Post-transcriptional effects of interleukin-3, interferon-gamma, erythropoietin and butyrate on in vitro hemoglobin chain synthesis in congenital hemolytic anemia

Background and Objectives. Various agents modulate hemoglobin synthesis. In vitro modulation of translation in hemoglobin chain synthesis was analyzed in patients with congenital hemolytic anemia (n = 32) and healthy controls (n = 17). Design and Methods. Enriched reticulocytes were co-incubated with H-3-leuci ne and cytokines or butyrate. Reversed-phase chromatography enabled separat ion of alpha-, beta- and gamma -globin chains. Globin chain synthesis was c alculated from measured 3H-leucine incorporation. Transferrin, erythropoiet in, interleukin-3 and interferon-gamma receptors were detected by flow cyto metry. Reverse-transcription polymerase chain reaction (RT PCR) was used to demonstrate changes of RNA stability. Results and Discussion. Interleukin-3, interferon-gamma and butyrate caused a significant 2-fold increase (range 1.8-2.4; p < 0.01) of alpha- and beta -chain synthesis in congenital hemolytic anemias. Analysis of gamma -globi n chain synthesis revealed a lower, i.e. 1.4 fold increase (range 1.32 to 1 .41; p < 0.03). The absolute amount of globin synthesis was calculated to b e 2.9 X 10(-12) g/reticulocyte/24h. After incubation with interleukin-3 the absolute additional synthesis of the oc-globin chain reached 1.31 X 10(-12 ) g/reticulocyte/24h, of the beta -globin chain, 1.31 10(-12) g/reticulocyt e/24h and of the gamma -globin chain, 0.26 X 10(-12) g/reticulocyte/24h. Bu tyrate and interferon-gamma had no or even an inhibiting effect on reticulo cytes from normal controls, while interleukin-3 stimulated alpha- and gamma -chain synthesis (1.4 and 2.4 fold, respectively; p < 0.03) suggesting an increase of fetal hemoglobin (HbF). Erythropoietin showed no stimulating in fluence. Membrane-associated interleukin-3 receptors were detected in 0.78 +/- 0.14%, and interferon-gamma receptors in 0.1 +/- 0.015% of the red cell s. Erythropoietin receptors were extremely rare (0.05 +/- 0.015%). The expr ession of transferrin receptors (CD71) correlated with the extent of globin chain stimulation. The alpha-, and beta -globin mRNA content of the reticu locytes after interleukin-3 incubation, as measured by RT-PCR, increased. Interpretation and Conclusions. Hemoglobin chain synthesis could be modulat ed post-transcriptionally by interleukin-3, interferon-gamma and butyrate. Transferrin receptor and globin RNA stability might be involved in this phe nomenon. (C) 2001, Ferrata Storti Foundation.