Context: Chlorogenic acid (CGA) and luteolin (Lut) are the predominant constituents of Caulis Lonicerae, which is usually used in the treatment for rheumatoid arthritis (RA). Objective: In this study, we investigated whether CGA and Lut could synergistically inhibit the proliferation of fibroblast-like synoviocytes (FLSs) in RA synovial tissues. Methods: Rat FLS cells (RSC-364) induced by interleukin (IL)-1β were treated by CGA, Lut or both of them. The apoptosis rates were detected by flow cytometer. Protein expression of key molecules of NF-κB and JAK/STAT signaling pathways were detected by Western blot. Results and discussion: Treatment with CGA and Lut inhibited the proliferation of RSC-364 cells stimulated by IL-1β significantly and induced cell apoptosis notably. The ratio of apoptosis in RSC-364 cells induced with IL-1β accompanied by both CGA and Lut increased approximately 7-fold compared with those incubated with IL-1β alone. The results of immunoblot analysis revealed that the key molecules involved in the NF-κB and JAK/STAT-signaling pathways, including NF-κB p50, p100, IKKα/β, gp103, JAK1 and STAT3, were decreased significantly in RSC-364 cells treated by IL-1β plus CAG and Lut compared with those incubated with IL-1β alone. Additionally, the amounts of phospho-IKKα/β and phospho-STAT3 were also decreased significantly in cells treated with CGA and Lut. Furthermore, the synergistic effect of CGA and Lut was superior to the effect of one of these two ingredients. Conclusion: Our finding suggested that the combination of CGA and Lut may be a potential therapeutic treatment for the inflammatory proliferation of synoviocytes in patients with RA.

Xian-Fang-Huo-Ming-Yin (XFHM), a traditional herbal formula, has been used to treat sores and carbuncles for hundreds of years in Asia. Nowadays, its clinical effects in treatment of rheumatoid arthritis (RA) have been validated. In this study, we want to study its possible molecular mechanisms of regulating the differentiation of lymphocytes and production of pro-inflammatory cytokines in collagen-induced arthritis (CIA) mice for RA treatment. A high performance liquid chromatography-electrospray ionization/mass spectrometer (HPLC-ESI/MSn) system was used to analyze the constituents of XFHM granules. An arthritics mouse model was induced by collagen and leflunomide (LEF) was used as a positive control medicine. Pathological changes at the metatarsophalangeal joint were studied through Safranin O and immunohistochemical staining. The differentiation of T, B and NK cells was examined by flow cytometry and pro-inflammatory cytokines were assayed using an Inflammation Antibody Array assay. The expression of key molecules of the nuclear factor κB (NF-κB) and Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathways in spleen were studied by western-blot analysis. In our study. 21 different dominant chemical constituents were identified in XFHM. Treatment with XFHM suppressed the pathological changes in arthrosis of CIA. Additionally, XFHM down-regulated the proliferation and differentiation of CD3+ T cells and CD3−CD19+ B cells significantly. However, XFHM had no significant effect on CD3−NK1.1+ NK cells. Further study showed that the production of pro-inflammatory cytokines had been suppressed by inhibiting the activation of NF-κB and JAK/STAT signaling. XFHM can regulate and maintain the immunologic balance of lymphocytic immunity and inhibit the production of pro-inflammatory cytokines, thus suppressing the pathological changes of RA. Therefore, XFHM may be used as an application of traditional medicine against RA in modern complementary and alternative therapeutics.

Angelicae Sinensis, Radix Astragali and Rhizoma Coptidis are all herbs of modified Danggui Buxue Tang (DGBX) and are extensively applied herbs in traditional Chinese medicine for the treatment of anemia and inflammation. In this study, immune-induced AA mice were used as an animal model, and the immunosuppressive agent, Ciclosporin A (CsA), was used as a positive control. Multiple pro-inflammatory cytokines were examined by bead-based multiplex flow cytometry. The T-cell subsets were assessed using a fluorescence-activated cell sorter (FACS). Western blot analysis was used to estimate the protein expression levels of specific transcription factors for T helper cells (Th1, Th2 and Th17) and key molecules of the Janus-activated kinase (Jak)/signal transducer and activator of transcription (Stat3) signaling pathway. DGBX treatment could significantly increase the production of whole blood cells in peripheral blood (PB); inhibit the expansion of Th1 and Th17 cells; increase the differentiation of Th2 and Tregs cells; regulate the expression levels of T-bet, GATA-3, RORγ and proinflammatory cytokines; and decrease the expression levels of key molecules in the Jak/Stat signaling pathway. These results indicate that DGBX can regulate the differentiation of T lymphocytes, resulting in immunosuppressive and hematogenic functions on AA mice. DGBX might be a good candidate for inclusion in a randomized study for AA with more data on the possible side effects and doses used in humans. Ultimately, it may be used for applications of traditional medicine against AA in modern complementary and alternative immunosuppressive therapeutics.

Background: Radix AstragaliandRadix Angelicae Sinensisare two herbs that compose Danggui Buxue Tang (an herbal formula for treatment of anemia diseases). In this study, we explored the molecular mechanism and effective targets to immune destruction of bone marrow (BM) cells treated withRadix Astragali,Radix Angelicae Sinensisor a combination of two agents. The potential synergic advantages of two herbs should also be explored.Methods: The constituents ofRadix AstragaliandRadix Angelicae Sinensiswere analyzed by high performance liquid chromatography-electrospray ionization/mass spectrometer system BM cells were separated from limbs of BALB/c mice, and immune destruction was induced with IFN-γ. The percentages of hematopoietic stem cells (HSCs) and CD3⁺T cells were detected by flow cytometry. The distribution of T-bet and changes in the combination of SAP and SLAM in BM cells were observed by immunofluorescence. Western blotting was used to assay the expression of key molecules of the eIF2 signaling pathway in BM cells.Results: Seven constituents ofRadix Astragaliand six constituents ofRadix Angelicae Sinensiswere identified. The percentages of HSCs increased significantly after treatment withRadix Angelicae Sinensis, especially at high concentrations. The percentages of CD3⁺T cells were significantly decreased afterRadix AstragaliandRadix Angelicae Sinensistreatment. However, the synergistic function of two-herb combinations was superior to that of the individual herbs alone. The distribution of T-bet in BM cells was decreased significantly afterRadix Angelicae Sinensistreatment. The number of SLAM/SAP double-stained cells was increased significantly afterRadix Astragalitreatment at low concentrations. The phosphorylation levels of eIF2α were also reduced afterRadix AstragaliandRadix Angelicae Sinensistreatment.Conclusions: Radix AstragaliandRadix Angelicae Sinensiscould intervene in the immunologic balance of T lymphocytes, inhibit the apoptosis of BM cells induced by immune attack, restore the balance of the T cell immune response network and recover the hematopoietic function of HSCs. The synergistic effects ofRadix AstragaliandRadix Angelicae Sinensiswere superior to those of each herb alone.

A derivative formula, DGBX, which is composed of three herbs (Radix astragali,Radix Angelicae sinensis, andCoptis chinensisFranch), is derived from a famous Chinese herbal formula, Danggui Buxue Tang (DBT) (Radix astragaliandRadix Angelicae sinensis). We aimed to investigate the effects of DGBX on the regulation of the balance between proliferation and apoptosis of hematopoietic stem cells (HSCs) due to the aberrant immune response in a mouse model of aplastic anemia (AA). Cyclosporine (CsA), an immunosuppressor, was used as the positive control. Our results indicated that DGBX could downregulate the production of IFNγin bone marrow cells by interfering with the binding between SLAM and SAP and the expressions of Fyn and T-bet. This herbal formula can also inhibit the activation of Fas-mediated apoptosis, interferon regulatory factor-1-induced JAK/Stat, and eukaryotic initiation factor 2 signaling pathways and thereby induce proliferation and attenuate apoptosis of HSCs. In conclusion, DGBX can relieve the immune-mediated destruction of HSCs, repair hematopoietic failure, and recover the hematopoietic function of HSCs in hematogenesis. Therefore, DGBX can be used in traditional medicine against AA as a complementary and alternative immunosuppressive therapeutic formula.