Non-obstructive Azoospermia Patients Research Articles

Parameters affecting the success rate of microscopic testicular sperm extraction in male patients with a solitary testis and non-obstructive azoospermia.

We aimed to compare the success rate of spermatozoa retrieval through microscopic testicular sperm extraction (mTESE) in non-obstructive azoospermic (NOA) men with a solitary testis with that of mTESE in NOA men with bilateral testes and the parameters affecting these rates. A retrospective cross-sectional study of factors contributing to infertility in NOA patients with a solitary testis and men with bilateral testes was carried out. In this multicenter study, 74 patients with NOA with a solitary testis were matched with 74 patients with bilateral testes in terms of age, duration of infertility, and volume of the solitary testis from 2770 patients with NOA with bilateral testes. Hormonal parameters, presence of varicocele, history of varicocelectomy, history of undescended testis and karyotype analysis results were compared. Spermatozoa were obtained from 40 (54.1%) patients with a solitary testis and 42 (56.76%) patients with bilateral testes. No differences were found regarding age, duration of infertility, or mean testicular volume between patients with a solitary testis and patients with bilateral testes. When serum follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels were compared regardless of sperm retrieval status, it was observed that both levels were greater in the group of patients with a solitary testis (p < 0.01). Patients with solitary and bilateral testes from whom spermatozoa were obtained had larger testes than those from whom spermatozoa could not be obtained (p < 0.05). Similarly, the serum levels of FSH and LH were significantly greater in patients with a solitary testis than in those with bilateral testes (p < 0.05). To the best of our knowledge, this is the first study in the literature to evaluate the parameters that influence mTESE outcome in NOA patients with a solitary testis and NOA patients with bilateral testes. Greater testicular volume was found to positively affect spermatozoa retrieval for patients with a solitary testis. The higher levels of FSH and LH in patients with a solitary testis than in patients with bilateral testes of similar testicular volume may be due to a compensatory mechanism developed by the hypothalamic-pituitary-gonadal axis. The fact that these hormones are higher in patients with a solitary testis does not mean that the number of spermatozoa obtained through mTESE will be decreased.

Testicular histopathology and follicular stimulating hormone to predict fertility in nonobstructive azoospermia.

To examine the ability of testicular histopathology in Non-obstructive azoospermia (NOA) in predicting sperm retrieval rate (SR), sperm quality and assisted reproductive technology success. A retrospective study recruited clinically diagnosed NOA patients between 2007 and 2015. Testicular biopsy and conventional sperm extraction (TESE) were done concomitantly. Correlation between pathological categories, SR rate, sperm quality and success of intracytoplasmic sperm injection (ICSI) was studied. FSH was measured as a predictor of fertility. One hundred eighteen patients were recruited. Histopathological classification was hypospermatogenesis (HS) 45 (38%), maturation arrest (MA) 22(19%), Sertoli cell only syndrome (SCOS) 34 (29%) and normal spermatogenesis (NS) 17 (14%). FSH value was above normal level in 34 (76%) of HS, 19 (86%) of MA, 32 (94%) of SCOS and 5 (29%) of NS. Positive SR was obtained in 108 (92%) patients. The highest SR rate was seen in NS group 100% and the lowest was in SCOS 26 (77%). The worst sperm quality was found in SCOS as type C represents 46%, followed by MA 40% and HS 24%. Patients had ICSI following TESE had variable success rate as success of ICSI was seen (9/15) for HS, (0/7) for MA, (5/15) for SCOS and (8/9) for NS. FSH is strongly correlated to SR, quality of sperm and success of ICSI as positive SR in normal FSH patients was obtained in 28 (100%) of normal FSH, 70 (97%) of high FSH and 10 (56%) of double high FSH (p value < 001). The success of ICSI significantly correlates with FSH value as normal FSH has 77% success ICSI rate, high FSH (52%) and double high FSH (0%) (p value < 0.001). Testicular biopsy and histopathology findings in NOA are strongly correlated SR rate, quality of sperms, and success of ICSI. FSH is a strong noninvasive predictor of fertility in NOA patients.

Proteomic biomarkers in seminal plasma as predictors of reproductive potential in azoospermic men.

Azoospermia, characterized by an absence of sperm in the ejaculate, represents the most severe form of male infertility. While surgical sperm retrieval in obstructive azoospermia (OA) is successful in the majority of cases, patients with non-obstructive azoospermia (NOA) show retrieval rates of only about 50% and thus frequently have unnecessary surgery. Surgical intervention could be avoided if patients without preserved spermatogenesis are identified preoperatively. This prospective study aimed to discover biomarkers in seminal plasma that could be employed for a non-invasive differential diagnosis of OA/NOA in order to rationalize surgery recommendations and improve success rates. All patients signed written informed consent, underwent comprehensive andrological evaluation, received human genetics to exclude relevant pathologies, and patients with azoospermia underwent surgical sperm retrieval. Using label-free LC-MS/MS, we compared the proteomes of seminal plasma samples from fertile men (healthy controls (HC), n=8) and infertile men diagnosed with 1) OA (n=7), 2) NOA with successful sperm retrieval (mixed testicular atrophy (MTA), n=8), and 3) NOA without sperm retrieval (Sertoli cell-only phenotype (SCO), n=7). Relative abundance changes of two candidate markers of sperm retrieval, HSPA2 and LDHC, were confirmed by Western Blot. We found the protein expression levels of 42 proteins to be significantly down-regulated (p ≤ 0.05) in seminal plasma from SCO NOA patients relative to HC whereas only one protein was down-regulated in seminal plasma from MTA patients. Analysis of tissue and cell expression suggested that the testis-specific proteins LDHC, PGK2, DPEP3, and germ-cell enriched heat-shock proteins HSPA2 and HSPA4L are promising biomarkers of spermatogenic function. Western blotting revealed a significantly lower abundance of LDHC and HSPA2 in the seminal plasma of men with NOA (SCO and MTA) compared to controls. The results indicate that certain testis-specific proteins when measured in seminal plasma, could serve as indicators of the presence of sperm in the testis and predict the success of sperm retrieval. Used in conjunction with conventional clinical assessments, these proteomic biomarkers may assist in the non-invasive diagnosis of idiopathic male infertility.

Systematic molecular analyses for 115 karyotypically normal men with isolated non-obstructive azoospermia.

Do copy-number variations (CNVs) in the azoospermia factor (AZF) regions and monogenic mutations play a major role in the development of isolated (non-syndromic) non-obstructive azoospermia (NOA) in Japanese men with a normal 46, XY karyotype? Deleterious CNVs in the AZF regions and damaging sequence variants in eight genes likely constitute at least 8% and approximately 8% of the genetic causes, respectively, while variants in other genes play only a minor role. Sex chromosomal abnormalities, AZF-linked microdeletions, and monogenic mutations have been implicated in isolated NOA. More than 160 genes have been reported as causative/susceptibility/candidate genes for NOA. Systematic molecular analyses were conducted for 115 patients with isolated NOA and a normal 46, XY karyotype, who visited our hospital between 2017 and 2021. We studied 115 unrelated Japanese patients. AZF-linked CNVs were examined using sequence-tagged PCR and multiplex ligation-dependent probe amplification, and nucleotide variants were screened using whole exome sequencing (WES). An optimized sequence kernel association test (SKAT-O), a gene-based association study using WES data, was performed to identify novel disease-associated genes in the genome. The results were compared to those of previous studies and our in-house control data. Thirteen types of AZF-linked CNVs, including the hitherto unreported gr/gr triplication and partial AZFb deletion, were identified in 63 (54.8%) cases. When the gr/gr deletion, a common polymorphism in Japan, was excluded from data analyses, the total frequency of CNVs was 23/75 (30.7%). This frequency is higher than that of the reference data in Japan and China (11.1% and 14.7%, respectively). Known NOA-causative AZF-linked CNVs were found in nine (7.8%) cases. Rare damaging variants in known causative genes (DMRT1, PLK4, SYCP2, TEX11, and USP26) and hemizygous/multiple-heterozygous damaging variants in known spermatogenesis-associated genes (TAF7L, DNAH2, and DNAH17) were identified in nine cases (7.8% in total). Some patients carried rare damaging variants in multiple genes. SKAT-O detected no genes whose rare damaging variants were significantly accumulated in the patient group. The number of participants was relatively small, and the clinical information of each patient was fragmentary. Moreover, the pathogenicity of identified variants was assessed only by in silico analyses. This study showed that various AZF-linked CNVs are present in more than half of Japanese NOA patients. These results broadened the structural variations of AZF-linked CNVs, which should be considered for the molecular diagnosis of spermatogenic failure. Furthermore, the results of this study highlight the etiological heterogeneity and possible oligogenicity of isolated NOA. This study was supported by Grants from the Japan Society for the Promotion of Science (21K19283 and 21H0246), the Japan Agency for Medical Research and Development (22ek0109464h0003), the National Center for Child Health and Development, the Canon Foundation, the Japan Endocrine Society, and the Takeda Science Foundation. The results of this study were based on samples and patient data obtained from the International Center for Reproductive Medicine, Dokkyo Medical University Saitama Medical Center, Koshigaya, Japan. The authors have no conflicts of interest to disclose. N/A.

Evaluation of human bone marrow mesenchymal stem cells in the treatment of non obstructive azoospermia.

Non-obstructive azoospermia (NOA) represents an infertility problem that is usually difficult to treat. Such patients usually have testicular biopsy of germ cell aplasia or spermatogenic arrest. In recent decades, Mesenchymal Stem Cells (MSCs) had been studied thoroughly and proved safe and effective regarding their capability for trans-differentiation into different cell types. The aim of this study was to evaluate the effect of MSCs local intratesticular injection in induction of spermatogenesis. The current study included 87 infertile non-obstructive azoospermic patients. Clinical assessment and repeated semen analysis with centrifugation were done to confirm azoospermia. Karyotyping and AZF study were done. Some of the patients had previous testicular biopsy proving a lack of sperm in the testes. Single intratesticular injection of purified MSCs suspension was done. 20.7% of patients showed sperm in their semen after variable period of time. Hormonal profile among treated patients showed significant improvement regardless success of treatment. Also most of the treated patients appreciated the improvement of their sexual function and libido. Bone marrow derived MSCs could be a new hope and therapeutic modality for treatment of refractory cases of NOA.

(278) The Loss of Histone Reader Phf7 Leads to Immune Pathways Activation via Endogenous Retroviruses During Spermiogenesis

Abstract Introduction PHF7 has been identified as playing a crucial role in germline development in animals. However, its correlation with human infertility remains ambiguous. Objective To ascertain if a germline mutation in the human PHF7 gene correlates with azoospermic patients and to further determine if this mutation is a potential cause of non-obstructive azoospermia. Methods Genetic screening of azoospermic patients for PHF7 mutations. Using Phf7 knockout mice models to simulate the human mutation, aiming to validate if these mutations are directly responsible for male infertility. Investigating the role of endogenous retroviruses related to Phf7 deletion and its link to activating the immune pathway. Results A germline mutation of the human PHF7 gene was identified in azoospermic patients. In the Phf7 knockout mice model, male infertility was observed, specifically stemming from an inhibited histone-to-protamine exchange process. The absence of spermatogenesis due to the Phf7 deletion was found to be associated with the activation of the immune pathway, mediated by endogenous retroviruses. PPARα was identified as a significant factor in the testis, which is suppressed by endogenous retroviruses. Conclusions PHF7 is an essential gene associated with human infertility, particularly in non-obstructive azoospermic patients. Its role as an epigenetic reader became evident, and the absence of PHF7 leads to immune pathway activation. The study also highlighted that the PPARα agonist, Astaxanthin, holds promise as a therapeutic agent to treat male infertility resulting from PHF7 mutations. Disclosure No.

(244) Optimization of Sperm Retrieval in Non-Obstructive Azoospermic Patients using Conventional TESE

Abstract Introduction Sperm retrieval in NOA patients remain to be the only hope for these patients for fatherhood using assisted reproductive techniques. Micro-TESE is the gold standard with the best retrieval rate. Microsurgical facilities are not available at every center and conventional TESE techniques are not standardized between different centers. Objective Our aim is To improve the spermatogenic function of NOA patients to enable the appearance of sperm in their ejaculate or to improve the chances of a successful retrieval from the testis for ICSI and To optimize and standardize our conventional TESE technique Methods Over last 7 years 94 cases of NOA patients were subjected to conventional TESE. We Exclude the cases of Obstructive Azoospermia, Cases with abnormal chromosomal analysis were referred for higher center for genetic counselling and diagnosis. Laboratory workup including at least two semen samples, hormonal valuation (FSH, LH, Prolactin, Testosterone) and chromosomal analysis (Karyotyping and Testing for Y chromosome micro-deletions. Pre-TESE medical Treatment: included Clomiphene citrate: 50 mg OD for 3–6 months, Anti-oxidant therapy (Vitamin E 200mg BID and Vitamin C, 1 gm, OD). 6 biopsies from each testis: upper pole 2(medial &amp;Lateral), mid zone 2 (medial &amp; Lateral), lower pole 2(medial &amp;Lateral). Separate biopsy from each testis was sent for definitive histopathological correlation. Experienced embryologist attends each procedure examining each biopsy until sperms are retrieved. Samples taken were centrifuged and re-examined at lab for any missed sperm Results sperm retrieval was successful in 38patients (40.4%). All cases were done as day case No single case of wound infection or intra-testicular hematoma.Post-operative pain was tolerable and managed by paracetamol ±NSAID Negative cases for sperms were continued on higher dose of CC and referred for microTESE Conclusions Though of low-evidence, empiric Pre-TESE enhancement of spermatogenesis should be attempted whenever possible and after proper patients counselling. This systematic anatomical approach for biopsies yielded high SRR compared to other conventional TESE. Exact localization of + ve testicular site and side If patient needed Re-do TESE for more sperms in the future. Although micro TESE remains to be the best modality for SR in NOA, in our hands and in canters without Micro-TESE facilities, applying this standardized technique with pre-operative hormonal manipulation yields accepted rate of sperm retrieval Disclosure No.

A detection model of testis-derived circular RNAs in serum for predicting testicular sperm retrieval rate in non-obstructive azoospermia patients.

Microdissection testicular sperm extraction is an effective method to retrieve sperm from non-obstructive azoospermia patients. However, its successful rate is less than 50%. To identify the predictive value of circular RNAs in serum for sperm retrieval rate in non-obstructive azoospermia patients. 180 non-obstructive azoospermia patients were recruited in this study, including 84 individuals with successful sperm retrieval and 96 individuals with failed sperm retrieval. Our study contained two phases. First, 20 patients, selected from the 180 patients, were included in screening cohort. In this cohort, the top 20 circular RNAs from our previous testicular circRNA profiles were verified between successful and failed sperm retrieval groups using real-time polymerase chain reaction. Six circular RNAs with the most significantly different expressions were selected for further verification. Second, the 180 patients were included as discovery cohort to verify the six circular RNAs. Circular RNAs were extracted from serum in each participant. Logistic regression analysis was further performed to identify the predictive value and the area under the curve analysis was used to evaluate diagnostic efficiency, sensitivity, and specificity. Six circular RNAs including hsa_circ_0058058, hsa_circ_0008045, hsa_circ_0084789, hsa_circ_0000550, hsa_circ_0007422, and hsa_circ_0004099 showed aberrant expressions between the successful and failed sperm retrieval group. In addition, both single-circular RNA panels and multi-circular RNA panels were finally verified to be significant in predicting sperm retrieval rate. Notably, multi-circular RNAs panels demonstrated better predictive abilities compared with single-circRNA panels, and the combined panel of six-circular RNAs (risk score=1.094×hsa_circ_0058058+0.697×hsa_circ_0008045+0.718×hsa_circ_0084789-0.591×hsa_circ_0000550-0.435×hsa_circ_0007422-1.017×hsa_circ_0004099-1.561) exhibited the best predictive ability in the present study with an AUC of 0.977, a sensitivity of 91.7% and a specificity of 86.5%. A higher risk score indicated a higher risk of failure in sperm retrieval. Our study was the first to report that testis-derived circular RNAs in serum have the ability to predict sperm retrieval rate in non-obstructive azoospermia patients, whether it is a single-circular RNA or a combination of multi-circular RNAs.

Microsurgical Testicular Sperm Extraction: Predictive Factors and Outcomes for Men with Nonobstructive Azoospermia

Background. Nonobstructive azoospermia (NOA) is a frequent cause of male infertility, and microsurgical testicular sperm extraction (micro-TESE) is an effective technique for sperm retrieval in NOA cases. However, the predictive factors for successful sperm retrieval and the outcomes of micro-TESE in NOA patients are not fully understood. Methods. In this retrospective study, we evaluated the predictive value of preoperative data and outcomes of micro-TESE in 172 men with NOA who underwent the procedure between April 2012 and February 2021. Patients’ age, medical history, testicular size, hormonal profile, testicular histology, and sperm retrieval were recorded. Results. 172 cases of micro-TESE were performed (99 unilateral and 73 bilateral) and testicular sperm retrieval rate (SRR) was 72.7%. There was no statistical difference in age, testicular size, and hormonal profiles between positive SRR and negative SRR groups. Clinical varicocele and previous orchitis did not significantly affect the result of micro-TESE. Y chromosome microdeletions were detected in seven patients. On histological examination, hypospermatogenesis was found in 39% of patients who underwent testicular biopsy. SRR was higher in patients with hypospermatogenesis than in other groups. Conclusions. Testicular microdissection is an effective and safe procedure for nonobstructive azoospermia patients, regardless of the different etiologies and preoperative data. Age, testicular size, and hormonal levels did not predict the success of micro-TESE. The presence of hypospermatogenesis was associated with a higher SRR. Clinical varicocele and previous orchitis were not significantly associated with the outcome of micro-TESE.

A step closer to parenthood with non-obstructive azoospermia: Unveiling the impact of microdissection testicular sperm extraction in Australia's largest single-centre study.

Non-obstructive azoospermia (NOA) diagnosis poses challenges for couples seeking parenthood. Microdissection testicular sperm extraction (MD-TESE) excels in retrieving testicular sperm cells for NOA cases. However, limited live birth data in Australian NOA patients hinders accurate counselling. This study aimed to determine the likelihood of infertile couples with a male partner diagnosed with NOA conceiving biological children using MD-TESE / intracytoplasmic sperm injection (ICSI). A retrospective cohort study included 108 NOA men treated at a public fertility unit and a private fertility centre (May 2009-May 2022). live birth rate (LBR); secondary outcomes: sperm retrieval rate, pregnancy rate, and neonatal outcomes. Among 108 patients undergoing MD-TESE, the positive sperm retrieval rate (PSRR) was 64.8% (70/108). Histology best predicted sperm retrieval success, with hypo-spermatogenesis yielding a 94.1% PSRR. Age, testicular volume, and hormonal parameters had no significant impact. Mean male age: 35.4 years; mean partner age: 32.7 years. Fertilisation rate: 50.7%. LBR per initiated cycle: 58.7% (37/63); per embryo transfer: 63.8% (37/58); per initially diagnosed NOA man: 34.3% (37/108). Cumulative LBR: 74.1% (43/58); twin rate: 10.8% (4/37). No neonatal deaths or defects were observed among 47 live offspring. This study provides valuable data for counselling NOA couples on the probability of conceiving biological offspring. MD-TESE and ICSI yielded favourable PSRR (64.8%) and LBR (63.8%). However, couples should be aware that once NOA is confirmed, the chance of taking home a baby is 34%.

(336) Sperm Retrieval Success During Tese: Prognostic Factors in Non-obstructive Azoospermia Patients

(336) Sperm Retrieval Success During Tese: Prognostic Factors in Non-obstructive Azoospermia Patients

Y chromosome polymorphisms contribute to an increased risk of non-obstructive azoospermia: a retrospective study of 32,055 Chinese men.

To investigate the prevalence of Y chromosome polymorphisms in Chinese men and analyze their associations with male infertility and female adverse pregnancy outcomes. The clinical data of 32,055 Chinese men who underwent karyotype analysis from October 2014 to September 2019 were collected. Fisher's exact test, chi-square test, or Kruskal-Wallis test was used to analyze the effects of Y chromosome polymorphism on semen parameters, azoospermia factor (AZF) microdeletions, and female adverse pregnancy outcomes. The incidence of Y chromosome polymorphic variants was 1.19% (381/32,055) in Chinese men. The incidence of non-obstructive azoospermia (NOA) was significantly higher in men with the Yqh- variant than that in men with normal karyotype and other Y chromosome polymorphic variants (p < 0.050). The incidence of AZF microdeletions was significantly different among thenormal karyotype and different Y chromosome polymorphic variant groups (p < 0.001). The detection rate of AZF microdeletions was 28.92% (24/83) in the Yqh- group and 2.50% (3/120) in the Y ≤ 21 group. The AZFb + c region was the most common AZF microdeletion (78.57%, 22/28), followed by AZFc microdeletion (7.14%,2/28) in NOA patients with Yqh- variants. There was no significant difference in the distribution of female adverse pregnancy outcomes among thenormal karyotype anddifferent Y chromosome polymorphic variant groups (p = 0.528). Patients with 46,XYqh- variant have a higher incidence of NOA and AZF microdeletions than patients with normal karyotype and other Y chromosome polymorphic variants. Y chromosome polymorphic variants do not affect female adverse pregnancy outcomes.

An integrated approach for improving clinical management of non-obstructive azoospermia.

Non-obstructive azoospermia is the most severe form of male infertility. A testicular biopsy is required for the diagnosis of non-obstructive azoospermia, and the causal factors for non-obstructive azoospermia remain unknown. To reduce the risk of multiple biopsies and identify factors that contribute to non-obstructive azoospermia, we proposed an integrated approach for the preoperative diagnosis and clinical management of non-obstructive azoospermia by applying the chromosome-spreading technique and whole-exome sequencing. Between July 2020 and December 2022, after ruling out definitive obstructive azoospermia and non-obstructive azoospermia patients with testicular volume<6mL, 20 patients with non-obstructive azoospermia who underwent preoperative testicular diagnostic biopsy using testicular sperm aspiration were subjected to retrospective analysis. Microscopic examination identified four patients with sperm cells, and 16 without sperm cells. Routine pathological analysis classified one patient as normal spermatogenesis, three as hypospermatogenesis, five as maturation arrest, nine as Sertoli cell-only, and two as unable to judge. With chromosome-spreading technology using routine cell suspension samples for microscopic examination, 18 patient diagnoses were validated, and two patients without a definitive diagnosis were supplemented. Detection of the Y chromosome and a well-organized whole-exome sequencing analysis revealed potential genetic factors. The full use of testicular biopsy is beneficial for the diagnosis of azoospermia, as it avoids the risk of multiple biopsies. Moreover, in combination with whole-exome sequencing, clinicians can obtain more information regarding the pathogenesis of non-obstructive azoospermia, which may guide treatment.

Microdissection testicular sperm extraction in non-obstructive azoospermic patients with solitary testis: a retrospective case-control study.

Obtaining sperm from the testis surgically and using these sperm with the intracytoplasmic sperm injection technique, has opened the way for the possibility of biological fathering in men with non-obstructive azoospermia (NOA). We aimed to evaluate our sperm retrieval rate (SRR) by microdissection testicular sperm extraction (micro-TESE) in NOA patients with solitary testis. In this retrospective case-control study, fortyfive patients with NOA who had a congenital or acquired solitary testis were included, between September 2003 and January 2022. These patients were randomly matched with patients with NOA who had bilateral testes, using a 1:3 matching ratio. We found that SRR by micro-TESE in patients with solitary testis was similar to NOA patients with bilateral testis (51.1% vs. 50.4%). Age, infertility period, ejaculate volume, serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH) and testosterone, history of varicocelectomy, history of orchiopexy, testicular stimulation therapy before micro-TESE, testicular volume, genetic status, TESE side, micro-TESE success, complications and histopathological evaluation results of both groups were evaluated, there was a statistically significant difference in only serum FSH and LH levels. There was no difference between the groups in terms of complications and hormonal effects in the early postoperative period. Micro-TESE in NOA patients with solitary testis has similar sperm retrieval and complication rates as NOA patients with bilateral testis.

Micromapping testicular sperm extraction: A new technique for microscopic testicular sperm extraction in nonobstructive azoospermia.

In microscopic testicular sperm extraction (mTESE) for nonobstructive azoospermia (NOA), sperm can be recovered relatively easily in some cases, and mTESE may be retrospectively considered excessive. However, mTESE is routinely performed in the majority of NOA patients because of the difficulty in predicting tissue status. A minimally invasive and comprehensive sperm retrieval method that allows on-the-spot tissue assessment is needed. We have developed and evaluated a novel sperm retrieval technique for NOA called micromapping testicular sperm extraction (MMTSE). MMTSE involves dividing the testis into four sections and making multiple small needle holes in the tunica albuginea to extract seminiferous tubules and retrieve sperm. The sperm-positive group by MMTSE (Group I) underwent additional tissue collection (ATC) via a small incision, whereas the sperm-negative group by MMTSE (Group 0) underwent mTESE. In total, 40 NOA participants underwent MMTSE. Group I included 15 patients and Group 0 included 25 patients. In Group 1, sperm were recovered from all patients by ATC. In Group 0, sperm were recovered in 4 of 25 cases using mTESE. MMTSE shows promise as a simple method that comprehensively searches testicular tissue and retrieves sperm using an appropriate method while minimizing patient burden.

DNM3OS Enhances the Apoptosis and Senescence of Spermatogonia Associated with Nonobstructive Azoospermia by Providing miR-214-5p and Decreasing E2F2 Expression.

Nonobstructive azoospermia (NOA) is a complex disease characterized by the spermatogenic dysfunction of testicular tissues. The roles played by long noncoding RNAs (lncRNAs) in NOA pathogenesis have not been extensively studied. Microarray assays were performed on samples of testicular biopsy tissue obtained from patients with NOA for the purpose of identifying differentially expressed lncRNAs and messenger RNA (mRNA) transcripts, and the results were verified by quantitative real-time polymerase chain reaction. Mouse-derived GC-1 spermatogonia (spg) cells undergoing treatment with Adriamycin (ADR) were used to investigate the biological functions of the selected lncRNAs in vitro. The target microRNAs (miRNAs) of lncRNAs and the target mRNAs of miRNAs were predicted by a bioinformatics analysis. Functional studies performed using the CCK-8 assay, EdU incorporation assay, apoptosis detection, and senescence-associated β-galactosidase (SA-β-Gal) staining were conducted using GC-1 spg cells. Totals of 2,652 lncRNAs and 2,625 mRNAs were found to be differentially expressed in the testicular tissue of NOA patients when compared with patients in a control group. Dynamin 3 opposite strand (DNM3OS) was a provider of pe-miR-214-5p that positively regulates miR-214-5p expression in GC-1 spg cells. The E2 factor (E2F) family of transcription factor 2 (E2F2) was initially predicted and subsequently verified to be a downstream gene of miR-214-5p. E2F2 expression was upregulated after DNM3OS knockdown in ADR-treated GC-1 spg cells. Moreover, knockdown of either DNM3OS or miR-214-5p significantly alleviated ADR-induced decreases in cellular activity and proliferation, as well as increases in apoptosis and senescence of mouse spermatogonial GC-1 spg cells. DNM3OS was found to regulate the apoptosis and senescence of spermatogonia by providing miR-214-5p and decreasing E2F2 expression, suggesting it as a novel target for gene therapy of male infertility.

Identification of ITPR1 gene as a novel target for hsa-miR-34b-5p in non-obstructive azoospermia: a Ca2+/apoptosis pathway cross-talk

MiR-34b-5p has been reported as a non-invasive diagnostic biomarker for infertility. However, no gene targets regulating the mechanism of cation of this miRNA are known. In this study, using gene set enrichment analysis the Inositol 1,4,5-Trisphosphate Receptor Type 1 (ITPR1) gene was identified as the sole target for hsa-miR-34b-5p, and found significantly overexpressed in non-obstructive azoospermia (NOA) patients. This finding was confirmed by qRT-PCR on fresh testicular tissues from NOA patients. Then, pathway enrichment analysis as well as the diagnostic value analysis of hsa-miR-34b-5p/ITPR1 indicated ITPR1 as a hub gene in the calcium (Ca2+)-apoptosis pathway, and a valuable predictive biomarker for NOA. Moreover, gene expression and histological assays showed the association of the effects of ITPR1’s increased expression on spermatogenesis failure through induction of apoptosis in NOA patients. These data suggested that the hsa-miR-34b-5p/ITPR1 axis could serve as a potential regulatory predictive biomarker for human spermatogenesis through the Ca2+-apoptosis pathway cross-talk.

AI Model Developed Using Machine Learning for Predicting Sperm Retrieval in Micro-TESE for Nonobstructive Azoospermia Patients

Azoospermia is a severe problem that prevents couples from having their own children through natural pregnancy. In nonobstructive azoospermia (NOA), microdissection testicular sperm extraction (micro-TESE) is required to collect sperm and, at 40%–60%, the sperm retrieval success rate is not very high. Previous studies identified no single clinical finding or investigation that could accurately predict the outcome of sperm retrieval. It would be very valuable to have a factor for predicting the possibility of sperm retrieval in patients with NOA before performing micro-TESE. We retrospectively obtained data from the medical records of 430 patients who underwent micro-TESE from 2011 to 2020. Parameters extracted were age, height, body weight, body mass index, luteal hormone, follicle-stimulating hormone, PRL, total testosterone, E2, T/E2, sperm retrieval, G-band, AZF, medical history, Rt testis, and Lt testis. Prediction One, which does not require coding, was used to create the AI prediction model for sperm retrieval. Prediction One makes the best prediction model using an artificial neural network with internal cross-validation. Prediction One also evaluates the “importance of variables” using a method based on permutation feature importance. The AUC for the AI model was 0.7246, which is acceptable. In addition, among the variables, T/E2 ratios contributed most to predicting whether sperm retrieval was possible or not. However, the difference in T/E2 between successful and unsuccessful sperm retrieval was not statistically significant. In addition, our analysis of data from 20 patients who underwent micro-TESE in 2021 found that in 85%, the actual result matched the result predicted using our novel AI model. We created an AI model for predicting sperm retrieval in patients with NOA before undergoing micro-TESE. In addition, we found that T/E2 ratios contributed most to predicting possibility of sperm retrieval in NOA using machine learning.

#337 : Six Characters of Seminiferous Tubules Appearances During Micro-Tese Related with Different Sperm Retrieval Rate

Background and Aims: For non-obstructive azoospermia (NOA) patients, micro-TESE is considered to have higher sperm retrieval rate (SRR) than traditional surgery. However, various clinical parameters are not reliable predictors for the presence of focal spermatogenesis and SRR. This study summarized the characters of seminiferous tubules appearances and investigated their relationship with SRR. Method: This was a retrospective study. From September 2014 to April 2023, 638 NOA patients treated with micro-TESE were divided into six groups according to the seminiferous tubules appearances (Group-A: homogeneously thick, Group-B: homogeneously thin, Group-C: seminiferous tubules were severe thin and loss the tubal appearance, Group-D: the testicles were filled with hyperplastic Leydig cells, seminiferous tubules were severe thin and rare, Group-E, thick tubules mixed with thin tubules, Group-F, extensive testicular microlithiasis). Age, testicular volume, serum FSH, testosterone level, SRR and the heterogeneous seminiferous tubules presenting rate (HSTPR) were compared. Results: Sperms were successfully retrieved in 333 cases (SRR=52.2%). There were no statistical differences in age, testicular volume, FSH and testosterone level between the patients who succeeded and failed to obtain sperm (Table 1, all [Formula: see text]&gt;0.05). The constituent ratio of etiology was different in the six groups (Table 2, [Formula: see text]&lt;0.05). Between the six groups, testicular volume, FSH, testosterone, SRR and HSTPR were statistically different (Table 3, all [Formula: see text]&lt;0.05). Conclusion: Age, testicular volume, FSH and testosterone were insufficient to predict the SRR of NOA patients. Micro-TESE is commended to terminated when sperm are retrieved in case that the seminiferous tubules are rare or too thin to present tubal appearance, thick tubules mix with thin tubules or extensive testicular microlithiasis present (Group C-F). If the tubules appear homogeneously thick or thin, micro-TESE would has less advantage comparing with conventional multiple TESE because of the difficulty to identify the differences between the tubules even if the sperm is finally retrieved (Group A-B).

#110 : Diagnostic Approach and Clinical Recommendations for Non-obstructive Azoospermia: Global Andrology Forum Survey Results

Background and Aims: Non-obstructive azoospermia (NOA) is the most common form of male infertility, characterized by an absence of sperm in ejaculate and is caused by testicular failure. While NOA afflicts 10-38.3% of infertile males, there is no specific recommendations in EAA/EAU guidelines on its clinical diagnosis or WHO guidelines on semen analysis. Therefore, we conducted a global survey to assess the standard approach of practitioners in the diagnosis and management of NOA. Method: The Global Andrology Forum (GAF) conducted a survey on 436 independent practicing entities from 55 countries. We compiled data from 336 respondents (andrologists, urologists, fertility specialists). Information collected included frequency of NOA, varicocele, frequency of obtaining semen, interval between obtaining semen samples, hormone evaluation (FSH, LH, prolactin, total/free testosterone, SHBG, estradiol, inhibin), genetic testing (karyotype, Y-chromosome deletion, CFTR, Kal1) and referral to genetic counsellors. Results: The majority of participants (80%) either reported &lt;10% or 10-25% NOA frequency in their population (Fig. 1A). 40.6% of participants stated that 10-25% of men with NOA cases have a clinically significant varicocele (Fig. 1B). For NOA diagnosis, majority of participants cited the requirement of at least two semen samples, with a period of 14-29 days as the interval between two semen analyses. To achieve high-quality semen, consideration must be given to an abstinence period (2-7days) and the complete collection of ejaculates. The survey reported FSH (97%), LH (86.9%) and total testosterone (92.9%) tests as most frequently used for NOA patients (Fig. 2A). The initial hormonal evaluation of a patient with suspected NOA should include total testosterone levels and LH, as these hormones are the primary regulators of spermatogenesis. Karyotype analysis (86.2%), Y-microdeletion (88.3%), CFTR (21.9%) and Kal1 (2.7%) were most routinely performed genetic tests (Fig. 2B). Referrals to genetic counsellors is recommended as these defects have the potential to be passed to future generations. Conclusion: This global study evaluates NOA frequency, clinical diagnosis and management by specialists, in an effort to fill the gap in NOA research. Concomitantly, we provide GAF expert recommendations to enhance our collective understanding of NOA, optimize patient care and improve the management of NOA.