Study of Expression of FLVCR in Childhood Thalassemic Patients Demonstrate the Importance of FLVCR in Effective Erythropoiesis.
Authors: Zhu Yiping , Zhang Yiqiong , Abkowitz Janis L. , Yang Zhantao , Yuan Lixing 
Published: April 2020

Abstract
Feline Leukemia Virus, Subgroup C (FLVCR) is a newly found cytoplasmic heme exporter. Retroviral interference with FLVCR display results in a loss of erythroid cells after the CFU-E/proerythroblast stage of differentiation and severe anemia in cats. We studied expression of FLVCR in thalassemia with relatively redundant free cytoplasmic heme, evaluated the correlation of FLVCR expression with clinical parameters, and explore the importance of FLVCR for erythropoiesis in human cells. Samples of bone marrow from 50 children with thalassemia (n=21), other hemolytic anemia(AIHA, G6PD Deficiency, n=17) and non-anemia controls (n=12) were collected. Expression of FLVCR protein and mRNA in mononuclear red cells was quantitated in CFU-E/proerythroblasts (CD71 bright, CD117+) and later erythroid precursors (CD71 bright, CD117 negative cells) by 3 color flow cytometry and fluorescence quantitative Real-time PCR. Expression of FLVCR is higher in early precursors than later erythroid cells: Specifically, the percentage of specimens with high levels of FLVCR protein expression in proerythroblasts and the mean fluorescence intensities(MFI) of thalassemia, other hemolytic anemia and non-anemias were 78.9%, 84.2%,74.85% and 306,347, 291.5 respectively; while these values in later erythroid precursors were 8.8%,10.6%,6.9% and 150,122,119 respectively, P&lt;0.01. Expression of FLVCR tended to be higher in proerythroblasts of thalassemia and other hemolytic anemia:Positive rate and MFI of FLVCR protein expression in thess two groups were 78.9%,306 and 84.2%,347 respectively, as compared with cells of non-anemias (74.85%,291.5), with no statistical differences, P&gt;0.05. Expression of FLVCR is significantly higher in later erythroid precursors of thalassemia:Positive rate and MFI of FLVCR protein expression in later erythroid precursors of thalassemia were 8.8%, and 150 respectively, with significantly higher levels of expression compared with non-anemias (6.9%,119) P&lt;0.05. Expression of FLVCR mRNA in proerythroblasts of thalassemia, other hemolytic anemia and non-anemias were 3.6,4.85,3.7 respectively; while in later erythroblast were 4.13,6.06,3.92 respectively, P&lt;0.01. Hemolytic anemia patients with higher expression of FLVCR had higher reticulocyte count, higher percentage of later erythroblast and lower LDH, P&lt;0.05. Thalassemic patients with higher expression of FLVCR had higher percentage of sideroblasts and serum ferritin, P&lt;0.05. Expression of FLVCR protein was remarkably upregulated in proerythroblasts, combined with previous studies in vitro and identification of FLVCR as a human heme exporter, supporting FLVCR plays an important role in developing erythroid cells to protect them from heme toxicity. There was no significantly up-regulation of FLVCR expression in proerythroblasts of thalassemia, indicating probably other pathways exist in regulating heme metabolic balance combined with FLVCR; Expression of FLVCR in later erythroid precursors of thalassemia was higher than that of non-anemias, which implies FLVCR up-regulates with compensation, this may help counteract the ineffectiveness of late erythropoiesis that results from the poor coordination of heme and globin synthesis. The disparity between FLVCR protein and mRNA may reflect posttranscriptional regulation of protein level. Clinical results also support effect of FLVCR on erythropoiesis. FLVCR expression has close relation with iron metabolic parameters.