Guizhi-Shaoyao-Zhimu decoction possesses anti-arthritic effects on type II collagen-induced arthritis in rats via suppression of inflammatory reactions, inhibition of invasion & migration and induction of apoptosis in synovial fibroblasts.

Rheumatoid arthritis (RA) is a known intractable chronic inflammatory disease of synovial joints characterized by hyperplasia and consecutive inflammation with a high prevalence.Guizhi-Shaoyao-Zhimu (GSZD) is the first choice for clinical treatment of RA in Chinese traditional medicine. This study is aimed to explore the possible pharmacological mechanisms of anti-arthritic effect of GSZD. Type II collagen-induced arthritis (CIA) rat model was used to study the anti-arthritic activity of GSZDin vivo, and toe swelling & arthritis score, serum levels of cytokines, and pathological examinations were carried out. In vitro, TNF-α induced MH7A cells were used to study the possible mechanisms of GSZD. The anti-proliferative effects of GSZD were determined by MMT assay, and pro-apoptotic activity of GSZD in MH7A cells was determined by flow cytometry analysis & DAPI staining. Furthermore, the adhesive and invasive abilities of MH7A cells were determined using cell adhesion and transwell assays. MMPs levels were determined by ELISA assays, and mRNA expressions of Caspase-3, -9, Bax, SOCS1, Bcl-2, JAK2, STAT-3 and -5 were determined using qRT-PCR analysis. Besides, the major chemical components in GSZD were analyzed by HPLC-QqQ-MS analysis. Our results showed GSZD reduced the toe swelling & arthritis score, and serum levels of TNF-α, IL-1β, IL-6 & IL-17a in CIA rats; pathological examination results indicated GSZD improved ankle joint injury in CIA rats.In vitro, GSZD showed significant anti-proliferative and pro-apoptotic effects on TNF-α stimulated MH7A cells. After GSZD treatment, the adhesive and invasive abilities of MH7A cells were reduced, and secretions of MMPs, IL-6 and IL-8 were also reduced. GSZD decreased the releases of TNF-α and IL-1β in LPS stimulated RAW 264.7 cells. Further studies showed GSZD up-regulated mRNA expressions of Caspase-3, -9, Bax, and SOCS1, whereas down-regulated mRNA expressions of Bcl-2, JAK2, STAT3 and STAT5. Besides, 13 major chemical components were identified in GSZD extracts through HPLC-QqQ-MS analysis. Our results suggested GSZD possesses an anti-rheumatic effect on CIA rats, and the possible mechanism is related to inhibiting inflammatory response, inhibiting invasion and migration of synovial fibroblasts, and inducing apoptosis in synovial fibroblasts.

Zhang Q ，Peng W ，Wei S ，Wei D ，Li R ，Liu J ，Peng L ，Yang S ，Gao Y ，Wu C ，Pu X ... -  《-》

Guizhi-Shaoyao-Zhimu decoction attenuates bone erosion in rats that have collagen-induced arthritis via modulating NF-κB signalling to suppress osteoclastogenesis.

Wei SJ ，Zhang Q ，Xiang YJ ，Peng LY ，Peng W ，Ren Q ，Gao YX ... -  《-》

Inducing Apoptosis and Suppressing Inflammatory Reactions in Synovial Fibroblasts are Two Important Ways for Guizhi-Shaoyao-Zhimu Decoction Against Rheumatoid Arthritis.

Guizhi-Shaoyao-Zhimu decoction (GSZD) is often applied to control rheumatoid arthritis (RA), gout, osteoarthritis, etc. In this study, bioinformatic analysis and experimental verification were used to uncover the integral mechanism profile of GSZD against RA. The chemical compositions of GSZD were identified by UPLC-QTOF-MS/MS. MH7A cell model was established to screen active compounds in GSZD, and potential targets of these compounds were predicted through online database retrieval. The differential expression genes (DEGs) in synovial tissue of RA patients and normal controls were retrieved from the GEO database. DEGs and the predicated compounds targets were overlapped, and the overlapped genes were subsequently enriched by GO and KEGG analysis. The pathways with significant enrichments were further experimentally verified. A total of 19 constituents were identified from GSZD, and 11 compounds showed obviously antiproliferative effects on MH7A cells with IC50 < 100 μg/mL. Bioinformatic analysis indicated that IL-1β, IL-6, MAPK8, JAK2, CXCL8, and CASP3 were the main targets of GSZD, and the integral pharmacological mechanisms profile of GSZD might be related to anti-inflammation and proapoptosis. GSZD can promote the loss of mitochondrial membrane potential (MOMP) and induce apoptosis in MH7A cells. Furthermore, in vitro experiments showed GSZD can not only downregulate mRNA expressions of IL-1β (p<0.05), IL-6 (p<0.05), MMPs (p<0.05) and CCL5 (p<0.05) but also inhibit the nuclear transcription of NF-κB. GSZD also reduced the expressions of Bcl-2 (p<0.05), JAK2 (p<0.05), STAT-3 (p<0.05), whereas increase Bax (p<0.05), Caspase-3 (p<0.05) and caspase-9 (p<0.05). Collectively, inducing synovial fibroblast apoptosis and inhibiting inflammatory response are two important ways for GSZD to RA, and our study proved bioinformatic analysis combined with experimental verification is a feasible method to explore the drug targets and mechanism of actions of TCMs.

Zhang Q ，Duan HX ，Li RL ，Sun JY ，Liu J ，Peng W ，Wu CJ ，Gao YX ... -  《Journal of Inflammation Research》

Guizhi-Shaoyao-Zhimu decoction attenuates rheumatoid arthritis partially by reversing inflammation-immune system imbalance.

Guo Q ，Mao X ，Zhang Y ，Meng S ，Xi Y ，Ding Y ，Zhang X ，Dai Y ，Liu X ，Wang C ，Li Y ，Lin N ... -  《Journal of Translational Medicine》

Tibetan medicine Ershiwuwei Lvxue Pill attenuates collagen-induced arthritis via inhibition of JAK2/STAT3 signaling pathway.

Ershiwuwei Lvxue Pill (ELP, མགྲིན་མཚལ་ཉེར་ལྔ།), a traditional Tibetan medicine preparation, has been used hundreds of years for the clinical treatment of rheumatoid arthritis (RA) in the highland region of Tibet, China. However, the underlying mechanism of its therapeutic effect remains unclear. The present study aimed to investigate the potential pharmacological mechanisms of anti-arthritic effect of ELP. The main chemical constituents of ELP were analyzed by ultra-performance liquid chromatography quadrupole-time-flight mass spectrometry (UPLC-Q-TOF-MS). Forty-eight male Wistar rats (220 ± 20 g) were randomly divided into six groups: normal group, collagen-induced arthritis (CIA) group, methotrexate group (1.05 mg/kg), ELP groups (115, 230 and 460 mg/kg). CIA rat models were assigned to evaluate the anti-RA activity of ELP by determining the paws swelling, arthritis score, organ coefficients of spleen and thymus, and histopathological analysis of knee joints of synovial tissues. The levels of TNF-α, IL-10, IL-6 and IL-17 in serum were measured by ELISA. In addition, mRNA and protein expression levels associated with JAK2/STAT3 signaling pathway in synovial tissues of CIA rats were detected by qRT-PCR, immunohistochemistry and Western blot analyses. Fourteen main chemical constituents of ELP were quantitatively determined by UPLC-Q-TOF-MS analysis. Treatment with ELP reduced the paw swelling, arthritis score and organ coefficients of spleen and thymus. Histopathological examination revealed the protective effects of ELP on CIA rats with knee joint injury. The levels of serum pro-inflammatory cytokines (TNF-α, IL-6 and IL-17) were markedly reduced while the anti-inflammatory cytokine IL-10 was significantly increased with the treatment of ELP. Further investigations showed ELP down-regulated the mRNA and protein expression levels of Bcl-2, whereas up-regulated Bax, SOCS1 and SOCS3. Meanwhile, the ratios of p-JAK2/JAK2 and p-STAT3/STAT3 proteins from synovial tissues were dramatically decreased with the treatment of ELP, whereas no changes of the mRNA and protein expression levels of JAK2 and STAT3 were observed. These results indicated that ELP reduced the severity of arthritis and joint swelling, suggesting an antirheumatic effect on CIA rats. The possible mechanism is related to inhibiting inflammatory response and inducing apoptosis in synovial tissues by regulating JAK2/STAT3 signaling pathway. However, further in vivo and in vitro investigations are still needed to clarify the underlying mechanism of ELP in treating RA.

Liu C ，Zhao Q ，Zhong L ，Li Q ，Li R ，Li S ，Li Y ，Li N ，Su J ，Dhondrup W ，Meng X ，Zhang Y ，Tu Y ，Wang X ... -  《-》