Edenfield R Clayton, Potter Samuel B, Crow Krista S, Cho In Ki, Easley Kristen F, Lara Nathalia L M, Waters Elizabeth S, Hedges Jason C, Lo Jamie O, Dobrinski Ina, Koval Michael, Easley Charles A
Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, Georgia; Regenerative Bioscience Center, University of Georgia, Athens, Georgia; Division of Reproductive and Developmental Sciences, Oregon National Primate Center, Oregon Health and Science University, Beaverton, Oregon.
Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, Georgia.
F S Sci. 2025 Jul 29. doi: 10.1016/j.xfss.2025.07.005.
This study aims to investigate the impact of alcohol on blood-testis barrier (BTB) integrity using a novel in vitro model and to elucidate potential nonhormonal mechanisms underlying alcohologenic reversible azoospermia.
Primary rhesus macaque Sertoli cells were exposed to ethanol to evaluate dose-dependent effects on BTB integrity. Barrier function was assessed through electrical resistance and permeability assays, with recovery evaluated after a 48-hour withdrawal period. Gene expression of Sertoli cells and tight junction markers and cytokine levels associated with barrier degradation were analyzed.
Primary rhesus macaque Sertoli cells were used.
Clinically relevant concentrations (10, 60, and 100 mM), equivalent to 1-2 drinks to requiring hospitalization, of ethanol were used.
The main outcome measures were BTB integrity and permeability, assessed via transepithelial electrical resistance and Dye Flux assays, recovery of barrier function after ethanol withdrawal, gene expression changes in Sertoli cells and tight junction markers, and cytokine levels associated with barrier impairment. These metrics evaluated the functional and molecular impacts of in vitro ethanol exposure on the BTB.
Ethanol exposure was associated with a dose-dependent decrease in BTB integrity, with partial recovery observed at lower concentrations (10 and 60 mM) after 48 hours of withdrawal, but not at 100 mM. Additionally, ethanol increased the expression of key Sertoli cells and tight junction genes such as SOX9 and CLDN3, and elevated cytokines associated with barrier degradation at higher concentrations.
Clinically relevant ethanol concentrations reversibly disrupt BTB function through a nonhormonal mechanism, with partial recovery at lower concentrations. These findings provide novel insights into the role of BTB dysfunction in alcohologenic reversible azoospermia.
本研究旨在使用一种新型体外模型研究酒精对血睾屏障（BTB）完整性的影响，并阐明酒精性可逆性无精子症潜在的非激素机制。
将原代恒河猴支持细胞暴露于乙醇中，以评估其对BTB完整性的剂量依赖性影响。通过电阻和通透性测定评估屏障功能，并在撤药48小时后评估恢复情况。分析支持细胞、紧密连接标志物的基因表达以及与屏障降解相关的细胞因子水平。
使用原代恒河猴支持细胞。
使用临床相关浓度（10、60和100 mM）的乙醇，相当于1 - 2杯酒至需要住院治疗的剂量。
主要观察指标为BTB完整性和通透性，通过跨上皮电阻和染料通量测定进行评估；乙醇撤药后屏障功能的恢复情况；支持细胞和紧密连接标志物的基因表达变化；以及与屏障损伤相关的细胞因子水平。这些指标评估了体外乙醇暴露对BTB的功能和分子影响。
乙醇暴露与BTB完整性的剂量依赖性降低相关，在撤药48小时后，较低浓度（10和60 mM）下观察到部分恢复，但100 mM时未恢复。此外，乙醇增加了关键支持细胞和紧密连接基因如SOX9和CLDN3的表达，并在较高浓度下升高了与屏障降解相关的细胞因子水平。
临床相关浓度的乙醇通过非激素机制可逆地破坏BTB功能，较低浓度下有部分恢复。这些发现为BTB功能障碍在酒精性可逆性无精子症中的作用提供了新的见解。
