Journal of Traditional Chinese Medicine ›› 2023, Vol. 43 ›› Issue (1): 68-77.DOI: 10.19852/j.cnki.jtcm.20220928.001
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Qingchi San (青赤散) treats ulcerative colitis in mice by inhibiting the nuclear factor-kappa B signaling pathway and Nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 inflammasome formation
ZHOU Zhenghua1,2(
), JI Jianbin1,2, WANG Hongxia1,2, YAN Lin1,2, KANG Hongchang1,2
- 1 Department of Gastroenterology, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin 300381, China
2 National Clinical Research Center for Chinese Medicine Acupuncture and Moxibustion, Tianjin 300381, China
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Received:2021-08-05Accepted:2021-11-21Online:2023-02-15Published:2023-01-10 -
Contact:ZHOU Zhenghua,JI Jianbin,WANG Hongxia,YAN Lin,KANG Hongchang -
About author:Prof. ZHOU Zhenghua, Department of Gastroenterology, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin 300381, China. 15202248272@139.com. Telephone:+86-15202248272
-
Supported by:The National Key Research and Development Program(Grant2018YFC1705403);Clinical Effect Evaluation of Chinese Medicine on Moderately Active Ulcerative Colitis; Natural Science Foundation of Tianjin(Grant18JCYBJC26000);Study on the Mechanism of Qingchi San Treating Ulcerative Colitis By Regulating Gut Microflora; Scientific Research Program of Tianjin Education Commission(Grant2017KJ155);based on γδT-NLRP3 Inflammasome-Caspase Pathway, Qingchi San Regulates Gut Microflora to Treat UC
Cite this article
ZHOU Zhenghua, JI Jianbin, WANG Hongxia, YAN Lin, KANG Hongchang. Qingchi San (青赤散) treats ulcerative colitis in mice by inhibiting the nuclear factor-kappa B signaling pathway and Nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 inflammasome formation[J]. Journal of Traditional Chinese Medicine, 2023, 43(1): 68-77.
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Table 1 Primer sequence used in RT-qPCR
| cDNA | Order | Annealing Temp (℃) | Sequence |
|---|---|---|---|
| COX2 | Sense | 59 | 5′-GTCAATACGGATCATATCCT-3′ |
| Antisense | 5′-ACTGCAACTGACCATTCTAC-3′ | ||
| iNOS | Sense | 58 | 5′-AGCAGTTTATTGACATAAATCAA-3′ |
| Antisense | 5′-CGTGAACTCGGTCTATGATTCA-3′ | ||
| GAPDH | Sense | 57 | 5′-AGAAGGCTGGGGCTCATTTG-3′ |
| Antisense | 5′-AGAAGGCTGGGGCTCATTTG-3′ |
Table 1 Primer sequence used in RT-qPCR
| cDNA | Order | Annealing Temp (℃) | Sequence |
|---|---|---|---|
| COX2 | Sense | 59 | 5′-GTCAATACGGATCATATCCT-3′ |
| Antisense | 5′-ACTGCAACTGACCATTCTAC-3′ | ||
| iNOS | Sense | 58 | 5′-AGCAGTTTATTGACATAAATCAA-3′ |
| Antisense | 5′-CGTGAACTCGGTCTATGATTCA-3′ | ||
| GAPDH | Sense | 57 | 5′-AGAAGGCTGGGGCTCATTTG-3′ |
| Antisense | 5′-AGAAGGCTGGGGCTCATTTG-3′ |
Figure 1 QCS significantly inhibited weight loss, decreased DAI score, and inhibited abnormal lesions in the colon in UC mice A: body weight; B: DAI score; C: length of colon; D: weight of colon. 3%DSS, mesalazine solution (0.82g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; QCS: Qingchi San; DAI: disease activity index; UC: ulcerative colitis. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure 1 QCS significantly inhibited weight loss, decreased DAI score, and inhibited abnormal lesions in the colon in UC mice A: body weight; B: DAI score; C: length of colon; D: weight of colon. 3%DSS, mesalazine solution (0.82g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; QCS: Qingchi San; DAI: disease activity index; UC: ulcerative colitis. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure 2 ELISA assay of TNF-α, IL-1β, IL-6, MPO and SOD A: TNF-α; B: IL-1β; C: IL-6; D: MPO; E: SOD. 3% DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11, DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; ELISA: enzyme-linked immunosorbent assay; TNF-α: tumor necrosis factor-α; IL-1β: interleukin-1β; IL-6: interleukin-6; MPO: myeloperoxidase; SOD: superoxide dismutase. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure 2 ELISA assay of TNF-α, IL-1β, IL-6, MPO and SOD A: TNF-α; B: IL-1β; C: IL-6; D: MPO; E: SOD. 3% DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11, DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; ELISA: enzyme-linked immunosorbent assay; TNF-α: tumor necrosis factor-α; IL-1β: interleukin-1β; IL-6: interleukin-6; MPO: myeloperoxidase; SOD: superoxide dismutase. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure 3 Protein relative expression of NF-κB, IκBα and p-IκBα A: relative expression of NF-κB, IκBα and p-IκBα by Western blotting; B: NF-κB expression level in the groups; C: p-IκBα expression level in the groups; D: IκBαexpression level in the groups. 3%DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; NF-κB: nuclear factor-kappa B, IκBα: inhibitor of NF-κBα; p-IκBα: phosphorylated inhibitor of NF-κBα. aP < 0.001, bP < 0.01, cP< 0.05, vs control.
Figure 3 Protein relative expression of NF-κB, IκBα and p-IκBα A: relative expression of NF-κB, IκBα and p-IκBα by Western blotting; B: NF-κB expression level in the groups; C: p-IκBα expression level in the groups; D: IκBαexpression level in the groups. 3%DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; NF-κB: nuclear factor-kappa B, IκBα: inhibitor of NF-κBα; p-IκBα: phosphorylated inhibitor of NF-κBα. aP < 0.001, bP < 0.01, cP< 0.05, vs control.
Figure 4 Protein relative expression of caspase-1, NLRP3 and ASC A: relative expression of caspase-1, NLRP3 and ASC by Western blotting; B: caspase-1 expression level in the groups; C: NLRP3 expression level in the groups; D: ASC expression level in the groups. 3%DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; NLRP3: nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain- containing 3; ASC: apoptosis-associated speck-like protein containing a caspase recruitment domain. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure 4 Protein relative expression of caspase-1, NLRP3 and ASC A: relative expression of caspase-1, NLRP3 and ASC by Western blotting; B: caspase-1 expression level in the groups; C: NLRP3 expression level in the groups; D: ASC expression level in the groups. 3%DSS, mesalazine solution (0.82 g·kg-1·d-1), QCS-L (19.50 g·kg-1·d-1), QCS-M (29.25 g·kg-1·d-1) and QCS-H (38.99 g·kg-1·d-1) were used for enema twice a day for 0-11 d, and mice were killed on day 11. DSS: dextran sulfates sodium; QCS-L: low-dose Qingchi San; QCS-M: middle-dose Qingchi San; QCS-H: high-dose Qingchi San; NLRP3: nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain- containing 3; ASC: apoptosis-associated speck-like protein containing a caspase recruitment domain. aP < 0.001, bP < 0.01, c P < 0.05, vs control.
Figure S1 HE staining of colon tissue (X10)
Figure S1 HE staining of colon tissue (X10)
Figure S2 mRNA relative expression of iNOS and COX2
Figure S2 mRNA relative expression of iNOS and COX2
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