β-Carotene Uptake and Bioconversion to Retinol Differ Between Human Melanocytes and Keratinocytes
Uppsala University, Uppsala, Uppsala, Sweden Nutrition and Cancer
(Impact Factor: 2.32).
02/2001; 39(2):300-6. DOI: 10.1207/S15327914nc392_21
beta-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 beta-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]beta-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 beta-carotene incubation. The conversion of [14C]beta-carotene to [14C]retinol peaked at 24 hours of incubation in keratinocytes and melanocytes. The results suggest that beta-carotene can function as a local supply of vitamin A in the skin and that melanocytes are especially likely to store beta-carotene.
Available from: Artur Cieslar-Pobuda
- "( 94030304 ; Sigma - Aldrich , St . Louis , MO , USA ) were cultured in Eagle ' s minimum essential medium ( EMEM ) GlutaMAX , supplemented with 50 IU / ml penicillin - G , 50 µg / ml streptomycin , and 10% fetal bovine serum ( all from Gibco , Paisley , UK ) . Melanocytes were kindly provided by Petra Wäster and cultured as described previously ( Andersson et al . , 2001 ) . Human cervical cancer cells HeLa ( CCL - 2 ; ATCC ) , breast cancer cells MDA - MB - 231 ( HTB - 26 ; ATCC ) and human colon cancer HCT - 116 ( CCL - 247 ; ATCC ) were cultured in Dulbecco ' s modified eagle medium GlutaMAX supplemented with 50 IU / ml penicillin - G , 50 µg / ml streptomycin and 10% fetal bovine serum . Cells were "
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ABSTRACT: Molecular tools for fluorescent imaging of cells and their components are vital for understanding the function and activity of cells. Here, we report an imidazole functionalized pentameric oligothiophene, p-HTIm, that can be utilized for fluorescent imaging of cells. p-HTIm fluorescence in normal cells appeared in a peripheral punctate pattern partially co-localized with lysosomes, whereas a one-sided perinuclear Golgi associated localization of the dye was observed in malignant cells. The uptake of p-HTIm was temperature dependent and the intracellular target was reached within 1 h after staining. The ability of p-HTIm to stain cells was reduced when the imidazole side chain was chemically altered, verifying that specific imidazole side-chain functionalities are necessary for achieving the observed cellular staining. Our findings confirm that properly functionalized oligothiophenes can be utilized as fluorescent tools for vital staining of cells and that the selectivity toward distinct intracellular targets are highly dependent on the side-chain functionalities along the conjugated thiophene backbone.
Available from: Petra Wäster
- "All experiments were performed according to the ethical principles of the Helsinki declaration and approved by the Ethical Committee at Linkö ping University, Linkö ping, Sweden. Melanocytes were obtained from Caucasian donors (0–3 years of age) by means of foreskin circumcisions and pure cultures were established as described previously (Andersson et al., 2001). The melanocytes were cultured in Medium 199 with 2% fetal bovine serum and supplemented with 50 U/ml penicillin, 50 mg/ml streptomycin, 50 mg/ ml fungizone, 10 ng/ml basic fibroblast growth factor, 10 mg/ml inositol (all from Invitrogen, Paisley, Scotland, UK), 10 mg/ml insulin, 0.1 nM cholera toxin, 0.4 mg/ml hydrocortisone, 1 nM triiodothyronine , 10 mg/ml transferrin (all from Sigma Aldrich, St Louis, MO), and 10 ng/ml epidermal growth factor (Roche Diagnostics, Mannheim, Germany) (Gilchrest et al., 1984). "
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ABSTRACT: We demonstrate UVA/B to induce apoptosis in human melanocytes through the mitochondrial pathway, displaying cytochrome c release, caspase-3 activation, and fragmentation of nuclei. The outcome of a death signal depends on the balance between positive and negative apoptotic regulators, such as members of the Bcl-2 protein family. Apoptotic melanocytes, containing fragmented nucleus, show translocation of the proapoptotic proteins Bax and Bid from the cytosol to punctate mitochondrial-like structures. Bcl-2, generally thought to be attached only to membranes, was in melanocytes localized in the cytosol as well. In the fraction of surviving melanocytes, that is, cells with morphologically unchanged nucleus, the antiapoptotic proteins Bcl-2 and Bcl-X(L) were translocated to mitochondria following UVA/B. The lysosomal proteases, cathepsin B and D, which may act as proapoptotic mediators, were released from lysosomes to the cytosol after UVA/B exposure. Proapoptotic action of the cytosolic cathepsins was confirmed by microinjection of cathepsin B, which induced nuclear fragmentation. Bax translocation and apoptosis were markedly reduced in melanocytes after pretreatment with either cysteine or aspartic cathepsin inhibitors. No initial caspase-8 activity was detected, excluding involvement of the death receptor pathway. Altogether, our results emphasize translocation of Bcl-2 family proteins to have central regulatory functions of UV-induced apoptosis in melanocytes and suggest cathepsins to be proapoptotic mediators operating upstream of Bax.
Available from: rero.ch
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ABSTRACT: L'épiderme humain contient des rétinoides endogènes et des caroténoïdes. Nous présentons une méthode d'évaluation de la pénétration et du métabolisme des rétinoïdes (acide rétinoïque, rétinaldéhyde, rétinol, palmitate-de-rétinyle) dans la peau humaine "ex-vivo" à partir de peau totale incubée dans des cellules de Franz. Les résultats montrent qu'il n'y a pas de métabolisme pour l'acide rétinoïque, forme active de la vitamine A qui lie les récepteurs nucléaires, alors qu'on observe un métabolisme d'oxydation et de réduction pour le rétinaldehyde, ainsi qu'une augmentation des rétinoïdes endogènes pour le rétinol et ses esters. Nous mettons en évidence pour la première fois que le β-carotène est un précurseur de la vitamine A dans l'épiderme humain. Nous montrons un effet filtre de la vitamine A (palmitate-de-rétinyle) "in vivo" chez l'homme qui contribue potentiellement à la protection de l'ADN contre les effets des UVB.
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