Solamargine Alleviated UVB-Induced Inflammation and Melanogenesis in Human Keratinocytes and Melanocytes via the p38 MAPK Signaling Pathway, a Promising Agent for Post-inflammatory Hyperpigmentation

Abstract

Post-inflammatory hyperpigmentation (PIH) is a common acquired pigmentary disorder occurring after skin inflammation or injury. Ultraviolet B irradiation could exaggerate PIH clinically due to its effect on promoting cutaneous inflammation and melanogenesis in keratinocytes and melanocytes, respectively. Solamargine (SM), a steroidal alkaloid glycoside extracted from Solanum undatum, significantly inhibits Ultraviolet B (UVB)-induced pro-inflammatory cytokines IL-1α, IL-1β, IL-8, and IFN-γ, as well as paracrine melanogenic factors ET-1, α-MSH, and bFGF in human keratinocytes. Additionally, SM significantly attenuated UVB-induced melanin synthesis in human epidermal melanocytes through down-regulation of tyrosinase activity and expression of MITF, TRP-1, TRP-2, and tyrosinase. SM exerted an anti-inflammatory effect in UVB-irradiated keratinocytes through the p38 MAPK/Nrf2/HO-1 signaling pathway. With its anti-inflammatory and whitening effect, SM may improve PIH through paracrine regulations of keratinocytes and direct action on melanocytes, making it a promising agent for PIH.

Keywords: HO-1; MAPK; Nrf2; SM (solamargine); inflammation; melanogenesis; p38; post-inflammatory hyperpigmentation (PIH).

FIGURE 1

Cell viability and melanin content of HaCaT and HEM cells after NB-UVB irradiation. The inhibition rate of HaCaT cells and HEM cells of different concentrations of SM. (A) The chemical structure of SM modified from https://pubchem.ncbi.nlm.nih.gov/compound/Solamargine. (B) Cell viability of HaCaT cells and HEM cells was detected by CCK-8 24 h after irradiation of NB-UVB with different fluences (100, 200, 300, and 500 mJ/cm²). Cell viability is presented as a eprcentage of control. (C) Melanin content of HEM cells after irradiation of NB-UVB with 250 mJ/cm² Melanin content is percentage as a percentage of control. (D,E) The IC50 value for HaCaT cells was 4.252 μg/ml and for HEM cells was 4.472 μg/ml. *P < 0.05, **P < 0.01, and ***P < 0.001.

FIGURE 2

Solamargine (SM) attenuated UVB-induced secretion of pro-inflammatory cytokines and melanogenic factors in HaCaT cells. Real-time PCR (24 h after incubation with SM) and ELISA assay (48 h after incubation with SM) demonstrated SM attenuated UVB-induced secretion of pro-inflammatory cytokines, IL-1α (A,E), IL-1β (B,F), IL-8 (C,G), and IFN-γ (D,H) in HaCaT cells. Treatment with SM after UVB irradiation significantly decreased the mRNA and protein levels of IL-1α, IL-1β, IL-8, and IFN-γ. The anti-inflammatory effect of SM is comparable to 150 μg/ml dexamethasone. Real-time PCR and ELISA assay demonstrated SM attenuated UVB-induced secretion of melanogenic factors, ET-1 (I,L), α-MSH (J,M), and bFGF (K,N) in HaCaT cells. Treatment with SM after UVB irradiation significantly decreased the mRNA and protein levels of ET-1, α-MSH, and bFGF. The anti-melanogenic effect of SM is comparable to 150 μg/ml Arbutin. The data were represented as mean ± SD from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 (vs. non-treated group); ^#P < 0.05, ^##P < 0.01; ^###P < 0.001 (vs. NB-UVB irradiated group).

FIGURE 3

Solamargine (SM) Inhibited UVB-induced melanogenesis in HEM cells. (A) Melanin content of HEM cells 48 h after the UVB irradiation with SM or arbutin treatment. (B) Tyrosinase activity of HEMs 24 h after the UVB irradiation with SM or arbutin treatment. Tyrosinase activity is presented as a percentage of control. Real-time PCR (24 h after incubation with SM) and Western Blot (48 h after incubation with SM) demonstrated SM attenuated UVB-induced secretion of melanogenic factors, TYR (C) Western blot assay of MITF, TYR,TRP-1 and TRP-2 after UVB exposure with/without SM or arbutin treatment. (D,H), MITF (E,I), TRP-1 (F,J), and TRP-2 (G,K) in HEM cells. Treatment with SM after UVB irradiation significantly decreased the mRNA and protein levels of TYR, MITF, TRP-1, and TRP-2. The anti-melanogenesis effect of SM is comparable to 150 μg/ml Arbutin. The data represent the mean ± S.D. from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 (vs. non-treated group); ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 (vs. NB-UVB irradiated group).

FIGURE 4

Solamargine (SM) exerting anti-inflammatory effect via the activation of p38 MAPK/Nrf2/HO-1 signaling pathways in HaCaT cells. After UVB irradiation of HaCaT cells, the protein level of Nrf-2 (A) and HO-1 (B) was significantly decreased, and p-p38/p38 (C) expression was significantly increased. This effect can be reversed by SM. The expression of Nrf-2 and HO-1 proteins in HaCaT cells was significantly decreased and p-p38/p38 expression was significantly increased in the UVB + SM + p38 inhibitor (SB203580) group compared with the UVB + SM group. After the addition of the p38 inhibitor, IL-1α (D) Western blot assay of Nrf-2, HO-1, p-p38 and p38 after UVB exposure with/without SM or SB203580 treatment. (E), IL-1β (F), IL-8 (G), and IFN-γ (H) in the cell supernatant were significantly increased. The data represent the mean ± S.D. from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 (vs. non-treated group); ^ΔP < 0.05, ^ΔΔ P < 0.01 (vs. NB-UVB irradiated group); ^##P < 0.01, ^###P < 0.001 (vs. NB-UVB irradiated and SM treated group).