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Vitamin A and ß-carotene metabolism and effects of UV irradiation in human keratinocytes and melanocytes
Linköping University, Department of Biomedicine and Surgery, Dermatology. Linköping University, Faculty of Health Sciences.
2002 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Retinoids (vitamin A and its derivatives) are modulators of proliferation and differentiation. Both retinol (ROH) and its metabolite 3,4-didehydroretinol (ddROH) can be converted to retinoic acid (RA) and 3,4-didehydroretinoic acid (ddRA), ligands for the nuclear receptors, which induce gene transcriptions. A perturbed ROH metabolism is observed in several dermatoses and iu non-melanoma skin cancer. Dietary ß-carotene has been considered to play a critical role in the natural defence against cancer. Whether ß-carotene is converted to ROH in the skin has been debated.

We have investigated ß-carotene and retinoid metabolism, retinoid binding proteins and retinoid receptors in human keratinocytes (KCs) and melanocytes (MCs) in vitro. Similar studies of vitamin A have been done in human malignant epithelial cells (HeLa) and malignant melanoma cells. The influence of ultraviolet radiation (UVR) on retinoid metabolism and receptor expression was specially focused upon this thesis. KCs and MCs contained high concentrations of ROH, ddROH, while HeLa- and melanoma cells contained lower levels. KCs contained the highest level of the retinoid-binding proteins CRBP I and CRABP II compared to MCs, HeLa and melanoma cells. High CRABP II levels showed a correlation with the ability to accumulate ddROH. In MCs, CRABP I was highly expressed, but in melanoma cells CRABP II dominated. The difference between MCs and melanoma cells in receptor levels was most pronounced for RARß, which was highly expressed in melanoma cells. Such dissimilarities between benign and malignant MCs might play a role in differentiation and growth regulation. The uptake of [3H]ROH, [3H]RA and ß-carotene was significantly higher in MCs than in KCs. We were able to demonstrate that [14C]ß-carotene was converted to [14C]ROH in both these cell types. This suggests that this local storage of ß-carotene might serve as au alternative supply for vitamin A in the skin.

A moderate dose of UVR reduced the concentration of ROH, ddROH and [3H]RA in KCs and MCs by 20-50%. The concentration returned to starting levels in 1-2 days, and could be explained by a retarded metabolism of RA, the biologically most active metabolite. When KCs and MCs were exposed to UVR, the mRNA and protein levels of the three nuclear retinoid receptors (RARα, RARγ and RXRα) decreased rapidly. In MCs these levels were close to normal 3 days postirradiation. In KCs only the RARα mRNA and protein levels returned to baseline within 3 days. This thesis has increased our knowledge of the effects of UVR on retinoid metabolism and retinoid receptors in human cells. Further studies are needed to understand the role of ß-carotene and retinoid signaling in UV induced skin cancer.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet , 2002. , 66 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 725
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-25546Local ID: 9993ISBN: 91-7373-169-2 (print)OAI: oai:DiVA.org:liu-25546DiVA: diva2:245876
Public defence
2002-07-01, Berzeliussalen, Universitetssjukhuset, Linköping, 13:00 (Swedish)
Opponent
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2012-09-14Bibliographically approved
List of papers
1. The metabolism of vitamin A to 3,4-didehydroretinol can be demonstrated in human keratinocytes, melanoma cells and HeLa cells, and is correlated to cellular retinoid-binding protein expression
Open this publication in new window or tab >>The metabolism of vitamin A to 3,4-didehydroretinol can be demonstrated in human keratinocytes, melanoma cells and HeLa cells, and is correlated to cellular retinoid-binding protein expression
1994 (English)In: Biochimica et Biophysica Acta. Molecular Cell Research, ISSN 0167-4889, E-ISSN 1879-2596, Vol. 1224, no 3, 349-354 p.Article in journal (Refereed) Published
Abstract [en]

Conversion of retinol to 3,4-didehydroretinol is probably a rate-limiting step in the formation of 3,4-didehydroretinoic acid, a candidate ligand for nuclear retinoid receptors in human epidermal keratinocytes. To investigate whether this metabolic pathway also exists in other cell systems, we compared the retinoid concentrations and the bioconversion of [3H]retinol to [3H]3,4-didehydroretinol in human primary keratinocytes, human cervical carcinoma (HeLa) cells, human melanoma (JKM86-4) cells, monkey kidney epithelium (CV-1) cells, and murine teratocarcinoma (F9) cells. The cellular retinol concentration ranged from 2.33 to 99.1 pmol/mg protein with the highest values observed in keratinocytes. 3,4-Didehydroretinol was only detected in cells of human origin and its concentration ranged from 0.24 pmol/mg in HeLa to 34.6 pmol/mg in the keratinocytes. Incubation with [3H]retinol for 1–24 h resulted in a rapid appearance of [3H]3,4-didehydroretinol in human keratinocytes, and to a lesser extent in HeLa and melanoma cells, but not in the other cells. Analysis of cellular retinol- and retinoic acid-binding protein concentrations showed a correlation to the cells' ability to accumulate 3,4-didehydroretinol, suggesting a role for these proteins in the 3,4-didehydro metabolic pathway. The combined results suggest that although 3,4-didehydroretinol is most typical for human keratinocytes, studies of its metabolism are also feasible in HeLa cells which contain low levels of retinoid-binding proteins.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-81440 (URN)10.1016/0167-4889(94)90267-4 (DOI)
Available from: 2012-09-14 Created: 2012-09-14 Last updated: 2017-12-07Bibliographically approved
2. β-Carotene Uptake and Bioconversion to Retinol Differ Between Human Melanocytes and Keratinocytes
Open this publication in new window or tab >>β-Carotene Uptake and Bioconversion to Retinol Differ Between Human Melanocytes and Keratinocytes
2001 (English)In: Nutrition and Cancer, ISSN 0163-5581, E-ISSN 1532-7914, Vol. 39, no 2, 300-306 p.Article in journal (Refereed) Published
Abstract [en]

β-Carotene is one of the carotenoids that has been considered to play a role in the natural defense against ultraviolet-induced skin cancer. It is not known whether epidermal cells are able to accumulate β-carotene and, subsequently, convert it to vitamin A. We used normal cultured human keratinocytes and melanocytes to study the uptake, and possible bioconversion to retinol, of authentic or [14C]β-carotene. The uptake was much higher in melanocytes than in keratinocytes, corresponding to a fivefold difference in the intracellular fraction after two days of incubation. An increased level of cellular retinol was noted after one day of β-carotene incubation. The conversion of [C]β-carotene to [14C]retinol peaked at 24 hours of incubation in keratinocytes and melanocytes. The results suggest that β-carotene can function as a local supply of vitamin A in the skin and that melanocytes are especially likely to store β-carotene.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-49078 (URN)10.1207/S15327914nc392_21 (DOI)
Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2017-12-12Bibliographically approved
3. Vitamin A metabolism and mRNA expression of retinoid-binding protein and receptor genes in human epidermal melanocytes and melanoma cells
Open this publication in new window or tab >>Vitamin A metabolism and mRNA expression of retinoid-binding protein and receptor genes in human epidermal melanocytes and melanoma cells
1997 (English)In: Melanoma research, ISSN 0960-8931, E-ISSN 1473-5636, Vol. 7, no 4, 267-274 p.Article in journal (Refereed) Published
Abstract [en]

Retinoids inhibit proliferation of melanocytes and melanoma cells and affect disorders of hypo- and hyperpigmentation. Such effects might involve retinoid-binding proteins, retinoid metabolites and nuclear retinoid receptors for transcriptional activation. We detected messenger RNA transcripts for the cellular retinol- and retinoic acid-binding proteins (CRBP, CRABP I and II) in cultured epidermal melanocytes. In the melanoma cell lines the major transcript was CRABP II. Nuclear retinoic acid (RA) receptor transcripts and the 9-cis-retinoic acid receptor transcript were detected in all cells. The endogenous concentrations of retinol (ROH) and its metabolite 3,4-didehydroretinol (ddROH) in melanocytes were five times those in melanoma cells. When cells were incubated with [3H]ROH the main metabolites in the melanocytes were [3H]ddROH (4%) and [3H]RA (0.4%). Formation of [3H]RA was only detected in one melanoma cell line. Both melanocytes and melanoma cells produced an unidentified metabolite when incubated with [3H]ROH and [3H]RA. Dissimilarities in the metabolism and endogenous concentration of retinoids between benign and malignant melanocytes might play a key role in differentiation and growth regulation.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-81443 (URN)10.1097/00008390-199708000-00001 (DOI)
Available from: 2012-09-14 Created: 2012-09-14 Last updated: 2017-12-07Bibliographically approved
4. Ultraviolet irradiation depletes cellular retinol and alters the metabolism of retinoic acid in cultured human keratinocytes and melanocytes
Open this publication in new window or tab >>Ultraviolet irradiation depletes cellular retinol and alters the metabolism of retinoic acid in cultured human keratinocytes and melanocytes
1999 (English)In: Melanoma research, ISSN 0960-8931, E-ISSN 1473-5636, Vol. 9, no 4, 339-346 p.Article in journal (Refereed) Published
Abstract [en]

Vitamin A is an intrinsic modulator of proliferation and differentiation in human epidermis, and may be destroyed by ultraviolet radiation (UVR) impinging on the skin. To identify the deleterious effects of a perturbed cellular vitamin A status, we investigated the endogenous retinoid concentrations and the metabolism of [3H]retinol and all-trans [3H]retinoic acid in cultured human keratinocytes and melanocytes exposed to UVR, using high performance liquid chromatography. Before UVR the retinoid content was similar in keratinocytes and melanocytes, but the uptake of [3H]retinol was three-fold higher and the uptake of [3H]retinoic acid was 10-fold higher in the melanocytes. In both cell types, UVR (UVA 360 mJ/cm2 plus UVB 140 mJ/cm2) instantaneously reduced the concentration of retinol by about 50% and that of 3,4-didehydroretinol by about 20%. The retinoid concentrations returned to normal within 1-2 days post-irradiation, despite there being no overt increase in the uptake of [3H]retinol or the biosynthesis of 3,4- didehydroretinol. However, in both types of irradiated cells, the accumulation of the biologically most active metabolite, all-trans [3H]retinoic acid, was about 60% higher than in control cells. Furthermore, the metabolism of authentically supplied [3H]retinoic acid was reduced, especially in irradiated keratinocytes, which probably contributed to the restoration of retinoid levels after UV exposure.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-24867 (URN)10504051 (PubMedID)9269 (Local ID)9269 (Archive number)9269 (OAI)
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2017-12-13Bibliographically approved
5. Differential effects of UV irradiation on the nuclear retinoid receptor levels of cultured keratinocytes and melanocytes
Open this publication in new window or tab >>Differential effects of UV irradiation on the nuclear retinoid receptor levels of cultured keratinocytes and melanocytes
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Skin cancer is the most common malignancy in man. A major risk factor is UV irradiation, which not only damages DNA but may also perturb cellular signaling, e.g. via the retinoid receptor system believed to be important for cancer protection. We used cultured normal human keratinocytes and melanocytes to study the effects of UV radiation on the expression of the retinoid receptors RARα, RARβ, RARγ and RXRα. By real-time PCR and Western blot technique, the mRNA and protein levels were monitored, before and up to 4 days following 50 mJ/cm2 UVB. In keratinocytes, UVB caused a rapid drop in all four mRNA levels (minus 50-70% the first 8 h) and protein levels dropped by 30-40% followed by a gradual increase, but full normalization was ouly reached for RARα within the study period. ln melanocytes, UVB caused a quick drop both in the receptor mRNA and protein levels (minus 50-60% after 4 h), followed by normalization of the protein levels for all receptors within 2-3 days. The UV-induced depletion of vitamin A and retinoid receptors might abrogate the retinoid signaling, which subsequently might promote tumor development.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-81447 (URN)
Available from: 2012-09-14 Created: 2012-09-14 Last updated: 2012-09-14Bibliographically approved

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