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- Edlundh-Rose, Esther, et al.
(författare)
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Gene expression analysis of human epidermal keratinocytes after N-acetyl L-cysteine treatment demonstrates cell cycle arrest and increased differentiation
- 2005
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Ingår i: Pathobiology (Basel). - : S. Karger AG. - 1015-2008 .- 1423-0291. ; 72:4, s. 203-212
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Tidskriftsartikel (refereegranskat)abstract
- Objectives: Several cancer prevention programmes have previously been executed using treatment of antioxidant compounds. The antioxidant N-acetyl L-cysteine (NAC), a membrane-permeable aminothiol, is a sulfhydryl reductant reducing oxidised glutathione, as well as being a precursor of intracellular cysteine and glutathione. A previous report based on the cellular response to NAC treatment showed that NAC induced a 10-fold more rapid differentiation in normal primary keratinocytes as well as a reversion of a colon carcinoma cell line from neoplastic proliferation to apical-basolateral differentiation. In order to investigate molecular events underlying the changes in proliferation and differentiation induced by NAC treatment, we performed global gene expression analysis of normal human epidermal keratinocytes in a time series. Methods: Treated samples were compared to untreated samples through a reference design using a spotted cDNA array comprising approximately 30,000 features. B statistics was used to identify differentially expressed genes, and RT-PCR of a selected set of genes was performed to verify differential expression. Results: The number of differentially expressed genes increased over time, starting with 0 at 30 min, 73 at 3 h and increasing to 952 genes at 48 h. Results of the expression analysis showed arrest of the cell cycle and an upregulation of cytoskeletal reorganisation, implicating increased differentiation. A comparison to gene ontology groups indicated downregulation of a large number of genes involved in cell proliferation and regulation of the cell cycle. Conclusions: A significant fraction of the differentially expressed genes could be classified according to their role in the differentiation process, demonstrating that NAC regulates the conversion from proliferation to differentiation at a transcriptional level.
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| 2. |
- Gustafsson, Anna, et al.
(författare)
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Global gene expression analysis in time series following N-acetyl L-cysteine induced epithelial differentiation of human normal and cancer cells in vitro
- 2005
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Ingår i: BMC Cancer. - : Springer Science and Business Media LLC. - 1471-2407. ; 5, s. 75-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Cancer prevention trials using different types of antioxidant supplements have been carried out at several occasions and one of the investigated compounds has been the antioxidant N-acetyl-L-cysteine (NAC). Studies at the cellular level have previously demonstrated that a single supplementation of NAC induces a ten-fold more rapid differentiation in normal primary human keratinocytes as well as a reversion of a colon carcinoma cell line from neoplastic proliferation to apical-basolateral differentiation [1]. The investigated cells showed an early change in the organization of the cytoskeleton, several newly established adherens junctions with E-cadherin/β-catenin complexes and increased focal adhesions, all features characterizing the differentiation process. Methods: In order to investigate the molecular mechanisms underlying the proliferation arrest and accelerated differentiation induced by NAC treatment of NHEK and Caco-2 cells in vitro, we performed global gene expression analysis of NAC treated cells in a time series (1, 12 and 24 hours post NAC treatment) using the Affymetrix GeneChip™ Human Genome U95Av2 chip, which contains approximately 12,000 previously characterized sequences. The treated samples were compared to the corresponding untreated culture at the same time point. Results: Microarray data analysis revealed an increasing number of differentially expressed transcripts over time upon NAC treatment. The early response (1 hour) was transient, while a constitutive trend was commonly found among genes differentially regulated at later time points (12 and 24 hours). Connections to the induction of differentiation and inhibition of growth were identified for a majority of up- and down-regulated genes. All of the observed transcriptional changes, except for seven genes, were unique to either cell line. Only one gene, ID-1, was mutually regulated at 1 hour post treatment and might represent a common mediator of early NAC action. The detection of several genes that previously have been identified as stimulated or repressed during the differentiation of NHEK and Caco-2 provided validation of results. In addition, real-time kinetic PCR analysis of selected genes also verified the differential regulation as identified by the microarray platform. Conclusion: NAC induces a limited and transient early response followed by a more consistent and extensively different expression at later time points in both the normal and cancer cell lines investigated. The responses are largely related to inhibition of proliferation and stimulation of differentiation in both cell types but are almost completely lineage specific. ID-1 is indicated as an early mediator of NAC action.
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| 3. |
- Kivi, Marten, et al.
(författare)
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Helicobacter pylori genome variability in a framework of familial transmission
- 2007
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Ingår i: BMC Microbiology. - : Springer Science and Business Media LLC. - 1471-2180. ; 7, s. 54-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Helicobacter pylori infection is exceptionally prevalent and is considered to be acquired primarily early in life through person-to-person transmission within the family. H. pylori is a genetically diverse bacterial species, which may facilitate adaptation to new hosts and persistence for decades. The present study aimed to explore the genetic diversity of clonal isolates from a mother and her three children in order to shed light on H. pylori transmission and host adaptation. Results: Two different H. pylori strains and strain variants were identified in the family members by PCR-based molecular typing and sequencing of five loci. Genome diversity was further assessed for 15 isolates by comparative microarray hybridizations. The microarray consisted of 1,745 oligonucleotides representing the genes of two previously sequenced H. pylori strains. The microarray analysis detected a limited mean number (+/- standard error) of divergent genes between clonal isolates from the same and different individuals (1 +/- 0.4, 0.1%, and 3 +/- 0.3, 0.2%, respectively). There was considerable variability between the two different strains in the family members (147 +/- 4, 8%) and for all isolates relative to the two sequenced reference strains (314 +/- 16, 18%). The diversity between different strains was associated with gene functional classes related to DNA metabolism and the cell envelope. Conclusion: The present data from clonal H. pylori isolates of family members do not support that transmission and host adaptation are associated with substantial sequence diversity in the bacterial genome. However, important phenotypic modifications may be determined by additional genetic mechanisms, such as phase-variation. Our findings can aid further exploration of H. pylori genetic diversity and adaptation.
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