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Development of hybrid-type modified chitosan derivative nanoparticles for the intracellular delivery of midkine-siR NA in hepatocellular carcinoma cells

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Hepatobiliary & Pancreatic Diseases International 2015年第1期14卷 82-89页

作者： Jing Zhong Hui-Lian Huang Jing Li Fu-Chu Qian Li-Qin Li Ping-Ping Niu Li-Cheng Dai Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital

BACKGROUND： Hepatocellular carcinoma（HCC） is one of the most common cancers worldwide. Most of the patients with HCC lose the surgical opportunity at the time of diagno sis. Some novel therapeutic modalities, like gene therapy, are promising for the treatment of HCC. However, the success of gene therapy depends on two aspects： efficient gene materials and gene delivery vectors. The present study was to develop new chitosan-based nanoparticles for a midkine-si RNA（anti HCC gene drug） delivery.METHODS： The novel gene delivery vector（MixN CH） was syn thesized by hybrid-type modification of chitosan with 2-chloro ethylamine hydrochloride and N, N-dimethyl-2-chloroethylamine hydrochloride. The chemical structure of Mix NCH was char acterized by FT-IR and 1HNMR. The cytotoxicity of Mix NCH was determined by MTS assay. The gene condensation ability and size, zeta potential and morphology of Mix NCH/MK si RNA nanoparticles were measured. The in vitro transfection and gene knockdown efficiency of midkine by Mix NCH/MK si RNA nanoparticles was detected by q RT-PCR and Western blotting. Gene knockdown effect at the molecule level on the proliferation of Hep G2 in vitro was determined by MTS assay.RESULTS： Mix NCH was successfully acquired by aminoalkyl ation modification of chitosan. The Mix NCH could condense MK-siR NA well above the weight ratio of 3. The average size of Mix NCH/MK-si RNA nanoparticles was 100-200 nm, and thesurface charge was about ＋5 m V. Morphologically, Mix NCH/MK-si RNA nanoparticles were in regular spherical shape-with no aggregation. Regarding to the in vitro transfection of nanoparticles, the MixN CH/MK-siR NA nanoparticles reduced MK m RNA level to 14.03%±4.03%, which were comparable to Biotrans（8.94%±3.77%）. Mix NCH/MK-si RNA effectively inhibited the proliferation of Hep G2 in vitro.-CONCLUSION： Mix NCH/MK-si RNA nanoparticles could be effective for the treatment of hepatocellular carcinoma.

关键词： chitosan gene vector nanoparticles midkine siRNA hepatocellular carcinoma

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Midkine translocated to nucleoli and involved in carcinogenesis

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World Journal of Gastroenterology 2009年第4期15卷 412-416页

作者： Li-Cheng Dai Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital Huzhou 313000 Zhejiang ProvinceChina

Midkine(MK) is a heparin-binding growth factor with its gene first identified in embryonal carcinoma cells at early stages of retinoic acid-induced differentiation. MK is frequently and highly expressed in a variety of human carcinomas.Furthermore,the blood MK level is frequently elevated with advance of human carcinomas,decreased after surgical removal of the tumors.Thus,it is expected to become a promising marker for evaluating the progress of carcinomas. There is mounting evidence that MK plays a significant role in carcinogenesis-related activities,such as proliferation,migration,anti-apoptosis,mitogenesis,transforming,and angiogenesis.In addition,siRNA and anti-sense oligonucleotides for MK have yielded great effects in anti-tumor activities.Therefore,MK appears to be a potential candidate molecular target of therapy for human carcinomas.In this paper,we review MK targeting at nucleoli in different tumor cells and its role in carcinogenesis to deepen our understanding of the mechanism of MK involved in carcinogenesis.

关键词：致癌作用癌细胞遗传基因肿瘤科

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Antisense oligonucleotide targeting p53 increased apoptosis of MCF-7 cells induced by ionizing radiation

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Acta Pharmacologica Sinica 2006年第11期27卷 1453-1458页

作者： Li-cheng DAI~2 Xiang WANG Xing YAO Li-shan MIN Fu-chu QIAN Jian-fang HE Huzhou Key Laboratory of Molecular Medicine,huzhou Central Hospital,huzhou 313000,China Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital Huzhou 313000 China. dlc@***

Aim:To investigate the effect of antisense compounds(AS)targeting human p53mRNA on radiosensitivity of MCF-7 cells.Methods:Western blotting and RT-PCR were used to analyze the protein content and mRNA level.Additionally,cellproliferation,cell cycle and cell apoptosis were all analyzed in irradiated or sham-irradiated cells.Results:Among the five antisense compounds(AS),AS3 wasidentified to efficiently inhibit p53 mRNA level and protein content.Interestingly,AS3 transfer has little effect on cell proliferation in DU-145 cells(mutant p53)afterionizing radiation(IR).In contrast,a marked increase of cell apoptosis and growthinhibition were observed in MCF-7 cells(wild-type p53),suggesting that AS3 canincrease radiosensitivity of MCF-7 cells.Additionally,it was also observed thatthe transfection of AS3 decreased the fraction of G1phase cells,and increased theproportion of S phase cells compared to untreated cells 24 h after IR in MCF-7 celllines.Conclusion:AS3 transfection increases MCF-7 cell apoptosis induced by5 Gy-radiation,and this mechanism may be closely associated with abrogation ofG1phase arrest.

关键词： antisense oligodeoxyribonucleotides MCF-7 p53 genes radiation tolerance cell cycle

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Upregulation of MiR-1280 Expression in Non-small Cell Lung Cancer Tissues

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Chinese Medical Journal 2015年第5期128卷 670-673页

作者： Li-Min Xu Li-Qin Li Jing Li Hong-Wei Li Qi-Bin Shen Jin-Liang Ping Zhi-Hong Ma Jing Zhong Li-Cheng Dai Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital Huzhou Zhejiang 313000 China Department of Thoracic Surgery Huzhou Central Hospital Huzhou Zhejiang 313000 China Department of Pathology Huzhou Central Hospital Huzhou Zhejiang 313000 China

Background：Non-small cell lung cancer （NSCLC） is a prolific and high-mortality disease with few effective treatments.Although the detection and surgical techniques for NSCLC continue to advance,the survival rate for the patients with NSCLC remains poor.Enhanced predictive biomarkers such as microRNAs （miRNAs） are needed at the time of diagnosis to better tailor therapies for patients.This study focused on the expression ofmiR-1280 in NSCLC tissues and distal normal tissues in order to explore the association between miR-1280 expression and NSCLC.Methods：A total of 72 newly diagnosed primary NSCLC patients were enrolled in this study.Quantitative real-time polymerase chain reaction （PCR） was performed to identify the expression level ofmiR-1280 in the NSCLC tissues and distal normal tissues of these patients.Results：The miR-1280 expression was significantly higher in the NSCLC tissues （0.084 ± 0.099） than distal normal tissues （0.014 ± 0.015,P =0.009）.In 54 patients （75%）,the miR-1280 expression in the NSCLC tissues was upregulated （2-△△ct 〉 2）,and no case showed a downregulation of miR-1280 expression.Conclusions：The expression level ofmiR-1280 could be regarded as a biomarker for NSCLC.

关键词： Biomarker miR-1280 Non-small Cell Lung Cancer Upregulation

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Allele polymorphisms of interleukin-lO and hepatitis B, C virus infection

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Chinese Medical Journal 2010年第10期123卷 1338-1344页

作者： LU Yong-liang WU Xiao HUANG Hui-lian DAI Li-cheng huzhou Teachers College School of medicine Huzhou Zhejiang 313000 China SIP Halo Biomaterial Technology Co. Ltd. Suzhou Jiangsu 215000 China Huzhou Key Laboratory of Molecular Medicine Affiliated Central Hospital of Huzhou Teachers College Huzhou Zhejiang 313000 China

Background Interleukin 10 （IL-10） is an important cytokine with anti-inflammatory, anti-immune and anti-fibrotic functions. This study aimed at evaluating the relationship between allele polymorphisms in the IL-10 promoter region and hepatitis B virus （HBV） or hepatitis C virus （HCV） infection. Methods The odds ratios （ORs） of IL-10 allele distributions in patients with HBV or HCV infection were analyzed against healthy controls. All the relevant studies in PubMed were identified, and poor qualified studies were excluded. The meta-analysis software REVMAN 4.2 was applied for investigating heterogeneity among individual studies and summarizing all the studies. The publication bias was also evaluated. Results This study demonstrated a significant association between the IL-10-592 A/C polymorphism and HBV infection in the Asian population under the overall effect size of allele A versus C. In our subgroup meta-analysis, we found a significant association of IL-10-592 A/C polymorphism to HCV infection susceptibility in Asian populations, although sensitivity analysis showed that the combined result was not associated with the worldwide population. Other IL-10 allele polymorphisms were not associated with HBV or HCV infection. Conclusion IL-10-592 A/C allele might be a risk factor for HBV or HCV in Asians but not in Europeans.

关键词： meta-analysis interleukin-lO hepatitis B virus hepatitis C virus

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Novel functional proteins interact with midkine in hepatic cancer cells

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Hepatobiliary & Pancreatic Diseases International 2012年第3期11卷 272-277页

作者： Qiang Yan, Hui-Lian Huang, Xing Yao, Jing Li, Li-Qin Li, Jing Zhong, Li-Shan Min, Li-Cheng Dai and Shu-Sen Zheng Department of Hepatobiliary and Pancreatic Surgery, huzhou Central Hospital (Yan Q and Yao X) Huzhou Key Laboratory of Molecular Medicine, Affiliated Central Hospital of huzhou Teachers College (Huang HL, Li J, Li LQ, Zhong J, Min LS and Dai LC), huzhou 313000, China Division of Hepatobiliary and Pancreatic Surgery, First Affiliated Hospital, Zhejiang University School of medicine, Hangzhou 310003, China (Yan Q and Zheng SS) Department of Hepatobiliary and Pancreatic Surgery Huzhou Central Hospital Huzhou 313000 China Division of Hepatobiliary and Pancreatic Surgery First Affiliated Hospital Zhejiang University School of Medicine Hangzhou 310003 China Huzhou Key Laboratory of Molecular Medicine Affiliated Central Hospital of Huzhou Teachers College Huzhou 313000 China

BACKGROUND: Midkine is a heparin-binding growth factor that promotes the proliferation, survival, migration and differentiation of various target cells. Midkine plays an important role in tumorigenesis and tumor progression, and is overexpressed in many human malignant tumors. Patients with high tumor midkine expression frequently have a worse prognosis than those with low expression. The present study was designed to investigate the interaction network of midkine in hepatic cancer cells, and to elucidate its role in hepatocellular carcinoma. METHODS: DNA encoding full-length midkine was cloned into pDBLeu vector to serve as bait in yeast two-hybrid screening to identify interacting proteins. Candidate proteins were examined on SC-Leu-Trp-His+3-AT (20 mmol/L) plates and assayed for X-gal activity, then sequenced and classified according to the GenBank. Finally, identified proteins were expressed by the in vitro expression system pCMVTnT, and protein interactions were confirmed by co-immunoprecipitation. RESULTS: Using the yeast two-hybrid system, we found 6 proteins that interacted with midkine: NK-kappa-B inhibitor alpha (I-κ-B-α), Dvl-binding protein naked cuticle 2, granulin, latent active TGF-β binding protein 3, latent active TGF-β binding protein 4, and phospholipid scramblase 1. In vitro co-immunoprecipitation demonstrated that all identified proteins directly interacted with midkine.CONCLUSION: The identification of midkine-interacting proteins in hepatic cancer cells indicates that midkine is a multifunctional factor that may participate in cell migration, differentiation, and proliferation, and is also associated with the multicellular response feedback during the development of hepatocellular carcinoma.

关键词： midkine yeast two-hybrid interaction hepatic cancer cell

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Clinicopathologic and prognostic implications of progranulin in breast carcinoma

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Chinese Medical Journal 2011年第13期124卷 2045-2050页

作者： LI Li-qin HUANG Hui-lian PING Jin-liang WANG Xiao-hong ZHONG Jing DAI Li-cheng Huzhou Key Laboratory of Molecular Medicine Affiliated Central Hospital of Huzhou Teachers College Huzhou Zhejiang 313000 China Department of Pathology Affiliated Central Hospital of Huzhou Teachers College Huzhou Zhejiang 313000 China Department of Breast Surgery Affiliated Central Hospital of Huzhou Teachers College Huzhou Zhejiang 313000 China

Background Progranulin is a newly discovered 88-kDa glycoprotein originally purified from the highly tumorigenic mouse teratoma-derived cell line PC. Its expression is closely correlated with the development and metastasis of several cancers. However, no immunohistochemical evidence currently exists to correlate progranulin expression with clinicopathologic features in breast carcinoma biopsies, and the role of progranulin as a new marker of metastatic risk and prognosis in breast cancer has not yet been studied. The aim of this study was to investigate the clinicopathologic and prognostic implications of progranulin expression in breast carcinoma and its correlation with tumor angiogenesis. Methods Progranulin expression was determined immunohistochemically in 183 surgical specimens from patients with breast cancer and 20 tissue samples from breast fibroadenomas. The tumor angiogenesis-related biomarker, vascular endothelial growth factor was assayed and microvessel density was assessed by counting vascular endothelial cells in tumor tissues labeled with endoglin antibody. The relationship between progranulin expression and the clinicopathologic data were analyzed. Results Progranulin proteins were overexpressed in breast cancer. The level of progranulin expression was significantly correlated with tumor size （P=0.004）, lymph node metastasis （P 〈0.001） and TNM staging （P 〈0.001）. High progranulin expression was associated with higher tumor angiogenesis, reflected by increased vascular endothelial growth factor expression （P〈0.001） and higher microvessel density （P=0.002）. Conclusion Progranulin may be a valuable marker for assessing the metastasis and prognosis of breast cancer, and could provide the basis for new combination regimens with antiangiogenic activity.

关键词： breast carcinoma progranulin endoglin angiogenesis immunohistochemistry

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In vitro and in vivo suppression of hepatocellular carcinoma growth by midkine-antisense oligonucleotide-loaded nanoparticles

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World Journal of Gastroenterology 2009年第16期15卷 1966-1972页

作者： Li-Cheng Dai Xing Yao Xiang Wang Shu-Qiong Niu Lin-Fu Zhou Fang-Fang Fu Shui-Xin Yang Jin-Liang Ping Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital Huzhou 313000 Zhejiang Province China Department of General Surgery Huzhou Central Hospital Huzhou 313000 Zhejiang Province China Zhejiang University School of medicine Hangzhou 310016 Zhejiang Province China Department of Pathology Huzhou Central Hospital Huzhou 313000 Zhejiang Province China

AIM:To synthesize antisense oligonucleotides (ASODNs) of midkine (MK), package the ASODNs with nanoparticles, and to inhibit hepatocellular carcinoma (HCC) growth using these nanoparticles.METHODS: HepG2 cell proliferation was analyzed in vitro using the 3-(4,5-dimethythiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2Htetrazolium, inner salt assay. The in vivo activity of nanoparticles delivering the MK-ASODNs was analyzed by histopathological and immunohistochemical staining and quantitative real time polymerase chain reaction (PCR). RESULTS: The in vitro proliferation of HepG2 cells was signif icantly inhibited by the nanoparticles packaged with MK-ASODNs (NANO-ASODNs). Furthermore, the NANO- ASODNs signif icantly inhibited the growth of HCC in the mouse model. CONCLUSION: NANO-ASODNs can significantly suppress the growth of HCC in vitro and in vivo.

关键词：反义寡核苷酸肝癌细胞生长纳米粒子中期因子体外增殖体内抑制 HepG2细胞增殖实时定量聚合酶链反应

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Antisense oligonucleotide targeting midkine suppresses in vivo angiogenesis

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World Journal of Gastroenterology 2007年第8期13卷 1208-1213页

作者： Li-Cheng Dai Xiang Wang Xing Yao Yong-Liang Lu Jin-Liang Ping Jian-Fang He Huzhou Key Laboratory of Molecular Medicine Huzhou Central Hospital Huzhou 313000 Zhejiang Province China Department of General Surgery Huzhou Central Hospital Huzhou 313000 Zhejiang Province China Department of Pathology Huzhou Central Hospital Huzhou 313000 Zhejiang Province China

AIM: To evaluate the effect of antisense oligonucleotide targeting midkine (MK-AS) on angiogenesis in chick chorioallantoic membrane (CAM) and in situ human hepatocellular carcinoma (HCC). METHODS: An in situ human hepatocellular carcinoma (HCC) model and CAM assay were used in this experiment. The effect of MK-AS on angiogenesis was evaluated by cell proliferation assay and hematoxylin- eosin (HE) staining. RESULTS: MK-AS significantly inhibited human umbilical vein endothelial cells (HUVEC) and in situ human HCC growth. At the same time, MK-AS suppressed the angiogenesis both in human hepatocellular carcinoma cell line (HEPG2)-induced CAM and in situ human HCC tissues. CONCLUSION: MK-AS is an effective antiangiogenesis agent in vivo.

关键词：中期因子肿瘤血管发生反义寡核苷酸抑制作用靶点

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Identification of Boseongicola sediminum sp.nov.,a Novel Decabromodiphenyl Ether(BDE-209)-tolerant Strain Isolated from Coastal Sediment in Xiamen,China

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Biomedical and Environmental Sciences 2021年第8期34卷 656-661页

作者： TANG Chen FAN Chun GUO Dong Bei MA Xiu Juan CAI Qing Tao CHEN Xiao Xuan ZHANG Min LI Jia Yao AN Qiu Ying ZHAO Ran State key laboratory of molecular Vaccinology and molecular Diagnotics School of Public HealthXiamen UniversityXiamen 361102FujianChina Department of Environ mental and Occupational medicine Shanghai Jinshan Disease Center for Prevention and ControlShanghai 201599China Department of Environmental and Occupational Health Huzhou Center for Disease Prevention and ControlHuzhou 313000ZhejiangChina

Polybrominated diphenyl ethers(PBDEs)are a group of aromatic organic bromine compounds,which have been used in a broad array of polymeric materials(plastics,foams,resins,and adhesives)as brominated flame retardants.They are used in commercial and household products,including textiles,electronic equipment,airplanes,and automobiles,especially in China and the USA,because they are inexpensive with excellent flame-retardant effects[1].During the past decade,over 14,000 tonnes of PBDEs have entered the market,and have become ubiquitous environmental contaminants[2].

关键词： Xiamen resins aromatic

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