Varicocele is a medical condition in which men’s scrotum veins widen, potentially interfering with testicular hemodynamics.^[1] It is estimated to affects 15% to 20% of the world’s male population, and is thought to be responsible for more than 40% of male infertility.^{[2, 3]} Varicocele has a significant impact on the quality of sperm and semen. This condition is caused by instability of the hemodynamics.^{[4, 5]} DNA fragmentation is also known to play a big role in the development of varicocele, which is caused by the process of apoptosis, packaging abnormal chromatin, as well as oxidative stress.^[6] In addition, varicocele is known to damage sperm DNA function through improved oxidative stress and hypoxic effects on the testes.^[7]

Various risk factors have been identified in the development of varicocele, such as damage to the venous valves, physical activity, weight, hormonal imbalance, smoking habits, and genetic factors.^{[8, 9]} Patients with varicocele are often underdiagnosed, either when presenting with pain or during fertility evaluations.^[10] Although most cases of varicocele are asymptomatic, some patients present with a feeling of heaviness in the scrotum or even testicular atrophy.^[1]

Mutation genetics, such as single nucleotide polymorphism (SNP), are also linked with varicocele and infertility.^[11] Previous research has shown that varicocele is the only component of strong genetics, with evidence that the condition can be naturally derived, indicating that genetics play an important role in the development of this disease.‌^{[12, 13]} One of the interesting findings is the improvement in the expression of the enzyme nitric oxide synthase (NOS) in patients’ varicocele, which causes relaxation in cells, muscles, and blood vessels.^[14] Furthermore, the development of varicoceles is influenced by endothelial vascular overload, which is mediated by prokineticin 2 (PK2) and a number of other genes that contribute to varicocele pathophysiology.^{[15, 16]}

Despite these findings, the genetic basis of varicocele remains poorly understood. Current studies have mainly focused on clinical manifestations and general pathophysiological mechanisms, while the role of specific genetic biomarkers, particularly genes involved in oxidative stress and angiogenesis, is still underexplored and not clearly validated.^[17] This lack of precise molecular understanding contributes to delayed diagnosis and limited options for targeted therapy. Therefore, further investigation into genetics associated with varicocele is essential to establish reliable biomarkers that may improve early detection, risk stratification, and personalized treatment strategies. Various genome database platforms are used to investigate single nucleotide polymorphisms (SNPs) that have the potential to regulate angiogenesis and oxidative stress, making them biomarkers and treatment targets.^[18–20]

While the results from this study are still limited, the findings provide valuable insights into the potential role of genetic biomarkers in the diagnosis and treatment of varicocele. The aim of this study was to look into genes that may influence the development of varicoceles in the hopes of laying the groundwork for future research. This could eventually lead to the discovery of new biomarkers for diagnosing varicocele that can be used as therapeutic targets in the future.

Varicocele is still a medical condition with unknown causes, though genetic factors are becoming more widely recognized as influencing its development.^[20] This study aimed to identify key genetic biomarkers associated with varicocele using integrated bioinformatics approaches, including STRING, MCODE, and CytoHubba, which have been effectively applied in related conditions such as male infertility and liver fibrosis.^{[21, 22]} Our analysis identified three significant candidate biomarkers: IL-1B, BCL2, and MMP-9.

Interleukin-1 beta (IL-1B) is a proinflammatory cytokine critically involved in vascular pathology, including varicocele, characterized by abnormal venous dilation in the scrotum. It mediates inflammatory signaling through the NF-κB pathway, contributing to endothelial dysfunction and vascular inflammation observed in related diseases like atherosclerosis.^[23] Such mechanisms plausibly underpin varicocele progression via chronic vascular inflammation and increased endothelial permeability, which may exacerbate venous damage.^[24] Additionally, IL-1B promotes oxidative stress by stimulating reactive oxygen species (ROS) production, impairing endothelial function and exacerbating venous hypertension typical of varicocele.^[25] Its role in angiogenesis, through upregulation of VEGF in macrophages, further implicates IL-1β in abnormal vessel formation.^[26] These multifaceted functions position IL-1B as a promising biomarker and potential therapeutic target, especially for inflammation-driven varicocele.

B-cell lymphoma 2 (Bcl-2), an anti-apoptotic protein, regulates cell survival and is linked to angiogenesis, oxidative stress, and autophagy. Increased BCL2 expression has been observed in spermatic veins of varicocele patients, suggesting enhanced endothelial cell survival that may contribute to pathological angiogenesis and venous abnormalities.^[27] The imbalance between BCL2 and pro-apoptotic BAX exacerbates oxidative damage to vascular and testicular tissues, potentially impairing sperm quality and fertility.‌^[28] Furthermore, BCL2’s interaction with hypoxia-inducible factors such as HIF-1α^[29] reflects a cellular adaptive response to hypoxic stress caused by disrupted blood flow, stimulating angiogenesis and cell proliferation.^[30] These insights highlight BCL2’s critical role in maintaining venous homeostasis under varicocele-associated stress.

MMP-9 is a matrix metalloproteinase involved in extracellular matrix remodeling critical for vascular integrity.‌^[31] Genetic polymorphisms in MMP-9 have been linked to varicose veins, a pathophysiologically related condition, and may similarly predispose individuals to varicocele through altered extracellular matrix degradation leading to vessel dilation and valve incompetence.^[32]MMP-9 activation by ROS generated under heat stress and venous congestion damages collagen and elastin fibers, reducing vascular elasticity and exacerbating varicocele progression.‌^[32] Moreover, dysregulated MMP-9 activity disrupts angiogenesis, leading to inefficient collateral vessel formation and persistent venous congestion. The imbalance between MMP-9 and its inhibitor TIMP2 further aggravates this pathological process, underscoring MMP-9’s central role in varicocele pathogenesis.

Previous studies have investigated the genetic underpinnings of male infertility, either through targeted analysis of specific polymorphisms, such as the NOS3 variants linked to oxidative stress in varicocele patients, or by identifying broader infertility-associated genes using genome-wide bioinformatics approaches.^[33] However, these efforts did not specifically isolate or prioritize genetic biomarkers uniquely relevant to varicocele as a distinct pathological entity. The bioinformatics study, for instance, focused on general male infertility without distinguishing varicocele-associated molecular mechanisms, identifying genes like TEX11 and SYCP3 primarily involved in meiosis and spermatogenesis. While previous studies have noted the involvement of these proteins in vascular disorders, our integrative bioinformatics approach uniquely prioritizes IL1B, BCL2, and MMP-9 as varicocele-specific biomarkers by combining multi-database gene screening with network and functional analyses. This provides novel insights into their potential interplay in varicocele development, bridging gaps between genetic predisposition and molecular mechanisms. However, this study’s reliance on in silico data limits direct clinical applicability without experimental validation.

This study’s limitations include its reliance solely on computational bioinformatics analysis without experimental validation, which limits the ability to confirm the actual expression or functional impact of the identified biomarkers IL1B, BCL2, and MMP-9 in varicoceles. The absence of patient-derived clinical samples and functional assays means that the biological roles of these genes in varicocele pathogenesis remain hypothetical. Additionally, the cross-sectional design limits interpretation of causality and dynamic gene expression changes over time or in response to treatment. To address these limitations, future studies should include molecular validation using patient tissues, assess biomarker specificity through case-control comparisons, and apply longitudinal designs to evaluate changes post-varicocelectomy. Incorporating predictive modeling tools such as ROC analysis and regression frameworks would also help determine the diagnostic and prognostic utility of these candidate genes in clinical practice.

This study successfully identified 79 varicocele-associated genes by integrating data from OpenTarget, DisGeNET, and NCBI databases. Subsequent protein-protein interaction network analysis and module detection pinpointed three key biomarkers, IL1B, BCL2, and MMP-9, which are critically involved in varicocele pathogenesis through mechanisms such as inflammation, oxidative stress, angiogenesis, and vascular damage. These biomarkers hold promise for improving varicocele diagnosis and developing targeted therapeutic strategies. Overall, our findings enhance the molecular understanding of varicocele and lay the groundwork for future research aimed at more precise and effective clinical interventions.

This research was conducted without any external financial support.

M.F. conceptualized and designed the study, conducted the majority of data analysis, interpreted the results, and wrote the original draft of the manuscript. H.F.A. assisted in data collection, contributed to the bioinformatics analysis, and participated in reviewing and editing the manuscript. Both authors read and approved the final manuscript and agree to be accountable for all aspects of the work.

The authors have declared that no competing interests exist.