PubMed Health⌕ Search

Biomedical subjects

R Blomhoff

Publications and source records attributed to R Blomhoff.

At least 55 records · Page 3Linked to original sources
PubMed1995

Tissue distribution of the receptor for plasma retinol-binding protein.

The tissue distribution of the retinol-binding-protein receptor has been studied by using a cell-free binding assay. High binding activity was found in placenta, retina pigment epithelial cells, bone marrow and kidneys. Specific binding activity was also found in the small intestines, spleen and liver, and to a lesser extent in lung. Scatchard analysis revealed that the difference in binding activity was due to variations in receptor level and not affinity changes. When the kidneys were separated into cortex and medulla we found that almost all the specific binding activity present in kidneys was recovered in the cortex. The choroid plexus, an important site in the delivery of nutrients to the cerebrospinal fluid, expressed very high binding activity. The pineal gland, which has been shown to store vitamin A, also showed high binding activity. Testes from immature animals showed higher binding activity than testes from mature rabbits. Cultured undifferentiated kidney keratinocytes showed about 40 times higher binding activity than differentiated cells. Skin fibroblasts demonstrated no binding activity. In conclusion, the data presented in this report show that the level of the retinol-binding-protein receptor varies considerably between cell types. The observed tissue distribution of the receptor agrees well with the present knowledge on retinol function and metabolism by various cells.

Animals↗
PubMed1995

Effect of a coffee lipid (cafestol) on regulation of lipid metabolism in CaCo-2 cells.

The influence of cafestol, a lipid component found in boiled coffee, on low density lipoprotein (LDL) and lipid metabolism was investigated in CaCo-2 cells cultured on filter membranes. The rate of uptake and degradation of 125I-labeled tyramine cellobiose-LDL was increased 50% in CaCo-2 cells incubated with cafestol (20 micrograms/ml, 63 microM) for 24 h, whereas in cells incubated with 25-hydroxycholesterol (10 micrograms/ml, 25 microM) the rate of uptake and degradation showed a 30% decrease. A mixture of kahweol and cafestol, both natural components of coffee beans, modestly enhanced the rate of LDL uptake and degradation, as compared to pure cafestol. Incubation of cafestol with CaCo-2 cells induced a 3-fold up-regulation of LDL receptor mRNA, as compared to control cells. In contrast, incubation of the cells with 25-hydroxycholesterol produced a 30% decrease of LDL receptor expression. CaCo-2 cells were transfected with a promoter region containing the sterol regulatory element-1 (SRE-1) coupled to the reporter gene chloramphenicol acetyltransferase (CAT). When cells transfected with SRE-1 promoter were incubated with cafestol, there was a 20% up-regulation of CAT activity, whereas 25-hydroxycholesterol abolished this activity. Cafestol contributed to a significantly lowered secretion of cholesteryl ester and triacylglycerol, regardless of the radiolabeled precursor used ([2-14C]acetic acid, [1,2,3-3H]glycerol, [3H]water, and [1-14C]oleic acid). This reduction in secretion of lipids was accompanied by an increase in trichloroacetic acid-soluble activity when radiolabeled oleic acid was used as a tracer. We conclude that cafestol promotes an enhanced rate of uptake and degradation of LDL, probably due to an increase in transcription of LDL receptor mRNA and a reduced secretion of cholesteryl ester and triacylglycerol in CaCo-2 cells.

Cell Line↗
PubMed1994

Effect of n-3 and n-6 fatty acids on proliferation and differentiation of promyelocytic leukemic HL-60 cells.

Promyelocytic leukemic HL-60 cells were incubated with different fatty acids. Arachidonic acid (AA; 20:4, n-6) and eicosapentaenoic acid (EPA; 20:5, n-3) were the most potent inhibitors of proliferation in a dose-dependent way. Retinoic acid (RA) was used as a positive control. Inhibitors of cyclooxygenase and lipoxygenase or addition of antioxidants did not influence the effect of EPA or AA on cell proliferation. Increased capacity to generate superoxide anions after phorbol ester treatment and a reduced serglycin messenger RNA level in cells treated with AA or EPA indicated that these fatty acids induced differentiation in HL-60 cells similar to that induced by RA. However, down-regulation of the c-myc mRNA level, also typical for differentiation with RA in HL-60 cells, was not observed in cells incubated with AA or EPA. Flow cytometric analyses showed that in cultures incubated with AA or EPA, the proportion of cells in the G1 phase of the cell cycle increased. Similar effects were observed with RA. By flow cytometry and light scatter analyses it could be shown that AA made 8% of the cells apoptotic and 7% necrotic. The corresponding numbers were 21% and 10% for RA-treated cells, and 19% and 32% for EPA-treated cells. The present study shows that AA and EPA reduce the proliferation rate of HL-60 cells. This is mediated by mechanisms independent of eicosanoids or lipid peroxidation products and is due to effects both on apoptosis/necrosis and cell differentiation.

Apoptosis↗
PubMed1994

All-trans retinoic acid directly inhibits granulocyte colony-stimulating factor-induced proliferation of CD34+ human hematopoietic progenitor cells.

In this study we examine the effects of retinoids on purified CD34+ human hematopoietic progenitor cells. All-trans retinoic acid inhibited granulocyte colony-stimulating factor (G-CSF)-induced proliferation of CD34+ cells in short-term liquid cultures in a dose-dependent fashion with maximal inhibition of 72% at a concentration of retinoic acid of 1 mumol/L. Although no significant effects were observed on granulocyte-macrophage CSF (GM-CSF)--interleukin-3--or stem cell factor (SCF)-induced proliferation, the combinations of G-CSF and each of these cytokines were all inhibited. Moreover, retinol (3 mumol/L) and chylomicron remnant retinyl esters (0.1 mumol/L) in concentrations normally found in human plasma also had inhibitory effects. Single-cell experiments showed that the effects of retinoic acid were directly mediated. Retinoids also significantly inhibited G-CSF-induced colony formation in semisolid medium, with 88% inhibition observed at a concentration of retinoic acid of 1 mumol/L. However, we did not observe any effects of retinoic acid on G-CSF-induced differentiation as assessed by morphology and flowcytometry. Similar to previous findings using total bone marrow mononuclear cells, we observed a stimulation of GM-CSF-induced colony formation after 14 days. We also observed a stimulatory effect of low doses of retinoic acid (30 nmol/L) on blast-cell colony formation on stromal cell layers. Taken together, the data indicate that vitamin A present in human plasma has inhibitory as well as stimulatory effects on myelopoiesis.

Adult↗
PubMed1994

Characterization of liver stellate cell retinyl ester storage.

The stellate cells of the liver are the main storage site of retinyl esters in the body. During cultivation in vitro of stellate cells isolated from rat and rabbit livers were observed that the cells rapidly loose their retinyl ester content. Freshly isolated stellate cells contain about 144 nmol of total retinol/mg of protein, while cells cultivated for 14 days contained below 0.1 nmol/mg of protein. When 3-day-old cultures were incubated for 6 h with 2 microM retinol, the cellular content increased from 5.6 to approx. 9.4 nmol of total retinyl esters/mg of protein. In contrast, little retinyl ester accumulated in 10-20-day-old cultures incubated with 2 microM retinol. At 50 microM retinol, however, the retinyl ester level did increase both with 3-day-old cultures and 10-20-day-old cultures. In parallel experiments with cultured fibroblasts esterification characteristics similar to those seen in older cultures of stellate cells were observed. When 10-day-old cultures of stellate cells were incubated with retinol alone, or in combination with palmitic acid, linoleic acid or oleic acid, the total storage of retinyl esters increased by 20-150%. In most cases, the fatty acid supplemented in the medium was found to be the dominant fatty acid esterified with retinol. Cultures of stellate cells were then exposed to a physiological concentration (1.3 microM) of radioactive retinol free in solution or bound to retinol-binding protein. With 3-day-old cultures, as well as older cultures, the cellular content of unesterified retinol was 10-20 times higher when free retinol was added compared with addition of retinol bound to retinol-binding protein. However, 2-3-fold as much radioactive retinyl esters were recovered in cells incubated with retinol-retinol-binding protein compared with retinol free in solution. These results show that retinol delivered to stellate cells from retinol-binding protein is preferentially esterified, and that the complex is handled differently to free retinol by the stellate cells.

Animals↗
PubMed1994

Uptake and storage of retinol and retinyl esters in bone marrow of children with acute myeloid leukemia treated with high-dose retinyl palmitate.

Twenty-one children with AML, who achieved complete remission with cytostatics, were treated with high doses of retinoids after remission was achieved. They were given 52 mumol retinyl palmitate/m2 (50,000 International Units/m2) daily for at least 2 years thereafter. Thirteen of the children are still in complete remission with a mean observation time of 103 months. Due to the positive effects of retinoids seen in the treatment of AML, we have studied uptake of lipoprotein-associated retinyl esters in bone marrow cells and peripheral leukocytes in vivo. An oral load of 104 mumol/m2 (100,000 International Units/m2) retinyl palmitate resulted in a doubling of the concentration of total retinol in bone marrow cells and peripheral leukocytes after 5 hours. However, in the fasting state no significant difference was observed between the content of total retinol in bone marrow cells from controls and from patients receiving retinyl palmitate daily for years. Our study suggests that bone marrow cells take up retinyl esters postprandially, but do not store retinoids.

Anticarcinogenic Agents↗
PubMed1993

[Recommendations of the Council for Nutrition on infant nutrition. Norwegian National Nutrition Council].

Because of the high frequency of nursing mothers there is concensus that, in general, the diet of Norwegian newborns is good for the first six months of life. However, several publications have questioned the recommendations concerning the babies' diet from the age of six to 12 months with respect to time of weaning and introduction of cow's milk, and intake of energy, fat-soluble vitamins, iron and essential fatty acids. The Norwegian National Nutrition Council has therefore evaluated the available data, and has adjusted some of the recommendations, including the recommendations for energy intake, energy percent from fat and essential polyunsaturated fatty acids.

Breast Feeding↗
PubMed1993

[Vitamin A and toxicity. Should pregnant women and small children reduce their intake of liver products and vitamin A?].

It has been recognized for many years that high doses of vitamin A-metabolites may be teratogenic. Recently, it was found that the content of vitamin A in liver and liver products was much higher than the value prescribed in dietary tables. Thus, in many countries, health personnel advised pregnant women to reduce their intake of liver and liver products. Officials in Norway, however, have not given similar advice to Norwegian women. The background for this point of view is discussed in the present paper.

Female↗
PubMed1993

The effect of retinoids and clofibric acid on the peroxisomal oxidation of palmitic acid and of 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid in rat and rabbit hepatocytes.

The effects of retinoids and the peroxisome proliferator clofibric acid on peroxisomal enzyme pathways were studied in hepatocytes from both rat and rabbit. Retinoic acid and retinol increased the activity of acyl-CoA oxidase in rabbit hepatocytes around 60% and around 30% in rat hepatocytes. Exposure to clofibric acid caused an increase in acyl-CoA oxidase activity of 115% in rat hepatocytes and of 40% in rabbit hepatocytes, indicating that rabbit is less sensitive to peroxisome proliferator than rat. Simultaneous exposure to clofibric acid and retinoids did not act additatively or synergistically. Both rabbit and rat hepatocytes expressed mRNA for the peroxisome proliferator activated receptor, (PPAR), although the transcript in rabbit was slightly smaller compared to that expressed in rat hepatocytes. The effect of retinoic acid in 7800 C1 Morris rat hepatoma cells, a cell line known to have an inducible peroxisomal beta-oxidation of fatty acids, was only slight with an increase of the acyl-CoA oxidase activity of 25% compared with control cells. As for clofibric acid, which gave a 2-fold induction of the acyl-CoA oxidase activity, the effect of retinoic acid was potentiated by dexamethasone. These cells also expressed mRNA for PPAR, with the same size as that found in rat hepatocytes. The oxidation of 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid (THCA), an intermediate in bile acid formation, in rat hepatocytes increased 110% by clofibric acid and around 80% by retinoic acid. In rabbit hepatocytes, clofibric acid increased the oxidation rate 75% and retinoic acid 100%. The results presented here show similarities in the effects of retinoids and clofibric acid on the acyl-CoA oxidase activity and the oxidation rate of THCA, since they increase these two peroxisomal activities in hepatocytes in vitro. A decrease in both these enzyme activities occurs during cultivation time in untreated primary hepatocyte cultures. The present data may therefore either be explained by an increased expression or an induced stability of the enzymes involved.

Animals↗
PubMed1993

Transfer of retinol-binding protein from HepG2 human hepatoma cells to cocultured rat stellate cells.

Rat liver stellate cells were cocultured with HepG2 human hepatoma cells, which are known to synthesize and secrete retinol-binding protein (RBP). Transfer of human RBP from HepG2 cells to stellate cells was studied by cryoimmunoelectron microscopy. In stellate cells, human RBP was found on the cell surface and within endosomes. The transfer of human RBP from HepG2 cells to stellate cells was blocked by addition of RBP antibodies to the culture medium. Very little uptake of RBP was observed when fibroblasts were cocultured with HepG2 cells. In a series of experiments, RBP was bound to its putative cell surface receptor at 4 degrees C, and the stellate cells were washed and then incubated at 37 degrees C in order to allow them to internalize a pulse of RBP. About 50% of the RBP was internalized after 6 min of incubation. The RBP-positive vesicles were initially (after 1-2 min) located close to the cell surface and later were found deeper in the cytoplasm. During the first 10 min, RBP was mainly observed in close association with membranes. After 2 hr, however, most RBP was localized in intracellular vesicles at a distance from the vesicular membranes, suggesting that RBP had been released from its receptor. Saturable binding of RBP to liver cells was demonstrated when cells were incubated with 125I-RBP at 4 degrees C and cell-associated radioactivity was determined. The calculated dissociation constant for the specific binding was 12.7 +/- 3.2 nM. A binding assay was also developed for determination of solubilized RBP receptor. Solubilized proteins from the nonparenchymal liver cells bound about 30 times more 125I-labeled RBP than did parenchymal cells (based on mass of cell protein). These data suggest that RBP mediates the paracrine transfer of retinol from hepatocytes to perisinusoidal stellate cells in liver and that stellate cells bind and internalize RBP by receptor-mediated endocytosis.

Animals↗
PubMed1993

Receptor-mediated endocytosis of retinol-binding protein by liver parenchymal cells: interference by radioactive iodination.

Retinol-binding protein (RBP) was iodinated directly by radio-iodine substitution on the tyrosyl residues by the sodium hypochlorite (NaOCl) or the Enzymobead (EB) methods, or indirectly by linkage of 125I-tyramine-cellobiose (TC) or 125I-N-succinimidyl-3-(4- hydroxyphenyl)propionic acid ester (SHPP) adduct on to free amino residues of RBP. Binding, uptake and degradation of iodinated RBP were studied in isolated rat and rabbit liver parenchymal cells. The amount of ligand bound to cells at 4 degrees C was dependent on the type of labelling, in that the 125I-TC ligand was bound to a lesser extent than NaClO-labelled 125I-RBP, EB-labelled 125I-RBP and 125I-SHPP-RBP. At 37 degrees C, the 125I-SHPP-RBP and the EB-labelled 125I-RBP became cell-associated more rapidly than the other two ligands. The higher cell association at 37 degrees C than at 4 degrees C suggests that internalization of the ligand occurred at the higher temperature. The degradation of the ligands was also different. The EB-labelled 125I-RBP, the 125I-TC-RBP and the 125I-SHPP-RBP showed an apparent lag phase before a steady increase in acid-soluble radioactivity was observed. Much less of EB-labelled 125I-RBP and 125I-TC-RBP were degraded (about 6%) than of the other two ligands (about 16%) after 120 min. About 50% of the acid-soluble radioactivity in these experiments could be accounted for by degradation in the medium, suggesting that about half of the degradation observed was intracellular. The present study therefore shows that the different labelling techniques yield varying estimates of the cellular handling of RBP. In addition, a rapid release of RBP was observed in experiments where cells were pulsed with radioactive RBP at 4 degrees C, washed and incubated further at 37 degrees C. Between 50% and 70% was released after 5 min of incubation. By increasing the temperature during the pulse to 37 degrees C, or by lowering the temperature during the chase to 4 degrees C, much less RBP was released from the cells. These data suggest that the release process represents recycling of internalized ligand from an early endosome.

Animals↗
PubMed1993

Retinol and retinyl esters in rabbit bone marrow and blood leukocytes.

Due to the well documented effects of retinoids on growth and differentiation of some leukaemic cells in vivo and in vitro, we measured the amount of retinol and retinyl esters in bone marrow, blood leukocytes, and liver in rabbits fed large doses of retinyl palmitate. Both Chinchilla rabbits and Watanabe Heritable Hyperlipidaemic rabbits which lack functional low density lipoprotein receptors, were fed 26 mumoles (25.000 IU) of retinyl palmitate daily for 8 weeks. The animals stored retinoids in large amounts in the liver, whereas only minor amounts were stored in bone marrow. More than 97% of the retinoids in the liver was esterified, while most of the retinoids in bone marrow were unesterified. We also studied the post-prandial increase in chylomicron associated retinyl esters in rabbit leukocytes in vivo. After administering an oral load of 26 mumoles retinyl palmitate, retinoids increased four-fold in blood leukocytes after 5 h. There was almost no difference in retinoid uptake in leukocytes in Watanabe Heritable Hyperlipidaemic rabbits compared to normal rabbits, suggesting that chylomicron remnant retinyl esters are taken up in peripheral blood leukocytes independently of the low density lipoprotein receptor. In conclusion, bone marrow does not store high amounts of retinoids, and retinyl ester transport and storage appear normal in absence of functional low density lipoprotein receptors.

Animals↗
PubMed1993

Vitamin A metabolism in rat liver: a kinetic model.

Vitamin A metabolism in the liver involves both hepatocytes and the nonparenchymal perisinusoidal stellate cells. To describe and quantitate the dynamic relationships between retinol in these cells and in plasma, we administered either chylomicrons labeled with [3H]retinyl esters or plasma containing [3H]retinol-retinol-binding protein-transthyretin to rats. Radioactivity and retinol masses were measured in plasma, liver, and isolated hepatocytes for 15 days; data were analyzed by model-based compartmental analysis. The resulting model predicts that: 1) approximately 20% of the total plasma turnover of retinol goes to the liver (vs. nonhepatic tissues) and approximately 20% of plasma retinol input is from liver (vs. nonhepatic tissues), 2) about one-half of the retinol recycling from plasma to liver is taken up by hepatocytes and about one-half by nonparenchymal cells, 3) retinyl esters in both cell types are derived preferentially from newly taken up retinol rather than from the main intracellular retinol pools, and 4) at least one-half of the retinol secreted by hepatocytes of rats consuming low levels of vitamin A is directly transferred to nonparenchymal cells. In addition, the data are compatible with the hypothesis that retinol-binding protein is the vehicle for transfer of retinol from hepatocytes to nonparenchymal stellate cells and between plasma and liver cells.

Animals↗
PubMed1992

Vitamin A is a key regulator for cell growth, cytokine production, and differentiation in normal B cells.

In the present paper we demonstrate that retinol-retinol-binding protein and chylomicron remnant retinyl esters in concentrations normally found in human plasma inhibit growth of normal human B lymphocytes. Physiological concentrations of retinoic acid (about 30 nM) were less active than physiological concentrations of retinol (about 3 microM). Pharmacological concentrations of retinol and retinoic acid were more active than the concentrations normally found in plasma. Retinol (3 microM) inhibited anti-IgM-mediated DNA synthesis as measured by [3H]thymidine uptake at 72 h by 78%. Furthermore, we found that the cells were blocked in the mid-G1 phase of the cell cycle. Thus, neither MYC up-regulation measured at 3 h nor the expression of the early activation antigen 4F2 was reduced by retinol, whereas the late activation markers (transferrin receptor expression and actinomycin D staining at 48 h of stimulation) were markedly inhibited. Retinol reduced the interleukin 6 production induced by anti-IgM and interleukin 4 after 48 h, whereas the induction of interleukin 6 and tumor necrosis factor by O-tetradecanoylphorbol-13-acetate and ionomycin was less affected. We also noted that the retinoids reduced the formation of plaque-forming cells (i.e. Ig synthesis). These data imply that vitamin A present in human plasma is a normal modulator of B cell function.

B-Lymphocytes↗
PubMed1992

Direct mobilization of retinol from hepatic perisinusoidal stellate cells to plasma.

We have studied the mechanism for mobilization of retinol from stellate cells. Our data show that perisinusoidal stellate cells isolated from liver contained retinol-binding protein (RBP) mRNA. By Western blot analysis we found that cultivated liver stellate cells secreted RBP into the medium. Cultivated stellate cells loaded in vitro with [3H]retinyl ester mobilized radioactive retinol as a complex with RBP. Furthermore, exogenous RBP added to the medium of cultured stellate cells increased the secretion of retinol to the medium. These data suggest that liver stellate cells in vivo mobilize retinol directly to the blood and that a transfer to parenchymal cells for secretion as holo-RBP is not required. The direct mobilization of retinol from liver stellate cells as retinol-RBP to blood is indirectly supported by the demonstration of RBP mRNA production and RBP secretion by lung stellate cells. The data suggest that the same mechanism for retinol mobilization may exist in hepatic and extrahepatic stellate cells. This is, vitamin A-storing stellate cells in liver, lungs, and probably also in other organs may synthesize their own RBP (or alternatively use exogenous RBP) and mobilize holo-RBP directly to the blood.

Animals↗
PubMed1992

McCollum Award Lecture, 1992: vitamin A absorption, transport, cellular uptake, and storage.

We discuss vitamin A with emphasis on its absorption, transport, cellular uptake, storage, and intracellular metabolism. Dietary retinyl esters are hydrolyzed to retinol in the intestinal lumen before absorption by enterocytes. Carotenoids are absorbed and then partially converted to retinol in the enterocytes. In enterocytes retinol is esterified before incorporation into chylomicrons together with triacylglycerols. Chylomicrons reach the general circulation by way of the intestinal lymph, and chylomicron remnants are formed in the blood capillaries. The remnants, which contain almost all the absorbed retinol, are cleared by the liver parenchymal cells, and to some extent also by cells in blood, bone marrow, adipose tissue, and spleen. The uptake is most probably mediated via surface receptors for low-density lipoproteins or a low-density lipoprotein-receptor-related protein. In the liver parenchymal cells the retinyl esters are rapidly hydrolyzed to retinol, which binds to retinol-binding protein. Normally, most of the absorbed retinol coming into the liver parenchymal cell is transferred on retinol-binding protein to stellate cells, which store retinol as retinyl esters in lipid droplets.

Awards and Prizes↗