Why Patients with 5q- Syndrome Have Thrombocythemia?
Authors: Campr Vit , Stritesky Jan , Fuchs Ota , Jonasova Anna , Zemanova Zuzana , Neuwirtova Radana 
Published: October 2019

Abstract
Introduction. Fli1 (Friend leukemia virus integration 1) together with other transcription factors induces the megakaryocytic differentiation of MEP (megakarycytic and erythroid progenitor). Refractory anemia and thrombocythemia is typical for 5q- syndrome. We found increased mRNA level of Fli1 in mononuclear bone marrow cells of 5q- syndrome patients in comparison with healthy controls (Neuwirtova et al., Ann Hematol 2013). The reason of the elevated Fli1 in 5q- syndrome is haploinsufficiency of microRNA-145, which targets Fli1 mRNA (Kumar et al., Blood 2011). Due to haploinsufficiency of RPS14 in 5q- syndrome non-consumed ribosomal proteins cause ribosomal stress and inactivate HDM2 in erythroblasts. E3 ubiquitin ligase HDM2 regulates p53 level by p53 degradation in proteasome. Inactivated HDM2 in erythroid precursors of 5q- syndrome leads to apoptosis of erythroblasts and to anemia. Why ribosomal stress does not cause thrombocytopenia and ineffective megakaryopoiesis as well? Our previous results support significant role of Fli1 in this process. Fli1 binds to promoter of the HDM2 gene and increases its transcription (Truong et al., Oncogene 2005). The increased activity of HDM2 in megakaryocytes inspite of ribosomal stress maintains p53 regulation and its degradation in proteasome. Megakaryopoiesis remains effective. Why it is not the case in erythroid precursors? To answer this question it was necessary to detect Fli1 as the protein and to determine in which cells Fli1 is present.
Material and Methods. Twenty-three control representative bone marrow trephine biopsies of patients from controls (8 negative staging biopsies in lymphoma) and of patients with various hematological diagnoses (7 MDS with normal chromosome 5, 4 MPN, 3 AML and 1 RARS-T) and from 15 patients with 5q- syndrome were examined. In 13 patients with 5q- syndrome, samples taken before and 6 months after lenalidomide (Revlimid) therapy were available. The expression of Fli1 protein was investigated by immunohistochemistry (IHC). Expression of Fli1 on erythroid precursors was studied by double staining IHC procedure utilizing antibodies against Fli1 and either glycophorin A or E-cadherin known as reliable markers for erythroid precursors.
Results. Nuclear expression of Fli1 was demonstrated in normal as well as in dysplastic megakaryocytes, in most cells of granulocytic series and lymphocytes. No staining for Fli1 was seen in erythroblasts and proerythroblasts visualized by expression of either glycophorin A or E-cadherin both in 5q- syndrome and controls. There were no significant differences in Fli1 expression between samples taken before and after lenalidomide treatment.The used IHC technique does not permit quantitative analysis of Fli1 protein levels. This fact could explain why we did not find any difference in Fli1 protein labeling in megakaryocytes before and after lenalidomide treatment while Fli1 mRNA level was decreased in majority of 5q- syndrome patients after six months of this therapy.
Conclusion. Fli1 expression was found in normal as well as in dysplastic megakaryocytes. However, no Fli1 positivity was found in erythroid precursors in both 5q- syndrome and controls. Negativity of Fli1 expression in erythroid precursors in 5q- syndrome support our hypothesis of protective role of Fli1 against apoptosis under ribosomal stress in megakaryocytes in contrast to erythroblasts lacking Fli1. This protective role of Fli1 in megakaryocytes consists in Fli1 potentiation of expression of the E3 ubiquitine ligase HDM2 (Truong et al., Oncogene 2005). The presence of increased Fli1 in megakaryocytes helps to explain effective megakaryopoiesis in 5q- syndrome and is the answer to the question in the title of our abstract.
Supported by Ministry of Health, Czech Republic-conceptual development of research organization Institute of Hematology and Blood Transfusion 00023736, RVO-VFN64165 and PRVOUK P-27/LF1/2.

Disclosures
No relevant conflicts of interest to declare.