Tujia medicine Toddalia asiatica improves synovial pannus in rats with collagen-induced arthritis through the PI3K/Akt signaling pathway

doi:10.12122/j.issn.1673-4254.2024.08.17

Abstract

Abstract:

Objective To investigate the therapeutic mechanism of Tujia medicine Toddalia asiatica alcohol extract (TAAE) for synovial pannus formation in rats with college-induced arthritis (CIA). Methods Sixty male SD rats were randomized into normal control group, CIA model group, TGT group, 3 TAAE treatment groups at low, medium and high doses (n=10). Except for those in the normal control group, all the rats were subjected to CIA modeling using a secondary immunization method and treatment with saline, TGT or TAAE by gavage once daily for 35 days. The severity of arthritis was assessed using arthritis index (AI) score, and knee joint synovium pathologies were examined with HE staining. Serum levels of TNF-α, IL-6, and IL-1β were detected with ELISA; the protein expressions of PI3K, Akt, p-PI3K, p-Akt, VEGF, endostatin, HIF-1α, MMP1, MMP3, and MMP9 in knee joint synovial tissues were determined using Western blotting, and the mRNA expressions of TNF‑α, IL-6, IL-1β, VEGF, HIF-1α, PI3K, and Akt were detected with RT-PCR. Results Treatment of CIA rat models with TAAE and TGT significantly alleviated paw swelling, lowered AI scores, and reduced knee joint pathology, neoangiogenesis, and serum levels of inflammatory factors. TAAE treatment obviously increased endostatin protein expression, downregulated p-PI3K, p-Akt, MMP1, MMP3, MMP9, VEGF, and HIF-1α proteins, and reduced TNF‑α, IL-6, IL-1β, PI3K, Akt, VEGF, and HIF-1α mRNA levels in the synovial tissues, and these changes were comparable between high-dose TAAE group and TGT group. Conclusion TAAE can improve joint symptoms and inhibit synovial pannus formation in CIA rats by regulating the expressions of HIF-1α, VEGF, endostatin, MMP1, MMP3, and MMP9 via the PI3K/Akt signalling pathway.

Key words: Toddalia asiatica, rheumatoid arthritis, pannus, inflammation, PI3K/Akt signaling pathway

Shan XIANG, Zongxing ZHANG, Lu JIANG, Daozhong LIU, Weiyi LI, Zhuoma BAO, Rui TIAN, Dan CHENG, Lin YUAN. Tujia medicine Toddalia asiatica improves synovial pannus in rats with collagen-induced arthritis through the PI3K/Akt signaling pathway[J]. Journal of Southern Medical University, 2024, 44(8): 1582-1588.

Figures/Tables 8

Tab.1 Scoring criteria for arthritis severity in rats

Score	Joint symptoms
0	No swelling or redness
1	Mild swelling and redness
2	Moderate swelling and redness with slight decrease in joint mobility
3	Severe swelling and redness with slight decrease in joint mobility
4	Severe redness and swelling with significant stiffness and limited joint mobility

Tab.2 Primers sequence for RT-qPCR

Name	Primer sequence (5'-3')
GAPDH	F:CTGGAGAAACCTGCCAAGTATG
GAPDH	R:GGTGGAAGAATGGGAGTTGCT
TNF-α	F:CCCTCACACTCACAAACCACC
TNF-α	R:CTTTGAGATCCATGCCGTTG
IL-1β	F:GCATCCAGCTTCAAATCTCGC
IL-1β	R:TGTTCATCTCGGAGCCTGTAGTG
IL-6	F:CCCCAATTTCCAATGCTCTCC
IL-6	R:CGCACTAGGTTTGCCGAGTA
PI3K	F:TGACAGGCACAACGACAACATC
PI3K	R:AGGTAAGCCCTAACGCAGACAT
Akt	F:CGACGTAGCCATTGTGAAGGAG
Akt	R:ATTGTGCCACTGAGAAGTTGTTG
HIF-1α	F:CCGCCACCACCACTGATGAATC
HIF-1α	R:CCGCCACCACCACTGATGAATC
VEGF	F:CCGCCACCACCACTGATGAATC
VEGF	R:CCGCCACCACCACTGATGAATC

Fig.1 Changes in foot swelling of the rats (Mean±SD, n=10). **P<0.01 vs control group; #P<0.05, ##P<0.01 vs model group.

Fig.2 Changes in joint index of the rats (Mean±SD, n=10). **P<0.01 vs control group; #P<0.05, ##P<0.01 vs model group.

Fig.3 Effects of TAAE and TGT on serum levels of TNF-α, IL-6 and IL-1β in CIA rats (Mean±SD, n=10). **P<0.01 vs control group; ##P<0.01 vs model group.

Fig.4 Histopathological changes of the joints in CIA rats (HE staining, original magnification: ×100).

Fig.5 Comparison of protein expressions in the synovial tissue of the rats in different groups (Mean±SD, n=6). **P<0.01 vs control group; #P<0.05, ##P<0.01 vs model group.

Fig.6 Comparison of expressions of pathway-related proteins in synovial tissue of rats in each group (Mean±SD, n=6). **P<0.01 vs control group; #P<0.05, ##P<0.01 vs model group.

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