Fang-Nong Lai

Oxidative stress induced by zearalenone in porcine granulosa cells and its rescue by curcumin in vitro.

Oxidative stress (OS), as a signal of aberrant intracellular mechanisms, plays key roles in maintaining homeostasis for organisms. The occurrence of OS due to the disorder of normal cellular redox balance indicates the overproduction of reactive oxygen species (ROS) and/or deficiency of antioxidants. Once the balance is broken down, repression of oxidative stress is one of the most effective ways to alleviate it. Ongoing studies provide remarkable evidence that oxidative stress is involved in reproductive toxicity induced by various stimuli, such as environmental toxicants and food toxicity. Zearalenone (ZEA), as a toxic compound existing in contaminated food products, is found to induce mycotoxicosis that has a significant impact on the reproduction of domestic animals, especially pigs. However, there is no information about how ROS and oxidative stress is involved in the influence of ZEA on porcine granulosa cells, or whether the stress can be rescued by curcumin. In this study, ZEA-induced effect on porcine granulosa cells was investigated at low concentrations (15 μM, 30 μM and 60 μM). In vitro ROS levels, the mRNA level and activity of superoxide dismutase, glutathione peroxidase and catalase were obtained. The results showed that in comparison with negative control, ZEA increased oxidative stress with higher ROS levels, reduced the expression and activity of antioxidative enzymes, increased the intensity of fluorogenic probes 2’, 7’-Dichlorodihydrofluorescin diacetate and dihydroethidium in flow cytometry assay and fluorescence microscopy. Meanwhile, the activity of glutathione (GSH) did not change obviously following 60 μM ZEA treatment. Furthermore, the underlying protective mechanisms of curcumin on the ZEA-treated porcine granulosa cells were investigated. The data revealed that curcumin pre-treatment significantly suppressed ZEA-induced oxidative stress. Collectively, porcine granulosa cells were sensitive to ZEA, which may induce oxidative stress. The findings from this study clearly demonstrate that curcumin is effective to reduce the dysregulation of cellular redox balance on porcine granulosa cells in vitro and should be further investigated for its protective role against ZEA in animals.

Di (2-ethylhexyl) Phthalate Exposure Impairs Growth of Antral Follicle in Mice

Di (2-ethylhexyl) phthalate (DEHP) is a widely used plastic additive. As an environmental endocrine disruptor, it has been shown to be harmful to the mammalian reproductive system. Previous studies indicated that DEHP inhibited the development of mouse ovarian follicles. However, the mechanisms by which DEHP affects ovarian antral follicle development during the pre-puberty stage are poorly understand. Thus, we investigated the effects of direct DEHP exposure on antral follicle growth in pre-pubescent mice by use of intraperitoneal injection. Our results demonstrated that the percentage of large antral follicles was significantly reduced when mice were exposed to 20 or 40 μg/kg DEHP every 5 days from postnatal day 0 (0 dpp) to 15 dpp. In 20 dpp, we performed microarray of these ovaries. The microarray results indicated that mRNA levels of apoptosis related genes were increased. The mRNA levels of the apoptosis and cell proliferation (negative) related genes Apoe, Agt, Glo1 and Grina were increased after DEHP exposure. DEHP induced the differential gene expression of Hsp90ab1, Rhoa, Grina and Xdh which may play an important role in this process. In addition, TUNEL staining and immunofluorescence showed that DEHP exposure significantly increased the number of TUNEL, Caspase3 and γH2AX positive ovarian somatic cells within the mouse ovaries. Flow cytometer analyses of redox-sensitive probes showed that DEHP caused the accumulation of reactive oxygen species. Moreover, the mRNA expression of ovarian somatic cell antioxidative enzymes was down-regulated both in vivo and in vitro. In conclusion, our data here demonstrated that DEHP exposure induced oxidative stress and ovarian somatic cell apoptosis, and thus may impact antral follicle enlargement during the pre-pubertal stage in mice.

Cutaneous applied nano-ZnO reduce the ability of hair follicle stem cells to differentiate

Abstract The ability of metal oxide nanoparticles to penetrate the skin has aroused a great deal of interest during the past decade due to concerns over the safety of topically applied sunscreens that contain physical UV-resistant metal particles, such as nano-Zinc oxide (nZnO). Previous studies demonstrate that metal oxide nanoparticles accumulate in skin furrows and hair follicles following topical application while little is known about the consequence of these nanoparticles on skin homeostasis. The current investigation tested the effects of nZnO (0.5 mg/day mouse) on hair follicle physiology. Topical application of Vaseline containing nZnO, bulk ZnO (bZnO), or ionized Zn to newborn mice vibrissa pad over a period of 7 consecutive days revealed that nZnO accumulated within hair follicles, and this induced the apoptosis of hair follicle stem cells (HFSCs). In vitro studies also indicated that nZnO exposure caused obvious DNA damage and induced apoptosis in HFSCs. Furthermore, it was found that nZnO exposure perturbed genes associated with HFSC apoptosis, cell communication, and differentiation. HFSCs transplantation assay demonstrated that the potential of HFSCs to differentiate was reduced. This investigation indicates a potential risk of topically applied ZnO nanoparticles on skin homeostasis.

Whole-genome scanning for the litter size trait associated genes and SNPs under selection in dairy goat ( Capra hircus )

Dairy goats are one of the most utilized domesticated animals in China. Here, we selected extreme populations based on differential fecundity in two Laoshan dairy goat populations. Utilizing deep sequencing we have generated 68.7 and 57.8 giga base of sequencing data, and identified 12,458,711 and 12,423,128 SNPs in the low fecundity and high fecundity groups, respectively. Following selective sweep analyses, a number of loci and candidate genes in the two populations were scanned independently. The reproduction related genes CCNB2, AR, ADCY1, DNMT3B, SMAD2, AMHR2, ERBB2, FGFR1, MAP3K12 and THEM4 were specifically selected in the high fecundity group whereas KDM6A, TENM1, SWI5 and CYM were specifically selected in the low fecundity group. A sub-set of genes including SYCP2, SOX5 and POU3F4 were localized both in the high and low fecundity selection windows, suggesting that these particular genes experienced strong selection with lower genetic diversity. From the genome data, the rare nonsense mutations may not contribute to fecundity, whereas nonsynonymous SNPs likely play a predominant role. The nonsynonymous exonic SNPs in SETDB2 and CDH26 which were co-localized in the selected region may take part in fecundity traits. These observations bring us a new insights into the genetic variation influencing fecundity traits within dairy goats.

Di (2-ethylhexyl) phthalate exposure impairs meiotic progression and DNA damage repair in fetal mouse oocytes in vitro.

Di (2-ethylhexyl) phthalate (DEHP), is the most common member of the class of phthalates that are used as plasticizers and have become common environmental contaminants. A number of studies have shown that DEHP exposure impacts reproductive health in both male and female mammals by acting as an estrogen analog. Here, we investigated the effects of DEHP on meiotic progression of fetal mouse oocytes by using an in vitro model of ovarian tissue culture. The results showed that 10 or 100 μM DEHP exposure inhibited the progression of oocytes throughout meiotic prophase I, specifically from the pachytene to diplotene stages. DEHP possibly impairs the ability to repair DNA double-strand breaks induced by meiotic recombination and as a consequence activates a pachytene check point. At later stages, such defects led to an increased number of oocytes showing apoptotic markers (TUNEL staining, expression of pro-apoptotic genes), resulting in reduced oocyte survival, gap junctions, and follicle assembly in the ovarian tissues. Microarray analysis of ovarian tissues exposed to DEHP showed altered expression of several genes including some involved in apoptosis and gonad development. The expression changes of some genes clustered in cell-cell communication and signal transduction, along with plasma membrane, extracellular matrix and ion channel function classes, were dependent on the DEHP concentration. Together, these results bring new support to the notion that exposure to DEHP during gestation might exert deleterious effects on ovary development, perturbing germ cell meiosis and the expression of genes involved in a wide range of biological processes including ovary development.

LncRNA as ceRNAs may be involved in lactation process

The main function of the mammary gland is to secret milk for newborn growth. Milk production process is regulated by hormones, growth factors, noncoding RNAs and other factors locally. Long non-coding RNAs (lncRNAs), one type of recently discovered non-coding RNA, have been found in mammary gland and some studies suggested lncRNA may play important roles in mammary gland development. Competing endogenous RNAs (ceRNAs) are emerging to compete for miRNA binding and, in turn, regulate each other. In the current study, we sequenced mRNA, miRNA and lncRNA in goat mammary tissue at 2 points in lactation (early and mature). All data were co-expressed together from the same samples. Our data showed that the ceRNAs up-regulated during the mature lactation phase were associated with lipid, protein, carbon and amino acid synthesis and metabolism. This correlates with the function of the mature lactation phase: i.e. the continuous production of large amounts of milk, rich in proteins, lipids, amino acids and other nutrients. Alternately, the ceRNAs up-regulated during early lactation were associated with PI3K-AKT pathways and ECM-receptor interactions; these fulfil the functional role of preparing the mammary gland for full lactation. Therefore, the results suggest that ceRNAs work synergistically during different developmental stages to regulate specific functions associated with lactation control. This study suggests that ceRNAs (lncRNA-mRNA) may be involved in lactation process.

Low-dose diethylhexyl phthalate exposure does not impair the expressive patterns of epigenetics-related genes and DNA methylation of breast cancer-related genes in mouse mammary glands

BackgroundsDi-(2-ethylhexyl) phthalate (DEHP), as an endocrine-disrupting chemical (EDC), is widely used in plasticizer and other productions. Ubiquitous human exposure to DEHP has been proposed to be a potential risk to public health. Developmental exposure to DEHP could alter epigenetic programming and result in adult-onset disease.MethodsIn this study, we investigated whether DEHP exposure to pregnant mice affected epigenetic changes as a result of increase in breast cancer incidence.ResultsOur results showed that the expression of total 143 epigenetics-related genes in mammary gland cells, have no significantly altered after short time and low-dose treated with DEHP from 0.5 days post-coitum (dpc) to 3.5 dpc of pregnant mice. DNA methylation status of some neoplastic development genes, such as EGFr, Esr1, Pgr, Fos and Rassf5 also had no obvious change.ConclusionThese finding showed no impact of DEHP on the expressive patterns of epigenetics-related genes and DNA methylation of breast cancer-related genes in pregnant mouse mammary gland cells.

Analysis of the SNP loci around transcription start sites related to goat fecundity trait base on whole genome resequencing

Genome flanking regions surrounding transcription start sites (TSSs) are critical for the regulation of gene expression, containing many translational regulatory elements. To investigate whether critical single nucleotide polymorphisms (SNPs) exist around TSSs in the dairy goat genome, we performed high throughput DNA sequencing to compare two dairy goat groups with discrepant litter sizes. After genome mapping, SNP calling, and annotation, we screened the SNPs within 2kb scales surrounding annotated TSSs in high fecundity (HF) and low fecundity (LF) groups, respectively. We attempted to identify distinct SNPs and motifs near the TSSs in both groups. The SNPs near the TSSs most were consistent; 318 new SNPs were uncovered in the HF group, of which 305 were heterozygote SNPs, 13 were homozygote SNPs, and majority of which were distributed on chromosome 2 and 29. After validation by Sanger sequencing we found that a SNP in CHI16: 27612330 C>A in the PSEN2 gene presented an A/A genotype in the HF group and an A/A or A/C genotype in the LF group. In conclusion, our study provides insightful information into the dairy goat genomic variations surrounding TSSs, which may contribute to enhanced litter size. Based on comparison studies of SNPs exist around transcription start sites between high fecundity group and low fecundity group. Our finding provides insights concerning the goat litter size phenotypic and will promote future goat breeding.