Tanshinone IIA Group Research Articles

High-Dose Tanshinone IIA Suppresses Migration and Proliferation While Promoting Apoptosis of Astrocytoma Cells Via Notch-1 Pathway.

Malignant astrocytoma is the most common malignant tumor with strong invasion in the central nervous system. Tanshinone IIA is an effective compound to suppress cell proliferation and promote cell apoptosis. However, there is little research about the role of tanshinone IIA in the treatment of astrocytoma. This study aimed to investigate the effect of tanshinone IIA on migration, proliferation and apoptosis of astrocytoma cells. The efficacy of tanshinone IIA on migration, proliferation and apoptosis of astrocytoma cells were evaluated by flow cytometry and the assays of plate clone formation, CCK-8, wound healing and transwell migration. The protein molecule and signaling pathway were detected by western blot. High-dose tanshinone IIA suppressed migration and proliferation of astrocytoma cells while promoting apoptosis of astrocytoma cells. The western blot results showed that there were high Notch-1 protein expression and low c-Myc, MMP-9 and Bcl-2 activation in the high-dose tanshinone IIA group compared with the control group. High-dose tanshinone IIA suppresses astrocytoma cell proliferation, migration while promoting apoptosis through Notch-1 pathway. Tanshinone IIA may be used to develop new drugs for the treatment of astrocytoma.

Tanshinone IIA attenuates paraquat‑induced acute lung injury by modulating angiotensin‑converting enzyme 2/angiotensin‑(1‑7) in rats.

Tanshinone IIA (TIIA) is an active compound that can be isolated from the Chinese herb, Salviamiltiorrhizae Bunge, also known as danshen. Previous studies have demonstrated that TIIA can effectively attenuate bleomycin‑induced pulmonary fibrosis in rats. However, it has not been determined whether TIIA can attenuate paraquat (PQ)‑induced acute lung injury (ALI). In the present study, the protective effects exhibited by TIIA on PQ‑induced ALI, as well as its underlying mechanisms, were investigated using Sprague‑Dawley (SD) rats. ALI animal models using rats were established via administration of PQ. Adult male SD rats were randomly divided into three groups: A control group, a PQ group and a PQ+TIIA group. Total cell count, total protein levels and lactic dehydrogenase (LDH) levels in bronchoalveolar lavage fluid (BALF), as well as myeloperoxidase (MPO) activity in lung tissues were determined. Lung histological alterations were also investigated. Angiotensin converting enzyme 2 (ACE2) and Angiotensin1‑7 [Ang‑(1‑7)] expression levels in the lung were also analyzed. The results demonstrated that administration of PQ induced marked histological alterations, and markedly increased neutrophil infiltration, lung wet/dry weight ratio, total cell count, protein content and LDH levels in BALF. In addition, PQ was revealed to significantly decrease ACE2 and Ang‑(1‑7) expression levels in lung tissues. However, it was demonstrated that TIIA attenuated these effects. Therefore, the results of the present study suggest that that TIIA may exhibit a therapeutic effect regarding PQ‑induced ALI in rats, and that ACE2 and Ang‑(1‑7) may be involved in the underlying mechanisms of this effect.

Tanshinone IIA mitigates peritoneal fibrosis by inhibiting EMT via regulation of TGF-β/smad pathway

Purpose : To explore the effects of tanshinone IIA (T-IIA) on Dianeal-N PD-4 (PDF)-induced expression of fibrogenic cytokines in human peritoneal mesothelial cells (HPMCs), and to elucidate the mechanisms of action involved. Methods : Seven groups of HPMCs were used in the study: control group, PDF group, T-IIA group, LY364947 group, and 2 transforming growth factor-β (TGF-β) groups (TGF-β+ 50 μM T-IIA and TGF-β+ 100 μM T- IIA). The expression levels of mRNA and protein of TGF-β, smad2, smad7, α-smooth muscle actin(α-SMA), fibronectin, collagen І, E-cadherin, N-cadherin, matrix metalloprotein-2(MMP-2), and MMP-9 in the various groups were determined by reverse transcription-polymerase chain reaction (RTPCR) and Western blotting as appropriate. Results : The expressions of α-SMA, fibronectin, collagen І, TGF-β and smad2 were significantly upregulated in HPMCs by PDF treatment, but smad7 was down-regulated, relative to the control group (p < 0.01).These PDF-induced effects were reversed by T-IIA (p < 0.05). Inhibition of TGF-β/smad pathway by LY364947 treatment led to significant decrease in the expressions of fibrosis-related proteins, when compared with PDF group (p < 0.05). TGF-β treatment also produced numerous spindleshaped HPMCs characteristic of epithelial-mesenchymal transition (EMT). However, this morphological transition was alleviated, and the expression levels of EMT-related proteins were significantly downregulated by exposure to the two doses of T-IIA (p < 0.05). Conclusion : Tanshinone IIA inhibits EMT in HPMCs by regulating TGF-β/smad pathway, thus mitigating peritoneal fibrosis. Therefore, T-IIA has promising potential as a new drug for the treatment of peritoneal dialysis (PD)-induced fibrosis. Keywords : Peritoneal dialysis, Peritoneal fibrosis, Tanshinone IIA, Epithelial-mesenchymal transition

Tanshinone IIA Attenuates Contrast-Induced Nephropathy via Nrf2 Activation in Rats

Background/Aims: Tanshinone IIA is a chemical compound extracted from Salvia miltiorrhiza Bunge, a perennial plant also known as red sage used in traditional Chinese medicine. Tanshinone IIA has been shown to protect against various organ injuries. In this study, we hypothesized that Tanshinone IIA could play an anti-oxidative role in contrast-induced nephropathy (CIN) through enhancing Nrf2/ARE activation. Methods: To test whether Tanshinone IIA can attenuate CIN, oxidative stress, and apoptosis, we utilized two models: an in vivo Sprague-Dawley rat model of ioversol-induced CIN and an in vitro cell model of oxidative stress in which HK2 cells, a human renal tubular cell line, are treated with hydrogen peroxide (H₂O₂). Rats were randomly assigned to 4 groups (n = 6 per group): control group, ioversol group (ioversol-induced CIN), vehicle group (ioversol-induced CIN rats pretreated with vehicle), and Tanshinone IIA group (ioversol-induced CIN rats pretreated with 25mg/kg Tanshinone IIA). Renal functions, renal injuries and apoptosis were evaluated by using serum creatinine, histological scoring, and TUNEL staning respectively. Malondialdehyde, 8-hydroxy-2’ –deoxyguanosine, and intracellular reactive oxygen species were used for oxidative stress assessment. Levels of Nrf2 and heme oxygenase-1 (HO-1) were measured in vivo and in vitro. Results: Tanshinone IIA attenuated renal tubular necrosis, apoptosis and oxidative stress in rats and oxidative stress in HK2 cells. Furthermore, Tanshinone IIA activated Nrf2, and up-regulated HO-1 expression in vivo and in vitro, resulting in a reduction in oxidative stress. Conclusion: Tanshinone IIA may protect against CIN through enhancing Nrf2/ARE activation.

Effect of tanshinone IIA on oxidative stress and apoptosis in a rat model of fatty liver.

Oxidative stress is a crucial factor associated with fatty liver disease, which raises the possibility of using antioxidants to improve liver steatosis. Tanshinone IIA (TSIIA) is a traditional Chinese medicine that has been reported to have antioxidant effects in vitro. The present study aimed to investigate whether TSIIA possesses antioxidant effects in vivo and whether TSIIA was able to improve liver steatosis. Hence, the ability of TSIIA to protect rats from liver disease was explored, particularly in regard to antioxidant activity. Rats were fed a high-lipid diet for 90 days, causing severe liver steatosis, both morphologically and biochemically. An increase in reactive oxygen species (ROS) in the liver was exhibited in addition to significantly elevated serum lipids and malondialdehyde (MDA). Furthermore, hepatocyte apoptosis was measured by Hoechst staining, reverse transcription-quantitative polymerase chain reaction and western blot analysis and an increase in hepatocyte apoptosis rate was indicated in mice on a high-fat diet. Following intraperitoneal injection of TSIIA (10 mg/kg/day), liver steatosis was significantly inhibited. In rats receiving TSIIA treatment, less ROS were indicated in the liver and significantly decreased levels of MDA (P<0.05) in serum were exhibited, whereas significantly increased activities of total superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-PX) were observed (P<0.05 and P<0.01, respectively). In addition, the rate of hepatocyte apoptosis was significantly decreased in the TSIIA group (P<0.01). However, TSIIA elicited no effect on serum lipid profiles. These results suggest that TSIIA attenuates oxidative stress by decreasing ROS and MDA production and enhancing the activity of T-SOD and GSH-PX, which may contribute to the inhibition of apoptosis and amelioration of liver steatosis.

Neuroprotective effect of combining tanshinone IIA with low-dose methylprednisolone following acute spinal cord injury in rats.

The present study compared the potential neuroprotective effect of tanshinone IIA (TIIA) monotherapy, methylprednisolone (MP) monotherapy and combined treatment in an adult acute spinal cord injury (ASCI) rat model. The current study used the weight-drop method (Allen's Impactor) in the rat model and the mechanical scratch method in primary spinal cord neuron culture to determine whether the combined treatment was able to reduce the required dosage of MP in the treatment of ASCI to produce a similar or improved therapeutic effect. In vivo male Sprague Dawley rats (n=60) were randomly divided into 5 groups, of which 12 rats were selected for the sham group and T9-T11 laminectomies, leading to ASCI, were performed on 48 of the 60 rats using a 10 g ×25 mm weight-drop at the level of T10 spinal cord. Therefore, the ASCI group (n=12) included the ‘laminectomy and weight-drop’. The remaining 36 ASCI model animals were subdivided into 3 groups (n=12 each group): TIIA group (30 mg/kg/day), MP group (30 mg/kg) and combined treatment group (TIIA 30 mg/kg/day + MP 20 mg/kg). Neuronal function following ASCI was evaluated using the Basso Beattie Bresnahan (BBB) locomotor rating scale. Levels of the anti-apoptotic factor B-cell lymphoma-2 (Bcl-2), the pro-apoptotic factors Bcl-2 associated protein X (Bax) and caspase-3, and the inflammatory associated factor nuclear factor-κB, were analyzed by western blot analysis. Immunohistochemistry was used to detect caspase-3. To investigate the underlying mechanism, the anti-oxidative effect of combination TIIA and MP treatment was assessed by measuring the activity of malondialdehyde (MDA) and superoxide dismutase (SOD) in ASCI. In agreement with the experiment in vivo, primary neurons were prepared from the spinal cord of one-day-old Sprague-Dawley rats' and co-cultured with astrocytes from the brain cortex. The injury of neurons was induced by mechanical scratch and levels of apoptosis factors were analyzed by western blot analysis. The results of the current study indicated that injured animals in the combined treatment group exhibited a significant increase in BBB scores (P<0.05). TIIA + MP combined treatment and MP treatment was observed to reduce the expression of pro-apoptotic factors and promote neuron survival in vivo and in vitro. Combined treatment may promote neuroprotection through reduced apoptosis and inflammation caused by ASCI, similar to MP alone. Combined treatment reversed the decrease of SOD and the increase of MDA level caused by ASCI. In addition, combined treatment decreased the expression of caspase-3 in the neurons following ASCI in rats, as indicated by immunofluorescence double labeling. Overall, the present study indicates that the combined treatment of TIIA and MP may protect the neurons by stimulating the rapid initiation of neuroprotection following ASCI and reduce the dosage of MP in the treatment of ASCI required to produce the same or improved neuroprotective effects in vivo and in vitro.

Protective effect of tanshinone IIA against cardiac hypertrophy in spontaneously hypertensive rats through inhibiting the Cys-C/Wnt signaling pathway.

The study aimed to investigate the protective effect of tanshinone IIA against cardiac hypertrophy in spontaneously hypertensive rats (SHRs) through the Cys-C/Wnt signaling pathway. Thirty SHRs were randomly divided into cardiac hypertrophy, low- and high-dose tanshinone IIA groups. Ten Wistar-Kyoto rats were selected as control group. The systolic blood pressure (SBP), heart weight (HW), left ventricular weight (LVW) and body weight (BW) of all rats were recorded. HE staining and qRT-PCR were applied to observe the morphology of myocardial tissue and mRNA expressions of COL1A1 and COL3A1. ELISA and Western blotting were used to measure the serum asymmetric dimethylarginine (ADMA), nitric oxide (NO) and cardiac troponin I (cTnI) levels, and the expressions of the Cys-C/Wnt signaling pathway-related proteins, eNOS and Nox4. Compared with the cardiac hypertrophy group, the SBP, HW/BW, LVW/BW, swelling degree of myocardial cells, COL1A1 and COL3A1 mRNA expressions, serum cTnI and ADMA levels, and the Cys-C/Wnt signaling pathway-related proteins and Nox4 expressions in the low- and high-dose tanshinone IIA groups were decreased, but the endothelial NO synthase (eNOS), phosphorylated eNOS (Ser1177) and NO expressions were increased. No significant difference was found between the low- and high-dose tanshinone IIA groups. Our study indicated a protective effect of tanshinone IIA against cardiac hypertrophy in SHRs through inhibiting the Cys-C/Wnt signaling pathway.

Progressive development of endometriosis and its hindrance by anti-platelet treatment in mice with induced endometriosis

We have recently shown that platelets drive smooth muscle metaplasia (SMM) and fibrogenesis in endometriosis through epithelial-mesenchymal transition (EMT) and fibroblast-to-myofibroblast transdifferentiation (FMT). To see whether this is true in vivo, this prospective, randomized, and serially evaluated mouse investigation was conducted. Endometriosis was induced in female Balb/C mice, which were then randomly divided into two groups: Tanshinone IIA (TAN) and control (CTL) groups. TAN mice were treated with TAN but CTL mice received none. Every week until the 6th week after induction, five mice from each group were killed. Lesion weight was measured and lesion samples were subjected to immunohistochemistry and histochemistry analysis of platelet aggregation (CD41), E-cadherin, TGF-β1, phosphorylated Smad3, α-SMA, collagen I, CCN2, LOX, desmin and SM-MHC, and the extent of fibrosis was evaluated by Masson trichrome staining. It was found that endometriotic lesions exhibited progressive cellular changes consistent with the progressive EMT, FMT, SMM, and fibrogenesis. TAN treatment resulted in significant hindrance of EMT, FMT, SMM and fibrogenesis, and reduced lesion weight (all P-values <0.05). These data corroborate the notion that endometriotic lesions undergo progressive EMT and FMT, giving rise to SMM and ultimately fibrosis. This understanding sheds new light onto the natural history of endometriosis.

Tanshinone IIA Prevent Tendon Adhesion in the Rat Achilles Tendon Model

Background: Tendon adhesion between the sheath and tendon surface is a common clinical problem. Orthopedist makes the improvement of repair techniques and rehabilitation to treat tendon adhesion, but it fails to cure completely. TSA was one of the major active phytochemicals because of its anti-inflammatory activity. We used tanshinone IIA (TSA) for the prevention of tendon adhesion in the rat Achilles tendon model and investigated the possible mechanisms, including microRNAs (miRNAs) and protein expression via TGF-β/Smad signaling pathway. Method: Sprague-Dawley (SD) rat Achilles tendons were half partial lacerated and sutured by a modified Kessler's technique, with TSA and normal saline for control. Macroscopic and histological evaluations were applied to examine the injured tendon six weeks after surgery. We evaluated the degree of adhesion in Gross observation and the remodeling of collagen fibers by observing microscopically and determining the amount of scar formation. The expression of microRNAs (miRNAs) was quantified by real-time PCR detection and protein expression were quantified by western blotting detection. Results: In gross evaluation of tendon adhesion, the TSA group had less adhesion appeared. No evidences of tendon rupture or local infection were observed. The content of collagen fibers in tendon tissue was decreased in TSA group compared with the control group, it indicated a significant difference from the control group, P = 0.0004. The expression of miRNAs including miR-155, miR-29b, miR21, miR-133b and let7 were detected in the repaired tendon tissue, and only miR-29b treated with TSA was observed significantly higher than control group, P &lt;0.0001. The protein expression of TGF-β1 and p-Smad3 treated with TSA was lower than control group. Conclusions: The usage of TSA may be an efficient approach for preventing tendon adhesion.

Effect of tanshinone IIA on microvessel density and the expression of basic fibroblast growth factor, vascular endothelial growth factor of human bladder cancer transplanted tumor in nude mice

Objective To study the effect of tanshinone ⅡA on microvessel density (MVD) and the expression of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) of human bladder cancer transplanted tumor in nude mice. Methods Thirty-six BALB/c nude mice were collected, and divided into model group, thiotepa group, and tanshinone ⅡA group after bladder cancer xenograft models were established. Four weeks later, the tumor growth, angiogenesis, and expression of apoptosis proteins were observed. Results At 4th week, bladder tumor weight, and wet weight index were significantly higher in tanshinone ⅡA group than in model group and thiotepa group, and the inhibitory rate in tanshinone ⅡA group was significantly higher than that of thiotepa group [(0.62±0.07) g vs. (0.91±0.09) g, (0.68±0.00) mg/g vs. (0.90±0.08) mg/g, and 53.38 vs. 31.58, t=4.243-8.981, P<0.05]. MVD, bFGF and VEGF mRNA expression levels in tanshinone ⅡA group were significantly lower than those in model group, and thiotepa group [(8.45±0.75 vs. 13.06±1.65), (35.52±4.29 vs. 65.14±7.21), and (41.39±4.77 vs. 68.32±7.24)] (t=7.601-15.732, P<0.05). The expression levels of p53, and B cell lymphoma/leukemia-2 associated X protein (bax) in tanshinone ⅡA group were significantly higher than those in model group and thiotepa group (1.25±0.26 vs. 0.64±0.07, 0.86±0.12 vs. 0.67±0.08), and those of B cell lymphoma/leukemia-2 (bcl-2), and Survivin were significantly lower than those in model group and thiotepa group (0.41±0.04 vs. 0.70±0.09, and 0.38±0.04 vs. 0.85±0.09, t=2.821-16.149, P<0.05). Conclusion Tanshinone II A can inhibit the growth of transplanted tumor of human bladder cancer in nude mice, reduce the density of blood vessels and the production of pro-angiogenesis molecules, and induce cell apoptosis. Key words: Bladder cancer; Tanshinone Ⅱ A; Transplanted tumor; Angiogenesis; Apoptosis proteins

Effects of albendazole combined with TSII-A (a Chinese herb compound) on optic neuritis caused by Angiostrongylus cantonensis in BALB/c mice.

BackgroundAngiostrongylus cantonensis (A. cantonensis) infection can lead to optic neuritis, retinal inflammation, damage to ganglion cells, demyelination of optic nerve and visual impairment. Combined therapy of albendazole and dexamethasone is a common treatment for the disease in the clinic, but it plays no role in vision recovery. Therefore, it has been necessary to explore alternative therapies to treat this disease. Previous studies reported the neuro-productive effects of two constituents of Danshen (a Chinese herb)-tanshinone II-A (TSII-A) and cryptotanshinone (CPT), and this study aims to evaluate the impacts of TSII-A or CPT combined with albendazole on optic neuritis caused by A. cantonensis infection in a murine model.MethodsTo assess the effects of TSII-A or CPT combined with albendazole on optic neuritis due to the infection, mice were divided into six groups, including the normal control group, infection group and four treatment groups (albendazole group, albendazole combined with dexamethasone group, albendazole combined with CPT group and albendazole combined with TSII-A group). The infection group and treatment groups were infected with A. cantonensisand the treatment groups received interventions from 14 dpi (days post infection), respectively. At 21 dpi, the visual acuity of mice in each group was examined by visual evoked potential (VEP). The pathologic alteration of the retina and optic nerve were observed by hematoxylin and eosin (H&E) staining and transmission electronic microscopy (TEM).ResultsInfection of A. cantonensis caused prolonged VEP latency, obvious inflammatory cell infiltration in the retina, damaged retinal ganglions and retinal swelling, followed by optic nerve fibre demyelination and a decreasing number of axons at 21 dpi. In treatment groups, albendazole could not alleviate the above symptoms; albendazole combined with dexamethasone lessened the inflammation of the retina, but was futile for the other changes; however, albendazole combined with CPT and albendazole combined with TSII-A showed obvious effects on the recovery of prolonged VEP latency, destruction and reduction of ganglion cells, optic nerve demyelination and axon loss. Compared with albendazole-CPT compound, albendazole combined with TSII-A was more effective.ConclusionsThe current study demonstrates that albendazole combined with TSII-A plays a more effective role in treating optic neuritis caused by A. cantonensis in mice than with dexamethasone, as applied in conventional treatment, indicating that albendazole combined with TSII-A might be an alternate therapy for this parasitic disease in the clinic.

Neuroprotective Capabilities of Tanshinone IIA against Cerebral Ischemia/Reperfusion Injury via Anti-apoptotic Pathway in Rats

Danshen, derived from the dried root or rhizome of Salviae miltiorrhizae BGE., has Tanshinone IIA (TSA) as one of its active ingredients. Recent reports have shown that TSA can inhibit the apoptosis induced by serum withdrawal or ethanol in cultured PC12 cells. However, whether TSA has any neuroprotective effect remains unknown. In this study, we investigated the effects of TSA on cerebral apoptosis induced by middle cerebral artery occlusion (MCAO) in which cerebral ischemia had been induced 2 h earlier. Twenty-four hours after reperfusion, the rats were assessed for infarct volume etc. Intraperitoneal administration of 25 and 40 mg/kg TSA 10 min after MCAO significantly diminished infarct volume and brain water content and improved neurological deficits in a dose-dependent manner. The 25 mg/kg dosage was more effective. Treatment with 25 mg/kg TSA significantly improved symptoms and reduce infarct volume at different points in time, of which 10 min after MCAO was the most significant. Nissl-staining and HE-staining of the 25 mg/kg TSA group were more appreciable in terms of improvement relative to the vehicle group in the infarct core. TSA of dosage 25 mg/kg significantly decreased the expression of cleaved caspase-3 protein and increased the expression of B-cell lymphoma 2 (bcl-2) protein in the ischemic cortex. Fewer terminal deoxyribonucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick-end labeling (TUNEL)-positive cells were found in the penumbra of the treated group, but they were significantly more common in the vehicle group. We here conclude that the neuroprotective effects of TSA against focal cerebral ischemic/reperfusion injury are likely to be related to the attenuation of apoptosis.

Effect of Tanshinone IIA on phosphorylated NMDA receptor 1 expression and intracellular free calcium concentration in neonatal SD rats with hypoxic ischemic brain damage

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. 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. (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). 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.

Effect of tanshinone IIA on the change of calcium current induced by beta-amyloid protein 25-35 in neurons of nucleus basalis of Meynert

To explore the effect of tanshinone IIA (TanIIA) on calcium current induced by beta-amyloid protein 25-35 (Abeta25-35) in neurons of nucleus basalis of Meynert (nbM). Cell acute dissociated technique and the whole-cell recording model of patch-clamp technique of single-cell were used. The voltage-dependent calcium current in neurons of nbM was recorded in SD rats first. Then the effect of TanIIA on the voltage-dependent calcium current in the neurons was assayed. The change of calcium current induced by Abeta25-35 as well as the effect of TanIIA on the change of calcium current induced by Abeta25-35 in neurons of nbM were analyzed. Extracellular fluid containing different concentrations of TanIIA was irrigated, respectively. The peak current did not change obviously. There was no difference in current density between the TanIIA group and the control group at 0 mV (P>0.05). Extracellular fluid containing 200 nmol/L Abeta25-35 was irrigated after the normal calcium current recorded under whole patch clamp, and the peak current changed obviously. There was distinct difference in the current density between the Abeta group and the control group at 0 mV (P<0.05). Extracellular fluid containing Abeta25-35 and different concentrations of TanIIA were irrigated after the normal calcium current was recorded under whole patch clamp, respectively, and the peak current did not change. There was no difference in current density between the TanIIA +Abeta group and the control group at 0 mV (P>0.05). In vitro, TanIIA could inhibit the calcium current amplification induced by Abeta25-35 in neurons of nbM. TanIIA may protect neurons against the toxicity of Abeta and decrease the inward flow of Ca(2+).

Growth inhibition and apoptosis induction of tanshinone II-A on human hepatocellular carcinoma cells.

To evaluate the effects of tanshinone II-A on inducing growth inhibition and apoptosis of human hepatocellular carcinoma (HCC) cells. The human hepatocellular carcinoma cell line SMMC-7721 was used for the study. The cells were treated with tanshinone II-A at different doses and different times. Cell growth and proliferation were measured by MTT assay, cell count and colony-forming assay. Apoptosis induction was detected by microscopy, DNA ladder electrophoresis and flow cytometry. In MTT assay, the inhibitory effect became gradually stronger with the passage of time, 24, 48, 72 and 96 h after treatment with tanshinone II-A, and the most significant effect was observed at 72 h. On the other hand, the increase of doses (0.125, 0.25, 0.5, 1.0 mg/L tanshinone II-A) resulted in enhanced inhibitory effect. The growth and proliferation of SMMC-7721 cells were obviously suppressed in a dose- and time-dependent manner. The results of cell count were similar to that of MTT assay. In colony-forming assay, the colony-forming rates were obviously inhibited by tanshinone II-A. In tanshinone II-A group, the morphology of cellular growth inhibition and characteristics of apoptosis such as chromatin condensation, crescent formation, margination and apoptotic body were observed under light and transmission electron microscopes. DNA ladder of cells was presented in electrophoresis. The apoptosis index (AI) was 16.9% (the control group was 4.6%) in flow cytometry. The cells were arrested in G(0)/G(1) phase, and the expressions of apoptosis-related genes bcl-2 and c-myc were down-regulated and fas, bax, p53 up-regulated. Tanshinone II-A could inhibit the growth and proliferation of HCC cell effectively in vitro by apoptosis induction, which was associated with up-regulation of fas, p53, bax, expression and down-regulation of bcl-2 and c-myc.