Activation of PKA, p38 MAPK and ERK1/2 by gonadotropins in cumulus cells is critical for induction of EGF-like factor and TACE/ADAM17 gene expression during in vitro maturation of porcine COCs

<p>Abstract</p> <p>Objectives</p> <p>During ovulation, it has been shown that LH stimulus induces the expression of numerous genes via PKA, p38 MAPK, PI3K and ERK1/2 in cumulus cells and granulosa cells. Our recent study showed that EGF-like factor and its protease (TACE/ADAM17) are required for the activation of EGF receptor (EGFR), cumulus expansion and oocyte maturation of porcine cumulus-oocyte complexes (COCs). In the present study, we investigated which signaling pathways are involved in the gene expression of EGF-like factor and in <it>Tace/Adam17 </it>expression in cumulus cells of porcine COC during <it>in vitro </it>maturation.</p> <p>Methods</p> <p><it>Areg</it>, <it>Ereg</it>, <it>Tace/Adam17</it>, <it>Has2</it>, <it>Tnfaip6 </it>and <it>Ptgs2 </it>mRNA expressions were detected in cumulus cells of porcine COCs by RT-PCR. Protein level of ERK1/2 phosphorylation in cultured cumulus cells was analyzed by westernblotting. COCs were visualized using a phase-contrast microscope.</p> <p>Results</p> <p>When COCs were cultured with FSH and LH up to 2.5 h, <it>Areg</it>, <it>Ereg </it>and <it>Tace/Adam17 </it>mRNA were expressed in cumulus cells of COCs. <it>Areg</it>, <it>Ereg </it>and <it>Tace/Adam17 </it>gene expressions were not suppressed by PI3K inhibitor (LY294002), whereas PKA inhibitor (H89), p38 MAPK inhibitor (SB203580) and MEK inhibitor (U0126) significantly suppressed these gene expressions. Phosphorylation of ERK1/2, and the gene expression of <it>Has2</it>, <it>Tnfaip6 </it>and <it>Ptgs2 </it>were also suppressed by H89, SB203580 and U0126, however, these negative effects were overcome by the addition of EGF to the medium, but not in the U0126 treatment group.</p> <p>Conclusion</p> <p>The results showed that PKA, p38 MAPK and ERK1/2 positively controlled the expression of EGF-like factor and TACE/ADMA17, the latter of which impacts the cumulus expansion and oocyte maturation of porcine COCs via the EGFR-ERK1/2 pathway in cumulus cells.</p

トウキョウ ノ フランケンシュタイン アル オーストリア ハンガリー ガイコウカン ノ トウキョウ チュウザイ 1911 13 ネン 1

研究ノー

Individually separated supramolecular polymer chains toward solution-processable supramolecular polymeric materials

Herein, we present a simple design concept for a monomer that affords individually separated supramolecular polymer chains. Random introduction of alkyl chains with different lengths onto a monomer prevented its supramolecular polymers from bundling, permitting the preparation of concentrated solutions of the supramolecular polymer without gelation, precipitation, or crystallization. With such a solution in hand, we succeeded in fabricating self-standing films and threads consisting of supramolecular polymers

センソウ ジョウタイカ コッコウ ダンゼツカ ダイイチジ セカイ タイセンチュウ ニオケル ニホン ト オーストリア ハンガリー ノ コクサイホウジョウ ノ カンケイ ニツイテ ノ ガイコウシテキ コウサツ

研究ノー

トウキョウ ノ フランケンシュタイン アル オーストリア ハンガリー ガイコウカン ノ トウキョウ チュウザイ 1911 13 ネン ニ

研究ノー

Activation of Toll-like receptor 7/8 encoded by the X chromosome alters sperm motility and provides a novel simple technology for sexing sperm

In most mammals, the male to female sex ratio of offspring is about 50% because half of the sperm contain either the Y chromosome or X chromosome. In mice, the Y chromosome encodes fewer than 700 genes, whereas the X chromosome encodes over 3,000 genes. Although overall gene expression is lower in sperm than in somatic cells, transcription is activated selectively in round spermatids. By regulating the expression of specific genes, we hypothesized that the X chromosome might exert functional differences in sperm that are usually masked during fertilization. In this study, we found that Toll-like receptors 7/8 (TLR7/8) coding the X chromosome were expressed by approximately 50% of the round spermatids in testis and in approximately 50% of the epididymal sperm. Especially, TLR7 was localized to the tail, and TLR8 was localized to the midpiece. Ligand activation of TLR7/8 selectively suppressed the mobility of the X chromosome–bearing sperm (X-sperm) but not the Y-sperm without altering sperm viability or acrosome formation. The difference in sperm motility allowed for the separation of Y-sperm from X-sperm. Following in vitro fertilization using the ligand-selected high-mobility sperm, 90% of the embryos were XY male. Likewise, 83% of the pups obtained following embryo transfer were XY males. Conversely, the TLR7/8-activated, slow mobility sperm produced embryos and pups that were 81% XX females. Therefore, the functional differences between Y-sperm and X-sperm motility were revealed and related to different gene expression patterns, specifically TLR7/8 on X-sperm.This work was supported in part by Livestock Promotional Funds of Japan Racing Association (JRA) and by Hiroshima Cryopreservation Service Co. (to MS)

The acceleration of reproductive aging in Nrg1flox/flox;Cyp19-Cre female mice

Irregular menstrual cycles, reduced responses to exogenous hormonal treatments, and altered endocrine profiles (high FSH/high LH/low AMH) are observed in women with increasing age before menopause. In this study, because the granulosa cell-specific Nrg1 knockout mice (gcNrg1KO) presented ovarian and endocrine phenotypes similar to older women, we sought to understand the mechanisms of ovarian aging and to develop anewstrategy for improving fertility in older women prior to menopause. In the ovary of 6-month-old gcNrg1KO mice, follicular development was blocked in bilayer secondary follicles and heterogeneous cells accumulated in ovarian stroma. The heterogeneous cells in ovarian stroma were distinguished as two different types: (i) the LH receptor-positive endocrine cells and (ii) actin-rich fibrotic cells expressing collagen. Both the endocrine and fibrotic cells disappeared following long-term treatment with a GnRH antagonist, indicating that the high levels of serum LH induced the survival of both cell types and the abnormal endocrine profile to reduce fertility. Moreover, follicular development to the antral stages was observed with reduced LH and the disappearance of the abnormal stromal cells. Mice treated with the GnRH antagonist regained normal, recurrent estrous cycles and continuously delivered pups for at least for 3 months. We conclude that endocrine and matrix alternations occur within the ovarian stroma with increasing age and that abolishing these alternations resets the cyclical release of LH. Thus, GnRH antagonist treatments might provide a new, noninvasive strategy for improving fertility in a subset of aging women before menopause.This work was supported in part by The Japan Society for the Promotion of Science (JSPS) KAKENHI, JP24688028, JP 16H05017 (to MS) and JP15J05331 (to TU), by Japan Agency for Medical Research and Development (AMED) 16gk0110015 h0001 (to MS), and by National Institute of Health (NIH)-HD-076980 (to JSR)