Packaging of Mammalian Paternal Genome and Male Pronuclear Formation

在体内观察了大鼠精子核在附睾成熟和穿透卵母细胞过程中的dna稳定性和硫醇二硫状态。精细胞和精子经醋酸固定后用吖啶橙（AO）染色，共聚焦显微镜下红/绿荧光比例下降，直至附睾头。另一方面，用rnBBr标记精子后，精子从睾丸经附睾运输过程中荧光强度逐渐降低。在穿透透明带和精子去浓缩过程中，红/绿AO荧光比例增加。将卵泡周围空间的精子用mBBr标记后检测荧光。这些结果表明，在附睾精子成熟过程中，DNA抗酸的稳定性似乎早于鱼精蛋白的氧化，并且大鼠基因组中解包装过程的初始事件开始于或仅仅在带渗透之后，但在渗透到卵浆之前。采用荧光高效液相色谱法检测大鼠精子穿透和原核形成过程中卵细胞中谷胱甘肽和其他硫醇的含量。还原谷胱甘肽（GSH）含量在未受精卵母细胞和精子核去致密的穿透卵子中高于原核阶段的卵子。各试验组间氧化谷胱甘肽（GSSG）含量无显著差异。在受精过程中，GSSG/GSH的比值没有变化。虽然还原半胱氨酸含量在实验组之间没有变化，但在精子去浓缩和原核形成之间，半胱氨酸的氧化形式增加了。受精过程中检测到半胱氨酸含量较低，但各处理组均未检测到半胱氨酸和γ -谷氨酰半胱氨酸。这些结果表明，大鼠卵子中的谷胱甘肽含量在精子去浓缩和原核形成之间下降，可能是由于γ -谷氨酰转肽酶活性增加。少

DNA-stability and thiol-disulfide status of rat sperm nuclei was observed during maturation in the epididymis and during penetration to the oocytes in vivo. When spermatids and spermatozoa were stained with acridine orange (AO) after fixation with acetic alcohol, the ratio of red/green fluorescence under a confocal microscope decreased until caput epididymis. On the other hand, when spermatozoa were labeled with rnBBr the fluorescence intensities gradually decreased during transportation of spermatozoa from testis through epididymis. The ratio of red/green AO fluorescence increased during penetration of zona pellucida and sperm decondensation. When spermatozoa in the perivitelline space were labeled with mBBr and examined fluorescence was detected. These results demonstrate that DNA stability against acid appears to be ahead of the oxidization of protamine during sperm maturation in the epididymis and that an initial event of the unpackaging process in rat genome begins during or just … More after zona-penetration but before penetration into the ooplasm.The contents of glutathione and other thiols in rat eggs were examined during sperm penetration and pronuclear formation by high-performance liquid chromatography with fluorescence detection. Reduced glutathione (GSH) content was higher in unfertilized oocytes and penetrated eggs with a decondensed sperm nucleus than eggs at the pronuclear stage. The content of oxidized glutathione (GSSG) was not different among experimental groups. The ratio of GSSG/GSH was not different during fertilization. Although reduced cysteinylglycine content did not changed among experimental groups, oxidized form of cysteinylglycine increased between sperm decondensation and pronuclear formation. Low contents of cystine were detected during fertilization but cysteine and gamma-glutamylcysteine were not detected in any treatment groups. These results demonstrate that GSH content in rat eggs decreases between sperm decondensation and pronuclear formation, probably due to the increased activity of gamma-glutamyl transpeptidase. Less