The aim of this study was to identify if cells obtained from periodontal granulation tissue possess embryonic stem cell properties and osteogenic capacities in vitro. Periodontal granulation tissue was removed from one furcation and one infrabony defect (FGTC/IGTC-furcation/infrabony defect derived granulation tissue cells) of six patients. The extracted tissues were treated with collagenase/dispase solution, cultured and passaged twice, while a fraction of them was bacteriologically analyzed. Upon reaching confluence, total RNA was extracted, followed by cDNA synthesis and real-time PCR analysis. Gene expression levels of collagen type I, alkaline phosphatase (ALP), and the embryonic stem cell markers Nanog, Oct-4, Rex-1 and Sox-2 were measured, calibrated against the housekeeping gene GAPDH. Further, osteogenic differentiation was induced. Mineralized matrix formation was confirmed by von Kossa staining, and ALP activity was measured colorimetrically. The total bacterial load amounted to 9.4 ± 14.6 × 106 counts/mg of tissue for IGTC, and 11.1 ± 6.1 × 106 counts/of tissue for FGTC. Among the embryonic stem cell markers (FGTC/IGTC), Nanog was most highly expressed (3.48 ± 1.2/5.85 ± 5.7), followed by Oct-4 (1.79 ± 0.69/2.85 ± 2.5), Sox-2 (0.66 ± 0.3/1.26 ± 1.4) and Rex-1 (0.06 ± 0.0/0.04 ± 0.0). The osteogenic differentiation process was positive in both FGTC and IGTC, judged by increased von Kossa staining, and elevated ALP activity and gene expression. This study provides evidence that infected periodontal granulation tissue harbors cells expressing embryonic stem cell markers, and exhibiting osteogenic capacities when in culture in vitro.