Male germline-specific thioredoxin domain containing 8 (TXNDC8; alias SPTRX3) accumulates indefective human spermatozoa. We assessed the efficiency of two-step semen purification inremoving spermatozoa carrying TXNDC8, and examined the relationship of TXNDC8 with theoutcomes of assisted reproductive therapy (ART), conventional semen parameters, and sperm DNA integrity in sperm chromatin structure assay (SCSA). Semen samples (n = 255) from 91 ART couples were screened in two independent trials, both including a two-step, gradient-and-swim-up separation procedure yielding A-samples (raw semen), B-samples (gradient separated), and C-samples (gradient-and-swim-up). The C-samples were used for intracytoplasmic sperm injection (ICSI) with morphologically selected spermatozoa (IMSSI). Percentage of TXNDC8-positive spermatozoaincreased progressively from A to B/C-samples in both trials. In the first trial (35 couples), the TXNDC8 correlated positively with sperm DNA fragmentation index (%DFI; r = 0.66) measured before separation, and negatively with sperm concentration (r = −0.57) and motility (r = −0.67), also taken before separation. The high DNA stainability index (%HDS) correlated with the percentage of spermatozoa lacking TXNDC8 (r = 0.68). Both SCSA and TXNDC8 parameters showed moderate correlations (r = 0.33–0.66) with blood serum levels of hCG on day 11 (Beta 1) and day13 (Beta 2) after oocyte retrieval. In the second trial (56 couples), fathers of multiplets had a significantly lower percentage of TXNDC8-positive spermatozoa in B-sample (gradient separationonly) compared to men who conceived a singleton pregnancy (p = 0.01) and those who produced no pregnancy (p = 0.02). Those multiplets’ fathers also had a significantly higher sperm concentration while their SCSA parameters did not differ from others. It is concluded that theTXNDC8 levels correlate with SCSA and conventional raw semen parameters, and are predictive of pregnancy outcome and multiple births after ART. Two-step purification does not efficiently remove TXNDC8 carrying spermatozoa.

Abbreviations

ART– assisted reproductive therapy; DFI- DNA fragmentation index; FC- flow cytometry (FC); hCG: human chorionic gonadotropin; HDS: high DNA stainability index; HEPES- (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid); HTF- human tubal fluid; ICSI- intracytoplasmic sperm injection; IgG- immunoglobulin G; IMSSI- ICSI with morphologically selected spermatozoa; IVF- in vitro fertilization; IU-: intrauterine insemination; NGS- normal goat serum; PBS- phosphate buffered saline; PVP- polyvinylpyrrolidone; SAB- spontaneous abortion; SCSA- sperm chromatin structure assay; SPTRX3- spermatid specific thioredoxin 3; SSS- synthetic serum substitute; TRITC- tetramethyl rhodamine isothiocyanate; TX-100- Triton X-100; TXNDC- thioredoxin domain-containing proteins; TXNDC8- thioredoxin domain containing 8; TUNEL- Terminal deoxynucleotidyl transferase dUTP nick end labeling

中文翻译：

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TXNDC8 的精子含量反映了精子染色质结构、妊娠建立和 ART 后多胞胎的发生率。

含有 8 个（TXNDC8；别名 SPTRX3）的雄性种系特异性硫氧还蛋白结构域会积累有缺陷的人类精子。我们评估了两步精液纯化去除携带 TXNDC8 的精子的效率，并检查了 TXNDC8 与辅助生殖治疗 (ART) 结果、常规精液参数和精子染色质结构测定 (SCSA) 中精子 DNA 完整性的关系。来自 91 对 ART 夫妇的精液样本（n = 255）在两个独立的试验中进行筛选，这两个试验都包括一个两步、梯度和向上分离程序，产生 A 样本（原始精液）、B 样本（梯度分离） , 和 C-samples (gradient-and-swim-up)。C 样本用于胞浆内单精子注射 (ICSI) 和形态学选择的精子 (IMSSI)。在两项试验中，TXNDC8 阳性精子的百分比从 A 到 B/C 样本逐渐增加。在第一次试验（35 对夫妇）中，TXNDC8 与分离前测量的精子 DNA 碎片指数（%DFI；r = 0.66）呈正相关，与精子浓度（r = -0.57）和运动性（r = -0.67）呈负相关，分离前也采取。高 DNA 染色指数 (%HDS) 与缺乏 TXNDC8 的精子百分比相关 (r = 0.68)。SCSA 和 TXNDC8 参数与取卵后第 11 天（Beta 1）和第 13 天（Beta 2）的血清 hCG 水平均显示出中等相关性（r = 0.33–0.66）。在二审（56对夫妇）中，与单胎妊娠的男性 (p = 0.01) 和未怀孕的男性 (p = 0.02) 相比，多胞胎的父亲在 B 样本（仅梯度分离）中具有显着较低的 TXNDC8 阳性精子百分比。这些多胞胎的父亲也有明显更高的精子浓度，而他们的 SCSA 参数与其他人没有区别。结论是 TXNDC8 水平与 SCSA 和传统的原始精液参数相关，并且可以预测 ART 后的妊娠结果和多胞胎。两步纯化不能有效去除携带精子的 TXNDC8。结论是 TXNDC8 水平与 SCSA 和传统的原始精液参数相关，并且可以预测 ART 后的妊娠结果和多胞胎。两步纯化不能有效去除携带精子的 TXNDC8。结论是 TXNDC8 水平与 SCSA 和传统的原始精液参数相关，并且可以预测 ART 后的妊娠结果和多胞胎。两步纯化不能有效去除携带精子的 TXNDC8。

缩写

ART——辅助生殖治疗；DFI-DNA片段化指数；FC-流式细胞术（FC）；hCG：人绒毛膜促性腺激素；HDS：高DNA染色指数；HEPES-(4-(2-羟乙基)-1-哌嗪乙磺酸)；HTF-人输卵管液；ICSI-卵胞浆内单精子注射；IgG-免疫球蛋白G；IMSSI-ICSI 与形态学选择的精子；IVF-体外施肥；IU-：宫内人工授精；NGS-正常山羊血清；PBS-磷酸盐缓冲盐水；PVP-聚乙烯吡咯烷酮；SAB-自然流产；SCSA-精子染色质结构测定；SPTRX3-精细胞特异性硫氧还蛋白3；SSS-合成血清替代品；TRITC-四甲基罗丹明异硫氰酸酯；TX-100-海卫X-100；TXNDC-含硫氧还蛋白结构域的蛋白质；TXNDC8-含8个硫氧还蛋白结构域；TUNEL-末端脱氧核苷酸转移酶dUTP缺口末端标记