A novel solution for freezing individual spermatozoa using a right angular cryopiece embedded in a grooved petri dish.

Herein, we introduced a novel individual sperm freezing device named SpermCD, which consists of a right angular cryopiece (RA-Cryopiece, or "C") and a grooved petri dish ("D"). SpermCD allows embryologists to transfer sperm and perform ICSI on the same focal plane. Thirty-five patients underwent single sperm cryopreservation using SpermCD, including four patients with non-obstructive azoospermia (NOA), 14 patients with virtual azoospermia and 17 patients with cryptozoospermia. One hundred and twenty-five cryopreserved spermatozoa from nine patients were thawed on the day of the oocyte retrieval and 121 spermatozoa were found, with a sperm recovery rate of 97.1 ± 4.6%. Sixty-five MII oocytes from their spouse were injected with thawed sperm. Normal fertilization and high-quality embryo rates were 68.0% ± 33.2% and 24.4% ± 22.2%. Nineteen transplantable embryos were formed after fertilization with frozen sperm, eight of which were transplanted in five couples, resulting in four successful deliveries. SpermCD is a simple and practical individual sperm freezing device.

N6-methyladenosine demethylase FTO suppresses clear cell renal cell carcinoma through a novel FTO-PGC-1α signalling axis.

The abundant and reversible N6-methyladenosine (m6A) RNA modification and its modulators have important roles in regulating various gene expression and biological processes. Here, we demonstrate that fat mass and obesity associated (FTO), as an m6A demethylase, plays a critical anti-tumorigenic role in clear cell renal cell carcinoma (ccRCC). FTO is suppressed in ccRCC tissue. The low expression of FTO in human ccRCC correlates with increased tumour severity and poor patient survival. The Von Hippel-Lindau-deficient cells expressing FTO restores mitochondrial activity, induces oxidative stress and ROS production and shows impaired tumour growth, through increasing expression of PGC-1α by reducing m6A levels in its mRNA transcripts. Our work demonstrates the functional importance of the m6A methylation and its modulator, and uncovers a critical FTO-PGC-1α axis for developing effective therapeutic strategies in the treatment of ccRCC.

Telbivudine can safely reduce mother-to-child transmission in chronic hepatitis B women after 12 weeks of gestation.

BACKGROUND: To evaluate the efficacy and safety of telbivudine in chronic hepatitis B women during the second and third trimesters of pregnancy. METHODS: The week 12-34 of pregnant women were screened in this prospective non-intervention study, with HBV DNA > 106 IU/mL and alanine aminotransferase > 50 IU/L. The patients were received telbivudine treatment as a treatment group or without antiviral treatment as a control group. All infants were received recombinant hepatitis B vaccine 10 µg within 12 h of birth, at week 4 and week 24, immunoglobulin G within 12 h of birth and were detected HBV markers at the range from 7 to 12 months after delivery. RESULTS: A total of 241 patients were finally enrolled, 139 patients in telbivudine group and 102 patients in control group. HBsAg negative rate of infants was 99.3% (135/136) in telbivudine group and was 91.9% (91/99) in control group after 7 months (P = 0.005), respectively. The incidence of undetectable HBV DNA levels (47.5%) was significantly lower in telbivudine-treated mothers than that in the controls (0%), and 75.5% patients alanine aminotransferase returned to normal in telbivudine group, and 51% in control group at delivery (P < 0.001), respectively. CONCLUSIONS: Telbivudine can safely reduce mother-to-child transmission in chronic hepatitis B women after 12 weeks of gestation.

Acetyl-CoA Synthetase 2 Promotes Cell Migration and Invasion of Renal Cell Carcinoma by Upregulating Lysosomal-Associated Membrane Protein 1 Expression.

BACKGROUND/AIMS: Reprogramming energy metabolism is an emerging hallmark of many cancers, and this alteration is especially evident in renal cell carcinomas (RCCs). However, few studies have been conducted on lipid metabolism. This study investigated the function and mechanism of lipid metabolism-related acetyl-CoA synthetase 2 (ACSS2) in RCC development, cell migration and invasion. METHODS: Quantitative real-time PCR (qRT-PCR) was used to determine the expression of ACSS2 in cancer tissue and adjacent tissue. The inhibition of ACSS2 expression was achieved by RNA interference, which was confirmed by qRT-PCR and Western blotting. Cell proliferation and apoptosis were detected by a CCK8 assay and a flow cytometry analysis, respectively. Cell migration and invasion were determined by the scratch and transwell assays. Following the knockdown of ACSS2 expression, the expression of the autophagy-related factor LAMP1 was measured by qRT-PCR and Western blotting. RESULTS: Compared to adjacent tissues, ACSS2 expression was upregulated in RCC cancer tissues and positively correlated with metastasis. Inhibition of ACSS2 had no effect on RCC cell proliferation or apoptosis. However, decreased ACSS2 expression was found to inhibit RCC cell migration and invasion. ACSS2 was determined to promote the expression of LAMP1, which can also promote cell migration. This pathway may be considered a potential mechanism through which ACSS2 participates in RCC development. CONCLUSION: These data suggest that ACSS2 is an important factor for promoting RCC development and is essential for cell migration and invasion, which it promotes by increasing the expression of LAMP1. Taken together, these findings reveal a potential target for the diagnosis and treatment of RCC.

Plac1 Expression Pattern at the Mouse Fetomaternal Interface and Involvement in Trophoblast Differentiation.

BACKGROUND/AIMS: It is well known that Plac1 is a placenta-specific gene; however, its spatiotemporal expression pattern and exact role at t h e mouse fetomaternal interface r e m a i n s unclear. METHODS: In situ hybridization (ISH) was used to localize the Plac1 mRNA at the mouse fetomaternal interface. A trophoblast stem cell (TS) differentiation model with Plac1 shRNA-overexpressing lentivirus was employed to investigate the possible role of Plac1 in placentation. Real-time RT-PCR was used to detect changes in gene expression. RESULTS: Plac1 was exclusively expressed in the ectoplacental cone (EPC) as well as in 8.5 and 9.5 days post-coitum (dpc) embryos. Subsequently, Plac1 expression was abundant in the spongiotrophoblast layer and moderately in the labyrinth layer until 13.5 dpc, and declined thereafter. Interestingly, Plac1 was also expressed by secondary trophoblast giant cells and glycogen trophoblast cells, but not in primary trophoblast giant cells. Plac1 transcription was increased during the TS differentiation (P < 0.01), and knockdown of Plac1 significantly impaired TS differentiation. CONCLUSION: Plac1 is abundantly expressed at the fetomaternal interface and in all trophoblast subtypes except in primary trophoblast giant cells. Plac1 knockdown retarded the progress of TS differentiation, indicating that Plac1 is necessary for normal trophoblast differentiation into various trophoblast subpopulations.

Are testicular sperms superior to ejaculated sperms in couples with previous ART failure due to high rate of fragmented embryos? A retrospective cohort study.

Objective: The aim was to clarify whether using testicular sperm reduces embryo fragmentation and improves cycle outcomes. Methods: Fragmented embryo was defined as an embryo in which fragments account for more than one third of the embryonic surface area. High rate of fragmented embryos was defined by a proportion of fragmented embryos higher than 50%. We recruited infertile couples who had undergone at least one ovarian stimulation cycle using ejaculated sperm but failed to conceive due to high rate of fragmented embryos in each previous cycle. After fully informed consent, the couples agreed to obtain testicular sperm by testicular puncture and use testicular sperm for intracytoplasmic sperm injection (ICSI). The normal fertilization rate, transferable embryo rate, fragmented embryo rate and cycle outcomes were compared between ejaculated sperm group (EJA-sperm group) and testicular sperm group (TESTI-sperm group). Results: Twenty-two couples who agreed to participate in our study underwent 32 ICSI cycles with ejaculated spermatozoa and 23 ICSI cycles with testicular spermatozoa. Embryo transfers were cancelled in 8 ejaculated cycles and 4 testicular cycles because of no transferable embryos. There were no significant differences in age, normal fertilization rate and high-quality embryo rate between ejaculated and testicular groups. The transferable embryo rate and implantation rate in TESTI-sperm group were significantly higher than those in EJA-sperm group (36.9% vs. 22.0%, p < 0.01; 34.2% vs. 0%, p < 0.001). The fragmented embryo rate in TESTI-sperm group was significantly lower than that in EJA-sperm group (61.2% vs. 75.7%, p < 0.05). Conclusion: Our small retrospective cohort study suggests that using testicular sperm may be a recommended option for couples with previous ART failure because of high rate of fragmented embryos. Large samples, multicenter studies or randomized controlled trial (RCT) are needed to further confirm the superiority of testicular sperm.

Acetate promotes SNAI1 expression by ACSS2-mediated histone acetylation under glucose limitation in renal cell carcinoma cell.

Metastasis is the main cause of cancer-associated deaths, yet this complex process is still not well understood. Many studies have shown that acetate is involved in cancer metastasis, but the molecular mechanisms remain to be elucidated. In the present study, we first measured the effect of acetate on zinc finger transcriptional repressor SNAI1 and acetyl-CoA synthetase 2 (ACSS2) under glucose limitation in renal cell carcinoma cell lines, 786-O and ACHN. Then, RNA interference and overexpression of ACSS2 were used to detect the role of acetate on SNAI1 expression and cell migration. Finally, chromatin immunoprecipitation assay (ChIP) was used to investigate the regulatory mechanism of acetate on SNAI1 expression. The results showed that acetate increased the expressions of SNAI1 and ACSS2 under glucose limitation. ACSS2 knockdown significantly decreased acetate-induced SNAI1 expression and cell migration, whereas overexpression of ACSS2 increased SNAI1 level and histone H3K27 acetylation (H3K27ac). ChIP results revealed that acetate increased H3K27ac levels in regulatory region of SNAI1, but did not increase ACSS2-binding ability. Our study identified a novel inducer, acetate, which can promote SNAI1 expression by ACSS2-mediated histone acetylation in partly. This finding has important implication in treatment of metastatic cancers.

Histone H3K9 demethylase JMJD1A is a co-activator of erythropoietin expression under hypoxia.

Erythropoietin (EPO) is a secreted hormone that stimulates the production of red blood cells, and the level of EPO is increased under hypoxia. The expression of EPO is regulated not only by the hypoxia-inducible factor (HIF) but also partly through epigenetic modifications, including histone acetylation and methylation. In this study, we report that histone H3K9 demethylase JMJD1 A is regulated by HIF-2α in HepG2 cells under hypoxia. Knockdown or over-expression of JMJD1 A can decrease or increase EPO expression, respectively. JMJD1 A can interact with HIF-2α to form a co-activator complex, which binds to the hypoxia response elements of EPO and increases EPO expression by catalyzing demethylation of H3K9me2, a transcription suppression marker. The results demonstrate that JMJD1 A is a co-activator of EPO expression.

Long noncoding RNA HOTAIR promotes metastasis of renal cell carcinoma by up-regulating histone H3K27 demethylase JMJD3.

Long Noncoding RNAs (lncRNAs) are a kind of non-protein coding transcripts longer than 200 nucleotides, and play important roles in diverse biological processes, such as embryonic development and apoptosis. Homeobox (HOX) transcript antisense intergenic RNA (HOTAIR) is a negative prognostic factor in a variety of human cancers, such as breast, liver and lung cancers. HOTAIR can promote cancer cell metastasis by reprogramming chromatin organization. In the present study, HOTAIR expression was elevated in tissues of renal cell carcinoma compared to adjacent normal tissues, and positively correlated with metastasis (P<0.05). The cell migration was inhibited in scratch test and transwell assay after HOTAIR knockdown (P<0.05). Further researches revealed that histone demethylase JMJD3 was reduced and its target gene Snai1 expression was down-regulated after HOTAIR suppression (P<0.05). Meanwhile, the level of histone methytransferase EZH2 target gene PCDHB5 was increased (P<0.05). Collectively, these data suggest that HOTAIR is an important promoter in metastasis of renal cell carcinoma and also plays a dual regulatory role in chromatin state by effecting both histone metylation and demethylation at different gene loci.