Gene expression in buccal keratinocytes with emphasis on carbonyl metabolism
Author: Staab, Claudia A
Date: 2008-10-31
Location: Hillarpsalen, Retzius väg 8, Karolinska Institutet, Campus Solna
Time: 09.00
Department: Institutionen för medicinsk biokemi och biofysik (MBB) / Department of Medical Biochemistry and Biophysics
View/ Open:
Thesis (1.472Mb)
Abstract
The inner lining of the cheek, the buccal mucosa, is a target for
air-borne, dietary and tobacco usage-derived carcinogens, but also
interesting from a drug delivery point of view. Cancer arising in the
buccal epithelium, buccal squamous cell carcinoma (BSCC), often diagnosed
at a late disease stage, is highly aggressive and recurrent, emphasizing
the need for novel approaches in diagnosis and therapy.
An in vitro model for human buccal carcinogenesis consisting of normal buccal keratinocytes (NBK) and two transformed cell lines of buccal origin was applied toexplore mechanisms of buccal carcinogenesis, tumor marker and drug target expression. Two-dimensional gel electrophoresis, DNA microarray analysis, and the application of three bioinformatics programs for data mining allowed for the identification of multiple established and potential novel markers for BSCC, including tumor promoter/suppressor genes. Furthermore, post-confluent culture of NBK in absence and presence of fetal bovine serum was successfully used to induce terminal squamous differentiation (TSD) to various extents and thus enrich for different strata of the epithelium. Here, expression and activity of carbonyl-metabolizing enzymes (CMEs) were assessed in view of their multiple roles in phase I biotransformation. The combined results put forward ADH3 transcription as a proliferation marker as well as ALDH4A1, AKR1A1 and HPGD transcription as markers for the onset of TSD. Finally, the findings suggested increased CME-mediated xenobiotic activity at the onset of TSD, with potential implications for drug response and BSCC susceptibility.
In addition to its function as a ubiquitous formaldehyde scavenger, including in the buccal mucosa, alcohol dehydrogenase 3 (ADH3) irreversibly reduces S-nitrosoglutathione (GSNO) which implies an important role in nitric oxide homeostasis. Assessment of ADH3 in terms of formaldehyde/GSNO competition provided indirect evidence for formaldehyde as a physiological trigger of ADH3-mediated GSNO depletion with possible direct implications for asthma, where formaldehyde is known for exacerbating and GSNO for protective effects. Moreover, GSNO reduction resulted in the glutathione-controlled formation of glutathione transferase inhibitors, thus possibly affecting phase II biotransformation under conditions of oxidative stress. In light of these ADH3-mediated adverse effects, several substrate analogues were tested as ADH3 inhibitors. The results provide guidelines for future design of a specific ADH3 inhibitor with potential direct clinical use for the prevention of asthmaexacerbating effects.
An in vitro model for human buccal carcinogenesis consisting of normal buccal keratinocytes (NBK) and two transformed cell lines of buccal origin was applied toexplore mechanisms of buccal carcinogenesis, tumor marker and drug target expression. Two-dimensional gel electrophoresis, DNA microarray analysis, and the application of three bioinformatics programs for data mining allowed for the identification of multiple established and potential novel markers for BSCC, including tumor promoter/suppressor genes. Furthermore, post-confluent culture of NBK in absence and presence of fetal bovine serum was successfully used to induce terminal squamous differentiation (TSD) to various extents and thus enrich for different strata of the epithelium. Here, expression and activity of carbonyl-metabolizing enzymes (CMEs) were assessed in view of their multiple roles in phase I biotransformation. The combined results put forward ADH3 transcription as a proliferation marker as well as ALDH4A1, AKR1A1 and HPGD transcription as markers for the onset of TSD. Finally, the findings suggested increased CME-mediated xenobiotic activity at the onset of TSD, with potential implications for drug response and BSCC susceptibility.
In addition to its function as a ubiquitous formaldehyde scavenger, including in the buccal mucosa, alcohol dehydrogenase 3 (ADH3) irreversibly reduces S-nitrosoglutathione (GSNO) which implies an important role in nitric oxide homeostasis. Assessment of ADH3 in terms of formaldehyde/GSNO competition provided indirect evidence for formaldehyde as a physiological trigger of ADH3-mediated GSNO depletion with possible direct implications for asthma, where formaldehyde is known for exacerbating and GSNO for protective effects. Moreover, GSNO reduction resulted in the glutathione-controlled formation of glutathione transferase inhibitors, thus possibly affecting phase II biotransformation under conditions of oxidative stress. In light of these ADH3-mediated adverse effects, several substrate analogues were tested as ADH3 inhibitors. The results provide guidelines for future design of a specific ADH3 inhibitor with potential direct clinical use for the prevention of asthmaexacerbating effects.
List of papers:
I. Staab CA, Ceder R, Jägerbrink T, Nilsson JA, Roberg K, Jörnvall H, Höög JO, Grafström RC (2007). Bioinformatics processing of protein and transcript profiles of normal and transformed cell lines indicates functional impairment of transcriptional regulators in buccal carcinoma. J Proteome Res. 6(9): 3705-17.
Pubmed
View record in Web of Science®
II. Nilsson JA, Hedberg JJ, Vondracek M, Staab CA, Hansson A, Höög JO, Grafström RC (2004). Alcohol dehydrogenase 3 transcription associates with proliferation of human oral keratinocytes. Cell Mol Life Sci. 61(5): 610-7.
Pubmed
View record in Web of Science®
III. Staab CA, Ceder R, Roberg K, Grafström RC, Höög JO (2008). Serum-responsive gene expression of carbonyl-metabolizing enzymes in cultured human buccal keratinocytes. Cell Mol Life Sci. [Accepted]
Pubmed
View record in Web of Science®
IV. Staab CA, Alander J, Brandt M, Lengqvist J, Morgenstern R, Grafström RC, Höög JO (2008). Reduction of S-nitrosoglutathione by alcohol dehydrogenase 3 is facilitated by substrate alcohols via direct cofactor recycling and leads to GSH-controlled formation of glutathione transferase inhibitors. Biochem J. 413(3): 493-504.
Pubmed
View record in Web of Science®
V. Staab CA, Hellgren M, Grafström RC, Höög JO (2008). Medium-chain fatty acids and glutathione derivatives as inhibitors of alcohol dehydrogenase 3-mediated GSNO reduction. [Submitted]
I. Staab CA, Ceder R, Jägerbrink T, Nilsson JA, Roberg K, Jörnvall H, Höög JO, Grafström RC (2007). Bioinformatics processing of protein and transcript profiles of normal and transformed cell lines indicates functional impairment of transcriptional regulators in buccal carcinoma. J Proteome Res. 6(9): 3705-17.
Pubmed
View record in Web of Science®
II. Nilsson JA, Hedberg JJ, Vondracek M, Staab CA, Hansson A, Höög JO, Grafström RC (2004). Alcohol dehydrogenase 3 transcription associates with proliferation of human oral keratinocytes. Cell Mol Life Sci. 61(5): 610-7.
Pubmed
View record in Web of Science®
III. Staab CA, Ceder R, Roberg K, Grafström RC, Höög JO (2008). Serum-responsive gene expression of carbonyl-metabolizing enzymes in cultured human buccal keratinocytes. Cell Mol Life Sci. [Accepted]
Pubmed
View record in Web of Science®
IV. Staab CA, Alander J, Brandt M, Lengqvist J, Morgenstern R, Grafström RC, Höög JO (2008). Reduction of S-nitrosoglutathione by alcohol dehydrogenase 3 is facilitated by substrate alcohols via direct cofactor recycling and leads to GSH-controlled formation of glutathione transferase inhibitors. Biochem J. 413(3): 493-504.
Pubmed
View record in Web of Science®
V. Staab CA, Hellgren M, Grafström RC, Höög JO (2008). Medium-chain fatty acids and glutathione derivatives as inhibitors of alcohol dehydrogenase 3-mediated GSNO reduction. [Submitted]
Issue date: 2008-10-10
Rights:
Publication year: 2008
ISBN: 978-91-7409-127-4
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