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Leukemia inhibitory factor (LIF) plays a key role in the survivability of mouse embryos during pre-implantation. In this study, we verified the role of LIF by detecting gene expression in morula stage embryos through DNA microarray. Our results showed that LIF knockdown affected expression of 369 genes. After LIF supplementation, the epidermal growth factor (EGF) is most affected by LIF expression. To observe the correlation between LIF and EGF, the LIF knockdown embryos were supplemented with various growth factors, including LIF, EGF, GM-CSF, TGF, and IGF II. Only LIF and EGF caused the rate of blastocyst development to recover significantly from 52% of control to 83% and 93%, respectively. All of the variables, including the diameter of blastocysts, the number of blastomeres, and cells in ICM and TE, were almost restored. Moreover, EGF knockdown also impaired blastocyst development, which was reversed by LIF or EGF supplementation. The treatment with various signaling suppressors revealed that both EGF and LIF promoted embryonic development through the JAK/STAT3 signaling pathway. These data suggest that the EGF and LIF can be compensatory to each other during early embryonic development, and at least one of them is necessary for sustaining the normal development of pre-implantation embryos.

This study determined the incidence of sperm nuclear DNA fragmentation, mitochondrial dysfunction, and chromosomal aneuploidy. The results were correlated with the semen analysis parameters and fertilization rates.Semen samples from 10 men showing oligoasthenoteratozoospermia (OAT) and undergoing ICSI treatment were analyzed. Another semen samples from 10 men showing normozoospermia and undergoing IVF treatment were analyzed for comparison. The samples were prepared using a two-step discontinuous Percoll gradient (80%-50%) and analyzed using a Hamilton-Thorne Integrated Visual Optical System (IVOS) Sperm Analyzer. DNA fragmentation was detected with a terminal deoxynucleotidyl transferase-mediated dUTP nick end label (TUNEL) assay. Functional integrity of mitochondria was detected using an Apoalert Mitochondrial Membrane Sensor Kit. Chromosomal aneuploidy was assayed by fluorescence in situ hybridization.Higher sperm DNA fragmentation rate (18.8% vs. 2.8%), mitochondrial dysfunction rate (24.9% vs. 5.7%), and chromosomal aneuploidy rate (0.12% vs. 0.06%) were found in the oligoasthenoteratozoospermic patients in comparison with the normozoospermic patients.The result indicates that spermatozoa from oligoasthenoteratozoospermic patients contain greater DNA fragmentation, mitochondrial dysfunction, and chromosomal aneuploidy. Because extremely poor semen samples are the indication for ICSI treatment, the result indicates the importance of selecting good quality sperm for oocyte injection.

Objective To quantify the mitochondrial DNA contents in human embryos with good or poor quality at the 8-cell stage. Design Prospective study. Setting Private infertility clinic. Patient(s) Five women aged 24 to 34 years. Intervention(s) Embryos obtained in standard superovulation and embryo culture procedures. Main outcome measure(s) Mitochondrial DNA (mtDNA) copy numbers in human embryos at cleavage stage were quantified by real-time polymerase chain reaction, in an effort to correlate with morphology. Result(s) The grade 8A embryos contained a mean mtDNA copy number at 1,163,937 (n = 8, from three patients); grade 8B embryos, at 939,345 (n = 5, from two patients); grade 8C + embryos, at 637,872 (n = 12, from 5 patients); and grade 8C + embryos derived from 3PN zygotes, at 300,429 (n = 3, from a single patient). Conclusion(s) Great variations were found among blastomeres from a single embryo and among embryos from a single patient. The native variations of mtDNA copy number may affect developmental ability irrespective of morphology.

DNA amplification fingerprinting (DAF) and nuclear ribosomal DNA (nrDNA) amplification were used to discriminate between two laboratory colonies of two closely related species of weevils: the rice weevil, Sitophilus oryzae (L.), and the maize weevil, S. zeamais Motschulsky. For DAF, three sets of primers (aldolase, prolactin receptor, and interleukin-1 β ) were used for identification by polymerase chain reaction (PCR) and agarose gel electrophoresis, and the highly similar patterns of the resultant amplicons reconfirmed that the two weevils are closely related. The fragments of nrDNA amplification showed that for S. oryzae and S. zeamais , the homologies of the nucleotide sequences of the internal transcribed spacer regions, ITS-1 and ITS-2, were unusually high, at 96% and 97%, respectively. Based on the ITS-1 and ITS-2 sequences, two species-specific primer sets were designed: with the primer set ITS3/So , the predicted 450 bp DNA fragment was yielded with S. oryzae genomic DNA after PCR amplification ( n =10), but no PCR product was obtained with S. zeamais ( n =10); with the primer set ITS3/Sz , the same 10 S. zeamais specimens yielded a 550 bp DNA fragment, but S. oryzae yielded no amplicons. In view of the difficulty of distinguishing between these two closely related species, the specificity and availability of these two primer sets might prove to be a useful tool for distinguishing between them. However, the nrDNA sequences of the ITS-1 and ITS-2 regions of geographically isolated populations of both weevils still need to be elucidated, and the applicability of this technique to different geographical populations will need to be confirmed by further study.