A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells

Aoyagi, Kazuhiko; Tatsuta, Takeshi; Nishigaki, Michiko; Akimoto, Shingo; Tanabe, Chikako; Omoto, Yoko; Hayashi, Shin Ichi; Sanada, Hiromi; Sakamoto, Michiie; Yoshida, Teruhiko; Terada, Masaaki; Sasaki, Hiroki

doi:10.1016/s0006-291x(02)02967-4

Cited by 85 publications

(57 citation statements)

References 14 publications

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“…Actually, GSDM protein was observed in the region (Figure 1a). To confirm GSDM mRNA expression in the pit cells, we carried out RT-PCR analysis combined with a laser-captured microdissection (LCM) procedure as described in our previous study (Aoyagi et al, 2003). With this procedure, the GSDM mRNA was detected predominantly in the pit region among the three regions of pit, isthmus/neck and gland (Figure 1b).…”

Section: Resultsmentioning

confidence: 95%

“…The cryostat sections (8 mm) were microdissected with a PixCell II LCM system (Arcturus Engineering, Mountain View, CA, USA). The mRNA was isolated and amplified by our previously developed method (Aoyagi et al, 2003). RT-PCR was carried out using primer sets designed for detecting the 3 0 side of cDNA of each gene: for GSDM, 5 0 -GATGGAACAGAACTTCCTGC-3 0 and 5 0 -CTGAAATGC CCTTAGGTGGT-3 0 , for LMO1, 5 0 -TGGCACCTTTGAAT CCCAAG-3 0 and 5 0 -AGAACTTCTCCGTCCTCGAT-3 0 , for RUNX3, 5 0 -CACGAGGAAAGGAAGACGTT-3 0 and 5 0 -CT GTACAAGCAAGTTGTGCG-3 0 , for RUNX2, 5 0 -CTTCGT CAGGATCCTATCAG-3 0 and 5 0 -TCATCCATTCTGCCAC TAGA-3 0 , for RUNX1, 5 0 -TCATGTCACTACTGTCCGTG 3 0 and 5 0 -CACTGTTCTGAAGTCCTGCT-3 0 .…”

Section: Iimmunohistochemistrymentioning

confidence: 99%

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GASDERMIN, suppressed frequently in gastric cancer, is a target of LMO1 in TGF-β-dependent apoptotic signalling

Saeki¹,

et al. 2007

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Defining apoptosis-regulatory cascades of the epithelium is important for understanding carcinogenesis, since cancer cells are considered to arise as a result of the collapse of the cascades. We previously reported that a novel gene GASDERMIN (GSDM) is expressed in the stomach but suppressed in gastric cancer cell lines. Furthermore, in this study, we demonstrated that GSDM is expressed in the mucus-secreting pit cells of the gastric epithelium and frequently silenced in primary gastric cancers. We found that GSDM has a highly apoptotic activity and its expression is regulated by a transcription factor LIM domain only 1 (LMO1) through a sequence to which Runt-related transcription factor 3 (RUNX3) binds, in a GSDM promoter region. We observed coexpression of GSDM with LMO1, RUNX3 and type II transforming growth factor-b receptor (TGF-bRII) in the pit cells, and found that TGF-b upregulates the LMO1-and GSDMexpression in the gastric epithelial cell line and induces apoptosis, which was confirmed by the finding that the apoptosis induction is inhibited by suppression of each LMO1-, RUNX3-and GSDM expression, respectively. The present data suggest that TGF-b, LMO1, possibly RUNX3, and GSDM form a regulatory pathway for directing the pit cells to apoptosis.

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Section: Resultsmentioning

confidence: 95%

Section: Iimmunohistochemistrymentioning

confidence: 99%

GASDERMIN, suppressed frequently in gastric cancer, is a target of LMO1 in TGF-β-dependent apoptotic signalling

Saeki¹,

et al. 2007

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“…Thus, the potential of DNA microarray-based microbial screening and diagnostic technologies is currently limited by front-end target-specific nucleic acid detection. The presence of an ubiquitous poly-adenylated tail at the 3 0 -end of eukaryotic messenger RNAs offers the possibility to convert minute amounts of RNA to micrograms of labeled material, with minimal effects on the respective abundance of the mRNA mixture [8,139]. Prokaryotic RNAs are not poly-adenylated and are thus more challenging to work with when starting material is scarce.…”

Section: Target Amplifications and Sensitivity Issuesmentioning

confidence: 99%

Introduction to Microarray‐Based Detection Methods

Schrenzel

Kostić²,

Bodrossy

et al. 2009

Detection of Highly Dangerous Pathogens

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Microarrays consist of an orderly arrangement of probes (oligonucleotides, DNA fragments, proteins, sugars or lectins) attached to a solid surface. The main advantages of microarray technology are high throughput, parallelism, miniaturization, speed and automation. Despite the fact that microarray analysis is a relatively novel technology, with the publication of the first microarray studies in 1995 [101,152], it is now broadly applied and the milestone of nearly 5000 published microarray papers was recorded in 2004 [80]. The scientific and technological background discussed here will be limited to DNA microarrays, excluding the new evolving fields of protein microarrays and glycomics [140].Analogous to antigen-antibody interactions on immunoarrays, DNA microarrays rely on sequence complementarity of the two strands. They put in practice the fundamentals of complementary base-pairing (hybridization) that were first described by Ed Southern [162]. In general, the strategy of microarray hybridization is reversed to that of a standard dot-blot, leading to recurring confusion in the nomenclature. Therefore, it has been suggested to describe tethered nucleic acid as the probe and free nucleic acid as the target [134].Earlier studies on duplex melting and reformation, carried out on DNA solutions, have provided the basic knowledge about the reaction kinetics. Furthermore, a method for computational determination of the melting point as a function of nucleic acid composition and salt concentration was established [6,148,149]. Much of the pioneering work can be linked to the use of nitrocellulose membranes [69], dot-blots [84] and Southern blots [162]. Development of cDNA or oligonucleotide arrays was possible by combined innovations in micro-engineering, molecular biology [33,120,123,156,163,164] and bioinformatics [59]. The real breakthrough in microarray technology was initiated by two key innovations: the use of non-porous solid supports (such as glass and silicon) and the development of methods for highdensity synthesis of oligonucleotides directly onto the microarray surface [59].

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“…From each core biopsy, sufficient total RNA was extracted for oligonucleotide array analysis and preliminary patterns predictive of sensitivity and resistance to specific treatments have been reported (Chang et al, 2003), where others report 45% (Buchholz et al, 2002) or as high as 93% (Ellis et al, 2002) of core biopsies to yield sufficient high-quality RNA for array analysis. Other investigators have reported faithful linear RNA amplification protocols using limited amounts of RNA from microdissected breast tissues (Aoyagi et al, 2003). Further work is essential in integrating amplification protocols into large-scale microarray analysis, and validating these pilot predictive expression patterns into independent patient cohorts.…”

Section: Microarray Analysis To Identify Predictive Biomarkersmentioning

confidence: 99%

Genomic approaches in the management and treatment of breast cancer

2005

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Breast cancer is the most common malignancy afflicting women from Western cultures. It has been estimated that approximately 211 000 women will be diagnosed with breast cancer in 2003 in the United States alone, and each year over 40 000 women will die of this disease. Developments in breast cancer molecular and cellular biology research have brought us closer to understanding the genetic basis of this disease. Unfortunately, this information has not yet been incorporated into the routine diagnosis and treatment of breast cancer in the clinic. Recent advancements in microarray technology hold the promise of further increasing our understanding of the complexity and heterogeneity of this disease, and providing new avenues for the prognostication and prediction of breast cancer outcomes. The most recent application of microarray genomic technologies to studying breast cancer will be the focus of this review. Mortality from breast cancer results from the ability of some tumours to metastasise to distant sites. Selecting patients with micrometastases at diagnosis is crucial for clinicians in deciding who should and who should not receive toxic and expensive adjuvant chemotherapy to eradicate these metastatic cells. Axillary nodal status, the best marker now available, is still an imperfect indicator, since about 25% of node-negative patients do harbour micrometastases and are destined to recur, while up to 25% of node-positive patients will not recur even without adjuvant treatment after many years of follow-up. In spite of extensive studies, expression of most individual genes has not proven powerful enough for routine clinical use to predict accurately distant metastases over the lifetime of an individual patient. However, recent developments in applying microarray technologies to breast tumour samples suggest that these new techniques may provide for the transition of molecular biological discoveries to clinical application, and will generate clinically useful genomic profiles that more accurately predict long-term outcome of individual breast cancer patients. NATURAL HISTORY OF BREAST CANCERBreast cancer is characterised by a very heterogeneous clinical course. A major goal of recent studies is to determine whether RNA microarray expression profiling, or DNA array gene amplification/gene loss patterns, can accurately predict an individual's long-term potential for recurrence from breast cancer, so that appropriate treatment decisions can be made. It is well established that some aspects of breast cancer heterogeneity is related to the different risk factors for diagnosis of this disease, such as race, diet, age, environmental factors, and cumulative exposure to the sex hormone oestrogen. The diversity in clinical course of breast cancer is undoubtedly related to differences in tumour growth rates, tumour invasiveness, metastatic potential, and other complex cellular growth signalling and survival pathways. It has long been held that knowledge of these various biological factors would help individualise patient t...

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A faithful method for PCR-mediated global mRNA amplification and its integration into microarray analysis on laser-captured cells

Cited by 85 publications

References 14 publications

GASDERMIN, suppressed frequently in gastric cancer, is a target of LMO1 in TGF-β-dependent apoptotic signalling

GASDERMIN, suppressed frequently in gastric cancer, is a target of LMO1 in TGF-β-dependent apoptotic signalling

Introduction to Microarray‐Based Detection Methods

Genomic approaches in the management and treatment of breast cancer

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