Metformin therapy in a hyperandrogenic anovulatory mutant murine model with polycystic ovarian syndrome characteristics improves oocyte maturity during superovulation

<p>Abstract</p> <p>Background</p> <p>Metformin, an oral biguanide traditionally used for the treatment of type 2 diabetes, is widely used for the management of polycystic ovary syndrome (PCOS)-related anovulation. Because of the significant prevalence of insulin resistance and glucose intolerance in PCOS patients, and their putative role in ovulatory dysfunction, the use of metformin was touted as a means to improve ovulatory function and reproductive outcomes in PCOS patients. To date, there has been inconsistent evidence to demonstrate a favorable effect of metformin on oocyte quality and competence in women with PCOS. Given the heterogeneous nature of this disorder, we hypothesized that metformin may be beneficial in mice with aberrant metabolic characteristics similar to a significant number of PCOS patients. The aim of this study was to gain insight into the <it>in vitro </it>and <it>in vivo </it>effects of metformin on oocyte development and ovulatory function.</p> <p>Methods</p> <p>We utilized metformin treatment in the transgenic <it>ob/ob </it>and <it>db/db </it>mutant murine models which demonstrate metabolic and reproductive characteristics similar to women with PCOS. Results: Metformin did not improve <it>in vitro </it>oocyte maturation nor did it have an appreciable effect on <it>in vitro </it>granulosa cell luteinization <it>(</it>progesterone production) in any genotype studied. Although both mutant strains have evidence of hyperandrogenemia, anovulation, and hyperinsulinemia, only <it>db/db </it>mice treated with metformin had a greater number of mature oocytes and total overall oocytes compared to control. There was no observed impact on body mass, or serum glucose and androgens in any genotype.</p> <p>Conclusions</p> <p>Our data provide evidence to suggest that metformin may optimize ovulatory performance in mice with a specific reproductive and metabolic phenotype shared by women with PCOS. The only obvious difference between the mutant murine models is that the <it>db/db </it>mice have elevated leptin levels raising the questions of whether their response to metformin is related to elevated leptin levels and/or if a subset of PCOS women with hyperleptinemia may be responsive to metformin therapy. Further study is needed to better define a subset of women with PCOS that may be responsive to metformin.</p

Magnetic and Cryogenic Design of the MICE Coupling Solenoid Magnet System

The Muon Ionization Cooling Experiment (MICE) will demonstrate ionization cooling in a short section of a realistic cooling channel using a muon beam at Rutherford Appleton Laboratory in the UK. The coupling magnet is a superconducting solenoid mounted around four 201MHz RF cavities, which produces magnetic field up to 2.6 T on the magnet centerline to keep muons within the iris of RF cavities windows. The coupling coil with inner radius of 750mm, length of 285mm and thickness of 102.5mm will be cooled by a pair of 1.5 W at 4.2 K small coolers. This paper will introduce the updated engineering design of the coupling magnet made by ICST in China. The detailed analyses on magnetic fields, stresses induced during the processes of winding, cool down and charging, and cold mass support assembly are presented as well

High-throughput gene discovery in the rat

The rat is an important animal model for human diseases and is widely used in physiology. In this article we present a new strategy for gene discovery based on the production of ESTs from serially subtracted and normalized cDNA libraries, and we describe its application for the development of a comprehensive nonredundant collection of rat ESTs. Our new strategy appears to yield substantially more EST clusters per ESTs sequenced than do previous approaches that did not use serial subtraction. However, multiple rounds of library subtraction resulted in high frequencies of otherwise rare internally primed cDNAs, defining the limits of this powerful approach. To date, we have generated >200,000 3′ ESTs from >100 cDNA libraries representing a wide range of tissues and developmental stages of the laboratory rat. Most importantly, we have contributed to ∼50,000 rat UniGene clusters. We have identified, arrayed, and derived 5′ ESTs from >30,000 unique rat cDNA clones. Complete information, including radiation hybrid mapping data, is also maintained locally at http://genome.uiowa.edu/clcg.html. All of the sequences described in this article have been submitted to the dbEST division of the NCBI

MicroRNA-15b regulates reversion-inducing cysteine-rich protein with Kazal motifs (RECK) expression in human uterine leiomyoma

Background: Human uterine leiomyoma (fibroids; LYO) are the most common benign neoplasms in reproductive-aged women. Dysregulated extracellular matrix and irregular LYO reversion-inducing cysteine-rich protein with Kazal motifs (RECK) expression are thought to be mediated by aberrant microRNA (miR) expression. The relationship of miR-15b and RECK expression in LYO has not been studied. Methods: The expression levels of miR-15b and RECK were determined by quantitative RT-PCR, Western blot, and immunohistochemistry in cultures derived from commercial primary leiomyoma (cpLYO) and myometrial (cpMYO) cell lines and leiomyoma (pLYO) and myometrium (pMYO) tissue from surgical samples respectively. The relationship between miR-15b and RECK expression in cpLYO and pLYO (compared to their respective myometrial controls) was evaluated following transfection of cell cultures with either miR-15b mimic or inhibitor. Results: Elevated levels of miR-15b were observed in cpLYO (2.82-fold; p = 0.04) and pLYO cell (1.30-fold; p = 0.0001) cultures respectively compared to corresponding MYO cell controls. Following transfection with miR-15b mimic, cpLYO cells (0.62-fold; p < 0.0001) and pLYO cells (0.68-fold; p < 0.0001) demonstrated reduced RECK protein expression. Following transfection with miR-15b inhibitor, cpLYO cells (1.20-fold; p < 0.0001) and pLYO cells (1.31-fold; p = 0.0007) demonstrated elevated RECK protein expression. RECK protein expression was reduced in pLYO tissues (0.73-fold; p < 0.0001) and pLYO (0.47-fold; p = 0.047) cells when compared to the corresponding MYO tissue controls. Conclusion: Our findings suggest that miR-15b negatively regulates RECK expression in LYO, and increased miR-15b and decreased RECK expression may contribute to the pathobiology of LYO. The functional significance of miR-15b and RECK expression warrants further investigation as potential therapeutic targets for the treatment of human LYO

Nuclear accumulation of glioma-associated oncogene 2 protein and enhanced expression of forkhead-box transcription factor M1 protein in human hepatocellular carcinoma

The hedgehog (Hh) signaling pathway has been reported to be crucial in human carcinogenesis and tumor progression. Glioma-associated oncogenes (Gli), are zinc finger transcription factors which mediate the transcriptional response to Hh signaling. To explore the role of Gli in the development and progression of hepatocellular carcinoma (HCC), we investigated the expression of Gli2 and FoxM1 (forkhead-box transcription factor M1) which is one of the Gli downstream target genes modulating cell cycle progression in 91 specimens of human HCCs with immunohistochemistry. These immunostaining results were compared with various clinicopathologic parameters. Immunoreactivity of Gli2 and FoxM1 was observed respectively in 84.6% (77/91) and 80.2% (73/91) cases of HCC tumor tissues, and this was considerably higher than expression in the peritumoral tissues. Distribution of Gli2 and FoxM1 proteins in tumor cells was nuclear with or without cytoplasmic staining, or cytoplasmic alone. Statistically, increased nuclear immunopositivity of Gli2 protein correlated significantly with poorer tumor differentiation (P<0.05), as well as with portal vein tumor thrombosis (P<0.05). In addition, overexpression of FoxM1 protein was significantly associated with increased tumor grade (P<0.01) and advanced tumor stage (P<0.05). Moreover, there was a significant association between the expressions of Gli2 and FoxM1 proteins in HCC (r=0.464, P=0.000). This is consistent with the concept that in human HCC, the Hh signaling pathway is involved in the differentiation and proliferation of tumor cells, in part through inducing nuclear accumulation of Gli2 protein and subsequent upregulation of FoxM1 protein

Expression and distribution of cytokeratin 8-18 intermediate filaments in bovine antral follicles and corpus luteum: An intrinsic mechanism of resistance to apoptosis?

Apoptosis is a mechanism of cell elimination during follicular atresia and luteal regression. Recent evidence suggests sensitivity to apoptosis in some cell types is partly dependent upon cytokeratin-containing intermediate filaments. Specifically, cytokeratin 8/18 (CK8/18) filaments are thought to impart resistance to apoptosis. Here, cytokeratin filament expression within bovine ovarian follicles and corpora lutea (CL) was characterized and the potential relationship between cellspecific CK8/18 expression and apoptosis explored. Immunoprecipitation and western blot analysis confirmed CK8 associates with CK18 to form CK8/18 heterodimeric filaments within bovine ovarian cells. Immunostaining revealed populations of CK18-positive (CK18+) cells in healthy growing follicles that increased in postovulatory follicles. Atretic follicles at all stages of atresia also contained some CK18+ cells. However, no CK18+ cells were detected in primordial or primary follicles. In CL, developing CL contained a higher proportion of CK18+ cells (~35%, range 30-70%) than mature CL (~16%) and regressing CL (~5%; P<0.05, n = 3-5 CL/stage), suggesting CK8/18 filament expression diminishes over time, as luteal cells become more susceptible to apoptosis. Dual-fluorescence labeling for CK18 and a cell death marker (TUNEL labeling) confirmed this view, demonstrating less death of CK18+ than CK18- luteal cells throughout the estrous cycle (P<0.05). The results indicate differential expression of CK8/18 filaments occurs in cells of bovine ovarian follicles and CL throughout the estrous cycle. The prevalence and cell-specific pattern of cytokeratin expression in these structures is consistent with the concept these filaments might impart resistance to apoptosis in ovarian cells as is seen in other cell types