Topflash assay conditions were described previously

ependence. It suggested that HBs exposure was able to induce apoptosis in sperm cells. The caspase Effects of HBs on Sperm Functions can ultimately affect sperm functions or result in the death of sperm cells, leading to loss of sperm motility and reduced fertilizing ability. Oxidative stress caused DNA damage and sperm chromosome aberrations Oxidative stress as one of the major caused of sperm DNA damage has been widely adopted. This kind of damage is characterized by single and double DNA strand breaks. Although DNA damage 20008854 of the first type may be repaired by the oocyte or the embryo, fertilization of an oocyte by a spermatozoon with extensive double-stranded DNA fragmentation is virtually not repairable and incompatible with normal embryo and fetal development. The double-stranded DNA breaks may cause chromosomal aberrations. It was reported by our laboratory that the higher frequency and various types of chromosome aberrations were observed in sperm cells from the patients with HBV chronic infection. These events were probably caused by the imbalance between ROS production and TAC induced by HBs, leading to the accumulation of the unrepaired DNA damage and formation of sperm chromosome aberrations. Taken together, our data provided the solid evidence that HBs exposure could cause a series of deleterious events in sperm cells such as induction of ROS generation and lipid peroxidation, reduction of total antioxidant capacity, PS externalization, activation of caspases, and DNA fragmentation, resulting in increased apoptosis of sperm cells, the loss of sperm membrane integrity and sperm dysfunctions. Effects of HBs on Sperm Functions Materials and Methods Ethical approval After the informed consent approval was obtained, the human sperm samples were collected from the healthy male donors who were explicitly informed about the research aims, their rights and interests in the research. All “25849133 the protocols used in the present study were approved by Institutional Ethical Review Board of Shantou University Medical College, and conformed to the ethical guidelines of the 2008 Declaration of Helsinki as reflected in a prior approval by the institution’s human research committee. was separated, then the absorbance was measured at 530 nm and expressed as units per 16106/ml sperm cells for MDA. The above experiment was repeated five times. Assessment of antioxidant status in sperm cells FRAP assay was performed to measure the TAC of sperm cells in the test and control groups using a commercially available assay kit in a Multimode Microplate Reader according to the manufacturer’s instructions. PR 619 Briefly, the sperm cells were homogenized in 200 ml of phosphate buffer and sonicated over ice, and then centrifuged at 12,0006g at 4uC for 5 min to collect the supernatant for assay of TAC. The values were calculated using optical density at 593 nm and expressed as units per 16106/ml sperm cells for TAC. The above experiment was repeated five times. Preparations of human spermatozoa Human sperm samples were obtained by masturbation after 3 days of sexual abstinence from the healthy men. Semen samples were kept in a CO2 incubator for 30 min to allow liquefaction. Motile spermatozoa were selected by the swim-up method as follows: in each test tube, the 0.5 ml liquefied semen sample layered gently under 2 ml of biggers-whittem-whittingham medium containing 0.3% bovine serum albumin and incubated at 37uC in a 5% CO2 incubator for 1 h. The supernatant collected fr