BACKGROUND:12-lipoxygenase(12-LOX) has been reported to be an important gene in cancer cell proliferation and survival,and tumor metastasis.However,its role in hepatocellular carcinoma(HCC) cells remains unknown.METHO...BACKGROUND:12-lipoxygenase(12-LOX) has been reported to be an important gene in cancer cell proliferation and survival,and tumor metastasis.However,its role in hepatocellular carcinoma(HCC) cells remains unknown.METHODS:Expression of 12-LOX was assessed in a diethylnitrosamine-induced rat HCC model,and in SMMC-7721,HepG2 and L-02 cells using immunohistochemical staining and reverse transcriptase-polymerase chain reaction(RT-PCR).GST-π and Ki-67 were determined in vivo by immunohistochemical staining.Apoptosis was evaluated by TUNEL assay.Cell viability and apoptosis were determined by MTT assay and flow cytometry,respectively.Apoptosis-related proteins in SMMC-7721 and HepG2 cells were detected by Western blotting.RESULTS:Immunohistochemical staining and RT-PCR showed that 12-LOX was over-expressed in rat HCC and two HCC cell lines,while the expression was inhibited by baicalein,a specific inhibitor of 12-LOX.Baicalein inhibited cell proliferation and induced apoptosis in rat HCC and both cell lines in a dose-and time-dependent manner.Our in vivo study demonstrated that baicalein also reduced neoplastic nodules.Mechanistically,baicalein reduced Bcl-2 protein expression coupled with a slight increase of the expression of Bax and activation of caspase-3.Furthermore,baicalein inhibited the activation of ERK-1/2(phosphorylated).Interestingly,the effects of baicalein were reversed by 12(S)-HETE,a metabolite of 12-LOX.CONCLUSIONS:Inhibition of 12-LOX leads to reduced numbers of HCC cells,partially caused by increased apoptosis.12-LOX may be a potential molecular target for HCC prevention and treatment.展开更多
The interplay between inflammation and cancer progression is a growing area of research. A combination of clinical, epidemiological, and basic science investigations indicate that there is a relationship between infla...The interplay between inflammation and cancer progression is a growing area of research. A combination of clinical, epidemiological, and basic science investigations indicate that there is a relationship between inflammatory changes in the pancreas and neoplastic progression. Diets high in ω-6 polyunsaturated fatty acids provide increased substrate for arachidonic acid metabolism by cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) to form eicosanoids. These eicosanoids directly contribute to pancreatic cancer cell proliferation. Both COX-2 and 5-LOX are upregulated in multiple cancer types, including pancreatic cancer. In vitro studies using pancreatic cancer cell lines have demonstrated upregulation of COX-2 and 5-LOX at both the mRNA and protein levels. When COX-2 and 5-LOX are blocked via a variety of mechanisms, cancer cell proliferation is abrogated both in vitro and in vivo. The mechanism of COX-2 has been shown to include effects on apoptosis as well as angiogenesis. 5-LOX has been implicated in apoptosis. The use of COX-2 and 5-LOX inhibitors in clinical studies in patients with pancreatic cancer has been limited. Patient enrollment has been restricted to those with advanced disease which makes evaluation of these drugs as chemopreventive agents difficult. COX-2 and 5-LOX expression have been shown to be present during the early neoplastic changes of pancreatic cancer, well before progression to invasive disease. This indicates that the ideal role for these interventions is early in the disease process as preventive agents, perhaps in patients with chronic pancreatitis or hereditary pancreatitis.展开更多
AIM: To evaluate the 5-lipoxygenases (Loxs) expression level in human colorectal cancer specimens in order to determine its clinicopathologic significance in human tumorigenesis. METHODS: The relative quantity of 5-Lo...AIM: To evaluate the 5-lipoxygenases (Loxs) expression level in human colorectal cancer specimens in order to determine its clinicopathologic significance in human tumorigenesis. METHODS: The relative quantity of 5-Lox mRNA in paired 91 colorectal tumor and adjacent normal mucosa samples was determined by real time quantitative PCR. Additionally, the expression of 5-Lox and cyclooxygenase (Cox)-2 proteins was also examined using immunohistochemical staining methods. RESULTS: There was a marked increase in 5-Lox mRNA levels in the tumor compared with paired normal mucosa samples (P < 0.0001). Sixty six (72.5%) tumors showed high 5-Lox mRNA levels. The positivity rate of 5-Lox and Cox-2 protein expression was 68.7% and 79.1% respectively. There was a significant association between tumoral 5-Lox mRNA level and tumor size (Rho = 0.392, P = 0.0002), depth or vessel invasion. CONCLUSION: These results suggest that 5-Lox is up-regulated in colorectal cancer and that inhibition of its expression might be valuable in the prevention and treatment of colorectal cancer.展开更多
OBJECTIVE Previously we demonstrated the neuroprotective effect of 5-lipoxygenase(5-LOX)inhibitor as well as cysteinyl leukotriene receptor 1(Cys LT1)antagoniston rotenone-induced microglial activation and neuronal de...OBJECTIVE Previously we demonstrated the neuroprotective effect of 5-lipoxygenase(5-LOX)inhibitor as well as cysteinyl leukotriene receptor 1(Cys LT1)antagoniston rotenone-induced microglial activation and neuronal death.In this study,we determined the effects of 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast on neurotoxicity induced by 1-methyl-4-phenylpyridine(MPP+)in an in vitro model of Parkinson disease(PD).METHODS The neurotoxicity of MPP+,a neurotoxin relevant to PD,on the PC12 cells was measured by MTT assay,lactate dehydrogenase(LDH)release and double fluorescence staining with Hoechst/propidiumiodide(PI).The protective effects of 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast were investigated by the above methods.RESULTS We found that exposure of PC12 cells to MPP+led to a reduced cell viability and an increased level of LDH in a concentration-dependent manner.Pretreatment with zileuton and montelukast significantly attenuated viability loss and LDH release in MPP+-treated PC12 cells.Furthermore,MPP+increasednecrotic cell death in PC12 cells.Administration of montelukast significantly decreased MPP+-induced cell necrosis in PC12 cells.CONCLUSION The 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast have a neuroprotective effects on MPP+-induced neurotoxicity in PC12 cells.The 5-LOX inhibitor and Cys LT1 antagonist might raise a possibility as potential therapeutic agent for PD and other inflammation-related the central nervous system disorders.展开更多
目的探讨12/15-脂氧合酶抑制剂黄芩素(Baicalein)对硝普钠(SNP)诱导的软骨细胞凋亡的抑制作用及可能机制。方法取8周雄性SD大鼠膝关节软骨,采用Ⅱ型胶原酶消化法提取软骨细胞并体外培养。设置对照组、SNP凋亡组和Baicalein给药组,以0.5 ...目的探讨12/15-脂氧合酶抑制剂黄芩素(Baicalein)对硝普钠(SNP)诱导的软骨细胞凋亡的抑制作用及可能机制。方法取8周雄性SD大鼠膝关节软骨,采用Ⅱ型胶原酶消化法提取软骨细胞并体外培养。设置对照组、SNP凋亡组和Baicalein给药组,以0.5 m M SNP作用24小时诱导软骨细胞凋亡,以5 M,25 M,50 M,100 M不同浓度的Baicalein作用细胞,确定最适浓度,对照组仅加入同体积溶剂(蒸馏水)。CCK8法检测药物对细胞毒性;Annexin V/PI双染法检测细胞凋亡;免疫荧光观测凋亡诱导因子(AIF)在软骨细胞中的定位。结果 Baicalein在各浓度下对软骨细胞无明显毒性,与对照组相比,凋亡组软骨细胞活性明显下降(<0.01),与凋亡组相比,给药组细胞活性浓度依赖性地升高(<0.01);流式检测显示对照组软骨细胞早期凋亡率3.16%,凋亡组27.8%,100 M Baicalein给药组14.1%;免疫荧光检测显示,凋亡组AIF出现核内移位,100 M Baicalein给药组AIF核移位受到抑制。结论 Baicalein通过抑制12/15-脂氧合酶活性,进而抑制AIF从线粒体中的释放及核转位,发挥抗软骨细胞凋亡作用。展开更多
Objective:To determine the pharmacodynamic material basis and mechanism of Danbei Yifei formula on pulmonary fibrosis.Methods:Starting with the clear absorbed components of Danbei Yifei formula or the potential effect...Objective:To determine the pharmacodynamic material basis and mechanism of Danbei Yifei formula on pulmonary fibrosis.Methods:Starting with the clear absorbed components of Danbei Yifei formula or the potential effective components in line with the five rules of Ribinsky,the network pharmacology method and technology of traditional Chinese medicine were used to predict and analyze the action targets of Danbei Yifei formula in vivo,such as Salvia miltiorrhiza,PINBEI,Taoren,etc.On the basis of enrichment analysis,the core pathway of Danbei Yifei formula in the treatment of pulmonary fibrosis was identified,and the binding energy of drug ligand and protein target was determined through molecular docking technology simulation and verification,and its affinity and stability were evaluated.To clarify the material basis and mechanism of Danbei Yifei formula in the treatment of pulmonary fibrosis.Result:The results of network pharmacology prediction of traditional Chinese medicine showed that Danbei Yifei formula contained 72 potential pharmacodynamic components and 26 corresponding targets,including CHRM1、MAPK14、CCL2、ADRB1、PTGS1、PPARG、ALOX5、Pde3a、CHRM2、Adrb2、TNF、JUN、Adora2a、LTA4H、CYP1A2、OPRD1、CHRM3、DRD2、OPRM1、ARG1、EDNRA、Il6st、TACR1、MMP1、MMP8、Ptgs2,which were related to pulmonary fibrosis and pulmonary fibrosis Lung related diseases are highly correlated.There were 26 Go items(P<0.05)in go functional enrichment analysis,including 22 biological process(BP),9 cellular component(CC)and 3 molecular function(MF)categories.The results of network pharmacology showed that many components,such as protocatechuic acid and aminosuccinic acid,had direct effects on known targets of pulmonary fibrosis.Conclusion:Danbei Yifei formula contains many effective components which have inhibitory effect on pulmonary fibrosis,and it may play its role through the mechanism of multi-component and multi-target synergistic effect.展开更多
Recovery of functional beta cell mass offers a biological cure for type 1 diabetes. However, beta cell mass is difficult to regain once lost since the proliferation rate of beta cells after youth is very low. Angiopoi...Recovery of functional beta cell mass offers a biological cure for type 1 diabetes. However, beta cell mass is difficult to regain once lost since the proliferation rate of beta cells after youth is very low. Angiopoietin like-protein 8 (ANGPTL8), a peptide that has a role in the regulation of lipoprotein lipase activity, was reported to increase beta cell proliferation in mice in 2013. Subsequent studies of human ANGPTL8 for short term (3 to 8 days) in non-diabetic mice showed little or no increase in beta cell proliferation. Here, we examined the effect of ANGPTL8 on glucose homeostasis in models that have not been examined previously. We expressed mouse ANGPTL8 using adenovirus in 2 mouse models of diabetes (streptozotocin and Non-Obese Diabetic (NOD) mice) over 2 weeks. Also, we tested ANGPTL8 in NOD mice deficient in leukocyte 12-lipoxygenase (12LO), an enzyme that contributes to insulitis and loss of beta cell function in NOD, in an effort to determine whether 12LO deficiency alters the response to ANGPTL8. Adenovirus-mediated expression of ANGPTL8 lowered blood glucose levels in streptozotocin treated mice without an increase in beta cell proliferation or serum insulin concentration. While ANGPTL8 did not reverse hyperglycemia in overtly hyperglycemic NOD mice or alter glucose homeostasis of non-diabetic NOD mice, ANGPTL8 reduced blood glucose levels in 12LOKO NOD mice. However, the lower glucose levels in 12LOKO NOD were not associated with higher serum insulin levels or beta cell proliferation. In summary, while mouse ANGPTL8 does not increase beta cell proliferation in NOD mice or streptozotocin treated mice in agreement with studies in non-diabetic mice, it lowers blood glucose levels in multiple low-dose streptozotocin induced diabetes and 12LO deficiency indicating that host factors influence the impact of ANGPTL8 on glucose homeostasis.展开更多
基金supported by grants from the National Natural Science Foundation of China(81000998)the Natural Science Foundation of Hubei Province,China(2007ABA248)the Foundation of the Ministry of Education of China for New Teachers(20090141120003)
文摘BACKGROUND:12-lipoxygenase(12-LOX) has been reported to be an important gene in cancer cell proliferation and survival,and tumor metastasis.However,its role in hepatocellular carcinoma(HCC) cells remains unknown.METHODS:Expression of 12-LOX was assessed in a diethylnitrosamine-induced rat HCC model,and in SMMC-7721,HepG2 and L-02 cells using immunohistochemical staining and reverse transcriptase-polymerase chain reaction(RT-PCR).GST-π and Ki-67 were determined in vivo by immunohistochemical staining.Apoptosis was evaluated by TUNEL assay.Cell viability and apoptosis were determined by MTT assay and flow cytometry,respectively.Apoptosis-related proteins in SMMC-7721 and HepG2 cells were detected by Western blotting.RESULTS:Immunohistochemical staining and RT-PCR showed that 12-LOX was over-expressed in rat HCC and two HCC cell lines,while the expression was inhibited by baicalein,a specific inhibitor of 12-LOX.Baicalein inhibited cell proliferation and induced apoptosis in rat HCC and both cell lines in a dose-and time-dependent manner.Our in vivo study demonstrated that baicalein also reduced neoplastic nodules.Mechanistically,baicalein reduced Bcl-2 protein expression coupled with a slight increase of the expression of Bax and activation of caspase-3.Furthermore,baicalein inhibited the activation of ERK-1/2(phosphorylated).Interestingly,the effects of baicalein were reversed by 12(S)-HETE,a metabolite of 12-LOX.CONCLUSIONS:Inhibition of 12-LOX leads to reduced numbers of HCC cells,partially caused by increased apoptosis.12-LOX may be a potential molecular target for HCC prevention and treatment.
文摘The interplay between inflammation and cancer progression is a growing area of research. A combination of clinical, epidemiological, and basic science investigations indicate that there is a relationship between inflammatory changes in the pancreas and neoplastic progression. Diets high in ω-6 polyunsaturated fatty acids provide increased substrate for arachidonic acid metabolism by cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) to form eicosanoids. These eicosanoids directly contribute to pancreatic cancer cell proliferation. Both COX-2 and 5-LOX are upregulated in multiple cancer types, including pancreatic cancer. In vitro studies using pancreatic cancer cell lines have demonstrated upregulation of COX-2 and 5-LOX at both the mRNA and protein levels. When COX-2 and 5-LOX are blocked via a variety of mechanisms, cancer cell proliferation is abrogated both in vitro and in vivo. The mechanism of COX-2 has been shown to include effects on apoptosis as well as angiogenesis. 5-LOX has been implicated in apoptosis. The use of COX-2 and 5-LOX inhibitors in clinical studies in patients with pancreatic cancer has been limited. Patient enrollment has been restricted to those with advanced disease which makes evaluation of these drugs as chemopreventive agents difficult. COX-2 and 5-LOX expression have been shown to be present during the early neoplastic changes of pancreatic cancer, well before progression to invasive disease. This indicates that the ideal role for these interventions is early in the disease process as preventive agents, perhaps in patients with chronic pancreatitis or hereditary pancreatitis.
文摘AIM: To evaluate the 5-lipoxygenases (Loxs) expression level in human colorectal cancer specimens in order to determine its clinicopathologic significance in human tumorigenesis. METHODS: The relative quantity of 5-Lox mRNA in paired 91 colorectal tumor and adjacent normal mucosa samples was determined by real time quantitative PCR. Additionally, the expression of 5-Lox and cyclooxygenase (Cox)-2 proteins was also examined using immunohistochemical staining methods. RESULTS: There was a marked increase in 5-Lox mRNA levels in the tumor compared with paired normal mucosa samples (P < 0.0001). Sixty six (72.5%) tumors showed high 5-Lox mRNA levels. The positivity rate of 5-Lox and Cox-2 protein expression was 68.7% and 79.1% respectively. There was a significant association between tumoral 5-Lox mRNA level and tumor size (Rho = 0.392, P = 0.0002), depth or vessel invasion. CONCLUSION: These results suggest that 5-Lox is up-regulated in colorectal cancer and that inhibition of its expression might be valuable in the prevention and treatment of colorectal cancer.
基金The project supported National Natural Science Foundation of China(81273491)the Zhejiang Provincial Natural Science Foundation(LY12H31010)
文摘OBJECTIVE Previously we demonstrated the neuroprotective effect of 5-lipoxygenase(5-LOX)inhibitor as well as cysteinyl leukotriene receptor 1(Cys LT1)antagoniston rotenone-induced microglial activation and neuronal death.In this study,we determined the effects of 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast on neurotoxicity induced by 1-methyl-4-phenylpyridine(MPP+)in an in vitro model of Parkinson disease(PD).METHODS The neurotoxicity of MPP+,a neurotoxin relevant to PD,on the PC12 cells was measured by MTT assay,lactate dehydrogenase(LDH)release and double fluorescence staining with Hoechst/propidiumiodide(PI).The protective effects of 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast were investigated by the above methods.RESULTS We found that exposure of PC12 cells to MPP+led to a reduced cell viability and an increased level of LDH in a concentration-dependent manner.Pretreatment with zileuton and montelukast significantly attenuated viability loss and LDH release in MPP+-treated PC12 cells.Furthermore,MPP+increasednecrotic cell death in PC12 cells.Administration of montelukast significantly decreased MPP+-induced cell necrosis in PC12 cells.CONCLUSION The 5-LOX inhibitor zileuton and Cys LT1 antagonist montelukast have a neuroprotective effects on MPP+-induced neurotoxicity in PC12 cells.The 5-LOX inhibitor and Cys LT1 antagonist might raise a possibility as potential therapeutic agent for PD and other inflammation-related the central nervous system disorders.
文摘目的探讨12/15-脂氧合酶抑制剂黄芩素(Baicalein)对硝普钠(SNP)诱导的软骨细胞凋亡的抑制作用及可能机制。方法取8周雄性SD大鼠膝关节软骨,采用Ⅱ型胶原酶消化法提取软骨细胞并体外培养。设置对照组、SNP凋亡组和Baicalein给药组,以0.5 m M SNP作用24小时诱导软骨细胞凋亡,以5 M,25 M,50 M,100 M不同浓度的Baicalein作用细胞,确定最适浓度,对照组仅加入同体积溶剂(蒸馏水)。CCK8法检测药物对细胞毒性;Annexin V/PI双染法检测细胞凋亡;免疫荧光观测凋亡诱导因子(AIF)在软骨细胞中的定位。结果 Baicalein在各浓度下对软骨细胞无明显毒性,与对照组相比,凋亡组软骨细胞活性明显下降(<0.01),与凋亡组相比,给药组细胞活性浓度依赖性地升高(<0.01);流式检测显示对照组软骨细胞早期凋亡率3.16%,凋亡组27.8%,100 M Baicalein给药组14.1%;免疫荧光检测显示,凋亡组AIF出现核内移位,100 M Baicalein给药组AIF核移位受到抑制。结论 Baicalein通过抑制12/15-脂氧合酶活性,进而抑制AIF从线粒体中的释放及核转位,发挥抗软骨细胞凋亡作用。
基金Applied Technology Research and Development Project of Heilongjiang Province(No.GA19C1080)。
文摘Objective:To determine the pharmacodynamic material basis and mechanism of Danbei Yifei formula on pulmonary fibrosis.Methods:Starting with the clear absorbed components of Danbei Yifei formula or the potential effective components in line with the five rules of Ribinsky,the network pharmacology method and technology of traditional Chinese medicine were used to predict and analyze the action targets of Danbei Yifei formula in vivo,such as Salvia miltiorrhiza,PINBEI,Taoren,etc.On the basis of enrichment analysis,the core pathway of Danbei Yifei formula in the treatment of pulmonary fibrosis was identified,and the binding energy of drug ligand and protein target was determined through molecular docking technology simulation and verification,and its affinity and stability were evaluated.To clarify the material basis and mechanism of Danbei Yifei formula in the treatment of pulmonary fibrosis.Result:The results of network pharmacology prediction of traditional Chinese medicine showed that Danbei Yifei formula contained 72 potential pharmacodynamic components and 26 corresponding targets,including CHRM1、MAPK14、CCL2、ADRB1、PTGS1、PPARG、ALOX5、Pde3a、CHRM2、Adrb2、TNF、JUN、Adora2a、LTA4H、CYP1A2、OPRD1、CHRM3、DRD2、OPRM1、ARG1、EDNRA、Il6st、TACR1、MMP1、MMP8、Ptgs2,which were related to pulmonary fibrosis and pulmonary fibrosis Lung related diseases are highly correlated.There were 26 Go items(P<0.05)in go functional enrichment analysis,including 22 biological process(BP),9 cellular component(CC)and 3 molecular function(MF)categories.The results of network pharmacology showed that many components,such as protocatechuic acid and aminosuccinic acid,had direct effects on known targets of pulmonary fibrosis.Conclusion:Danbei Yifei formula contains many effective components which have inhibitory effect on pulmonary fibrosis,and it may play its role through the mechanism of multi-component and multi-target synergistic effect.
文摘Recovery of functional beta cell mass offers a biological cure for type 1 diabetes. However, beta cell mass is difficult to regain once lost since the proliferation rate of beta cells after youth is very low. Angiopoietin like-protein 8 (ANGPTL8), a peptide that has a role in the regulation of lipoprotein lipase activity, was reported to increase beta cell proliferation in mice in 2013. Subsequent studies of human ANGPTL8 for short term (3 to 8 days) in non-diabetic mice showed little or no increase in beta cell proliferation. Here, we examined the effect of ANGPTL8 on glucose homeostasis in models that have not been examined previously. We expressed mouse ANGPTL8 using adenovirus in 2 mouse models of diabetes (streptozotocin and Non-Obese Diabetic (NOD) mice) over 2 weeks. Also, we tested ANGPTL8 in NOD mice deficient in leukocyte 12-lipoxygenase (12LO), an enzyme that contributes to insulitis and loss of beta cell function in NOD, in an effort to determine whether 12LO deficiency alters the response to ANGPTL8. Adenovirus-mediated expression of ANGPTL8 lowered blood glucose levels in streptozotocin treated mice without an increase in beta cell proliferation or serum insulin concentration. While ANGPTL8 did not reverse hyperglycemia in overtly hyperglycemic NOD mice or alter glucose homeostasis of non-diabetic NOD mice, ANGPTL8 reduced blood glucose levels in 12LOKO NOD mice. However, the lower glucose levels in 12LOKO NOD were not associated with higher serum insulin levels or beta cell proliferation. In summary, while mouse ANGPTL8 does not increase beta cell proliferation in NOD mice or streptozotocin treated mice in agreement with studies in non-diabetic mice, it lowers blood glucose levels in multiple low-dose streptozotocin induced diabetes and 12LO deficiency indicating that host factors influence the impact of ANGPTL8 on glucose homeostasis.
基金国家自然科学青年基金（81000300）国家自然科学基金（81170669）+1 种基金国家教育部博士点基金（2010006120024）吉林大学白求恩B计划项目.志谢 感谢美国Beckman Research Institute of City of Hope糖尿病研究中心Dr.N.Rama友情提供的稳定高表达12-LO基因的MC（pcDNA-12-LO）及对照组MC（pcDNA）
