The transcriptome of corona radiata cells from individual MII oocytes that after ICSI developed to embryos selected for transfer: PCOS women compared to healthy women

BACKGROUND: Corona radiata cells (CRCs) refer to the fraction of cumulus cells just adjacent to the oocyte. The CRCs are closely connected to the oocyte throughout maturation and their gene expression profiles might reflect oocyte quality. Polycystic ovary syndrome (PCOS) is a common cause of infertility. It is controversial whether PCOS associate with diminished oocyte quality. The purpose of this study was to compare individual human CRC samples between PCOS patients and controls. METHODS: All patients were stimulated by the long gonadotropin-releasing hormone (GnRH) agonist protocol. The CRC samples originated from individual oocytes developing into embryos selected for transfer. CRCs were isolated in a two-step denudation procedure, separating outer cumulus cells from the inner CRCs. Extracted RNA was amplified and transcriptome profiling was performed with Human Agilent® arrays. RESULTS: The transcriptomes of CRCs showed no individual genes with significant differential expression between PCOS and controls, but gene set enrichment analysis identified several cell cycle- and DNA replication pathways overexpressed in PCOS CRCs (FDR < 0.05). Five of the genes contributing to the up-regulated cell cycle pathways in the PCOS CRCs were selected for qRT-PCR validation in ten PCOS and ten control CRC samples. qRT-PCR confirmed significant up-regulation in PCOS CRCs of cell cycle progression genes HIST1H4C (FC = 2.7), UBE2C (FC = 2.6) and cell cycle related transcription factor E2F4 (FC = 2.5). CONCLUSION: The overexpression of cell cycle-related genes and cell cycle pathways in PCOS CRCs could indicate a disturbed or delayed final maturation and differentiation of the CRCs in response to the human chorionic gonadotropin (hCG) surge. However, this had no effect on the in vitro development of the corresponding embryos. Future studies are needed to clarify whether the up-regulated cell cycle pathways in PCOS CRCs have any clinical implications. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13048-014-0110-6) contains supplementary material, which is available to authorized users

A comparative analysis of the intestinal metagenomes present in guinea pigs (Cavia porcellus) and humans (Homo sapiens)

<p>Abstract</p> <p>Background</p> <p>Guinea pig (<it>Cavia porcellus</it>) is an important model for human intestinal research. We have characterized the faecal microbiota of 60 guinea pigs using Illumina shotgun metagenomics, and used this data to compile a gene catalogue of its prevalent microbiota. Subsequently, we compared the guinea pig microbiome to existing human gut metagenome data from the MetaHIT project.</p> <p>Results</p> <p>We found that the bacterial richness obtained for human samples was lower than for guinea pig samples. The intestinal microbiotas of both species were dominated by the two phyla <it>Bacteroidetes</it> and <it>Firmicutes</it>, but at genus level, the majority of identified genera (320 of 376) were differently abundant in the two hosts. For example, the guinea pig contained considerably more of the mucin-degrading <it>Akkermansia</it>, as well as of the methanogenic archaea <it>Methanobrevibacter</it> than found in humans. Most microbiome functional categories were less abundant in guinea pigs than in humans. Exceptions included functional categories possibly reflecting dehydration/rehydration stress in the guinea pig intestine. Finally, we showed that microbiological databases have serious anthropocentric biases, which impacts model organism research.</p> <p>Conclusions</p> <p>The results lay the foundation for future gastrointestinal research applying guinea pigs as models for humans.</p

Persistent Organic Pollutant Exposure Leads to Insulin Resistance Syndrome

International audienceBackground: the incidence of the insulin resistance syndrome has increased at an alarming rate worldwide, creating a serious challenge to public health care in the 21st century. Recently, epide-miological studies have associated the prevalence of type 2 diabetes with elevated body burdens of persistent organic pollutants (POPs). However, experimental evidence demonstrating a causal link between POPs and the development of insulin resistance is lacking. Objective: We investigated whether exposure to POPs contributes to insulin resistance and meta-bolic disorders. Methods: Sprague-Dawley rats were exposed for 28 days to lipophilic POPs through the con-sumption of a high-fat diet containing either refined or crude fish oil obtained from farmed Atlantic salmon. In addition, differentiated adipocytes were exposed to several POP mixtures that mimicked the relative abundance of organic pollutants present in crude salmon oil. We measured body weight, whole-body insulin sensitivity, POP accumulation, lipid and glucose homeostasis, and gene expres-sion and we performed micro array analysis. Results: Adult male rats exposed to crude, but not refined, salmon oil developed insulin resis-tance, abdominal obesity, and hepatosteatosis. The contribution of POPs to insulin resistance was confirmed in cultured adipocytes where POPs, especially organochlorine pesticides, led to robust inhibition of insulin action. Moreover, POPs induced down-regulation of insulin-induced gene-1 (Insig-1) and Lpin1, two master regulators of lipid homeostasis. Conclusion: Our findings provide evidence that exposure to POPs commonly present in food chains leads to insulin resistance and associated metabolic disorder

Optimizing Staining Protocols for Laser Microdissection of Specific Cell Types from the Testis Including Carcinoma In Situ

Microarray and RT-PCR based methods are important tools for analysis of gene expression; however, in tissues containing many different cells types, such as the testis, characterization of gene expression in specific cell types can be severely hampered by noise from other cells. The laser microdissection technology allows for enrichment of specific cell types. However, when the cells are not morphologically distinguishable, it is necessary to use a specific staining method for the target cells. In this study we have tested different fixatives, storage conditions for frozen sections and staining protocols, and present two staining protocols for frozen sections, one for fast and specific staining of fetal germ cells, testicular carcinoma in situ cells, and other cells with embryonic stem cell-like properties that express the alkaline phosphatase, and one for specific staining of lipid droplet-containing cells, which is useful for isolation of the androgen-producing Leydig cells. Both protocols retain a morphology that is compatible with laser microdissection and yield RNA of a quality suitable for PCR and microarray analysis

Sucrose Counteracts the Anti-Inflammatory Effect of Fish Oil in Adipose Tissue and Increases Obesity Development in Mice

BACKGROUND: Polyunsaturated n-3 fatty acids (n-3 PUFAs) are reported to protect against high fat diet-induced obesity and inflammation in adipose tissue. Here we aimed to investigate if the amount of sucrose in the background diet influences the ability of n-3 PUFAs to protect against diet-induced obesity, adipose tissue inflammation and glucose intolerance. METHODOLOGY/PRINCIPAL FINDINGS: We fed C57BL/6J mice a protein- (casein) or sucrose-based high fat diet supplemented with fish oil or corn oil for 9 weeks. Irrespective of the fatty acid source, mice fed diets rich in sucrose became obese whereas mice fed high protein diets remained lean. Inclusion of sucrose in the diet also counteracted the well-known anti-inflammatory effect of fish oil in adipose tissue, but did not impair the ability of fish oil to prevent accumulation of fat in the liver. Calculation of HOMA-IR indicated that mice fed high levels of proteins remained insulin sensitive, whereas insulin sensitivity was reduced in the obese mice fed sucrose irrespectively of the fat source. We show that a high fat diet decreased glucose tolerance in the mice independently of both obesity and dietary levels of n-3 PUFAs and sucrose. Of note, increasing the protein∶sucrose ratio in high fat diets decreased energy efficiency irrespective of fat source. This was accompanied by increased expression of Ppargc1a (peroxisome proliferator-activated receptor, gamma, coactivator 1 alpha) and increased gluconeogenesis in the fed state. CONCLUSIONS/SIGNIFICANCE: The background diet influence the ability of n-3 PUFAs to protect against development of obesity, glucose intolerance and adipose tissue inflammation. High levels of dietary sucrose counteract the anti-inflammatory effect of fish oil in adipose tissue and increases obesity development in mice