Korean J Physiol Pharmacol.  2023 Jul;27(4):357-364. 10.4196/kjpp.2023.27.4.357.

Shikonin ameliorates salivary gland damage and inflammation in a mouse model of Sjögren’s syndrome by modulating MAPK signaling pathway

• Guo W
• Wang X
• Sun C
• Wang J
• Wang T

• KMID: 2544137
• DOI: http://doi.org/10.4196/kjpp.2023.27.4.357

Abstract

Sjögren syndrome (SS) is a systemic inflammatory autoimmune disease that involves exocrine glands. Shikonin is extracted from comfrey, which is conventionally used as an anti-tumor, antibacterial, and antiviral drug in China. However, the application of Shikonin in SS remains unreported. This study aimed to verify the potential functions of Shikonin in SS progression. Firstly, non-obese diabetic mice were used as the SS mouse model, with C57BL/6 mice serving as the healthy control. It was demonstrated that the salivary gland damage and inflammation were aggravated in the SS mouse model. Shikonin improved salivary gland function decline and injury in the SS mouse model. Moreover, Shikonin reduced inflammatory cytokines and immune infiltration in the SS mouse model. Further experiments discovered that Shikonin attenuated the MAPK signaling pathway in the SS mouse model. Lastly, inhibition of the MAPK signaling pathway combined with Shikonin treatment further alleviated the symptoms of SS. In conclusion, Shikonin ameliorated salivary gland damage and inflammation in a mouse model of SS by modulating the MAPK signaling pathway. Our findings indicate that Shikonin may be a useful drug for SS treatment.

Keyword

Inflammation; Salivary gland; Shikonin; Sjogren’s syndrome

Figure

• 

  Fig. 1 Shikonin improves salivary gland function decline and injury in the SS mouse model. Groups were divided into five groups: C57BL/6, NOD, NOD + 12.5 mg/kg Shikonin, NOD + 25 mg/kg Shikonin, and NOD + 50 mg/kg Shikonin (n = 6 for each group). (A) Salivary flow (mg/10 min) was measured. ***p < 0.001 vs. the C57BL/6 group, ###p < 0.001 vs. the NOD group. (B) Salivary gland index (mg/g) and spleen index (mg/g) were determined. ***p < 0.001. Values are presented as mean ± SD. SS, Sjögren syndrome; NOD, non-obese diabetic; ns, not significant.

• 

  Fig. 2 Shikonin reduces inflammatory cytokines in the SS mouse model. Groups were divided into five groups: C57BL/6, NOD, NOD + 12.5 mg/kg Shikonin, NOD + 25 mg/kg Shikonin, and NOD + 50 mg/kg Shikonin. (A) The levels of TNF-α, IL-6, IL-17, IL-4 in the blood were measured using ELISA. (B) The protein expressions of p-NF-κB, NF-κB in the blood were analyzed using Western blot. *p < 0.05, ***p < 0.001. Values are presented as mean ± SD. SS, Sjögren syndrome; NOD, non-obese diabetic; ns, not significant.

• 

  Fig. 3 Shikonin reduces immune infiltration in the SS mouse model. Groups were divided into five groups: C57BL/6, NOD, NOD + 12.5 mg/kg Shikonin, NOD + 25 mg/kg Shikonin, and NOD + 50 mg/kg Shikonin. (A) Flow cytometry was used to detect the percentages of CD4+, CD8+, and B220+CD19+ cells. (B, C) The cell viability of B and T cells was measured using the CCK-8 assay. *p < 0.05, **p < 0.01, ***p < 0.001. Values are presented as mean ± SD. SS, Sjögren syndrome; NOD, non-obese diabetic; ns, not significant.

• 

  Fig. 4 Shikonin modulates the MAPK signaling pathway. Groups were divided into five groups: C57BL/6, NOD, NOD + 12.5 mg/kg Shikonin, NOD + 25 mg/kg Shikonin, and NOD + 50 mg/kg Shikonin. Western blot was used to examine the protein expressions of p-ERK, ERK, p-JNK, JNK, p-P38, and P38. ***p < 0.001. Values are presented as mean ± SD. NOD, non-obese diabetic.

• 

  Fig. 5 Inhibition of the MAPK signaling pathway alleviates the symptoms of SS. Groups were divided into four groups: C57BL/6, NOD, NOD + 5 mg/kg SB202190, NOD + 5 mg/kg SB202190 + 50 mg/kg Shikonin. (A) Salivary flow (mg/10 min) was examined. ***p < 0.001 vs. the C57BL/6 group, ###p < 0.001 vs. the NOD group, ^^^p < 0.001 vs. the NOD + 5 mg/kg SB202190 group. (B) The salivary gland index (mg/g) and spleen index (mg/g) were measured. (C) ELISA was used to verify the levels of TNF-α, IL-6, IL-17, and IL-4. (D) Flow cytometry was used to detect the percentages of CD4+, CD8+, and B220+CD19+ cells. *p < 0.05, **p < 0.01, ***p < 0.001. Values are presented as mean ± SD. SS, Sjögren syndrome; NOD, non-obese diabetic.

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