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.

Effect of retinoids and 1,25(OH)2 vitamin D3 bound to their plasma transport proteins on growth and differentiation of HL-60 cells.

We have compared the effect of physiological and pharmacological concentrations of retinoids and 1,25(OH)2 vitamin D3 bound to their plasma transport proteins upon the proliferation and differentiation of HL-60 cells. Concentrations of chylomicron remnant retinyl ester similar to that obtained in plasma after a vitamin A-rich meal reduced the proliferation in more than 50% of HL-60 cells. Pharmacological concentrations of chylomicron remnant retinyl ester completely blocked the proliferation of the cells, and induced differentiation in 60% of the cells after 5 days. Physiological and pharmacological concentrations of retinoic acid bound to albumin had comparable effects. In contrast to earlier published data, which have been obtained with retinoids dissolved in ethanol, our results suggest that physiological and pharmacological concentrations of retinol (i.e. retinyl esters in chylomicron remnants) are as active as retinoic acid in reduction of proliferation and induction of differentiation of HL-60 cells. Physiological concentrations of 1,25(OH)2 vitamin D3 bound to vitamin D-binding protein (DBP) and retinol bound to retinol-binding protein had only a small effect on differentiation and proliferation of HL-60 cells.

Cytotoxic effect of TNF-alpha and abnormal regulation of cAMP in the retinoid resistant cell line U937/GTB.

We have studied the effect of retinoids, forskolin and TNF-alpha on two sublines of U937 cells, U937/clone 4 and U937/GTB. Retinoids induced differentiation of U937/clone 4, while the U937/GTB cells were resistant to induction of differentiation by retinoids. Retinoids effectively reduced the proliferation of both U937/clone 4 and U937/GTB, and decreased the steady state levels of c-myc mRNA in both sublines. Furthermore, the retinoid resistant U937/GTB cells were hypersensitive to the cytotoxic effect of TNF-alpha. Forskolin increased the cAMP level in the U937/clone 4 cells 14 times above basal level, but did not change the cAMP concentration in the U937/GTB cells.

Effects of antiepileptics on the hepatic storage of retinol.

The effect of various antiepileptics on the retinol storage in rat liver was tested. We observed a dose- and time-dependent reduction in the hepatic storage of retinol after the administration of several of these drugs. Administration for 11 weeks of phenobarbital, primidone and carbamazepine in doses comparable to those used in humans reduced the retinol concentration in the Liver by 17-33% compared to control rats. The plasma retinol levels remained unaffected in all the rats. Plasma retinol from 31 epileptic children had plasma levels between 0.7 and 2.4 nmol/ml, which is regarded as normal.