Current bioinformatics tools in genomic biomedical research (Review)Teufel, Andreas ; Krupp, Markus ; Weinmann, Arndt ; Galle, Peter R.
doi: 10.3892/ijmm.17.6.967pmid: N/A
On the advent of a completely assembled human genome, modern biology and molecular medicine stepped into an era of increasingly rich sequence database information and high-throughput genomic analysis. However, as sequence entries in the major genomic databases currently rise exponentially, the gap between available, deposited sequence data and analysis by means of conventional molecular biology is rapidly widening, making new approaches of high-throughput genomic analysis necessary. At present, the only effective way to keep abreast of the dramatic increase in sequence and related information is to apply biocomputational approaches. Thus, over recent years, the field of bioinformatics has rapidly developed into an essential aid for genomic data analysis and powerful bioinformatics tools have been developed, many of them publicly available through the World Wide Web. In this review, we summarize and describe the basic bioinformatics tools for genomic research such as: genomic databases, genome browsers, tools for sequence alignment, single nucleotide polymorphism (SNP) databases, tools for ab initio gene prediction, expression databases, and algorithms for promoter prediction.
Adiponectin receptor expression in the human adrenal cortex and aldosterone-producing adenomasRossi, Gian Paolo; Sticchi, Daniele Paolo; Giuliani, Luisa Paolo; Bernante, Paolo Paolo; Zavattiero, Silvia Paolo; Pessina, Achille C.; Nussdorfer, Gastone G.
doi: 10.3892/ijmm.17.6.975pmid: N/A
Adiponectin is an adipocyte-derived circulating peptide that plays an important role in adipose tissue metabolism, insulin sensitivity and cardiovascular disease. The adrenal gland, by secreting glucorticoid and mineralocorticoid hormones, intervenes in cardiovascular and glucose metabolism regulation and is surrounded by adipose tissue. Hence, we investigated the hypothesis that adiponectin receptor types 1 and 2 (adipo-R1 and adipo-R2) are expressed in the human adrenal gland and in adrenocortical zona glomerulosa cell-derived aldosterone-producing adenoma (APA) tissue. We used real-time reverse transcription-polymerase chain reaction to demonstrate the mRNA of adipo-R1 and adipo-R2 in 10 histologically normal human adrenal cortexes that were obtained from patients with renal cancer undergoing nephrectomy with ipsilateral adrenalectomy and in 10 APAs. Melting curve analysis and sequencing were used to confirm the specificity of the amplicons obtained. Results consistently showed the expression of specific mRNAs of adiponectin receptors in all histologically normal human adrenal cortexes and APAs. This novel finding suggests that adiponectin could play a regulatory role in adrenocortical function and growth in humans.
An image analysis of the spatial distribution of perivascular mast cells in human melanomaGuidolin, Diego ; Crivellato, Enrico ; Nico, Beatrice ; Andreis, Paola G.; Nussdorfer, Gastone G.; Ribatti, Domenico G.
doi: 10.3892/ijmm.17.6.981pmid: N/A
A mutual spatial and functional relationship occurs between mast cells (MCs) and endothelial cells and the density of MCs is highly correlated with the extent of tumor angiogenesis. The aim of this study was to investigate the pattern of MCs around the blood vessels in melanoma samples by means of an approach derived from spatial statistics, based on the analysis of the distribution of the distances of MCs from vessels to objectively establish if the two structures (MCs and vessels) are distributed independently over the studied area or if they displayed any kind of spatial association. Results showed that a higher number of vessels and MCs can be observed in melanoma as compared with samples from common acquired nevi (control group). The percent of area covered by vessel profiles was significantly higher in the melanoma group than the control group and the MC density was also significantly different; the melanoma group showing a number of MCs per unit area twice as high as the number measured in the control group. Furthermore, in the melanoma group, MCs were closer to each other and to the vessels. In fact, both the mean distance from vessels and the mean distance from the nearest cell profile were significantly lower than in the control group. This close association between MCs and the endothelium does not necessarily imply a participation of MCs in angiogenic processes, but might rather indicate that MCs are involved in the maintenance reaction necessary for the long lasting functional integrity of the endothelium.
Expression of thioredoxin and thioredoxin-binding protein-2 in the liver of patients with chronic hepatitis C as a predictor of response to interferon therapyHamano, Kenichi ; Seo, Yasushi ; Kato, Hirotaka ; Kato, Miyuki ; Yano, Yoshihiko ; Ninomiya, Toshiaki ; Kasuga, Masato
doi: 10.3892/ijmm.17.6.989pmid: N/A
Oxidative stress contributes to the pathogenesis of various hepatic injuries. Thioredoxin (TRX) is an indicator of oxidative stress, reported to be increased in the serum of patients with chronic hepatitis C with the progression of fibrosis. The aim of this study was to evaluate the clinical significance of the expression of TRX and thioredoxin-binding protein-2 (TBP-2), which is a negative regulator of TRX function, in the liver of patients with chronic hepatitis C and the relationship of this to the efficacy of interferon (IFN) treatment. A retrospective study was performed using the liver biopsy specimens obtained before IFN treatment from 69 patients with chronic serotype 1 hepatitis C virus (HCV) infection. TRX and TBP-2 mRNA levels in the liver biopsy specimens were amplified by real-time RT-PCR. The serum TRX protein level was estimated with a sandwich enzyme-linked immunosorbent assay kit, and the expression of TRX protein in the liver was examined immunohistochemically in 19 patients. There was no association between the serum TRX level and the TRX level in the liver. There was a significant correlation between the expression level of TRX protein in the liver and the TRX mRNA level in the liver. TRX and TBP-2 levels in the liver tended to decrease slightly with increased fibrosis stage, although not significantly. The TRX level in the liver tended to increase with hepatitis activity index, although not significantly. TBP-2 mRNA levels in the liver were significantly higher in responders than non-responders to the IFN therapy (p<0.05). Among patients who had a high viral load of >850 KIU/ml, the TRX level in the livers of non-responders was significantly lower than that in the livers of responders (p<0.05). TRX and TBP-2 mRNA levels in the liver before IFN therapy may predict the outcome of IFN therapy in patients with chronic serotype 1 HCV infection.
Polysiphonia japonica extract suppresses the Wnt/β-catenin pathway in colon cancer cells by activation of NF-κBGwak, Jungsug ; Park, Seoyoung ; Cho, Munju ; Song, Taeyun ; Cha, Seon-Heui ; Kim, Dong-Eun ; Jeon, You-Jin ; Shin, Jae-Gook ; Oh, Sangtaek
doi: 10.3892/ijmm.17.6.1005pmid: N/A
Abnormal activation of the Wnt/β-catenin pathway and subsequent up-regulation of β-catenin response transcription (CRT) are associated with the development of colon cancer. Thus, the Wnt/β-catenin pathway is an attractive target for chemoprevention and treatment of this cancer. We used a cell-based screen to identify a methanol extract of Polysiphonia japonica (EPJ) that suppresses the Wnt/β-catenin pathway without altering the level of β-catenin protein and reduces the expression of cyclin D1, which is a known β-catenin/T cell factor (TCF)-dependent gene. EPJ inhibited the growth of various colon cancer cells. In addition, EPJ induced the nuclear translocation of nuclear factor-κB (NF-κB) in SW480 colon cancer cells. Our findings suggest that EPJ attenuates Wnt/β-catenin signaling via activation of NF-κB and can potentially be used as a chemopreventive agent against colon cancer.
Method for efficient transfection of in vitro-transcribed mRNA into SK-N-AS and HEK293 cells: Difference in the toxicity of nuclear EGFP compared to cytoplasmic EGFPEjeskär, Katarina ; Fransson, Susanne ; Zaibak, Faten ; Ioannou, Panayiotis A.
doi: 10.3892/ijmm.17.6.1011pmid: N/A
Here we report a method for efficient transfection of in vitro-transcribed mRNA into two different types of human adherent cells, the neuroblastoma cell line SK-N-AS, and the transformed kidney cell line HEK293. By using newly trypsinized adherent cells in suspension and Lipofectamine™ 2000, we detected a transfection efficiency of 80-90% in both cell lines and a cell viability of 90% in SK-N-AS and 60% in HEK293, 24 h after transfection when using cytoplasmic enhanced green fluorescent protein (EGFP)-mRNA. We have evaluated the different effects of the generally used EGFP that mainly localizes to the cytoplasm and nuclear EGFP, where the nuclear EGFP are more toxic to the cells than the cytoplasmic EGFP. In order to develop a null experiment, we constructed a short non-functional mRNA including a nuclear localization signal and evaluated the concentrations at which mRNA encoding nuclear proteins can be added without a general toxicity, depending on the fact that the proteins are localized to the nucleus. For both SK-N-AS and HEK293 cells, a concentration of up to 100 ng mRNA in 105 cells, encoding a nuclear protein with no other function, did not affect the cells. For evaluation of the method, we screened four different human mRNAs, PDG, DFFA, CORT and PEX14, for their ability to affect cell proliferation in these cells. PEX14 was the only gene that significantly (p=0.03) reduced cell proliferation for both cell types, DFFA significantly (p=0.04) reduced cell proliferation in SK-N-AS but not in HEK293 cells. PGD and CORT did not have any effect on cell proliferation. We have developed an easy method for efficient delivery of in vitro-transcribed mRNA into the adherent cell lines, SK-N-AS and HEK293. This method is useful for a quick screening of how different genes affect cell proliferation.
A novel MLH1 mutation harbored as a germ line aberration by a young woman of an HNPCC-like family and exhibited by a CML patient when occurring prior to the initiation of the blast phase concomitant with a c-MYC amplificationAmikam, Dorit ; Leshanski, Lucy ; Sagi, Michal
doi: 10.3892/ijmm.17.6.1023pmid: N/A
Germ line mutations in the MLH1 and MSH2 genes account for the majority of hereditary nonpolyposis colorectal cancer (HNPCC) families. Here, we describe a family that does not meet the international criteria for HNPCC, of which a young woman harbors a missense mutation (D132H). This novel germ line mutation has not previously been reported. Of the mismatch repair (MMR) genes, MLH1 has been shown to play an important role in hematologic malignancies. The novel mutation was also revealed to be a somatic aberration occurring prior to the initiation of the blast phase in a chronic myelogenous leukemia (CML) patient. Among the possible MLH1 partners involved in signaling MMR or apoptosis is the proto-oncogene c-MYC, which is closely related to cellular proliferation. We further revealed a concomitant c-MYC dramatic amplification in the CML-MLH1-mutation carrier patient, also occurring at the pre-blast phase. Our data contribute further to characterizing the mutational spectrum of the MLH1 gene. Furthermore, given the role of c-MYC and its interaction with MLH1, taken together with the mutational status of both genes revealed at the pre-blast phase in the CML patient, a plausible increased genetic instability might be expected to take place, possibly contributing to blast triggering. Our results may provide additional insight into the complex interplay between the MMR system and other cellular pathways.
The differential expression of SPARC in esophageal squamous cell carcinomaChe, Yiqun ; Luo, Aiping ; Wang, Hai ; Qi, Jun ; Guo, Jian ; Liu, Zhihua
doi: 10.3892/ijmm.17.6.1027pmid: N/A
Secreted protein, acidic and rich in cysteine (SPARC) is a matricellular protein that modulates cell adhesion and growth. The aim of this study was to assess the prevalence of SPARC mRNA and protein expression in esophageal squamous cell carcinoma (ESCC) and to investigate the role of SPARC in proliferation and metastasis of ESCC. The differential expression of SPARC between esophageal squamous cell carcinoma and its corresponding normal esophageal mucosa was analyzed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, as well as Western blotting. The expression of SPARC in ESCC was significantly higher than that in the corresponding normal esophageal mucosa as shown by semi-quantitative RT-PCR, immunohistochemistry and Western blotting. Meanwhile, SPARC protein was also significantly up-regulated in ESCC compared with the corresponding normal esophageal mucosa. SPARC protein was located in the cytoplasm and nuclei of ESCC cells. The results of Western blotting showed that out of 18 paired samples, SPARC was up-regulated in 13 cases, no significant differences were found for the other 5 cases. These findings revealed that SPARC was up-regulated in ESCC and was closely associated with ESCC metastasis. This study provides new insights into nuclei location of SPARC.