Single-cell RNA sequencing reveals transcriptomic landscape and potential targets for human testicular ageing.

STUDY QUESTION: What is the molecular landscape underlying the functional decline of human testicular ageing? SUMMARY ANSWER: The present study provides a comprehensive single-cell transcriptomic atlas of testes from young and old humans and offers insights into the molecular mechanisms and potential targets for human testicular ageing. WHAT IS KNOWN ALREADY: Testicular ageing is known to cause male age-related fertility decline and hypogonadism. Dysfunction of testicular cells has been considered as a key factor for testicular ageing. STUDY DESIGN, SIZE, DURATION: Human testicular biopsies were collected from three young individuals and three old individuals to perform single-cell RNA sequencing (scRNA-seq). The key results were validated in a larger cohort containing human testicular samples from 10 young donors and 10 old donors. PARTICIPANTS/MATERIALS, SETTING, METHODS: scRNA-seq was used to identify gene expression signatures for human testicular cells during ageing. Ageing-associated changes of gene expression in spermatogonial stem cells (SSCs) and Leydig cells (LCs) were analysed by gene set enrichment analysis and validated by immunofluorescent and functional assays. Cell-cell communication analysis was performed using CellChat. MAIN RESULTS AND THE ROLE OF CHANCE: The single-cell transcriptomic landscape of testes from young and old men was surveyed, revealing age-related changes in germline and somatic niche cells. In-depth evaluation of the gene expression dynamics in germ cells revealed that the disruption of the base-excision repair pathway is a prominent characteristic of old SSCs, suggesting that defective DNA repair in SSCs may serve as a potential driver for increased de novo germline mutations with age. Further analysis of ageing-associated transcriptional changes demonstrated that stress-related changes and cytokine pathways accumulate in old somatic cells. Age-related impairment of redox homeostasis in old LCs was identified and pharmacological treatment with antioxidants alleviated this cellular dysfunction of LCs and promoted testosterone production. Lastly, our results revealed that decreased pleiotrophin signalling was a contributing factor for impaired spermatogenesis in testicular ageing. LARGE SCALE DATA: The scRNA-seq sequencing and processed data reported in this paper were deposited at the Genome Sequence Archive (https://ngdc.cncb.ac.cn/), under the accession number HRA002349. LIMITATIONS, REASONS FOR CAUTION: Owing to the difficulty in collecting human testis tissue, the sample size was limited. Further in-depth functional and mechanistic studies are warranted in future. WIDER IMPLICATIONS OF THE FINDINGS: These findings provide a comprehensive understanding of the cell type-specific mechanisms underlying human testicular ageing at a single-cell resolution, and suggest potential therapeutic targets that may be leveraged to address age-related male fertility decline and hypogonadism. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research and Development Program of China (2022YFA1104100), the National Natural Science Foundation of China (32130046, 82171564, 82101669, 82371611, 82371609, 82301796), the Natural Science Foundation of Guangdong Province, China (2022A1515010371), the Major Project of Medical Science and Technology Development Research Center of National Health Planning Commission, China (HDSL202001000), the Open Project of NHC Key Laboratory of Male Reproduction and Genetics (KF202001), the Guangdong Province Regional Joint Fund-Youth Fund Project (2021A1515110921, 2022A1515111201), and the China Postdoctoral Science Foundation (2021M703736). The authors declare no conflict of interest.

[Compound Chamomile and Lidocaine Hydrochloride Gel applied at different time-windows for premature ejaculation: A prospective single-center randomized controlled study].

OBJECTIVE: To investigate the efficacy and safety of Compound Chamomile and Lidocaine Hydrochloride Gel (CCLH) (Kamistad) applied at different time-windows on premature ejaculation (PE). METHODS: This prospective study included 72 PE patients treated by application of CCLH to the glans and penile body in our hospital from February to October 2021. According to the time of drug administration before insertion into the vagina, we randomly divided the patients into a 5-minute group (n = 39) and a 15-minute group (n = 33). Before and after 1 and 2 weeks of treatment, we compared the intravaginal ejaculation latency time (IELT), PE diagnostic tool (PEDT) score, quality of life, and adverse reactions between the two groups of patients. RESULTS: Totally 62 of the patients completed the follow-up, 35 in the 5-minute group and 27 in the 15-minute group, and all showed significant improvement in IELT (P < 0.01) and PEDT score (P < 0.05) after treatment compared with the baseline. No allergic reactions, such as redness and swelling, developed at the application site in any of the patients, and no adverse significant effect was observed on the erectile hardness in 61 of the cases. Six cases showed increased erectile hardness instead. Fifty-seven of the patients experienced no obvious penile numbness or reduced sexual satisfaction, and all could complete their sexual activities. CONCLUSION: Compound Chamomile and Lidocaine Hydrochloride Gel applied at different time-windows is effective on PE, with a 5-minute rapid onset of action before intercourse, and no obvious adverse effects.

[Expression of the NMDA receptor in the rat model of orchialgia].

OBJECTIVE: To explore the expression of the N-methyl-D-aspartate (NMDA) receptor in the rat model of orchialgia and its possible mechanisms. METHODS: According to Yoshioka's method, the male rats in the control group were injected with 0.2 ml saline, and those in the experimental group with 0.2 ml 2% acetic acid solution. Then we tested the behavioral responses of the rats and determined the expressions of the subunits NR1 and NR2B of the NMDA receptor in the dorsal root ganglion and spinal dorsal horn by Western blot, RT-qPCR and immunofluorescence staining. RESULTS: The withdrawal latency was decreased in the model rats, reaching the lowest value at 4 hours after modeling, significantly lower than in the controls (ï¼»4.15 ± 0.84ï¼½ vs ï¼»12.32 ± 1.05ï¼½, P < 0.05). Compared with the controls, the model rats showed remarkably increased mRNA and protein expressions of NR2B in the dorsal root ganglion (P < 0.05) but not in the spinal dorsal horn at 4 hours. However, no statistically significant difference was found in the expression of NR1 either in the dorsal root ganglion or in the spinal dorsal horn between the two groups (P > 0.05). CONCLUSIONS: The NMDA receptor plays an important role in pathogenesis of orchialgia in rats. In the early stage of pain, upregulating the expression of the subunit NR2B of the NMDA receptor can mediate peripheral hyperalgesia and consequently orchialgia.

Subinguinal microsurgical varicocelectomy with intraoperative microvascular Doppler ultrasound leads to the pain-free outcome after surgery.

OBJECTIVE: To determine efficacy of intraoperative microvascular Doppler ultrasound in assisting subinguinal microsurgical varicocelectomy for pain relief in the treatment of painful varicoceles, compared to the microsurgery without Doppler ultrasound. METHODS: Total 153 patients underwent microsurgical varicocelectomy were randomly allocated to two groups: Groups 1 and 2 included 82 and 71 patients monitored with and without using intraoperative microvascular Doppler ultrasound, respectively. The assessments were compared between two groups, including intraoperative parameters (vessel numbers and operative time) and postoperative outcomes (pain resolution, complications and recurrence). RESULTS: The average numbers of internal spermatic veins ligated (13.87±6.43 vs 11.72±5.66) and arteries preserved (1.96±0.87 vs 1.73±0.86) were significantly greater in Group 1 with Doppler ultrasound. Precisely, the smaller size of the internal spermatic veins was ligated and the more encircled arteries were also preserved in Group 1. In two groups with and without using Doppler ultrasound, 56 (68.3%) and 36 (50.7%) patients experienced a complete resolution of pain, 21 (25.6%) and 29 (40.9%) patients experienced partial resolution, whereas 5 (6.1%) and 6 (8.5%) patients experienced no change in the chronic pain, respectively. Thus, patients in Group 1 had a better outcome in chronic pain resolution (P = 0.033). The operative time, complications and recurrence rate were not different between the two groups. CONCLUSIONS: Subinguinal microsurgical varicocelectomy is an effective method to treat painful varicoceles. With the assistance of Doppler Ultrasound monitoring, greater numbers of vessels were identified and a better outcome of pain resolution was achieved.

[Animal models of male genital tract chlamydia infection: An update].

Chlamydia trachomatis infection is one of the most prevalent sexually transmitted diseases in males and studies on its pathogenic and immunologic mechanisms are rather limited. Animal models play an important role in studying the pathogenesis, course and treatment of human diseases, and those of male genital tract chlamydial infection are relatively few and not well developed. This article focuses on the chlamydia species, animal species, infection route, infected organ, and infection process of chlamydia, as well as its impact on reproduction, aiming to provide some help for further studies of male genital tract chlamydial infection.

[Expressions of TRPV1 and TRPA1 in the dorsal root ganglion in the rat model of orchialgia].

OBJECTIVE: To explore the expressions of transient receptor potential vanilloid 1 (TRPV1) and TRP ankyrin 1 (TRPA1) in the dorsal root ganglion (DRG) and their action mechanisms in the rat model of orchialgia. METHODS: The models of orchialgia were established in male SD rats by injection of 2% acetic acid into the testis. Then the number of spontaneous pain responses and withdrawal latency in the model rats were recorded by behavioral tests and the expressions of TRPV1 and TRPA1 in T13－L1 DRGs determined by RT-qPCR, Western blot and immunofluorescence staining. RESULTS: Compared with the normal control rats, the orchialgia models showed a significant increase in the number of spontaneous pain responses (0.13 ± 0.35 vs 22.63 ± 3.42, P<0.01) and a decrease in the withdrawal latency at 4 hours after injection (ï¼»12.75 ± 1.50ï¼½ vs ï¼»4.85 ± 1.00ï¼½ s, P<0.05). The mRNA expressions of both TRPV1 and TRPA1 were observed in the membrane of the neurons in the DRG, the former increased by 1.77 times and the latter by 1.75 times that of the control (P<0.05). CONCLUSIONS: The expressions of TRPV1 and TRPA1 were up-regulated in the DRG of the rat models of orchialgia, which may be involved in the allodynia and hyperalgesia of the rats.

[Updated diagnosis and management of chronic orchialgia].

Chronic orchialgia (CO) is a common complaint in urology or andrology. Due to its complicated pathogenesis, the diagnosis and treatment of CO are quite challenging. Based on different etiologies, CO can be idiopathic or secondary. Idiopathic CO accounts for approximately 50% of the cases and is probably associated with Wallerian degeneration in the spermatic cord nerves and peripheral sensitization. Secondary CO can be attributed to direct causes and its treatment focuses on the pathologic condition identified. The main methods for the treatment of CO include conservative and surgical strategies, among which microsurgical spermatic cord denervation ( MSCD) is an effective and minimally invasive option, while orchiectomy is but the last alternative when no other means is left.

[Roles of pericytes in the urological and reproductive systems].

Pericyte, also known as mural cell or Rouget cell, is one of the main cells that make up the wall of capillaries. Pericytes play important roles not only in the maturation, stability, and function maintenance of blood vessels, but also in the growth and development of tissues and organs, wound repair, and other physiological and pathological processes. Researches on the functions of pericytes are mainly concentrated on their multipotency, adjustment of vascular functions, and process of fibrosis, as well as on renal fibrosis, renal blood flow regulation, and glomerular filtration in urology, but are quite insufficient in andrology. This article reviews the location, origin, distribution, morphology, markers, and functions of pericytes, aiming to induce further studies of pericytes in andrology.

[Yimusake Tablet: safe and efficacious for premature ejaculation].

OBJECTIVE: To objectively evaluate the efficacy and safety of Yimusake Tablet in the treatment of premature ejaculation (PE) through a multi-centered large-sample trial. METHODS: We conducted a multi-centered, open, fixed-dose, and self-compared clinical trial among 300 patients with diagnosed PE. The trial lasted 12 weeks, including 4 weeks without any medication and 8 weeks of treatment with Yimusake Tablet, 2 pills (1 g) per night. We observed the intravaginal ejaculation latency time (IELT) before and after treatment, evaluated the safety of medication, and performed a questionnaire investigation on the patients' satisfaction. RESULTS: Of the 300 PE patients, 288 accomplished the clinical trial. The patients ranged in age from 22 to 60 years, averaging at 31.6 years. The mean IELT of the patient was 62.5 seconds at baseline, 168.9 seconds after 4 weeks of treatment with Yimusake Tablet, and 222.2 seconds after 8 weeks of medication. Among the 157 patients with normal erectile function (IIEF >21), the mean IELT was 71.4 seconds before treatment, 147.4 seconds after 4 weeks of medication, and 172.5 seconds after 8 weeks of medication. The patients' satisfaction was significantly increased after treatment. Those complicated by mild to moderate erectile dysfunction achieved different degrees of improvement in the IIEF-5 score, with a mean increase of 3.8. Only a few patients experienced mild adverse events, including constipation, dry mouth, nose bleeding, abdominal pain, and lumbosacral pain, which were all relieved without drug withdrawal. CONCLUSION: Yimusake Tablet is a safe and effective medicine for the treatment of PE.

[Microsurgical management of male infertility in china: 15-year development and prospects].

Male infertility is a common and complex disease in urology and andrology, and for many years there has been no effective surgical treatment. With the emergence of microsurgery and assisted reproductive medicine (IVF/ICSI), rapid development has been achieved in the treatment of male infertility. The Center for Male Reproductive Medicine and Microsurgery at Weill Cornell Medical College of Cornell University has been playing an important leading role in developing microsurgical techniques for the management of male infertility. The development of microsurgical treatment of male infertility in China has experienced the 3 periods of emerging, making, and boosting ever since its systematic introduction from Weill Cornell Medical College 15 years ago. At present, many Chinese hospitals have adopted microsurgery in the management of male infertility, which has contributed to the initial establishment of a microsurgical treatment system for male infertility in China. However, some deficiencies do exist concerning microsurgical treatment of male infertility, as in normalized technical training programs for competent surgeons, unified criteria for evaluation of surgical outcomes, and detailed postoperative follow-up data. This article presents an overview on the 15-year development of microsurgical management of male infertility in China, points out the existing deficiencies, and offers some propositions for the promotion of its development.

Cell-specific expression of Cre recombinase in rat noradrenergic neurons via CRISPR-Cas9 system.

Noradrenergic neurons play a crucial role in the functioning of the nervous system. They formed compact small clusters in the central nervous system. To target noradrenergic neurons in combination with viral tracing and achieve cell-type specific functional manipulation using chemogenetic or optogenetic tools, new transgenic animal lines are needed, especially rat models for their advantages in large body size with facilitating easy operation, physiological parameter monitoring, and accommodating complex behavioral and cognitive studies. In this study, we successfully generated a transgenic rat strain capable of expressing Cre recombinase under the control of the dopamine beta-hydroxylase (DBH) gene promoter using the CRISPR-Cas9 system. Our validation process included co-immunostaining with Cre and DBH antibodies, confirming the specific expression of Cre recombinase. Furthermore, stereotaxic injection of a fluorescence-labeled AAV-DIO virus illustrated the precise Cre-loxP-mediated recombination activity in noradrenergic neurons within the locus coeruleus (LC). Through crossbreeding with the LSL-fluorescence reporter rat line, DBH-Cre rats proved instrumental in delineating the position and structure of noradrenergic neuron clusters A1, A2, A6 (LC), and A7 in rats. Additionally, our specific activation of the LC noradrenergic neurons showed effective behavioral readout using chemogenetics of this rat line. Our results underscore the effectiveness and specificity of Cre recombinase in noradrenergic neurons, serving as a robust tool for cell-type specific targeting of small-sized noradrenergic nuclei. This approach enhances our understanding of their anatomical, physiological, and pathological roles, contributing to a more profound comprehension of noradrenergic neuron function in the nervous system.

HEF1 promotes epithelial mesenchymal transition and bone invasion in prostate cancer under the regulation of microRNA-145.

The principal problem arising from prostate cancer (PCa) is its propensity to metastasize to bones, and it's crucial to understand the mechanism of tumor progression to metastasis in order to develop therapies that may reduce the morbidity and mortality of PCa patients. Although we had identified that microRNA(miR)-145 could repress bone metastasis of PCa via regulating epithelial-mesenchymal transition (EMT) in previous study, it is still unknown how miR-145 regulated EMT. In the present study, we constructed a luciferase reporter system and identified HEF1 as a direct target of miR-145. More importantly, HEF1 was shown to promote migration, invasion and EMT of PC-3 cells, a human PCa cell line originated from a bone metastatic PCa specimen. And HEF1 was also shown to partially mediate miR-145 suppression of EMT and invasion. Furthermore, inhibition of HEF1 repressed bone invasion of PC-3 cells in vivo. Expression of HEF1 was negatively correlated with miR-145 in primary PCa and bone metastatic specimens, but HEF1 was higher in samples which were more likely to commit to bone metastasis or those with higher free prostate-specific antigen (fPSA) levels and Gleason scores. Taken together, these findings indicate that HEF1 promotes EMT and bone invasion in prostate cancer by directly targeted by miR-145, and miR-145 suppresses EMT and invasion, at least in part, through repressing HEF1.

Transurethral bipolar plasma kinetic resection of ejaculatory duct for treatment of ejaculatory duct obstruction.

To evaluate the efficacy of transurethral bipolar plasma kinetic resection of ejaculatory duct for ejaculatory duct obstruction. The clinical information of 42 cases of ejaculatory duct obstruction was analyzed between July 2008 and June 2012. The diagnostic criteria included semen analysis, fructose and neutral α-glucosidase measurement in seminal plasma, transrectal ultrasonography, magnetic resonance imaging and vasography necessarily. Endoscopic procedure with bipolar plasma kinetic resection of ejaculatory duct was performed in all patients. Among these cases followed up 6 ≈ 24 months after operation, 38 patients (90.5%) had improved semen parameters, 23 azoospermic patients (60.5%) had sperm in the semen and 13 patients' wife (31%) achieved pregnancies in 42 cases of bipolar plasma kinetic resection of ejaculatory duct. Postoperative complications ensued as epididymitis in 1 case, watery ejaculate in 1, but no serious complication was observed. Bipolar plasma kinetic resection of ejaculatory duct appears to represent a promising endoscopic treatment alternative for ejaculatory duct obstruction patients, with high efficacy, less complications, quicker recovery and satisfactory follow-up parameters.

[Qilin pills combined with levofloxacin for asthenospermia accompanied with accessory sex gland infection: a clinical observation].

OBJECTIVE: To observe the efficacy of the combination of Qilin Pills and levofloxacin in the treatment of asthenospermia accompanied with accessory sex gland infection. METHODS: We randomly assigned 80 asthenospermia patients with accessory sex gland infection to groups 1 and 2 of equal number, the former treated with Qilin Pills + levofloxacin, and the latter with levofloxacin only. Qilin Pills were administered at the dose of 6 g tid for 30 days, and levofloxacin at the dose of 0.5 g qd for 20 days. We obtained semen parameters, including the percentage of progressively motile sperm and peroxidase-positive white blood cell (WBC) count, before and after medication, and compared the clinical effects between the two groups. RESULTS: All the patients accomplished the clinical trial. The therapeutic effectiveness rates in improving progressive sperm motility were 60% in group 1 and 17.5% in group 2, with statistically significant differences between the two groups (P < 0.05). As for the peroxidase-positive WBC count in semen, the effectiveness rates were 87.5% and 82.5%, respectively, with no significant differences between the two groups (P > 0.05). CONCLUSION: For the treatment of asthenospermia accompanied with accessory sex gland infection, Qilin Pills combined with levofloxacin is evidently better than levofloxacin alone in improving sperm motility, and it has no obvious adverse effects.

Precise Correction of Lhcgr Mutation in Stem Leydig Cells by Prime Editing Rescues Hereditary Primary Hypogonadism in Mice.

Hereditary primary hypogonadism (HPH), caused by gene mutation related to testosterone synthesis in Leydig cells, usually impairs male sexual development and spermatogenesis. Genetically corrected stem Leydig cells (SLCs) transplantation may provide a new approach for treating HPH. Here, a novel nonsense-point-mutation mouse model (LhcgrW495X ) is first generated based on a gene mutation relative to HPH patients. To verify the efficacy and feasibility of SLCs transplantation in treating HPH, wild-type SLCs are transplanted into LhcgrW495X mice, in which SLCs obviously rescue HPH phenotypes. Through comparing several editing strategies, optimized PE2 protein (PEmax) system is identified as an efficient and precise approach to correct the pathogenic point mutation in Lhcgr. Furthermore, delivering intein-split PEmax system via lentivirus successfully corrects the mutation in SLCs from LhcgrW495X mice ex vivo. Gene-corrected SLCs from LhcgrW495X mice exert ability to differentiate into functional Leydig cells in vitro. Notably, the transplantation of gene-corrected SLCs effectively regenerates Leydig cells, recovers testosterone production, restarts sexual development, rescues spermatogenesis, and produces fertile offspring in LhcgrW495X mice. Altogether, these results suggest that PE-based gene editing in SLCs ex vivo is a promising strategy for HPH therapy and is potentially leveraged to address more hereditary diseases in reproductive system.

Percutaneous ultrasound-guided radiofrequency ablation treatment and genetic testing for renal cell carcinoma with Von Hippel-Lindau disease.

Percutaneous ultrasound-guided radiofrequency ablation is increasingly being studied in the treatment of renal tumors. Because percutaneous ultrasound-guided radiofrequency ablation is a minimally invasive and nephron-sparing procedure, it is ideally suited for patients with a single kidney, multiple tumors, or contraindications to conventional surgery. We report on a patient with Von Hippel-Lindau (VHL) disease who had multicentric tumors in the single kidney that was successfully treated with percutaneous ultrasound-guided radiofrequncy ablation. The one-year follow-up showed that there was no local recurrence or metastasis. And genetic testing showed the patient had a T to G heterozygotic missense mutation at nucleotide 515 of VHL gene exon 1.

Vessel-Sparing Microsurgical Longitudinal Intussusception Vasoepididymostomy to Treat Epididymal Obstructive Azoospermia.

The epididymis is a common site of obstruction in obstructive azoospermia (OA). Vasoepididymostomy has become an important method for the treatment of epididymal OA since 2000. There are two challenges in classic microscopic vasoepididymostomy. First, anastomosis of the vas deferens and epididymis is performed with double-needle sutures. However, there is a lack of good-quality and cost-effective double-needle sutures in China, which leads to increased difficulty and poor success rates of anastomosis. Second, the separation of the vas deferens does not retain vasculature, although the vas deferens vasculature plays an important role in the blood supply to the vas deferens, epididymis, and testis. This affects the blood supply to the anastomotic area and epididymis. Therefore, this team has made innovative improvements to address these problems. Good-quality, cost-effective, single-needle sutures, which are easy to purchase in China and other countries, were used in microsurgical longitudinal intussusception vasoepididymostomy. This can optimize the operation procedure and shorten the operation time while ensuring the success rate of the anastomosis. The surgical method of preserving the vas deferens vessels was innovatively proposed because the etiology of epididymal OA is mostly inflammatory in China. The protection of the blood supply to the vas deferens and epididymis is maximized using microsurgical forceps to separate and protect the vasculature. Patency reached 81.7% in the postoperative follow-up, indicating a better surgical treatment effect.

"Seminal testosterone", rising viewpoint of local spermatogenesis in nonobstructive azoospermia: One center long-term bidirectional cohort study.

Objective: Reproductive hormones are a traditional good method to evaluate spermatogenesis but might not accurately represent local spermatogenesis. To find a more accurate method, seminal reproductive hormones were studied. Methods: A bidirectional cohort study was performed. A total of 126 infertile men from 2018 to 2019 were retrospectively analyzed. They were divided into nonobstructive azoospermia (NOA), oligozoospermia (OLZ) and normal (NOR) groups. A prospective study was conducted on patients in the NOA and OLZ groups for 2 years. Microscopic testicular sperm extraction was performed for NOA patients, who were divided into a focal spermatogenesis group (FS) and an idiopathic azoospermia group (IA). Drug treatment was for OLZ patients, who were divided into a valid group (VA) and an invalid group (IN). The differences in sperm parameters and reproductive hormones were compared. ANOSIM analysis was used between and within groups. Pearson correlation analysis, CO inertia analysis and Proctor's analysis were for relationships. ROC curve for the specificity and sensitivity. Time series analysis was for the trends between hormones and time. Results: The b-FSH, b-LH, s-T and ΔT in the NOA group were significantly higher than those in the OLZ and NOR groups. However, the s-FSH, s-E2, s-P, ΔFSH, ΔLH, ΔP and ΔE2 were lower. Thirty-one NOA patients underwent MTSE, of whom 12 had sperm (FS) and 19 had no sperm (IA). The s-FSH and s-E2 of the FS group were higher than those of the IA group. Twenty-six OLZ patients completed 30 days of treatment, of which 11 had an improved sperm count (VA) and 15 had no (IN). The ΔT of the VA group was higher than that of the IN group. After follow-up for 2 years, 18 patients' results showed that b-FSH, b-LH and s-T were different over time, with delays of 19, 3 and -19 days. SC is closely related to pH, s-FSH, s-LH, s-E2, s-P, s-T, b-FSH, b-LH, ΔFSH, ΔLH, ΔP, ΔE2 and ΔT. There were complex common trends and relationships between different kinds of hormones. s-FSH, s-LH, s-E2, s-P, s-T, b-FSH and b-LH were useful to judge spermatogenesis, of which s-T, b-FSH and b-LH were more sensitive. If s-T, b-FSH and b-LH reached 64.4, 9.4 and 4.7, respectively, their prediction performance was the strongest. Conclusion: Seminal testosterone is sensitive for judging local spermatogenesis in nonobstructive azoospermia patients, which may be the direction of local spermatogenesis in nonobstructive azoospermia. Clinical trial registration: http://www.chictr.org.cn/index.aspx, identifier ChiCTR2200060463.

AAV-mediated gene therapy produces fertile offspring in the Lhcgr-deficient mouse model of Leydig cell failure.

Leydig cell failure (LCF) caused by gene mutation results in testosterone deficiency and infertility. Serum testosterone levels can be recovered via testosterone replacement; however, established therapies have shown limited success in restoring fertility. Here, we use a luteinizing hormone/choriogonadotrophin receptor (Lhcgr)-deficient mouse model of LCF to investigate the feasibility of gene therapy for restoring testosterone production and fertility. We screen several adeno-associated virus (AAV) serotypes and identify AAV8 as an efficient vector to drive exogenous Lhcgr expression in progenitor Leydig cells through interstitial injection. We observe considerable testosterone recovery and Leydig cell maturation after AAV8-Lhcgr treatment in pubertal Lhcgr-/- mice. Of note, this gene therapy partially recovers sexual development, substantially restores spermatogenesis, and effectively produces fertile offspring. Furthermore, these favorable effects can be reproduced in adult Lhcgr-/- mice. Our proof-of-concept experiments in the mouse model demonstrate that AAV-mediated gene therapy may represent a promising therapeutic approach for patients with LCF.

Transplantation of encapsulated human Leydig-like cells: A novel option for the treatment of testosterone deficiency.

Previous studies have demonstrated that the transplantation of alginate-poly-ʟ-lysine-alginate (APA)-encapsulated rat Leydig cells (LCs) provides a promising approach for treating testosterone deficiency (TD). Nevertheless, LCs have a limited capacity to proliferate, limiting the efficacy of LC transplantation therapy. Here, we established an efficient differentiation system to obtain functional Leydig-like cells (LLCs) from human stem Leydig cells (hSLCs). Then we injected APA-encapsulated LLCs into the abdominal cavities of castrated mice without an immunosuppressor. The APA-encapsulated cells survived and partially restored testosterone production for 90 days in vivo. More importantly, the transplantation of encapsulated LLCs ameliorated the symptoms of TD, such as fat accumulation, muscle atrophy and adipocyte accumulation in bone marrow. Overall, these results suggest that the transplantation of encapsulated LLCs is a promising new method for testosterone supplementation with potential clinical applications in TD.