An oral environment is constantly exposed to environmental factors and microorganisms. The periodontal ligament (PDL) fibroblasts within this environment are subject to bacterial infection and allergic reaction. However, how these condition affect PDL fibroblasts has yet to be elucidated. PDL fibroblasts were isolated from healthy donors. We examined using reverse transcription-polymerase chain reaction and measuring the intracellular Ca2+ concentration ([Ca2+]i). This study investigated the receptors activated by exogenous bacterial pathogens (Lipopolysaccharide and peptidoglycan) and allergens (German cockroach extract and house dust mite) as well as these pathogenic mediators-induced effects on the intracellular Ca2+ signaling in human PDL fibroblasts. Moreover, we evaluated the expression of pro-inflammatory cytokines (interleukin (IL)-1β, IL-6, and IL-8) and bone remodeling mediators (receptor activator of NF-κB ligand and osteoprotegerin) and intracellular Ca2+-involved effect. Bacterial pathogens and allergic mediators induced increased expression of pro-inflammatory cytokines, and these results are dependent on intracellular Ca2+. However, bacterial pathogens and allergic mediators did not lead to increased expression of bone remodeling mediators, except lipopolysaccharide-induced effect on receptor activator of NF-κB ligand expression. These experiments provide evidence that a pathogens and allergens-induced increase in [Ca2+]i affects the inflammatory response in human PDL fibroblasts.