As a part of innate immune defense, the role of mast cells during viral replication has been incompletely understood. In this study, we characterized and compared the responses of the human mast cell line, LAD2, and human lung epithelial cell line, Calu-3, against three influenza A virus strains; A/PR/8/34 (H1N1), A/WS/33 (H1N1) and A/HK/8/68 (H3N2). We found that there were strain-dependent mast cell responses, and different profiles of cytokine, chemokine and antiviral gene expression between the two cell types. All three strains did not induce histamine or β-hexosaminidase release in LAD2. A/HK/8/68 induced release of prostaglandin D2 in LAD2, whereas A/PR/8/34 and A/WS/33 did not. We found that, among those examined, only CCL4 (by A/PR/8/34) was statistically significantly released from LAD2 cells. Furthermore, there was increased mRNA expression of viral recognition receptors (RIG-I and MDA5) and antiviral protein, viperin, but levels and kinetics of the expression were different among the cell types, as well as by the strains examined. Our findings highlight the variability in innate response to different strains of influenza A virus in two human cell types, indicating that further investigation is needed to understand better the role of mast cells and epithelial cells in innate immunity against influenza A viruses.