Adipocytes were recently identified as an important source of endocrine and paracrine mediators, regulating the metabolism and activity of various cell types and body functions. 3T3-L1 preadipocytes are useful model for physiological, pharmacological and cell signaling studies. Differentiation of 3T3-L1 murine fibroblasts into adipocyte-like cells was conducted in presence of IBMX, dexamethasone and insulin and demonstrated by Oil Red O staining of accumulated lipid droplets. Using Inflammatory Multi- Analyte Cytokines ELISArray Kit we investigated the release of cytokines under basal conditions, after PGF_2α treatment for 24 hours to induce pro-inflammatory phenotype, and after PGF_2α treatment and incubation in the presence of L-C-Propargylglycine (PGG, 1 mmol/l), a selective inhibitor of cystathionine-gamma-lyase (CSE). The last combination was used to explore the role of H₂S, released from CSE, for cytokine and H₂O₂ release. We found that PGF_2α strongly increased TNF_α secretion from differentiated adipocytes, the latter effect being antagonized by PGG. The CSE inhibitor enhanced IL-6 production and suppressed IL-10 secretion. PGG enhanced H₂O₂ production of in PGF_2α-treated cells. It is concluded that pro-inflammatory phenotype of differentiated 3T3-L1 adipocyte-like cells, induced by PGF_2α is characterized by enhanced TNF_α production which critically depends on the ability of CSE to produce H₂S.