Korean J Physiol Pharmacol.  2012 Aug;16(4):281-285. 10.4196/kjpp.2012.16.4.281.

Comparison of Neurite Outgrowth Induced by Erythropoietin (EPO) and Carbamylated Erythropoietin (CEPO) in Hippocampal Neural Progenitor Cells

• Oh DH
• Lee IY
• Choi M
• Kim SH
• Son H

• KMID: 2011163
• DOI: http://doi.org/10.4196/kjpp.2012.16.4.281

Abstract

A previous animal study has shown the effects of erythropoietin (EPO) and its non-erythropoietic carbamylated derivative (CEPO) on neurogenesis in the dentate gyrus. In the present study, we sought to investigate the effect of EPO on adult hippocampal neurogenesis, and to compare the ability of EPO and CEPO promoting dendrite elongation in cultured hippocampal neural progenitor cells. Two-month-old male BALB/c mice were given daily injections of EPO (5 U/g) for seven days and were sacrificed 12 hours after the final injection. Proliferation assays demonstrated that EPO treatment increased the density of bromodeoxyuridine (BrdU)-labeled cells in the subgranular zone (SGZ) compared to that in vehicle-treated controls. Functional differentiation studies using dissociated hippocampal cultures revealed that EPO treatment also increased the number of double-labeled BrdU/microtubule-associated protein 2 (MAP2) neurons compared to those in vehicle-treated controls. Both EPO and CEPO treatment significantly increased the length of neurites and spine density in MAP2(+) cells. In summary, these results provide evidences that EPO and CEPO promote adult hippocampal neurogenesis and neuronal differentiation. These suggest that EPO and CEPO could be a good candidate for treating neuropsychiatric disorders such as depression and anxiety associated with neuronal atrophy and reduced hippocampal neurogenesis.

Keyword

Carbamylated erythropoietin; Erythropoietin; Hippocampus; Neurogenesis

MeSH Terms

Adult
Animals
Anxiety
Atrophy
Bromodeoxyuridine
Dendrites
Dentate Gyrus
Depression
Erythropoietin
Hippocampus
Humans
Male
Mice
Neurites
Neurogenesis
Neurons
Spine
Stem Cells
Bromodeoxyuridine
Erythropoietin

Figure

• 

  Fig. 1 Effect of erythropoietin (EPO) administration on cell proliferation in the dentate gyrus of adult mice. (A) BrdU labeling was used to assess cell proliferation. BrdU(+) cells were observed in the SGZ of the dentate gyrus (DG). (B) Stereological three-dimensional counts revealed that EPO treatment produced a significant increase (saline, 3,440±175 cells/mm3 vs. EPO, 4,228±286 cells/mm3, n=4 animals in each group) in BrdU(+) cells compared with those in saline treatment (control group, CTL) when analyzed 12 hours after treatment completion. *p<0.05. Scale bar: 200 µm.

• 

  Fig. 2 Effects of EPO treatment on differentiation of cultured adult hippocampal progenitor cells. (A) Cells were prepared as described in Methods. (B) Representative microscopic images showing that BrdU(+) cells differentiated into neuronal cells (MAP2(+) in green) (arrows) in the presence of EPO (10 U). (C) Quantitative analysis of BrdU/MAP2 doubly-labeled cells among total BrdU(+) cells. The results indicate that the percentage of BrdU(+) cells labeled for MAP2 was significantly increased by EPO treatment compared to that in the control group (CTL). *p<0.05, **p<0.01. Scale bar: 200 µm.

• 

  Fig. 3 EPO and CEPO increase the lengths of neurites. The lengths of neurites extending from the cell soma (arrows) were counted in neurons immunostained for MAP2 (n=40 cells per treatment). (A) Cells were prepared as described in Methods. (B, C) Treatment with EPO and EPO significantly (p<0.05) increased neurite outgrowth compared with that in the control group (CTL). *p<0.05, **p<0.01. Scale bar: 50 µm.

• 

  Fig. 4 EPO and CEPO increase the spine density. (A) Representative images of high-magnification Z-stack projections of dendrites of MAP2(+) hippocampal cells. (B) The density of dendritic spines (arrows) was significantly increased by EPO and CEPO (***p<0.001). n=9-15 MAP2(+) neurons per each group. The data were expressed as the number of spines per 10 µm. Scale bar: 10 µm.

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