Differential Gene Expression of Ca2+ Channel-Mediated Signaling Pathways in DPSCs under Hypoxic Condition

JOHN T. TRAN
Dental pulp stem cell (DPSC) survival, differentiation into odontoblast and deposition of matrix for mineralization is essential for vital pulp therapy (VPT) and regenerative endodontic therapy (RET). Calcium homeostasis in DPSC plays a critical role for its survival, longevity, and differentiation processes. Cells use this external source of signal calcium ions by activating various entry channels with different properties. The objective of this research is to investigate key gene candidates involved with calcium homeostasis in human dental pulp tissue and DPSCs.

DPSCs isolated from human dental pulp were cultured in hypoxic chamber (3%) for 21 days at 37°C in differentiation media and controls were cultured under normal conditions. Later, DPSCs collected were extracted for mRNA isolation. The mRNA was sequenced using NextSeq Illunima and was analyzed using the ERGOTM transcription tool using statistical methods such as Limma (Linear Model) and DESeq2 (Negative Binomial).

There were a total of 24 biomarkers identified with a 6-fold higher over expressed in hypoxic conditions compared to normoxia. These include key genes such as CALM2, MYLK, FGR1, ASPH, PDGFRA, ATP2B4, ATP2B1, PDGFRB, CALM1, and PPP3CA. The genes and pathways were identified in KEGG pathways overlaying expression values.

The release of calcium ions from internal stores and the influx of calcium ions from the environment cause a dramatic and rapid increase in cytoplasmic calcium concentration, which has been exploited for signal transduction. In combination with power statistical analysis such as DESeq2 and Limma, key pathways can be identified that are responsible for calcium homeostasis and calcium channel signaling as key diagnostic markers for VPT and RET.