Regulatory effects of transcription factor PAX3 on human dental pulp stem cells

Dental pulp stem cells (DPSCs) are promising for regenerative dentistry but undergo replicative senescence during in vitro expansion, leading to diminished proliferative capacity and osteogenic potential. This study aimed to investigate the role of the transcription factor PAX3 in regulating the senescence process of human DPSCs (hDPSCs).

A replicative senescence model of hDPSCs was established through serial passaging (P3 as young cells vs. P11 as senescent cells). The expression of PAX3 and senescence markers (p53, p21) was assessed. The functional role of PAX3 was investigated by knocking it down (si-PAX3) or overexpressing it (OE-PAX3) in hDPSCs, followed by evaluations of senescence-associated β-galactosidase (SA-β-gal) activity, cell proliferation, and osteogenic differentiation.

PAX3 expression was significantly downregulated in senescent P11 hDPSCs compared to young P3 cells. Functionally, PAX3 knockdown was associated with accelerated cellular senescence, as evidenced by increased SA-β-gal activity, upregulation of p53 and p21 mRNA levels, impaired cell proliferation, and reduced osteogenic differentiation. Conversely, PAX3 overexpression was associated with an attenuated senescent phenotype, resulting in decreased SA-β-gal activity and expression of p53/p21, enhanced proliferation, and improved osteogenic capacity.

The transcription factor PAX3 may play a critical regulatory role in mitigating hDPSCs senescence, as downregulation of PAX3 was associated with increased senescence markers and its overexpression with reduced senescence markers. The observed correlation with p53/p21 mRNA levels suggests that this pathway may be involved, but mechanistic validation at the protein level is still required. These findings indicate that PAX3 represents a candidate for further investigation as a potential target to delay senescence and maintain the functional properties of hDPSCs, warranting further investigation for its potential utility in tissue engineering and clinical stem cell-based therapies.