9505 to 0.9991) and could be used to accurately predict growth rates. The quadratic polynomial model was validated, and the predicted growth rate values were in good agreement with the experimental values. The polynomial model was found to be an appropriate secondary model for growth rate (GR) and lag time (LT) based on the correlation of determination (r(2) = 0.9899 for GR, 0.9782 for LT), bias factor (B(f) = 1.006 for GR, 0.992 for LT), and accuracy factor (A(f) = 1.024 for GR, 1.011 for LT). Thus, this model holds great promise for use in predicting the growth of B. cereus in fresh wet noodles using only the bacterial concentration, an important contribution to the manufacturing of safe products.     

PCR identification of ruminant tissue in raw and heat-treated meat meals

To control the spread of bovine spongiform encephalopathy in cattle through contaminated animal feedstuffs, screening of feed products is essential. We designed five pairs of primers to identify specifically raw and heat-treated tissue from cattle, sheep, goat, deer, and ruminants in general. A forward common primer was designed based on a conserved DNA sequence in the mitochondrial 12S rRNA-tRNA(val)-16S rRNA gene, and reverse primers were designed to hybridize with a species-specific DNA sequence for each species considered. All primers were developed to create a specific PCR product small enough (less than 200 bp) to be suitable for heat-treated material. To evaluate the effect of heat treatment, a severe sterilization condition (133 degrees C at 300 kPa for 20 min) was chosen. Species-specific amplicons were obtained from all types of heat-treated meat meals. Analysis of laboratory-contaminated vegetable meals revealed that the detection limit of the assay was 0.05% for each species analyzed. This PCR-based analysis can be used as a routine method for detecting banned animal-derived ingredients in raw and heat-treated feedstuffs.     

Modified Fluoroquinolones as Antimicrobial Compounds Targeting Chlamydia trachomatis

Chlamydia trachomatis causes the most common sexually transmitted bacterial infection and trachoma, an eye infection. Untreated infections can lead to sequelae, such as infertility and ectopic pregnancy in women and blindness. We previously enhanced the antichlamydial activity of the fluoroquinolone ciprofloxacin by grafting a metal chelating moiety onto it. In the present study, we pursued this pharmacomodulation and obtained nanomolar active molecules (EC₅₀) against this pathogen. This gain in activity prompted us to evaluate the antibacterial activity of this family of molecules against other pathogenic bacteria, such as Neisseria gonorrhoeae and bacteria from the ESKAPE group. The results show that the novel molecules have selectively improved activity against C. trachomatis and demonstrate how the antichlamydial effect of fluoroquinolones can be enhanced.     

Single-Step Fast Tissue Clearing of Thick Mouse Brain Tissue for Multi-Dimensional High-Resolution Imaging

Recent advances in optical clearing techniques have dramatically improved deep tissue imaging by reducing the obscuring effects of light scattering and absorption. However, these optical clearing methods require specialized equipment or a lengthy undertaking with complex protocols that can lead to sample volume changes and distortion. In addition, the imaging of cleared tissues has limitations, such as fluorescence bleaching, harmful and foul-smelling solutions, and the difficulty of handling samples in high-viscosity refractive index (RI) matching solutions. To address the various limitations of thick tissue imaging, we developed an Aqueous high refractive Index matching and tissue Clearing solution for Imaging (termed AICI) with a one-step tissue clearing protocol that was easily made at a reasonable price in our own laboratory without any equipment. AICI can rapidly clear a 1 mm thick brain slice within 90 min with simultaneous RI matching, low viscosity, and a high refractive index (RI = 1.466), allowing the imaging of the sample without additional processing. We compared AICI with commercially available RI matching solutions, including optical clear agents (OCAs), for tissue clearing. The viscosity of AICI is closer to that of water compared with other RI matching solutions, and there was a less than 2.3% expansion in the tissue linear morphology during 24 h exposure to AICI. Moreover, AICI remained fluid over 30 days of air exposure, and the EGFP fluorescence signal was only reduced to ~65% after 10 days. AICI showed a limited clearing of brain tissue >3 mm thick. However, fine neuronal structures, such as dendritic spines and axonal boutons, could still be imaged in thick brain slices treated with AICI. Therefore, AICI is useful not only for the three-dimensional (3D) high-resolution identification of neuronal structures, but also for the examination of multiple structural imaging by neuronal distribution, projection, and gene expression in deep brain tissue. AICI is applicable beyond the imaging of fluorescent antibodies and dyes, and can clear a variety of tissue types, making it broadly useful to researchers for optical imaging applications.     

Characterization of the Secretome of a Specific Cell Expressing Mutant Methionyl-tRNA Synthetase in Co-Culture Using Click Chemistry

Co-culture system, in which two or more distinct cell types are cultured together, is advantageous in that it can mimic the environment of the in vivo niche of the cells. In this study, we presented a strategy to analyze the secretome of a specific cell type under the co-culture condition in serum-supplemented media. For the cell-specific secretome analysis, we expressed the mouse mutant methionyl-tRNA synthetase for the incorporation of the non-canonical amino acid, azidonorleucine into the newly synthesized proteins in cells of which the secretome is targeted. The azidonorleucine-tagged secretome could be enriched, based on click chemistry, and distinguished from any other contaminating proteins, either from the cell culture media or the other cells co-cultured with the cells of interest. In order to have more reliable true-positive identifications of cell-specific secretory bodies, we established criteria to exclude any identified human peptide matched to bovine proteins. As a result, we identified a maximum of 719 secreted proteins in the secretome analysis under this co-culture condition. Last, we applied this platform to profile the secretome of mesenchymal stem cells and predicted its therapeutic potential on osteoarthritis based on secretome analysis.     

L－Fuzzy集上（G）Fuzzy积分特性

给出ＴＬ－ｆｕｚｚｙ集上（Ｇ）ｆｕｚｚｙ积分的定义．讨论了这类广义ｆｕｚｚｙ积分的基本特性，得到了积分转化定理及可积性充要条件，最后指出（Ｇ）ｆｕｚｚｙ积分的两个收敛定理．