Oligoasthenozoospermia is one of the most common forms of male reproductive dysfunction. In rodent studies, delivering electroacupuncture (EA) consistently across repeated sessions poses a technical challenge, particularly when combining abdominal midline and hind-limb acupoints, where stable electrode contact is critical. Zhongji (CV3), Guanyuan (CV4), Zusanli (ST36), and Sanyinjiao (SP6) were selected because these points are frequently used in acupuncture protocols for infertility and semen abnormalities, and the abdominal midline plus hind-limb configuration suits standardized electrode pairing. This report describes a practical protocol for establishing an adenine-induced rat model of oligoasthenozoospermia and for administering isoflurane-assisted EA with paired stimulation of CV3-CV4 and ST36-SP6. Male Sprague-Dawley rats received adenine by daily gavage for 28 days to induce reproductive impairment. Subsequently, EA was administered daily for 28 days under inhalational anesthesia, using 3% isoflurane for induction and 1.5% for mask maintenance, to enable stable needle placement and electrode pairing (CV3-CV4 and ST36-SP6) with defined stimulation parameters (2 Hz, 1.5 mA, 30 min). Treatment effects were evaluated via computer-assisted semen analysis for sperm count and motility, supplemented by testicular hematoxylin and eosin (HE) histology. Representative results indicate that adenine exposure reduced sperm count and motility and disrupted seminiferous tubule architecture, while EA was associated with partial recovery in both semen parameters and testicular morphology. This protocol offers a ready reference for studies requiring consistent EA delivery and semen/histological verification in a rat model of oligoasthenozoospermia.

Post-translational modifications (PTMs) of histone variants are the key mechanisms driving epigenetic regulation. They play a crucial role in modulating chromatin structure and gene activity. As epigenetics has become a foundational aspect of molecular biology, the precise characterization of histones is essential for understanding gene regulation, cellular plasticity, and the molecular basis of diseases.Mass spectrometry in combination with liquid chromatography has emerged as the leading technology for mapping and quantification of histone peptidoforms. It offers exceptional sensitivity and resolution to unravel the complexity of the epigenetic landscape. Despite its advantages, the inherent diversity of histone isoforms and PTMs presents analytical challenges, and no universal workflow currently enables comprehensive and unbiased profiling of the overall state of histone modifications.This chapter presents key methodologies for histone analysis using bottom-up proteomics, including protocols for histone extraction, enzymatic digestion, and chemical derivatization prior to LC-MS/MS. It also provides details of specific aspects of LC-MS/MS analysis and outlines strategies for data evaluation and interpretation. The clinical potential of histone proteomics is illustrated through a case study focusing on the characterization of histones in human sperm in the context of male infertility.

The human microbiome, a dynamic endocrine organ, exerts profound systemic influence through the production of bioactive metabolites. While the microbiome-gut-brain axis is well-established, the direct conduit between the gut microbiota and the reproductive system, the Microbiome-Gut-Gonad Axis, remains an emerging paradigm. This review explored cutting-edge evidence to construct a comprehensive model of the Microbiome-Gut-Gonad axis, focusing on the mechanistic roles of specific microbial metabolites in both physiological reproductive function and the pathogenesis of endocrine disorders. We move beyond mere correlation to elucidate how gut-derived molecules, such as short-chain fatty acids (SCFAs), secondary bile acids, and indole derivatives, directly and indirectly modulate the hypothalamic-pituitary-gonadal (HPG) axis by modulating the production of neuropeptides and hormones (Gonadotropin-releasing hormone (GnRH)) that regulate reproductive functions and also steroidogenesis and gametogenesis. We examine novel mechanisms including: the epigenetic regulation of steroidogenic enzymes by butyrate; the modulation of enterohepatic circulation of estrogens by β-glucuronidase-producing bacteria; and the role of tryptophan metabolites as ligands for aryl hydrocarbon receptor (AhR) in ovarian and testicular function. Furthermore, we critically appraise the disruptive potential of dysbiosis-driven metabolite shifts in PCOS, endometriosis, and male infertility, highlighting microbial metabolite signatures as promising exploratory biomarkers that require standardized, multi-center clinical validation before diagnostic use. At present, these signatures should be considered candidate biomarkers only, because external validation cohorts, assay reproducibility, and clinically meaningful estimates of sensitivity, specificity, predictive values, and clinical utility have not yet been established. Therapeutically, we evaluate innovative interventions, including precision probiotics, postbiotics, and dietary strategies targeting specific bacterial guilds, but these approaches remain investigational because current human evidence is still limited and heterogeneous. Finally, by integrating microbial endocrinology into reproductive medicine, this review establishes a new framework for understanding the etiology of reproductive endocrine disorders and paves the way for microbiome-targeted therapeutic avenues. Importantly, the evidence base is tiered: mechanistic statements in this review are drawn primarily from in vitro and animal studies, human disease links are described separately as observational evidence, and interventional claims are limited to early clinical studies and randomized trial summaries.

BackgroundCouples experiencing infertility often seek medical help to achieve a pregnancy using assisted reproductive technology, which might cause stress and affect their quality of life.ObjectivesThe purpose of the current study was to investigate the mutual effects of optimism and stress on fertility-related quality of life among women and their husbands during the process of in vitro fertilization (IVF), as well as the success of the treatment.DesignA cross-sectional study with a prospective outcome assessment design aimed at assessing psychological variables, with a short-term prospective follow-up, among 51 couples undergoing IVF treatment.MethodsParticipants completed questionnaires on fertility-related stress, optimism, and fertility-related quality of life. Two weeks after embryo transfer they were contacted by a researcher to check for pregnancy. The dyadic associations were analyzed using the Actor-Partner Interdependence Mediation Model (APIM).ResultsWomen's fertility-related quality of life correlated negatively with their stress (r=-.67; p<.01) and positively with their husbands' fertility-related quality of life (r=.31; p<.05). Husbands' fertility-related quality of life correlated positively with their optimism (r=.35; p<.05) and negatively with their stress (r=-.56; p<.01) and their wife's stress (r=-.33; p<.01). A positive correlation was found between women's optimism and their husbands' optimism. Women's stress was higher and their fertility-related quality of life lower than those of their husbands. The APIM analysis revealed actor (B=.22, p<.001) and partner (B=.12, p=.02) effects of optimism on fertility-related quality of life. The only predictor of pregnancy was younger age.ConclusionsBoth spouses experience stress during IVF treatments, therefore psychological support should be offered to both during this time, including stress management interventions and couple-based support aimed at strengthening dyadic coping. The positive correlations between women's fertility-related quality of life and that of their husbands, and between women's fertility-related quality of life and their husbands' optimism, highlight the importance of spouses' support for each other.

This study aimed to establish Diagnostic criteria for kidney Yang deficiency syndrome in male infertility(referred to as "this criteria"). A preliminary questionnaire item pool was constructed through literature retrieval, in combination with male infertility-related textbooks, existing national standards, guidelines, and expert consensus documents, as well as expert interviews and group discussions. Two rounds of questionnaire surveys were conducted using the Delphi method to standardize the diagnostic criteria item pool and complete item supplementation and screening. Expert questionnaires were collected using the analytic hierarchy process(AHP) to statistically determine item weights and diagnostic thresholds. Finally, a diagnostic test was conducted by enrolling 105 participants to evaluate the diagnostic criteria, with sensitivity, specificity, accuracy, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio used as indicators of diagnostic efficacy at the established threshold. Based on the results of expert discussions, this criteria was established. After systematic literature screening, 66 articles were included. By incorporating textbooks, national standards, and consensus documents, all items were summarized, merged, and classified, resulting in an initial diagnostic criteria item pool of 39 items. The first round of the Delphi method retained 29 important items, and the second round retained 18 important items. AHP identified three levels of item weights, i.e., high, medium, and low. The primary symptoms for diagnosing kidney Yang deficiency syndrome in MI were finally determined as aversion to cold, soreness and weakness of the lower back and knees, a pale and swollen tongue, and a thin white tongue coating. The secondary symptoms were decreased libido, cold sensation in the lower abdomen or genital region, increased nocturnal urination, and a deep and weak pulse or a deep and slow pulse. Patients presenting all primary symptoms were diagnosed with this syndrome; patients presenting two primary symptoms together with any two manifestations from the secondary symptoms, physical signs, or pulse conditions were also diagnosed with this syndrome. After establishment of this criteria, 105 MI patients from the Department of Andrology, Xiyuan Hospital of China Academy of Chinese Medical Sciences were enrolled. Eligible participants were diagnosed using both the evaluated this criteria and the reference diagnostic criteria. The diagnostic efficacy of this criteria showed a sensitivity of 0.91, specificity of 0.88, and accuracy of 0.89. This study established Diagnostic criteria for kidney Yang deficiency syndrome in male infertility and completed clinical validation, providing a reference basis for the clinical diagnosis of MI with kidney Yang deficiency syndrome.