Impact of Skin Exposure to Benzo[a]pyrene in Rat Model: Insights into Epidermal Cell Function and Draining Lymph Node Cell Response

Abstract

The skin is a direct target of the air pollutant benzo[a]pyrene (BaP). While its carcinogenic qualities are well-studied, the immunotoxicity of BaP after dermal exposure is less understood. This study examines the immunomodulatory effects of a 10-day epicutaneous BaP application, in environmentally/occupationally relevant doses, by analyzing ex vivo skin immune response (skin explant, epidermal cells and draining lymph node/DLN cell activity), alongside the skin's reaction to sensitization with experimental hapten dinitrochlorobenzene (DNCB). The results show that BaP application disrupts the structure of the epidermal layer and promotes immune cell infiltration in the dermis. BaP exposure led to oxidative stress in epidermal cells, characterized by decreased reduced glutathione and increased AHR and Cyp1A1 expression. Production and gene expression of proinflammatory cytokines (TNF, IL-1β) by epidermal cells decreased, while IL-10 response increased. Decreased spontaneous production of IFN-γ and IL-17, along with unchanged IL-10, was observed in DLC cells, whereas ConA-stimulated production of these cytokines was elevated. Local immunosuppression caused by BaP application seems to reduce the skin's response to an additional stimulus, evidenced by decreased effector activity of DLN cells three days after sensitization with DNCB. These findings provide new insight into the immunomodulatory effects and health risks associated with skin exposure to BaP.

Keywords: draining lymph nodes cell activity; epicutaenous benzo[a]pyrene application; epidermal cell activity; rats; skin explants.

Figure 1

Histology of skin following epicutaneous BaP application. (A) Control skin with normal histological structure. Epidermal layer and upper dermis with thin collagen fibrils parallel arranged in control skin (insert). (B) Collagen bundles in different directions. (C) Mast cells appearance in deep dermis of control skin. (D) BaP 20 ng/cm² application induced increased skin thickness, appearance of epidermal cells with pyknotic nuclei (top insert), disturbed epidermal stratification (bottom insert) and dermal infiltration (inserts). (E) Increased cellularity, disarrangement of collagen bundles and mild hyperemia of dermal blood vessels (insert), as well as (F) more numerous and densely stained mast cells in dermis following BaP 20 ng/cm² application. (G) BaP 100 ng/cm² application resulted in patchy desquamation (bottom insert) and pronounced keratinocytes hyperplasia with atypical nuclei and loss of ordered stratification (top insert). (H) More pronounced dermal cellularity and (I) mast cells number, mostly located in deep dermis and along blood vessels, in process of degranulation (insert) following BaP 100 ng/cm² application.

Figure 2

Inflammatory response in skin following epicutaneous BaP application. (A) TNF and (B) IL-1β production by skin explants. Results are presented as mean (±SD). Significance at: * p < 0.05 vs. control animals (BaP dose 0 ng/cm²).

Figure 3

Stress response in epidermal cells following epicutaneous BaP application. (A) Intracellular GSH. Epidermal cells mRNA expression of (B) AhR and (C) Cyp1A1. Results are presented as mean (±SD). Significance at: * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control animals (BaP dose 0 ng/cm²).

Figure 4

Effect of epicutaneous BaP application on epidermal cells’ cytokine response. (A,B) TNF, (C,D) IL-1β and (E,F) IL-10 mRNA expression and spontaneous production, respectively. Results are presented as mean (±SD). Significance at: * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control animals (BaP dose 0 ng/cm²).

Figure 5

Effect of epicutaneous BaP application on draining lymph node cells’ cytokine response. (A–C) IFN-γ, (D–F) IL-17 and (G–I) IL-10 mRNA expression, spontaneous and ConA-stimulated production, respectively. Results are presented as mean (±SD). Significance at: * p < 0.05, ** p < 0.01 vs. control animals (BaP dose 0 ng/cm²).