circ_0042103/TAF15/NER Axis Drives DNA Damage in Pulpitis

Study Background and Research Question

Pulpitis, a prevalent dental inflammatory disease, results from persistent infection and immune activation in dental pulp tissues. Chronic inflammation in pulpitis is closely associated with DNA damage, leading to progressive tissue dysfunction and pain. However, the molecular regulators connecting inflammation, DNA damage response (DDR), and tissue degeneration have remained incompletely characterized. Circular RNAs (circRNAs), a class of covalently closed non-coding RNAs, have emerged as key players in epigenetic regulation, yet their specific function in pulpitis and in controlling DNA repair pathways has been unclear (source: Lai et al., 2026).

This study addresses a central question: How does the circular RNA circ_0042103 regulate DNA damage and inflammatory signaling in the context of pulpitis, and what are the mechanistic links to the nucleotide excision repair (NER) pathway?

Key Innovation from the Reference Study

The principal innovation lies in the identification and functional characterization of a novel regulatory axis—circ_0042103/TAF15/NER—that orchestrates DNA damage and inflammation in dental pulp stem cells (DPSCs) during pulpitis. The research demonstrates that circ_0042103 binds to the RNA-binding protein TAF15, subsequently downregulating NER pathway components (ERCC1 and PCNA), which exacerbates DNA double-strand breaks (DSBs) and amplifies inflammatory cytokine expression. This mechanistic insight provides a direct molecular link between circRNA regulation, DNA repair impairment, and inflammatory progression in dental tissue (source: Lai et al., 2026).

Methods and Experimental Design Insights

The study employed a comprehensive suite of molecular and cellular techniques to interrogate RNA profiles, DNA damage markers, and protein-protein/RNA interactions:

• Microarray and Single-cell RNA Sequencing: Used to profile circRNA expression and DNA damage response genes in inflamed pulp tissues and derived stem cells.
• Quantitative RT-PCR and Western Blot: Deployed for quantifying the DNA double-strand break marker γ-H2AX and inflammatory cytokines (e.g., IL-6, IL-8, TNF-α) in pulpitis samples.
• Bioinformatics Analysis: Enabled identification of differentially expressed circRNAs specifically upregulated during inflammation.
• Functional Assays: Lipopolysaccharide (LPS) was used to induce inflammation and DNA damage in DPSCs in vitro, followed by knockdown and overexpression experiments targeting circ_0042103.
• RNA FISH, Pulldown, and Nuclear-Cytoplasmic Fractionation: These assays elucidated the subcellular localization and direct interaction between circ_0042103 and TAF15.
• siRNA-mediated Knockdown: Targeting circ_0042103 and TAF15 enabled dissection of their roles in modulating the NER pathway and downstream phenotypes.

Collectively, this multimodal approach validated both the correlation and causality between circ_0042103 activity, DNA repair inhibition, and inflammation in human dental pulp cells.

Protocol Parameters

• RNA extraction from pulp tissue | ~50–100 mg tissue per sample | Ensures adequate RNA yield for downstream assays | Standard for tissue-level RNA profiling | paper
• qRT-PCR for γ-H2AX, ERCC1, PCNA, cytokines | 10–50 ng cDNA per reaction | Detects changes in DNA damage and repair gene expression | Enables sensitive quantification of inflammatory and DNA repair markers | paper
• LPS stimulation of DPSCs | 1 μg/mL for 24 h | Models inflammatory injury in vitro | Recapitulates clinically relevant inflammatory insult | paper
• circRNA knockdown/overexpression | 50–100 nM siRNA/plasmid per well | Functional validation of regulatory axis | Allows mechanistic dissection | paper
• RNase R treatment for circRNA enrichment | 1–2 U/μg RNA, 30 min at 37°C | Enriches circular RNAs, depletes linear RNA | Facilitates specific circRNA detection and analysis | workflow_recommendation

Core Findings and Why They Matter

Key findings from the study include:

• DDR Activation in Pulpitis: Both inflamed pulp tissues and DPSCs exhibited marked activation of DNA damage response pathways, as evidenced by elevated γ-H2AX expression.
• Positive Correlation Between DNA Damage and Inflammation: Levels of DNA double-strand breaks correlated with proinflammatory cytokine expression, indicating intertwined pathophysiology.
• circ_0042103 as a Central Regulator: Upregulation of circ_0042103 intensified LPS-induced DNA damage and cytokine release, while its silencing mitigated both outcomes.
• TAF15 Interaction and NER Pathway Inhibition: circ_0042103 physically binds TAF15, sequestering it and reducing the expression of NER pathway proteins (ERCC1, PCNA), thus impeding effective DNA repair and amplifying cell injury.
• Definition of the circ_0042103/TAF15/NER Axis: The newly defined axis mechanistically explains how circRNA dysregulation can drive both genomic instability and inflammation in pulpitis (source: Lai et al., 2026).

These results provide a foundation for targeting circ_0042103 or its downstream effectors as a strategy to ameliorate pulpitis by restoring DNA repair and dampening inflammatory signaling.

Comparison with Existing Internal Articles

The present findings align with and extend prior investigations into the role of circular RNAs in inflammatory and DNA repair processes. Notably, the internal article "circ_0042103/TAF15/NER Axis Links Circular RNA to DNA Damage in Pulpitis" offers a concise overview, underscoring the centrality of the circ_0042103/TAF15/NER axis in driving pathological DNA damage and inflammation. The current reference study provides more comprehensive mechanistic evidence and direct functional validation in human DPSCs.

Further, internal resources such as "The circ_0042103/TAF15/NER Axis Drives Inflammation in Pulpitis" reinforce the significance of circular RNA-mediated regulatory networks in dental tissue pathophysiology. Complementary articles on Ribonuclease R (20 U/μL) highlight the technical tools that enable specific enrichment and analysis of circular RNAs, which are instrumental in studies like the present one.

Limitations and Transferability

While the study robustly demonstrates the circ_0042103/TAF15/NER mechanism in vitro and in human dental pulp samples, several limitations should be considered:

• In Vivo Validation: The findings are primarily based on cell culture and ex vivo tissue assays; in vivo confirmation in animal models or clinical cohorts would strengthen the translational relevance.
• Context-Specificity: The regulatory axis may exhibit cell-type or tissue-specific effects, and its role in other forms of inflammatory or DNA repair-related diseases remains to be determined.
• Complexity of circRNA Networks: Although circ_0042103 is highlighted, other circular RNAs may contribute to pulpitis pathogenesis and warrant further investigation.

Nevertheless, the mechanistic insights provided are highly transferable to the development of targeted therapeutics or biomarker strategies within the field of dental regenerative medicine.

Research Support Resources

For researchers aiming to dissect circular RNA function or perform similar RNA structure analysis, the use of highly processive exoribonucleases such as Ribonuclease R (RNase R) (20 U/μL) (SKU K3061, APExBIO) is recommended. RNase R enables selective degradation of linear RNAs, thereby enriching for circular RNA species and facilitating downstream functional or quantitative assays (workflow_recommendation). This enzymatic approach supports robust circular RNA enrichment, validation, and pathway analysis in inflammation and RNA stability studies.