Many transgenic domestic animals have been developed to produce therapeutic proteins in the mammary gland, and this approach is one of the most important methods for agricultural and biomedical applications. However, expression and secretion of a protein varies because transgenes are integrated at random sites in the genome. In addition, distal enhancers are very important for transcriptional gene regulation and tissue-specific gene expression. Development of a vector system regulated accurately in the genome is needed to improve production of therapeutic proteins. The objective of this study was to develop a knock-in system for expression of human fibroblast growth factor 2 (FGF2) in the bovine β-casein gene locus. The F2A sequence was fused to the human FGF2 gene and inserted into exon 3 of the β-casein gene. We detected expression of human FGF2 mRNA in the HC11 mouse mammary epithelial cells by RT-PCR and human FGF2 protein in the culture media using western blot analysis when the knock-in vector was introduced. We transfected the knock-in vector into bovine ear fibroblasts and produced knock-in fibroblasts using the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system. Moreover, the CRISPR/Cas9 system was more efficient than conventional methods. In addition, we produced knock-in blastocysts by somatic cell nuclear transfer using the knock-in fibroblasts. Our knock-in fibroblasts may help to create cloned embryos for development of transgenic dairy cattle expressing human FGF2 protein in the mammary gland via the expression system of the bovine β-casein gene.

We previously demonstrated that the chronic consumption of a high-fat diet (HFD) promotes lung and liver metastases of 4T1 mammary carcinoma cells in obesity-resistant BALB/c mice. To examine early transcriptional responses to tumour progression in the liver and lungs of HFD-fed mice, 4-week-old female BALB/c mice were divided into four groups: sham-injected, control diet (CD)-fed; sham-injected, HFD-fed (SH); 4T1 cell-injected, CD-fed (TC); 4T1 cell-injected, HFD-fed (TH). Following 16 weeks of either a CD or HFD, 4T1 cells were injected into the mammary fat pads of mice in the TC and TH groups and all mice were continuously fed identical diets. At 14 d post-injection, RNA was isolated from hepatic and pulmonary tissues for microarray analysis of mRNA expression. Functional annotation and core network analyses were conducted for the TH/SH Unique gene set. Inflammation in hepatic tissues and cell mitosis in pulmonary tissues were the most significant biological functions in the TH/SH Unique gene set. The biological core networks of the hepatic TH/SH Unique gene set were characterised as those genes involved in the activation of acute inflammatory responses (Orm1, Lbp, Hp and Cfb), disordered lipid metabolism and deregulated cell cycle progression. Networks of the pulmonary Unique gene set displayed the deregulation of cell cycle progression (Cdc20, Cdk1 and Bub1b). These HFD-influenced alterations may have led to favourable conditions for the formation of both pro-inflammatory and pro-mitotic microenvironments in the target organs that promote immune cell infiltration and differentiation, as well as the infiltration and proliferation of metastatic tumour cells.

A metal-supported solid oxide fuel cell (SOFC) using Ce0.8Sm0.2O2 (Sm-doped ceria, SDC) buffer layer and La0.9Sr0.1Ga0.8Mg0.2O3 (LSGM) electrolyte films showed a small degradation in the cell performance after a long-term operation because of La migration from the electrolyte to the buffer layer, resulted in a formation of a less conductive phase. Thus, various ceramic materials such as doped ceria and perovskite-related oxides were investigated for an effective buffer layer with respect to fabricating reliable metal-supported SOFCs using a LSGM electrolyte film. In particular, La-doped CeO2 (LDC) and Pr-doped LaCrO3 (LPCr) were investigated as buffer layer material since the materials showed chemical compatibility with the LSGM and anode materials. The cell using a LDC buffer layer showed a prior stability during the operation for 100 h at 973 K, while the power density of the cell was slightly low owing to the low electrical conductivity of LDC compared with that of SDC or LPCr. In contrast, the cell using a LPCr buffer layer revealed significantly low open circuit voltage (OCV) and power density, which were attributed to Pr decomposition in the LPCr caused by the reactivity with water vapor. However, the metal-supported cell with a multilayer electrolyte film including LSGM/LPCr/SDC layers showed an almost theoretical OCV and reasonably high power density with no degradation after a long-term operation for 100 h at 973 K, suggesting that the LPCr layer effectively prevented La migration and the SDC layer led to avoid the Pr decomposition. Thus, a LPCr is an effective buffer layer material for reliable metal-supported SOFCs using a LSGM electrolyte thin film.

Oxidative stress may be affected by lead exposure as well as antioxidants, yet little is known about the interaction between dietary antioxidants and blood lead levels (BLL) on oxidative stress level. We investigated the interaction between dietary antioxidants and BLL on oxidative stress level. As part of the Biomarker Monitoring for Environmental Health conducted in Seoul and Incheon, Korea, between April and December 2005, we analysed data from 683 adults (female = 47·4 %, mean age 51·4 (sd 8·4) years) who had complete measures on BLL, dietary intakes and oxidative stress marker (urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG)). Dietary intakes were assessed by a validated semi-quantitative FFQ, BLL was measured using atomic absorption spectrophotometry, and 8-OHdG by ELISA. Multivariate linear regression analyses were used to evaluate the influence of BLL on the association between dietary antioxidants and 8-OHdG. Geometric means of BLL and 8-OHdG concentrations were 4·1 (sd 1·5) μg/dl and 5·4 (sd 1·9) μg/g creatinine, respectively. Increases of vitamins C and E were significantly associated with the decrease of log10 8-OHdG in the adults from the lowest quartile of the BLL group ( ≤ 3·18 μg/dl, geometric mean = 2·36 μg/dl) than those of the highest quartile BLL group (>5·36 μg/dl, geometric mean = 6·78 μg/dl). Regarding antioxidant-related foods, vegetables excluding kimchi showed a higher inverse relationship with 8-OHdG in the lowest quartile BLL group than the highest group. These findings suggest a rationale for lowering the BLL and increasing the intake of dietary antioxidants in the urban population in Korea.

Electroneuronography (ENoG) has become a useful test for estimating the degree of facial nerve degeneration and predicting the prognosis in patients with facial nerve palsy. Test results may be influenced by several factors, including the electrode positions, skin resistance, stimulus magnitude, and possible artifacts. Regarding recording electrode positions, different groups have used two different locations, the nasolabial fold and nasal ala. The authors compared the waveforms recorded from these two locations in ENoG recordings to obtain the optimal waveform. Twenty healthy volunteers and 25 patients with unilateral facial nerve palsy were included in this study. Recordings were carried out with the recording electrode placed on the nasolabial fold, followed by placement on the nasal ala after 10 minutes. The following parameters were assessed: (1) the supramaximal threshold, (2) amplitude and shape of the waveform, (3) interside difference, and (4) test-retest variability. There was no significant difference in the amplitude of the waveform, interside difference, and test-retest variability between the two groups. However, when the electrode was placed on the nasal ala, the threshold was significantly lower, an ideal biphasic configuration was present in almost all cases (97.5 per cent) of normal volunteers and it was easier to identify the waveform. Placement of the recording electrode on the nasal ala would be the preferred method.