OBJECTIVETo compare the anaphylactoid effect of Danshen injection and its components on guinea pig.

METHODApplying active systemic anaphylaxis (ASA) tests, the corresponding experimental injections were administrated to guinea pigs to sensitized, and allergen with double doses was injected to stimulate in the 11 days after the last sensitized. The anaphylaxis situation of guinea pigs was observed.

RESULTDanshen injection and its components are suspicion on guinea pigs, while negative reaction was observed on guinea pigs which injected by the liquid excipients of Danshen injection.

CONCLUSIONDanshen injection using the ultrafiltration method still have some antigenic impurities which cannot be removed completely, and this may be one of the reasons for anaphylactic reaction.

Anaphylaxis ; chemically induced ; Animals ; Drugs, Chinese Herbal ; toxicity ; Female ; Guinea Pigs ; Injections ; methods ; Male ; Phenanthrolines ; toxicity ; Salvia miltiorrhiza ; chemistry ; toxicity

OBJECTIVETo compare the anaphylactoid effect of old Shengmai injection and new Shengmai injection on Cynomolgus monkey.

METHODCynomolgus monkeys were randomly divided into 4 groups, and were respectively injected with 5% glucose injection, old Shengmai injection, new Shengmai injection, positive control drug. The changes of each monkey were observed from injection before until 24 hours after injection, and the response level was determined according to the severity of the symptoms. Blood samples were collected before injection and at 10 min after injection for measuring histamine content in plasma. Blood pressure and heart rates were detected before injection and at 10 min after injection. Sensitization of the injection was comprehensively determined by combined the response level of symptoms and the histamine level.

RESULTThe Cynomolgus monkeys administrated with old Shengmai injection showed typical symptoms of anaphylactoid reactions and the content of serum histamine is not more than doubled. The Cynomolgus monkeys administrated with new Shengmai injection showed untypical symptoms of anaphylactoid reactions and the content of serum histamine did not rise.

CONCLUSIONThe old Shengmai injection can induce typical anaphylactoid reactions on Cynomolgus monkeys, and the sensitization ability is strong. The symptoms of anaphylactoid reactions induced by the new Shengmai injection appeared later and showed lesser degree with the sensitization lower.

Anaphylaxis ; chemically induced ; Animals ; Drugs, Chinese Herbal ; administration & dosage ; toxicity ; Injections ; methods ; Macaca fascicularis ; Male

OBJECTIVETo evaluate the sensitization effect of different components of Shengmai injection (new production process) on Beagle dogs.

METHODBeagle dogs were randomly divided into 7 groups, 3 in each group. Each group was respectively injected with 5% glucose injection, Ginseng Radix et Rhizoma Rubra extract, Ophiopogonis Radix extract, Schisandrae Chinensis Fructus extract, Schisandrae Chinersis Fructus distillate, Shengmaifang, 0.2% tween 80. The changes of each dog were observed from injection before until 24 hours after injection, and the response level was determined according to the severity of the symptoms. Blood samples were collected before injection and at 10 min after injection for measuring histamine content in plasma by ELISA. Sensitization of the injection was comprehensively determined by combined the response level of symptoms and the histamine level.

RESULTOne dog of Ginseng Radix et Rhizoma Rubra extract group showed untypical symptoms of anaphylactoid reactions, and serum histamine of two dogs increased more than doubled. The Beagle dogs administrated with 0.2% tween 80 showed typical symptoms of anaphylactoid reactions, while there was no significant increase of serum histamine. Other groups were observed with no typical anaphylactoid reactions.

CONCLUSIONThe sensitization effect of Shengmai injection (new production process) may be associated with Ginseng Radix et Rhizoma Rubra extract and 0.2% tween 80.

Anaphylaxis ; chemically induced ; Animals ; Dogs ; Drugs, Chinese Herbal ; administration & dosage ; chemistry ; toxicity ; Female ; Injections ; Male ; Random Allocation

Objective:To evaluate the effect of sufentanil on activation of Schwann cells after peripheral nerve injury in mice.Methods:Eighty healthy pathogen-free male Balb/c mice, aged 6-8 weeks, weighing 18-22 g, were divided into 4 groups ( n=20 each) using a random number table method: peripheral nerve injury group (group PNI), high dose sufentanil group (group H), medium dose sufentanil group (group M) and low dose sufentanil group (group L). The model of unilateral sciatic nerve transaction was established in ketamine-anesthetized mice.Immediately after establishment of the model, sufentanil 10, 5 and 2.5 μg/kg was injected intraperitoneally once a day for 3 consecutive days in H, M and L groups, respectively, while the equal volume of normal saline was given instead in group PNI.Sciatic function index (SFI) was calculated at 4, 8 and 12 weeks after establishment of the model.At 2, 4, 8 and 12 weeks, 5 mice in each group were sacrificed, and segments of the injuried ipsilateral sciatic nerve were removed for examination of the ultrastructure of the sciatic nerve (with a transmission electron microscope) and for detection of the expression of glial fibrillary acidic protein (GFAP) of sciatic nerve (by immunohistochemistry). Results:Compared with group PNI, SFI was significantly increased, and the expression of GFAP was up-regluated at each time point after establishment of the model in H and M groups ( P<0.05) and no significant change was found in SFI and GFAP expression after establishment of the model in group L ( P>0.05). Compared with group L, SFI was significantly increased, and GFAP expression was up-regluated in H and M groups ( P<0.05). There was no significant difference in SFI and GFAP expression between group H and group M ( P>0.05). The thickness of myelin lamellae was dense, and the proliferation of Schwann cells was not marked in H and M groups.The thickness of myelin lamellae was thin, and the proliferation of Schwann cells was marked in L and MO groups. Conclusion:The mechanism by which sufentanil improves repair after peripheral nerve injury may be related to promoting activation of Schwann cells in mice.

Objective To investigate the release kinetics of Zn2+ from nZCP-loaded polylactic acid/hydroxyapatite(PLA/HA)composite scaffold(PHZ)and determine the optimal nZCP content in the scaffold.Methods The particle size of nZCP was measured by DLS measurement,and PXRD,FTIR,and SEM were used to characterize the scaffolds and nZCP distribution;EDS was used to analyze element composition of the scaffold.Compression strength of the scaffold was determined,and ion release profile was investigated using ICP-MS.The biocompatibility of the materials was evaluated by CCK-8 assay and dead/alive staining of rat bone marrow stem cells(BMSCs)incubated with their aqueous extracts.ALP staining,alizarin red staining,RT-qPCR,and Western blotting were used to assess the osteogenic potential of the treated cells.In a rat model of bilateral ovariectomy(OVX)with femoral condylar bone defect,PHZ-1,PHZ-2,PHZ-3 or PLA/HA scaffold was implanted into the bone defect,and bone repair was observed using a microCT scanner and histological staining at 6 and 12 weeks.Results DLS,PXRD,SEM,FTIR,and EDS confirmed successful synthesis of 10-nm ZCP and efficient nZCP loading in the scaffold.PHZ-2 and PHZ-3 had significantly greater compression strength than PLA/HA.ICP-MS showed that Zn2+ release from PHZ-1,PHZ-2 and PHZ-3 were all optimal for promoting osteogenesis.In rat BMSCs,all the 4 scaffolds showed good biocompatibility,and their extracts enhanced ALP activity and extracellular matrix mineralization and promoted expressions of ALP,RUNX2,and OCN in the cells.In the rat models,nZCP in the implants improved bone graft integration at 6 weeks,and PHZ-2 and PHZ-3 more effectively induced new bone formation at 12 weeks(P<0.05).Conclusion PHZ scaffold is capable of stable Zn2+ release to promote osteoporotic bone defect healing,and PHZ-2 and PHZ-3 scaffolds with nZCP mass fraction of 4.5%-7.5%have better osteogenic activity.

Objective:To investigate the effect of total alkaloid of harmaline (TAH) on inducing cellular autophagy and degradating of neurotoxic proteins Tau and α-synuclein (α-Syn).Methods:(1) The in vitro cultured PC12 cells were divided into blank control group, and 1, 2.5, 5, 10, 20 and 50 μg/mL TAH groups, respectively; and they were treated with 0, 1, 2.5, 5, 10, 20 and 50 μg/mL TAH for 24 h; cell morphology and number were observed, and cell survival rate was determined by MTT assay. (2) PC12 cells were divided into blank control group, rapamycin group, and 1, 2.5, 5, 10 and 20 μg/mL TAH groups; these cells were treated with same amount of solvent, 50 nmol/L autophagy activator rapamycin, and 1, 2.5, 5, 10 and 20 μg/mL TAH for 4 h, respectively, and the number of autophagosomes was detected by immunofluorescent staining. (3) PC12 cells were divided into blank control group, rapamycin group, and 10 μg/mL TAH group; these cells were treated with same amount of solvent, 50 nmol/L rapamycin, and 10 μg/mL TAH for 4 h; the protein expression levels of p62 and microtubule-associated protein 1 light chain 3 II (LC3-II) was detected by Western blotting. (4) PC12 cells were divided into blank control group, chloroquine group, TAH group, and TAH+chloroquine group; these PC12 cells were treated with 50 nmol/L autophagy inhibitor chloroquine, 10 μg/mL TAH, and 10 μg/mL TAH+50 nmol/L chloroquine for 4 h, respectively; the LC3-II protein expression was detected by Western blotting. (5) PC12 cells were divided into TAH group and blank control group; 10 μg/mL TAH and same amount of solvent were given to each group for 4 h, and then, the phosphorylated mammalian target of rapamycin (p-mTOR) and phosphorylated 70-KD ribosomal protein S6 kinase (p-P70S6K) protein expression levels were detected by Western blotting. (6) Tet on HEK293 cells with Tau-green fluorescent protein (GFP) overexpression were divided into blank control group, TAH group, doxycycline group, doxycycline+TAH group, doxycycline+TAH+3-MA group, and doxycycline+TAH+chloroquine group. Cells in the later 4 groups were treated with 200 ng/mL Tet system inducer doxycycline for 24 h; cells in the blank control group were treated with same amount of solvent, those in the TAH group were treated with 10 μg/mL TAH, and cells in the latter 3 groups were treated with 10 μg/mL TAH, 10 μg/mL TAH+5 mmol/L 3-MA, and 10 μg/mL TAH+50 nmol/L chloroquine, respectively, for 24 h; the changes of green fluorescence intensity of these cells were observed under laser confocal microscope. The Tau-GFP and LC3-II protein expression levels were detected by Western blotting. (7) HEK293 cells with stable α-Syn expression were divided into blank control group, chloroquine group, TAH group and TAH+chloroquine group; these cells were treated with same amount of solvent, 50 nmol/L chloroquine, 10 μg/mL TAH and 10 μg/mL TAH+50 nmol/L chloroquine for 24 h, respectively; the α-Syn and LC3-II protein expression levels were detected by Western blotting. Results:(1) As compared with that in the blank control group, the cell survival rate in 20 and 50 μg/mL TAH groups was significantly lower, and that in the 50 μg/mL TAH group was statistically lower than that in 20 μg/mL TAH group ( P<0.05). (2) As compared with that in the blank control group, the number of autophagosomes in rapamycin group, and 10 and 20 μg/mL TAH groups was significantly increased, and that in 10 μg/mL TAH group was statistically higher than that in 20 μg/mL TAH group ( P<0.05); 10 μg/mL TAH group was selected for subsequent experiments. (3) As compared with the blank control group, the rapamycin group and TAH group had significantly decreased P62 protein expression and significantly increased LC3-II protein expression ( P<0.05). (4) As compared with that in the blank control group, the LC3-II protein expression in the chloroquine group, TAH group and TAH+chloroquine group was significantly increased, and LC3-II protein expression in TAH+chloroquine group was statistically higher than that in chloroquine group ( P<0.05). (5) The p-mTOR and p-p70S6K expression levels in the TAH group were significantly decreased as compared with those in the blank control group ( P<0.05). (6) The Tau-GFP protein expression in doxycycline group was significantly increased as compared with that in the blank control group ( P<0.05); that in doxycycline+TAH group was significantly decreased as compared with that in the doxycycline group ( P<0.05); that in the doxycycline+TAH+3-MA group and doxycycline+TAH+chloroquine group was statistically increased as compared with that in doxycycline+TAH group ( P<0.05). The LC3-II protein expression in the TAH group was significantly increased as compared with that in the control group, that in the doxycycline+TAH group was significantly increased as compared with that in the doxycycline group, that in the doxycycline+TAH+3-MA group was significantly decreased as compared with that in the doxycycline+TAH group, and that in doxycycline+TAH+ chloroquine group was significantly increased as compared with that in the doxycycline+TAH group ( P<0.05). Conclusion:TAH may activate autophagy by inhibiting the mTOR/p70S6K signaling pathway, which in turn promotes the degradation of neurotoxic proteins Tau and α-Syn.

Objective To investigate the release kinetics of Zn2+ from nZCP-loaded polylactic acid/hydroxyapatite(PLA/HA)composite scaffold(PHZ)and determine the optimal nZCP content in the scaffold.Methods The particle size of nZCP was measured by DLS measurement,and PXRD,FTIR,and SEM were used to characterize the scaffolds and nZCP distribution;EDS was used to analyze element composition of the scaffold.Compression strength of the scaffold was determined,and ion release profile was investigated using ICP-MS.The biocompatibility of the materials was evaluated by CCK-8 assay and dead/alive staining of rat bone marrow stem cells(BMSCs)incubated with their aqueous extracts.ALP staining,alizarin red staining,RT-qPCR,and Western blotting were used to assess the osteogenic potential of the treated cells.In a rat model of bilateral ovariectomy(OVX)with femoral condylar bone defect,PHZ-1,PHZ-2,PHZ-3 or PLA/HA scaffold was implanted into the bone defect,and bone repair was observed using a microCT scanner and histological staining at 6 and 12 weeks.Results DLS,PXRD,SEM,FTIR,and EDS confirmed successful synthesis of 10-nm ZCP and efficient nZCP loading in the scaffold.PHZ-2 and PHZ-3 had significantly greater compression strength than PLA/HA.ICP-MS showed that Zn2+ release from PHZ-1,PHZ-2 and PHZ-3 were all optimal for promoting osteogenesis.In rat BMSCs,all the 4 scaffolds showed good biocompatibility,and their extracts enhanced ALP activity and extracellular matrix mineralization and promoted expressions of ALP,RUNX2,and OCN in the cells.In the rat models,nZCP in the implants improved bone graft integration at 6 weeks,and PHZ-2 and PHZ-3 more effectively induced new bone formation at 12 weeks(P<0.05).Conclusion PHZ scaffold is capable of stable Zn2+ release to promote osteoporotic bone defect healing,and PHZ-2 and PHZ-3 scaffolds with nZCP mass fraction of 4.5%-7.5%have better osteogenic activity.

Lysosomes are degradation and signaling centers within the cell, and their dysfunction impairs a wide variety of cellular processes. To understand the cellular effect of lysosome damage, we screened natural small-molecule compounds that induce lysosomal abnormality using Caenorhabditis elegans (C. elegans) as a model system. A group of vobasinyl-ibogan type bisindole alkaloids (ervachinines A-D) were identified that caused lysosome enlargement in C. elegans macrophage-like cells. Intriguingly, these compounds triggered cell death in the germ line independently of the canonical apoptosis pathway. In mammalian cells, ervachinines A-D induced lysosomal enlargement and damage, leading to leakage of cathepsin proteases, inhibition of autophagosome degradation and necrotic cell death. Further analysis revealed that this ervachinine-induced lysosome damage and lysosomal cell death depended on STAT3 signaling, but not RIP1 or RIP3 signaling. These findings suggest that lysosome-damaging compounds are promising reagents for dissecting signaling mechanisms underlying lysosome homeostasis and lysosome-related human disorders.
Alkaloids ; pharmacology ; Animals ; Caenorhabditis elegans ; cytology ; drug effects ; metabolism ; Cell Death ; drug effects ; Cell Survival ; drug effects ; HeLa Cells ; Humans ; Lysosomes ; drug effects ; pathology ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; drug effects