Fsk and IBMX inhibit proliferation and proapoptotic of glioma stem cells via activation of cAMP signaling pathway.

OBJECTIVE: We aimed to find out the underlying mechanism of forskolin (Fsk) and 3-isobutyl-1-methylxanthine (IBMX) on glioma stem cells (GSCs). METHODS: The expression of cAMP-related protein CREB and pCREB as well as apoptosis-related proteins were detected through Western blot analysis. The level of proliferation and growth rate of human GSCs was measured through thiazolyl blue tetrazolium bromide assay and stem cells forming sphere assay. The apoptosis-related gene expression was measured through reverse transcription-polymerase chain reaction. RESULTS: cAMP signaling pathway was activated in GSCs with Fsk-IBMX administration. Fsk-IBMX could inhibit the proliferation as well as invasion and promote the apoptosis of U87 cells. Besides, U0126 could inhibit MAPK signaling pathway to increase the sensitivity of GSCs to cAMP signaling pathway. As a result, Fsk-IBMX combined with U0126 had more negative effect on GSCs. CONCLUSIONS: The relationship of cAMP and MAPK signaling pathway in GSCs may provide a potential therapeutic strategy in glioma.

Simultaneous determination of ferulic acid and gastrodin of Tianshu Capsule in rat plasma by ultra-fast liquid chromatography with tandem mass spectrometry and its application to a comparative pharmacokinetic study in normal and migraine rats.

Tianshu Capsule, consisting of Ligusticum chuanxiong Hort and Gastrodia elata Blume, is a widely used Traditional Chinese Medicine preparation for the treatment of migraine. Ferulic acid and gastrodin are main active constituents in Ligusticum chuanxiong Hort and Gastrodia elata Blume, and have been used as marker components for quality control of Tianshu Capsule. In this study, a selective, sensitive, and reliable ultra-fast liquid chromatography with tandem mass spectrometry method was developed for simultaneous determination of ferulic acid and gastrodin in rat plasma using geniposide as internal standard. The plasma samples were extracted by protein precipitation with methanol after acidification and separated on a Shim-Pack XR-ODS C18 column (75 × 3.0 mm, 2.2 µm) using gradient elution with a mobile phase consisting of water (containing 0.1% formic acid) and acetonitrile at a flow rate of 0.6 mL/min. Detection was performed on 3200 QTRAP mass spectrometry equipped with turbo ion spray source in negative ionization mode. Validation parameters were within acceptable ranges. The validated method was applied to compare the pharmacokinetic profiles of ferulic acid and gastrodin in normal and migraine rats. Our results showed that there were remarkable differences in the pharmacokinetic properties of the analytes between the normal and migraine groups.

Schidandrin B kills tumor cells by initiating apoptosis in glioma SHG-44 cells.

OBJECTIVE: To observe the proliferation inhibition, cell cycle, and apoptosis of human glioma cell line SHG-44 treated with different concentration of Schidandrin B and explore the effect of Schidandrin B on glioma SHG-44 cells. METHODS: Glioma SHG-44 cells were treated with Schidandrin B (0, 50, 100 or 200 µg/mL) for 24, 48, 72 and 96 h, and cells were treated with vehicle as control. Viability of cells were detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis; cell cycle was examined with flow cytometry assay; apoptosis was detected with annexin V assay. Bax and caspase-3 proteins expression were checked by Western blot. RESULTS: MTT analysis showed that viability of glioma SHG-44 cells significantly decreased after exposure to Schidandrin B for the indicated time. Flow cytometry revealed that the number of cells in the sub G1 phase was increased, however, the number of cells in G0/G1, S and G2/M phases were decreased after treatment with 50, 100 or 200 µg/mL Schidandrin B, compared with the respective control group. Annexin V analysis confirmed that apoptosis rates of the control group, 50, 100, and 200 µg/mL Schidandrin B group were 1.76%±0.47%, 13.98%±5.05%, 19.64%±5.53% and 63.28%±6.88% respectively, apoptotic rate increased significantly with dose-dependent manner, and apoptosis of cells were observed under the inverted microscope after 100 µg/mL Schidandrin B treatment. Bax and caspase-3 protein were highly expressed in Schidandrin B group compared with the control group. CONCLUSION: The apoptosis could be induced by different concentration of Schidandrin B on glioma SHG-44 cells, and the mechanism may be directly excited by Schidandrin B in glioma SHG-44 cells through activating mitochondrial pathway.

[Efficacy of Schizandra chinesis polysaccharide on cyclophosphamide induced dyszoospermia of rats and its effects on reproductive hormones].

OBJECTIVE: To explore the efficacy of Schizandra Chinensis polysaccharide (SCP) on cyclophosphamide (CTX) induced dyszoospermia of rats and its effects on reproductive hormones. METHODS: SCP was extracted by ethanol-alkali solution. Fifty male Wistar rats were randomly divided into 5 groups, i.e., the normal control group, the model group, the low dose SCP group (100 mg/kg), the middle dose SCP group (200 mg/kg), and the high dose SCP group (400 mg/kg). Except the normal control group, the dyezoospermia rat model was established by peritoneal injection of CTX at the daily dose of 80 mg/kg, once daily for 5 successive days. After modeling, SCP was intragastrically administered at corresponding dose to the three SCP groups. Equal volume of normal saline was given to rats in the normal control group and the model group by gastrogavage. All the medication was performed once daily for 60 successive days. The blood serum and testis were withdrawal 24 h after the last intragastric administration. The levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), and testosterone (T) in the testis homogenate were determined by enzyme linked immunosorbent assay. The sperm count, the motility rate, and the teratospermia rate were compared. The morphology of the testis was observed using HE staining. RESULTS: Compared with the normal control group, the sperm count and the motility rate decreased, the teratospermia rate increased, the serum levels of FSH and LH increased, the T content in the testis homogenate decreased in the model group, showing statistical difference (P <0.01). Compared with the model group, the sperm count and the motility rate increased, the teratospermia rate decreased, the serum levels of FSH and LH decreased, the T content in the testis homogenate increased in the model group, showing statistical difference (P <0.01, P <0.05). All the indices showed dose-dependent manner in the SCP groups. The histological results showed the pathological injury in the testicular tissue was improved in all SCP groups. CONCLUSION: SCP showed obvious therapeutical effects on CTX induced dyszoospermia in rats, and its mechanisms might be correlate with recovering the regulation function of hypothalamus-hypophysis-gonad axis.

Synthesis and anticonvulsant activity of N-(2-hydroxy-ethyl)amide derivatives.

A series novel of N-(2-hydroxyethyl)amide derivatives was synthesized and screened for their anticonvulsant activities by the maximal electroshock (MES) test, and their neurotoxicity was evaluated by the rotarod test (Tox). The maximal electroshock test showed that N-(2-hydroxyethyl)decanamide 1g, N-(2-hydroxyethyl)palmitamide 1l, and N-(2-hydroxyeth-yl)stearamide 1n were found to show a better anticonvulsant activity and also had lower toxicity than the marked anti-epileptic drug valproate. In the anti-MES potency test, these compounds exhibited median effective doses (ED50) of 22.0, 23.3, 20.5 mg/kg, respectively, and median toxicity doses (TD50) of 599.8, >1000, >1000 mg/kg, respectively, resulting in a protective index (PI) of 27.5, >42.9, >48.8, respectively. This is a much better protective index than that of the marked anti-epileptic drug valproate (PI = 1.6). To further investigate the effects of the anticonvulsant activity in several different models, compounds 1g, 1l, and 1n were tested having evoked convulsions with chemical substances, including pentylenetetrazloe, isoniazide, 3-mercaptopropionic acid, bicuculline, thiosemicarbazide, and strychnine.