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OBJECTIVE To explore whether diterpenoid 12-deoxyphorbol-13-palmitate （DP） from Euphorbia fischeriana can exert anti-leukemia effects through the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt） signal pathway， and to provide experimental evidence for developing it into a new anti-leukemia drug. METHODS Using LY294002 （PI3K specific inhibitor） as tool drug， the effects of 24 h DP treatment on the proliferation and apoptosis of human myeloid leukemia HL60 cells were detected by MTT method， Annexin Ⅴ-FITC/PI staining and AO-EB staining. ELISA method was used to detect lactic dehydrogenase （LDH） release and the activities of cysteinyl aspartate specific proteinase 3 （caspase-3） and caspase-9. The transcriptional level of caspase-3， caspase-9， forkhead box O3a （FoxO3a） and B cell lymphoma 2 interacting mediator of cell death （Bim） mRNA were detected by real-time quantitative polymerase chain reaction （qRT-PCR）. The protein expression of phosphorylated FoxO3a （p- FoxO3a） and phosphorylated Akt （p-Akt） were detected by Western blot method. The nuclear translocation of FoxO3a protein was detected by immunostaining combined with laser confocal microscopy. RESULTS 10 μmol/L DP and 10 μmol/L DP+LY294002 could inhibit the proliferation and induce the apoptosis of HL60 cells （P＜0.01）. After treatment of 5， 10， 20 μmol/L DP， HL60 cells showed typical morphological characteristics of apoptosis； DP could significantly increase the levels of LDH release and the activities of caspase-3 and caspase-9 （P＜0.05 or P＜0.01）， in dose-dependent manner. After treatment of 10 μmol/L DP and 10 μmol/L DP+LY294002， the transcriptional levels of caspase-3， caspase-9 and Bim mRNA were increased significantly （P＜0.05 or P＜0.01）， and transcriptional level of FoxO3a mRNA and protein expressions of p-FoxO3a and p-Akt were decreased significantly （P＜0.05 or P＜0.01）. Nuclear translocation changes were observed in FoxO3a protein in 10 μmol/L DP+LY294002 group， and the change was more significant than that of LY294002 group. CONCLUSIONS DP can inhibit the proliferation and induce the apoptosis of HL60 cells via inhibiting PI3K/Akt signaling pathway.
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Microglia differ from macrophages with unique origin and role. In glioblastoma, microglia plays an important role in regulating tumor immune status, promoting tumor angiogenesis, destroying the blood-brain barrier, and reducing the sensitivity of treatment. Therefore, the combination of microglial therapy in the radiotherapy, chemotherapy and immunotherapy of glioblastoma has also become a clinically promising treatment.
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OBJECTIVE：To study the improvement eff ects and p otential mechanism of chelidonine on CCl 4-induced hepatic fibrosis model rats. METHODS ：According to the random number table method ，a total of 48 rats were randomly divided into normal control group ，model group ，chelidonine low-dose ，middle-dose and high-dose groups （0.125，0.25，0.50 mg/kg），positive control group （Liver-protecting tablet ，0.42 g/kg），with 8 rats in each group. Except for normal control group ，other groups were given CCl 4-olive oil solution intraperitoneally for 8 weeks to induce hepatic fibrosis model. From the fifth week of modeling ， normal control group and model group were given water intragastrically ；administration groups were given relevant medicine intragastrically，once a day ，for consecutive 10 weeks. After last intragastric administration ，hepatic index of rats was calculated. The levels of aspartate aminotransferase （AST），alanine aminotransferase （ALT）and hyaluronic acid （HA）in serum and the level of hydroxyproline （Hyp）in liver tissue were determined. The staining of collagen fibrin in rat liver was observed. The protein expression of α-smooth muscle actin （α-SMA），microtubule-associated protein 1 light chain 3（LC3）and p 62 as well as the phosphorylation level of phosphoinositide 3 kinase（PI3K），protein kinase B （Akt）and mammalian target of rapamycin （mTOR）in liver tissue were determined ；mRNA expression corresponding to above protein were also determined. RESULTS ：Compared with normal control group ，the hepatic index ，the serum levels of AST ，ALT，HA and Hyp ，the percentage of positive staining area for collagen fibrin ，the mRNA and protein expression of α-SMA and LC 3- Ⅱ were increased significantly （P＜0.05）. Protein expression of p 62，phosphorylation levels of PI 3K，Akt and mTOR as well as mRNA expression of p 62，PI3K，Akt and mTOR were significantly down-regulated （P＜0.05）. Compared with model group ，phosphorylation levels of PI 3K and mTOR were decreased significantly in chelidonine low-dose group （P＜0.05）. The changes of above indexes in chelidonine middle-dose and high-dose groups （except for liver index ， HA level in middle-dose group ） were reversed significantly （P＜0.05）. CONCLUSIONS：Chelidonine can attenuate CCl 4-induced liver fibrosis in rats ；the mechanism of it may be associated with activating PI 3K/Akt/mTOR signaling pathway and inhibiting autophagy.
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Aim To observe the ameliorating effects of sophocarpine on mice with neuropathic pain induced by spared nerve injury(SNI), and to explore the analgesic mechanism.Methods Sixty male ICR mice were randomly divided into six groups: sham-operated, neuropathic-pain model(SNI), SNI+Pregabalin(Pre) 10 mg·kg-1, SNI+SC 40 mg·kg-1, 20 mg·kg-1 and 10 mg·kg-1 group.Neuropathic-pain mouse models were established by SNI of the sciatic nerve.The paw withdrawal mechanical threshold(PWMT), paw withdrawal thermal latency(PWTL), and cold withdrawal times(CWT) were detected to assess the analgesic effects of sophocarpine on mice with neuropathic pain induced by spared nerve injury.The expressions levels of TLR4, p38 MAPK, IL-1β and TNF-α mRNA and protein in spinal cord tissue were detected by reverse transcription-polymerase chain reaction(RT-PCR) and Western blot methods.Results Compared with sham group, the SNI group′s PWMT decreased, PWTL was shorten, CWT increased, and the expression levels of p38 MAPK, TLR4, TNF-α and IL-1β mRNA and protein in spinal cord tissue were up-regulated.Compared with SNI model group, the sophocarpine 40 mg·kg-1 group′s PWMT increased, PWTL was lengthened, CWT decreased, and the expression levels of p38 MAPK, TLR4, TNF-α and IL-1β mRNA and protein in spinal cord tissue were down-regulated.Conclusion Sophocarpine has the analgesic effects on neuropathic pain induced by SNI, and its mechanism may be related to the down-regulation of TLR4, p38 MAPK, IL-1β and TNF-α expression levels.