Gonadectomy in Mito-Ob mice revealed a sex-dimorphic relationship between prohibitin and sex steroids in adipose tissue biology and glucose homeostasis

Background

Recently, we have developed a novel transgenic mouse model by overexpressing prohibitin (PHB) in adipocytes, which developed obesity due to upregulation of mitochondrial biogenesis in adipocytes, hence named “Mito-Ob.” Interestingly, only male Mito-Ob mice developed obesity-related impaired glucose homeostasis and insulin sensitivity, whereas female Mito-Ob mice did not. The observed sex differences in metabolic dysregulation suggest a potential involvement of sex steroids. Thus, the main aim of this study is to investigate the role of sex steroids on the overall phenotype of Mito-Ob mice through gonadectomy, as well as direct effect of sex steroids on adipocytes from Mito-Ob mice in vitro.

Methods

Mito-Ob mice and wild-type CD-1 mice were gonadectomized at 12 weeks of age. Age- and sex-matched sham-operated mice were used as controls. Body weight, white adipose tissue, glucose tolerance, and insulin sensitivity were analyzed 3 months post-surgery. Differentiation of adipocytes isolated from female and male Mito-Ob mice were studied with and without sex steroids.

Results

Gonadectomy significantly reduced body weight in Mito-Ob mice compared with sham-operated mice, whereas the opposite trend was observed in wild-type mice. These changes occurred independent of food intake. A corresponding decrease in adipose tissue weight was found in gonadectomized Mito-Ob mice, but depot-specific differences were observed in male and female. Gonadectomy improved glucose tolerance in male wild-type and Mito-Ob mice, but the effect was more pronounced in wild-type mice. Gonadectomy did not alter insulin sensitivity in male Mito-Ob mice, but it was improved in male wild-type mice. In primary cell cultures, testosterone inhibited adipocyte differentiation to a lesser extent in male Mito-Ob preadipocytes compared with the wild-type mice. On the other hand, preadipocytes from female wild-type mice showed better differentiation potential than those from female Mito-Ob mice in the presence of 17β-estradiol.

Conclusions

PHB requires sex steroids for the development of obese phenotype in Mito-Ob mice, which differentially affect glucose homeostasis and insulin sensitivity in male and female. It appears that PHB plays sex- and adipose depot-specific roles and involves additional factors. In vitro studies suggested that PHB differently influenced adipocyte differentiation in the presence and absence of sex steroids. Overall, this study along with available information in the literature indicated that a multifaceted relationship exists between PHB and sex steroids, which may work in a cell/tissue type- and sex-specific manner.

     

Effects of Zinc Supplementation During In Vitro Maturation on Meiotic Maturation of Oocytes and Developmental Capacity in Yak

Zinc (Zn) is an essential trace element that is required during mammalian developmental processes. The objective of this study was to investigate the effects of Zn supplementation during in vitro maturation (IVM) on the developmental capacity of yak (Bos grunniens) oocytes. Cumulus expansion, nuclear maturation, intracellular glutathione (GSH), reactive oxygen species (ROS) levels, superoxide dismutase (SOD) activity, subsequent embryonic development, and the expression of Zn transporters (ZnTs) and Zrt and Irt-like proteins (ZiPs) were evaluated. The Zn concentrations in yak plasma and follicular fluid were 0.740?±?0.012 and 0.382?±?0.009 μg/mL, respectively. The cumulus expansion did not show significant differences in COCs after matured with or without Zn supplementation (P?>?0.05). The intracellular GSH was higher in oocytes matured with 1 or 2 mg/L Zn than in control group (0 mg/L) (P?<?0.05). However, ROS levels of oocytes matured with 1 or 2 mg/L Zn were reduced significantly compared with the control and 0.5 mg/L groups (P?<?0.05). The SOD activity was increased significantly after Zn supplementation. The cleavage rate was not significantly different after Zn supplementation (P?>?0.05). Percentages of matured oocytes that developed into the blastocyst stage after IVF were 47.9, 50.5, 60.4, and 58.9% for 0, 0.5, 1, and 2 mg/L Zn groups, respectively. Gene expression analysis revealed that the expression patterns associated with Zn were changed after Zn supplementation. In conclusion, Zn supplementation to IVM improved yak oocyte maturation and subsequent development by increasing GSH and SOD activity, decreasing ROS in oocytes.     

秦艽体细胞愈伤组织诱导及继代研究

本文报道了秦艽取叶、幼苗为外植体诱导愈伤组织试验。结果表明,取幼苗６７－Ⅴ培养基,２４－Ｄ２５ｍｇ／Ｌ,ＮＡＡ０５ｍｇ／Ｌ,诱导频率最优,可高达１００％。继代培养,以６７％培养基,ＮＡＡ０５ｍｇ／Ｌ,２４－Ｄ（１－１５）ｍｇ／Ｌ最优     

继代十七年的玉米花粉胚性细胞系的异常核和异常分裂观察

对继代17年的玉米花粉胚性细胞系核形态和细胞分裂情况进行了观察分析,结果表明:随着继代培养时间的延长,异形核细胞比率增加,异形核类型和异常分裂增多。分析认为异形核和异常分裂现象出现和增多是导致胚性细胞系分化率降低、异形分化和非整倍体细胞产生的主要原因。并提出来自玉米农家种“八趟白”的部分胚性细胞系能长期保持倍性稳定和胚胎发生并再生,与不断挑选和继代培养中不加2,4-D有关。     

Estimation of mean molecular weight of Enterobacteriaceae lipopolysaccharides using gas chromatography for determination of fatty acids

In the paper, we propose a method for estimation of the mean molecular weight of lipopolysaccharide, which is important for accuracy of endotoxin activity investigation. In our study, it was assumed that lipid A portion in Enterobacterial lipopolysaccharide is substituted by four 3-hydroxytetradecanoic acid residues. Lipopolysaccharides of S, Ra, Rc and Re chemotypes being laboratory preparations as well as purchased from Sigma were investigated. Fatty acids were determined by of gas chromatography as methyl esters according to the procedure described by Wollenweber and Rietschel. Mean molecular weight was calculated by the formula: MMW = [formula: see text]. A high agreement between the estimated and the theoretical molecular weight values was demonstrated in the case of Salmonella minnesota R595 (Re) LPS preparation. As expected, LPS heterogeneity increase together with enlargement of polysaccharide chain length which is visible in electrophoregrams also. Except for LPS mean molecular weight estimation, the method allows its detection in various preparations and samples, distinguishing of R and S LPS forms as well as the determination of mean length of O-specific chain in lipopolysaccharides which structures are known.     

Application of denaturing high-performance liquid chromatography for rice variety identification and seed purity assessment

Recent DNA sequencing projects and the establishment of high-throughput assays have provided an abundance of sequence information and data on nucleotide polymorphisms in rice. Based on previously identified single-nucleotide polymorphisms (SNPs) and insertions/deletions, we employed denaturing high-performance liquid chromatography (DHPLC) to genotype rice varieties using a chromatographic pattern-based strategy. In this study, 12 amplicons harboring multiple and informative SNPs were screened. PCR products of the 12 amplicons from 47 rice varieties were analyzed by DHPLC and DNA sequencing. Each homozygous sample with a single peak pattern in the initial DHPLC analysis was mixed with zhenshan97 for a second DHPLC analysis. The 12 amplicons were found to be polymorphic across the hybrids, and mixed homozygous samples with 43 distinct DHPLC elution profiles detected. Sequence analysis confirmed that the distinct DHPLC patterns corresponded to different DNA sequences. A set of distinct characteristic profiles in six amplicons differentiated between all of the hybrids, inbred lines, and restorer lines and produced unique a fingerprint for these lines. In addition, we found that the DHPLC pattern of the hybrid was in accordance with the results obtained by DHPLC analysis of a mixed sample of the two parents. These results demonstrate that DHPLC can be efficiently applied for the rapid and automated identification of diverse rice varieties and could possibly be utilized for seed genetic purity testing on a high-throughput scale.     

p62 links the autophagy pathway and the ubiqutin–proteasome system upon ubiquitinated protein degradation

The ubiquitin–proteasome system (UPS) and autophagy are two distinct and interacting proteolytic systems. They play critical roles in cell survival under normal conditions and during stress. An increasing body of evidence indicates that ubiquitinated cargoes are important markers of degradation. p62, a classical receptor of autophagy, is a multifunctional protein located throughout the cell and involved in many signal transduction pathways, including the Keap1–Nrf2 pathway. It is involved in the proteasomal degradation of ubiquitinated proteins. When the cellular p62 level is manipulated, the quantity and location pattern of ubiquitinated proteins change with a considerable impact on cell survival. Altered p62 levels can even lead to some diseases. The proteotoxic stress imposed by proteasome inhibition can activate autophagy through p62 phosphorylation. A deficiency in autophagy may compromise the ubiquitin–proteasome system, since overabundant p62 delays delivery of the proteasomal substrate to the proteasome despite proteasomal catalytic activity being unchanged. In addition, p62 and the proteasome can modulate the activity of HDAC6 deacetylase, thus influencing the autophagic degradation.