OBJECTIVETo study the oxidative damage to human sperm mitochondrial tRNA LeuUUR gene by reactive oxygen species (ROS) in vitro.

METHODSSpermatozoa of normal physiological function selected from semen samples by Percoll gradient centrifugation technique were used as normal sperm models, which were divided into two groups of 20 cases each, a damage group and a control group, the former treated with hypoxanthine xanthine oxidase system and the latter left untreated, both incubated at 37 degrees C in aerobic environment for 60 minutes. Sperm DAN was extracted, and digestion by the enzymes fpg and ligation-mediated PCR ( LM-PCR) was performed to map the damage to mitochondrial tRNA LeuUUR gene. The spermatozoa were labeled with specific fluorescent probe of Rhodamine 123 to measure mitochondrial membrane potential ( MMP) by flow cytometry and observe sperm function.

RESULTSCompared with the control group, after the normal spermatozoa were incubated with ROS, MMP of the spermatozoa significantly decreased ( 116. 27+/-11.72 vs 64.00+/-4. 88) , P <0.05. Digestion by the enzymes fpg and LM-PCR showed damage to mitochondrial tRNA LeuUUR, gene.

CONCLUSIONReactive oxygen species may inflict oxidative damage on sperm mitochondrial tRNA LeuUUR gene and thus affect sperm function ( as shown by significant decrease in MMP), resulting in infertility.