FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2022670053> ?p ?o ?g. }

• W2022670053 endingPage "726" @default.
• W2022670053 startingPage "722" @default.
• W2022670053 abstract "The epidermis has a requirement for fatty acids in order to synthesize cellular membranes and the extracellular lipid lamellar membranes in the stratum corneum. Despite high endogenous production of fatty acids the transport of exogenous essential fatty acids into the epidermis is an absolute requirement. Fatty acid uptake by keratinocytes has been shown to be mediated by a transport system. In this study we determined in murine epidermis and human cultured keratinocytes the expression of three putative fatty acid transport related proteins and fatty acyl CoA synthase, an enzyme that facilitates the uptake of fatty acids by promoting their metabolism. In cultured human keratinocytes, mRNA for fatty acid transport protein (FATP), plasma membrane fatty acid binding protein (FABP-pm), and fatty acyl CoA synthase (FACS) were detectable. Differentiation, induced by high calcium, did not affect FATP mRNA levels, but resulted in an ≈50% increase in FACS mRNA, while decreasing FABP-pm mRNA by 50%. Fatty acid translocase (FAT) mRNA was not detected in cultured human keratinocytes. In murine epidermis, FATP, FABP-pm, FACS, and FAT mRNA were all present. Barrier disruption by either tape stripping or acetone treatment increased FAT mRNA levels by ≈2-fold without affecting FATP, FABP-pm, or FACS. Occlusion with an impermeable membrane immediately after barrier disruption completely blocked the increase in FAT mRNA levels, indicating that this increase is related to barrier disruption rather than a nonspecific injury effect. In summary, this study demonstrates that several putative fatty acid transport related proteins as well as fatty acyl CoA synthase are expressed in keratinocytes and epidermis, and that the expression of these proteins may be regulated by differentiation and/or barrier disruption. The epidermis has a requirement for fatty acids in order to synthesize cellular membranes and the extracellular lipid lamellar membranes in the stratum corneum. Despite high endogenous production of fatty acids the transport of exogenous essential fatty acids into the epidermis is an absolute requirement. Fatty acid uptake by keratinocytes has been shown to be mediated by a transport system. In this study we determined in murine epidermis and human cultured keratinocytes the expression of three putative fatty acid transport related proteins and fatty acyl CoA synthase, an enzyme that facilitates the uptake of fatty acids by promoting their metabolism. In cultured human keratinocytes, mRNA for fatty acid transport protein (FATP), plasma membrane fatty acid binding protein (FABP-pm), and fatty acyl CoA synthase (FACS) were detectable. Differentiation, induced by high calcium, did not affect FATP mRNA levels, but resulted in an ≈50% increase in FACS mRNA, while decreasing FABP-pm mRNA by 50%. Fatty acid translocase (FAT) mRNA was not detected in cultured human keratinocytes. In murine epidermis, FATP, FABP-pm, FACS, and FAT mRNA were all present. Barrier disruption by either tape stripping or acetone treatment increased FAT mRNA levels by ≈2-fold without affecting FATP, FABP-pm, or FACS. Occlusion with an impermeable membrane immediately after barrier disruption completely blocked the increase in FAT mRNA levels, indicating that this increase is related to barrier disruption rather than a nonspecific injury effect. In summary, this study demonstrates that several putative fatty acid transport related proteins as well as fatty acyl CoA synthase are expressed in keratinocytes and epidermis, and that the expression of these proteins may be regulated by differentiation and/or barrier disruption. epidermal fatty acid binding protein fatty acid binding protein plasma membrane fatty acid binding protein psoriasis-associated fatty acidbinding protein fatty acid translocase fatty acid transport protein transepidermal water loss The epidermis has a requirement for fatty acids in order to synthesize cellular membranes and the extracellular lipid enriched lamellar membranes in the stratum corneum that are responsible for the barrier to water transit (Schurer and Elias, 1991Schurer N.Y. Elias P.M. The biochemistry and function of stratum corneum lipids.Adv Lipid Rev. 1991; 24: 27-50Crossref PubMed Google Scholar). Previous studies by our laboratory and others have shown that the epidermis is a very active site of de novo fatty acid synthesis (Feingold, 1991Feingold K.R. The regulation and role of epidermal lipid synthesis.Adv Lipid Res. 1991; 24: 57-82Crossref PubMed Google Scholar). Moreover, disruption of the permeability barrier results in a 2–3-fold increase in de novo fatty acid synthesis in the epidermis (Feingold, 1991Feingold K.R. The regulation and role of epidermal lipid synthesis.Adv Lipid Res. 1991; 24: 57-82Crossref PubMed Google Scholar;Proksch et al., 1993Proksch E. Holleran W.M. Menon G.K. Elias P.M. Feingold K.R. Barrier function regulates epidermal lipid and DNA synthesis.Br J Dermatol. 1993; 128: 473-482Crossref PubMed Scopus (193) Google Scholar). Furthermore, inhibition of this increase in fatty acid synthesis delays barrier repair. Despite the high endogenous production of fatty acids in the epidermis, the transport of exogenous fatty acids into the epidermis is an absolute requirement as the epidermis requires essential fatty acids and arachidonic acid (Schurer and Elias, 1991Schurer N.Y. Elias P.M. The biochemistry and function of stratum corneum lipids.Adv Lipid Rev. 1991; 24: 27-50Crossref PubMed Google Scholar). Essential fatty acid deficiency is characterized by epidermal hyperplasia, scaly skin, and increased transepidermal water loss (TEWL) (Prottey, 1976Prottey C. Essential fatty acids and the skin.Br J Dermatol. 1976; 94: 549-587Google Scholar;Elias and Brown, 1978Elias P.M. Brown B.E. The mammalian cutaneous permeability barrier: defective barrier function in essential fatty acid deficiency correlates with abnormal intercellular lipid composition.Lab Invest. 1978; 39: 574-583PubMed Google Scholar). In the stratum corneum linoleic acid is present in ω-esterified ceramides 1 and 4, which are localized to the lamellar bilayers in the stratum corneum and are required for normal barrier function (Schurer and Elias, 1991Schurer N.Y. Elias P.M. The biochemistry and function of stratum corneum lipids.Adv Lipid Rev. 1991; 24: 27-50Crossref PubMed Google Scholar;Downing, 1992Downing D.T. Lipid and protein structures in the permeability barrier of mammalian epidermis.J Lipid Res. 1992; 33: 301-313Abstract Full Text PDF PubMed Google Scholar). In essential fatty acid deficiency, oleic acid substitutes for linoleate in acyl ceramides, resulting in the disruption of normal membrane structures and an incompetent barrier (Wertz et al., 1983Wertz P.W. Cho E.S. Downing D.T. Effect of essential fatty acid deficiency on the epidermal sphingolipids of the rat.Biochim Biophys Acta. 1983; 753: 350-355Crossref PubMed Scopus (115) Google Scholar). In addition to linoleic acid, the epidermis also requires exogenous arachidonic acid (C20:4, n-6), as the enzyme Δ6 desaturase required for arachidonic acid synthesis is not present in the epidermis (Chapkin and Ziboh, 1984Chapkin R.S. Ziboh V.A. Inability of skin enzyme preparations to biosynthesize arachidonic acid from linoleic acid.Biochem Biophys Res Commun. 1984; 124: 784-792Crossref PubMed Scopus (62) Google Scholar). Arachidonic acid is the required substrate for eicosanoid synthesis (Sprecher, 1981Sprecher H.W. Biochemistry of essential fatty acids.Prog Lipid Res. 1981; 20: 13-22Crossref PubMed Scopus (260) Google Scholar). Essential fatty acids are not synthesized by mammalian cells and therefore must be acquired from dietary sources. Fatty acid uptake by keratinocytes has been demonstrated to be mediated by a transport system that is temperature sensitive, has saturable kinetics, and can be reduced by prior treatment with trypsin, which indicates that plasma membrane proteins mediate fatty acid uptake (Schurer et al., 1994Schurer N.Y. Stremmel W. Grundmann J.U. Schliep V. Kleinert H. Bass W.M. Williams M.L. Evidence for a novel keratinocyte fatty acid uptake mechanism with preference for linoleic acid: comparison of oleic and linoleic acid uptake by cultured human keratinocytes, fibroblasts, and a human hepatoma cell line.Biochim Biophys Acta. 1994; 1211: 51-60Crossref PubMed Scopus (50) Google Scholar). Moreover, keratinocyte fatty acid uptake demonstrated a higher specificity for the essential fatty acids, i.e., linoleic acid and arachidonic acid, than for nonessential fatty acids, such as oleic acid (C18:1). These transport characteristics are not shared with other cell types, such as hepatocytes, HepG2 cells, and dermal fibroblasts (Schurer et al., 1994Schurer N.Y. Stremmel W. Grundmann J.U. Schliep V. Kleinert H. Bass W.M. Williams M.L. Evidence for a novel keratinocyte fatty acid uptake mechanism with preference for linoleic acid: comparison of oleic and linoleic acid uptake by cultured human keratinocytes, fibroblasts, and a human hepatoma cell line.Biochim Biophys Acta. 1994; 1211: 51-60Crossref PubMed Scopus (50) Google Scholar), which transport nonessential and essential fatty acids with similar kinetics. The identity of the proteins responsible for fatty acid uptake in keratinocytes has not been established. Recently, there have been a number of advances in our understanding of fatty acid transport into cells. In other cell types, three distinct proteins have been shown to mediate the uptake of long chain fatty acids across plasma membranes (Schaffer and Lodish, 1995Schaffer J.E. Lodish H.F. Molecular mechanism of long chain fatty acid uptake.Trends Cardiovas Med. 1995; 5: 218-224Abstract Full Text PDF PubMed Scopus (45) Google Scholar). Plasma membrane fatty acid binding protein (FABP-pm) is a 43 kDa protein that is identical to mitochondrial aspartate aminotransferase (Isola et al., 1995Isola L.M. Zhou S.L. Kiang C.L. Stump D.D. Bradbury M.W. Berk P.D. 3T3 fibroblasts transfected with a cDNA for mitochondrial aspartate aminotransferase express plasma membrane fatty acid binding protein and saturable fatty acid uptake.Proc Natl Acad Sci. 1995; 92: 9866-9870Crossref PubMed Scopus (137) Google Scholar). This protein is expressed in tissues with high fatty acid uptake, such as the liver and adipose tissue, and levels of FABP-pm expression correlate with fatty acid transport (Stremmel et al., 1985Stremmel W. Strohmeyer G. Borchard F. Kochwa S. Berk P.D. Isolation and partial characterization of fatty acid binding protein in rat liver plasma membranes.Proc Natl Acad Sci. 1985; 82: 4-8Crossref PubMed Scopus (281) Google Scholar;Zhou et al., 1995Zhou S.L. Stump D. Kiang C.L. Isola L.M. Berk P.D. Mitochondrial aspartate aminotransferase expressed on the surface of 3T3–L1 adipocytes mediates saturable fatty acid uptake.Proc Soc Exp Biol Med. 1995; 208: 263-270Crossref PubMed Scopus (48) Google Scholar;Berk et al., 1997Berk P.D. Zhou S.L. Kiang C.L. Stump D. Bradbury M. Isola L.M. Uptake of long chain free fatty acids is selectively up-regulated in adipocytes of Zucker rats with genetic obesity and non-insulin dependent diabetes mellitus.J Biol Chem. 1997; 272: 8830-8835Crossref PubMed Scopus (120) Google Scholar). Moreover, antibodies to FABP-pm inhibit fatty acid transport into cells (Stremmel et al., 1985Stremmel W. Strohmeyer G. Borchard F. Kochwa S. Berk P.D. Isolation and partial characterization of fatty acid binding protein in rat liver plasma membranes.Proc Natl Acad Sci. 1985; 82: 4-8Crossref PubMed Scopus (281) Google Scholar,Stremmel et al., 1986Stremmel W. Strohmeyer G. Berk P.D. Hepatocellular uptake of oleate is energy dependent, sodium linked, and inhibited by an antibody to a hepatocyte plasma membrane fatty acid binding protein.Proc Natl Acad Sci. 1986; 83: 3584-3588Crossref PubMed Scopus (184) Google Scholar). Fatty acid translocase (FAT) is an 88 kDa glycoprotein that has high homology to human CD36 (Abumrad et al., 1993Abumrad N.A. el-maghrabi M.R. Amri E.Z. Lopez E. Grimaldi P.A. Cloning of a rat adipocyte membrane protein implicated in binding or transport of long chain fatty acids that is induced during preadipocyte differentiation. Homology with human CD36.J Biol Chem. 1993; 268: 17665-17668Abstract Full Text PDF PubMed Google Scholar). FAT is associated with the plasma membrane and is present in tissues with high fatty acid transport, such as the muscle, heart, intestine, and adipose tissue (Abumrad et al., 1993Abumrad N.A. el-maghrabi M.R. Amri E.Z. Lopez E. Grimaldi P.A. Cloning of a rat adipocyte membrane protein implicated in binding or transport of long chain fatty acids that is induced during preadipocyte differentiation. Homology with human CD36.J Biol Chem. 1993; 268: 17665-17668Abstract Full Text PDF PubMed Google Scholar;Van Nieuwenhoven et al., 1995Van Nieuwenhoven F.A. Verstijnen C.P. Abumrad N.A. Williamson P.H. Van Eys G.J. Van der Vusse G.J. Glatz J.F. Putative membrane fatty acid translocase and cytoplasmic fatty acid binding protein are co-expressed in rat heart and skeletal muscles.Biochem Biophys Res Commun. 1995; 207: 747-752Crossref PubMed Scopus (128) Google Scholar). The level of FAT expression in these tissues correlates with fatty acid transport. Moreover, overexpression of FAT in cells increases fatty acid uptake (Ibrahimi et al., 1996Ibrahimi A.Z. Sfeir Z. Magharaic H. Amri E.Z. Grimaldi P. Abumrad N.A. Expression of the CD36 homolog (FAT) in fibroblast cells: effects on fatty acid transport.Proc Natl Acad Sci. 1996; 93: 2646-2651Crossref PubMed Scopus (201) Google Scholar). Lastly, fatty acid transport protein (FATP) is a 63 kDa protein that has been shown to be present in 3T3-L-1 adipocytes, adipose tissue, heart, and muscle, with low levels in kidney, liver, brain, and lung (Schaffer and Lodish, 1994Schaffer J.E. Lodish H.F. Expression cloning and characterization of a novel adipocyte long chain fatty acid transport protein.Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (711) Google Scholar). FATP is localized to the plasma membrane and overexpression of this protein increases the uptake of long chain fatty acids (Schaffer and Lodish, 1994Schaffer J.E. Lodish H.F. Expression cloning and characterization of a novel adipocyte long chain fatty acid transport protein.Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (711) Google Scholar). FATP expression increases with differentiation in 3T3-L-1 adipocytes and is negatively regulated by insulin (Schaffer and Lodish, 1994Schaffer J.E. Lodish H.F. Expression cloning and characterization of a novel adipocyte long chain fatty acid transport protein.Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (711) Google Scholar;Man et al., 1996Man M.Z. Hui T.Y. Schaffer J.E. Lodish H.F. Bernlohr D.A. Regulation of the murine adipocyte fatty acid transporter gene by insulin.Molec Endocrinol. 1996; 10: 1021-1028Crossref PubMed Scopus (67) Google Scholar). Whether any of these three distinct, putative fatty acid transport proteins are expressed in keratinocytes is not known. In addition to these transport proteins, the enzyme fatty acyl CoA synthase (FACS) is also thought to play an important role in cellular fatty acid uptake (Schaffer and Lodish, 1995Schaffer J.E. Lodish H.F. Molecular mechanism of long chain fatty acid uptake.Trends Cardiovas Med. 1995; 5: 218-224Abstract Full Text PDF PubMed Scopus (45) Google Scholar). This enzyme catalyzes the activation of long chain fatty acids to acyl-CoA esters, which are required for both the oxidation of fatty acids and the esterification of fatty acids to glycerol to form triglycerides or phospholipids (Kornberg and Pricer, 1953Kornberg A. Pricer Jr, We Enzymatic synthesis of the coenzyme A derivatives of long chain fatty acids.J Biol Chem. 1953; 204: 329-343Abstract Full Text PDF PubMed Google Scholar). The formation of acyl-CoA by preventing the efflux of fatty acids, assures that transport is a unidirectional process. Furthermore, expression cloning studies in adipose cells have shown that FACS is a protein that increases fatty acid uptake into cells (Schaffer and Lodish, 1994Schaffer J.E. Lodish H.F. Expression cloning and characterization of a novel adipocyte long chain fatty acid transport protein.Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (711) Google Scholar). The purpose of this study was to determine (i) which of these proteins FABP-pm, FAT, FATP, and FACS are expressed in murine epidermis and in human keratinocytes grown in culture; (ii) if differentiation changes the expression of these proteins in cultured keratinocytes; and (iii) if the expression of these proteins is altered by disruption of the permeability barrier, a condition that we have previously shown markedly upregulates de novo fatty acid synthesis in epidermis. Hairless male mice (Crl:SKHI-hrBR) were purchased from Charles River Laboratories (Wilmington, MA). Molecular biology grade chemicals were purchased from Sigma (St. Louis, MO) and Fischer Scientific (Fairlawn, NJ). [a-32P] dCTP (3000 Ci per mmol, 10 mCi per ml) was purchased from NEN Research Products (Boston, MA). The Multiprime Labeling System was purchased from Amersham International (Amersham, U.K.). Mini-spin columns (G-50) were purchased from Worthington Biochemical (Freehold, NJ). Oligo(dT)-cellulose, type 77F, was purchased from Pharmacia LKB Biotechnology AB (Uppsala, Sweden). Nytran Plus membrane was purchased from Schleicher and Schuell (Keene, NH). Spin-X centrifuge filters were purchased from Corning Costar (Cambridge, MA). cDNA for rat FATP was kindly provided by J.E. Schaffer (Washington University, St Louis, MO). cDNA for rat FABP-pm was kindly provided by J.R. Mattingly (University of Missouri, Kansas City, MO). cDNA for rat FAT was kindly provided by N. Abumrad (SUNY, Stony Brook, NY). cDNA for rat cyclophilin cDNA (pCD15:8–1) was kindly provided by Dr. G. Strewler, (Harvard Medical School, MA). cDNA for FACS was kindly provided by Dr. P. Smith (Abbott Labs, Columbus, OH). Fuji RX film was used for autoradiography. Human foreskin keratinocytes, second passage, were seeded and maintained in 0.07 mM Ca++ KGM. When the cells attached, the culture medium was changed to either 0.03 mM or 1.2 mM Ca++. In the presence of 1.2 mM Ca++ the keratinocytes were induced to differentiate. Cells were harvested when 80% confluent and 100% confluent, respectively. The skin of 6–8 wk old male hairless mice was treated either by gently applying acetone-soaked cotton balls for 5–10 min or by sequential applications of cellophane tape (Tesa Tuck, New Rochelle, NY) as described previously (Menon et al., 1985Menon G.K. Feingold K.R. Moser A.H. Brown B.E. Elias P.M. De novo sterologenesis in the skin: fate and function of newly synthesized lipids.J Lipid Res. 1985; 26: 418-427Abstract Full Text PDF PubMed Google Scholar;Proksch et al., 1990Proksch E. Elias P.M. Feingold K.R. Regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity in murine epidermis: modularion of enzyme content and activation state by barrier requirements.J Clin Invest. 1990; 85: 874-882Crossref PubMed Scopus (94) Google Scholar). Controls for acetone perturbation of the barrier were treated with cotton balls soaked in 0.9% (wt/vol) sodium chloride. Untreated animals served as controls for tape stripping. The TEWL was measured immediately after treatment using a Meeco electrolytic water analyzer, as described previously (Menon et al., 1985Menon G.K. Feingold K.R. Moser A.H. Brown B.E. Elias P.M. De novo sterologenesis in the skin: fate and function of newly synthesized lipids.J Lipid Res. 1985; 26: 418-427Abstract Full Text PDF PubMed Google Scholar). Animals with a TEWL rate greater than 6 mg per cm2 per h (normal < 0.3 mg per cm2 per h) after barrier disruption were included in this study. In some experiments mice were covered immediately after barrier disruption by a finger from a latex glove to occlude the disrupted area as described previously (Grubauer et al., 1989Grubauer G. Elias P.M. Feingold K.R. Transepidermal water loss: the signal for recovery of barrier structure and function.J Lipid Res. 1989; 30: 323-334Abstract Full Text PDF PubMed Google Scholar). Four hours following barrier disruption, the animals were killed by Isoflurane anaethesia (Abbot Laboratories, Chicago, IL) and the skin was excised. The subcutaneous fat was removed by scraping with a scalpel blade. To separate whole epidermis, skin was placed in 10 mM ethylenediamine tetraacetic acid in calcium and magnesium free PBS pH 7.4 for 35 min at 37°C (Jackson et al., 1992Jackson S.M. Wood L.C. Lauer S. Taylor J.M. Cooper A.D. Elias P.M. Feingold K.R. Effect of cutaneous permeability barrier disruption on HMG CoA reductase, LDL receptor and apoprotein E mRNA levels in the epidermis of hairless mice.J Lipid Res. 1992; 33: 1307-1314Abstract Full Text PDF PubMed Google Scholar). The skin was blotted dry and the epidermis was removed by scraping with a scalpel blade. FACS activity was measured by a radio-isotopic assay in keratinocytes by the method ofTanaka et al., 1979Tanaka T. Hosaka K. Hoshimaru M. Numa S. Purification and properties of long chain acyl-CoA synthetase from rat liver.Eur J Bioc. 1979; 98: 165-172Crossref PubMed Scopus (172) Google Scholar. Keratinocytes were homogenized in a buffer (0.25 M sucrose, 1 mM dithiothreitol, 1 mM ethylenediamine tetraacetic acid) and the homogenates were centrifuged at 800 × g for 5 min. The resultant supernatants were used for measuring enzyme activity. As reported by others (Tanaka et al., 1979Tanaka T. Hosaka K. Hoshimaru M. Numa S. Purification and properties of long chain acyl-CoA synthetase from rat liver.Eur J Bioc. 1979; 98: 165-172Crossref PubMed Scopus (172) Google Scholar), our initial experiments also showed that the rate of reaction is linear up to 10 min and plateaued by 15 min. Hence, in all subsequent assays the reaction time was kept at 5 min. The assay mixture contained 0.1 M Tris-HCl buffer pH 8.0, 5 mM dithiothreitol, 0.15 M KCl, 15 mM MgCl2, 1.6 mM triton X-100, 10 mM ATP, 1 mM coenzyme A, and 1 mM [U-14C] palmitic acid in a final reaction volume of 0.2 ml. The assay was initiated by addition of 5–10 μl of enzyme suspension (5 μg protein) and the reaction was carried out at 37°C for 5 min. The reaction was terminated by adding 2.5 ml of a mixture containing isopropanol:heptane:sulfuric acid 1 M (40:10:1 vol/vol/vol). The unreacted palmitic acid was extracted with heptane and the radioactivity in the aqueous phase containing palmitoyl-CoA was measured. Control reactions containing no protein were run in parallel to correct for the background. FACS activity is expressed as nmoles palmitoyl-CoA formed per mg protein per min. Protein was assayed by the method of Bradford (Bio-Rad laboratories). Poly (A)+ RNA was isolated using a variation of the proteinase-K extraction method. Briefly, cells from two 100 mm dishes were washed with phosphate-buffered saline and scraped into 5 ml solution A (0.5 M NaCl, 10 mM Tris pH 7.5, 1 mM ethylenediamine tetraacetic acid, 1% sodium dodecyl sulfate, and 200 mg proteinase-K per ml). The viscosity of the solution was reduced by passing through a 25-gauge needle, and then incubating for 1 h at 37°C. Oligo(dT)-cellulose, 7.5 mg, was added to each sample and incubated for 1 h. The oligo(dT)-cellulose was washed and the poly (A)+ RNA eluted. Total RNA was prepared by a variation of the guanidinium thiocyanate method, as described previously (Jackson et al., 1992Jackson S.M. Wood L.C. Lauer S. Taylor J.M. Cooper A.D. Elias P.M. Feingold K.R. Effect of cutaneous permeability barrier disruption on HMG CoA reductase, LDL receptor and apoprotein E mRNA levels in the epidermis of hairless mice.J Lipid Res. 1992; 33: 1307-1314Abstract Full Text PDF PubMed Google Scholar;Wood et al., 1992Wood L.C. Jackson S.M. Elias P.M. Grunfeld C. Feingold K.R. Cutaneous barrier perturbation stimulates cytokine production in the epidermis of mice.J Clin Invest. 1992; 90: 482-487Crossref PubMed Scopus (387) Google Scholar). Total RNA was purified and added to oligo(dT)-cellulose to obtain poly (A)+ RNA. Northern blots were prepared as described previously. The uniformity of sample applications was checked by ultraviolet visualization of the acridine orange-stained gel before transfer to Nytran membranes. In order to avoid any potential problems with RNA transfer from the gels, the exposure to acridine orange was kept to a minimum by staining the gels for only 1 min followed by 1 h of destaining. The blots were exposed to X-ray films for various durations to ensure that measurements were done on the linear portion of the curve, and the bands were quantitated by densitometry (Bio-Rad Laboratories, Hercules, CA). Quantitation of film, exposed in the linear range of sensitivity, was achieved using a Biorad laboratories (Hercules, CA) densitometer. The densitometry quantitation was adjusted for cyclophilin levels on the same blot and expressed as fold of control, with the control as one. Statistical significance was determined using a two-tailed unpaired Student’s t test. Results are expressed as mean ± SEM. As shown in Figure 1(a), FATP, FABP-pm, and FACS mRNA were detectable in human keratinocytes. The sizes of the mRNA transcripts for FATP, FABP-pm, and FACS in keratinocytes were identical to those previously reported in other tissues (Abumrad et al., 1993Abumrad N.A. el-maghrabi M.R. Amri E.Z. Lopez E. Grimaldi P.A. Cloning of a rat adipocyte membrane protein implicated in binding or transport of long chain fatty acids that is induced during preadipocyte differentiation. Homology with human CD36.J Biol Chem. 1993; 268: 17665-17668Abstract Full Text PDF PubMed Google Scholar;Schaffer and Lodish, 1994Schaffer J.E. Lodish H.F. Expression cloning and characterization of a novel adipocyte long chain fatty acid transport protein.Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (711) Google Scholar;Zhou et al., 1995Zhou S.L. Stump D. Kiang C.L. Isola L.M. Berk P.D. Mitochondrial aspartate aminotransferase expressed on the surface of 3T3–L1 adipocytes mediates saturable fatty acid uptake.Proc Soc Exp Biol Med. 1995; 208: 263-270Crossref PubMed Scopus (48) Google Scholar;Berk et al., 1997Berk P.D. Zhou S.L. Kiang C.L. Stump D. Bradbury M. Isola L.M. Uptake of long chain free fatty acids is selectively up-regulated in adipocytes of Zucker rats with genetic obesity and non-insulin dependent diabetes mellitus.J Biol Chem. 1997; 272: 8830-8835Crossref PubMed Scopus (120) Google Scholar). In contrast, FAT mRNA was not observed by northern blotting in human keratinocytes. The effect of differentiation was compared by growing human keratinocyte cultures in either 0.03 or 1.2 mM calcium. In high calcium, FAT mRNA was again not detected in human keratinocytes. Differentiation did not effect FATP mRNA levels and resulted in an ≈50% decrease in FABP-pm mRNA levels Figure 1b. In contrast, FACS mRNA levels increased by 70% under high calcium conditions Figure 1b. Additionally, FACS activity was increased 45% in differentiated keratinocytes (low calcium 108.3 ± 7.0 versus high calcium 157.0 ± 11.8 nmoles per mg per min, p < 0.01, n = 6). FATP, FABP-pm, FACS, and FAT mRNA were all present in murine epidermis Figure 2. The transcript sizes of these mRNA were identical to those previously reported in other tissues and in keratinocytes. To determine whether the expression of the mRNA for these proteins in the epidermis is regulated by conditions that increase epidermal fatty acid requirements, we next measured the mRNA levels in the epidermis following barrier disruption. We have previously shown that barrier disruption increases de novo fatty acid synthesis and that inhibition of this increase in synthesis delays barrier recovery (Grubauer et al., 1987Grubauer G. Feingold K.R. Elias P.M. The relationship of epidermal lipogenesis to cutaneous barrier function.J Lipid Res. 1987; 28: 746-752Abstract Full Text PDF PubMed Google Scholar;Man et al., 1993Man M.Q. Elias P.M. Feingold K.R. Fatty acids are required for epidermal barrier function.J Clin Invest. 1993; 92: 792-798Google Scholar;Ottey et al., 1995Ottey K.A. Wood L.C. Grunfeld C. Elias P.M. Feingold K.R. Cutaneous permeability barrier disruption increases fatty acid synthetic enzyme activity in the epidermis of hairless mice.J Invest Dermatol. 1995; 104: 401-405Crossref PubMed Scopus (45) Google Scholar). Barrier disruption by acetone treatment did not significantly affect FATP, FABP-pm, or FACS mRNA levels Figure 3; however, FAT mRNA levels increased ≈2-fold Figure 3. FAT has both 2.9 and 4.8 kb transcripts and both transcripts of FAT increase following barrier disruption. To determine if the increase in FAT mRNA was due to barrier disruption or a nonspecific effect of acetone, we next disrupted the barrier by an entirely different method, tape stripping. As found with acetone treatment, tape stripping did not alter FATP, FABP-pm, or FACS mRNA levels (data not shown). As observed following acetone treatment, however, tape stripping increased both the 2.9 and the 4.8 kb mRNA transcripts for FAT Figure 4. To further link the increase in FAT mRNA levels with barrier disruption we next artificially provided a permeability barrier by immediately occluding the disrupted skin with an impermeable membrane. As shown in Figure 4, occlusion completely blocked the increase in both the 2.9 and the 4.8 kb FAT mRNA transcripts that occur following tape stripping. These data indicate that the increase in FAT mRNA that occurs after barrier disruption is not a nonspecific effect secondary to injury, but rather is related to barrier disruption per se. In this study, we demonstrate that two of the three putative fatty acid transporters, FABP-pm and FATP, are expressed in human keratinocytes in culture. In contrast, FAT expression was not found i" @default.
• W2022670053 created "2016-06-24" @default.
• W2022670053 creator A5020684279 @default.
• W2022670053 creator A5048434963 @default.
• W2022670053 creator A5050608918 @default.
• W2022670053 creator A5051847291 @default.
• W2022670053 creator A5054659472 @default.
• W2022670053 creator A5055517539 @default.
• W2022670053 date "1998-11-01" @default.
• W2022670053 modified "2023-09-30" @default.
• W2022670053 title "Expression and Regulation of mRNA for Putative Fatty Acid Transport Related Proteins and Fatty Acyl CoA Synthase in Murine Epidermis and Cultured Human Keratinocytes" @default.
• W2022670053 cites W1512516272 @default.
• W2022670053 cites W1556697438 @default.
• W2022670053 cites W1796779887 @default.
• W2022670053 cites W180788089 @default.
• W2022670053 cites W1872675327 @default.
• W2022670053 cites W1963927869 @default.
• W2022670053 cites W1985698954 @default.
• W2022670053 cites W1987203691 @default.
• W2022670053 cites W1991713765 @default.
• W2022670053 cites W1993839824 @default.
• W2022670053 cites W1995502632 @default.
• W2022670053 cites W1998222131 @default.
• W2022670053 cites W1999047621 @default.
• W2022670053 cites W2000699501 @default.
• W2022670053 cites W2000833589 @default.
• W2022670053 cites W2001496481 @default.
• W2022670053 cites W2011455390 @default.
• W2022670053 cites W2028385745 @default.
• W2022670053 cites W2028957084 @default.
• W2022670053 cites W2030814140 @default.
• W2022670053 cites W2032740831 @default.
• W2022670053 cites W2036720565 @default.
• W2022670053 cites W2045479963 @default.
• W2022670053 cites W2049223762 @default.
• W2022670053 cites W2056392613 @default.
• W2022670053 cites W2059435029 @default.
• W2022670053 cites W2066156775 @default.
• W2022670053 cites W2074392865 @default.
• W2022670053 cites W2091708390 @default.
• W2022670053 cites W2092448980 @default.
• W2022670053 cites W2094091679 @default.
• W2022670053 cites W2106511392 @default.
• W2022670053 cites W2119201362 @default.
• W2022670053 cites W2120030741 @default.
• W2022670053 cites W2126394796 @default.
• W2022670053 cites W2129471415 @default.
• W2022670053 cites W2166021357 @default.
• W2022670053 cites W2168359294 @default.
• W2022670053 cites W2254937121 @default.
• W2022670053 cites W2321228822 @default.
• W2022670053 cites W4234167212 @default.
• W2022670053 cites W48316491 @default.
• W2022670053 cites W75618519 @default.
• W2022670053 doi "https://doi.org/10.1046/j.1523-1747.1998.00383.x" @default.
• W2022670053 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/9804328" @default.
• W2022670053 hasPublicationYear "1998" @default.
• W2022670053 type Work @default.
• W2022670053 sameAs 2022670053 @default.
• W2022670053 citedByCount "22" @default.
• W2022670053 countsByYear W20226700532012 @default.
• W2022670053 countsByYear W20226700532013 @default.
• W2022670053 countsByYear W20226700532014 @default.
• W2022670053 countsByYear W20226700532016 @default.
• W2022670053 crossrefType "journal-article" @default.
• W2022670053 hasAuthorship W2022670053A5020684279 @default.
• W2022670053 hasAuthorship W2022670053A5048434963 @default.
• W2022670053 hasAuthorship W2022670053A5050608918 @default.
• W2022670053 hasAuthorship W2022670053A5051847291 @default.
• W2022670053 hasAuthorship W2022670053A5054659472 @default.
• W2022670053 hasAuthorship W2022670053A5055517539 @default.
• W2022670053 hasBestOaLocation W20226700531 @default.
• W2022670053 hasConcept C104317684 @default.
• W2022670053 hasConcept C105580179 @default.
• W2022670053 hasConcept C105702510 @default.
• W2022670053 hasConcept C112243037 @default.
• W2022670053 hasConcept C153911025 @default.
• W2022670053 hasConcept C181199279 @default.
• W2022670053 hasConcept C2776458125 @default.
• W2022670053 hasConcept C2776834966 @default.
• W2022670053 hasConcept C48289651 @default.
• W2022670053 hasConcept C543025807 @default.
• W2022670053 hasConcept C55493867 @default.
• W2022670053 hasConcept C86803240 @default.
• W2022670053 hasConcept C95444343 @default.
• W2022670053 hasConceptScore W2022670053C104317684 @default.
• W2022670053 hasConceptScore W2022670053C105580179 @default.
• W2022670053 hasConceptScore W2022670053C105702510 @default.
• W2022670053 hasConceptScore W2022670053C112243037 @default.
• W2022670053 hasConceptScore W2022670053C153911025 @default.
• W2022670053 hasConceptScore W2022670053C181199279 @default.
• W2022670053 hasConceptScore W2022670053C2776458125 @default.
• W2022670053 hasConceptScore W2022670053C2776834966 @default.
• W2022670053 hasConceptScore W2022670053C48289651 @default.
• W2022670053 hasConceptScore W2022670053C543025807 @default.
• W2022670053 hasConceptScore W2022670053C55493867 @default.
• W2022670053 hasConceptScore W2022670053C86803240 @default.
• W2022670053 hasConceptScore W2022670053C95444343 @default.

• next