To explore methods using modified tissue enzymatic separations for culturing primary hDPSCs in vitro and further identify the cells produced.

Primary hDPSCs were cultured using the modified tissue enzymatic separation method, and cells were identified by morphology, cell surface markers, and differentiation potential and evaluated using flow cytometry and growth curves.

The hDPSCs were successfully isolated using the modified tissue enzymatic separation method. The morphology of these cells was similar to that of fibroblasts and mesenchymal stem cells, and the growth curve was "S" -type. The results of cell phenotype analysis indicated that the cells were positive for surface markers of mesenchymal stem cells, including CD29, CD44, and CD90, and negative for markers of hematopoietic stem cells, including CD34, CD45, and CD106. The cells were capable of differentiating into multiple cell types.

The modified tissue enzymatic separation method can successful be used to culture primary hDPSCs in vitro.

To analyze the role of TLR4 in innate immune response of dental pulp by comparing the locations and expressions of TLR4 in healthy dental pulp tissue and dental pulp tissue affected by deep caries.

Healthy teeth and teeth affected by deep caries were demineralized and stained with hematoxylin and eosin (HE) to observe the morphology of dental pulp. Immunohistochemistry staining was performed to observe the expressions of TLR4.

Observed under HE staining, dentin tubules of teeth affected by deep caries were damaged with a lot bacteria mass. The expression of TLR4 were located in the odontoblast layer and near the blood vessels in both groups. Positive staining of TLR4 in deep caries pulps (2.10±0.74) were significantly higher than that in healthy teeth (1.25 ± 0.46).

Expression of TLR4 in deep caries pulp is stronger than that in healthy pulp. It suggests that TLR4 may play a role in the innate immune response of deep caries.