The authors declare that there is no conflict of interest.
Accumulation of a repulsive axonal guidance molecule RGMa in amyloid plaques: a possible hallmark of regenerative failure in Alzheimer's disease brains
Version of Record online: 25 JAN 2013
© 2012 The Authors. Neuropathology and Applied Neurobiology © 2012 British Neuropathological Society
Neuropathology and Applied Neurobiology
Volume 39, Issue 2, pages 109–120, February 2013
How to Cite
Satoh, J., Tabunoki, H., Ishida, T., Saito, Y. and Arima, K. (2013), Accumulation of a repulsive axonal guidance molecule RGMa in amyloid plaques: a possible hallmark of regenerative failure in Alzheimer's disease brains. Neuropathology and Applied Neurobiology, 39: 109–120. doi: 10.1111/j.1365-2990.2012.01281.x
- Issue online: 25 JAN 2013
- Version of Record online: 25 JAN 2013
- Accepted manuscript online: 14 MAY 2012 04:59PM EST
- Received 6 December 2011; Accepted after revision 4 May 2012; Published online Article Accepted on 14 May 2012
Figure S1. RGMa, NEO1, NTN1 and TGFβ1 mRNA expression in human brain tissues and cultured neural cells. The expression of (a) RGMa, (b) NEO1, (c) NTN1, (d) TGFβ1 and (e) G3PDH mRNAs was determined by RT-PCR. The lanes represent (1) the human frontal cerebral cortex (CBR) without inclusion of the reverse transcription (RT) step, (2) CBR with inclusion of the RT step, (3) cultured astrocytes (AS), (4) cultured neuronal progenitor (NP) cells, (5) NTera2 teratocarcinoma-derived neurones (NTera2N), (6) SK-N-SH neuroblastoma, (7) IMR-32 neuroblastoma, (8) U-373MG astrocytoma, (9) HMO6 microglia and (10) peripheral blood mononuclear cells (PBMC). The 100 bp ladder marker is shown on the left.
Figure S2. The specificity of anti-RGMa antibody, anti-neogenin antibody and recombinant protein probes for tissue protein overlay. (A) The specificity of anti-RGMa antibody (AF2459) and anti-neogenin (NEO1) antibody (sc-15337) was validated by Western blot of the recombinant RGMa protein tagged with V5 and the recombinant NEO1 tagged with Xpress, both of which were expressed in HEK293 cells. The panels represent (a) RGMa, (b) V5, (c) heat shock protein 60 (HSP60), an internal control of protein loading, (d) NEO1, (e) Xpress and (f) HSP60. The lanes represent (1, 3) non-transfected cells and (2, 4) the cells transfected with the vector expressing (a–c) RGMa or (d–f) NEO1. (B) The protein extract (50 μg protein/lane) of (lanes 5, 7) NTera2N and (lanes 6, 8) U-373MG was processed for Western blot for detection of (g) RGMa, (h) NEO1 and (i) HSP60. (C) The vector expressing the Flag-tagged CTFβ of APP or Flag-tagged GFP was cotransfected with the vector expressing Myc-tagged RGMa in HEK293 cells. The cellular protein extract was processed for immunoprecipitation (IP) with (j, l) anti-Flag antibody or (k, m) anti-Myc antibody. Then, the immunoprecipitates were processed for Western blot for detection of (j, l) Myc or (k, m) Flag. The lanes indicate (9, 12) input control, (10, 13) IP with anti-Flag or anti-Myc antibody and (11, 14) IP with normal mouse or rabbit IgG. (D) The recombinant Xpress-tagged RGMa or LacZ protein was processed for Western blot for detection of (o) Xpress or (p) RGMa. The lanes indicate (15) Xpress-tagged LacZ and (16) Xpress-tagged RGMa.
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