Betulinic acid inhibits high-fat diet-induced obesity and improves energy balance by activating AMPK.

BACKGROUND AND AIM: Metabolic syndromes are prevalent worldwide and result in various complications including obesity, cardiovascular disease and type II diabetes. Betulinic acid (BA) is a naturally occurring triterpenoid that has anti-inflammatory properties. We hypothesized that treatment with BA may result in decreased body weight gain, adiposity and hepatic steatosis in a diet-induced mouse model of obesity. METHODS AND RESULTS: Mice fed a high-fat diet and treated with BA showed less weight gain and tissue adiposity without any change in calorie intake. Gene expression profiling of mouse tissues and cell lines revealed that BA treatment increased expression of lipid oxidative genes and decreased that of lipogenesis-related genes. This modulation was mediated by increased AMP-activated protein kinase (AMPK) phosphorylation, which facilitates energy expenditure, lipid oxidation and thermogenic capacity and exerts protective effects against obesity and nonalcoholic fatty liver disease. Overall, BA markedly inhibited the development of obesity and nonalcoholic fatty liver disease in mice fed a high-fat diet, and AMPK activation in various tissues and enhanced thermogenesis are two possible mechanisms underlying the antiobesity and antisteatogenic effects of BA. CONCLUSIONS: The current findings suggest that treatment with BA is a potential dietary strategy for preventing obesity and nonalcoholic fatty liver disease.

Genome-wide expression analysis of a rice mutant line under salt stress.

Salinity is a major environmental stress to plants. In this study, the ability of plants to tolerate salt was investigated by studying growth, physiological characteristics, and expression levels of genes related to the salt-stress response in the salt-tolerant rice mutant (Till-II-877), which was derived from γ-ray irradiation. Compared to plants grown under normal conditions, the height and root length of wild type (WT) were reduced by approximately 40 and 29% following exposure to salt stress for 3 weeks, whereas Till-II-877 line showed 29 and 23% reductions in plant height and root length, respectively. No significant changes were observed in total chlorophyll content, and the malondialdehyde content of the mutant increased less than that of the WT under salt treatment. Gene expression was compared between the WT and mutant lines using microarray analysis. An unbiased analysis of the gene expression datasets allowed us to identify the pathways involved in salt-stress responses. Among the most significantly affected pathways, changes in gene expression were observed in α-linolenic acid and linoleic acid metabolism (in lipid metabolism), fructose and mannose metabolism and glycolysis-gluconeogenesis (in carbohydrate metabolism), cysteine and methionine metabolism (in amino acid metabolism), and carbon fixation (in the energy metabolism of photosynthetic organisms) under salt stress. These results show that the differential response of plants subjected to salt stress was due to changes in multiple metabolic pathways. These findings increase our understanding of the effects of salt stress in rice and may aid in the development of salt-tolerant rice cultivars.

Comparative genomic hybridization analysis of rice dwarf mutants induced by gamma irradiation.

Comparative genomic hybridization (CGH) is a powerful tool used to analyze changes in copy number, polymorphisms, and structural variations in the genome. Gene copy number variation (CNV) is a common form of natural diversity in the genome, which can create new genes and alter gene structure. Thus, CNVs may influence phenotypic variation and gene expression. In this study, to detect CNVs, we irradiated rice seeds with gamma rays (300 Gy) and selected two dwarf mutagenized plants, GA-III-189 and -1052, in the M3 generation. These plants were subjected to CGH analysis using Agilent's RICE CGH array. Most of the CNVs identified were less than 10 kb in length. We detected 90 amplified and 18 deleted regions in GA-III-189, and 99 amplified and 11 deleted regions in GA-III-1052. Of note, CNVs were located on chromosome 12 in both GA-III-189 and -1052, which contained 39 commonly amplified regions in 29 genes. The commonly amplified genes included six genes encoding F-box domain-containing proteins. Alterations in these F-box domain-containing genes were confirmed by quantitative RT-PCR. Integration of CGH and gene expression data identified copy number aberrations and novel genes potentially involved in the dwarf phenotype. These CGH and gene expression data may be useful for uncovering the mechanisms underlying the dwarf phenotype.

Factors related to number of present teeth in Korean elderly adults aged 55-84 years.

OBJECTIVES: The aim of this study was to determine the associations between the number of present teeth (NT) and socio-economic, demographic and oral health behavioural factors among Korean adults aged 55-84 years. METHODS: The total subjects comprised 3767 individuals who were examined and who answered the questions on socio-economic status and oral health behaviour from the fourth Korean National Health and Nutrition Examination Survey conducted from 2007 to 2009. The dependent variable was NT, with binary status divided by the median. Socio-economic and demographic factors included gender, educational level, parent's educational levels, region of residence, household income, type of health insurance and mother's economic activity. Oral health behaviours were as follows: daily toothbrushing frequency, smoking status, recent dental visit and illegal dental treatment. Multivariate logistic regression models were applied to explain the associations between NT and other variables. RESULTS: In a model adjusted by socio-economic, demographic and oral health behavioural variables, subjects who lived in urban areas were more likely to have larger NT compared to those in suburban areas (OR: 1.22, P = 0.025). Males were more likely to have larger NT (OR: 1.90, P < 0.001), and daily toothbrushing frequency was associated with NT (OR = 1.25, P = 0.023). Non-smokers (OR: 2.44, P < 0.001) and past smokers (OR: 1.70, P < 0.001) were more likely to have lager NT compared to current smokers. Subjects without illegal dental treatments were more likely to have lager NT compared to those with illegal dental treatments (OR = 2.21, P < 0.001). CONCLUSIONS: Interventions aiming to preserve present teeth in elderly adults should consider socio-economic, demographic and oral health behavioural factors.

Proteomic characterization of Kunitz trypsin inhibitor variants, Tia and Tib, in soybean [Glycine max (L.) Merrill].

The soybean Kunitz trypsin inhibitor (KTi) has several polymorphic variants. Of these, Tia and Tib, which differ by nine amino acids, are the two main types. In this study, differences in KTi proteome between Tia and Tib were investigated using three soybean cultivars and three mutant lines. Two cultivars, Baekwoon (BW) and Paldal (PD), and one mutant line, SW115-24, were Tia type, whereas one soybean cultivar, Suwon115 (SW115), and two mutant lines, BW-7-2 and PD-5-10, were Tib type. Protein from the six soybean lines was extracted and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), non-denaturing polyacrylamide gel electrophoresis (non-denaturing PAGE), and two-dimensional polyacrylamide gel electrophoresis (2-DE). By SDS-PAGE, there was no difference between soybean cultivars and mutant lines, except for SW115-24. Western blot analysis revealed that, in comparison with Tia, Tib type accumulated relatively low amounts of KTi. By non-denaturing PAGE, the three soybean lines of Tib type were characterized by slower mobility than the three soybean lines of Tia type. Zymography detected eight distinct zones of trypsin inhibitory activity among which Tia and Tib lacked the fifth and sixth zone, respectively. By two-dimensional native polyacrylamide gel electrophoresis (2-DN), the spots related to trypsin inhibitory activity showed different mobilities, whereas only one KTi (21.5 kDa) spot was resolved by 2-DE. By two-dimensional zymography (2-DZ), Tib showed a broader activity zone (pI 4-7) in comparison with Tia (pI 4-5). The results indicate that the genotypes with a different type of KTi present different proteomic profiles and trypsin inhibitory activities.

Overexpression of stathmin1 in the diffuse type of gastric cancer and its roles in proliferation and migration of gastric cancer cells.

BACKGROUND: Stathmin1 is a microtubule-regulating protein that has an important role in the assembly and disassembly of the mitotic spindle. The roles of stathmin1 in carcinogenesis of various cancers, including prostate and breast cancer, have been explored. However, its expression and roles in gastric cancer have not yet been described. METHODS: Stathmin1 expression in paraffin-embedded tissue sections from 226 patients was analysed by immunohistochemistry. Roles of stathmin1 were studied using a specific small interfering RNA (siRNA). RESULTS: The expression of stathmin1 was positively correlated with lymph node metastasis, TNM stages and vascular invasion, and negatively with recurrence-free survival, in the diffuse type of gastric cancer. The median recurrence-free survival in patients with a negative and positive expression of stathmin1 was 17.0 and 7.0 months, respectively (P=0.009). When the expression of stathmin1 was knocked down using siRNA, the proliferation, migration and invasion of poorly differentiated gastric cancer cells in vitro were significantly inhibited. Moreover, stathmin1 siRNA transfection significantly slowed the growth of xenografts in nude mice. CONCLUSION: These results suggest that stathmin1 can be a good prognostic factor for recurrence-free survival rate and is a therapeutic target in diffuse-type gastric cancer.

Identification of Kunitz trypsin inhibitor mutations using SNAP markers in soybean mutant lines.

The Kunitz trypsin inhibitor (KTi) in soybean has several polymorphic types that are controlled by multiple alleles, which behave in a co-dominant fashion. Of these, Tia and Tib, which differ by nine amino acids, are the predominant types. In order to develop a single nucleotide amplified polymorphism (SNAP) marker for the classification of the predominant KTi types, Tia and Tib, and evaluate KTi activities by differing KTi type total 451 soybean mutant lines (M(12)-M(16) generation) were incorporated in this study. Among 451 soybean mutants, 144 and 13 mutant lines showed decreased and increased trypsin inhibitor activity when compared with the original cultivars, respectively. To identify the KTi type, we designed a SNAP marker. Among 451 mutant lines from 12 soybean cultivars and landraces, 8 mutant lines derived from cvs. Baekwoon, Paldal and Suwon115 showed a change in KTi type when compared with the original cultivars using the SNAP marker. Five mutant lines in Suwon115 changed from Tib to Tia, while two mutant lines derived from cv. Baekwoon and one mutant line derived from cv. Paldal were changed from Tia to Tib. These changes of KTi types were confirmed by sequencing of the KTi genes and non-denaturing polyacrylamide gel electrophoresis of the KTi proteins. To identify the effect of KTi activity based on the change in KTi type, we measured the KTi activity using the three cultivars and eight mutant lines that showed changes in KTi type. Two mutant lines (BW-1 and 7-2) derived from cv. Baekwoon and one mutant line (PD-5-10) from cv. Paldal that changed from Tia to Tib showed lower activity than the original cultivar. In cv. Suwon115, five mutant lines that changed from Tib to Tia showed higher activity than the original cultivar. These results indicate that the designed SNAP marker was capable of identifying the KTi type and that Tia activity was higher than Tib activity in soybean.

Fracture in long bones stabilised by telescopic intramedullary rods in patients with osteogenesis imperfecta.

We investigated the fracture-free survival of long bones stabilised by a telescopic intramedullary rod (TIMR) in patients with osteogenesis imperfecta with respect to the remodelling status of fracture or osteotomy sites and TIMR regions, in order to identify risk factors for fracture. A total of 44 femora and 28 tibiae in 25 patients with a mean age of 5.0 years (1.9 to 10.5) at presentation were studied. There were six patients with Sillence type I, five with type III, 13 with type IV and one with type V osteogenesis imperfecta. All received bisphosphonate treatment at the same stage during the mean follow-up of 7.3 years (0.5 to 18.1). The fracture-free survival was estimated at 6.2 years (95% confidence interval 5.1 to 7.3) by Kaplan-Meier analysis. More than half the fracture or osteotomy sites remained in a less-remodelled state at the latest follow-up or time of fracture. Of the 33 fractures, 29 (87.9%) occurred in long bones containing a less-remodelled site, and these fractures were located at this site. The relative fracture risk at the rod tip was significantly greater than in any other TIMR region (p < 0.001), and this was higher in bone segments having a less-remodelled site. This study shows a persistent fracture risk in TIMR-stabilised long bones, especially at less-remodelled fracture or osteotomy sites and at the rod tip.

Skeletal phenotype of growing transgenic mice that express a function-perturbing form of beta1 integrin in osteoblasts.

Skeletal modeling entails the deposition of large amounts of extracellular matrix (ECM) to form structures tailored to withstand increasing mechanical loads during rapid growth. Specific ECM molecules bind to integrin receptors on the cell surface, thereby triggering a cascade of signaling events that affect critical cell functions. To evaluate the role of integrins during skeletal growth, transgenic mice were engineered to express a function-perturbing fragment of beta1 integrin consisting of the transmembrane domain and cytoplasmic tail under the control of the osteocalcin promoter (TG mice). Thus, transgene expression was targeted to mature cells of the osteoblast lineage, and herein we show that cultured cells resembling osteocytes from 90-day-old TG mice display impaired adhesion to collagen I, a ligand for beta1 integrin. To determine the influence of beta1 integrin on bones that are responsible for providing structural support during periods of rapid growth, we examined the phenotype of the appendicular skeleton in TG mice compared to wild type (WT) mice. According to radiographs, bones from mice of both genotypes between 14 and 90 days of age appeared similar in gross structure and density, although proximal tibiae from 35-90 days old TG mice were less curved than those of WT mice (72-92% TG/WT). Although there were only mild and transient differences in absolute bone mass and strength, once normalized to body mass, the tibial dry mass (79.1% TG/WT females), ash mass (78.5% TG/WT females), and femoral strength in torsion (71.6% TG/WT females) were reduced in TG mice compared to WT mice at 90 days of age. Similar effects of genotype on bone mass and curvature were observed in 1-year-old retired breeders, indicating that these phenotypic differences between TG and WT mice were stable well into adulthood. Effects of genotype on histomorphometric indices of cancellous bone turnover were minimal and evident only transiently during growth, but when present they demonstrated differences in osteoblast rather than osteoclast parameters. Together, these results suggest that integrin signals generated during growth enhance the acquisition of a skeletal mass, structure, and strength to withstand the mechanical loads generated by weight-bearing.

Measurement method for electric fields based on stark spectroscopy of argon atoms

We report the development of a method for the measurement of electric fields in glow discharge plasmas, based on Stark spectroscopy of argon atoms. The method is based on laser excitation of transitions in atomic argon. The key feature of the method is that the electric field is determined by matching experimentally obtained absorption spectra to theoretically calculated spectra. The dependence of the positions of energy levels of argon atoms on the strength of the electric field was calculated by solving the Schrodinger equation for the argon atom. Measurements of Stark spectra were made in the sheath region of a glow discharge using laser optogalvanic spectroscopy. The wavelength of the laser radiation was tuned to the transitions 4s-->nf (n=7,8,ellipsis,14) of the argon atom. For n=11, the lower limit for electric field measurements was estimated to be 14 V/mm.