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ABSTRACT: MicroRNAs (miRNAs) have been implicated in the maintenance of the cancer stem cell (CSC) phenotype via their ability to affect expression of genes and proteins that regulate cell proliferation and/or cell death. Thus, identification of CSC-related miRNAs would provide information for a better understanding of CSCs. Here, we compared the miRNA profiles of CD133+ and CD133- primary hepatocellular carcinoma (HCC) subpopulations and found upregulation of 5 miRNAs in CD133- subpopulations, including hsa-miR-150, which may be involved in maintenance of the CD133+ liver CSC phenotype. We also show that miR-150 interacts with the 3'UTR of c-Myb mRNA and overexpression of miR-150 downregulates c-Myb protein levels. Furthermore, overexpression of miR-150 lead to a significant reduction of CD133+ cells, accompanied by significant inhibition of cell growth and tumorsphere formation. In addition, overexpression of miR-150 induces cell cycle arrest and apoptosis in CD133+ cells. Consistent with the outcome of cell cycle arrest and cell apoptosis, Western blotting results demonstrate that the cell cycle regulator cyclin D1 and cell survival regulator Bcl-2 are decreased in cells transfected with miR-150. Collectively, our findings demonstrate for the first time that miR-150 may be involved in liver CSC self-renewal, potentially via modulation of the downstream target c-Myb.
International Journal of Oncology 03/2012; 40(3):747-56. · 2.40 Impact Factor
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ABSTRACT: Antibodies have been proved to be effective in cancer treatment. Peroxiredoxin I (Prx I) is a potential target for cancer radiotherapy. The aim of this article is to investigate the effect of a novel phage display single-chain variable fragment (scFv) antibody targeting Prx I on human lung carcinoma cell line A549 radiosensitivity and the underlying mechanisms.
A549 cell radiosensitivity was measured by colony-forming assay; cell cycle, cell apoptosis, and intracellular reactive oxygen species (ROS) level were determined by flow cytometer; RAD51 and γ-H2AX expression was evaluated by Western blot; and caspase 3 expression was determined by immunocytochemistry. A nude mouse bearing A549 tumor was established, and radiation sensitivity was measured to verify the in vitro data.
Prx I scFv incubation significantly increased A549 cell radiosensitivity, and this might be through enhanced intracellular ROS level and caspase 3 expression. In addition, protein expression of radiosensitivity-related proteins, RAD51 and γ-H2AX, was also modulated. The nude mouse xenograft model bearing A549 tumor treated with Prx I scFv also exibited enhanced radiosensitivity compared with the PBS group.
These results suggested that Prx I scFv could become a new therapy candidate for lung cancer radiotherapy.
Cancer Biotherapy & Radiopharmaceuticals 10/2011; 27(5):307-16. · 1.44 Impact Factor
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ABSTRACT: The present paper is aimed to detect superparamagnetic iron oxide labeled c-erbB2 oncogene antisense oligonucleotide probe (magnetic antisense probe) connected with SK-Br-3 oncocyte mRNA nucleotide by high resolution atomic force microscope (AFM). We transfected SK-Br-3 oncocyte with magnetic antisense probe, then observed the cells by AFM with high resolution and detected protein expression and magnetic resonance imagine (MRI). The high resolution AFM clearly showed the connection of the oligonucleotide remote end of magnetic antisense probe with the mRNA nucleotide of oncocyte. The expression of e-erbB2 protein in SK-Br3 cells were highly inhibited by using magnetic antisense probe. We then obtained the lowest signal to noise ratio (SNR) of SK-Br-3 oncocyte transfected with magnetic antisense probe by MRI (P<0.05). These experiments demonstrated that the high resolution AFM could be used to show the binding of magnetic antisense probe and SK-Br-3 mRNA of tumor cell nuclear.
Sheng wu yi xue gong cheng xue za zhi = Journal of biomedical engineering = Shengwu yixue gongchengxue zazhi 06/2011; 28(3):442-5.
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ABSTRACT: Baicalin-polyvinylpyrrolidone (PVP) coprecipitate was prepared by the solvent method of solid dispersion technology to improve the dissolution rate of baicalin. The coprecipitate was characterized using differential scanning calorimetry (DSC), X-ray powder diffraction (XRD), infrared spectrometry (IR) and dissolution testing. Furthermore, AFM·IPC-208B high-resolution atomic force microscopy (AFM) was utilized to characterize the molecular morphology of baicalin within its carrier and the interaction between baicalin and its carrier. The results of DSC and XRD indicated that baicalin resided in PVP polymers in an amorphous or molecular phase, dissolution test results demonstrated that the dissolution rate of the coprecipitate was 21.4 times that of the active pharmaceutical ingredient (API). The results of IR indicated the possibility of the formation of intermolecular hydrogen bonds. The AFM·IPC-208B findings revealed that baicalin was dispersed in PVP polymers with a molecular size of 2 nm and either wrapped or surrounded by approximately 0.4 nm of a five-membered ring of PVP arranged along the carbon chain sequentially. An intermolecular hydrogen bond was formed between the 4-OH of the glucuronide of baicalin and the O of the carbonyl group from PVP in addition to the formation of intramolecular hydrogen bonds within baicalin.
International journal of pharmaceutics 02/2011; 408(1-2):91-6. · 2.96 Impact Factor
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ABSTRACT: The protein kinase LKB1 is a crucial regulator of cell growth/proliferation and cell polarity and is the causative gene in the cancer-predisposing disease Peutz-Jeghers syndrome (PJS). The activity of LKB1 is greatly enhanced following its association with the Ste20-like adapter protein STRAD. Unlike LKB1 however, mutations in STRAD have not been identified in PJS patients and thus, the key tumour suppressive role(s) of LKB1 might be STRAD independent. Here, we report that Caenorhabditis elegans strd-1/STRAD mutants recapitulate many phenotypes typical of par-4/LKB1 loss of function, showing defects during early embryonic and dauer development. Interestingly, although the growth/proliferation defects in severe par-4 and strd-1 mutant dauers are comparable, strd-1 mutant embryos do not share the polarity defects of par-4 embryos. We demonstrate that most of par-4-dependent regulation of germline stem cell (GSC) quiescence occurs through AMPK, whereby PAR-4 requires STRD-1 to phosphorylate and activate AMPK. Consistent with this, even though AMPK plays a major role in the regulation of cell proliferation, like strd-1 it does not affect embryonic polarity. Instead, we found that the PAR-4-mediated phosphorylation of polarity regulators such as PAR-1 and MEX-5 in the early embryo occurs in the absence of STRD-1. Thus, PAR-4 requires STRD-1 to phosphorylate AMPK to regulate cell growth/proliferation under reduced insulin signalling conditions, whereas PAR-4 can promote phosphorylation of key proteins, including PAR-1 and MEX-5, to specify early embryonic polarity independently of STRD-1. Our results therefore identify a key strd-1/STRAD-independent function of par-4/LKB1 in polarity establishment that is likely to be important for tumour suppression in humans.
Development 02/2010; 137(4):661-70. · 6.60 Impact Factor
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ABSTRACT: The Peroxiredoxin I (Prx I) is a member of the Peroxiredoxin family, which is overexpressed in many diverse tumor types and is an anti-apoptosis protein for tumor cell proliferation and survival. Therapeutic strategies targeting the Prx I may therefore be effective broad-spectrum anticancer agents. We constructed a phage display single-chain variable fragment (scFv) antibody library and sieve out the fully human, lung adenocarcinoma-sepcific monoclonal antibodies. The selection on Prx I was performed using above-mentioned lung adenocarcinoma-sepcific monoclonal antibodies with high affinity to Prx I overexpressing lung adenocarcinoma cells. The candidate scFv sequences, based on enzyme-linked immunosorbent assay (ELISA) screening data, were chosen for soluble expression, and a 30 kDa band was observed on polyacrylamide gel electrophoresis as predicted. The purified antibodies were characterized by immunoblotting and showed high specificity to Prx I-overexpressing lung adenocarcinoma cells A549. Radioimmunoimaging was taken to evaluate specificity and distribution of antibodies in vivo. The radiolocalization index (RI) of tumor/serum and tumor/muscle gradually increased, reaching its peak (4.06 +/- 0.13 and 5.17 +/- 0.97, respectively) at 48 h postadministration. Single photon emission computed tomography (SPECT) imaging showed the radioactivity was aggregated in tumor locations and tumor imaging was clearly observed. The internalized scFv resulted in antibody-mediated cell apoptosis and downregulation of Prx I expression. These results demonstrate that the scFv possesses strong antitumor activity on lung adenocarcinoma and may therefore be an effective therapeutic candidate for the treatment of cancers that are dependent on Prx I for growth and survival.
Cancer biology & therapy 08/2009; 8(14):1369-77. · 2.64 Impact Factor
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ABSTRACT: Molecular imaging of tumor antisense gene techniques have been applied to the study of magnetic resonance (MR) gene imaging associated with malignant tumors. In this study, we designed, synthesized, and tested a novel molecular probe, in which the antisense oligodeoxynucleotide (ASODN) was labeled with superparamagnetic iron oxide (SPIO), and its efficiency was examined by in vitro MR imaging after SK-Br-3 mammary carcinoma cell lines (oncocytes) transfection. The SPIO-labeled ASODN probe was prepared through SPIO conjugated to ASODN using a chemical cross linking method. Its morphology and size were detected by atomic force microscope, size distribution were detected by laser granulometer, the conjugating rate and biological activity were determined by high performance liquid chromatography, and the stability was determined by polyacrylamide gel electrophoresis. After that, the probes were transfected into the SK-Br-3 oncocytes, cellular iron uptake was analyzed qualitatively at light and electron microscopy and was quantified at atomic absorption spectrometry, and the signal change of the transfected cells was observed and measured using MR imaging. The morphology of the SPIO-labeled ASODN probe was mostly spherical with well-distributed scattering, and the diameters were between 25 and 40 nm (95%) by atomic force microscope and laser granulometer, the conjugating rate of the probe was 99%. Moreover, this probe kept its activity under physiological conditions and could conjugate with antisense oligodeoxynucleotide. In addition, light microscopy revealed an intracellular uptake of iron oxides in the cytosol and electron microscopic studies revealed a lysosomal deposition of iron oxides in the transfected SK-Br-3 oncocytes by antisense probes, some of them gathered stacks, and the iron content of the group of transfected SK-Br-3 oncocytes by antisense probe is significantly higher (18.37 +/- 0.42 pg) than other contrast groups, the MR imaging showed that transfected SK-Br-3 oncocytes by antisense probe had the lowest signal of all. The SPIO-labeled ASODN probe shows unique features including well-distributed spherical morphology, high conjugating rate and loading efficiency, and the signal intensity of SPIO-labeled ASODN-transfected SK-Br-3 oncocytes is reduced in MR imaging. These results indicate that the SPIO-labeled ASODN probe is potentially useful as a MR targeting contrast enhancing agent to specifically diagnose tumors which had over-expression of the c-erbB2 oncogene at an early stage.
Annals of biomedical engineering 05/2009; 37(6):1240-50. · 2.41 Impact Factor
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ABSTRACT: The Notch signalling pathway is conserved among higher metazoans and is used repeatedly throughout development to specify distinct cell fates among populations of equipotent cells. Mounting evidence suggests that Notch signalling may also be crucial in neuronal function in postmitotic, differentiated neurons. Here, we demonstrate a novel role for the canonical Notch signalling pathway in postmitotic neurons during a specialised ;diapause-like' post-embryonic developmental stage in C. elegans called dauer. Our data suggest that cell signalling downstream of the developmental decision to enter dauer leads to the activation of Notch-responding genes in postmitotic neurons. Consistent with this, we demonstrate that glp-1, one of the two C. elegans Notch receptors, and its ligand lag-2 are expressed in neurons during the dauer stage, and both genes are required to maintain this stage in a daf-7/TGFbeta dauer constitutive background. Our genetic data also suggest that a second Notch receptor, lin-12, functions upstream of, or in parallel with, insulin-like signalling components in response to replete growth conditions to promote dauer recovery. Based on our findings, cues associated with the onset of dauer ultimately trigger a glp-1-dependent Notch signalling cascade in neurons to maintain this developmental state. Then, as growth conditions improve, activation of the LIN-12 Notch receptor cooperates with the insulin-like signalling pathway to signal recovery from the dauer stage.
Development 09/2008; 135(15):2583-92. · 6.60 Impact Factor
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ABSTRACT: To explore the possibility of the c-erbB2 oncogene antisense probe labeled with superparamagnetic iron oxide (SPIO) nanoparticles as a target contrast agent for magnetic resonance (MR) imaging whose morphology was observed with atomic force microscopy (AFM), and its efficiency was examined by MR imaging.
The c-erbB2 oncogene antisense probe labeled with SPIO was synthesized by a chemical cross-linking approach. Its morphology was observed with AFM.
The chemical constitution of c-erbB2 oncogene antisense probes can be observed with AFM. The molecular structure of probes is easily visualized under AFM. Probes with diameters of 25-40 nm are in order, follow uniformity and the arrangement rule, can be separated from each other, and appear as cubes with a rugged surface morphology. Strong, low signals of the probes in transfected cells were observed by MR cellular imaging.
AFM is ideal for morphological observation and for analyzing the molecular structure of synthesized c-erbB2 oncogene antisense probes.
Molecular vision 02/2008; 14:114-7. · 2.20 Impact Factor
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ABSTRACT: The formation of a complex multicellular organism requires the precise specification of many diverse cell types at the correct time and position throughout development. This may be achieved by coordinating cell fate specification processes with progression through the cell cycle. Here, we show that the extra distal tip cells (DTCs) associated with the loss of cki-1, a Caenorhabditis elegans homologue of the cyclin-dependent kinase inhibitor p27, do not arise from duplications of pre-existing DTCs, but that they are formed from another cell type within the somatic gonad. Results from our laser microsurgery experiments suggest that the extra DTCs are caused by aberrant somatic gonadal precursor cell divisions in the absence of cki-1, resulting in abnormal daughter cell fates. cki-1(RNAi) animals also possess extra anchor cells and ectopic gonad arms with variable sheath cell numbers and positioning. In addition, cki-1(RNAi) animals display an endomitotic oocyte (Emo) phenotype. Our results uncover a novel role of this CKI in cell fate acquisition, either by directly influencing specification, or through a more conventional role in appropriately linking cell cycle phase with this process.
Developmental Biology 12/2003; 263(2):242-52. · 4.07 Impact Factor
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ABSTRACT: The innexins represent a highly conserved protein family, the members of which make up the structural components of gap junctions in invertebrates. We have isolated and characterized a Caenorhabditis elegans gene inx-6 that encodes a new member of the innexin family required for the electrical coupling of pharyngeal muscles. inx-6(rr5) mutants complete embryogenesis without detectable abnormalities at restrictive temperature but fail to initiate postembryonic development after hatching. inx-6 is expressed in the pharynx at all larval stages, and an INX-6::GFP fusion protein showed a punctate expression pattern characteristic of gap junction proteins localized to plasma membrane plaques. Video recording and electropharyngeograms revealed that in inx-6(rr5) mutants the anterior pharyngeal (procorpus) muscles were electrically coupled to a lesser degree than the posterior metacorpus muscles, which caused a premature relaxation in the anterior pharynx and interfered with feeding. Dye-coupling experiments indicate that the gap junctions that link the procorpus to the metacorpus are functionally compromised in inx-6(rr5) mutants. We also show that another C. elegans innexin, EAT-5, can partially substitute for INX-6 function in vivo, underscoring their likely analogous function.
Molecular Biology of the Cell 08/2003; 14(7):2630-44. · 4.94 Impact Factor
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ABSTRACT: Consider the problem of predicting the occurrence of an event, the onset of diabetes mellitus, say, from a vector of continuous and discrete predictors. We propose a new algorithm for the construction of a tree-structured predictor for the event of interest, which uses a new approach for dealing with continuous predictors. The novelty is that the tree uses splits for continuous variables. This means that at each node an individual goes to the right branch with a certain probability, function of a predictor. The predictor as well as the particular shape of the function is chosen from the data by the proposed algorithm. We evaluate its performance on several real data sets, in particular comparing it with a standard tree-growing algorithm. We also present an analysis of a well-known data set, the Pima Indian diabetes data set, to illustrate the application of the method in biostatistics.
Statistics in Medicine 05/2002; 21(8):1145-65. · 1.88 Impact Factor
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Shaolin Li
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ABSTRACT: Initiation of postembryonic development is an important event for normal C. elegans development. Extrinsic factors affect development as well as intrinsic developmental cues. In order to investigate the molecular basis of initiation of postembryonic development, a genetic screen was performed to identify temperature-sensitive mutants that cannot initiate the cell divisions associated with postembryonic development at the restrictive temperature. Hydroxyurea (HU), a DNA replication inhibitor, was used as a tool to select against worms that initiate postembryonic cell divisions and/or the developmental program. 1,600,000 haploid genomes were screened, and 20 mutants have been isolated. 6 of them have been mapped to a relatively small genetic interval, and one inx-6 has been cloned and encodes an innexin family protein. Mutation of inx-6 caused abnormalities in pharyngeal pumping, resulting in worms that could not feed. The functions of a cyclin B homologue (ZC168.4) in postembryonic development have also been studied since cyclin B mutants also have postembryonic developmental arrest phenotype. Results indicate that zygotic expression of cyclin B is absolutely required for normal postembryonic development. Moreover, we found a novel function of this cyclin B homologue, which demonstrates an uncommon paternal effect required for spermatogenesis and/or fertilization.
