A panel of extracellular vesicle long noncoding RNAs in seminal plasma for predicting testicular spermatozoa in nonobstructive azoospermia patients.

study questionWhether the testis-specific extracellular vesicle (EV) long noncoding RNAs (lncRNAs) in seminal plasma could be utilized to predict the presence of testicular spermatozoa in nonobstructive azoospermia (NOA) patients? SUMMARY ANSWER: Our findings indicate that the panel based on seminal plasma EV lncRNAs was a sensitive and specific method in predicting the presence of testicular spermatozoa and may improve clinical decision-making of NOA. WHAT IS KNOWN ALREADY: The adoption of sperm retrieval techniques, especially microdissection testicular sperm extraction (mTESE), in combination with ICSI has revolutionized treatment for NOA. However, there are no precise and noninvasive methods for predicting whether there are testicular spermatozoa in NOA patients before mTESE. STUDY DESIGN, SIZE, DURATION: RNA sequencing was performed on seminal plasma EVs from 6 normozoospermic men who underwent IVF due to female factor and 5 idiopathic NOA patients who failed to obtain testicular spermatozoa by mTESE and were diagnosed as having Sertoli cell-only syndrome by postoperative pathology. A biomarker panel of lncRNAs was constructed and verified in 96 NOA patients who underwent mTESE. Decision-making process was established based on the panel in seminal plasma EVs from 45 normozoospermia samples, 43 oligozoospermia samples, 62 cryptozoospermia samples, 96 NOA samples. PARTICIPANTS/MATERIALS, SETTING,

methodsRNA sequencing was done to examine altered profiles of EV lncRNAs in seminal plasma. Furthermore, a panel consisting of EV lncRNAs was established and evaluated in training set and validation sets. MAIN RESULTS AND THE ROLE OF CHANCE: A panel consisting of nine differentially expressed testis-specific lncRNAs, including LOC100505685, SPATA42, CCDC37-DT, GABRG3-AS1, LOC440934, LOC101929088 (XR_927561.2), LOC101929088 (XR_001745218.1), LINC00343 and LINC00301, was established in the training set and the AUC was 0.986. Furthermore, the AUC in the validation set was 0.960. Importantly, the panel had a unique advantage when compared with models based on serum hormones from the same group of NOA cases (AUC, 0.970 vs 0.723; 0.959 vs 0.687, respectively). According to the panel of lncRNAs, a decision-making process was established, that is when the score of an NOA case exceeds 0.532, sperm retrieval surgery may be recommended. LIMITATIONS, REASONS FOR CAUTION: In the future, the sample size needs to be further expanded. Meanwhile, the regulatory functions and mechanism of lncRNAs in spermatogenesis also need to be elucidated. WIDER IMPLICATIONS OF THE

findingsWhen the score of our panel is below 0.532, subjecting the NOA patients to ineffective surgical interventions may not be recommended due to poor sperm retrieval rate. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (81871110, 81971314 and 81971759); the Guangdong Special Support Plan-Science and Technology Innovation Youth Top Talents Project (2016TQ03R444); the Science and Technology Planning Project of Guangdong Province (2016B030230001 and 201707010394); the Key Scientific and Technological Program of Guangzhou City (201604020189); the Pearl River S&T Nova Program of Guangzhou (201806010089); the Transformation of Scientific and Technological Achievements Project of Sun Yat-sen University (80000-18843235) and the Youth Teacher Training Project of Sun Yat-sen University (17ykpy68 and 18ykpy09). There are no competing interests related to this study. TRIAL REGISTRATION NUMBER: N/A.