Preparation and Effect of (3-aminopropyl)triethoxysilane-Coated LiNi0.5Co0.2Mn0.3O2 Cathode Material for Lithium Ion Batteries.

Surface coating using (3-aminopropyl)triethoxysilane (APTES) has been applied to improve the electrochemical properties of LiNi0.5Co0.2Mn0.3O2 (NCM523) cathode materials. The APTES coating layer on the surface of NCM523 protects the direct contact area between the cathode material and the electrolyte, and facilitates the presence of electrons through the abundance of electron-rich amine groups, thereby improving electrochemical performance. X-ray photoelectron spectroscopy confirmed the existence of APTES coating layers on the surface of NCM523 cathode materials, revealing three peaks-N1s, O1s, and Si1s-that were not identified in bare NCM523. In addition, the discharge capacities of the bare electrode and the APTES-coated NCM523 electrode were 121.06 mAh/g and 156.43 mAh/g, respectively. To the best of our knowledge, the use of an APTES coating on NCM523 cathode materials for lithium-ion batteries has never been reported.

Role of Hydrophilic APTES-PP Separator in Enhancing the Electrochemical Performance of Ni-Rich Cathode for Li-Ion Battery.

Polypropylene (PP) separators essentially have poor compatibility with normal liquid electrolytes, including EC/DEC, due to the low surface energies of their hydrophobic surfaces. Therefore, they have a poor ability to retain electrolyte solutions within the separators because of low absorption capacity for the liquid electrolytes, which could directly damage the AC impedance and C-rate performance of LIBs. This study aims to improve the hydrophilicity and adhesion properties using (3-aminopropyl)triethoxysilane (APTES) coating on hydrophobic PP separators. Their surfaces were treated with a thin and stable silane layer, using APTES with -NH2 of the hydrophilic group. Hydrophilic PP separator surfaces with pore structures were fabricated by a facile solution-immersion method. The contact angle of the APTES-PP separator decreased from 102±2.5° to 60±1.5°. The electrochemical measurement results indicate that the cell using the modified PP separator showed a better initial discharge capacity of 165.79 mAh g-1 during the first cycle, at a current density of 0.1 C, as compared with the initial discharge capacity (141.61 mAh g-1) of the cell with a bare PP separator. The performances of all cells with coated PP separators were improved with regard to interfacial resistance, discharge capacity and C-rate capacity, compared to the uncoated PP separator.

In vitro development and gene expression of frozen-thawed 8-cell stage mouse embryos following slow freezing or vitrification.

OBJECTIVE: This study was performed to compare the efficiency of slow freezing and vitrification based on survival, development to blastocysts, and cell numbers of blastocysts. Changes in embryonic gene expression in fresh and frozen-thawed embryos were also examined. METHODS: Eight-cell stage embryos were collected from superovulated female BDF1 mice. The collected embryos were randomly divided into three groups. One group was maintained as fresh controls (n=42), one was frozen by slow freezing (n=43), and one was cooled by vitrification (n=43). After thawing or cooling, survival rates, development to blastocyst, and cell numbers and inner cell mass (ICM) cell numbers of blastocysts were compared with those of the control group. The expressions of eight genes (Rbm3, Birc5, Sod1, Sod2, Cirbp, Caspase3, Trp53, Hsp70.1) were examined by real time-quantitative polymerase chain reaction in the fresh and frozen-thawed embryos. RESULTS: There were no significant differences in the slow freezing and vitrification groups' survival rate after thawing (88.4% vs. 88.4%), development to blastocyst (100% vs. 97.4%), cell numbers (107.0±21.0 vs. 115.0±19.7), or ICM cell numbers of blastocysts (11.3±5.2 vs. 11.1±3.7). Cell numbers of blastocysts were significantly (p<0.05) lower in the frozen-thawed embryos than the fresh embryos. There were no significant differences in the slow freezing and the vitrification groups' expressions of the eight genes. The expressions of CirbP and Hsp70.1 were higher in the frozen-thawed embryos than in the fresh embryos but there were no significant differences. CONCLUSION: These results suggest that there were no significant differences between embryos that underwent slow freezing and vitrification.

Identification of mouse blastocyst genes that are downregulated by double-stranded RNA-mediated knockdown of Oct-4 expression.

Oct-4 is an essential transcription factor involved in the differentiation of the inner cell mass (ICM) in mouse blastocysts, and is thought to be the pluripotent gene of embryonic stem cells. However, downstream genes of Oct-4 and the mechanism by which it regulates the transcription machinery remain unclear. Here, we specifically knocked down Oct-4 gene expression in mouse blastocysts by double-stranded RNA (dsRNA) interference. A recently developed method, the annealing control primer (ACP) technique, was then used to identify the downstream genes of Oct-4. By using 120 arbitrary ACP, 10 clones were found to be differentially expressed in the knocked down embryos and the cloned genes were analyzed by DNA sequencing and BLAST searching. Quantitative real time reverse transcription (RT)-polymerase chain reaction (PCR) confirmed that the expression of these genes is altered by Oct-4 knockdown. Of the 10 genes, 8 (Atp6ap2, GK003, Ddb1, hRscp, Dppa1, Dpp3, Sap18, and Rent1) were downregulated and 2 (Rps14 and ETIF2B) were upregulated in Oct-4 dsRNA-injected blastocysts. One of the downregulated genes is developmental pluripotency associated-1 (Dppa1), which has already been identified as being an Oct-4 downstream gene. Two other genes, Rent1 and Sap18, were found to be Oct-4 downstream genes for the first time. The genes identified here will provide insights into the roles played by Oct-4 during embryonic development.

Identification of differentially expressed genes in murine embryos at the blastocyst stage using annealing control primer system.

The identification of embryo-specific genes would provide insights into early embryonic development. However, the current methods employed to identify the genes that are expressed at a specific developmental stage are labor intensive and suffer from high rates of false positives. Here we employed a new and accurate reverse transcription-polymerase chain reaction (RT-PCR) method that involves annealing control primers (ACPs) to identify the genes that are specifically or prominently expressed in mouse blastocysts compared to 4-cell stage embryos. Using 120 ACPs, we identified and sequenced 74 of these differentially expressed genes (DEGs). Basic Local Alignment Search Tool (BLAST) searches revealed that 53 were known genes, 9 encoded ribosomal proteins, and 12 were unknown genes. Of the known genes, 14 were selected and further characterized using real-time quantitative PCR to assess their stage-specific expression in mouse embryos. This analysis suggests that the ACP system is a very good method for the identification of stage-specific genes in small numbers of mouse embryos. Further analysis of the differentially expressed blastocyst genes we have identified will provide insights into the molecular basis of preimplantation development.

Oocyte-selective expression of MT transposon-like element, clone MTi7 and its role in oocyte maturation and embryo development.

Previously, we found MT transposon-like element, clone MTi7 (MTi7) is highly expressed in the mouse ovary. Here, we show that the MTi7 is expressed in the oocyte from the primordial to the preovulatory follicles. For RNA interference (RNAi), double stranded RNAs (dsRNAs) were prepared for MTi7 and c-mos, a control gene with known functions. Each dsRNA was microinjected into germinal vesicle (GV) stage oocytes or zygotes with pronuclei (PN), after which developmental changes, mRNA expression, and nuclear and microtubular organization were analyzed. We found a 43.4-53% GV arrest in the microinjected oocytes with a concomitant decrease in targeted mRNA expression. In MTi7 dsRNA-injected early and late PN zygotes, a 92.9% 1-cell arrest and 76.9% 2-cell arrest were observed, respectively. This is the first report of an oocyte-selective expression of MTi7 mRNA, and our results strongly suggest that MTi7 involved in the nuclear membrane breakdown during oocyte maturation and embryo development.

Identification of differentially regulated genes in bovine blastocysts using an annealing control primer system.

The identification of embryo-specific genes would provide insights into early embryonic development. However, the current methods employed to identify the genes that are expressed at a specific developmental stage are labor intensive and suffer from high rates of false positives. Here we employed a new and accurate reverse transcription-polymerase chain reaction (RT-PCR) technology that involves annealing control primers (ACPs) to identify the genes that are specifically or prominently expressed in bovine early blastocysts and hatched blastocysts produced in vitro. Using these techniques, a total of nine expressed sequence tags (ESTs) of genes that were differentially expressed in hatched blastocysts, as compared to blastocyst embryos, were cloned and sequenced. The cloned genes or ESTs (C1-C9) all exhibited significant sequence similarity with known bovine genes (99-100%; FTL, RPS12, LAPTM4a, and RPL12) or ESTs (80-94%; AIBP, CULLIN-1, HDLP, COX5a, and RECS1) of other species. As revealed by real time RT-PCR, these genes were regulated upstream in the hatched blastocyst stage during early implantation. These results suggest that this new, PCR-based differential display RT-PCR technique is a very useful tool for the identification of stage-specific genes of preimplantation embryos.

Bovine oocyte cytoplasm supports nuclear remodeling but not reprogramming of murine fibroblast cells.

Nuclear transfer (NT) is used to elucidate fundamental biological issues such as reversibility of cell differentiation and interactions between the cytoplasm and nucleus. To obtain an insight into interactions between the somatic cell nucleus and oocyte cytoplasm, nuclear remodeling and gene expression were compared in bovine oocytes that had received nuclei from bovine and mouse fibroblast cells. While the embryos that received nuclei from bovine fibroblast cells developed into blastocysts, those that received nuclei from mouse fibroblasts did not develop beyond the 8-cell stage. Similar nuclear remodeling procedures were observed in oocytes reconstructed with mouse and bovine fibroblast cells. Foreign centrosomes during NT were introduced into embryos reconstructed with both fibroblast cell types. A number of housekeeping mouse genes (hsp70, bax, and glt-1) were abnormally expressed in embryos that had received nuclei from mouse fibroblast cells. However, development-related genes, such as Oct-4 and E-cad, were not expressed. The results collectively suggest that the bovine oocyte cytoplasm supports nuclear remodeling, but not reprogramming of mouse fibroblast cells.

Maternal gamma (gamma)-tubulin is involved in microtubule reorganization during bovine fertilization and parthenogenesis.

In this study, gamma-tubulin distribution was determined chronologically in conjunction with microtubule dynamics during bovine fertilization and parthenogenesis. In unfertilized bovine oocytes, gamma-tubulin was identified in the cytoplasm, mainly in the cortex and concentrated in the meiotic spindle. Following sperm penetration, gamma-tubulin in the cytoplasm was recruited by a sperm component. During pronuclear apposition, gamma-tubulin was localized as spots at the spindle poles. gamma-tubulin spots were observed in blastomeres of embryos cleaved in vitro. Following electrical stimulation, gamma-tubulin and microtubule matrix were noted in oocyte cortex. In the late pronuclear stage, considerably less gamma-tubulin and microtubules were detected in the cytoplasm. At the mitotic metaphase of parthenotes, gamma-tubulin was recruited to the condensed chromatin and concentrated in the spindle. The gamma-tubulin spots were not detected until the 8-cell stage of parthenotes. This suggests that maternal gamma-tubulin is recruited by a sperm component to reconstitute the zygotic centrosome. In the absence of sperm components, the cell cycle-related assembly of gamma-tubulin organizes microtubule nucleation for positioning the pronucleus and spindle protein of mitotic metaphase during the first cell cycle of bovine parthenotes.

Nuclear and microtubule reorganization in nuclear-transferred bovine embryos.

We studied the nuclear and microtubule dynamics in nonactivated and pre-activated chromatin-removed oocytes following transfer of nuclei from bovine fibroblast cells. Immediately after fusion between membranes of oocytes and fibroblasts, a microtubule aster containing a gamma-tubulin spot was seen near the transferred nucleus in most oocytes regardless of activation conditions. Most fibroblast nuclei transferred into nonactivated oocytes underwent premature chromosome condensation (PCC) and divided into two masses of chromosomes. In contrast, fibroblast nuclei in pre-activated oocytes rarely underwent PCC, but formed a swelled pronuclear-like structure. Under nonactivation condition, microtubular spindles surrounded condensed chromosomes during the division of two nuclear structures. Gamma-tubulins were detected in the vicinity of condensed chromosomes, suggesting transient spindle formation. Two pronuclear-like structures near the microtubular aster containing gamma-tubulin spot(s) later formed a syngamy-like nuclear structure. While 20% of reconstructed oocytes under nonactivated conditions developed to morulae and blastocysts, only 4% of reconstructed oocytes under pre-activated conditions developed to morulae and blastocysts. These results suggest introduction of a foreign centrosome during somatic cell nuclear transfer, which probably plays a role in nuclear remodeling and subsequent development.