Tumor necrosis factor-alpha (TNF-α) is an important modulator of neuroinflammation, secreted from activated glial cells in response to intraocular stress. The purpose of this study was to investigate the clinical factors associated with elevated TNF-α and its level in aqueous humor of patients with open-angle glaucoma (OAG). Aqueous humor was collected from 73 OAG eyes, and TNF-α level was analyzed using the singleplex bead immunoassay method. Patients were divided into TNF-α-positive and TNF-α-negative groups according to the TNF-α level of 10 pg/mL, and baseline clinical characteristics were compared. The TNF-α-positive group showed higher baseline IOP, greater IOP fluctuation, and higher systolic blood pressure than the TNF-α-negative group (p = 0.007, p < 0.001, and p = 0.009, respectively). In the multivariate logistic regression analysis, IOP fluctuation (p = 0.037) and systolic blood pressure (p = 0.016) were all independently associated with positive TNF-α level. In normal-tension glaucoma (NTG) patients, presence of central scotoma (p = 0.029) was significantly associated with positive TNF-α level. In conclusion, positive TNF-α level in OAG patients was associated with greater IOP fluctuation and higher systolic blood pressure. In NTG patients, positive TNF-α level was associated with the presence of central scotoma. IOP factors and vascular factors, including blood pressure and presence of central scotoma, may indicate glaucoma pathogenesis related to TNF-α elevation in OAG patients.