[13] Analysis of water-soluble compounds: glucuronides

Ž. Ž . Absorption and fluorescence studies of retinol vitamin A alcohol and retinol palmitate vitamin A palmitate Ž . intercalated in phosphatidylcholine PC liposomes show that these compounds are bound to the lipid bilayer. It is further found that retinol binds liposomes with greater affinity as compared to retinol palmitate. In addition, the delivery of liposome-incorporated retinoids to the blood has also been studied and it is found that these systems reduce blood viscosity and cause less lysis of red blood cells than retinoid compounds not complexed in liposomes.

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“…Steady state fluorescence studies were performed on a Spex Spectrofluorolog instrument equipped with a 16810. 22 …”

Section: Materials and General Proceduresmentioning

confidence: 99%

Liposome encapsulated vitamin A compounds exhibit greater stability and diminished toxicity

Singh

Das

1998

Biophysical Chemistry

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“…Whereas the human keratinocytes were cultured in a serum-free medium [23,48], the murine cell culture medium contained 10% fetal calf serum. Even the latter is subphysiological with regard to its content of ROL and RA, which in both serum and skin is assumed to be in the range of 1 µM for ROL and 150-200 times less for RA [49]. In both cases, strong transactivation of the reporter gene could, however, be achieved when the transfected cells were exposed to 0.1 µM RA [23,48] (and this paper).…”

Section: Discussionmentioning

confidence: 95%

“…The activation could be inhibited by citral, thus implicating ROL→RA converting enzyme activity under low-Ca 2+ conditions [51]. Because the ROL content of skin is assumed to be in the micromolar range [49], transcription of RARE-controlled genes in basal epidermal cells can a priori not be excluded. RA-dependent responses in epidermal cells are, however, fully induced by maximal activation of the ROL→RA converting enzyme system upon entry of the cells into differentiation.…”

Section: Data)mentioning

confidence: 99%

Retinoic acid signaling cascade in differentiating murine epidermal keratinocytes: Alterations in papilloma- and carcinoma-derived cell lines

et al. 1997

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The retinoic acid (RA) signaling pathway was investigated by transient transfection of a chloramphenicol acetyltransferase (CAT) reporter gene construct containing the RA response element (RARE) of the murine (m) RARbeta2 gene into murine primary epidermal keratinocytes (PEK), papilloma-derived SP1 cells, and carcinoma-derived 3P2 cells. Murine PEK transfected in a low-Ca2+ medium (0.05 mM Ca2+) exhibited a strong transactivation of the CATgene after exposure of the cells to 0.1 microM RA. Transactivation of the CATgene could, however, also be achieved by shifting RAREbeta2-transfected low-Ca2+ PEK to high-Ca2+ conditions (0.15-1.2 mM Ca2+). Concomitantly, the Ca2+ raise also led to the induction of both cellular retinol (ROL)-binding protein I (CRBPI) and cellular RA-binding protein II (CRABPII), whereas expression of cellular RA-binding protein I (CRABPI) was not observed. Moreover, induction of in vitro differentiation also activated the ROL-->RA converting enzyme system in PEK. These findings suggest the following sequence of events involved in the high Ca2+-mediated activation of RAREbeta2. First, high Ca2+ induces the synthesis of mCRBPI, which binds ROL released from retinyl ester stores and makes it accessible to the ROL-RA converting enzyme system. Enzymatically generated RA is taken over by mCRABPII and transported to the nucleus, where it acts as ligand for nuclear receptors, which complex with RAREbeta2 to activate the reporter gene. This hypothetical cascade of RA signaling was supported by our findings that inhibition of the ROL-->RA converting enzyme system by citral abolished the Ca2+-mediated transactivation of the CAT gene in a nontoxic manner. Studies in transformed murine cell lines revealed that Ca2+-induced activation of RAREbeta2 was essentially maintained in papilloma-derived SP1 cells, although all parameters of the Ca2+-dependent RAREbeta2 activation cascade were induced to a much lower extent. In contrast, strong RAREbeta2 activity was already observed in low-Ca2+ carcinoma-derived 3P2 cells. Low-Ca2+ 3P2 cells also expressed high levels of both mCRBPI and mCRABPII and possessed a highly active ROL-->RA converting enzyme system. Again, inhibition of the enzyme by citral abolished RAREbeta2 activity in low-Ca2+ 3P2 cells. Our data show that Ca2+-induced differentiation in cultured murine PEK entails a series of events that ultimately lead to the activation of RARE-containing genes. These properties are maintained in transformed epidermal keratinocytes. However, with increasing malignant potential of the cells, the respective signaling pathway becomes independent from a differentiation stimulus and leads to constitutive activation of RARE-controlled genes.

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“…Retinoids were resolved by reverse-phase high performance liquid chromatography (HPLC) using a 3.9 ϫ 300-mm C18 Novapak reverse-phase analytical column. The mobile phase consisted of a 20-min linear gradient of 10 mM ammonium acetate in methanol:water (68:32, v:v.) to methanol:dichloromethane (4:1, v:v), followed by a 10-min hold (Barua, 1991). The flow rate was 1.0 ml/min.…”

Section: Retinoid Analysismentioning

confidence: 99%

“…Cells were washed twice with isotonic phosphate prior to lipid extraction (Barua, 1991). Retinoids were resolved by reverse-phase high performance liquid chromatography (HPLC) using a 3.9 ϫ 300-mm C18 Novapak reverse-phase analytical column.…”

Section: Retinoid Analysismentioning

confidence: 99%

Epidermal growth factor signaling pathway influences retinoid metabolism by reduction of retinyl ester hydrolase activities in normal and malignant keratinocytes

Jurukovski

Simon

2000

J. Cell. Physiol.

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The effects of EGFR signaling on retinol metabolism were evaluated in the squamous cell carcinoma cell lines defective in LRAT. In a 24-h incubation, the presence of EGF resulted in a 20-25% increase in retinyl ester accumulation. Assessment of retinol esterification and retinyl ester utilization (hydrolysis), in cell cultures and in cell homogenates, revealed that the increase in retinyl ester mass was the result of a reduction in retinyl ester hydrolysis. When grown in the absence of EGF, the cultures used about 40% of their retinyl esters, compared to about 21% in cultures grown with EGF. This effect of EGF was blocked by an EGF receptor-neutralizing antibody, an EGF receptor tyrosine-kinase inhibitor (PD153035), and a specific inhibitor of MEK kinase influencing the mitogen-activated protein kinase (MAPK) cascade (PD98059). Both transcription and translation were required, suggesting that signaling from the EGF receptor through the MAPK cascade controls the expression of modulators or inhibitors of the retinyl ester hydrolase(s). Thus EGFR signaling can alter the intracellular concentration of retinol by suppressing the access to the retinyl ester pool. Similar EGF effects were seen in cultures of normal keratinocytes.

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[13] Analysis of water-soluble compounds: glucuronides

Cited by 30 publications

References 13 publications

Liposome encapsulated vitamin A compounds exhibit greater stability and diminished toxicity

Liposome encapsulated vitamin A compounds exhibit greater stability and diminished toxicity

Retinoic acid signaling cascade in differentiating murine epidermal keratinocytes: Alterations in papilloma- and carcinoma-derived cell lines

Epidermal growth factor signaling pathway influences retinoid metabolism by reduction of retinyl ester hydrolase activities in normal and malignant keratinocytes

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