1.Effect of Kanglaite Injection on Proliferation and Apoptosis of Human Breast Cancer MCF-7 Cells
Jun CHEN ; Yan MA ; Yu LI ; Long QIN ; Li YAO
Chinese Journal of Information on Traditional Chinese Medicine 2014;(11):48-51
Objective To investigate effects of Kanglaite injection on proliferation, cycle and apoptosis of human breast cancer MCF-7 cells;To discuss its relevant mechanism. Methods Logarithmic growth phase cells were divided into control group and Kanglaite-treatment group (10, 20, 40μL/mL). Cells were cultured in RPMI-1640 for 24 h before drug treatment. The inhibition rate of Kanglaite injection on proliferation of human breast cancer MCF-7 cells was detected by MTT assay. Apoptosis and cell cycle of MCF-7 cells were detected by flow cytometry. Changes in cell nucleus were determined by Hochest staining assay. Protein expressions of Bcl-2 and Bax were detected by ELISA and Western blot. Results Kanglaite injection for 12 h, 24 h or 48 h resulted in a significant inhibition of MCF-7 cells proliferation (P<0.05, P<0.01);Compared with the control group, Kanglaite injection-treated cells showed increased percentage in G2/M and G0/G1 phases (P<0.001, P<0.01), but showed decreased percentage in S phase (P<0.01), and apoptosis rate increased (P<0.05, P<0.001). Kanglaite injection significantly decreased protein expression of Bcl-2, and enhanced protein expression of Bax of MCF-7 cells (P<0.01, P<0.001). Conclusion Kanglaite injection can inhibit the proliferation of human breast cancer MCF-7 cells, decrease cell cycle and induce apoptosis, the mechanism is related with decreasing protein expression of Bcl-2 and enhance the protein expression of Bax.
2.Influence of chronic fluorosis on expression of phospho-Elk-1 in rat brains
Yan-jie, LIU ; Qin, GAO ; Yi-guo, LONG ; Yan-ni, YU ; Zhi-zhong, GUAN
Chinese Journal of Endemiology 2011;30(3):251-255
Objective To investigate the expression and distribution of the downstream substrate of extracellular regulated protein kinase(ERK1/2) pathway, ternary complex factor phospho-Elk-1, in rat brains with chronic fluorosis, and reveal the mechanism of the impaired learning and memory ability caused by chronic fluorosis. Methods Seventy-two SD rats, weighing 100 - 120 g, were randomly divided into 3 groups, 24 in each group (half male and half female). The rats in control group were fed with tap water (fluoride < 0.5 mg/L); low- and high-dose fluoride groups were fed with tap water with different concentrations of NaF(5.0,50.0 mg/L F-, respectively). After 6 months, body weight was weighed, dental fluorosis was determined by observation and urinary fluoride and bone fluoride were detected by fluorine ion-selective electrode; the learning ability of rats was measured by navigation test of Morris water maze, and memory ability by spatial probe test in Morris water maze; the expression and distribution of phospho-Elk-1 in different brain regions were detected by immunohistochemistry method. Results In low- and high-fluoride groups, the body weight of rat[(449.2 ± 77.1), (312.8 ± 89.7)g] was significantly decreased than that of control [(635.5 ± 76.2 )g, all P< 0.05], the varying degrees of dental fluorosis were observed(x2 = 7.83, P<0.05), urinary fluoride[(2.56 ±0.91),(5.73 ±3.14)mg/L] and bone fluoride[(709.2 ± 37.4) ,(1306.3 ± 102.4) mg/kg] were significantly higher than those in controls[(0.92 ± 0.30)mg/L,(348.5 ± 89.2)mg/kg, all P< 0.05]. The escape latency of low- and high-fluoride groups[ (7.4 ± 4.1), (12.2 ± 5.7)s] was longer than that of control [(4.8 ± 2.7 )s, all P < 0.05] and the escape latency in high-fluoride group was significantly longer than that in other groups (all P < 0.05); in spatial probe test, the time of first crossing platform was longer in rats with fluorosis [(4.18 ± 1.10),(5.89 ± 0.56)s] as compared to control[(1.17 ± 0.75)s, all P< 0.05]. Expressions of phospho-Elk-1 in the hippocampus CA1(167.4 ± 8.3,163.2 ± 9.4), CA2(175.7 ± 5.0,183.3 ± 4.2), CA3(165.2 ± 11.6,162.9 ± 4.4), CA4(168.7± 6.9,169.5 ±5.3), fascia dentate (185.2 ±4.0,193.1 ±6.1) and caudate putamen( 181.4 ± 3.8, 179.8 ± 5.5) in low- and high-fluoride groups were higher than those of controls(142.4 ± 8.1,144.9 ± 8.4,143.6 ± 5.8, 116.8 ± 9.1,140.2 ± 7.8,163.1 ± 13.1, all P< 0.05). Conclusion Chronic fluorosis can cause increased expression of phospho-Elk-1 in the hippocampus and caudate putamen region of rat brains, which might be related to the mechanisms of decreased learning and memory ability of rats overexposed to fluoride.
3.Effects of S100B on the expressions of dopamine receptors and synthesis, metabolism of neurotransmitters dopamine and 5-hydroxytryptamine
Jialin LIU ; Fang ZHENG ; Yan LONG ; Yuan ZHENG ; Yan LIN ; Xiaoqing LIU ; Chuan QIN
Chinese Journal of Neurology 2016;49(9):697-702
Objective To investigate the effects of S100B on the expressions of dopamine receptors and the synthesis,metabolism of neurotransmitters dopamine,5-hydroxytryptamine which are related to the abnormal motor coordination of Parkinson's disease (PD).Methods The hS100B transgenic mice were established.The mice were divided into S100B transgenic group (TG,n =14),S100B knockout group (KG,n =14) and the non-transgenic control group (CG,n =14).The motor coordination ability of mice was measured by the Rota-rod test.The expressions of dopamine D1 receptor (D1DR),dopamine D2 receptor (D2DR),tyrosine hydroxylase (TH) and phosphorylated TH at Ser19,Ser31,Ser40 in brain tissue were detected by reverse transcription polymerase chain reaction and Western blotting.The levels of Tyr,levodopa,dopamine,homovanillic acid,Trp,5-hydroxytryptamine and 5-hydroxyindoleacetic acid in mesencephalon were measured by high performance liquid chromatography with fluorescence detection.Results Compared with CG,the motor coordination ability of mice (s) showed progressive decline in TG (3 months:4.60±0.30vs4.25±0.21,q =5.194;6 months:4.52±0.31 vs4.07±0.22,q =6.139;9 months:4.43 ± 0.25 vs 3.60 ± 0.18,q =13.484;all P < 0.05),the expressions of D2DR mRNA and protein decreased (1.34 ± 0.13 vs 0.48 ± 0.07,q =21.578;1.05 ± 0.15 vs 0.69 ± 0.10,q =8.063,both P<0.05) and phosphorylated TH at Serl9 and Ser40 increased (0.95 ±0.10 vs 1.14-0.13,q =4.972;0.94 ± 0.12 vs 1.17 ± 0.14,q=5.382,both P< 0.05),the levels of levodopa,dopamine and homovanillic acid were elevated (87.04 ± 11.77 vs 115.28 ± 16.80,q =4.764;56.66 ± 9.87 vs 72.96 ± 11.02,q=3.923;26.58 ± 8.11 vs 38.65 ± 6.67,q=3.981,all P< 0.05),the leve1 of 5-hydroxytryptamine was reduced (925.50 ± 74.26 vs 637.87 ± 56.76,q =11.084,P < 0.05),the ratios of homovanillic acid/dopamine and 5-hydroxyindoleacetic acid/5-hydroxytryptamine increased (0.45 ± 0.05 vs 0.54±0.08,q =3.325;0.94±0.07 vs 1.42±0.12,q =12.367,both P<0.05) in the brain of TG at the age of 9 months old.There was no significant difference of detection indexes between KG and CG.Conclusions S100B plays an important role in the development of PD and the brain-specific S100B transgenic mice can be used to investigate the function of S100B gene on the development of PD.
4.A Primary Study of the Subgroups of T Lymphocytes in MHV-3 Induced Chronic Viral Hepatitis
Jiang-guo, ZHANG ; Xiao-min, QIN ; Xiao-jing, WANG ; Wei-ming, YAN ; Chuan-long, ZHU ; Xiao-ping, LUO ; Qin, NING
Virologica Sinica 2007;22(5):339-346
To study the contribution of T cell subsets in the pathogenesis of Murine hepatitis virus Type3 (MHV-3) induced chronic viral hepatitis in C3H/Hej mice, ninety C3H/Hej mice were chosen to individually receive 10 plaque forming units (PFU) of MHV-3 intraperitoneally. The changes of virus titer and pathology in liver tissue were examined by standard plaque assay and by the hematoxylin/eosin (HE) staining method from 2 days post MHV-3 infection. The ratios of T cell subsets including CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4-CD8-, CD3+CD4+CD25+, CD3+CD4+CD25- and CD3+CD4-CD25+ T lymphocyte of total T lymphocytes in blood, spleen and liver were examined at 0, 2, 4, 6,8, 10, 12, 15, 20, 25, 30, 40 days post MHV-3 infection by flow cytosorting. We observed that the virus titer raised and showed persistent virus duplications and inflammatory changes in the livers of C3H/Hej mice from 2 days post MHV-3 infection. The double negative T cell (DN Treg cell) and CD4+CD25+ T cell ratios increased significantly from 2 days post MHV-3 infection in C3H/Hej mice, and CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4+CD25- and CD3+CD4-CD25+ T cell ratios decreased accordingly. In conclusion, the changes of virus titer and pathology in the livers of C3H/Hej mice post MHV-3 suggest their contribution to viral persistence. Further characterizations of DN Treg cells are that infection indicates that MHV-3 could induce the chronic inflammation in livers of C3H/Hej mice.The increase of the DN Treg cell and CD4+CD25+ T cell ratios in C3H/Hej mice post MHV-3 infection suggests that DN Treg cells and CD4+CD25+ T cells may both have important suppressive immunomodulation functions in the development of chronic viral hepatitis and have important roles in the virus persistent infection. Further characterizations of DNT cell and CD4+CD25+ T cell are under investigation.
5.Effect of reactive oxygen species induced by paraquat on neutrophil apoptosis.
Kai-xiu QIN ; Chun-wen LI ; Yan FANG ; Lei YU ; Xiao-long WANG
Chinese Journal of Applied Physiology 2015;31(2):111-114
OBJECTIVETo investigate the effect of paraquat (PQ) on reactive oxygen species (ROS) and neutrophil apoptosis and its possible signal transduction pathways.
METHODSCultured neutrophils were treated with different concentrations of PQ for 6-24 h. The apoptosis rate of neutrophils and ROS content were determined by flow cytometry. The exoressions of nuclear factor kappa B (NF-κB) and Caspase 3 were detected by Western blot. These parameters were checked again after NF-κB and Caspase 3 antagonist were applied.
RESULTSPQ could boost ROS generation and depress neutrophil apoptosis significantly. At the same time PQ could enhance the expression of NF-κB and inhibit the expression of Caspase 3. These effects could be reversed by ROS inhibitor diphenyleneiodonium (DPI) and NF-κB inhibitor pyrrolidinedithiocarbamate (PDTC).
CONCLUSIONPQ is a potent inducer of ROS and can inhibit neutrophil apoptosis by activating NF-κB and surpressing Caspase 3 activity.
Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cells, Cultured ; NF-kappa B ; antagonists & inhibitors ; metabolism ; Neutrophils ; cytology ; drug effects ; Paraquat ; toxicity ; Pyrrolidines ; pharmacology ; Reactive Oxygen Species ; metabolism ; Signal Transduction ; Thiocarbamates ; pharmacology
6.Role of JNK signal pathway in brain injury after resuscitation in a rat model of asphyxia cardiac arrest
Long CHEN ; Mingzhang ZUO ; Gongjian LIU ; Xiyan CHEN ; Yan ZHANG ; Qin CHENG ; Maoyin ZHANG
Chinese Journal of Anesthesiology 2012;32(1):93-95
Objective To evaluate the role of JNK signal pathway in brain injury after resuscitation in a rat model of asphyxia cardiac arrest.Methods Forty healthy male SD rats 'weighing 300-350 g were randomly divided into 4 groups ( n =10 each):sham operation group (group SH) ; cardiac arrest group (group CA) ; group SP600125-JNK inhibitor (group SP) and dimethyl sulfexide (DMSO) group.The rats were anesthetized with intraperitoneal pentobarbital 45 mg/kg,tracheostomized and mechanically ventilated.PETCO2 was maintained at 35-45 mm Hg.Femoral artery and vein were cannulated for BP monitoring and fluid infusion.Cardiac arrest was induced by clamping tracheal tube until ECG activity disappeared and MAP < 10 mm Hg.Resuscitation was started at 3 min after cardiac arrest.MAP > 60 mm Hg and HR > 250 bpm were considered to be signs of successful resuscitation.SP600125 20 mg/kg and DMSO 0.2 ml were injected iv as soon as chest compression was started in groups SP and DMSO respectively.The animals were sacrificed at 5 h after successful resuscitation and their brains were removed for determination of wet/dry (W/D) weight ratio and microscopic examination of hippocampus.Neuronal apoptosis was detected by TUNEL.Results Cardiac arrest significantly increased W/D ratio and the number of apoptotic cells in group CA.SP600125 iv significantly attenuated the cardiac arrest-induced increase in W/D ratio and the number of apoptotic cells but DMSO did not.Conclusion JNK signal pathway is involved in the brain injury after resuscitation in a rat model of asphyxia cardiac arrest.
7.Monitoring Urine Thiodiglycol after Rabbit Skin Exposure to Sulfur Mustard Based on Highly-sensitive Isotope Dilution Gas Chromatography-Mass Spectrometry
Zhiyong NIE ; Yajiao ZHANG ; Bidong WU ; Long YAN ; Jianlin FENG ; Qin LIU ; Jianwei XIE
Chinese Journal of Analytical Chemistry 2014;(7):980-984
A sensitive determination method for sulfur mustard ( HD ) metabolites thiodiglycol ( TDG ) in rabbit urine was established and validated using isotope dilution negative ion chemical ionization ( NICI) gas chromatography-mass spectrometry ( GC-MS ) , in which deuterated thiodiglycol ( TDG-d8 ) was used as internal standard. Two solid-phase extraction ( SPE) steps were established and optimized in order to reduce the interfering backgrounds, one was used to extract thiodiglycol ( TDG ) from urine with self-assemblied Florisil SPE cartridges, another cleaning treatment of the by-products after pentafluorobenzoyl chloride (PFBZ) derivatization. The results showed that the limits of detection quantitation of this method were 0. 1 and 0. 3 μg/L, respectively. The exposure time-response relationship and exposure dose-response relationship of TDG in rabbit urine were studied after rabbit skin exposure to sulfur mustard (HD, 0. 02-0. 15 LD50). The TDG levels in the rabbit urine increased rapidly during the first day after application and then decreased over time for all dosage groups. A secondary release was also noted for the high-dose group, and the duration of high TDG excretion levels was correlated positively with the HD dosage levels. We thus concluded that abnormally high levels of TDG in urine could be used as a clear diagnostic indicator of HD exposure.
8.Modified expression of extracellular signal-regulated protein kinase signal transducfion in rat braing and changed capacity of learning and memory of rats with chronic fluorosis
Yan-jie, LIU ; Qin, GAO ; Chang-xue, WU ; Yi-guo, LONG ; Zhi-zhong, GUAN
Chinese Journal of Endemiology 2009;28(1):32-35
Objective To investigate the expression of extraeellular signal-regulated protein kinase (ERK1/2)pathway in rat brains with fluorosis and the effects of fluoride on learning and memory of the rats,and to reveal the mechanisms of damaged nervous system resulted from the toxicity of the ion.Methods Seventy-two SD rats were divided into 3 groups and 24 rats were in each group.Three groups were fed respectively with different concentrations of fluoride(NaF)for 6 months to establish rat models with fluorosis.Controls were fed with tap water (NaF<0.5 mg/L):lower and higher concentration group were fed with water containing NaF(5,50 ms/L).Animals are sacrificed after 6 months of treatment with fluoride and the dissected brains were kept for analysis.The protein levels of ERK1/2 in rat brains were detected by Western-blotting and the mRNA level by RT-PCR. The spatial learning and memorizing ability was measured by Morris water maze test. Results The ERK1/2 protein in control group,lower and higher concentration group was 0.944±0.10,1.253±0.02,1.953±0.07,the differece being statistieally sighificant between any two groups (P < 0.05). The phospho-ERKl/2 protein in control group,lower and higher concentration group was 0.73±0.08,0.77±0.07,1.28±0.11,the differece being statistieally sighificant between any two groups(P < 0.05);the activation rate of phospho-ERK1/2 in lower and higher concentration group [(68.4± 3.8)%,(64.1±3.2)%] was decreased compared to control group[ (82.3±10.7)%],the differece being significant(P < 0.05). In the navigation trial,longer escape latencies of lower concentration group on the second, the third,the fifth and the sixth day were observed[ (46.0±8.0),(24.0±2.7),(8.9±5.3),(7.4±4.1 )s] compared to the control[ (39.3±6.9),(19.1±9.1 ),(8.3±3.4),(4.8±2.7)s],the differece being significant (P < 0.05 or < 0.01 );the similar results were also observed in the higher concentration group[ (36.9±16.8),(37.7±12.9), (19.7±7.6),(12.2±5.7 )s],and the escape latencies of the higher concentration group on the third,the fifth and the sixth day were longer than that in lower concentration group. In the probe test,the rats took more time to reach the first cross in lower and higher concentration group[(1.17±0.75),(4.18±1.10)s] than control group[ (5.89± 0.56 ) s ],the differece being significant (P < 0.05 or < 0.01 ) ;stayed shorter [ ( 17.05±4.25 ),(18.20±4.57 ) s ] than control [(25.37±5.65 )s ] in platform area (P < 0.01 );the activation rates of ERK1/2 were directly correlated with the time taken to reach the first cross platform located in the probe test(r = 0.364,P < 0.05) and the activation rates were also directly correlated with the escape latencies on the sixth day(r = 0.497,P < 0.05). Conclusion Long-term exposure of excessive fluoride induces the change of expression and activating rate of the ERK1/2 in rat brains,leading to the decreased capacity of learning and memory.
9.Level of oxidative stress in rat brains and learning and memory function of rats with chronic fluorosis
Qin, GAO ; Yan-jie, LIU ; Chang-xue, WU ; Yi-guo, LONG ; Zhi-zhong, GUAN
Chinese Journal of Endemiology 2008;27(4):371-373
Objective To investigate the changes of oxidative stress level in brain tissues and serum, and learning and memory in rats with oxidative stress level in nerve damage in chronic fluorosis. Methods The rats were randomly divided into 3 groups according to the body weight, eight rats in each group, i.e., control group, drinking water containing less than 0.5 mg/L of fluoride; lower fluoride exposure group, drinking water containing 5 mg/L of fluoride; higher fluoride exposure group, drinking water containing 50 mg/L of fluoride. The animals were examined six months after initiating the experiment. The total antioxidant capacity (T-AOC) and malondialdehyde (MDA), as well as learning and memory, were measured. Results Escape latency in higher fluoride exposed group[ (14.37±3.48)s] was significantly higher than that of controls[ (5.84±1.87)s] and exposed te lower fluoride [ (7.18±1.42)s], the difference being statistically signifieant(P<0.05). As compared with controls[ (2.17±0.11)× 103 U/L , (0.79±0.11)×103 U/g Pr] ,the rats exposed to higher fluoride and lower fluoride exhibited lower levels of T-AOC [(1.37±0.27)×103 U/L,(0.24±0.06)×103 U/g Prand (1.20±0.14) x 103 U/L,(0.41 ~ 0.10)×103 U/g Pr], the difference being statistically signifieant(P<0.05). As compared with controls[ (2.34±0.16) mmoL/L, (2.97±0.11)mmol/g Pr] and low fluoride exposed group[ (2.68±0.33)mmoL/L, (3.38±0.21)mmol/g Pr], higher level of MDA were observed in higher fluoride exposed group[ (3.72±0.59)retool/L, (4.01±0.21)mmol/g Pr], the difference being statistically significant(P<0.05). Conclusion The results indicated that higher amount of fluoride induced an increased level of oxidation, which might result in the decreased capacity of intelligence of rats with fluorosis.
10.Long-Term Therapeutic Effects of Different Therapeutic Programs in Ch ildren with Helicobacter Pylori Infection and Approach to Retreatment after Fail ure of Treatment
lan-ying, WU ; long-qin, CHANG ; fu-tang, YANG ; yan, XU
Journal of Applied Clinical Pediatrics 2004;0(09):-
Objective To evaluate the eradication rate and long-term therapeutic effect of a triple therapy consisted of cla-(rithromycin) (CLA), amoxicillin (AMO)and omeperazole on Hp infection,and explore the alternative therapeutic programs and their effects after first therapeutic failure.Methods A total of 92 children with Hp infection were divided into two groups: 70 children were given the triple therapy for one week (CLA group);Twenty-two children were given another triple therapy composed of metronida-(zoole) (MET), AMO and omeperazole for two weeks (MET group).All of the children were followed up for 1-30 months after the therapies ended.Children of the two groups who were therapeutic failure were given retreatment as follows.CLA triple therapy were given for one week to the children who were failure after MET triple therapy;increased doses of CLA with longer treatment course was given to the children who were failure after CLA triple therapy . A tetra therapy consisted of colloidal bismuth subcitrate (CBS), furazolidone (FUR) ,omeperazole and AMO was given to children in whom the retreatment failed.Results The Hp eradication rate of CLA group was 91.4%(64/70),and the Hp eradication rate of MET group was 72.7%(16/22).There was significant difference between eradication rate of the two groups(?~2=5.16 P