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ABSTRACT: Sample pretreatment is a critical series of processing steps in most protocols involving microarrays. It usually involves
a labor-intensive, time-consuming process, which also requires bulky and costly apparatus to ensure a high-quality RNA extraction.
Furthermore, an efficient hybridization process is also critical for practical applications in microarrays. The present study,
therefore, reports a new microfluidic system capable of automatically performing the sample pretreatment and hybridization
processes for microarrays. Process steps including cell lysis, extraction of messenger ribonucleic acid (mRNA), reverse transcription
(RT), purification of complementary deoxyribonucleic acid (cDNA), and hybridization, are all performed automatically in this
miniature system. When compared to the conventional methods, which usually require well-trained personnel and time consuming
processes, this proposed microfluidic system can perform the entire experimental protocol automatically within 6h. This developed
system may provide a useful platform for subsequent genetic analysis and diagnostic applications.
KeywordscDNA extraction–Cell lysis–Hybridization–mRNA extraction–Reverse transcription–Microfluidics–Microarray–MEMS
Microfluidics and Nanofluidics 04/2012; 10(5):999-1009. · 3.37 Impact Factor
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ABSTRACT: Curcumin, a yellow component of turmeric or curry powder, has been demonstrated to exhibit anti-carcinogenic effects in vitro, in vivo, and in human clinical trials. One of its molecular targets is protein kinase C (PKC) which has been reported to play essential roles in apoptosis, cell proliferation, and carcinogenesis. In this study, PKC mRNA expression was significantly inhibited in curcumin-treated human hepatocellular carcinoma (HCC) Hep 3B cells identified using a kinase cDNA microarray. Furthermore, curcumin decreased total protein expression of all PKCs in a time-related manner by immunoblotting of whole cell lysates, nuclear, membrane, and cytosolic fractions. In cytosolic fraction, the expression of PKC-α was totally inhibited by curcumin. In contrast, the expression levels of PKC-ζ and -μ were dramatically increased. Increases in expression of PKC-δ and PKC-ζ in the membrane and nucleus, and PKC-ι in the membrane were detected. In summary, the changes in expression and distribution of subcellular PKC isoforms in curcumin-treated Hep 3B cells suggest possible PKC-associated anti-tumor mechanisms of curcumin and provide alternative therapies for human HCC.
Materiae Vegetabiles 05/2011; 66(2):136-42. · 2.51 Impact Factor
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Chen-Yun Yeh,
Shin-Mei Shin,
Hsuan-Heng Yeh,
Tsung-Jung Wu, Jyh-Wei Shin,
Tsuey-Yu Chang,
Giri Raghavaraju,
Chung-Ta Lee,
Jung-Hsien Chiang,
Vincent S Tseng,
Yuan-Chii G Lee,
Cheng-Huang Shen,
Nan-Haw Chow,
Hsiao-Sheng Liu
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ABSTRACT: A cross-talk between different receptor tyrosine kinases (RTKs) plays an important role in the pathogenesis of human cancers.
Both NIH-Met5 and T24-Met3 cell lines harboring an inducible human c-Met gene were established. C-Met-related RTKs were screened by RTK microarray analysis. The cross-talk of RTKs was demonstrated by Western blotting and confirmed by small interfering RNA (siRNA) silencing, followed by elucidation of the underlying mechanism. The impact of this cross-talk on biological function was demonstrated by Trans-well migration assay. Finally, the potential clinical importance was examined in a cohort of 65 cases of locally advanced and metastatic bladder cancer patients.
A positive association of Axl or platelet-derived growth factor receptor-alpha (PDGFR-α) with c-Met expression was demonstrated at translational level, and confirmed by specific siRNA knock-down. The transactivation of c-Met on Axl or PDGFR-α in vitro was through a ras- and Src-independent activation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK/ERK) pathway. In human bladder cancer, co-expression of these RTKs was associated with poor patient survival (p < 0.05), and overexpression of c-Met/Axl/PDGFR-α or c-Met alone showed the most significant correlation with poor survival (p < 0.01).
In addition to c-Met, the cross-talk with Axl and/or PDGFR-α also contributes to the progression of human bladder cancer. Evaluation of Axl and PDGFR-α expression status may identify a subset of c-Met-positive bladder cancer patients who may require co-targeting therapy.
BMC Cancer 01/2011; 11:139. · 3.01 Impact Factor
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Pin-Yuan Chen,
Hao-Li Liu,
Mu-Yi Hua,
Hung-Wei Yang,
Chiung-Yin Huang,
Po-Chun Chu,
Lee-Ang Lyu,
I-Chou Tseng,
Li-Ying Feng,
Hong-Chieh Tsai, [......],
Yu-Jen Lu,
Jiun-Jie Wang,
Tzu-Chen Yen,
Yunn-Hwa Ma,
Tony Wu,
Jyh-Ping Chen,
Jih-Ing Chuang, Jyh-Wei Shin,
Chuen Hsueh,
Kuo-Chen Wei
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ABSTRACT: Malignant glioma is a common and severe primary brain tumor with a high recurrence rate and an extremely high mortality rate within 2 years of diagnosis, even when surgical, radiological, and chemotherapeutic interventions are applied. Intravenously administered drugs have limited use because of their adverse systemic effects and poor blood-brain barrier penetration. Here, we combine 2 methods to increase drug delivery to brain tumors. Focused ultrasound transiently permeabilizes the blood-brain barrier, increasing passive diffusion. Subsequent application of an external magnetic field then actively enhances localization of a chemotherapeutic agent immobilized on a novel magnetic nanoparticle. Combining these techniques significantly improved the delivery of 1,3-bis(2-chloroethyl)-1-nitrosourea to rodent gliomas. Furthermore, the physicochemical properties of the nanoparticles allowed their delivery to be monitored by magnetic resonance imaging (MRI). The resulting suppression of tumor progression without damaging the normal regions of the brain was verified by MRI and histological examination. This noninvasive, reversible technique promises to provide a more effective and tolerable means of tumor treatment, with lower therapeutic doses and concurrent clinical monitoring.
Neuro-Oncology 10/2010; 12(10):1050-60. · 5.72 Impact Factor
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Kuo-Chen Wei,
Chiung-Yin Huang,
Pin-Yuan Chen,
Li-Ying Feng,
Tai-Wei Erich Wu,
Shu-Mei Chen,
Hong-Chieh Tsai,
Yu-Jen Lu,
Ngan-Ming Tsang,
Chen-Kan Tseng,
Ping-Ching Pai, Jyh-Wei Shin
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ABSTRACT: Glioblastoma and astrocytoma are the most common brain tumors affecting adults 45-60 years of age. The poor prognosis for glioblastoma patients results from recurrence after treatment. There is therefore an urgent need to develop diagnostic and prognostic markers as well as new therapies.
Microarray analyses of clinical specimens from glioblastoma patients were used to identify potential tumor markers. Expression of candidate genes was analyzed by real-time reverse transcription-polymerase chain reaction and by immunoblotting and immunohistochemistry.
Five potential markers (CD44 antigen (CD44), growth arrest and DNA-damage-inducible, alpha (GADD45A), fibronectin 1 (FN1), CD63 antigen (CD63) and secreted phosphoprotein 1 (SPP1)) showed expression patterns that correlated significantly with malignant glioma. In particular, expression of the CD44 antigen was elevated in more severe tumor types, and higher in tumor cores than in peripheral regions. However, lower levels of CD44 expression surprisingly correlated with lower survival.
The CD44 antigen is a promising candidate for further development as a prognostic and therapeutic tool.
Anticancer research 01/2010; 30(1):253-9. · 1.73 Impact Factor
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ABSTRACT: Intrapleural fibrinolytic therapy (IFT) provides clinical benefit in the treatment of complicated pleural parapneumonic effusion (CPE). Whether IFT influences the proinflammatory cytokines production and fibrinlytic activity is currently unclear. Therefore, we collected pleural effusion samples from CPE patients with IFT (study group) and patients without IFT (control group). A membrane human inflammatory cytokines array kit was used to compare the difference of targeted cytokine production between these two groups. Enzyme-linked immunosorbent assay (ELISA) methods were used for quantitative analysis of targeted cytokines and fibrinolytic enzymes. The results showed there were no significant differences between the study (n = 16) and control (n = 14) groups in patients' demographic data. After fibrinolytic therapy, the patients in the study group had significant lower plasminogen activator inhibitor (PAI) level (732.36+/-254.09 ng/mL vs 1,509.36+/-1,340.11 ng/mL, p<0.05) and higher urokinase plasminogen activator (u-PA) level (75.56+/-41.70 ng/mL vs 6.87+/-5.07 ng/mL, p<0.05) than they did before treatment. Moreover, the tissue inhibitors of metalloproteinase-2 (TIMP-2) (1,560.03+/-403.49 pg/mL vs 3,686.45+/-1,263.83 pg/mL, p<0.05) and inflammatory chemokine, regulated on activation normal T-cell expressed and secreted/chemokine (C-C motif) ligand 5 (RANTES), (293.58+/-212.93 pg/mL vs 749.27+/-53.79 pg/mL, p<0.05), were also significantly lower in the study group after fibrinolytic therapy, but not in the control group. In conclusion, intrapleural fibrinolytic treatment with urokinase could enhance fibrinolytic activity and decrease TIMP-2 and RANTES production.
Inflammation 09/2009; 32(6):410-8. · 1.75 Impact Factor
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Wei-Chen Lin,
Sung-Chou Li,
Wen-Chang Lin, Jyh-Wei Shin,
Song-Nian Hu,
Xiao-Min Yu,
Ting-Yun Huang,
Shih-Chieh Chen,
Hua-Chien Chen,
Shu-Jen Chen,
Po-Jung Huang,
Richie Ruei-Chi Gan,
Cheng-Hsun Chiu,
Petrus Tang
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ABSTRACT: MicroRNAs (miRNAs) are a class of small noncoding RNAs that have important regulatory roles in multicellular organisms. However, miRNA has never been identified experimentally in protist. Direct cloning of 438 expressed miRNA tags by microRNA serial analysis of gene expression from the parasitic protist Trichomonas vaginalis identified nine candidate miRNAs. Bioinformatics analysis of the corresponding genomic region revealed that these miRNA candidates contain a classical stem-loop-stem structure of pre-microRNAs. Analysis of the 20 nt long mature tva-miR-001 showed that it is an intergenic miRNA located at the scaffold DS113596. Tva-miR-001 was differentially expressed in the trophozoite, pseudocyst and amoeboid stages. Based on the experimental results of the present study, we provided solid evidence that protist possesses a miRNA regulating network comparable with multicellular organisms for the first time.
Genomics 06/2009; 93(5):487-93. · 3.02 Impact Factor
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ABSTRACT: This paper presents a new integrated microfluidic chip that automatically performs ribonucleic acid (RNA) extraction and reverse transcription (RT) processes. The microfluidic system consists of a microfluidic control module and a magnetic bio-separator. The microfluidic control module can perform pumping and mixing of small amount of fluids and subsequent purification and concentration of RNA samples by incorporating with the magnetic bio-separator consisting of 2-dimension twisted microcoils. Notably, the magnetic bio-separators are developed either to generate the required magnetic field to perform the separation of magnetic beads or to work as a micro-heater to control the temperature field for the following RT process. Experimental results show that the total RNA can be successfully purified and extracted by using magnetic beads and the subsequent RT processing of the RNA can be performed automatically. Total RNA is successfully extracted and purified from T98 cells utilizing the microfluidic system, which is comparable with the conventional methods. The whole automatic procedure of RNA sample extraction only takes 35 min, which is much faster than the conventional method (more than 2 h). As a whole, the developed microfluidic system may provide a powerful platform for rapid RNA extraction and RT processes for further biomedical applications.
Biomedical Microdevices 12/2008; 11(2):339-50. · 3.03 Impact Factor
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ABSTRACT: There has been a rapid increase in cross-border female marriage immigrants in Taiwan. In this study, 1,434 Vietnamese female marriage immigrants arriving between July 1998 and June 2001 were examined for intestinal parasites. Most (77.9%) of these women were 20-29 years of age. The prevalence of intestinal parasite infection was 37.7%, and the trend increased from 1999 to 2001 (statistically significant, p < 0.0001), but decreased among age subgroups (p < 0.0001). Among the 20 species of intestinal parasites found in this study, 10 species (27.8%) were transmitted via the fecal-oral route, 5 (14.6%) via the soil-mediated route, and 5 (0.7%) by food-borne infection. The prevalence of blastocystosis (20.4%) and hookworm (9.7%) remained high among this population. The results provide unprecedented information on intestinal parasitic infection among these immigrants in southern Taiwan and recommend that appropriate health care be given after parasite infection is confirmed in these migrant communities.
The Kaohsiung journal of medical sciences 04/2006; 22(4):166-70. · 0.61 Impact Factor
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ABSTRACT: Abundant information about gene products is stored in online searchable databases such as annotation or literature. To efficiently obtain and digest such information, there is a pressing need for automated information-summarization and functional-similarity clustering of genes.
We have developed a novel method for semantic measurement of annotation and integrated it with a biomedical literature summarization system to establish a platform, GeneLibrarian, to provide users well-organized information about any specific group of genes (e.g. one cluster of genes from a microarray chip) they might be interested in. The GeneLibrarian generates a summarized viewgraph of candidate genes for a user based on his/her preference and delivers the desired background information effectively to the user. The summarization technique involves optimizing the text mining algorithm and Gene Ontology-based clustering method to enable the discovery of gene relations.
GeneLibrarian is a Java-based web application that automates the process of retrieving critical information from the literature and expanding the number of potential genes for further analysis. This study concentrates on providing well organized information to users and we believe that will be useful in their researches. GeneLibrarian is available on http://gen.csie.ncku.edu.tw/GeneLibrarian/.
BMC Bioinformatics 02/2006; 7:392. · 2.75 Impact Factor
