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Using a suppression subtractive hybridization method, we have previously identified genes differentially expressed in erythroid cells heterozygous for large deletions in beta-like globin cluster. Herein, we investigated the expression of four newly detected red cell-related genes in erythroid differentiation. ARID1B and TSPYL1, genes with chromatin remodeling properties, presented similar patterns of expression with an upregulation after erythropoietin (EPO) addition, similar to previous data found in reticulocytes. ZHX2, a transcriptional repressor, was downregulated, and a redoxin-related gene, SH3BGRL2, had higher levels of expression on differentiation. These are the first investigations of these newly described genes in erythroid differentiation and demonstrate that the expression of these genes is affected by EPO stimulation. These genes may participate in globin regulation and may be important in the normal physiology of erythrocytes.

1. The major effect associated with hydroxyurea (HU) treatment of sickle cell anaemia (SCA) patients is an increase in fetal haemoglobin (HbF) synthesis, which inhibits the polymerization of haemoglobin S. 2. Hydroxyurea improves clinical symptoms by reducing the frequency of pain and vaso-occlusive crises, acute chest syndrome, transfusion requirements and hospitalization. 3. The molecular mechanisms responsible for HU-mediated induction of fetal globin transcription are not completely understood. Therefore, the aim of the present study was to identify differentially expressed genes participating in these mechanisms. 4. We established two suppression subtractive hybridization (SSH) libraries from reticulocytes obtained from SCA patients either not on or on HU treatment. The gene expression of some of the genes identified was subsequently evaluated by real-time polymerase chain reaction (PCR). 5. Genes identified with altered expression included SUDS3, FZD5 and PHC3, which may be associated with the regulation of globin expression. 6. This is the first demonstration of an association between HU treatment and the expression of genes identified in erythroid cells.

Lipid Bodies are cytoplasmic inclusions mainly formed by triglycerides and cholesterol esters, being also intracellular deposits of arachidonic acid (AA), which can be metabolized for eicosanoids generation. PGE2 is a major AA metabolite produced by epithelial cells and can modulate restoration of epithelium homeostasis after injury. We studied lipid body biogenesis and their role in AA metabolic pathway in epithelial cells. A rat cell line derived from normal intestinal epithelium, IEC‐6, was used. IEC‐6 in sub‐confluent cultures presented 15–20 lipid bodies/cell after staining with by OsO4. Lipid bodies were virtually absent in IEC‐6 upon reaching confluency or after FBS withdraw. Stimulation of confluent IEC‐6 with IL‐1β, PAF, or PMA was unable to induce lipid bodies. In contrast, AA dose‐dependently induced lipid body formation and proliferation foci in confluent cultures independently of its metabolism to PGE2. AA induction of lipid bodies was dependent on signaling through p38, PKC, and PI3K, but not ERK 1/2 or JNK. Lipid body biogenesis facilitates AA release after ionophore activation without modifying cPLA2α expression. Our results suggest a strong relationship between lipid body biogenesis and epithelial cell proliferation, and indicate that these organelles are induced in a stimulus‐specific manner and facilitate AA mobilization in epithelial cells.Financial support: CNPq, FAPERJ, CAPES

Título em inglês : "Differentially expressed genes in hydroxyurea treated erythroid cells and potentially involved in the reactivation of fetal hemoglobin"

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