Sperm DNA damage is recognized as an important biomarker of male infertility. To investigate this, sperm DNA damage was assessed by the sperm chromatin dispersion (SCD) test in semen and motile spermatozoa harvested by combined density gradient centrifugation (DGC) and swim-up in 161 couples undergoing in vitro fertilization (IVF). Semen analysis and sperm DNA damage results were compared between couples who did or did not achieve pregnancy. The sperm DNA damage level was significantly different between the two groups (P < 0.05) and was negatively correlated with IVF outcomes. Logistic regression analysis confirmed that it was an independent predictor for achieving clinical pregnancy. The effects of different levels of sperm DNA damage on IVF outcomes were also compared. There were significant differences in day 3 embryo quality, blastocyst formation rate, and implantation and pregnancy rates (P < 0.05), but not in the basic fertilization rate between the two groups. Thus, sperm DNA damage as measured by the SCD appears useful for predicting the clinical pregnancy rate following IVF.
Adult ; Chromatin/chemistry* ; DNA Damage ; Embryo Implantation ; Embryonic Development ; Female ; Fertilization in Vitro ; Humans ; Male ; Predictive Value of Tests ; Pregnancy ; Pregnancy Outcome ; Pregnancy Rate ; Semen Analysis ; Sperm Injections, Intracytoplasmic ; Sperm Motility ; Spermatozoa/ultrastructure*

OBJECTIVETo investigate the influence of the DNA integrity of optimized sperm on the embryonic development and clinical outcomes of in vitro fertilization and embryo transfer (IVF-ET).

METHODSThis study included 605 cycles of conventional IVF-ET for pure oviductal infertility performed from January 1, 2013 to December 31, 2014. On the day of retrieval, we examined the DNA integrity of the sperm using the sperm chromatin dispersion method. According to the ROC curve and Youden index, we grouped the cycles based on the sperm DNA fragmentation index (DFI) threshold value for predicting implantation failure, early miscarriage, and fertilization failure, followed by analysis of the correlation between DFI and the outcomes of IVF-ET.

RESULTSAccording to the DFI threshold values obtained, the 605 cycles fell into four groups (DFI value < 5%, 5-10%, 10-15%, and ≥ 15%). Statistically significant differences were observed among the four groups in the rates of fertilization, cleavage, high-quality embryo, implantation, clinical pregnancy, early miscarriage, and live birth (P < 0.05), but not in the rates of multiple pregnancy, premature birth, and low birth weight (P > 0.05). DFI was found to be correlated negatively with the rates of fertilization (r = -0.32, P < 0.01), cleavage (r = -0.19, P < 0.01), high-quality embryo (r = -0.40, P < 0.01), clinical pregnancy (r = -0.20, P < 0.01), and live birth (r = -0.09 P = 0.04), positively with the rate of early miscarriage (r = 0.23, P < 0.01), but not with the rates of multiple pregnancy (r = -0.01, P = 0.83), premature birth (r = 0.04, P = 0.54), and low birth weight (r = 0.03, P = 0.62).

CONCLUSIONThe DNA integrity of optimized sperm influences fertilization, embryonic development, early miscarriage, and live birth of IVF-ET, but its correlation with premature birth and low birth weight has to be further studied.

Abortion, Spontaneous ; Chromatin ; ultrastructure ; DNA Fragmentation ; Embryo Implantation ; Embryo Transfer ; Embryonic Development ; Female ; Fertilization ; Fertilization in Vitro ; Humans ; Infertility, Female ; Male ; Pregnancy ; Pregnancy Outcome ; ROC Curve ; Spermatozoa ; cytology

Axonal transport of mitochondria is critical for neuronal survival and function. Automatically quantifying and analyzing mitochondrial movement in a large quantity remain challenging. Here, we report an efficient method for imaging and quantifying axonal mitochondrial transport using microfluidic-chamber-cultured neurons together with a newly developed analysis package named "MitoQuant". This tool-kit consists of an automated program for tracking mitochondrial movement inside live neuronal axons and a transient-velocity analysis program for analyzing dynamic movement patterns of mitochondria. Using this method, we examined axonal mitochondrial movement both in cultured mammalian neurons and in motor neuron axons of Drosophila in vivo. In 3 different paradigms (temperature changes, drug treatment and genetic manipulation) that affect mitochondria, we have shown that this new method is highly efficient and sensitive for detecting changes in mitochondrial movement. The method significantly enhanced our ability to quantitatively analyze axonal mitochondrial movement and allowed us to detect dynamic changes in axonal mitochondrial transport that were not detected by traditional kymographic analyses.
Animals ; Axonal Transport ; physiology ; Cerebral Cortex ; cytology ; metabolism ; Drosophila melanogaster ; cytology ; metabolism ; Embryo, Mammalian ; Gene Expression ; Lab-On-A-Chip Devices ; Microscopy, Confocal ; Mitochondria ; metabolism ; ultrastructure ; Motor Neurons ; metabolism ; ultrastructure ; Movement ; Mutation ; Primary Cell Culture ; RNA-Binding Protein FUS ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Software

The aim of this study was to investigate the effect of Bu-Shen-An-Tai recipe (BSATR) and its two components (Bushen recipe, and Huoxue recipe) on endometrial morphology during peri-implantation in superovulated mice. Mice were randomly divided into five groups, including the normal (N), model (M), Bushen (BS), Huoxue (HX) and Bu-Shen-An-Tai (BH) groups. The uteri were collected on day 4 of pregnancy, and the endometrium thickness, microvessel density (MVD) and number of pinopodes observed. Compared with the M group, the endometrial thickness in the BS, HX and BH groups was significantly increased and there was a significant difference in endometrial thickness between the BS and the BH groups. The mean MVD was significantly lower in the M group than in the N group, and there was a significant increase in MVD in the BS, HX and BH groups as compared with the M group. Compared with the M group, the pinopode scores in the endometrium were significantly increased in the HX and BH groups; and the BS group had significantly higher pinipode scores than the HX and BH groups. In conclusion, the results of the present study demonstrated that the recipes (Bushen, Huoxue and BSATR) could improve the endometrial environment by regulating the endometrial thickness, MVD and the number of pinopodes at the window of implantation. Moreover, the Huoxue recipe and the BSATR were more efficient than the Bushen recipe, with the BSATR tending to have the most beneficial effects.
Animals ; Antigens, CD34 ; metabolism ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Embryo Implantation ; physiology ; Endometrium ; drug effects ; physiology ; ultrastructure ; Female ; Immunohistochemistry ; Mice ; Microscopy, Electron, Scanning ; Microvessels ; metabolism ; physiology ; Ovulation ; physiology ; Pregnancy ; Random Allocation ; Time Factors

This study was conducted to establish an in vitro maturation (IVM) system by selection of efficient porcine serum during porcine in vitro production. To investigate the efficient porcine serum (PS), different types of PS [newborn pig serum, prepubertal gilt serum (PGS), estrus sow serum, and pregnancy sow serum] were used to supplement IVM media with or without gonadotrophin (GTH) and development rates of parthenogenetic activation (PA) and in vitro fertilization (IVF) embryos were then compared. The maturation rates of the PGS group was significantly higher when GTH was not added. Additionally, during development of PA embryos without GTH, the PGS group showed significantly higher cleavage and blastocyst formation rates. Moreover, the cleavage rates of IVF embryos were significantly higher in the PGS group, with no significant differences in the blastocyst formation. However, when GTH was supplemented into the IVM media, there were no significant differences among the four groups in the cleavage rates, development rates of the blastocyst, and cell number of the blastocyst after PA and IVF. In conclusion, PGS is an efficient macromolecule in porcine IVM, and GTH supplementation of the IVM media is beneficial when PS is used as macromolecule, regardless of its origin.
Animals ; Blastocyst/*drug effects ; Embryo, Mammalian/drug effects/*embryology/physiology/ultrastructure ; Fertilization in Vitro/veterinary ; Gonadotropins/administration & dosage/*metabolism ; In Vitro Oocyte Maturation Techniques/*methods/veterinary ; Parthenogenesis/*drug effects ; Sus scrofa/*embryology

Asoprisnil, a member of the selective progesterone receptor modulators, exerts high progesterone receptor selectivity, endometrial targeted advantages and significant anti-implantation effect in rats. The purpose of this study was to confirm the anti-implantation effect of asoprisil, investigate the ultrastructural changes of the peri-implantation endometrium in mice and explore the effect of asoprisnil on endometrial receptivity and its targeted contraceptive proficiency. Post-coitus mice were administered with different dosages (0.2, 0.1, 0.05 mg·g(-1)·day(-1)) of asoprisnil from day 1 of pregnancy to day 3. Then 3 animals in each group were killed on day 5 of pregnancy, and uteri were collected to examine the ultrastructural changes of endometria under a transmission electron microscope (TEM). A total of 80 animals were sacrificed on day 8 of pregnancy, and the uterine horns were examined for the presence or absence of nidation sites and the number of implantation embryos. The results showed that the implantation rate and the average number of implantation embryos in asoprisnil groups were statistically significantly decreased as compared with the vehicle control group (P<0.05). The TEM results revealed that, in vehicle control group, the tight junction between the luminal epithelia cells was short and straight, the gap was wide; the luminal epithelia cells were covered with plenty of short, clavate and neatly arranged microvilli; the endometril stromal cells were large with plenty of cytoplasm, and showed significant decidual change; there was more than one nucleus in stromal cells, and the karyotheca was integrity. In low dosage and high dosage asoprisnil groups, the tight junction was longer and more curve than in the vehicle control group; microvilli were uneven and asymmetrically distributed in luminal epithelia; the stromal cells were small and the decidual change was not significant; there were karyopyknosis and karyolysis in stromal cells; there were abnormal thick-wall vessels in the endometrium. It was suggested that asoprisnil changed the ultrastructure of the endometrium in implantation window, disturbed the endometrial receptivity and finally resulted in embryo implantation failure.
Animals ; Contraception, Postcoital ; methods ; Embryo Implantation, Delayed ; drug effects ; physiology ; Endometrium ; drug effects ; physiology ; ultrastructure ; Estrenes ; administration & dosage ; Female ; Mice ; Oximes ; administration & dosage ; Oxytocics ; administration & dosage ; Pregnancy ; Pregnancy, Animal ; Treatment Outcome

The integrity of blood vessels controls vascular permeability and extravasation of blood cells, across the endothelium. Thus, the impairment of endothelial integrity leads to hemorrhage, edema, and inflammatory infiltration. However, the molecular mechanism underlying vascular integrity has not been fully understood. Here, we demonstrate an essential role for A-kinase anchoring protein 12 (AKAP12) in the maintenance of endothelial integrity during vascular development. Zebrafish embryos depleted of akap12 (akap12 morphants) exhibited severe hemorrhages. In vivo time-lapse analyses suggested that disorganized interendothelial cell-cell adhesions in akap12 morphants might be the cause of hemorrhage. To clarify the molecular mechanism by which the cell-cell adhesions are impaired, we examined the cell-cell adhesion molecules and their regulators using cultured endothelial cells. The expression of PAK2, an actin cytoskeletal regulator, and AF6, a connector of intercellular adhesion molecules and actin cytoskeleton, was reduced in AKAP12-depleted cells. Depletion of either PAK2 or AF6 phenocopied AKAP12-depleted cells, suggesting the reduction of PAK2 and AF6 results in the loosening of intercellular junctions. Consistent with this, overexpression of PAK2 and AF6 rescued the abnormal hemorrhage in akap12 morphants. We conclude that AKAP12 is essential for integrity of endothelium by maintaining the expression of PAK2 and AF6 during vascular development.
A Kinase Anchor Proteins/*genetics/metabolism ; Animals ; Blood Vessels/abnormalities/*embryology/metabolism ; Cell Cycle Proteins/genetics/metabolism ; Down-Regulation ; Embryo, Nonmammalian/abnormalities/*blood supply/embryology/metabolism ; Gene Deletion ; *Gene Expression Regulation, Developmental ; Hemorrhage/*embryology/genetics/metabolism ; Human Umbilical Vein Endothelial Cells ; Humans ; Intercellular Junctions/genetics/metabolism/ultrastructure ; Kinesin/genetics/metabolism ; Myosins/genetics/metabolism ; Zebrafish/*embryology/genetics ; p21-Activated Kinases/genetics/metabolism

OBJECTIVETo evaluate the value of endometrial microvessel density (MVD) in assessing the endometrial receptivity during the peri-implantation period.

METHODSA total of 104 patients undergoing in vitro fertilization and embryo transfer (IVF-ET) treatment were analyzed retrospectively. The subjects were divided into clinical pregnancy group (50 cases) and nonpregnant group (54 cases) according to the IVF-ET outcome. Endometrial tissues were collected 7 days after the natural ovulation prior to IVF-ET for measurement of the endometrial MVD using electron microscopy, which was analyzed in relation to the clinical outcome of the treatment.

RESULTSThe endometrial MVD was significantly higher in the clinical pregnancy group than in the nonpregnant group [(4.12∓1.84)% vs (3.46∓1.26)%, t=-2.127, P=0.036). ROC curve analysis showed that the MVD had an area under the curve slightly over 0.5 (0.598) for predicting clinical pregnancy, suggesting a poor specificity in predicting the clinical outcome of the treatment.

CONCLUSIONIn IVF-ET cycles, the endometrial MVD during the peri-implantation period is helpful for assessing the endometrial receptivity, but the specificity remains low.

Adult ; Embryo Implantation ; physiology ; Embryo Transfer ; Endometrium ; blood supply ; physiology ; Female ; Fertilization in Vitro ; Humans ; Infertility, Female ; diagnostic imaging ; physiopathology ; therapy ; Microvessels ; ultrastructure ; Retrospective Studies ; Ultrasonography

The plaque-forming characteristics of Newcastle disease viruses of chickens and geese source were compared on various cells. The result showed that there were obvious differences of plaque formation between F48E9 and NA-1 on Vero cells, chicken embryo fibroblast cells (CEF) and goose embryo fibroblast cells (GEF). The plaque-forming ability of NA-1 was higher than F48E9 on GEF, but lower than F48E9 on CEF. On Vero cells, the plaque-forming ability of NA-1 was slightly stronger than F48E9. It demonstrated that the plaque-forming characteristics were consistent with host tropism of virus. The morphogenesis of F48E9 and NA-1 on Vero cells was observed with transmission electron microscope. There were different replication processes between F48E9 and NA-1 on cells at different stages. NA-1 had stronger adaptability to host than F48E9 according to budding processes and envelope integrity.
Animals ; Cercopithecus aethiops ; Chick Embryo ; Chickens ; Geese ; Host-Pathogen Interactions ; Newcastle Disease ; virology ; Newcastle disease virus ; growth & development ; isolation & purification ; physiology ; ultrastructure ; Poultry Diseases ; virology ; Vero Cells ; Viral Plaque Assay

Cell apoptosis induced by duck reovirus (DRV)in duck embryo fibroblasts (DEF) was ascertained by light microscope and electron microscopy, DNA Ladder, flow cytometry and fluorescent microscopy. Typical morphological apoptotic features including cell shrinkage and condensation, margination of nuclear chromatin were observed under light microscope and the formation of apoptotic bodies by electron microscopy. DNA ladder was shown by DNA fragment analysis at 24-144h post infection. Flow cytometry showed that the cell apoptosis appeared at 24h and reached it's crest-time at 72-96h, decreased at 144h. Fluorescent microscopy showed that the apoptotic cells which showed green fluorescence appeared at 24h, the number of dead cells which showed red fluorescence increased with the time went by. The results above confirmed that the apoptosis of DEF was successfully induced by DRV.
Animals ; Apoptosis ; physiology ; Cell Nucleus ; ultrastructure ; Cells, Cultured ; DNA Fragmentation ; Ducks ; Embryo, Nonmammalian ; cytology ; Fibroblasts ; cytology ; ultrastructure ; virology ; Flow Cytometry ; Host-Pathogen Interactions ; Microscopy, Electron, Transmission ; Reoviridae ; physiology