Pathway of PPAR-gamma coactivators in thermogenesis: a pivotal traditional Chinese medicine-associated target for individualized treatment of rheumatoid arthritis

Abstract

Traditional Chinese medicine (TCM) syndromes have been regarded as the crucial clinical manifestations for individualized diagnosis and treatment of complex diseases, including rheumatoid arthritis (RA) and cancer. Especially, RA patients are classified into cold and hot syndromes with different clinical manifestations, interventions and molecular mechanisms. Better effectiveness of a classic cold syndrome-specific herbal formula Wu-tou decoction (WTD) has been achieved. To explore molecular mechanisms of syndrome-specific formulae is of great clinical significance to improve the effectiveness and pertinence of treatment for the complex diseases with personalized conditions. However, the scientific basis of WTD treatment on RA with the cold syndrome remains unclear. Here, we predicted the putative targets for composite compounds contained in WTD using drugCIPHER-CS and constructed a WTD herbs-putative targets-RA related genes network. Next, a list of major WTD targets was identified based on their topological features, including the degree, node betweenness, closeness and k-coreness in the above pharmacological network. Importantly, pathway enrichment analysis revealed that these major WTD targets were significantly associated with the pathway of peroxisome proliferator-activated receptor (PPAR)-gamma (PPAR-γ) coactivators in thermogenesis. These computational findings were subsequently verified by experiments on a rat model of collagen-induced arthritis (CIA) with cold or hot syndromes, and on human fibroblast-like synoviocytes-rheumatoid arthritis (HFLS-RA) cell line. In conclusion, the pathway of PPAR-γ coactivators in thermogenesis might be one of the potential pharmacological targets of WTD to alleviate RA with the TCM cold syndrome. These findings may open new avenues for designing individualized treatment regimens for RA patients.

Keywords: PPAR-gamma; individualized treatment; network pharmacology; rheumatoid arthritis; traditional Chinese medicine syndrome.

Figure 1. Similarities and differences between RA patients with hot and cold syndromes with regard to the clinical manifestations, therapeutic strategies and molecular mechanisms

Red marks denote the hot syndrome, and blue marks denote the cold syndrome.

Figure 2. A schematic diagram of the systematic strategies for uncovering the pharmacological mechanisms of the herbal formula Wu-tou decoction acting on RA with the Traditional Chinese Medicine cold syndrome

Figure 3

A. Network based on the direct interactions among five herbs (Radix Aconiti, Herba Ephedrae, Radix Astragali, Radix Glycytthizae and Raidix Paeoniae Alba), putative targets of WTD and known RA-related targets (herbs-putative targets-known RA targets network). Circular nodes refer to the putative targets of WTD; Triangle nodes refer to the known RA-related targets; Nodes with red highlights refer to the overlap between the putative targets of WTD and known RA-related targets; The size of the nodes were ordered according to their degree in the network. B. Network based on the direct interactions among the five herbs (Radix Aconiti, Herba Ephedrae, Radix Astragali, Radix Glycytthizae and Raidix Paeoniae Alba), major nodes of herbs-putative targets-known RA targets network and their associated pathways (herbs-major putative targets-pathway network). Nodes with yellow highlights refer to the major components involved into the corresponding pathways.

Figure 4. Schematic diagram of the “Role of PPAR-gamma Coactivators in Obesity and Thermogenesis.”

Figure 5. Effects of WTD on the severity of arthritis in CIA rats

A. Macroscopic performance of arthritis, including erythema and swelling, was clearly observed in the CIA CIA-cold/hot model groups, whereas doses of 3.75 g/(kg•day) WTD significantly ameliorated the development and severity of arthritis in SD rats in the CIA-cold model groups; B. Doses of 3.75 and 7.5 g/(kg•day) WTD significantly decreased the mean arthritis score in the CIA-cold model group; C. Low-high doses of WTD significantly decreased the arthritis incidence in a dose-dependent manner in the CIA-cold model groups; D. Doses of 3.75 and 7.5 g/(kg•day) WTD significantly delayed the time when arthritis first appeared in the CIA-cold model groups; E. Low-high doses of WTD significantly decreased the percentage of arthritis limbs in the CIA-cold model groups; WTD could also decrease the severity of arthritis in CIA rats in the CIA-hot model group without statistical significance. Data are represented as the mean±S.D. (n=16). *, **, and ***, P<0.05, P<0.01, and P<0.001, comparison with the control group. ^#, ^##, ^###, P<0.05, P<0.01, and P<0.001, comparison with the CIA model group. ^@, ^@@, ^@@@, P<0.05, P<0.01, and P<0.001, comparison with the CIA-cold/hot model groups.

Figure 6. Effects of WTD on the temperature of the articular surface and the blood flow volume in CIA rats

A. The temperature of the articular surface in CIA, CIA-cold/hot model groups were markedly increased compared with the control group, while doses of 3.75 g/(kg•day) WTD significantly decreased the temperature in both the CIA-cold /hot model groups; B. Blood flow volume in the joints tend to increase in the CIA, CIA-cold/hot model groups compared with the control groups, and doses of 3.75 g/(kg•day) WTD increased the blood flow volume in both the CIA-cold/hot model groups; C. The temperature of the articular surface were markedly up-regulated in the CIA, CIA-cold/hot model groups, which were markedly reversed by low-high doses of WTD in the CIA-cold groups. No statistical significance was observed in the CIA-hot model groups; D. WTD could increase the blood flow volume in the joints in both the CIA-cold/hot model groups without a significant difference. Data are represented as the mean±S.D (n=16). *, **, and ***, P<0.05, P<0.01, and P<0.001, comparison with the control group. ^#, ^##, ^###, P<0.05, P<0.01, and P<0.001, comparison with the CIA model group. ^@, ^@@, ^@@@, P<0.05, P<0.01, and P<0.001, comparison with the CIA-cold/hot model groups.

Figure 7. Effect of WTD on radiological changes and histologic lesions in CIA rats

A. histologic observations of the joints in rats (HE staining); B. displays the clinical manifestation of CIA rats on day 21 after immunization, doses of 3.75 g/(kg•day) WTD improved paw swelling in the CIA-cold/hot model groups; C. D. and E. bone erosion, joint space and the degree of cartilage damage in joints, respectively, as described in the methods section. Data are represented as the mean±S.D (n=16). *, **, and ***, P<0.05, P<0.01, and P<0.001, comparison with the control group. ^#, ^##, ^###, P<0.05, P<0.01, and P<0.001, comparison with the CIA model group. ^@, ^@@, ^@@@, P<0.05, P<0.01, and P<0.001, comparison with the CIA-cold/hot model groups.

Figure 8

Effect of WTD on the expression of PPAR-γ A. RXR-α B. MED1 C. NCOA1 D. NCOA2 E. and CBP F. proteins in the joint part of CIA rats detected using western blotting analysis. Data are represented as the mean±S.D (n=16). *, **, and ***, P<0.05, P<0.01, and P<0.001, comparison with the control group. ^#, ^##, ^###, P<0.05, P<0.01, and P<0.001, comparison with the CIA model group. ^@, ^@@, ^@@@, P<0.05, P<0.01, and P<0.001, comparison with the CIA-cold/hot model groups.

Figure 9

Effect of WTD on the expression of PPAR-γ A. RXR-α B. MED1 C. NCOA1 D. NCOA2 E. and CBP F. proteins in HFLS-RA. Data are represented as the mean±S.D. *, **, and ***, P<0.05, P<0.01, and P<0.001, comparison with the control group. ^#, ^##, ^###, P<0.05, P<0.01, and P<0.001, comparison with the model group.