Upregulation of the Anti-Apoptotic Protein, Survivin, in Hematopoietic Cells in Sickle Cell Anemia and Effects of Hydroxyurea Therapy.
Authors: Lanaro Carolina , Franco-Penteado Carla Fernanda , Mello Mariana R. B. , Fertrin Kleber Yotsumoto , Bezerra Marcos André C , Albuquerque Dulcinéia M. , Conran Nicola , Saad Sara T.O. , Araujo Aderson S. , Costa Fernando F. 
Published: October 2019

Abstract
Abstract 1532
Poster Board I-555
Survivin (BIRC5) is a member of the inhibitors of apoptosis family implicated in both prevention of cell death and control of mitosis. Although the actions of survivin in control of cancer cell division and apoptosis have been studied, its role in nonneoplastic diseases is not elucidated. Chronic inflammation is associated with STAT-3 upregulation, which can induce survivin production. Sickle cell anemia (SCA) has been characterized as a chronic inflammatory state and growing evidence indicates that inflammatory stress within the microvasculature may play a significant role in the vasoocclusion that is characteristic of SCA. Long-term treatment with hydroxyurea (HU) has been shown to reduce the production of inflammatory cytokines in SCA patients and leukocyte number. Since enhanced survivin expression has been reported in leukocytes under inflammatory conditions, and during hematopoietic cell survival and proliferation, the aim of this study was to investigate changes in survivin levels during erythroid differentiation, and determine expression in neutrophils (NS), mononuclear cells (MC) and red blood cell (RBC) in steady-state SCA patients (n≥10), SCA patients on HU therapy (n≥16), and healthy controls (HC, n≥5). Survivin and STAT-3 gene expression were determined by qRT-PCR analysis in primary human erythroblasts cultures for 7, 10 and 13 days and leukocytes separated from peripheral blood samples. Survivin protein expression was determined by flow cytometry with survivin-specific antibodies. Survivin gene expression was significantly increased during erythroid differentiation, but survivin mRNA levels showed similar patterns between SCA and HC (7d: 0.8±0.1 × 0.7±0.08; 10d: 1.7±0.3 × 1.6±0.2; 13d: 2.2± 0.27 × 1.8±0.19,U.A.,P>0.05,respectively). However, protein levels of survivin in mature RBC (glicophorin A +) was significantly higher in SCA patients compared to HC (41.90± 2.9 × 25.76±1.9, P=0.0006, respectively). BIRC-5 gene expression in MC was significantly higher in SCA patients compared to HC (0.9±0.1 × 0.5±0.2, P=0.04, respectively). Survivin protein levels in MC from SCA was significantly increased to compared to HC (51.7±3.2 × 39.7±1.7, MFI, P=0.01,respectively). Survivin protein levels are elevated in NS of SCA patients compared to HC (28.4±1.6 × 21.9±1.5, MFI, P=0.02,respectively). No significant alterations in the mRNA levels of the gene encoding STAT-3 were found during erythroid differentiation (7d: 1.1±0.04 × 1.1±0.08; 10d: 0.6±0.07 × 0.8±0.08; 13d: 0.6±0.07 × 0.9±0.1, P>0.05,respectively) or MC cells (1.2±0.1 × 1.1± 0.1, P>0.05,respectively) in SCA patients compared to HC. Patients on HU therapy demonstrated lower survivin MC gene expressions and protein levels compared to non-treated patients (0.6±0.3 × 0.9±0.1; 37.9±1.5 × 51.7±3.3, P=0.02; P<0.0001,respectively), but no difference was shown in STAT-3 gene expressions (1.1±0.04 × 1.2 ±0.1, respectively). Survivin protein levels were not significantly different in NS and RBC in patients on HU therapy compared to SCA (27.1±1.8 × 28.4± 1.6; 45.9± 3.2× 41.9± 2.9, MFI, P>0.05, respectively). Our data showed that survivin gene and protein expression are upregulated in MC in SCA patients, independently of STAT-3 expression. In addition, a high protein expression was observed in NS and RBC in these patients. HU therapy was associated with lower survivin expression in MC, but not NS and RBC, indicating that the beneficial effect that HU has on the inflammatory state, may participate in the reduced levels of survivin. In conclusion, the exact importance of survivin in SCA vasooclusion is not clear, but data indicates a high expression of this protein in leukocytes and RBC of SCA patients and may imply a role for this protein in leukocytosis and RBC proliferation in SCA.

Disclosures
No relevant conflicts of interest to declare.