Network pharmacology and experimental validation to reveal the pharmacological mechanisms of Sini decoction against renal fibrosis.

OBJECTIVE: To investigate the mechanism by which Sini decoction (, SND) improves renal fibrosis (Rf) in rats based on transforming growth factor ß1/Smad (TGF-ß1/Smad) signaling pathway. METHODS: Network pharmacology was applied to obtain potentially involved signaling pathways in SND's improving effects on Rf. The targets of SND drug components and the targets of Rf were obtained by searching databases, such as the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCSMP) and GeenCard. The intersection targets of two searches were obtained and underwent signaling pathway analysis using a Venn diagram. Then experimental pharmacology was utilized to prove and investigate the effects of SND on target proteins in the TGF-ß1/Smad signaling pathway. The Rf rat model was established by unilateral ureteral occlusion (UUO). The expression levels of transforming growth factor, matrix metalloproteinase-9 (MMP-9), matrix metal protease-2 (MMP-2), connective tissue growth factor (CTGF), and tissue inhibitor of metalloproteinase-1 (TIMP-1) were determined by Masson staining of rat renal tissue, and immunohistochemical methods. The expression levels of Smad3, Smad2, and Smad7 in renal tissue were determined by Western blotting (WB). The mechanism of the improving effects of SND on Rf was investigated based on TGF-ß1/Smad signaling pathway. RESULTS: A total of 12 drug components of Fuzi (Radix Aconiti Lateralis Preparata), 5 drug components of Ganjiang (Rhizoma Zingiber), and 9 drug components of Gancao (Radix Glycy et Rhizoma) were obtained from the database search, and 207 shared targets were found. A total of 1063 Rf targets were found in the database search. According to the Venn diagram, in total, 96 intersection targets were found in two database searches. The metabolic pathways involved included TGF-ß signaling pathway, phosphatidylinositol-3-kinase/serine-threonine protein kinase signaling (PI3K/Akt) pathway, and hypoxia-inducible factor-1 (HIF-1) signaling pathway. Masson staining analysis showed that compared with the model group, the renal interstitial collagen deposition levels in the SSN and SND groups were significantly lower (P < 0.05). Immunohistochemical analysis, compared with the control group, the positive cell area expression levels of MMP-9/TIMP-1 and MMP-2/TIMP-1 in the kidney tissue of the model group were significantly decreased (P < 0.05, P < 0.01), and the positive cell area expression levels of CTGF and TGF-ß1 were significantly increased (P < 0.01). Compared with the model group, the positive cell area expression levels of MMP-9/TIMP-1 and MMP-2/TIMP-1 in the kidney tissue of the SSN and SND groups were significantly increased (P < 0.05, P < 0.01), and the positive cell area expression levels of CTGF and TGF-ß1 in the kidney tissue were significantly decreased (P < 0.05, P < 0.01). WB results showed that the SSN group and the SND group could reduce the expression of Smad2 and Smad3 (P < 0.05) and increase the expression of Smad7 (P < 0.05).

Shikonin derivatives protect immune organs from damage and promote immune responses in vivo in tumour-bearing mice.

Shikonin, a major component of Lithospermum erythrorhizon and Arnebia euchroma, exhibits antiinflammatory, immunomodulatory and antitumour activities. Although many recent studies have focused on the antitumour effects of shikonin, the exact mechanisms underlying its antitumour and immunomodulatory effects in tumour-bearing mice remain unclear. The aim of the present study was to investigate the antitumour and immunomodulatory effects of shikonin derivatives (ShD) in tumour-bearing mice. Swiss mice inoculated with hepatoma HepA(22) or sarcoma 180 (S(180)) cells were treated with ShD or 5-fluorouracil (5Fu). Survival time, immune organs, natural killer cell activity, lymphocytes, lymphocyte transformation and interleukin (IL)-2 production were analysed. ShD significantly prolonged the survival (median survival time prolonged by >7 days) of tumour-bearing mice in a dose-dependent manner, inhibited the growth of transplantable neoplasms (inhibitory rate, > 33%), and recovered (at [ShD] = 2.5 mg/kg/day) or increased (at [ShD] > 5 mg/kg/day) the number of CD3- and CD19-positive cells. ShD also played a role in protecting the immune organs from damage and reversed or enhanced immune responses, as noted by the nearly normal thymic structure; enlarged splenic corpuscles; and improved natural killer cell activity, lymphocyte transformation and IL-2 production in ShD-treated mice. ShD reduced the tumour load of tumour-bearing mice and protected the immune organs against tumour-induced damage and immune function impairment.