Exploring the Contribution of Interleukin-12A Genetic Polymorphisms to Pterygium Risk

Preliminary analyses demonstrated that the mean age of individuals with pterygium was comparable to that of the control group, with no statistically significant differences observed ( p =0.9006). This finding remained consistent when age was stratified using 60 years as a threshold ( p =0.2509), indicating that age was unlikely to confound the observed associations. Moreover, sex was deliberately matched at the recruitment stage to control for potential confounding. As a result, the proportions of male and female participants were equivalent between cases and controls, yielding no detectable difference ( p =1.0000). Collectively, these design strategies enhanced the internal validity of the study and minimized the occurrence of selection bias. The genotype distribution patterns of IL-12A rs568408 and rs2243115 polymorphisms in 160 pterygium patients and 320 control subjects are presented in Table II . In the control group, both polymorphisms conformed to Hardy-Weinberg equilibrium ( p =0.2165 and 0.5533, respectively; Table II), indicating no significant deviation from expected genotype frequencies. For IL-12A rs568408, the distributions of GG, AG, and AA genotypes among pterygium patients were 71.2%, 24.4%, and 4.4%, respectively, which were comparable to 74.7%, 22.5%, and 2.8% observed in the control group, with no statistically significant difference detected ( p for trend=0.5727). Although the proportions of the AG heterozygous and AA homozygous variant genotypes were modestly elevated in cases, these differences did not reach statistical significance (OR=1.14 and 1.63, 95%CI=0.72-1.78 and 0.59-4.49, p =0.6603 and 0.4950, respectively). Furthermore, analyses based on recessive and dominant genetic models also failed to demonstrate any significant association (OR=1.58 and 1.19, 95%CI=0.58-4.33 and 0.78-1.82, p =0.5292 and 0.4870, respectively; Table II, top section). Similarly, the genotype distribution of IL-12A rs2243115 did not differ significantly between cases and controls ( p for trend=0.5471). Consistent with this observation, neither recessive nor dominant model analyses revealed any significant relationship between rs2243115 variants and susceptibility to pterygium (Table II). Collectively, these findings suggest that neither IL-12A rs568408 nor rs2243115 polymorphisms are significantly associated with the risk of developing pterygium in this study population. N: Number; OR: odds ratio; CI: confidence interval; aBased on Chi-square test with Yates’ correction; bBased on Fisher’s exact test; p HWE : whether the frequencies in controls are consistent with Hardy-Weinberg Equilibrium. To further corroborate the observations reported in Table II, allelic frequency analyses were conducted for IL-12A rs568408 and rs2243115, with the results summarized in Table III . Consistent with the genotype-based findings, these analyses provided no evidence supporting an association between these polymorphisms and susceptibility to pterygium (OR=1.21 and 1.27, 95%CI=0.84-1.76 and 0.81-2.01, p =0.3527 and 0.3581, respectively; Table III). Specifically, the frequency of the variant A allele at rs568408 was 16.6% in patients, compared with 14.1% in controls. Similarly, the G allele frequency at rs2243115 was 10.3% among cases and 8.3% among controls. These comparable distributions between the two groups further reinforce the conclusion that neither IL-12A rs568408 nor rs2243115 polymorphisms significantly influence the risk of developing pterygium in the investigated population. N: Number; OR: odds ratio; CI: confidence interval; a Based on Chi-square with Yates’ correction test.

Recruitment of pterygium cases and matched controls. Participant recruitment for individuals with and without pterygium was carried out in accordance with a study protocol approved by the Institutional Review Board of China Medical University Hospital (CMUH111-REC1-176). The overall study framework, including its research objectives, underlying hypotheses, and experimental procedures, was subjected to comprehensive ethical and methodological evaluation prior to implementation. Written informed consent was obtained from all participants before enrollment. A total of 160 patients with clinically confirmed pterygium were recruited, alongside 320 control subjects with no prior history of the disease. All participants were of Taiwanese ethnicity and voluntarily completed a standardized questionnaire designed to collect demographic characteristics and lifestyle-related information. In parallel, peripheral blood specimens were obtained from each individual for subsequent genetic analyses. To ensure the validity and comparability of the control cohort, strict exclusion criteria were enforced. Individuals with a documented history of pterygium, endometriosis, uterine leiomyoma, or any malignancy were excluded from the control group. The demographic features such as age and gender of all participants are summarized in Table I . N, Number; SD, standard deviation; a Based on unpaired Student’s t-test; b Based on 2 X 2 Chi-square test with Yates’ correction. Genotyping of IL-12A polymorphisms. Genomic DNA was extracted from peripheral blood leukocytes of pterygium patients and control subjects using a commercially available DNA isolation kit (Blossom, Taipei, Taiwan, ROC), following established biomedical protocols ( 40 - 42 ). Genotyping of the IL-12A rs2243115 and IL-12A rs568408 polymorphisms was performed using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methodology, and the primers, PCR protocols, restriction enzyme selections were as previous published ( 43 , 44 ). All the genotyping process was conducted independently by at least two researchers in a double-blind manner to ensure accuracy. Notably, all genotyping results achieved a 100% success rate, with complete concordance across all samples. Statistical analysis of genotype and allele distributions. The genotype distributions of each IL-12A polymorphism among pterygium cases and control subjects were evaluated for conformity with Hardy-Weinberg equilibrium using the chi-square goodness-of-fit test. Differences in demographic characteristics, as well as genotypic and allelic frequencies of IL-12A variants between groups were analyzed using Pearson’s chi-square test (2×2 or 2×3 contingency tables, as appropriate). To quantify the relationship between IL-12A polymorphisms and susceptibility to pterygium, odds ratios (ORs) and corresponding 95% confidence intervals (CIs) were calculated through unconditional logistic regression analysis. Statistical significance was defined as a two-tailed p -value of less than 0.05.

No artificial intelligence tools, including large language models or machine learning software, were used in the preparation, analysis, or presentation of this manuscript.

Conceptualization: Yang YC, Bau DT and Tsai CW; Collection: Hsia NY and Hu PS; Data curation: Yang YC and Hsia TC; Genotyping: Chang WS, Yang YC and Tsai CW; Statistics: Yang YC and Tsai CW; Project administration: Yang YC and Bau DT; Supervision: Bau DT, Tsai CW and Chang WS; Validation: Hsia TC and Bau DT; Original draft: Yang YC, Bau DT and Tsai CW; Review and editing: Bau DT and Tsai CW; All Authors have read and agreed to the published version of the manuscript.

Notably, pterygium demonstrates several tumor-like features, including dysregulated fibrovascular proliferation, extension into adjacent corneal tissue, extensive extracellular matrix remodeling, heightened inflammatory responses, and a pronounced propensity for postoperative recurrence ( 45 - 47 ). Its characteristic wedge-shaped extension across the corneoscleral junction further mimics the invasive growth patterns commonly observed in solid tumors ( 48 ). Given these neoplasm-like properties, a number of molecular epidemiological investigations have explored the relationship between genetic variants of IL-12 and susceptibility to various malignancies. In particular, IL-12A polymorphisms, including rs568408 and rs2243115, have been implicated in the risk of multiple cancers such as breast cancer ( 49 ), oral cancer ( 50 ), nasopharyngeal carcinoma ( 51 ), thyroid cancer ( 52 ), and colorectal cancer ( 53 ). Notably, a recent study by Chen and his colleagues on the contribution of IL-12A to nasopharyngeal carcinoma has further demonstrated that IL-12A genetic variants are significantly associated with susceptibility to nasopharyngeal carcinoma risk in a Taiwanese population ( 51 ). These findings support the notion that IL-12A polymorphisms may closely related to determining personal disease susceptibility through modulation of host immune responses. Despite this growing body of evidence, the potential contribution of IL-12 genetic variants to pterygium development has not been previously addressed. To bridge this knowledge gap, we conducted a hospital-based case-control study to evaluate the role of IL-12A rs568408 and rs2243115 polymorphisms in determining susceptibility to pterygium. Our findings indicated that individuals harboring either heterozygous or homozygous variant genotypes at these loci exhibited a modest, yet statistically non-significant, elevation in risk, with odds ratios ranging from 1.14 to 2.07 (Table II). Furthermore, allelic frequency analyses corroborated these observations, demonstrating that neither polymorphism serves as a reliable molecular biomarker for predicting pterygium susceptibility (Table III). Pterygium and dye eye disease frequently coexist and share overlapping pathogenic mechanisms ( 54 , 55 ). A central link between these two conditions lies in chronic conjunctival inflammation driven by environmental stressors, including prolonged ultraviolet exposure, low-humidity conditions, strong winds, and airborne particulates ( 24 ). These external insults not only promote fibrovascular proliferation characteristic of pterygium but also exacerbate tear film instability and ocular surface damage, thereby intensifying dry eye symptoms ( 56 ). Although direct evidence demonstrating significant alterations of IL-12 during pterygium progression remains limited, insights from dry eye disease research suggest a potentially elevated expression of IL-12 in the tears of patients ( 57 ). Functional changes in dry eye have been shown to enhance neuro-immune crosstalk through upregulation of innate immune pathways, including TLR7/9, STING, and S100A8/A9 signaling cascades, accompanied by increased production of pro-inflammatory cytokines such as IL-12, interferons, and IL-23 ( 58 ). These findings imply that IL-12-mediated immune responses may contribute, at least indirectly, to the inflammatory microenvironment shared by both conditions. Nevertheless, the precise role of IL-12 in pterygium pathogenesis, particularly at the protein and mRNA expression levels, remains to be elucidated. Furthermore, whether genetic variations in IL-12A influence individual susceptibility to pterygium, as well as how broader immune regulatory networks participate in disease initiation and progression, are questions that warrant further investigation. Addressing these gaps will require integrated efforts from both clinical and translational research to clarify the immunological underpinnings of pterygium. In conclusion, our study suggests that the variant alleles at IL-12A rs568408 orrs2243115 polymorphisms may not serve as reliable biomarkers for predicting pterygium susceptibility. Future studies incorporating larger sample sizes, functional assays, and multi-omics approaches are warranted to further elucidate the immunological mechanisms underlying pterygium development.

Pterygium is a prevalent degenerative disorder of the ocular surface that occurs predominantly in regions characterized by intense solar radiation, arid climates, and frequent exposure to airborne particulates, particularly in areas near the equator ( 1 ). Although the global prevalence has been estimated to be approximately 12% ( 2 ), substantial geographic and ethnic variability has been documented, with reported rates ranging from as low as 1% to over 30% ( 3 ). Despite its high prevalence, the precise molecular and cellular mechanisms governing its pathogenesis, as well as the factors contributing to postoperative recurrence, remain incompletely understood ( 4 ). Epidemiological evidence suggests that both environmental exposures and host-related characteristics contribute synergistically to disease development. Among these, prolonged ultraviolet radiation represents a major etiological factor ( 5 - 7 ), while additional contributors include a positive family history ( 8 , 9 ), male sex ( 10 , 11 ), increasing age ( 10 , 12 ), engagement in outdoor or rural occupations ( 12 ), and chronic exposure to environmental irritants such as dust and sand ( 13 , 14 ). These factors collectively indicate a multifactorial origin involving cumulative external insults and intrinsic susceptibility. From a mechanistic perspective, immune-related processes play a central role in the initiation and progression of pterygium ( 15 , 16 ). Chronic inflammation, angiogenesis, and fibrovascular proliferation are key pathological features of pterygium, and accumulating evidence supports the notion that pterygium may arise from a state of localized immune dysregulation ( 17 - 19 ). Histopathological analyses frequently reveal infiltration of inflammatory cells, including lymphocytes, accompanied by elevated production of cytokines, which in turn promote excessive fibrovascular tissue growth in pterygium ( 20 ). Furthermore, sustained ultraviolet exposure can disrupt the structural integrity of the conjunctiva and trigger aberrant activation of the local immune system ( 21 ). This process induces a persistent, wound-healing-like response with inflammatory characteristics, ultimately leading to the formation of hyperplastic tissue ( 22 , 23 ). Although pterygium is not classified as a classical autoimmune disease, alterations in the ocular surface immune microenvironment, such as those associated with Dry eye disease, can exacerbate chronic inflammation and accelerate disease progression ( 24 ). Collectively, these observations underscore a critical link between immune imbalance and the pathophysiology of pterygium. Interleukin 12 (IL-12) is a heterodimeric cytokine consisting of two distinct subunits, p35 and p40 ( 25 ). Functioning as a pivotal regulator of immune responses, IL-12 is critically involved in shaping adaptive immunity ( 26 ). Evidence derived from murine models has demonstrated that exogenous administration of IL-12 can effectively suppress tumor progression by attenuating cellular proliferation, invasion, and metastatic potential ( 27 - 31 ). IL-12 is predominantly secreted by antigen-presenting cells, including dendritic cells, monocytes, macrophages, and B lymphocytes ( 32 ). Mechanistically, the effects of IL-12 are largely mediated through the induction of effector cells, such as natural killer (NK) cells, natural killer T (NKT) cells, and T lymphocytes, to produce pro-inflammatory cytokines, including interferon (IFN) and tumor necrosis factor (TNF) ( 33 ). The post-surgery administration of IFN- 2b can effectively prevent pterygium recurrence ( 34 - 36 ). In addition, IL-12 enhances cytotoxic activity mediated by NK cells and T lymphocytes, particularly CD8⁺ T cells, even within immunosuppressive tumor microenvironments ( 37 ). It also contributes to T cell proliferation and promotes the differentiation and adhesion of CD4⁺ T helper 1 (Th1) cells ( 38 , 39 ). Given its potent immunoregulatory properties, IL-12 has emerged as a promising candidate for cancer immunotherapy, supported by both preclinical and clinical investigations. In the present study, we conducted a pilot epidemiological analysis to assess the potential association of IL-12A genetic polymorphisms, including rs568408 and rs2243115 (illustrated in Figure 1 ), with susceptibility to pterygium in a Taiwanese population. The study cohort comprised 160 patients diagnosed with pterygium and 320 age- and sex-matched control subjects. Physical map for IL-12A rs568408 and rs2243115 polymorphic sites.

The Authors declare no conflicts of interest regarding this study.

The Authors would like to acknowledge the Tissue-Bank of China Medical University Hospital for their invaluable technical support. Furthermore, the authors would like to extend their gratitude to all the study participants, as well as the doctors, nurses, and colleagues who contributed to the study. This study is supported by the grant from Asia University and China Medical University Hospital (ASIA-112-CMUH-10).