In-Vivo Study of Erythropoietic Pathway Genes Regulation and Differentiation by Prolonged Simultaneous Administration of Glucocorticoids with Epoietin in Animal Modules
Authors: Agheli Aref , Avezbakiyev Boris , Steier William , Kalavar Madhumati , Chen Chi , He Zili 
Published: October 2019

Abstract
Abstract 4798

Objectives:
The role of steroids in mammalian erythropoiesis has not well defined. We have previously reported our observation on three human cases in which there was a synergism and accelerated response to the Erythropoietic Stimulating Agents (ESA) with simultaneous low and physiologic dose administration of glucocorticoids. In the current study, we investigated the additive effects of different dose schedules of steroids on hematopoietic effects of ESA in animal modules.

Methods:
A total of 74, four-weeks old male Sprague-Dawley rats were randomized to 6 groups; (A) control, (B) therapeutic doses of either erythropoietin, [Procrit Epoetin Alfa, 100 UI/kg], or (C) dexamethasome (300 mcg/kg), as well as combination of erythropoietin (Epoetin Alfa, 100 UI/kg) with (D) low, [25 mcg/kg], (E) physiologic, [300 mcg/kg], and (F) high, [2.5 mg/kg] doses of dexamethasone through abdominal hypodermal injection three times a week for a total of four weeks. At the conclusion of the study, peripheral blood sample, and Bone marrow mononuclear cells were collected through femur flushing. The samples were lysed and stored in RNA denaturation buffer at –80°C until use. Expressions of multiple hematopoietic major genes were assessed by real-time RT-PCR. Amplification data were processed using ΔΔCt method. Hemoglobin concentration and other CBC parameters were measured at the reference lab.

Results:
Mean hemoglobin concentrations were significantly higher in groups D (20.76 g/dl, 95% CI 20.08–21.45), E (20.45 g/dl, 95% CI, 19.97–20.94), and F (20.99 g/dl, 95% CI 20.55–21.42), compared to the controlled groups A, B, C (14.57, 15.68, 19.23 g/dl respectively) with two-tailed p-value of <.0001. (Figure-1) Real time RT-PCR based gene expression profiling of major hematopoietic regulators revealed robot increases of JAK2 gene expression in groups of animals treated with EPO only, or even higher increase with EPO plus either low or physiologic doses of dexamethasome. Similarly, GATA-1 levels are increased in groups treated with EPO only, or EPO with low or physiologic doses of dexamethasome. c-kit and NFkB1 expression levels are markedly higher in EPO plus dexamethasome groups. In contrast, the levels of EPOR are generally reduced in all groups receiving ESA. (Figure -2)

Conclusion:
The findings in this study is suggestive that simultaneous administration of ESAs with glucocorticoids is associated with significant additive elevation of the hemoglobin concentration; however, higher dose of dexamethasone is associated with more frequent adverse side effects such as significant weight loss. It is also suggested that the erythropoietic effect of steroid is concerted by up-regulation of the multiple erythropoietic gene expressions, such as JAK2, GATA-1, c-Kit, and NFkB1, while down regulations of EPOR is uniformly seen in the Epo-treated groups. This novel finding could be clinically utilized to accelerate the erythropoietic response of the ESA in selected cases.

Disclosures:
No relevant conflicts of interest to declare.