INTRODUCTIONThe treatment of human epithelial malignancies islimited by drug resistance and toxic and side effects,which results in the failure in the treatment ofmajority of advanced cancer victims.To seek for anew,a...INTRODUCTIONThe treatment of human epithelial malignancies islimited by drug resistance and toxic and side effects,which results in the failure in the treatment ofmajority of advanced cancer victims.To seek for anew,and specific antineoplastic therapy willprovide hope for tumor treatment.Althoughdisordered intermediary metabolism in cancer cellshas been known for many years,much of the展开更多
Objective:Our aim was to test the hypothesis that fatty acid synthase(FASN)expression contributes to radioresistance of nasopharyngeal carcinoma(NPC)cells and that inhibiting FASN enhances radiosensitivity.Methods:Tar...Objective:Our aim was to test the hypothesis that fatty acid synthase(FASN)expression contributes to radioresistance of nasopharyngeal carcinoma(NPC)cells and that inhibiting FASN enhances radiosensitivity.Methods:Targeting FASN using epigallocatechin gallate(EGCG)or RNA interference in NPC cell lines that overexpress endogenous FASN was performed to determine their effects on cellular response to radiationin vitro using MTT and colony formation assays,andin vivo using xenograft animal models.Western blot,immunohistochemistry,real-time PCR arrays,and real-time RT-PCR were used to determine the relationship between FASN and frizzled class receptor 10(FZD10)expression.FZD10 knockdown and overexpression were used to determine its role in mediating FASN function in cellular response to radiation.Immunohistochemical staining was used to determine FASN and FZD10 expressions in human NPC tissues,followed by analysis of their association with the overall survival of patients.Results:FASN knockdown or inhibition significantly enhanced radiosensitivity of NPC cells,bothin vitro andin vivo.There was a positive association between FASN and FZD10 expression in NPC cell lines grown as monolayers or xenografts,as well as human tissues.FASN knockdown reduced FZD10 expression,and rescue of FZD10 expression abolished FASN knockdown-induced enhancement of radiosensitivity.FASN and FZD10 were both negatively associated with overall survival of NPC patients.Conclusions:FASN contributes to radioresistance,possiblyvia FZD10 in NPC cells.Both FZD10 and FASN expressions were associated with poor outcomes of NPC patients.EGCG may sensitize radioresistance by inhibiting FASN and may possibly be developed as a radiosensitizer for better treatment of NPCs.展开更多
The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that are critical for animal growth and development. Animals need to obtain BCAAs from their diet because they cannot syn...The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that are critical for animal growth and development. Animals need to obtain BCAAs from their diet because they cannot synthesize them. Plants are the ultimate source of these amino acids. Acetolactate synthase (ALS) is the first common enzyme in the biosynthesis of BCAAs. The metabolic control of BCAA biosynthesis involves allosteric regulation of ALS by the end-products of the pathway, i.e., valine, leucine and isoleucine. ALS holoenzyme seems to consist of two large catalytic subunits and two small regulatory subunits. In a previous study, using homologous recombination dependent gene targeting we created rice plants in which W548Land S627I mutations were induced into the endogenous gene encoding the ALS catalytic subunit. These two amino acid substitutions conferred hypertolerance to the ALS-inhibiting herbicide bispyripac-sodium. In this study, we revealed that feedback regulation by valine and leucine was reduced by these two amino acid substitutions. Furthermore, in leaves and seeds of ALS mutants with W548Land/or S627I substitution, a 2- to 3-fold increase in BCAAs was detected. Our results suggest that the ALS catalytic subunit is also involved in feedback regulation of ALS, and that judicious modification of the regulatory and catalytic subunits of ALS-coding genes by gene targeting can lead to the efficient accumulation of BCAA in plants.展开更多
Objective To explore the effects of zinc-α2-glycoprotein(ZAG) on body weight and body fat in high-fat-diet(HFD)-induced obesity in mice and the possible mechanism.Methods Thirty-six male mice were fed with standard f...Objective To explore the effects of zinc-α2-glycoprotein(ZAG) on body weight and body fat in high-fat-diet(HFD)-induced obesity in mice and the possible mechanism.Methods Thirty-six male mice were fed with standard food(SF)(n=9) and HFD(n=27),respectively.Five weeks later,9 mice fed with HFD were subjected to ZAG expression plasmid DNA transfection by liposome transfection method,and another 9 mice to negative control plasmid transfection.Two weeks later,serum ZAG level in the mice was assayed by Western blot,and the effects of ZAG over-expression on body weight,body fat,serum biochemical indexes,and adipose tissue of obese mice were evaluated.The mRNA expressions of fatty acid synthase(FAS) and hormone-sensitive lipase(HSL) in liver tissue were determined by reverse transcription-polymerase chain reaction.Results Serum ZAG level significantly lowered in simple HFD-fed mice in comparison to SF-fed mice(0.51±0.10 AU vs.0.75±0.07 AU,P<0.01).Further statistical analysis demonstrated that ZAG level was negatively correlated with body weight(r =-0.56,P<0.001),epididymal fat mass(r=-0.67,P<0.001),percentage of epididymal fat(r=-0.65,P<0.001),and increased weight(r=-0.57,P<0.001) in simple SF-and HFD-fed mice.ZAG over-expression in obese mice reduced body weight and the percentage of epididymal fat.Furthermore,FAS mRNA expression decreased(P<0.01) and HSL mRNA expression increased(P<0.001) in the liver in ZAG over-expressing mice.Conclusions ZAG is closely related to obesity.Serum ZAG level is inversely correlated with body weight and percentage of body fat.The action of ZAG is associated with reduced FAS expression and increased HSL expression in the liver of obese mice.展开更多
Objective:To determine fatty acid synthase(FAS)expression in human multiple myeloma and verify its potential as a therapeutic target in multiple myeloma.Methods:FAS expression was determined by immunohistochemistry,re...Objective:To determine fatty acid synthase(FAS)expression in human multiple myeloma and verify its potential as a therapeutic target in multiple myeloma.Methods:FAS expression was determined by immunohistochemistry,re- verse-transcription polymerase chain reaction(RT-PCR)and immunoblot analysis in bone marrow samples obtained from 27 patients with multiple myeloma(MM patients)and peripheral blood mononuclear cells(PBMCs)obtained from 12 healthy donors. In parallel,additional analyses were performed on 2 human multiple myeloma cell lines,U266 and RPMI8226.U266 cells were treated with cerulenin at various concentrations(5 to 320μg/ml)for 24 h,and metabolic activity was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assays.Apoptosis was evaluated by dual Annexin V/PI (propidium iodide)labeling and flow cytometry(FCM)in U266 cells treated with 20μg/ml cerulenin for 12 h or 24 h.Results:By immunohistochemistry,we found that 19 of 27 bone marrow samples obtained from MM patients expressed significantly high levels of FAS.Similarly,by RT-PCR,22 of 27 bone marrow samples obtained from MM patients,U266 and RPMI8226 showed FAS expression,whereas PBMC samples from 12 healthy donors did not express detectable level of FAS.FAS protein expression was confirmed by immunoblot analysis in 16 of 27 bone marrow samples obtained from MM patients,U266 and RPMI8226 cell lines,and no FAS protein expression was detected in PBMC samples from 12 healthy donors.U266 cells were highly sensitive to cerulenin treatment,with a dosage-related effect on metabolic activity,as a measure for cell proliferation.U266 cells treated with 20μg/ml cerulenin for 12 and 24 h also showed early sign of apoptosis with 56.9%and 69.3%Annexin V+/PI ?cells,and late apoptotic and necrotic cells with 3.2%and 17.6%Annexin V+/PI +cells.Conclusion:Increased FAS expression existed in multiple myeloma samples and human myeloma cell lines.Cerulenin greatly inhibited metabolic activity/cell proliferation of U266 cells and induced apoptosis,suggesting that FAS is an effective target for pharmacological therapy in human multiple myeloma.展开更多
BACKGROUND: Nitric oxide synthase (NOS) inhibrtors have been widely used to investigate the role of NO on cerebral ischemic injury, but the results are controversial. Moreover, it has been considered to aggravate the ...BACKGROUND: Nitric oxide synthase (NOS) inhibrtors have been widely used to investigate the role of NO on cerebral ischemic injury, but the results are controversial. Moreover, it has been considered to aggravate the ischemic neuronal damage with the release of excessively excitatory amino acids (EAA) during cerebral ischemia. On the other hand, some inhibitory amino acid is suggested to be important for the neuronal protection against ischemic brain damage. Our study has recently showed that treatment with the NOS inhibitor NG-nitro-L-arginine (L-NA) reduced focal cerebral ischemic damage. The effect of L-NA on the contents of excitatory and inhibitory amino acid in the rat brain following cerebral ischemia is still unclear.OBJECTIVE: By evaluating the effect of NOS inhibitor, L-NA on the contents of aspartate, glutamate, glycine and γ-aminobutyric acid (GABA) in striatum, hippocampus and cortex in the rat brain following cerebral ischemia respectively, to investigate the beneficial effect of L-NA on cerebral ischemic injury and the possible mechanism. DESIGN: A randomized and controlled experiment.SETTING: Department of Pharmacology, Hebei Academy of Medical Sciences.MATERIALS: A total of 42 male healthy SD rats (grade Ⅱ, weighting 250-300 g) were provided by the Experimental Animal Center of Hebei Province (Certification: 04036). Aspartate, glutamate, glycine, GABA, L-NA and 2,3,5-triphenyltetrazolium chloride (TTC) were obtained from Sigma Chemicals Co, St Louis, MO, USA. HPLC-ultraviolet detector system consisted of Agilent 1100 HPLC.METHODS: The experiment was carried out in Department of Pharmrcology, Hebei Academy of Medical Sciences from June 2005 to June 2006. Rats were randomly divided into three groups: sham-operated group (n = 6), ischemic group (n = 18), L-NA group (n = 18). The model of focal cerebral ischemia in rat was prepared with intraluminal line occlusion methods. In sham-operated rats, the external carotid artery was surgically prepared, but the filament was not inserted. Each group was further divided into 3 subgroups (n = 6 for each): drugs were administrated at 2, 6 and 12 hours after the middle cerebral artery occlusion (MCAO) respectively. L-NA (20 mg/kg, ip) was administrated, twice a day, for 3 consecutive days. Same volume of normal saline was administrated in ischemic and sham operation groups. The changes of infarcted volume and the contents of amino acids were respectively assayed. Image analysis software was used for the measurement of the infarcted area. The results were expressed as a percentage of the infarcted volume of cerebral/volume of whole brain (IV%) in order to control for edema formation. The contents of aspartate, glutamate, glycine and GABA in striatum, hippocampus and cortex in the rat brain following cerebral ischemia were respectively measured by HPLC method. All data were analyzed with one-way ANOVA and Dunnett’s test.MAIN OUTCOME MEASURES: ① The volume of cerebral infarction; ② The contents of aspartate, glutamate, glycine and GABA in brain tissue after cerebral ischemia.RESULTS: All 42 rats were involved in the final analysis. ① Infarcted volume: Volume was 0 in sham-operated group. When L-NA was administrated at 2 and 6 hours after MCAO, the infarcted volume was (20.13±3.59)% and (23.12±5.84)% in L-NA group, which was not similar to that in ischemic group [(22.10±3.98)%, (25.38±5.37)%, P > 0.05]. However, the infarcted volume was markedly decreased compared with that of ischemic group when L-NA was administrated at 12 hours after MCAO [(26.11±3.55)% and (37.15±3.58)%, P < 0.01]. ② Changes of amino acid content: At 2 and 6 hours after ischemia, the contents of aspartate, glutamate, glycine and GABA in striatum, hippocampus and cortex in ischemic group were significantly increased compared with those in sham-operated group ( P < 0.05-0.01). However, contents in L-NA group were similar to those in ischemic group (P > 0.05). At 12 hours after ischemia, the contents of aspartate [(0.21±0.06), (0.36±0.05), (0.29±0.12) mg/g] and glutamate [(0.55±0.06), (0.78±0.10), (0.52±0.10) mg/g] in striatum, hippocampus and cortex in L-NA group were significantly decreased compared with those in ischemic group [(0.49±0.17), (0.63±0.03), (0.51±0.15) mg/g; (0.98±0.30), (1.15±0.15), (0.93±0.15) mg/g, P < 0.05-0.01]. Glycine in hippocampus was (0.40±0.07) mg/g, which was higher than that in ischemic group [(0.21±0.07) mg/g, P < 0.05]. GABA in striatum, hippocampus and cortex was (0.93±0.10), (0.62±0.12) and (0.81±0.10) mg/g, respectively, which was higher than that in ischemic group [(0.60±0.08), (0.37±0.17), (0.59±0.10) mg/g, P < 0.05-0.01]. CONCLUSION: It may be concluded that L-NA have beneficial effect on ischemic cerebral injury in ischemic later stage in rats. The possible mechanism is that L-NA can decrease the contents of aspartate and glutamate, increase the contents of glycine and GABA.展开更多
Background: The liver is the corner stone in lipid metabolism, free fatty acid uptake, synthesizing, storing and exporting lipids;non-alcoholic fatty liver disease (NAFLD) develops if there is any interruption or dera...Background: The liver is the corner stone in lipid metabolism, free fatty acid uptake, synthesizing, storing and exporting lipids;non-alcoholic fatty liver disease (NAFLD) develops if there is any interruption or derangements in lipid metabolim. Fatty acid synthase (FAS) is the major enzyme in lipogenesis, and its circulating level is a bi-omarker of metabolically demanding human diseases. Aim of the Work: To evaluate the level of circulating FAS in NAFLD patients and to correlate it to serum lipid pa-rameters. Materials and Methods: The study included forty NAFLD patients and forty age and sex-matched healthy subjects as controls. Results: FAS levels were signifi-cantly higher in NAFLD patients compared to their level in the controls (P < 0.05). Ad-ditionally, a positive correlation was found between the levels of FAS and BMI (r = 0.57), and between FAS levels and triglycerides and low density lipoprotein cholesterol levels in NAFLD patients (r = 0.79 & 0.53, respectively). Conclusion: Elevated levels of circulating FAS can be considered as a biomarker of fatty liver disease.展开更多
To examine the role of glycogen synthase kinase 3(GSK-3) in the apoptosis of pancreatic β-cells to better understand the pathogenesis and to find new approach to the treatment of type 2 diabetes,apoptosis was induced...To examine the role of glycogen synthase kinase 3(GSK-3) in the apoptosis of pancreatic β-cells to better understand the pathogenesis and to find new approach to the treatment of type 2 diabetes,apoptosis was induced by oleic acid(OA) in INS-1 cells and the activity of GSK-3 was inhibited by LiCl.The PI staining and flow cytometry were employed for the evaluation of apoptosis.The phosphorylation level of GSK-3 was detected by Western blotting.The results showed that OA at 0.4 mmol/L could cause conspicuous apoptosis of INS-1 cells and the activity of GSK-3 was significantly increased.After the treatment with 24 mmol/L of LiCl,a inhibitor of GSK-3,the OA-induced apoptosis of INS-1 cells was lessened and the phosphorylation of GSK-3 was increased remarkably.It is concluded that GSK-3 activation plays an important role in OA-induced apoptosis in pancreatic β-cells and inhibition of the GSK-3 activity can effectively protect INS-1 cells from the OA-induced apoptosis.Our study provides a new experimental basis and target for the clinical treatment of type-2 diabetes.展开更多
BACKGROUND:Clinical practice and modern pharmacology have confirmed that chlorogenic acid can ameliorate learning and memory impairments. OBJECTIVE: To observe the effects of chlorogenic acid on neuronal nitric oxide ...BACKGROUND:Clinical practice and modern pharmacology have confirmed that chlorogenic acid can ameliorate learning and memory impairments. OBJECTIVE: To observe the effects of chlorogenic acid on neuronal nitric oxide synthase (nNOS)-positive neurons in the mouse hippocampus,and to investigate the mechanisms underlying the beneficial effects of chlorogenic acid on learning and memory. DESIGN,TIME AND SETTING: The present randomized,controlled,neural cell morphological observation was performed at the Institute of Neurobiology,Central South University between January and May 2005. MATERIALS: Forty-eight female,healthy,adult,Kunming mice were included in this study. Learning and memory impairment was induced with an injection of 0.5 μL kainic acid (0.4 mg/mL) into the hippocampus. METHODS: The mice were randomized into three groups (n=16): model,control,and chlorogenic acid-treated. At 2 days following learning and memory impairment induction,intragastric administration of physiological saline or chlorogenic acid was performed in the model and chlorogenic acid-treated groups,respectively. The control mice were administered 0.5 μL physiological saline into the hippocampus,and 2 days later,they received an intragastric administration of physiological saline. Each mouse received two intragastric administrations (1 mL solution once) per day,for a total of 35 days. MAIN OUTCOME MEASURES: Detection of changes in hippocampal and cerebral cortical nNOS neurons by immunohistochemistry; determination of spatial learning and memory utilizing the Y-maze device. RESULTS: At day 7 and 35 after intervention,there was no significant difference in the number of nNOS-positive neurons in the cerebral cortex between the model,chlorogenic acid,and control groups (P > 0.05). Compared with the control group,the number of nNOS-positive neurons in the hippocampal CA1-4 region was significantly less in the model group (P<0.05). However,the control group was not different from the chlorogenic acid-treated group (P > 0.05). At day 7 following intervention,the number of correct responses in the Y-maze test was greater in the chlorogenic acid-treated group than in the model group. CONCLUSION: Chlorogenic acid protects kainic acid-induced injury to nNOS-positive neurons in the hippocampal CA1-4 regions,thereby ameliorating learning and memory impairment.展开更多
Objective: Functional significance of NO and central inhibitory neurotransmitter γ-aminobutyric(GABA) during seizures were investigated morphorlogically. Methods: A kainate-induced complex partialseizure model was us...Objective: Functional significance of NO and central inhibitory neurotransmitter γ-aminobutyric(GABA) during seizures were investigated morphorlogically. Methods: A kainate-induced complex partialseizure model was used in our experiment. Twenty SD rats were randomly divided into KA 30, 60, 90, 200min and control groups. The brain sections were stained by NADPh (nicotinamide adenine dinucleotide phosphate ) diaphorase (Nd ) histochemically, and were further stained by GABA immunohistochemically.Results: Histological and immunohistochemical study revealed that in KA groups the number of Nd and GABA-positive double labelled neurons in CA3 region, CA3 region and dentate gyms was significantly reduced,compared with the control group. Conclusion: Nd coexisted with GABA in the brain. Reduction of GABA release led to relief of GABA-ergic inhibition and in the same way, reduction of NO release weakened its negative feedback modulation. Therefore neuronal synchronous paroxysmal discharges increased. GABA and NO,both having antiepileptic action, acted through different ways or different link in the same way. NO may involve in the effect of GABA-ergic neurons and play cooperative antiepileptic action with GABA.展开更多
基金the Applied Science Foundation of Jiangsu Province,China,No.BJ97071
文摘INTRODUCTIONThe treatment of human epithelial malignancies islimited by drug resistance and toxic and side effects,which results in the failure in the treatment ofmajority of advanced cancer victims.To seek for anew,and specific antineoplastic therapy willprovide hope for tumor treatment.Althoughdisordered intermediary metabolism in cancer cellshas been known for many years,much of the
基金This work was supported by grants from the National Natural Science Foundation of China(Grant Nos.81572588,81872147)Shantou University Medical College Clinical Trial Uplift Program(Grant No.201423)+4 种基金the Medical Scientific Research Foundation of Guangdong Province,China(Grant No.B2018222)the Traditional Chinese Medicine Research Project from Traditional Chinese Medicine Bureau of Guangdong Province(Grant No.20191182)the Youth Research Grant from Shantou University Medical College Cancer Hospital(Grant No.2018A001,2018A008)the key Project of Science and Technology of Shantou[Grant No.(2018)37]and the Natural Science Foundation of Guangdong Province of China(Grant No.2020A1515010094).
文摘Objective:Our aim was to test the hypothesis that fatty acid synthase(FASN)expression contributes to radioresistance of nasopharyngeal carcinoma(NPC)cells and that inhibiting FASN enhances radiosensitivity.Methods:Targeting FASN using epigallocatechin gallate(EGCG)or RNA interference in NPC cell lines that overexpress endogenous FASN was performed to determine their effects on cellular response to radiationin vitro using MTT and colony formation assays,andin vivo using xenograft animal models.Western blot,immunohistochemistry,real-time PCR arrays,and real-time RT-PCR were used to determine the relationship between FASN and frizzled class receptor 10(FZD10)expression.FZD10 knockdown and overexpression were used to determine its role in mediating FASN function in cellular response to radiation.Immunohistochemical staining was used to determine FASN and FZD10 expressions in human NPC tissues,followed by analysis of their association with the overall survival of patients.Results:FASN knockdown or inhibition significantly enhanced radiosensitivity of NPC cells,bothin vitro andin vivo.There was a positive association between FASN and FZD10 expression in NPC cell lines grown as monolayers or xenografts,as well as human tissues.FASN knockdown reduced FZD10 expression,and rescue of FZD10 expression abolished FASN knockdown-induced enhancement of radiosensitivity.FASN and FZD10 were both negatively associated with overall survival of NPC patients.Conclusions:FASN contributes to radioresistance,possiblyvia FZD10 in NPC cells.Both FZD10 and FASN expressions were associated with poor outcomes of NPC patients.EGCG may sensitize radioresistance by inhibiting FASN and may possibly be developed as a radiosensitizer for better treatment of NPCs.
文摘The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that are critical for animal growth and development. Animals need to obtain BCAAs from their diet because they cannot synthesize them. Plants are the ultimate source of these amino acids. Acetolactate synthase (ALS) is the first common enzyme in the biosynthesis of BCAAs. The metabolic control of BCAA biosynthesis involves allosteric regulation of ALS by the end-products of the pathway, i.e., valine, leucine and isoleucine. ALS holoenzyme seems to consist of two large catalytic subunits and two small regulatory subunits. In a previous study, using homologous recombination dependent gene targeting we created rice plants in which W548Land S627I mutations were induced into the endogenous gene encoding the ALS catalytic subunit. These two amino acid substitutions conferred hypertolerance to the ALS-inhibiting herbicide bispyripac-sodium. In this study, we revealed that feedback regulation by valine and leucine was reduced by these two amino acid substitutions. Furthermore, in leaves and seeds of ALS mutants with W548Land/or S627I substitution, a 2- to 3-fold increase in BCAAs was detected. Our results suggest that the ALS catalytic subunit is also involved in feedback regulation of ALS, and that judicious modification of the regulatory and catalytic subunits of ALS-coding genes by gene targeting can lead to the efficient accumulation of BCAA in plants.
基金Supported by the National Natural Science Foundation of China (30771026)Beijing Natural Science Foundation (7082079)
文摘Objective To explore the effects of zinc-α2-glycoprotein(ZAG) on body weight and body fat in high-fat-diet(HFD)-induced obesity in mice and the possible mechanism.Methods Thirty-six male mice were fed with standard food(SF)(n=9) and HFD(n=27),respectively.Five weeks later,9 mice fed with HFD were subjected to ZAG expression plasmid DNA transfection by liposome transfection method,and another 9 mice to negative control plasmid transfection.Two weeks later,serum ZAG level in the mice was assayed by Western blot,and the effects of ZAG over-expression on body weight,body fat,serum biochemical indexes,and adipose tissue of obese mice were evaluated.The mRNA expressions of fatty acid synthase(FAS) and hormone-sensitive lipase(HSL) in liver tissue were determined by reverse transcription-polymerase chain reaction.Results Serum ZAG level significantly lowered in simple HFD-fed mice in comparison to SF-fed mice(0.51±0.10 AU vs.0.75±0.07 AU,P<0.01).Further statistical analysis demonstrated that ZAG level was negatively correlated with body weight(r =-0.56,P<0.001),epididymal fat mass(r=-0.67,P<0.001),percentage of epididymal fat(r=-0.65,P<0.001),and increased weight(r=-0.57,P<0.001) in simple SF-and HFD-fed mice.ZAG over-expression in obese mice reduced body weight and the percentage of epididymal fat.Furthermore,FAS mRNA expression decreased(P<0.01) and HSL mRNA expression increased(P<0.001) in the liver in ZAG over-expressing mice.Conclusions ZAG is closely related to obesity.Serum ZAG level is inversely correlated with body weight and percentage of body fat.The action of ZAG is associated with reduced FAS expression and increased HSL expression in the liver of obese mice.
基金Project supported by the Medicine and Health Research Fund of Zhejiang Province(No.2007B091)the Office of Education of Zhejiang Province,China(No.20070104)
文摘Objective:To determine fatty acid synthase(FAS)expression in human multiple myeloma and verify its potential as a therapeutic target in multiple myeloma.Methods:FAS expression was determined by immunohistochemistry,re- verse-transcription polymerase chain reaction(RT-PCR)and immunoblot analysis in bone marrow samples obtained from 27 patients with multiple myeloma(MM patients)and peripheral blood mononuclear cells(PBMCs)obtained from 12 healthy donors. In parallel,additional analyses were performed on 2 human multiple myeloma cell lines,U266 and RPMI8226.U266 cells were treated with cerulenin at various concentrations(5 to 320μg/ml)for 24 h,and metabolic activity was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assays.Apoptosis was evaluated by dual Annexin V/PI (propidium iodide)labeling and flow cytometry(FCM)in U266 cells treated with 20μg/ml cerulenin for 12 h or 24 h.Results:By immunohistochemistry,we found that 19 of 27 bone marrow samples obtained from MM patients expressed significantly high levels of FAS.Similarly,by RT-PCR,22 of 27 bone marrow samples obtained from MM patients,U266 and RPMI8226 showed FAS expression,whereas PBMC samples from 12 healthy donors did not express detectable level of FAS.FAS protein expression was confirmed by immunoblot analysis in 16 of 27 bone marrow samples obtained from MM patients,U266 and RPMI8226 cell lines,and no FAS protein expression was detected in PBMC samples from 12 healthy donors.U266 cells were highly sensitive to cerulenin treatment,with a dosage-related effect on metabolic activity,as a measure for cell proliferation.U266 cells treated with 20μg/ml cerulenin for 12 and 24 h also showed early sign of apoptosis with 56.9%and 69.3%Annexin V+/PI ?cells,and late apoptotic and necrotic cells with 3.2%and 17.6%Annexin V+/PI +cells.Conclusion:Increased FAS expression existed in multiple myeloma samples and human myeloma cell lines.Cerulenin greatly inhibited metabolic activity/cell proliferation of U266 cells and induced apoptosis,suggesting that FAS is an effective target for pharmacological therapy in human multiple myeloma.
基金the Natural Sci-ence Foundation of HebeiProvince, No. C2005000840
文摘BACKGROUND: Nitric oxide synthase (NOS) inhibrtors have been widely used to investigate the role of NO on cerebral ischemic injury, but the results are controversial. Moreover, it has been considered to aggravate the ischemic neuronal damage with the release of excessively excitatory amino acids (EAA) during cerebral ischemia. On the other hand, some inhibitory amino acid is suggested to be important for the neuronal protection against ischemic brain damage. Our study has recently showed that treatment with the NOS inhibitor NG-nitro-L-arginine (L-NA) reduced focal cerebral ischemic damage. The effect of L-NA on the contents of excitatory and inhibitory amino acid in the rat brain following cerebral ischemia is still unclear.OBJECTIVE: By evaluating the effect of NOS inhibitor, L-NA on the contents of aspartate, glutamate, glycine and γ-aminobutyric acid (GABA) in striatum, hippocampus and cortex in the rat brain following cerebral ischemia respectively, to investigate the beneficial effect of L-NA on cerebral ischemic injury and the possible mechanism. DESIGN: A randomized and controlled experiment.SETTING: Department of Pharmacology, Hebei Academy of Medical Sciences.MATERIALS: A total of 42 male healthy SD rats (grade Ⅱ, weighting 250-300 g) were provided by the Experimental Animal Center of Hebei Province (Certification: 04036). Aspartate, glutamate, glycine, GABA, L-NA and 2,3,5-triphenyltetrazolium chloride (TTC) were obtained from Sigma Chemicals Co, St Louis, MO, USA. HPLC-ultraviolet detector system consisted of Agilent 1100 HPLC.METHODS: The experiment was carried out in Department of Pharmrcology, Hebei Academy of Medical Sciences from June 2005 to June 2006. Rats were randomly divided into three groups: sham-operated group (n = 6), ischemic group (n = 18), L-NA group (n = 18). The model of focal cerebral ischemia in rat was prepared with intraluminal line occlusion methods. In sham-operated rats, the external carotid artery was surgically prepared, but the filament was not inserted. Each group was further divided into 3 subgroups (n = 6 for each): drugs were administrated at 2, 6 and 12 hours after the middle cerebral artery occlusion (MCAO) respectively. L-NA (20 mg/kg, ip) was administrated, twice a day, for 3 consecutive days. Same volume of normal saline was administrated in ischemic and sham operation groups. The changes of infarcted volume and the contents of amino acids were respectively assayed. Image analysis software was used for the measurement of the infarcted area. The results were expressed as a percentage of the infarcted volume of cerebral/volume of whole brain (IV%) in order to control for edema formation. The contents of aspartate, glutamate, glycine and GABA in striatum, hippocampus and cortex in the rat brain following cerebral ischemia were respectively measured by HPLC method. All data were analyzed with one-way ANOVA and Dunnett’s test.MAIN OUTCOME MEASURES: ① The volume of cerebral infarction; ② The contents of aspartate, glutamate, glycine and GABA in brain tissue after cerebral ischemia.RESULTS: All 42 rats were involved in the final analysis. ① Infarcted volume: Volume was 0 in sham-operated group. When L-NA was administrated at 2 and 6 hours after MCAO, the infarcted volume was (20.13±3.59)% and (23.12±5.84)% in L-NA group, which was not similar to that in ischemic group [(22.10±3.98)%, (25.38±5.37)%, P > 0.05]. However, the infarcted volume was markedly decreased compared with that of ischemic group when L-NA was administrated at 12 hours after MCAO [(26.11±3.55)% and (37.15±3.58)%, P < 0.01]. ② Changes of amino acid content: At 2 and 6 hours after ischemia, the contents of aspartate, glutamate, glycine and GABA in striatum, hippocampus and cortex in ischemic group were significantly increased compared with those in sham-operated group ( P < 0.05-0.01). However, contents in L-NA group were similar to those in ischemic group (P > 0.05). At 12 hours after ischemia, the contents of aspartate [(0.21±0.06), (0.36±0.05), (0.29±0.12) mg/g] and glutamate [(0.55±0.06), (0.78±0.10), (0.52±0.10) mg/g] in striatum, hippocampus and cortex in L-NA group were significantly decreased compared with those in ischemic group [(0.49±0.17), (0.63±0.03), (0.51±0.15) mg/g; (0.98±0.30), (1.15±0.15), (0.93±0.15) mg/g, P < 0.05-0.01]. Glycine in hippocampus was (0.40±0.07) mg/g, which was higher than that in ischemic group [(0.21±0.07) mg/g, P < 0.05]. GABA in striatum, hippocampus and cortex was (0.93±0.10), (0.62±0.12) and (0.81±0.10) mg/g, respectively, which was higher than that in ischemic group [(0.60±0.08), (0.37±0.17), (0.59±0.10) mg/g, P < 0.05-0.01]. CONCLUSION: It may be concluded that L-NA have beneficial effect on ischemic cerebral injury in ischemic later stage in rats. The possible mechanism is that L-NA can decrease the contents of aspartate and glutamate, increase the contents of glycine and GABA.
文摘Background: The liver is the corner stone in lipid metabolism, free fatty acid uptake, synthesizing, storing and exporting lipids;non-alcoholic fatty liver disease (NAFLD) develops if there is any interruption or derangements in lipid metabolim. Fatty acid synthase (FAS) is the major enzyme in lipogenesis, and its circulating level is a bi-omarker of metabolically demanding human diseases. Aim of the Work: To evaluate the level of circulating FAS in NAFLD patients and to correlate it to serum lipid pa-rameters. Materials and Methods: The study included forty NAFLD patients and forty age and sex-matched healthy subjects as controls. Results: FAS levels were signifi-cantly higher in NAFLD patients compared to their level in the controls (P < 0.05). Ad-ditionally, a positive correlation was found between the levels of FAS and BMI (r = 0.57), and between FAS levels and triglycerides and low density lipoprotein cholesterol levels in NAFLD patients (r = 0.79 & 0.53, respectively). Conclusion: Elevated levels of circulating FAS can be considered as a biomarker of fatty liver disease.
基金This project was supported by a grant from National Natural Sciences Foundation of China (No 30672262)
文摘To examine the role of glycogen synthase kinase 3(GSK-3) in the apoptosis of pancreatic β-cells to better understand the pathogenesis and to find new approach to the treatment of type 2 diabetes,apoptosis was induced by oleic acid(OA) in INS-1 cells and the activity of GSK-3 was inhibited by LiCl.The PI staining and flow cytometry were employed for the evaluation of apoptosis.The phosphorylation level of GSK-3 was detected by Western blotting.The results showed that OA at 0.4 mmol/L could cause conspicuous apoptosis of INS-1 cells and the activity of GSK-3 was significantly increased.After the treatment with 24 mmol/L of LiCl,a inhibitor of GSK-3,the OA-induced apoptosis of INS-1 cells was lessened and the phosphorylation of GSK-3 was increased remarkably.It is concluded that GSK-3 activation plays an important role in OA-induced apoptosis in pancreatic β-cells and inhibition of the GSK-3 activity can effectively protect INS-1 cells from the OA-induced apoptosis.Our study provides a new experimental basis and target for the clinical treatment of type-2 diabetes.
文摘BACKGROUND:Clinical practice and modern pharmacology have confirmed that chlorogenic acid can ameliorate learning and memory impairments. OBJECTIVE: To observe the effects of chlorogenic acid on neuronal nitric oxide synthase (nNOS)-positive neurons in the mouse hippocampus,and to investigate the mechanisms underlying the beneficial effects of chlorogenic acid on learning and memory. DESIGN,TIME AND SETTING: The present randomized,controlled,neural cell morphological observation was performed at the Institute of Neurobiology,Central South University between January and May 2005. MATERIALS: Forty-eight female,healthy,adult,Kunming mice were included in this study. Learning and memory impairment was induced with an injection of 0.5 μL kainic acid (0.4 mg/mL) into the hippocampus. METHODS: The mice were randomized into three groups (n=16): model,control,and chlorogenic acid-treated. At 2 days following learning and memory impairment induction,intragastric administration of physiological saline or chlorogenic acid was performed in the model and chlorogenic acid-treated groups,respectively. The control mice were administered 0.5 μL physiological saline into the hippocampus,and 2 days later,they received an intragastric administration of physiological saline. Each mouse received two intragastric administrations (1 mL solution once) per day,for a total of 35 days. MAIN OUTCOME MEASURES: Detection of changes in hippocampal and cerebral cortical nNOS neurons by immunohistochemistry; determination of spatial learning and memory utilizing the Y-maze device. RESULTS: At day 7 and 35 after intervention,there was no significant difference in the number of nNOS-positive neurons in the cerebral cortex between the model,chlorogenic acid,and control groups (P > 0.05). Compared with the control group,the number of nNOS-positive neurons in the hippocampal CA1-4 region was significantly less in the model group (P<0.05). However,the control group was not different from the chlorogenic acid-treated group (P > 0.05). At day 7 following intervention,the number of correct responses in the Y-maze test was greater in the chlorogenic acid-treated group than in the model group. CONCLUSION: Chlorogenic acid protects kainic acid-induced injury to nNOS-positive neurons in the hippocampal CA1-4 regions,thereby ameliorating learning and memory impairment.
文摘Objective: Functional significance of NO and central inhibitory neurotransmitter γ-aminobutyric(GABA) during seizures were investigated morphorlogically. Methods: A kainate-induced complex partialseizure model was used in our experiment. Twenty SD rats were randomly divided into KA 30, 60, 90, 200min and control groups. The brain sections were stained by NADPh (nicotinamide adenine dinucleotide phosphate ) diaphorase (Nd ) histochemically, and were further stained by GABA immunohistochemically.Results: Histological and immunohistochemical study revealed that in KA groups the number of Nd and GABA-positive double labelled neurons in CA3 region, CA3 region and dentate gyms was significantly reduced,compared with the control group. Conclusion: Nd coexisted with GABA in the brain. Reduction of GABA release led to relief of GABA-ergic inhibition and in the same way, reduction of NO release weakened its negative feedback modulation. Therefore neuronal synchronous paroxysmal discharges increased. GABA and NO,both having antiepileptic action, acted through different ways or different link in the same way. NO may involve in the effect of GABA-ergic neurons and play cooperative antiepileptic action with GABA.