This study evaluated cell viability and cytokine release in immortalized human oral fibroblasts (hTERT-hNOFs) and keratinocytes (IHOK) exposed to a dental-impregnated gingival retraction cord. To prepare the extracts, dental gingival retraction cords impregnated with aluminum chloride hexahydrate were immersed in a cell culture medium for 24 h at 37 °C. hTERT-hNOFs and IHOK were cultured for 24 h. The cell culture medium was removed and extracts of the dental gingival retraction cords were added. After incubation with the extract solution, cell viability was evaluated using an MTT assay. The levels of the cytokines IL-1α and IL-8 were measured in the supernatants of each cell type. The cell viability after exposure to the extract solution for 10 min exceeded 70 % in both cell types. The ET50 values for hTERT-hNOF and IHOK were 35.75 and 28.98 min, respectively. For IHOK, the IL-1α level was (5.35 ± 5.22) pg/mL at 10 min, (3.58 ± 5.38) pg/mL at 20 min, and (2.85 ± 4.28) pg/mL at 60 min of exposure (p > 0.05). The IL-8 level in IHOK was (67.16 ± 18.70) pg/mL at 10 min, (78.36 ± 7.50) pg/mL at 20 min, and (111.9 ± 26.10) pg/mL at 60 min of exposure (p > 0.05). Cytokine release was not observed from hTERThNOFs. Based on these results, cell viability and cytokine release were confirmed in cells exposed to the impregnated gingival retraction cord. In addition, the application of the extracts to hTERT-hNOF and IHOK during the actual contact time and determination of ET50 may be beneficial for evaluating the biocompatibility of dental-impregnated gingival retraction cords.