[Ulcerative colitis. Complications and sequelae in relation to the extent of the disease. A 10-year material]. / Ulcerøs kolitt. Komplikasjoner og følgetilstander i relasjon til utbredelse. Et tiårsmateriale.

Patients with ulcerative colitis in our catchment area were followed over a ten-years period with the focus on the relation between the extent of inflammation and complications. Total colitis was found in 180 out of 334 patients with known extension, while in 154 of the patients the inflammation was classified as left-sided colitis. The frequency of severe colitis, colectomy and cancer was more or less the same as stated in earlier reports. We observed a difference, however, between the patients with extensive colitis and those with left-sided colitis as regards the course of the disease and the complications related to ulcerative colitis. The risk of liver disease, severe colitis and colectomy were significantly higher among the patients suffering from extensive colitis. Extraintestinal manifestations, on the other hand, were equally distributed among both groups of patients. The reasons for the observed differences are unknown.

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.

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.

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.

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.

Uptake of chylomicron remnant retinyl esters in human leukocytes in vivo.

Retinoids have been successfully used in the treatment of some forms of leukaemia, suggesting that such cells have an efficient uptake mechanism for circulating retinoids. Therefore, we have studied the uptake of lipoprotein-associated retinyl esters in human leukocytes in vivo. After an oral load of 100 mumol retinyl palmitate (30,000 retinol equivalents) per square meter given to healthy adults, the concentration of retinoids in circulating leukocytes was determined. A peak was measured after 5 h, which coincided with a peak of retinyl esters in plasma. To test whether low-density lipoprotein receptors are necessary for the postprandial uptake of retinoids, we studied retinoid uptake in leukocytes from two patients homozygous for familial hypercholesterolaemia. After an oral load of retinoids we found that leukocytes from these patients took up at least as much retinoid as leukocytes in normal individuals, suggesting that uptake of chylomicron remnant retinyl esters may proceed independent of the low-density lipoprotein receptor. The expression of mRNA for the low density lipoprotein receptor-related protein, which is a putative chylomicron remnant receptor, was similar in leukocytes from a patient homozygous for familial hypercholesterolaemia and normal individuals. Six hours after vitamin A administration, recovery of unesterified retinol was 71% in normal leukocytes, however, only 9% unesterified retinol was recovered in leukocytes from the two patients with familial hypercholesterolaemia. Thus, the apparent rate of retinyl ester hydrolysis was markedly reduced in leukocytes from these patients, indicating different intracellular traffic of chylomicron remnants in normal individuals and patients homozygous for familial hypercholesterolaemia.

Retinyl esters in chylomicron remnants inhibit growth of myeloid and lymphoid leukaemic cells.

We have studied the effects of retinyl esters in chylomicron remnants on cell growth and differentiation of myeloid and lymphoid leukaemic cells. Ten mumol l-1 retinyl ester in chylomicron remnants effectively reduced proliferation of the myeloid leukaemic cell lines HL60, U937 and KG-1, and induced differentiation of 68% and 53% of the HL60 and U937 cells, respectively, in 5 days. While no effect on cell growth of the lymphoid cell lines Daudi, Raji and SOS was observed, 10 mumol 1-1 retinyl esters in chylomicron remnants reduced the growth of the B lymphoid cell line Reh by more than 50%. Primary cell cultures from six patients with acute leukaemia (four non-lymphocytic and two lymphocytic) were incubated with chylomicron remnant retinyl esters and proliferation was measured by means of thymidine incorporation. Among the myeloid leukaemic cells, the monomyelocytic, the two promyelocytic and the monoblastic leukaemic cells were growth inhibited. Chylomicron remnants had no effect on the growth of the c-ALL primary culture, but reduced proliferation of the T-ALL primary culture by approximately 20% after 48 h. These data suggest that high doses of retinol may be used in the treatment of some forms of acute leukaemia.

Cultivation of HL-60 cells in a serum-free medium containing granulocyte/macrophage colony-stimulating factor.

In order to develop a defined cultivation medium for HL-60 cells, we cultivated these cells in a serum-free suspension medium and tested the effect of various growth factors. Of the factors tested, granulocyte/macrophage colony-stimulating factor was most active in growth stimulation. A much lower effect was obtained with granulocyte colony-stimulating factor and transferrin. No effect was found with interleukin-3 and insulin. Granulocyte colony-stimulating factor was the only growth factor tested that also induced differentiation as judged by the nitroblue tetrazolium test. Growth of HL-60 cells in medium containing granulocyte/macrophage colony-stimulating factor (125 U/ml) and transferrin (5 micrograms/ml) as the only protein factors was similar to growth in medium containing 10% serum. No increase in spontaneous differentiation of HL-60 cells in this defined medium was observed. Physiological concentrations of retinol bound to retinol-binding protein and retinyl ester in chylomicron remnants reduced proliferation as well as the level of c-myc oncoprotein and induced differentiation of HL-60 cells cultivated in defined medium. Hence, this defined medium may be useful when studying the function of retinoids in HL-60 cells.

Metabolism of 24-ethyl-4-cholesten-3-one and 24-ethyl-5-cholesten-3 beta-ol (sitosterol) after intraperitoneal injection in the rat.

14C-labeled C29- and C27-steroids were injected in rats, which were killed after 14 days. Phytosterols (C29) were excreted mainly as such, whereas C27-steroids were recovered essentially as water soluble metabolites. The total 14C-excretion was lower from 3-oxo, delta 4-steroids (5 alpha-stanol precursors) than from 3 beta-hydroxy,delta 5-steroids. 14C-Phytosterols were accumulated more than C27-steroids in liver, serum (mainly in HDL) and especially in adrenal glands and ovaries. In relation to serum, particularly the 5 alpha-stanols were enriched in the adrenal glands and ovaries. No striking lysosomal accumulation of any of the steroids was found.

The presence of 5 alpha-sitostanol in the serum of a patient with phytosterolemia, and its biosynthesis from plant steroids in rats with bile fistula.

The presence of 5 alpha-sitostanol (24-ethyl-5 alpha-cholestan-3 beta-ol) in serum of a patient with the rare genetic disease phytosterolemia was confirmed. This study aimed at clarifying the pathway(s) for the formation of 5 alpha-sitostanol, by use of rats with bile fistula. 5 alpha-Sitostanol was formed only slowly from sitosterol, but readily from 24-ethyl-4-cholesten-3-one. Some conversion was also obtained with 7 alpha-hydroxysitosterol as precursor. In view of the low rate of 7 alpha-hydroxylation of sitosterol, however, a pathway from sitosterol to 5 alpha-sitostanol involving 7 alpha-hydroxysitosterol as intermediate is probably of small physiological importance. Intestinal microorganisms are not essential for the above conversions, since the 5 alpha-sitostanol was found in bile from bile fistula rats. 5 alpha-Sitostanol was converted to water soluble metabolites (bile acids) much more slowly than was cholestanol (5 alpha-cholestan-3 beta-ol), and was accumulated serum to a much larger extent.