Inhibitory Effect Of Tanshinone IIA Research Articles

Inhibitory Effect of Tanshinone IIA Extracted from Radix Salviae miltiorrhizae on Melanoma B16F10 Cells

In this study, we investigate the inhibitory effects of Tanshinone IIA on the proliferation and migration of B16F10 melanoma cells and its potential mechanism of action. We cultured B16F10 melanoma cells in vitro and treated them with varying concentrations of tanshinone IIA. The inhibitory effect of tanshinone IIA on B16F10 cells was assessed using the CCK-8 assay, employing methyl thiazolyl tetrazolium. Melanin content was determined by the NaOH cleavage method, while expression levels of melanin-related proteins were analyzed by Western blotting, and migration-related proteins were quantified using ELISA. When the concentration of Tanshinone IIA reached 20 and 40 μg/mL, there was a decrease in the survival rate of B16F10 melanoma cells. The proliferation inhibition rate of B16F10 melanoma cells gradually increased with the duration of drug action and the concentration of Tanshinone IIA. After 48 hours, there was an increase in melanin content. A comparison of melanin synthesis at the three concentrations yielded an F value of 61.820 and P <0.001. The relative expression levels of TYR, TRP-1, TRP-2, and MITF increased when treated with different concentrations of Tanshinone IIA (P < 0.05). MMP-2 and MMP-9 were downregulated when the concentration of Tanshinone IIA was at 40 and 80 μg/mL respectively (P <0.05). In conclusion, our study demonstrated the inhibitory effects of Tanshinone IIA on growth, proliferation, migration, and melanin synthesis in B16F10 melanoma cells. However, further research is needed to elucidate the specific signal transduction pathway activated by Tanshinone IIA.

Tanshinone IIA Alleviates Traumatic Brain Injury by Reducing Ischemia‒Reperfusion via the miR-124-5p/FoxO1 Axis.

Cerebral ischemia-reperfusion injury is a common complication of ischemic stroke that affects the prognosis of patients with ischemic stroke. The lipid-soluble diterpene Tanshinone IIA, which was isolated from Salvia miltiorrhiza, has been indicated to reduce cerebral ischemic injury. In this study, we investigated the molecular mechanism of Tanshinone IIA in alleviating reperfusion-induced brain injury. Middle cerebral artery occlusion animal models were established, and neurological scores, tetrazolium chloride staining, brain volume quantification, wet and dry brain water content measurement, Nissl staining, enzyme-linked immunosorbent assay, flow cytometry, western blotting, and reverse transcription-quantitative polymerase chain reaction were performed. The viability of cells was measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assays, while cell damage was measured by lactate dehydrogenase release in the in vitro oxygen glucose deprivation model. In addition, enzyme-linked immunosorbent assay, flow cytometry, western blotting, and reverse transcription-quantitative polymerase chain reaction were used to evaluate the therapeutic effect of Tanshinone IIA on ischemia/reperfusion (I/R) induced brain injury, as well as its effects on the inflammatory response and neuronal apoptosis, in vivo and in vitro. Furthermore, this study validated the targeting relationship between miR-124-5p and FoxO1 using a dual luciferase assay. Finally, we examined the role of Tanshinone IIA in brain injury from a molecular perspective by inhibiting miR-124-5p or increasing FoxO1 levels. After treatment with Tanshinone IIA in middle cerebral artery occlusion-reperfusion (MCAO/R) rats, the volume of cerebral infarction was reduced, the water content of the brain was decreased, the nerve function of the rats was significantly improved, and the cell damage was significantly reduced. In addition, Tanshinone IIA effectively inhibited the I/R-induced inflammatory response and neuronal apoptosis, that is, it inhibited the expression of inflammatory cytokines IL-1β, IL-6, TNF-α, decreased the expression of apoptotic protein Bax and Cleaved-caspase-3, and promoted the expression of antiapoptotic protein Bcl-2. In vitro oxygen-glucose deprivation/reoxygenation (OGD/R) cell model, Tanshinone IIA also inhibited the expression of inflammatory factors in neuronal cells and inhibited the occurrence of neuronal apoptosis. In addition, Tanshinone IIA promoted the expression of miR-124-5p. Transfection of miR-124-5p mimic has the same therapeutic effect as Tanshinone IIA and positive therapeutic effect on OGD cells, while transfection of miR-124-5p inhibitor has the opposite effect. The targeting of miR-124-5p negatively regulates FoxO1 expression. Inhibition of miR-124-5p or overexpression of FoxO1 can weaken the inhibitory effect of Tanshinone IIA on brain injury induced by I/R, while inhibition of miR-124-5p and overexpression of FoxO1 can further weaken the effect of Tanshinone IIA. Tanshinone IIA alleviates ischemic-reperfusion brain injury by inhibiting neuroinflammation through the miR-124-5p/FoxO1 axis. This finding provides a theoretical basis for mechanistic research on cerebral ischemia-reperfusion injury.

Inhibitory effect of tanshinone IIA, resveratrol and silibinin on enterovirus 68 production through inhibiting ATM and DNA-PK pathway

BackgroundHuman enterovirus 68 (EV68) is a primary etiological agent for respiratory illnesses, while no effective drug has yet used in clinics largely because the pathogenesis of EV68 is not clear. DNA damage response (DDR) responds to cellular DNA breaks and is also involved in viral replication. Three DDR pathways includes ataxia telangiectasia mutated (ATM), ATM and Rad3-related (ATR), and DNA-dependent protein kinase (DNA-PK). Natural products proved to be an excellent source for the discovery and isolation of novel antivirals. Among them, tanshinone IIA, resveratrol, silibinin, rutin and quercetin are reported to target DDR, therefore their roles in anti-EV68 are investigated in this study. PurposeThis study investigated the anti-EV68 ability of various natural compounds related to DDR. Study design and methodsThe methods include cell counting, flow cytometry, western blot, Immunofluorescence staining, comet assays, quantitative real-time RT PCR and short interfering RNAs (siRNAs) for analysis of cell number, cell cycle, protein expression, protein location, DNA damage, mRNA level and knock down target gene, respectively. ResultsEV68 infection induced DDR. Down-regulation or inhibition of ATM or DNA-PK lowered DDR in EV68-infected cells and mitigated viral protein expression, however, down-regulation or inhibition of ATR unexpectedly up-regulated DDR, and promoted viral protein expression. Meanwhile tanshinone IIA, resveratrol, and silibinin inhibited ATM and/or DNA-PK activation and decreased viral proliferation, while rutin and quercetin inhibited ATR activation and promoted viral production. The role of them in ATM, DNA-PK and ATR activation was consistent with previous reports. ConclusionTanshinone IIA, resveratrol and silibinin inhibited EV68 proliferation through inhibiting ATM and/or DNA-PK activation, and they were effective anti-EV68 candidates.

Tanshinone IIA attenuates the stemness of breast cancer cells via targeting the miR-125b/STARD13 axis

BackgroundTanshinone II A is an effective component extracted from Salvia miltiorrhiza and the roles of Tanshinone IIA in regulating the stemness of tumor cells remain unclear. This work aims to explore the roles and underlying mechanisms of Tanshinone IIA in breast cancer stemness.MethodsIn vitro mammary spheroid formation, flow cytometry assay on CD24−/CD44+ sub-population, ALDH activity detection, cell viability assay and western blot analysis, and in vivo tumor-initiating analysis were performed to examine the effects of Tanshinone IIA on the stemness of breast cancer cells. MiRNAs-based transcriptome sequencing and data analysis, online dataset analysis, luciferase reporter assay combined with rescuing experiments were constructed to explore the underlying mechanisms.ResultsTanshinone IIA attenuated the stemness of breast cancer cells, evident by downregulating the expression of stemness markers, hindering the capacity of spheroid formation, decreasing the CD24−/CD44+ sub-population in a concentration-dependent manner and reducing the tumor-initiating ability of breast cancer cells. Additionally, Tanshinone IIA enhanced adriamycin sensitivity and attenuated adriamycin resistance of breast cancer cells. Combined with miRNAs-based transcriptome sequencing assay, it was found that Tanshinone IIA downregulated miR-125b level and upregulated its target gene STARD13 (StAR-related lipid transfer protein 13) level, thus inactivating the miR-125b/STARD13 axis, which had been previously confirmed to promote breast cancer progression. Notably, miR-125b overexpression enhanced the stemness of breast cancer cells, and miR-125b overexpression or STARD13 knockdown impaired the inhibitory effects of Tanshinone IIA on the stemness of breast cancer cells.ConclusionsTanshinone IIA could attenuate the stemness of breast cancer cells via targeting the miR-125b/STARD13 axis.

Tanshinone IIA: New Perspective on the Anti-Tumor Mechanism of A Traditional Natural Medicine.

The search for natural and efficacious antineoplastic drugs, with minimal toxicity and side effects, is an important part of antitumor drug research and development. Tanshinone IIA is the most evaluated lipophilic active component of Salvia miltiorrhiza. Tanshinone IIA is a path-breaking traditional drug applied in cardiovascular treatment. It has also been found that tanshinone IIA plays an important role in the digestive, respiratory and circulatory systems, as well as in other tumor diseases. Tanshinone IIA significantly inhibits the proliferation of several types of tumors, blocks the cell cycle, induces apoptosis and autophagic death, in addition to inhibiting cell migration and invasion. Among these, the regulation of tumor-cell apoptosis signaling pathways is the key breakthrough point in several modes of antitumor therapy. The PI3K/AKT/MTOR signaling pathway and the JNK pathway are the key pathways for tanshinone IIA to induce tumor cell apoptosis. In addition to glycolysis, reactive oxygen species and signal transduction all play an active role with the participation of tanshinone IIA. Endogenous apoptosis is considered the main mechanism of tumor apoptosis induced by tanshinone IIA. Multiple pathways and targets play a role in the process of endogenous apoptosis. Tanshinone IIA can protect chemotherapy drugs, which is mainly reflected in the protection of the side effects of chemotherapy drugs, such as neurotoxicity and inhibition of the hematopoietic system. Tanshinone IIA also has a certain regulatory effect on tumor angiogenesis, which is mainly manifested in the control of hypoxia. Our findings indicated that tanshinone IIA is an effective treatment agent in the cardiovascular field and plays a significant role in antitumor therapeutics. This paper reviews the pharmacological potential and inhibitory effect of tanshinone IIA on cancer. It is greatly anticipated that tanshinone IIA will be employed as an adjuvant in the treatment of various cancers.

In vitro and computational insights revealing the potential inhibitory effect of Tanshinone IIA against influenza A virus

Seasonal human influenza is a serious respiratory infection caused by influenza viruses that can be found all over the world. Type A influenza is a contagious viral infection that, if left untreated, can lead to life-threatening consequences. Fortunately, the plant kingdom has many potent medicines with broad-spectrum antiviral activity. Herein, six plant constituents, namely Tanshinone IIA 1, Carnosic acid 2, Rosmarinic acid 3, Glycyrrhetinic acid 4, Baicalein 5, and Salvianolic acid B 6, were screened for their antiviral activities against H1N1 virus using in vitro and in silico approaches. Hence, their anti-influenza activities were tested in vitro to determine inhibitory concentration 50 (IC50) values after measuring their CC50 values using MTT assay on MDCK cells. Interestingly, Tanshinone IIA (TAN) 1 was the most promising member with CC50 = 9.678 μg/ml. Moreover, the plaque reduction assay carried on TAN 1 revealed promising viral inhibition percentages of 97.9%, 95.8%, 94.4%, and 91.7% using concentrations 0.05 μg/μl, 0.025 μg/μl, 0.0125 μg/μl, and 0.006 μg/μl, respectively. Furthermore, in silico molecular docking disclosed the superior affinities of Salvianolic acid B (SAL) 6 towards both surface glycoproteins of influenza A virus (namely, hemagglutinin (HA) and neuraminidase (NA)). The docked complexes of both SAL and TAN inside HA and NA receptor pockets were selected for 100 ns MD simulations followed by MM-GBSA binding free energy calculation to confirm the docking results and give more insights regarding the stability of both compounds inside influenza mentioned receptors, respectively. The selection criteria of the previously mentioned complexes were based on the fact that SAL showed the highest docking scores on both viral HA and NA glycoproteins whereas TAN achieved the best inhibitory activity on the other hand. Finally, we urge more advanced preclinical and clinical research, particularly for TAN, which could be used to treat the human influenza A virus effectively.

Tanshinone IIA inhibits gastric cancer cell stemness through inducing ferroptosis.

Tanshinone IIA is the active constituent extracted from Salvia Miltiorrhza. Numerous studies have shown that Tanshinone IIA could inhibit tumor proliferation and metastasis, including gastric cancer. However, the effect of Tanshinone IIA on gastric cancer cell stemness stays unclear. Here, we found that Tanshinone IIA could reduce gastric cancer cell stemness through detecting spheroid-forming, flow cytometry analysis, and the expression of stemness markers (OCT3/4, ALDH1A1, and CD44). Mechanistically, Tanshinone IIA increased the level of lipid peroxides and decreased glutathione level in gastric cancer cells, both of which are the markers of ferroptosis. Similarly, ferroptosis inducers (erastin, sulfasalazine, and sorafenib) reduced gastric cancer cell stemness. Additionally, the inhibitory effects of Tanshinone IIA on GC cell stemness were reversed by ferroptosis inhibitor (Fer-1) or overexpression of SLC7A11, which is a critical ferroptosis inhibitor. Therefore, we revealed that Tanshinone IIA inhibited the stemness of gastric cancer cells partly through inducing ferroptosis.

Tanshinone IIA regulates microRNA‑125b/foxp3/caspase‑1 signaling and inhibits cell viability of nasopharyngeal carcinoma

Nasopharyngeal carcinoma (NPC) is a common disease with high prevalence worldwide, affecting hundreds of thousands of patients every year. Although its progress can be inhibited by concurrent chemoradiotherapy and platinum-based agents, there is also a need for novel drugs to treat NPC. The present study identified tanshinone IIA as a potent drug that could suppress the proliferation of HK1 cells by enhancing pyroptosis via regulation of the miR-125b/foxp3/caspase-1 signaling pathway. Firstly, the effects of tanshinone IIA on HK1 cells were assessed and it was confirmed that treatment with tanshinone IIA significantly decreased the proliferation of HK1 cells, with increased activity of caspase-3 and caspase-9. Then, the pyroptosis levels after tanshinone IIA administration were detected. The results showed that tanshinone IIA enhanced pyroptosis in a dose-dependent manner. Furthermore, the mechanism underlying the effects of tanshinone IIA on HK1 cells were explored. It was found that transfection with a microRNA (miR)-125b agomir and a small interfering RNA (si)-foxp3 plasmid reversed the inhibitory effect induced by tanshinone IIA, accompanied by an increase in reactive oxygen species levels and lactate dehydrogenase release, indicating a critical role of miR-125b/foxp3 signaling in pyroptosis in HK1 cells. In conclusion, the present study demonstrates that tanshinone IIA enhances pyroptosis and inhibits the proliferation of HK1 cells by modulating miR-125b/foxp3/caspase-1/GSDMD signaling. It is the first study to reveal the inhibitory effect of tanshinone IIA on HK1 cells and to demonstrate the critical role of miR-125b/foxp3 signaling in mediating these effects, providing robust evidence for the treatment of NPC.

Inhibitory effects of tanshinone IIA from Salvia miltiorrhiza Bge on human bladder cancer BIU-87 cells and xenograft in nude mice

Abstract The objective of this study was to investigate the inhibitory effects of tanshinone IIA on human bladder cancer BIU-87 cells and the xenograft in nude mice. BIU-87 cells were treated with tanshinone IIA with concentration of 0.5, 1, 2 and 4 g/L. The cell proliferation, cycle and apoptosis were detected. The nude mice with BIU-87 cell xenograft were treated with normal saline (control), 5-fluorouracil (5-FU), 200, 400 and 600 mg/kg tanshinone IIA, respectively. The animal body weight and growth of tumor were measured. The expressions of B-cell lymphoma-2 (Bcl-2), Caspase-3 and proliferating cell nuclear antigen (PCNA) protein in xenograft were detected. The apoptosis index of cells was determined. Results showed that, tanshinone IIA inhibited the proliferation of BIU-87 cells, promoted their apoptosis, and arrested more cells in G0/G1 phase. Tanshinone IIA significantly inhibited the growth of xenograft in nude mice and promoted the apoptosis of tumor cells in xenograft. Tanshinone IIA down-regulated Bcl-2 and PCNA expression and up-regulated Caspase-3 expression in xenograft. In conclusion, tanshinone IIA has inhibitory effects on human bladder cancer BIU-87 cells, and can inhibit the BIU-87 cells xenograft in nude mice.

The Synergistic Antitumor Effect of Tanshinone IIA Plus Adriamycin on Human Hepatocellular Carcinoma Xenograft in BALB/C Nude Mice and Their Influences on Cytochrome P450 CYP3A4 In Vivo.

Objective Hepatocellular carcinoma is one of the most common diseases that seriously threaten human life and health. In this study, we evaluated the inhibitory effect of tanshinone IIA (Tan IIA) combined with adriamycin (ADM) on human hepatocellular carcinoma and developed a platform to assess the function if Chinese herbal ingredients combined with chemotherapy drugs have synergistic antitumor effects in vivo. Methods Established animal model of human hepatocarcinoma HepG2 cell in nude mice. Mice were divided into model control group, Tan IIA group, ADM group, and Tan IIA + ADM group. The changes from general condition, weight, tumor volume, and inhibition rate were observed. The data were gathered from serum AST level and histopathological changes. The content and activity of cytochrome P450 were determined by spectrophotometric analysis. CYP3A4 protein expression was analyzed by western blotting. The binding model crystal structure of Tan IIA and ADM with pregnane X receptor (PXR) was evaluated by Discovery Studio 2.1. Results A combination of Tan IIA with ADM could improve life quality by relieving ADM toxicity, decreasing tumor volume, declining serum AST level, and improving liner pathological section in tumor-bearing mice. The inhibitory rates of Tan IIA, ADM, and cotreatment were 32.77%, 60.96%, and 73.18%, respectively. The Tan IIA group significantly enhanced the content of cytochrome b5, P450, and erythromycin-N-demethylase activity. CYP3A4 protein expression was enhanced obviously by the Tan IIA + ADM group. Virtual molecular docking showed that both Tan IIA and ADM could be stably docked with the same binding site of PXR but different interactions. Conclusions Tan IIA in combination with ADM could improve the life quality in tumor-bearing mice and enhance the antitumor effect. The Tan IIA group increased the concentration of cytochrome P450 enzymes and activity. Combined Tan IIA with ADM could upregulate the CYP3A4 protein expression and make relevant interaction with protein PXR by virtual docking.

Inhibitory effect of Tanshinone IIA on the proliferation of human retinal pigment epithelial cells in hypoxia and its mechanism

Objective To investigate the effect of Tanshinone ⅡA on the proliferation and the signaling pathway of human retinal pigment epithelial (RPE) cells in hypoxia. Methods CoCl2 (150 μmol/L) was used to simulate hypoxic condition and the ARPE-19 cells cultured in vitro were divided into blank control group, hypoxia control group, Tanshinone ⅡA group and hypoxia-inducible factor-1α (HIF-1α) inhibitor group.The different doses of Tanshinone ⅡA were used to treat ARPE-19 for 24, 48 and 72 hours, respectively.The inhibitory rate of cell proliferation of different groups were detected by MTT after 24, 48 and 72 hours of administration cultured, and the apoptosis rate and the cell cycle distribution of cells in the hypoxia were analyzed by flow cytometry.Real-time PCR and Western blot were used to detect the expressions of mRNA and protein of HIF-1α and vascular endothelial growth factor (VEGF). Results MTT assay showed that Tanshinone ⅡA could inhibit the proliferation of ARPE-19 cells in a dose- and time-dependent manner, and the proliferation inhibitory rate gradually increased in the 1, 5 and 10 mg/L Tanshinone ⅡA groups, with significant differences between any two groups (all at P 0.05). Conclusions Tanshinone ⅡA can inhibit the proliferation of RPE cells, induce apoptosis by arresting cells at G0/G1 phase.The mechanism is related to the suppression of HIF-1α/VEGF signaling pathway. Key words: Tanshinone ⅡA; Retinal pigment epithelial cells; Hypoxia-inducible factor-1α; Vascular endothelial growth factor; Proliferation

Inhibitory effect of tanshinone IIA on inflammatory response in rheumatoid arthritis through regulating β-arrestin 2.

The current study aimed to investigate the inhibitory effect of tanshinone IIA (Tan IIA) on the inflammatory response in patients with rheumatoid arthritis (RA) and explore its mechanism. A total of 50 patients with RA were randomly separated into the control group (15 cases) and the research group (35 cases). The tumor necrosis factor (TNF)-α and interleukin (IL)-6 levels in serum were determined, and peripheral blood mononuclear cells (PBMCs) were separated from patients with RA and cultured in vitro. The effects of the β-arrestin 2 small interfering (si)RNA incubation, lipopolysaccharide (LPS) stimulation or Tan IIA incubation on TNF-α and IL-6 levels, and the expression levels of β-arrestin 2, NAD-dependent protein deacetylase sirtuin-1 (SIRT1) and transcription factor p65 (p65) proteins were investigated. Prior to treatment, no significant differences in TNF-α and IL-6 levels in serum of patients with RA were identified between the research and control groups. Following treatment, the TNF-α and IL-6 levels in the serum of patients with RA in the research group were significantly lower compared with those in the research group prior to treatment and those in the control group following treatment (P<0.05). Tan IIA inhibited the LPS-induced secretion of TNF-α and IL-6, upregulated the LPS-inhibited expression of the β-arrestin 2 and SIRT1 proteins, and downregulated the LPS-induced expression of the p65 protein in the PBMCs of patients with RA. The β-arrestin 2 small interfering (si)RNA significantly upregulated the secretion of TNF-α and IL-6, inhibited the expression of the SIRT1 protein and upregulated the expression of the p65 protein in PBMCs of patients with RA. Tan II A effectively increased the weight of rats with rheumatoid arthritis, and reduced the circumference of the left posterior ankle, the posterior plantar metatarsal thickness, and the content of serum TNF-α and IL-6. Tan IIA did not significantly reverse these β-arrestin 2 siRNA-induced changes. Tan IIA inhibited the inflammatory response in PBMCs of patients with RA by upregulating β-arrestin 2 expression.

Inhibitory effect of Tanshinone IIA on inverted formin-2 protects HaCaT cells against oxidative injury via regulating mitochondrial stress

Context: Epidermal cells play an important role in regulating the regeneration of skin after burns and wounds.Objective: The aim of our study is to explore the role of Tanshinone IIA (Tan IIA) in the apoptosis of epidermal HaCaT cells induced by H2O2, with a focus on mitochondrial homeostasis and inverted formin-2 (INF2).Materials and methods: Cellular viability was determined using the MTT assay, TUNEL staining, western blot analysis and LDH release assay. Adenovirus-loaded INF2 was transfected into HaCaT cells to overexpress INF2 in the presence of Tan IIA treatment. Mitochondrial function was determined using JC-1 staining, mitochondrial ROS staining, immunofluorescence and western blotting.Results: Oxidative stress promoted the death of HaCaT cells and this effect could be reversed by Tan IIA. At the molecular levels, Tan IIA treatment sustained mitochondrial energy metabolism, repressed mitochondrial ROS generation, stabilized mitochondrial potential, and blocked the mitochondrial apoptotic pathway. Furthermore, we demonstrated that Tan IIA modulated mitochondrial homeostasis via affecting INF2-related mitochondrial stress. Overexpression of INF2 could abolish the protective effects of Tan IIA on HaCaT cells viability and mitochondrial function. Besides, we also reported that Tan IIA regulated INF2 expression via the ERK pathway; inhibition of this pathway abrogated the beneficial effects of Tan IIA on HaCaT cells survival and mitochondrial homeostasis.Conclusions: Overall, our results indicated that oxidative stress-mediated HaCaT cells apoptosis could be reversed by Tan IIA treatment via reducing INF2-related mitochondrial stress in a manner dependent on the ERK signaling pathway.

Tanshinone IIA Inhibits High Glucose-Induced Collagen Synthesis via Nuclear Factor Erythroid 2-Related Factor 2 in Cardiac Fibroblasts

Background/Aims: Diabetes is associated with increased incidence of myocardial dysfunction, which is partly characterized by interstitial and perivascular fibrosis. Cardiac fibroblasts have been identified as an important participant in the development of cardiac fibrosis. Exposure of cultured cardiac fibroblasts to high glucose resulted in increased collagen synthesis. Tanshinone IIA can alleviate the ventricular fibrosis that develops in a number of different experimental conditions. However, whether tanshinone IIA can prevent high glucose-induced collagen synthesis in cardiac fibroblasts remains unknown. The aim of this study was to evaluate the effects of tanshinone IIA on high glucose-induced collagen synthesis in cardiac fibroblasts. Methods: Rat cardiac fibroblasts were cultured in high glucose (25 mM) media in the absence or presence of tanshinone IIA and the changes in collagen synthesis, transforming growth factor-β1 (TGF-β1) production and related signaling molecules were assessed by ³H-proline incorporation, quantitative polymerase chain reaction, enzyme linked immunosorbent assay, and Western blotting. Results: The results indicate cardiac fibroblasts exposed to high glucose condition show increased cell proliferation and collagen synthesis and these effects were abolished by tanshinone IIA treatment. Furthermore, the inhibitory effect of tanshinone IIA on high glucose induced cell proliferation and collagen synthesis may be associated with its activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) and the inhibition of TGF-β1 production and Smad2/3 phosphorylation. Conclusion: In summary, our results highlights the critical role tanshinone IIA plays as an antioxidant in attenuating high glucose-mediated collagen synthesis through inhibiting TGF-β1/Smad signaling in cardiac fibroblasts which provide a mechanistic basis for the clinical application of tanshinone IIA in the treating diabetic-related cardiac fibrosis.

Tanshinone IIA suppresses the progression of atherosclerosis by inhibiting the apoptosis of vascular smooth muscle cells and the proliferation and migration of macrophages induced by ox-LDL.

ABSTRACTThe profound inhibitory effect of Tanshinone IIA (Tan IIA) on atherosclerosis has been demonstrated, whereas the latent mechanism is not completely cleared. This study aimed to investigate the cellular and molecular mechanisms underlying Tan IIA protecting against atherosclerosis. Oil Red O staining and ELISA assay showed that Tan IIA suppressed the progress of atherosclerosis and reduced the levels of inflammatory cytokines in serum of apolipoprotein E deficient (ApoE–/–) mice. Flow cytometry assay revealed that Tan IIA inhibited oxidized LDL (ox-LDL)-induced apoptosis of VSMCs. MTT and transwell assay indicated that Tan IIA suppressed the ox-LDL-stimulated proliferation and migration of RAW264.7 cells. Moreover, Tan IIA ameliorated inflammatory cytokine upregulation elicited by ox-LDL in RAW264.7 cells. Additionally, Tan IIA inhibited the apoptosis of VSMCs and decreased the levels of MMP-2, MMP-9 in ApoE–/– mice. In conclusion, our study demonstrated Tan IIA suppressed the progression of atherosclerosis by inhibiting vascular inflammation, apoptosis of VSMCs and proliferation and migration of macrophages induced by ox-LDL.

Inhibitory effect of tanshinone IIA on rat hepatic stellate cells.

BackgroundAnti-inflammation via inhibition of NF-κB pathways in hepatic stellate cells (HSCs) is one therapeutic approach to hepatic fibrosis. Tanshinone IIA (C19H18O3, Tan IIA) is a lipophilic diterpene isolated from Salvia miltiorrhiza Bunge, with reported anti-inflammatory activity. We tested whether Tan IIA could inhibit HSC activation.Materials and MethodsThe cell line of rat hepatic stellate cells (HSC-T6) was stimulated with lipopolysaccharide (LPS) (100 ng/ml). Cytotoxicity was assessed by MTT assay. HSC-T6 cells were pretreated with Tan IIA (1, 3 and 10 µM), then induced by LPS (100 ng/ml). NF-κB activity was evaluated by the luciferase reporter gene assay. Western blotting analysis was performed to measure NF-κB-p65, and phosphorylations of MAPKs (ERK, JNK, p38). Cell chemotaxis was assessed by both wound-healing assay and trans-well invasion assay. Quantitative real-time PCR was used to detect gene expression in HSC-T6 cells.ResultsAll concentrations of drugs showed no cytotoxicity against HSC-T6 cells. LPS stimulated NF-κB luciferase activities, nuclear translocation of NF-κB-p65, and phosphorylations of ERK, JNK and p38, all of which were suppressed by Tan IIA. In addition, Tan IIA significantly inhibited LPS-induced HSCs chemotaxis, in both wound-healing and trans-well invasion assays. Moreover, Tan IIA attenuated LPS-induced mRNA expressions of CCL2, CCL3, CCL5, IL-1β, TNF-α, IL-6, ICAM-1, iNOS, and α-SMA in HSC-T6 cells.ConclusionOur results demonstrated that Tan IIA decreased LPS-induced HSC activation.

Tanshinone IIA inhibits the growth, attenuates the stemness and induces the apoptosis of human glioma stem cells

Glioma stem cells (GSCs) are believed to contribute to glioblastoma multiforme(GBM) propagation and treatment resistance. TanshinoneIIA possesses anticancer and anti-inflammatory activities. This study aimed to determine the inhibitory effect of tanshinoneIIA on human GSCs invitro and invivo and to explore the underlying mechanisms. In the present study, human GBM neurospheres (GBMS) were isolated from adherent GBM cells in serum-free medium, and the cells from the GBMS displayed characteristics of GSCs. Results from the MTT, neurosphere formation and invivo inhibition assays revealed that tanshinoneIIA had a significant inhibitory effect on human GSCs invitro and invivo. Furthermore, tanshinoneIIA increased the expression of differentiation and neural lineage markers including GFAP and β-tubulin, decreased expression of GSC markers including CD133 and nestin, and induced GSC apoptosis invitro and invivo in a dose‑dependent manner. Inflammatory cytokines and signaling pathways are believed to play key roles in maintaining the stem-like properties in human glioma cells. In the present study, inflammatory cytokine interleukin6 (IL6) and its downstream activated signal transducer and activator of transcription 3 [phospho-STAT3(tyrosine705) and phospho-STAT3(serine727)] were downregulated after tanshinoneIIA treatment invitro and invivo. This result indicated that disturbance of the IL6/STAT3 signaling axis by tanshinoneIIA is closely related to the growth inhibition of GSCs. Taken together, our results indicate that tanshinoneIIA has the potential to target and kill GSCs through suppression of proliferation, attenuation of stemness and induction of apoptosis. Its mechanism of activity may be associated with attenuation of the IL6/STAT3 signaling pathway.

Tanshinone IIA inhibits lipopolysaccharide-induced MUC1 overexpression in alveolar epithelial cells

The anti-inflammatory function of tanshinone IIA (TIIA), an active natural compound from Chinese herbal medicine Danshen, has been well recognized, and therefore TIIA has been widely used to treat various inflammatory conditions associated with cardiac and lung diseases. Mucin 1 (Muc1) plays important anti-inflammatory roles in resolution of acute lung inflammation. In this study, we investigated the effects of TIIA on LPS-induced acute lung inflammation, as well as its relationship to Muc1 expression in mouse lung and MUC1 in human alveolar epithelial cells. TIIA pretreatment significantly inhibited LPS-induced pulmonary inflammation in both Muc1 wild-type (Muc1(+/+)) and knockout (Muc1(-/-)) mice, as manifested by reduced neutrophil infiltration and reduced TNF-α and keratinocyte chemoattractant levels in bronchoalveolar lavage fluid. The inhibitory effects of TIIA on airway inflammation were associated with reduced expression of Muc1 in Muc1(+/+) mouse lung. Moreover, pretreatment with TIIA significantly inhibited LPS-induced MUC1 expression and TNF-α release in A549 alveolar epithelial cells. TNF-α upregulated MUC1 mRNA and protein expression in A549 cells, which was inhibited by pretreatment with TIIA. The LPS-induced MUC1 expression was blocked when A549 cells were transfected with siRNA targeting for TNF-α receptor 1. Furthermore, TIIA inhibited LPS-induced nuclear translocation of NF-κB and upregulation of Toll-like receptor 4 in A549 cells. Taken together, these results demonstrate that TIIA suppressed LPS-induced acute lung inflammation regardless of the presence of Muc1, and TIIA inhibited LPS- and TNF-α-induced MUC1/Muc1 expression in airway epithelial cells, suggesting that MUC1/Muc1 does not account for the mechanisms of the anti-inflammatory effects of TIIA in the airway.

The anti-atherosclerotic effect of tanshinone IIA is associated with the inhibition of TNF-α-induced VCAM-1, ICAM-1 and CX3CL1 expression

Tanshinone IIA is one of the major diterpenes in Salvia miltiorrhiza. The inhibitory effect of Tanshinone IIA on atherosclerosis has been reported, but the underlying mechanism is not fully understood. The present study aimed to study the anti-atherosclerosis effect of Tanshinone IIA on the adhesion of monocytes to vascular endothelial cells and related mechanism. Results showed that Tanshinone IIA, at the concentrations without cytotoxic effect, dose-dependently inhibited the adhesion of THP-1 monocytes to the TNF-α-stimulated human vascular endothelial cells. The expressions of cell adhesion molecules including VCAM-1, ICAM-1 and E-selectin were induced by TNF-α in HUVECs at both the mRNA and protein levels. The mRNA and protein expressions of VCAM-1 and ICAM-1, but not E-selectin, were both significantly suppressed by Tanshinone IIA in a dose dependent manner. In addition, the TNF-α-induced mRNA expression of fractalkine/CX3CL1 and the level of soluble fractalkine were both reduced by Tanshinone IIA. We also found that Tanshinone IIA significantly inhibited TNF-α-induced nuclear translocation of NF-κB which was resulted from the inhibitory effect of Tanshinone IIA on the TNF-α-activated phosphorylation of IKKα, IKKβ, IκB and NF-κB. As one of the major components of Salvia miltiorrhiza, Tanshinone IIA alone exerted more potent effect on inhibiting the adhesion of monocytes to vascular endothelial cells when compared with Salvia miltiorrhiza. All together, these results demonstrate a novel underlying mechanism for the anti-inflammatory effect of Tanshinone IIA by modulating TNF-α-induced expression of VCAM-1, ICAM-1 and fractalkine through inhibition of TNF-α-induced activation of IKK/NF-κB signaling pathway in human vascular endothelial cells.

Tanshinone IIA, a constituent of Danshen, inhibits the release of glutamate in rat cerebrocortical nerve terminals

Ethnopharmacological relevanceDanshen is a commonly used traditional Chinese medicine and has received considerable attention due to their beneficial effects on the health, including prevention of cardiovascular disease, and cancer. Tanshinone IIA, a major active constituent of Danshen, has been reported to have a neuroprotective profile. Aim of the studyAn excessive release of glutamate is considered to be related to neuropathology of several neurological diseases. In this study, we investigated whether tanshinone IIA could affect endogenous glutamate release and explored the possible mechanism. Materials and methodsThe experimental model was the isolated nerve terminals (synaptosomes) purified from the rat cerebral cortex. The release of glutamate was evoked by the K+ channel blocker 4-aminopyridine (4-AP) and measured by one-line enzyme-coupled fluorometric assay. We also used a membrane potential-sensitive dye to assay nerve terminal excitability and depolarization, and a Ca2+ indicator, Fura-2-acetoxymethyl ester, to monitor cytosolic Ca2+ concentrations ([Ca2+]C). ResultsTanshinone IIA inhibited the release of glutamate evoked by 4-AP in a concentration-dependent manner. Inhibition of glutamate release by tanshinone IIA was prevented by the chelating the extracellular Ca2+ ions, and by the vesicular transporter inhibitor bafilomycin A1. However, the glutamate transporter inhibitor DL-threo-beta-benzyl-oxyaspartate did not have any effect on the action of tanshinone IIA. Tanshinone IIA decreased the depolarization-induced increase in [Ca2+]C, whereas it did not alter the resting synaptosomal membrane potential or 4-AP-mediated depolarization. Furthermore, the effect of tanshinone IIA on evoked glutamate release was prevented by the Cav2.2 (N-type) and Cav2.1 (P/Q-type) channel blocker ω-conotoxin MVIIC, but not by the ryanodine receptor blocker dantrolene or the mitochondrial Na+/Ca2+ exchanger blocker CGP37157. Mitogen-activated protein kinase (MEK) inhibition also prevented the inhibitory effect of tanshinone IIA on evoked glutamate release. ConclusionThese results show that tanshinone IIA inhibits glutamate release from cortical synaptosomes in rats through the suppression of presynaptic voltage-dependent Ca2+ entry and MEK signaling cascade.