OBJECTIVE: To examine the mechanism of action of tanshinone II A (Tan II A) in promoting chemosensitization of osteosarcoma cells to cisplatin (DDP). METHODS: The effects of different concentrations of Tan II A (0-80 µ mol/L) and DDP (0-2 µ mol/L) on the proliferation of osteosarcoma cell lines (U2R, U2OS, 143B, and HOS) at different times were examined using the cell counting kit-8 and colony formation assays. Migration and invasion of U2R and U2OS cells were detected after 24 h treatment with 30 µ mol/L Tan II A, 0.5 µ mol/L DDP alone, and a combination of 10 µ mol/L Tan II A and 0.25 µ mol/L DDP using the transwell assay. After 48 h of treatment of U2R and U2OS cells with predetermined concentrations of each group of drugs, the cell cycle was analyzed using a cell cycle detection kit and flow cytometry. After 48 h treatment, apoptosis of U2R and U2OS cells was detected using annexin V-FITC apoptosis detection kit and flow cytometry. U2R cells were inoculated into the unilateral axilla of nude mice and then the mice were randomly divided into 4 groups of 6 nude mice each. The 4 groups were treated with equal volume of Tan II A (15 mg/kg), DDP (3 mg/kg), Tan II A (7.5 mg/kg) + DDP (1.5 mg/kg), and normal saline, respectively. The body weight of the nude mice was weighed, and the tumor volume and weight were measured. Cell-related gene and signaling pathway expression were detected by RNA sequencing and Kyoto Encyclopedia of Genes and Genomes pathway analysis. p38 MAPK signaling pathway proteins and apoptotic protein expressions were detected by Western blot. RESULTS: In vitro studies have shown that Tan II A, DDP and the combination of Tan II A and DDP inhibit the proliferation, migration and invasion of osteosarcoma cells. The inhibitory effect was more pronounced in the Tan II A and DDP combined treatment group (P<0.05 or P<0.01). Osteosarcoma cells underwent significantly cell-cycle arrest and cell apoptosis by Tan II A-DDP combination treatment (P<0.05 or P<0.01). In vivo studies demonstrated that the Tan II A-DD combination treatment group significantly inhibited tumor growth compared to the Tan II A and DDP single drug group (P<0.01). Additionally, we found that the combination of Tan II A and DDP treatment enhanced the p38 MAPK signaling pathway. Western blot assays showed higher p-p38, cleaved caspase-3, and Bax and lower caspase-3, and Bcl-2 expressions with the combination of Tan II A and DDP treatment compared to the single drug treatment (P<0.01). CONCLUSION Tan II A synergizes with DDP by activating the p38/MAPK pathway to upregulate cleaved caspase-3 and Bax pro-apoptotic gene expressions, and downregulate caspase-3 and Bcl-2 inhibitory apoptotic gene expressions, thereby enhancing the chemosensitivity of osteosarcoma cells to DDP.
Abietanes/therapeutic use* ; Osteosarcoma/enzymology* ; Cisplatin/therapeutic use* ; Humans ; Cell Line, Tumor ; Animals ; Apoptosis/drug effects* ; Mice, Nude ; Cell Proliferation/drug effects* ; Cell Movement/drug effects* ; p38 Mitogen-Activated Protein Kinases/metabolism* ; MAP Kinase Signaling System/drug effects* ; Bone Neoplasms/enzymology* ; Cell Cycle/drug effects* ; Xenograft Model Antitumor Assays ; Mice ; Drug Resistance, Neoplasm/drug effects* ; Neoplasm Invasiveness ; Mice, Inbred BALB C

Salvianolic acid B(Sal B), tanshinone Ⅱ_A(TSN Ⅱ_A), and glycyrrhetinic acid(GA) lipid emulsion(GTS-LE) was prepared by the high-speed dispersion method combined with ultrasonic emulsification.The preparation process of the emulsion was optimized by single-factor method and D-optimal method with appearance, centrifugal stability, and particle size of the emulsion as evalua-tion indexes, followed by verification.In vitro release of Sal B, TSN Ⅱ_A, and GA in GTS-LE was performed by reverse dialysis.In vivo pharmacokinetic evaluation was carried out in mice.The acute liver injury model was induced by acetaminophen.The effect of oral GTS-LE on the acute liver injury was investigated by serum liver function indexes and pathological changes in liver tissues of mice.The results showed that under the optimal preparation process, the average particle size of GTS-LE was(145.4±9.25) nm and the Zeta potential was(-33.6±1.45) mV.The drug-loading efficiencies of Sal B, TSN Ⅱ_A, and GA in GTS-LE were above 95%, and the drug release in vitro conformed to the Higuchi equation.The pharmacokinetic results showed that the C_(max) of Sal B, TSN Ⅱ_A, and GA in GTS-LE was 3.128, 2.7, and 2.85 times that of the GTS-S group, and AUC_(0-t) of Sal B, TSN Ⅱ_A, and GA in GTS-LE was 3.09, 2.23, and 1.9 times that of the GTS-S group.After intragastric administration of GTS-LE, the activities of alanine aminotransferase and aspartate aminotransferase were significantly inhibited, the content of malondialdehyde was reduced, and the structure of hepatocytes recovered to normal.In conclusion, GTS-LE can delay the release of Sal B and promote the release of TSN Ⅱ_A and GA.The encapsulation of three drug components in the emulsion can improve the oral bioavailability to varying degrees and can effectively prevent the acute liver injury caused by acetaminophen.
Abietanes/therapeutic use* ; Acetaminophen/therapeutic use* ; Alanine Transaminase/metabolism* ; Animals ; Antipyretics/therapeutic use* ; Aspartate Aminotransferases/metabolism* ; Benzofurans/therapeutic use* ; Chemical and Drug Induced Liver Injury/prevention & control* ; Depsides/therapeutic use* ; Emulsions ; Glycyrrhetinic Acid/therapeutic use* ; Liver/drug effects* ; Malondialdehyde ; Mice

OBJECTIVE: To explore the therapeutic mechanism of tanshinone IIA in the treatment of pulmonary arterial hypertension (PAH) in rats. METHODS: A total of 100 male SD rats were randomized into 5 groups (n=20), and except for those in the control group with saline injection, all the rats were injected with monocrotaline (MCT) on the back of the neck to establish models of pulmonary hypertension. Two weeks after the injection, the rat models received intraperitoneal injections of tanshinone IIA (10 mg/kg), phosphatidylinositol 3 kinase (PI3K) inhibitor (1 mg/kg), both tanshinone IIA and PI3K inhibitor, or saline (model group) on a daily basis. After 2 weeks of treatment, HE staining and α-SMA immunofluorescence staining were used to evaluate the morphology of the pulmonary vessels of the rats. The phosphorylation levels of PI3K, protein kinase B (PKB/Akt) and endothelial nitric oxide synthase (eNOS) in the lung tissue were determined with Western blotting; the levels of eNOS and NO were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: The results of HE staining and α-SMA immunofluorescence staining showed that tanshinone IIA effectively inhibited MCT-induced pulmonary artery intimamedia thickening and muscularization of the pulmonary arterioles (P < 0.01). The results of Western blotting showed that treatment with tanshinone IIA significantly increased the phosphorylation levels of PI3K, Akt and eNOS proteins in the lung tissue of PAH rats; ELISA results showed that the levels of eNOS and NO were significantly decreased in the rat models after tanshinone IIA treatment (P < 0.01). CONCLUSION Treatment with tanshinone IIA can improve MCT-induced pulmonary hypertension in rats through the PI3K/Akt-eNOS signaling pathway.
Abietanes ; Animals ; Hypertension, Pulmonary/drug therapy* ; Male ; Monocrotaline/toxicity* ; Nitric Oxide Synthase Type III/therapeutic use* ; Phosphatidylinositol 3-Kinase/pharmacology* ; Phosphatidylinositol 3-Kinases/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Pulmonary Artery ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

Tanshinone IIA is a pharmacologically active compound isolated from Danshen (Salvia miltiorrhiza), a traditional Chinese herbal medicine for the management of cardiac diseases and other disorders. But its underlying molecular mechanisms of action are still unclear. The present investigation utilized a data mining approach based on network pharmacology to uncover the potential protein targets of Tanshinone IIA. Network pharmacology, an integrated multidisciplinary study, incorporates systems biology, network analysis, connectivity, redundancy, and pleiotropy, providing powerful new tools and insights into elucidating the fine details of drug-target interactions. In the present study, two separate drug-target networks for Tanshinone IIA were constructed using the Agilent Literature Search (ALS) and STITCH (search tool for interactions of chemicals) methods. Analysis of the ALS-constructed network revealed a target network with a scale-free topology and five top nodes (protein targets) corresponding to Fos, Jun, Src, phosphatidylinositol-4, 5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA), and mitogen-activated protein kinase kinase 1 (MAP2K1), whereas analysis of the STITCH-constructed network revealed three top nodes corresponding to cytochrome P450 3A4 (CYP3A4), cytochrome P450 A1 (CYP1A1), and nuclear factor kappa B1 (NFκB1). The discrepancies were probably due to the differences in the divergent computer mining tools and databases employed by the two methods. However, it is conceivable that all eight proteins mediate important biological functions of Tanshinone IIA, contributing to its overall drug-target network. In conclusion, the current results may assist in developing a comprehensive understanding of the molecular mechanisms and signaling pathways of in a simple, compact, and visual manner.
Abietanes ; pharmacology ; therapeutic use ; Data Mining ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Heart Diseases ; drug therapy ; Humans ; Pharmacology ; Phytotherapy ; Proteins ; metabolism ; Salvia miltiorrhiza ; chemistry

AIM: This study was designed to evaluate the anti-cancer actions of tanshinone I and tanshinone IIA, and six derivatives of tanshinone IIA on normal and cancerous colon cells. Structure activity relationship (SAR) analysis was conducted to delineate the significance of the structural modifications of tanshinones for improved anti-cancer action. METHOD: Tanshinone derivatives were designed and synthesized according to the literature. The cytotoxicity of different compounds on colon cancer cells was determined by the MTT assay. Apoptotic activity of the tanshinones was measured by flow cytometry (FCM). RESULTS: Tanshinone I and tanshinone IIA both exhibited significant cytotoxicity on colon cancer cells. They are more effective in p53(+/+) colon cancer cell line. It was also noted that the anti-cancer activity of tanshinone I was more potent and selective. Two of the derivatives of tanshinone IIA (N1 and N2) also exhibited cytotoxicity on colon cancer cells. CONCLUSION The anti-colon cancer activity of tanshinone I was more potent and selective than tanshinone IIA, and is p53 dependent. The derivatives obtained by structural modifications of tanshinone IIA exhibited lower cytotoxicity on both normal and colon cancer cells. From steric and electronic characteristics point of view, it was concluded that structural modifications of ring A and furan or dihydrofuran ring D on the basic structure of tanshinones influences the activity. An increase of the delocalization of the A and B rings could enhance the cytotoxicity of such compounds, while a non-planar and small sized D ring region would provide improved anti-cancer activity.
Abietanes ; chemistry ; pharmacology ; therapeutic use ; Antineoplastic Agents, Phytogenic ; chemistry ; pharmacology ; therapeutic use ; Cell Line ; Colon ; drug effects ; Colonic Neoplasms ; drug therapy ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; therapeutic use ; HCT116 Cells ; HT29 Cells ; Humans ; Phytotherapy ; Salvia miltiorrhiza ; chemistry ; Structure-Activity Relationship

OBJECTIVE: To determine the effect of Tanshinone IIA (TanIIA) on the phosphory-lated NMDA receptor 1 at Serine 897 site (phospho-NR1 S897) and intracellular free calcium concentration ([Ca(2+)](i)) in neonatal SD rats with hypoxic ischemic brain damage (HIBD), and to explore the neuroprotective mechanism of TanIIA in HIBD. METHODS: Neonatal SD rats were randomly divided into a normal control, and an HIBD and TanIIA+HIBD group. Rice-Vannucci method was used for HIBD animal model. Time points were: 3, 6, 12, and 24 h after HIBD (n=10 in each group at each time point). TanIIA was intraperitoneally given at 1 μg/g every 12 h. Fura-2AM was used to mark the fluorescent calcium probe and [Ca(2+)](i) was measured by a Hitachi F-4500 Fluorescence Spectrophometer. Fluorescent immunohisotichemical study was used for the expression of phospho-NR1 S897. RESULTS: (1) Compared with the normal control group, both the [Ca(2+)](i) absolute number and ipsi-/contra-lateral ratio were increased at each time point with statistical significance (P<0.05). Compared with the HIBD group, the [Ca(2+)](i) in the HIBD+ TanIIA group was decreased at each time point. At 24 h after HIBD, the ipsi-/contra-lateral ratio of HIBD+ TanIIA group was 24.9% less than that of HIBD group with statistical significance (P<0.05). (2) In the normal control group, abundant phospho-NR1 S897 positive cells were nicely distributed in the cortex. Compared with the normal control group, at each time point, both the absolute number of phospho-NR1 S897 positive cells and the fluorescent intensity of phospho-NR1 S897 in the ipsilateral cortex of the HIBD group were decreased with statistical significance (P<0.05). Compared with the HIBD group, both the absolute number of phospho-NR1 S897 positive cells and the fluorescent intensity of phospho-NR1 S897 in the ipsilateral cortex of HIBD+ TanIIA were increased. There was significant difference at 3 and 12 h after the HIBD (P<0.05). CONCLUSION TanIIA reduced the HIBD-caused down-regulation of phospho-NR1 S897 and the HIBD-caused [Ca(2+)](i) elevation in the cortex. The neuroprotective effect of TanIIA may be related to influencing NMDA receptor expression and decreasing intracellular free calcium aggregation.
Abietanes ; pharmacology ; therapeutic use ; Animals ; Animals, Newborn ; Calcium ; metabolism ; Female ; Hypoxia-Ischemia, Brain ; drug therapy ; metabolism ; Male ; Neuroprotective Agents ; pharmacology ; therapeutic use ; Phosphorylation ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate ; genetics ; metabolism