Epoxy resin, which demonstrates a shape memory effect, is reinforced by chopped carbon fibers (CCFs) to improve the thermal and mechanical properties. The interfacial interactions between 2-mm-long CCFs and epoxy make an impact on not only molecular motion but also the physical behaviors of CCFs/epoxy composites. In particular, shape recovery ability of CCFs/epoxy composites is enhanced with an increase in thermal conductivity generated by crossing CCFs in the epoxy system, although CCFs/epoxy composites containing small amounts of CCFs, such as 1 or 3 phr (parts per hundred rubber), show slower recovery rates than those of raw epoxy specimens due to the difficulty of making heat bridges in composites. With these results, it is confirmed that for specific time-dependent purpose, the shape recovery vector of CCFs/epoxy can be controlled using the amount of CCFs.

Spermatogonial stem cells (SSCs; also known as Asingle [As] spermatogonia in mice) divide to self-renew or to produce progenitor cells known as Apaired(Apr) spermatogonia in basal compartment of seminiferous tubules of mammalian testis. These characterized cells are the finally differentiated product of a developmental process referred to as “spermatogenesis.” In the development of SSCs it is critical to maintain a balance between self-renewal and differentiation. because an excess of either process will lead to infertility. these two processes are tightly controlled by intrinsic signals of SSCs and extrinsic signals from the microenvironment, known as the SSC niche. The SSC niche is formed by Sertoli cells, the only somatic cells found inside the seminiferous tubules. The WNT/β-catenin pathway is known to regulate Sertoli cell functions critical to their capacity to support spermatogenesis in the postnatal testis, but The mechanisms and factors of the pathway are not well known. We found a factor TLE3 (Transducin Like Enhancer Of Split 3). The transcriptional co-repressor TLE family is known to function as transcription co-repressors within the context of Wnt signaling by interacting with histone deacetylase HDAC2. We examined the expression level of TLE3 in various mouse tissues. As a result of RT-PCR, TLE3 showed significantly higher expression in testis than that in other tissues. Immunofluorescent analysis revealed that TLE3 and HDAC2 expression are differentially regulated in the mouse testis during postnatal development. In adult testis, TLE3 and HDAC2 were co-expressed in Sertoli cells. TLE3 and HDAC2 protein are also located in nucleus in mouse TM4 Sertoli cells. Taken together, TLE3 may play a role in regulating WNT/β-catenin pathway via interaction with HDAC2 in Sertoli cell. Futher studies are needed to look into factors that regulated by siTLE3 in Sertoli cell and interated with TLE3 in WNT/β-catenin pathway.

The Hippo signaling pathway is essential for regulating proliferation, differentiation, and apoptosis in mammalian cells. Hippo signaling pathway exists in most body tissues and organs, where it controls the size of organs and tissues by keeping cell growth in check and promoting cell death as needed. It has been reported that the members of Hippo signaling pathway are highly expressed in mammalian ovaries and uteri. However, the regulatory mechanism of this pathway in the uterus during estrous cycle regulation remains unclear. Serine/Threonine Protein Kinase 4 (STK4, also known as MST1, a homolog of Hippo in Drosophila) is a major factor of Hippo signaling pathway. However STK4 in the mouse uterus has not yet been examined. The purpose of our study was to determine the expression of STK4 during the estrous cycle and regulation by estrogen in the mouse uterus. We found that STK4 was dynamically expressed in uterine endometrium during the estrous cycle. STK4 highly expressed at the estrus, diestrus, and were found to dramatically decrease as it progressed to the proestrus, metestrus stage of uterus during the estrous cycle. Expression of STK4 was dominant in glandular epithelial and luminal epithelial of proestrus, estrus, and diestrus stage, whereas in metestrus stage, expression of gene intensity was faint. Estrogen or estrogen receptor antagonist ICI 182,780 treatment, in ovariectomized mouse uterus, Expression of STK4 and its downstream genes were increased by estrogen. Our results show that the Hippo signaling pathway is estrogen-dependent in the mouse uterus. These informations will give us on sights to understand uterine dynamics during the estrous cycle.

Primary oocytes that are arrested in first meiotic prophase for years enter maturation process to meet a critical precondition for successful fertilization. During maturation, oocyte finishes meiosis I and progresses to the metaphase II stage, achieving meiotic maturity. Although importance of oocyte maturation for oocyte quality has been recognized, it is not fully understood for molecular mechanisms underlying oocyte maturation. Here, we found that dexamethasone-induced Ras-related protein 1 (RASD1), a member of RAS superfamily of small GTPases, was expressed in the mouse ovary. Immunohistochemical analysis revealed that Rasd1 expression was dominant in oocyte cytoplasm. Real-time PCR and RT-PCR analyses showed that Rasd1 mRNA was steadily expressed in germinal vesicle (GV), germinal vesicle break down (GVBD), metaphase I (MI) oocytes, but decreased in metaphase II(MII) oocytes during oocyte maturation. Konckdown of Rasd1 using RNAi system in the GV oocytes suppressed oocyte maturation through disruption of meiotic spindle and formation of misarranged chromosomes. Taken together, Rasd1 is a critical factor for MI-MII transition of oocyte and is involved in the regulation of spindle formation during oocyte maturation. Further study is needed to examine relationship between Rasd1 and spindle formation in MI-MII transition.

The spermatogenesis and oogenesis-specific helix-loop-helix transcription factor 2 (Sohlh2) is exclusively expressed in germ cells of male and female gonad. Sohlh2 acts as a transcriptional factor via its specific DNA binding site, E-box to regulate target genes such as Lhx8, Zp genes, Ngn3. Sohlh2 localize in the female oocyte and in the male spermatogonia. However, the regulatory mechanism of Sohlh2 was poorly understood. In this study, we examine the patterns interacting with Sohlh2. First, we performed immunoprecipitation with the antibody against Sohlh2 protein extracts from the testis. Two-dimensional SDS-gel showed sexual distinguishable protein including Fkbp12, 13, 3, 59. Among them, Fkbp3 is a member of the immunophilin protein family, which play a role in immunoregulation and basic cellular processes involving protein folding and trafficking. Protein is a cis-trans prolyl isomerase that binds the immunosuppressants FK506 and rapamycin. It has a higher affinity for rapamycin than for FK506 and thus may be an important target molecule for immunosuppression by rapamycin. In the expression analysis of Fkbp3 is detected in the multiple tissues; intestine, stomach, kidney, spleen, liver, heart, brain, lung, uterus, ovary, testis. Here, we identified that Fkbp3 mRNA is detected in the ovary and testis, kidney, liver, heart, brain, lung. Immunostaning assay shows that Fkbp3 is localized at the spermatogonia in testis. In further studies, in order to confirm the interaction between Fkbp3 and Sohlh2, we will perform immunoprecipitation.

Tdrd family members contain Tudor domain repeat which is found in polar granules in Drophila. Tdrd12 is one of Tdrd family members. Tdrd12 contains a DEAD-box and a Tudor domain. However, the molecular mechanism and physiological function of Tdrd12 has not been described. To examine the expression pattern of Tdrd12, RT-PCR and Northern blot analysis were performed using total RNAs extracted from tissues; liver, intestine, heart, brain, kidney, lung, brain, uterus, ovary, and testis. The full-length of Tdrd12 was amplified from total RNA from mouse testis and cloning into the cloning vector. Cloned PCR products were purified,sequenced and analyzed using the ABI Prism Sequencer 3130XL. To look into Tdrd12 protein location, rabbit antibodies against mouse Tdrd12 were made using two epitopes: 1st epitope: (318~334)- SQRPNEKPLRLTEKKDC and 2nd epitope: (737~750)- LEAKEDKKARRPLC, and its specificity was tested using tissue extracts including the gonad. Here, we identified that Tdrd12 mRNA is detected in the ovary and testis, but not in other tissues. The size of its transcript is about 4.5kb on the northern blot. Antibody against Tdrd12 detects about 150 kDa protein on the western blot analysis. Immunostaining assay shows that Tdrd12 is localized at the spermatid in the seminiferous tubules. The current study is the first to investigate Tdrd12 expression is limited in the gonad. Thissuggest that Tdrd12 plays a role in the gonad like other known Tdrd family members, Tdrd1, Tdrd6, Tdrd7, and Tdrd9.

The spermatogenesis and oogenesis-specific helix-loop-helix transcription factor 2 (Sohlh2) is exclusively expressed in germ cells of male and female gonad. Sohlh2 acts as a transcriptional factor via its specific DNA binding site, E-box to regulate target genes such as Lhx8, Zp genes, Ngn3. Sohlh2 localize in the female oocyte and in the male spermatogonia. In recent studies, Sohlh2 knockout (KO) mice occurs abnormal spermagoenesis resulting in sperm defect. Sohlh2 KO male mice were infertility due to disruption of numerous gene expression. However, the gene profiles of Sohlh2 KO testes were not characterized and the regulatory mechanism of Sohlh2 was poorly understood. In this study, we analyzed the gene profiles and examined the possible mechanism of Sohlh2 in the spermatogenesis. First, we performed histological analysis such as Hematoxylin and eosin stain, Tunel assay, and Immunohistochemistry to show the onset of disruption of Sohlh2 KO testes. These results showed that Sohlh2 KO testes have atrophic seminiferous tubule due to increased apoptosis at 2 weeks old. And then we analyzed the whole gene profiles in the Sohlh2 KO testes at 2 weeks old. We found that 91 genes were regulated at least 5-fold in knockout testes. Among these, several genes are involved in meiotic process. Quantitative-PCR results are shown that several meiotic factors are significantly down-regulated in 2-weeks-old Sohlh2 KO testes compared with that of wild type mice. Through chromosome spreading assay, we observed that the formation of synaptonemal complex of homologous chromosome during the meiosis in Sohlh2 KO testes was not completed. These suggest that Sohlh2 is critical for regulation of numerous factors including meiotic factors either directly or indirectly. Therefore, mis-regulation of meiotic factors at prophase I of meiosis during spermatogenesis leads to disruption of spermatogenesis in Sohlh2 KO testes. Further studies are needed to look at the mechanism of Sohlh2 for regulation of target genes in detail.

Lhx8 is a member of the LIM-homeobox transcription factor family expressed in the mouse ovary. We discovered that Lhx8 knockout females lose oocytes within 7 days after birth. Lhx8–/–ovaries fail to maintain the primordial follicles and growing follicles. Lhx8–/–ovaries misexpress numerous oocyte-specific genes such as H1foo and Nlrp14. The molecular mechanism of there gulation of Lhx8 in the oocyte has not been described. We examined to characterize Lhx8 DNA binding elements and to identify its direct target genes in the oocyte. CAST was performed using glutathione transferase Lhx8 homeodomain fusion protein (GST-LHX8HD). A 15-bp random sequence flanked by 20-bp of fixed sequences were incubated with purified GST-LHX8HD protein. Unbound DNA was washed with binding reaction buffer. Bound DNA was eluted and re-amplified by PCR for the next round of CAST. Final PCR products were cloned and sequenced to derive consensus binding sequence. EMSA was performed using 32P-labeled oligomers. Binding reactions were conducted by incubating 32P-labeled probes with purified protein. Dual luciferase assays were carried out with extracts of total HEK293 cell which was transfected by the pGL4-promoter vector containing three artificial repeats of LBE(3xLBE-Luc) and overexpression vector carrying the Lhx8 homeodomain as recommended by Promega. We identified several cis-acting sites, TGATTG as Lhx8 DNA binding elements (LBE) using a library of randomly generated oligonucleotides by CAST. EMSA reslut shows that Lhx8 preferentially binds to the oligomer including Lhx8 binding element (TGATTG) with high affinity. In addition, we found that the relative luciferase activity of reporter construct containing three copies of TGATTG was increased by 2.3-fold with Lhx8 overexpression. These results suggest that Lhx8 preferentially binds Lhx8 DNA binding element, TGATTG, and can transactivate reporter genes through the LBE. The transcription of Lhx8 target gene in oocytes directly might be regulated by its during early folliculogenesis.