Signal Transduction and Targeted Therapy | DKK3 in Keratinocytes Regulates Radiation-Induced Skin Hyperplasia and Fibrosis

This study reveals the critical role of DKK3 in keratinocytes during radiation-induced skin injury, demonstrating that it drives epidermal hyperplasia and skin fibrosis through the TGF-β-Wnt signaling axis. Targeting DKK3 significantly alleviates radiation dermatitis and fibrosis, offering a novel strategy for mitigating side effects of radiotherapy.

 

Literature Overview

The article titled 'Wnt-associated DKK3 in keratinocytes mediates radiation-induced hyperplasia, dermatitis and skin fibrosis,' published in the journal Signal Transduction and Targeted Therapy, reviews and summarizes the regulatory mechanisms of DKK3 in radiation-induced skin injury. Using a combination of in vitro and in vivo models—including 3D human skin models, cell type-specific Dkk3 knockout mice, and irradiated human skin samples—the study systematically reveals that upregulation of DKK3 in keratinocytes is a central driver of radiation-induced epidermal hyperplasia, inflammation, and skin fibrosis. This work not only clarifies the role of the Wnt signaling pathway in radiation-induced skin pathologies but also identifies DKK3 as a promising therapeutic target, holding significant translational value.

Background Knowledge

Radiation therapy is a cornerstone of cancer treatment, yet it often causes acute or chronic skin toxicities such as radiation dermatitis and skin fibrosis, severely impacting patients' quality of life. These pathological processes involve complex intercellular communication, including immune cell infiltration, fibroblast activation, and extracellular matrix (ECM) deposition. Currently, no effective treatments exist, and clinical management remains largely supportive. The Wnt signaling pathway plays a key role in tissue repair and fibrosis; its aberrant activation promotes fibroblast differentiation into myofibroblasts, driving excessive ECM deposition. Dickkopf family proteins (DKKs) are important regulators of Wnt signaling, with DKK3 exhibiting tissue-specific functions—capable of either activating or inhibiting Wnt signaling. Previous studies have implicated DKK3 in renal fibrosis and cardiomyopathy, but its role in radiation-induced skin injury remains unclear. Furthermore, TGF-β is a well-established master regulator of fibrosis and engages in extensive crosstalk with the Wnt pathway. Therefore, exploring whether DKK3 acts as an upstream regulator of the Wnt-TGF-β axis in radiation-induced skin fibrosis holds significant scientific and clinical relevance. By employing tissue-specific gene knockout mouse models and validating findings in human skin samples, this study fills a critical knowledge gap and identifies a novel target for anti-fibrotic interventions.

 

 

Research Methods and Experiments

The study employed multiple complementary approaches to investigate the role of DKK3 in radiation-induced skin injury. First,全身 Dkk3 knockout (DKK3-/-) and wild-type (WT) mice were subjected to a single 20 Gy dose of thoracic or hindlimb irradiation to assess skin toxicity phenotypes, including hair loss, epidermal hyperplasia, and fibrosis. Histological staining (Goldner’s, Masson’s trichrome), immunohistochemistry, and qPCR were used to analyze ECM deposition, proliferation markers, and inflammatory cell infiltration. To determine the cell-specific role of DKK3, conditional knockout mice were generated using K14-Cre (keratinocytes), CSF1R-Cre (macrophages), and PDGFR-Cre (fibroblasts). In vitro, immortalized human keratinocytes (N/TERT-1) were used with siRNA-mediated knockdown or inducible overexpression of DKK3, followed by clonogenic assays, flow cytometry, and β-galactosidase staining to evaluate radiation sensitivity and cellular senescence. A 3D human skin model was employed to confirm the impact of DKK3 on epidermal thickness and proliferation. In situ hybridization and immunohistochemistry were used to analyze DKK3 expression in both human and mouse skin tissues. A dual-reporter mouse model (DKK3 promoter-mCherry and Wnt-GFP) was utilized to monitor dynamic changes in Wnt activity post-irradiation. ROS detection, TGF-β pathway inhibitors, and neutralizing antibodies were applied to dissect the DKK3-Wnt-TGF-β signaling axis. Finally, human skin biopsy samples and public single-cell RNA-seq datasets were analyzed to validate DKK3 expression patterns in chronic radiation dermatitis and other fibrotic skin disorders.

Key Conclusions and Perspectives

• Radiation significantly upregulates DKK3 expression in keratinocytes, particularly in the basal layer, and this upregulation is commonly observed across multiple fibrotic skin diseases
• Loss of DKK3 (either globally or specifically in keratinocytes) markedly reduces radiation-induced epidermal hyperplasia, ECM deposition, and skin fibrosis, whereas deletion in macrophages or fibroblasts confers no protective effect
• DKK3 does not significantly affect the intrinsic radiosensitivity of keratinocytes but promotes radiation-induced cell cycle arrest and senescence
• Radiation-induced ROS upregulates DKK3 expression, which subsequently activates the canonical Wnt pathway; DKK3 knockdown blocks this process
• DKK3 activates Wnt signaling via autocrine TGF-β signaling, and inhibition of TGF-β receptor kinase or neutralization of TGF-β reverses DKK3-driven Wnt activation and epidermal hyperplasia
• Overexpression of DKK3 alters the secretory profile of keratinocytes, promoting macrophage polarization toward a CD163^highCD206^high M2-like phenotype, an effect dependent on direct cell-cell contact
• Macrophages conditioned by DKK3-overexpressing keratinocytes activate the TGF-β-SMAD pathway in fibroblasts, enhancing expression of fibrotic genes such as α-SMA and collagen
• Analysis of human skin samples shows significantly elevated DKK3 expression in basal keratinocytes in chronic radiation dermatitis, scars, lichen sclerosus, and morphea—consistent with scRNA-seq data

Research Significance and Prospects

This study systematically elucidates the central regulatory role of keratinocyte-derived DKK3 in radiation-induced skin fibrosis, revealing a signaling axis: 'ROS→DKK3→TGF-β→Wnt→epidermal hyperplasia→M2 macrophage polarization→fibroblast activation→fibrosis,' providing a novel perspective on the molecular mechanisms underlying radiation-induced skin injury. Traditionally, keratinocytes have been viewed primarily as barrier cells; however, this study highlights their active role in driving inflammation and fibrosis through signaling regulation, expanding our understanding of skin microenvironment crosstalk.

Targeting DKK3 may offer a new strategy for preventing or treating radiation dermatitis and fibrosis. Given that DKK3 is upregulated in multiple fibrotic diseases, its inhibitors may have broad-spectrum anti-fibrotic potential. Future studies could develop DKK3-neutralizing antibodies or small-molecule inhibitors and evaluate their efficacy and safety in large animal models. Additionally, exploring the role of DKK3 in radiation injury to other tissues (e.g., lung, intestine) could broaden its clinical applicability. This finding also offers new intervention strategies for other Wnt-related fibrotic diseases.

 

 

Conclusion

This study reveals the critical role of DKK3 in keratinocytes during radiation-induced skin injury. Ionizing radiation upregulates DKK3 expression in keratinocytes via ROS, which in turn activates the canonical Wnt pathway through autocrine TGF-β signaling, driving epidermal hyperplasia and cellular senescence. Keratinocytes with high DKK3 expression secrete specific factors that promote macrophage polarization toward an M2-like phenotype. These polarized macrophages activate fibroblasts through direct cell contact, ultimately leading to skin fibrosis. In multiple mouse models, keratinocyte-specific—but not macrophage- or fibroblast-specific—loss of DKK3 significantly alleviates radiation-induced skin lesions. Clinical sample analysis further confirms that DKK3 is significantly upregulated in the basal keratinocytes of patients with chronic radiation dermatitis and other fibrotic skin diseases. In summary, DKK3 serves as a key molecular node linking radiation injury to skin fibrosis. Targeting DKK3 holds promise as a novel therapeutic strategy to mitigate radiotherapy side effects, with significant translational value. This study provides new insights into the mechanisms of skin fibrosis and identifies a potential target for anti-fibrotic drug development.

 

Li Li, Ramon Lopez Perez, Khuram Shehzad, Roger Sandhoff, and Peter E Huber. Wnt-associated DKK3 in keratinocytes mediates radiation-induced hyperplasia, dermatitis and skin fibrosis. Signal Transduction and Targeted Therapy.