Tung-Chou Tsai

Lactoferrin as a natural regimen for selective decontamination of the digestive tract: Recombinant porcine lactoferrin expressed in the milk of transgenic mice protects neonates from pathogenic challenge in the gastrointestinal tract

BACKGROUND Nosocomial infection with antibiotic-resistant strains is a major threat to critical care medicine. Selective decontamination of the digestive tract (SDD) is one of the strategies used to reduce ventilator-associated pneumonia and sepsis in critically ill patients. In the present study, we performed pathogenic challenges of the digestive tract in a transgenic milk-fed animal model to test whether porcine lactoferrin (pLF) is an effective SDD regimen. METHODS Transgenic mice expressing recombinant pLF in their milk at a mean+/-SD concentration of 120+/-13.6 mg/L during the lactation stage fed normal CD-1 mice pups for 4 weeks. The pups were subsequently challenged with pathogenic Escherichia coli, Staphylococcus aureus, and Candida albicans. RESULTS Compared with the control groups fed wild-type (normal) milk, the groups fed pLF-enriched milk demonstrated statistically significant improvements in weight gain; lower bacterial numbers in intestinal fluid, blood, and liver; healthier microvilli in the small intestine; and alveoli in the lungs. CONCLUSIONS Our results showed that oral administration of pLF-enriched milk to mice led to broad-spectrum antimicrobial activity in the digestive tract and protected the mucosa of the small intestine from injury, implying that pLF can be used as an effective SDD regimen.

Effects of osteoporosis and nutrition supplements on structures and nanomechanical properties of bone tissue

In this study, the bone structures, nanomechanical properties and fracture behaviors in different groups of female C57BL/6 mice (control, sham operated, ovariectomized, casein supplemented, and fermented milk supplemented) were examined by micro-computed tomography, scanning and transmission electron microscopy, and nanoindentation. The control and sham operated mice showed dense bone structures with high cortical bone mineral densities of 544 mg/cm(3) (average) and high hardness of 0.9-1.1 GPa; resistance to bone fracture was conferred by microcracking, crack deflections and ligament bridging attributed to aligned collagen fibers and densely packed hydroxyapatite crystals. Bone mineral density, hardness and fracture resistance in ovariectomized mice markedly dropped due to loose bone structure with randomly distributed collagens and hydroxyapatites. The acidic casein supplemented mice with blood acidosis exhibited poor mineral absorption and loose bone structure, whereas the neutralized casein or fermented milk supplemented mice were resistant to osteoporosis and had high bone mechanical properties.

Identification of sex-specific polymorphic sequences in the goat amelogenin gene for embryo sexing

Amelogenin (AMEL) is a conserved gene located on the sex chromosomes of mammals. It is involved in the formation of enamel, which is the hard, white material that forms the protective outer layer of each tooth. In this study, we first cloned and determined the intron sequences of the goat AMELX and AMELY genes from female and male ear tissues. The polymorphic AMEL alleles were further analyzed by PCR-based RFLP and Southern blot hybridization analyses. Results showed that intron 5 nucleotide sequences of the goat AMELY gene contains multiple deletions/insertions and shares only 48.5% identity to intron 5 of the goat AMELX gene. Based on the polymorphic AMEL intron sequences, a set of sex-specific triplex primers was designed to PCR amplify a single fragment of 264 bp from the X chromosome of female goats and 2 fragments of 264 and 206 bp from the X and Y chromosomes, respectively, of male goats. An increased sensitivity for sex determination was reached with a single blastomere at the blastula stage isolated from goat embryos. A total of 43 goat embryos were used to estimate a 100% accuracy rate of this method confirmed by chromosomal karyotyping and live births. The embryo sexing technique has been successfully applied in different strains of goats including Alpine, Saanen, Nubian, and Taiwan goats.

Application of high-frequency ultrasound for the detection of surgical anatomy in the rodent abdomen

Rats are used extensively in abdominal disease research. To monitor disease progress in vivo, high-frequency ultrasound (HFU) can be a powerful tool for obtaining high-resolution images of biological tissues. However, there is a paucity of data regarding the correlation between rat anatomy and corresponding HFU images. Twenty-four adult male Sprague-Dawley (SD) rats underwent abdominal scans using HFU (40 MHz) surgical procedures to identify abdominal organs and major vessels as well as in situ scanning to confirm the imaging results. The results were compared with those of human abdominal organs in ultrasonographic scans. The rat liver, paired kidneys, stomach, intestines, and major blood vessels were identified by HFU and the ultrasonic morphologies of the liver and kidneys showed clear differences between rats and humans. Clinically relevant anatomical structures were identified using HFU imaging of the rat abdomen, and these structures were compared with the corresponding structures in humans. Increased knowledge with regard to identifying the anatomy of rat abdominal organs by ultrasound will allow scientists to conduct more detailed intra-abdominal research in rodents.

Kefir peptides prevent high-fructose corn syrup-induced non-alcoholic fatty liver disease in a murine model by modulation of inflammation and the JAK2 signaling pathway

Objective:In recent years, people have changed their eating habits, and high-fructose-containing bubble tea has become very popular. High-fructose intake has been suggested to be a key factor that induces non-alcoholic fatty liver disease (NAFLD). Kefir, a fermented milk product composed of microbial symbionts, has demonstrated numerous biological activities, including antibacterial, antioxidant and immunostimulating effects. The present study aims to evaluate the effects of kefir peptides on high-fructose-induced hepatic steatosis and the possible molecular mechanism.Results:An animal model of 30% high-fructose-induced NAFLD in C57BL/6J mice was established. The experiment is divided into the following six groups: (1) normal: H2O drinking water; (2) mock: H2O+30% fructose; (3) KL: low-dose kefir peptides (50 mg kg−1)+30% fructose; (4) KM: medium-dose kefir peptides (100 mg kg−1)+30% fructose; (5) KH: high-dose kefir peptides (150 mg kg−1)+30% fructose; and (6) CFM: commercial fermented milk (100 mg kg−1)+30% fructose. The results show that kefir peptides improve fatty liver syndrome by decreasing body weight, serum alanine aminotransferase, triglycerides, insulin and hepatic triglycerides, cholesterol, and free fatty acids as well as the inflammatory cytokines (TNF-α, IL-6 and IL-1β) that had been elevated in fructose-induced NAFLD mice. In addition, kefir peptides markedly increased phosphorylation of AMPK to downregulate its targeted enzymes, ACC (acetyl-CoA carboxylase) and SREBP-1c (sterol regulatory element-binding protein 1), and inhibited de novo lipogenesis. Furthermore, kefir peptides activated JAK2 to stimulate STAT3 phosphorylation, which can translocate to the nucleus, and upregulated several genes, including the CPT1 (carnitine palmitoyltransferase-1) involved in fatty acid oxidation.Conclusion:Our data have demonstrated that kefir peptides can improve the symptoms of NAFLD, including body weight, energy intake, inflammatory reaction and the formation of fatty liver by activating JAK2 signal transduction through the JAK2/STAT3 and JAK2/AMPK pathways in the high-fructose-induced fatty liver animal model. Therefore, kefir peptides may have the potential for clinical application for the prevention or treatment of clinical metabolic syndrome.

Nanomechanics Insights into the Performance of Healthy and Osteoporotic Bones

In situ nanoscopic observations of healthy and osteoporotic bone nanopillars under compression were performed. The structural-mechanical property relationship at the atomic scale suggests that cortical bone performance is correlated to the feature, arrangement, movement, distortion, and fracture of hydroxyapatite nanocrystals. Healthy bone comprising tightly bound mineral nanocrystals shows high structural stability with nanoscopic lattice distortions and dislocation activities. On the other hand, osteoporotic bone exhibits brittleness owing to the movements of dispersed minerals in and intergranular fracture along a weak organic matrix.

Oral immunotherapy with the ingestion of house dust mite extract in a murine model of allergic asthma

BackgroundAllergen-specific immunotherapy (ASIT) has the potential to modify allergic diseases, and it is also considered a potential therapy for allergic asthma. House dust mite (HDM) allergens, a common source of airborne allergen in human diseases, have been developed as an immunotherapy for patients with allergic asthma via the subcutaneous and sublingual routes. Oral immunotherapy with repeated allergen ingestion is emerging as another potential modality of ASIT. The aim of this study was to evaluate the therapeutic efficacy of the oral ingestion of HDM extracts in a murine model of allergic asthma.MethodsBABL/c mice were sensitized twice by intraperitoneal injection of HDM extracts and Al(OH)3 on day 1 and day 8. Then, the mice received challenge to induce airway inflammation by intratracheal instillation of HDM extracts on days 29–31. The treatment group received immunotherapy with oral HDM extracts ingestion before the challenge. All the mice were sacrificed on day 32 for bronchoalveolar inflammatory cytokines, mediastinal lymph node T cells, lung histology, and serum HDM-specific immunoglobulins analyses.ResultsUpon HDM sensitization and following challenge, a robust Th2 cell response and eosinophilic airway inflammation were observed in mice of the positive control group. The mice treated with HDM extracts ingestion had decreased eosinophilic airway inflammation, suppressed HDM-specific Th2 cell responses in the mediastinal lymph nodes, and attenuated serum HDM-specific IgE levels.ConclusionsOral immunotherapy with HDM extracts ingestion was demonstrated to have a partial therapeutic effect in the murine model of allergic asthma. This study may serve as the basis for the further development of oral immunotherapy with HDM extracts in allergic asthma.

Dimorphic Goat Amelogenin Alleles on Sex Chromosomes for Gender Determination of Preimplantation Embryos

 Abstract—Amelogenin (AMEL) is a conserved gene located on the sex chromosomes of mammals. It is involved in the formation of enamel, which is the hard, white material that forms the protective outer layer of each tooth. In this study, we first cloned the coding sequences of the goat AMELX and AMELY transcripts from female and male lamb enamel tissues during tooth development. All 207 amino acids of the putative AMEL proteins from the cDNA coding regions of both sex chromosomes were the same, but the AMEL sequences of the 3’-untranslated regions (UTR) were different. The results showed that the nucleotide sequences of intron 5 of the goat AMELX and AMELY genes contained multiple deletions/insertions and had only 48.5% identity. Based on the dimorphic AMEL intron sequences, a set of sex-specific primers was successfully applied to goat gender determination. A high sensitivity for sex determination was reached with a minimal amount of template, such as a trace amount of genomic DNA from a single blastomere isolated from embryos at the blastula stage among different strains of goats.

Abstract 2380: Curcumin reduces pulmonary tumorigenesis in vascular endothelial growth factor (VEGF)-overexpressing transgenic mice

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Transgenic animal techniques that provide animal models of human cancers have been widely used in biomedical research. Pulmonary cancers are among the most common cancers in the world. Angiogenesis is an important factor in the formation of tumors, tumor growth, invasion, and metastases. The vascular endothelial growth factor promotes the initial formation of blood vessels (vasculogenesis) and plays a vital role in the growth and expansion of existing blood vessels (angiogenesis). The study is to provide a method for manufacturing an animal model for researching a pulmonary tumor and serve as an animal model for researching the regulatory mechanism of the pulmonary adenocarcinoma. We investigated an lung-specific hVEGF-A165 overexpressed transgenic mice model which constructed mccsp-Vegf-A165-sv40 poly(A) transgene. The results showed that hVegf-A165 mRNA can be expressed specifically in the lung tissue of the transgenic mice. In the histopathologic slides of the lung tissue, it seemed that hVEGF-A165 overexpressed transgenic mice can induce bronchial epithelium flattened, abnormalproliferation of cells on bronchial epithelium, inflammation, fibrosis, adenoma, and cyst in the transgenic mice. In pathological section, we referred specimen to different levels of lung lesion, which showed a positive correlation with the expression levels of hVEGF by IHC, PCR, and western blot. Moreover, we performed cDNA microarray to exam gene expressions on lung samples of transgenic mice. The relative gene clusters of cell proliferation, cell cycle, cell metastasis, carcinogenesis, and angiogenesis were chosen to evaluate by RT-PCR, Q-PCR, and Western blots. Curcumin is a major active polyphenol compound of turmeric and exhibits remarkable effects on cancers such as brain, colon, bladder, breast, prostate and cervical cancers. In this study, we found that curcumin significantly eliminated hVEGF-A165 overexpression to normal, specifically in clara cells of the lungs of transgenic mice, and suppressed the formation of tumors. In addition, we demonstrated a relationship between curcumin treatment and the expression of VEGF, EGFR, ERK2, and Cyclin A at the transcriptional and translational levels. We also noticed a reduction of Cyclin A and Cyclin B treatment with curcumin that was an effect on the cell cycle. Curcumin-induced inhibition of Cyclin A and Cyclin B likely results in decreased progression through the S and G2/M phases. These results demonstrated that the expression of proteins involved in the S to M phase transition in transgenic mice is suppressed by curcumin. This result suggests that a blockade of the cell cycle may be a critical mechanism for the observed effects on vasculogenesis and angiogenesis following treatment with curcumin. This study additionally provides an animal model for analyzing the mechanisms of the regulation and proliferation of pulmonary adenocarcinoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2380. doi:10.1158/1538-7445.AM2011-2380