1.Establishment and validation of a dose-response curve for semi-automatic analysis of dicentric chromosomes
Qianqian MENG ; Zhongxin ZHANG ; Yue REN ; Xiaozhen LI ; Zeyu MIAO ; Chao WANG ; Hongyan LIU ; Ruifeng ZHANG
China Occupational Medicine 2024;51(1):70-74
ObjectiveTo establish a dose-effect curve for semi-automatic analysis of dicentric chromosomes(DC) based on an automatic chromosome analysis system. Methods A total of three healthy volunteers were recruited as the study subjects, and their peripheral blood was collected and stimulated by X-ray at doses of 0.00, 0.10, 0.25, 0.50, 0.75, 1.00, 2.00, 3.00, 4.00, and 5.00 Gy, with the absorbed dose rate of 1.0 Gy/min. Images of DC in the mid-stage of cell division were collected using a high-throughput automatic chromosome analysis system. The DCScore software was used to automatically analyze DC aberrations, and a dose-effect curve for semi-automatic analysis of DC was fitted after manual confirmation. The fitted dose-effect curve for semi-automatic analysis of DC was validated for accuracy using three proficiency test samples from the national quality assessment of biological dose. Results The incidence of DC increased with increasing irradiation doses in the range of 0.00-5.00 Gy (P<0.01). The dose-effect curve for the fitted semi-automatic analysis of DC was ŷ =0.000 8 (±0.000 2) +0.009 2(±0.000 9) D+0.014 2(±0.000 4) D2 (R2= 0.999 8). The relative deviation between the estimated dose and the actual dose of the three test samples was about 20.00%, indicating curve applicability for biological dose estimation. Moreover, excluding the time spent on manual analysis, the semi-automatic analysis method increased the analysis efficiency by 26.0 times. Conclusion The semi-automatic analysis dose-effect curve for DC stimulated by X-ray is constructed for biological dose estimation, which can reduce the manual analysis time, and holds great potential for application in nuclear emergency response to large-scale radiation accidents.
2.Apigenin Induces Apoptosis of Human Colorectal Cancer CL187 Cells via PI3K/Akt and MAPK Signaling Pathways
Si LIN ; Huizhen QIN ; Lingyu DENG ; Zeyu LI ; Fengfeng XIE ; Miao ZHANG ; Hua ZHU
Chinese Journal of Experimental Traditional Medical Formulae 2022;28(19):97-104
ObjectiveTo study the effect of apigenin on the proliferation and apoptosis of human colon cancer CL187 cells and the underlying mechanisms. MethodHuman colorectal cancer CL187 cells were treated with different concentrations of apigenin (0, 30, 45, 60 mg·L-1) according to the results of the preliminary experiment. The proliferation of CL187 cells was detected by methyl thiazolyl tetrazolium (MTT) and colony formation assays, and the apoptosis was observed via Hoechst 33258 staining. Real-time fluorescence quantitative PCR was conducted to determine the mRNA levels of cysteine protease-3 (Caspase-3), B-cell lymphoma-2 (Bcl-2), and Bcl-2-associated X protein (Bax) in the CL187 cells treated with apigenin. Western blot was employed to measure the protein levels of Caspase-3, Bcl-2, and Bax associated with apoptosis, protein kinase B (Akt) and phosphorylated Akt (p-Akt) in phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway, and extracellular signal-regulated kinases 1/2 (ERK1/2), p-ERK1/2, c-Jun N-terminal kinase (JNK), p-JNK, p38 mitogen-activated protein kinase (MAPK), and p-p38 MAPK protein in MAPK pathway. ResultCompared with the blank group, the apigenin groups had low cell survival rates and high inhibition rates on cell proliferation (P<0.01). Apigenin decreased the cell clone number and clone formation rate, and increased the inhibition rate on clone formation (P<0.01). After CL187 cells were treated with different concentrations of apigenin for 48 h, typical apoptosis characteristics such as nuclear pyknosis, chromatin condensation, and enhanced fluorescence reaction were observed. Compared with blank group, 45, 60 mg·L-1 apigenin treatments down-regulated the mRNA level of anti-apoptotic gene Bcl-2 (P<0.01) and all the apigenin treatments up-regulated those of the pro-apoptotic genes Bax and Caspase-3 (P<0.05, P<0.01). Similarly, apigenin treatments down-regulated the protein level of Bcl-2 (P<0.05, P<0.01) and up-regulated those of Caspase-3 (P<0.05, P<0.01) and Bax (P<0.01, 45, 60 mg·L-1). The blank group had higher protein level of Akt than the 60 mg·L-1 apigenin group (P<0.01), higher protein levels of p-Akt, ERK1/2, and p-ERK1/2 than the 45, 60 mg·L-1 apigenin groups (P<0.01), and higher protein levels of JNK and p-JNK than the apigenin groups (P<0.05, P<0.01). Compared with blank group, 60 mg·L-1 apigenin up-regulated the protein level of p38 MAPK (P<0.05), and all the apigenin groups up-regulated that of p-p38 MAPK (P<0.01). Furthermore, apigenin lowered the p-Akt/Akt ratio (P<0.05, P<0.01) and p-ERK1/2/ERK1/2 ratio (P<0.01), while it increased the p-JNK/JNK ratio (45, 60 mg·L-1; P<0.05, P<0.01) and p-p38 MAPK/p38 MAPK ratio (P<0.05, P<0.01). ConclusionApigenin can inhibit the proliferation and promote the apoptosis of CL187 cells by inhibiting the PI3K/Akt signaling pathway and regulating the expression of proteins in the MAPK signaling pathway.
3.Association study of LIS1 and TSNAX genes with bipolar disorder in Chinese Han population.
Xuan LI ; Lijie GUAN ; Yin LIN ; Xiaofei ZHANG ; Wenhao DENG ; Zhenxing YANG ; Xiaohong MA ; Guohui LAO ; Biyu YE ; Weijie HUANG ; Zeyu JIANG ; Guodong MIAO ; Guiyun XU ; Wentao LIU ; Yingcheng WANG ; Tao LI ; Liping CAO
Chinese Journal of Medical Genetics 2014;31(3):357-361
OBJECTIVETo assess the association of neural development-related genes LIS1and TSNAX with bipolar disorder in a Chinese Han population.
METHODSThree hundred and eight five patients (including 188 males and 197 females) from Guangzhou Brain Hospital with bipolar disorder meeting the Diagnostic and Statistic Manual of Bipolar Disorder (BDI) (Fourth Edition) criteria and 475 healthy controls from the local community were recruited. Ten single nucleotide polymorphisms (SNPs) of the LIS1 and TSNAX genes were genotyped by GoldenGate genotyping assay on an Illumina Beadstation 500 machine. Association analyses of SNPs and haplotypes were performed with Plink 1.07 software.
RESULTSAnalysis of the total sample has failed to find any association of SNP or haplotype of the two genes with BDI (P> 0.05). When patients were divided into subgroups with or without psychotic symptom, no significant association of the two genes was found with psychotic BDI or non-psychotic BDI (P> 0.05). No significant association was found between any SNP and haplotype of two genes and female BDI or male BDI, nor were significant association found between age of onset and LIS1 and TSNAX gene polymorphisms.
CONCLUSIONOur results indicated that LIS1 and TSNAX genes are not associated with susceptibility to bipolar I disorder in Chinese Han population.
1-Alkyl-2-acetylglycerophosphocholine Esterase ; genetics ; Adolescent ; Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; ethnology ; genetics ; Bipolar Disorder ; ethnology ; genetics ; Case-Control Studies ; DNA-Binding Proteins ; genetics ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Microtubule-Associated Proteins ; genetics ; Middle Aged ; Polymorphism, Single Nucleotide ; Young Adult
4.Research progress of pharmacological action and mechanism of piperine
Si LIN ; Huizhen QIN ; Lingyu DENG ; Miao ZHANG ; Fengfeng XIE ; Zeyu LI ; Hua ZHU
China Pharmacy 2022;33(13):1653-1659
Piperine is a kind of amide alkaloids presenting in Piper nigrum L.,which has the pharmacological action such as protecting cardiovascular system ,regulating glucose and lipid metabolism ,anti-tumor,improving nervous system diseases , anti-inflammation and so on. This paper summarized the pharmacological action and mechanisms of piperine in recent years and found that piperine ,as the main active ingredient of P. nigrum ,could protect the cardiovascular system by reducing inflammation and oxidative stress ;improve mitochondrial function through anti-inflammatory and antioxidant effects ,thereby regulate glucose and lipid metabolism ;play an anti-tumor role by mediating the signaling pathways of Wnt/β-catenin,NF-κB/Nrf-2/KeAP-1/HO-1, PI3K/Akt,TGF-β1/Smad2/ERK1/2;improve neurological diseases by inhibiting autophagy ,relieving inflammation ,improving antioxidant,inhibiting neuronal apoptosis and regulating the expression of related proteins in neurons ;play an anti-inflammatory effect by inhibiting the activity of NF-κB and other signaling pathways and reducing the expression of inflammation-related proteins. However,the mechanism of action of piperine is not perfect ,and most of the studies have been confined to the pharmacological level or a certain signaling pathway and a certain target ,without being able to elucidate the interconnection between the relevant signaling pathway and the specific target from a holistic perspective. In the follow-up ,the specific targets of piperine can be identified and clinical trials can be carried out to provide support for the clinical application of piperine.
5.Effect of Stemona tuberosa Alkaloids on Apoptosis and PI3K/Akt and JNK/MAPK Signaling Pathways of Human Lung Cancer A549 Cells
Si LIN ; Huizhen QIN ; Zeyu LI ; Liba XU ; Lingyu DENG ; Jing LUO ; Fengfeng XIE ; Miao ZHANG ; Hua ZHU ; Xiaoxun WANG
Chinese Journal of Experimental Traditional Medical Formulae 2023;29(4):69-76
ObjectiveTo investigate the effect of Stemona tuberosa alkaloids (STA) on apoptosis and phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) and c-Jun N-terminal kinase/p38 mitogen-activated protein kinase (JNK/p38 MAPK) signaling pathways in human lung cancer A549 cells. MethodA549 cells were classified into blank group and STA groups (100, 150, 200, 250, 300 mg⋅L-1). Thiazole blue (MTT) assay and colony formation assay were used to evaluate the proliferation of A549 cells. Apoptosis was observed based on Hoechst 33258 staining, flow cytometry, and Annexin V-FITC/PI staining. Western blot was employed to detect the expression of apoptosis-related proteins cysteine-aspartic acid protease-3 (Caspase-3), B-cell lymphoma-2 (Bcl-2)-associated X protein (Bax), and Bcl-2, and the expression of PI3K, phosphorylated (p)-PI3K, Akt, p-Akt, JNK, p-JNK, p38 MAPK, and p-p38 MAPK. ResultCompared with the blank group, STA groups (150, 200, 250, 300 mg⋅L-1) demonstrated the increase in inhibition rate of cell proliferation (P<0.01) and cell clone inhibition rate, and decrease in cell clone formation rate (P<0.01). In comparison with the blank group, STA groups showed typical characteristics of apoptosis, such as chromatin condensation and enhanced fluorescence reaction. The apoptosis rate of STA groups was significantly higher than that of the blank group (P<0.01). Compared with the blank group, STA (150, 200, 250, 300 mg⋅L-1) significantly up-regulated the protein expression of Caspase-3 and Bax (P<0.05, P<0.01) and down-regulated the expression of Bcl-2 protein (P<0.01). Compared with the blank group, STA had no significant influence on the total protein expression of PI3K, Akt, JNK, and p38 MAPK. However, STA (150, 200, 250, 300 mg⋅L-1) significantly decreased the levels of p-PI3K and p-Akt (P<0.05, P<0.01) and increased the level of p-p38 MAPK (P<0.05, P<0.01). Compared with the blank group, STA (200, 250, 300 mg⋅L-1) significantly raised the level of p-JNK (P<0.05, P<0.01). ConclusionSTA can inhibit the proliferation and induce the apoptosis of A549 cells by inhibiting PI3K/Akt signaling pathway and activating JNK/p38 MAPK signaling pathway.
6.Isoliquiritigenin induces HMOX1 and GPX4-mediated ferroptosis in gallbladder cancer cells.
Zeyu WANG ; Weijian LI ; Xue WANG ; Qin ZHU ; Liguo LIU ; Shimei QIU ; Lu ZOU ; Ke LIU ; Guoqiang LI ; Huijie MIAO ; Yang YANG ; Chengkai JIANG ; Yong LIU ; Rong SHAO ; Xu'an WANG ; Yingbin LIU
Chinese Medical Journal 2023;136(18):2210-2220
BACKGROUND:
Gallbladder cancer (GBC) is the most common malignant tumor of biliary tract. Isoliquiritigenin (ISL) is a natural compound with chalcone structure extracted from the roots of licorice and other plants. Relevant studies have shown that ISL has a strong anti-tumor ability in various types of tumors. However, the research of ISL against GBC has not been reported, which needs to be further investigated.
METHODS:
The effects of ISL against GBC cells in vitro and in vivo were characterized by cytotoxicity test, RNA-sequencing, quantitative real-time polymerase chain reaction, reactive oxygen species (ROS) detection, lipid peroxidation detection, ferrous ion detection, glutathione disulphide/glutathione (GSSG/GSH) detection, lentivirus transfection, nude mice tumorigenesis experiment and immunohistochemistry.
RESULTS:
ISL significantly inhibited the proliferation of GBC cells in vitro . The results of transcriptome sequencing and bioinformatics analysis showed that ferroptosis was the main pathway of ISL inhibiting the proliferation of GBC, and HMOX1 and GPX4 were the key molecules of ISL-induced ferroptosis. Knockdown of HMOX1 or overexpression of GPX4 can reduce the sensitivity of GBC cells to ISL-induced ferroptosis and significantly restore the viability of GBC cells. Moreover, ISL significantly reversed the iron content, ROS level, lipid peroxidation level and GSSG/GSH ratio of GBC cells. Finally, ISL significantly inhibited the growth of GBC in vivo and regulated the ferroptosis of GBC by mediating HMOX1 and GPX4 .
CONCLUSION
ISL induced ferroptosis in GBC mainly by activating p62-Keap1-Nrf2-HMOX1 signaling pathway and down-regulating GPX4 in vitro and in vivo . This evidence may provide a new direction for the treatment of GBC.
Animals
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Mice
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Carcinoma in Situ
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Chalcones/pharmacology*
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Ferroptosis
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Gallbladder Neoplasms/genetics*
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Glutathione Disulfide
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Kelch-Like ECH-Associated Protein 1
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Mice, Nude
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NF-E2-Related Factor 2/genetics*
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Reactive Oxygen Species
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Humans