To determine the roles of secretory phospholipase A2-IIa (sPLA2-IIa) in the inflammatory responses of the compromised ocular surface. Conjunctival impression cytology (IC) samples and tears were collected from patients with mild to severe non-Sjogren's dry eye disease (DED) and normal controls. The IC samples were analyzed for transcription of sPLA2-IIa and inflammatory cytokine/chemokine genes using quantitative real-time RT-PCR (qRT2-PCR) and pathway-focus PCR-array. The tear samples were analyzed for 13 inflammatory cytokines and chemokines with Millipore 13-Plex kit. Finally, sPLA2-IIa-treated human conjunctival epithelial cell (HCjE) cultures were analyzed with a pathway-focused PCR array. Transcription of sPLA2-IIa was significantly increased in severe DED patients as compared to those of mild DED patients and normal controls. The transcription of inflammatory cytokines (IL-1β, IL-4, IL-6, IL-17, TNF-α, IFN-γ), chemokines (IL-8, CXCL10, CXCL11, CXCL-14, CCR6, LTB) and matrix metalloproteinase 9 (MMP9) were simultaneously increased in the same IC samples of DED. Concentrations of IL-6 and IL-8 in tears were significantly higher in DED patients than those of the controls and positively correlated to DED severity scores. On the other hand, IL-2, IL-4, IL-10, IL-12 and IFN-γ were significantly lower in DED patients than those in the controls and inversely correlated to DEWS scores. Single treatment of sPLA2-IIa, IL-1β or TNF-α of HCjE cells induced minimal to no PGE2 production. When sPLA2-IIa was added to HCjE cells that were pre-treated with pro-inflammatory cytokines (TNF-α or IL-1β), significant stimulation of PGE2 production was observed, concurrent with the extensive transcriptional changes of many inflammatory cytokines/chemokines and their receptors. sPLA2-IIa activity was elevated and not only associated with inflammatory changes in DED patient samples, but was also found to cooperate with TNF- α and IL-1β to induce inflammatory response in human conjunctival epithelial cells. Understanding the roles of sPLA2-IIa in ocular surface inflammation may lead to better strategies for the treatment of chronic inflammation associated with DED and other ocular inflammatory conditions.