Contributions
Abstract: S277
Type: Oral Presentation
Session title: Focus on iron metabolism
Background
Chronic Kidney Disease (CKD) is characterized by progressive kidney failure, low erythropoietin (EPO) levels, inflammation and iron deficiency, factors that contribute to anemia development.
Current EPO-based therapies, often with iron supplementation, are effective but frequently lead to side effects, mainly cardio-vascular. Thus, more specific approaches are needed.
Transferrin Receptor 2 (TFR2) in hepatocytes regulates hepcidin and iron homeostasis, and in erythroid cells interacts with EPO receptor, adjusting EPO sensitivity. In mice, total and hepatic Tfr2 inactivation causes iron overload, while its ablation in the erythroid compartment boosts erythropoiesis.
Aims
We investigated whether Tfr2 may be a therapeutic target for renal anemia. Our approach is expected to simultaneously increase iron availability and erythroid EPO responsiveness. Moreover, given the restricted expression of TFR2, off-target effects should be limited.
Methods
Feeding mice an adenine-rich diet for 8 weeks we induced CKD in:
(1) mice with bone marrow (BM)-specific deletion of Tfr2 (Tfr2BMKO) and relative controls, generated through BM transplantation and fed the adenine diet after complete recovery;
(2) mice with reduced Tfr2 expression in the liver, obtained treating wild type mice twice a week for 6 weeks with antisense oligonucleotides anti-Tfr2 (Tfr2-ASO) after renal damage development;
(3) mice with germline Tfr2 inactivation in the whole organism (Tfr2KO) and relative controls (Tfr2WT).
Results
Renal damage and iron restriction were comparable between Tfr2BMKO and control mice, but Tfr2BMKOshowed enhanced erythropoiesis, due to the increased EPO responsiveness of erythroid cells lacking Tfr2, as suggested by the over-activation of the EPO-EPOR signaling pathway (despite comparable EPO levels). However, anemia was only delayed, since at the end of the 8-week-long protocol iron availability was not adequate to sustain the enhanced erythropoiesis.
Having the proof-of-principle that targeting erythroid Tfr2 stimulates erythropoiesis in CKD, we moved to Tfr2-ASO treatment, which downregulates Tfr2 in the liver (95-97%), but not in the erythroid compartment. Circulating iron levels were increased in Tfr2-ASO mice, that showed also higher red blood cells (RBC) count and hemoglobin (Hb) levels during the first 2 weeks of treatment. However, Hb reverted to control levels before the end of the protocol. Thus, we confirmed that increased iron availability alone, due to hepatic Tfr2 downregulation, is not sufficient to boost erythropoiesis for a long period.
Finally, despite comparable renal damage, Tfr2KO mice showed higher RBC count and Hb levels compared to Tfr2WT littermates, until the end of the 8-week-long protocol.
Conclusion
The concomitant targeting of hepatic and erythroid Tfr2, here obtained through a total genetic inactivation, is necessary to ameliorate anemia of CKD. Indeed, on one side, selective Tfr2 inactivation in the BM enhances EPO responsiveness of erythroid cells and stimulates erythropoiesis, without increasing EPO levels per se. On the other side, hepatic Tfr2 targeting increases iron availability. Therefore, a specific approach able to inhibit both hepatic and erythroid Tfr2 could adjust iron availability according to the enhanced erythropoiesis, correcting both the drivers of anemia development in CKD. In the future, we will compare this approach to the available therapies and possibly move to a pharmacologic targeting.
Keyword(s): Anemia, Chronic renal failure, Erythropoieisis, Transferrin receptor
Abstract: S277
Type: Oral Presentation
Session title: Focus on iron metabolism
Background
Chronic Kidney Disease (CKD) is characterized by progressive kidney failure, low erythropoietin (EPO) levels, inflammation and iron deficiency, factors that contribute to anemia development.
Current EPO-based therapies, often with iron supplementation, are effective but frequently lead to side effects, mainly cardio-vascular. Thus, more specific approaches are needed.
Transferrin Receptor 2 (TFR2) in hepatocytes regulates hepcidin and iron homeostasis, and in erythroid cells interacts with EPO receptor, adjusting EPO sensitivity. In mice, total and hepatic Tfr2 inactivation causes iron overload, while its ablation in the erythroid compartment boosts erythropoiesis.
Aims
We investigated whether Tfr2 may be a therapeutic target for renal anemia. Our approach is expected to simultaneously increase iron availability and erythroid EPO responsiveness. Moreover, given the restricted expression of TFR2, off-target effects should be limited.
Methods
Feeding mice an adenine-rich diet for 8 weeks we induced CKD in:
(1) mice with bone marrow (BM)-specific deletion of Tfr2 (Tfr2BMKO) and relative controls, generated through BM transplantation and fed the adenine diet after complete recovery;
(2) mice with reduced Tfr2 expression in the liver, obtained treating wild type mice twice a week for 6 weeks with antisense oligonucleotides anti-Tfr2 (Tfr2-ASO) after renal damage development;
(3) mice with germline Tfr2 inactivation in the whole organism (Tfr2KO) and relative controls (Tfr2WT).
Results
Renal damage and iron restriction were comparable between Tfr2BMKO and control mice, but Tfr2BMKOshowed enhanced erythropoiesis, due to the increased EPO responsiveness of erythroid cells lacking Tfr2, as suggested by the over-activation of the EPO-EPOR signaling pathway (despite comparable EPO levels). However, anemia was only delayed, since at the end of the 8-week-long protocol iron availability was not adequate to sustain the enhanced erythropoiesis.
Having the proof-of-principle that targeting erythroid Tfr2 stimulates erythropoiesis in CKD, we moved to Tfr2-ASO treatment, which downregulates Tfr2 in the liver (95-97%), but not in the erythroid compartment. Circulating iron levels were increased in Tfr2-ASO mice, that showed also higher red blood cells (RBC) count and hemoglobin (Hb) levels during the first 2 weeks of treatment. However, Hb reverted to control levels before the end of the protocol. Thus, we confirmed that increased iron availability alone, due to hepatic Tfr2 downregulation, is not sufficient to boost erythropoiesis for a long period.
Finally, despite comparable renal damage, Tfr2KO mice showed higher RBC count and Hb levels compared to Tfr2WT littermates, until the end of the 8-week-long protocol.
Conclusion
The concomitant targeting of hepatic and erythroid Tfr2, here obtained through a total genetic inactivation, is necessary to ameliorate anemia of CKD. Indeed, on one side, selective Tfr2 inactivation in the BM enhances EPO responsiveness of erythroid cells and stimulates erythropoiesis, without increasing EPO levels per se. On the other side, hepatic Tfr2 targeting increases iron availability. Therefore, a specific approach able to inhibit both hepatic and erythroid Tfr2 could adjust iron availability according to the enhanced erythropoiesis, correcting both the drivers of anemia development in CKD. In the future, we will compare this approach to the available therapies and possibly move to a pharmacologic targeting.
Keyword(s): Anemia, Chronic renal failure, Erythropoieisis, Transferrin receptor