Tamoxifen (TMX) is a selective estrogen receptor modulator that can mimic the neuroprotective effects of estrogen but lacks its systemic adverse effects. We found that TMX (1 mg/day) significantly improved the motor recovery of partially paralyzed hind limbs of male adult rats with thoracic spinal cord injury (SCI), thus indicating a translational potential for this cancer medication given its clinical safety and applicability and the lack of currently available treatments for SCI. To shed light on the mechanisms underlying the beneficial effects of TMX for SCI, we used proteomic analyses, Western blots and histological assays, which showed that TMX treatment spared mature oligodendrocytes/increased myelin levels and altered reactive astrocytes, including the upregulation of the water channels aquaporin 4 (AQP4), a novel finding. AQP4 increases in TMX-treated SCI rats were associated with smaller fluid-filled cavities with borders consisting of densely packed AQP4-expressing astrocytes that closely resemble the organization of normal glia limitans externa (in contrast to large cavities in control SCI rats that lacked glia limitans-like borders and contained reactive glial cells). Based on our findings, we propose that TMX is a promising candidate for the therapeutic treatment of SCI and a possible intervention for other neuropathological conditions associated with demyelination and AQP4 dysfunction.
Get full access to this article
View all access options for this article.
References
1.
AmtulZ., WangL., WestawayD., and RozmahelR. F. (2010). Neuroprotective mechanism conferred by 17beta-estradiol on the biochemical basis of Alzheimer's disease. Neuroscience, 169, 781–786.
2.
BourqueM., DluzenD. E., and Di PaoloT. (2009). Neuroprotective actions of sex steroids in Parkinson's disease. Front Neuroendocrinol., 30, 142–157.
3.
Garcia-SeguraL. M. and MelcangiR. C. (2006). Steroids and glial cell function. Glia, 54, 485–498.
4.
LebesgueD., ChevaleyreV., ZukinR. S., and EtgenA. M. (2009). Estradiol rescues neurons from global ischemia-induced cell death: multiple cellular pathways of neuroprotection. Steroids, 74, 555–561.
5.
SamantarayS., SribnickE. A., DasA., ThakoreN. P., MatzelleD., YuS. P., RayS. K., WeiL., and BanikN. L. (2010). Neuroprotective efficacy of estrogen in experimental spinal cord injury in rats. Ann. N. Y. Acad. Sci., 1199, 90–94.
6.
MhyreA. J. and DorsaD. M. (2006). Estrogen activates rapid signaling in the brain: role of estrogen receptor alpha and estrogen receptor beta in neurons and glia. Neuroscience, 138, 851–858.
7.
HosliE., JurasinK., RuhlW., LuthyR., and HosliL. (2001). Colocalization of androgen, estrogen and cholinergic receptors on cultured astrocytes of rat central nervous system. Int. J. Dev. Neurosci., 19, 11–19.
8.
PlataniaP., LaureantiF., BellomoM., GiuffridaR., Giuffrida-StellaA. M., CataniaM. V., and SortinoM. A. (2003). Differential expression of estrogen receptors alpha and beta in the spinal cord during postnatal development: localization in glial cells. Neuroendocrinology, 77, 334–340.
9.
BrzozowskiA. M., PikeA. C., DauterZ., HubbardR. E., BonnT., EngstromO., OhmanL., GreeneG. L., GustafssonJ. A., and CarlquistM. (1997). Molecular basis of agonism and antagonism in the oestrogen receptor. Nature, 389, 753–758.
10.
PaigeL. A., ChristensenD. J., GronH., NorrisJ. D., GottlinE. B., PadillaK. M., ChangC. Y., BallasL. M., HamiltonP. T., McDonnellD. P., and FowlkesD. M. (1999). Estrogen receptor (ER) modulators each induce distinct conformational changes in ER alpha and ER beta. Proc. Natl. Acad. Sci. U. S. A., 96, 3999–4004.
11.
BelandiaB. and ParkerM. G. (2003). Nuclear receptors: a rendezvous for chromatin remodeling factors. Cell, 114, 277–280.
12.
KlingeC. M. (2000). Estrogen receptor interaction with co-activators and co-repressors. Steroids, 65, 227–251.
13.
McKennaN. J. and O'MalleyB. W. (2002). Combinatorial control of gene expression by nuclear receptors and coregulators. Cell, 108, 465–474.
14.
NorrisJ. D., PaigeL. A., ChristensenD. J., ChangC. Y., HuacaniM. R., FanD., HamiltonP. T., FowlkesD. M., and McDonnellD. P. (1999). Peptide antagonists of the human estrogen receptor. Science, 285, 744–746.
15.
Marin-HusstegeM., MuggironiM., RabanD., SkoffR. P., and Casaccia-BonnefilP. (2004). Oligodendrocyte progenitor proliferation and maturation is differentially regulated by male and female sex steroid hormones. Dev. Neurosci., 26, 245–254.
16.
Garcia-SeguraL. M., Cardona-GomezP., NaftolinF., and ChowenJ. A. (1998). Estradiol upregulates Bcl-2 expression in adult brain neurons. Neuroreport, 9, 593–597.
17.
de MedinaP., FavreG., and PoirotM. (2004). Multiple targeting by the antitumor drug tamoxifen: a structure-activity study. Curr. Med. Chem. Anticancer Agents, 4, 491–508.
18.
AmrollahiZ., RezaeiF., SalehiB., ModabberniaA. H., MaroufiA., EsfandiariG. R., NaderiM., GheblehF., Ahmadi-AbhariS. A., SadeghiM., TabriziM., and AkhondzadehS. (2011). Double-blind, randomized, placebo-controlled 6-week study on the efficacy and safety of the tamoxifen adjunctive to lithium in acute bipolar mania. J. Affect. Disord., 129, 327–331.
19.
MaoJ., YuanJ., WangL., ZhangH., JinX., ZhuJ., LiH., XuB., and ChenL. (2013). Tamoxifen inhibits migration of estrogen receptor-negative hepatocellular carcinoma cells by blocking the swelling-activated chloride current. J. Cell Physiol., 228, 991–1001.
20.
KimelbergH. K. (2005). Astrocytic swelling in cerebral ischemia as a possible cause of injury and target for therapy. Glia, 50, 389–397.
21.
ArevaloM. A., Diz-ChavesY., Santos-GalindoM., BelliniM. J., and Garcia-SeguraL. M. (2012). Selective oestrogen receptor modulators decrease the inflammatory response of glial cells. J. Neuroendocrinol., 24, 183–190.
22.
IsmailoğluO., OralB., GörgülüA., SütçüR., DemirN. (2010). Neuroprotective effects of tamoxifen on experimental spinal cord injury in rats. J. Clin Neurosci., 17, 1306–1310.
23.
TianD. S., LiuJ. L., XieM. J., ZhanY., QuW. S., YuZ. Y., TangZ. P., PanD. J., and WangW. (2009). Tamoxifen attenuates inflammatory-mediated damage and improves functional outcome after spinal cord injury in rats. J. Neurochem., 109, 1658–1667.
24.
Durham-LeeJ. C., WuY., MokkapatiV. U., Paulucci-HolthauzenA. A., and NesicO. (2012). Induction of angiopoietin-2 after spinal cord injury. Neuroscience, 202, 454–464.
25.
KisangaE. R., GjerdeJ., SchjottJ., MellgrenG., and LienE. A. (2003). Tamoxifen administration and metabolism in nude mice and nude rats. J. Steroid Biochem. Mol. Biol., 84, 361–367.
26.
RobinsH. I., WonM., SeiferheldW. F., SchultzC. J., ChoucairA. K., BrachmanD. G., DemasW. F., and MehtaM. P. (2006). Phase 2 trial of radiation plus high-dose tamoxifen for glioblastoma multiforme: RTOG protocol BR-0021. Neuro. Oncol., 8, 47–52.
27.
GreavesP., GoonetillekeR., NunnG., TophamJ., and OrtonT. (1993). Two-year carcinogenicity study of tamoxifen in Alderley Park Wistar-derived rats. Cancer Res., 53, 3919–3924.
28.
BassoD. M., BeattieM. S., and BresnahanJ. C. (1995). A sensitive and reliable locomotor rating scale for open field testing in rats. J. Neurotrauma, 12, 1–21.
29.
PretzerE. and WiktorowiczJ. E. (2008). Saturation fluorescence labeling of proteins for proteomic analyses. Anal. Biochem., 374, 250–262.
30.
StraubC., PazdrakK., YoungT. W., StaffordS. J., WuZ., WiktorowiczJ. E., HaagA. M., EnglishR. D., SomanK. V., and KuroskyA. (2009). Toward the Proteome of the Human Peripheral Blood Eosinophil. Proteomics. Clin. Appl., 3, 1151–1173.
31.
GuptarakJ., WanchooS., Durham-LeeJ., WuY., ZivadinovicD., Paulucci-HolthauzenA., and NesicO. (2013). Inhibition of IL-6 signaling:Anovel therapeutic approach to treating spinal 3cord injury pain. Pain, 154, 1115–1128.
32.
PapkoffJ., RubinfeldB., SchryverB., and PolakisP. (1996). Wnt-1 regulates free pools of catenins and stabilizes APC-catenin complexes. Mol. Cell Biol., 16, 2128–2134.
33.
McMahonS. S. and McDermottK. W. (2002). Morphology and differentiation of radial glia in the developing rat spinal cord. J. Comp Neurol., 454, 263–271.
34.
BenjaminiY. and HochbergY. (1995). Controlling the false discovery rate: a practical and powerful approach to multiple testing. J. R. Statist. Soc. B., 57, 289–300.
35.
R Development Core Team R: A language and environment for statistical computing. R Foundation for Statistical Computing: Vienna, Austria.
36.
FudgeM. A., KavaliersM., BairdJ. P., and OssenkoppK. P. (2009). Tamoxifen produces conditioned taste avoidance in male rats: an analysis of microstructural licking patterns and taste reactivity. Horm. Behav., 56, 322–331.
37.
GrayJ. M., SchrockS., and BishopM. (1993). Estrogens and antiestrogens: actions and interactions with fluphenazine on food intake and body weight in rats. Am. J. Physiol., 264, R1214–R1218.
38.
LopezM., LelliottC. J., TovarS., KimberW., GallegoR., VirtueS., BlountM., VazquezM. J., FinerN., PowlesT. J., O'RahillyS., SahaA. K., DieguezC., and Vidal-PuigA. J. (2006). Tamoxifen-induced anorexia is associated with fatty acid synthase inhibition in the ventromedial nucleus of the hypothalamus and accumulation of malonyl-CoA. Diabetes, 55, 1327–1336.
39.
WadeG. N. and HellerH. W. (1993). Tamoxifen mimics the effects of estradiol on food intake, body weight, and body composition in rats. Am. J. Physiol, 264, R1219–R1223.
40.
Kasama-YoshidaH., TohyamaY., KuriharaT., SakumaM., KojimaH., and TamaiY. (1997). A comparative study of 2′,3′-cyclic-nucleotide 3′-phosphodiesterase in vertebrates: cDNA cloning and amino acid sequences for chicken and bullfrog enzymes. J. Neurochem., 69, 1335–1342.
41.
LiuA., StadelmannC., MoscarelloM., BruckW., SobelA., MastronardiF. G., and Casaccia-BonnefilP. (2005). Expression of stathmin, a developmentally controlled cytoskeleton-regulating molecule, in demyelinating disorders. J. Neurosci., 25, 737–747.
42.
NielsenJ. A., MaricD., LauP., BarkerJ. L., and HudsonL. D. (2006). Identification of a novel oligodendrocyte cell adhesion protein using gene expression profiling. J. Neurosci., 26, 9881–9891.
43.
MondalS., DirksP., and RutkaJ. T. (2010). Immunolocalization of fascin, an actin-bundling protein and glial fibrillary acidic protein in human astrocytoma cells. Brain Pathol., 20, 190–199.
44.
YamasakiM., YamadaK., FuruyaS., MitomaJ., HirabayashiY., and WatanabeM. (2001). 3-Phosphoglycerate dehydrogenase, a key enzyme for l-serine biosynthesis, is preferentially expressed in the radial glia/astrocyte lineage and olfactory ensheathing glia in the mouse brain. J. Neurosci., 21, 7691–7704.
45.
BergR. W., ChenM. T., HuangH. C., HsiaoM. C., and ChengH. (2009). A method for unit recording in the lumbar spinal cord during locomotion of the conscious adult rat. J. Neurosci. Methods, 182, 49–54.
46.
FalgairolleM., de SezeM., JuvinL., MorinD., and CazaletsJ. R. (2006). Coordinated network functioning in the spinal cord: an evolutionary perspective. J. Physiol. Paris, 100, 304–316.
47.
FussB., MallonB., PhanT., OhlemeyerC., KirchhoffF., NishiyamaA., and MacklinW. B. (2000). Purification and analysis of in vivo-differentiated oligodendrocytes expressing the green fluorescent protein. Dev. Biol., 218, 259–274.
48.
NesicO., LeeJ., YeZ., UnabiaG. C., RafatiD., HulseboschC. E., and Perez-PoloJ. R. (2006). Acute and chronic changes in aquaporin 4 expression after spinal cord injury. Neuroscience, 143, 779–792.
49.
TatorC. H., HashimotoR., RaichA., NorvellD., FehlingsM. G., HarropJ. S., GuestJ., AarabiB., and GrossmanR. G. (2012). Translational potential of preclinical trials of neuroprotection through pharmacotherapy for spinal cord injury. J. Neurosurg. Spine, 17, 157–229.
50.
GuestJ., HarropJ. S., AarabiB., GrossmanR. G., FawcettJ. W., FehlingsM. G., and TatorC. H. (2012). Optimization of the decision-making process for the selection of therapeutics to undergo clinical testing for spinal cord injury in the North American Clinical Trials Network. J. Neurosurg. Spine, 17, 94–101.
51.
O'BrianC. A., LiskampR. M., SolomonD. H., and WeinsteinI. B. (1985). Inhibition of protein kinase C by tamoxifen. Cancer Res., 45, 2462–2465.
52.
O'BrianC. A., HouseyG. M., and WeinsteinI. B. (1988). Specific and direct binding of protein kinase C to an immobilized tamoxifen analogue. Cancer Res., 48, 3626–3629.
53.
ZarateC. A., Jr., SinghJ. B., CarlsonP. J., QuirozJ., JolkovskyL., LuckenbaughD. A., and ManjiH. K. (2007). Efficacy of a protein kinase C inhibitor (tamoxifen) in the treatment of acute mania: a pilot study. Bipolar Disord., 9, 561–570.
54.
KisangaE. R., GjerdeJ., Guerrieri-GonzagaA., PigattoF., Pesci-FeltriA., RobertsonC., SerranoD., PelosiG., DecensiA., and LienE. A. (2004). Tamoxifen and metabolite concentrations in serum and breast cancer tissue during three dose regimens in a randomized preoperative trial. Clin. Cancer Res., 10, 2336–2343.
55.
LochabS., PalP., KanaujiyaJ. K., TripathiS. B., KapoorI., BhattM. L., SanyalS., BehreG., and TrivediA. K. (2012). Proteomic identification of E6AP as a molecular target of tamoxifen in MCF7 cells. Proteomics, 12, 1363–1377.
56.
Durham-LeeJ. C., MokkapatiV. U., JohnsonK. M., and NesicO. (2011). Amiloride improves locomotor recovery after spinal cord injury. J. Neurotrauma, 28, 1319–1326.
57.
CaoQ., HeQ., WangY., ChengX., HowardR. M., ZhangY., DeVriesW. H., ShieldsC. B., MagnusonD. S., XuX. M., KimD. H., and WhittemoreS. R. (2010). Transplantation of ciliary neurotrophic factor-expressing adult oligodendrocyte precursor cells promotes remyelination and functional recovery after spinal cord injury. J. Neurosci., 30, 2989–3001.
58.
CroweM. J., BresnahanJ. C., ShumanS. L., MastersJ. N., and BeattieM. S. (1997). Apoptosis and delayed degeneration after spinal cord injury in rats and monkeys. Nat. Med., 3, 73–76.
59.
LiuX. Z., XuX. M., HuR., DuC., ZhangS. X., McDonaldJ. W., DongH. X., WuY. J., FanG. S., JacquinM. F., HsuC. Y., and ChoiD. W. (1997). Neuronal and glial apoptosis after traumatic spinal cord injury. J. Neurosci., 17, 5395–5406.
60.
ShumanS. L., BresnahanJ. C., and BeattieM. S. (1997). Apoptosis of microglia and oligodendrocytes after spinal cord contusion in rats. J. Neurosci. Res., 50, 798–808.
61.
LeeJ. Y., ChoiS. Y., OhT. H., and YuneT. Y. (2012). 17beta-Estradiol inhibits apoptotic cell death of oligodendrocytes by inhibiting RhoA-JNK3 activation after spinal cord injury. Endocrinology, 153, 3815–3827.
62.
LiQ. M., TepC., YuneT. Y., ZhouX. Z., UchidaT., LuK. P., and YoonS. O. (2007). Opposite regulation of oligodendrocyte apoptosis by JNK3 and Pin1 after spinal cord injury. J. Neurosci., 27, 8395–8404.
63.
ArevaloM. A., Santos-GalindoM., LagunasN., AzcoitiaI., and Garcia-SeguraL. M. (2011). Selective estrogen receptor modulators as brain therapeutic agents. J. Mol. Endocrinol., 46, R1–R9.
64.
CerciatM., UnkilaM., Garcia-SeguraL. M., and ArevaloM. A. (2010). Selective estrogen receptor modulators decrease the production of interleukin-6 and interferon-gamma-inducible protein-10 by astrocytes exposed to inflammatory challenge in vitro. Glia, 58, 93–102.
65.
SudaS., Segi-NishidaE., NewtonS.S., DumanR.S. (2008). A postpartum model in rat: behavioral and gene expression changes induced by ovarian steroid deprivation. Biol. Psychiatry, 64, 311–319.
66.
RutkowskyJ.M., WallaceB.K., WiseP.M., O'DonnellM.E. (2011). Effects of estradiol on ischemic factor-induced astrocyte swelling and AQP4 protein abundance. Am. J. Physiol. Cell Physiol., 301, C204–12.
67.
NesicO., GuestJ. D., ZivadinovicD., NarayanaP. A., HerreraJ. J., GrillR. J., MokkapatiV. U., GelmanB. B., and LeeJ. (2010). Aquaporins in spinal cord injury: the janus face of aquaporin 4. Neuroscience, 168, 1019–1035.
68.
KimuraA., HsuM., SeldinM., VerkmanA. S., ScharfmanH. E., and BinderD. K. (2010). Protective role of aquaporin-4 water channels after contusion spinal cord injury. Ann. Neurol., 67, 794–801.
69.
NielsenS., NagelhusE. A., Amiry-MoghaddamM., BourqueC., AgreP., and OttersenO. P. (1997). Specialized membrane domains for water transport in glial cells: high-resolution immunogold cytochemistry of aquaporin-4 in rat brain. J. Neurosci., 17, 171–180.
70.
HolenT. (2011). The ultrastructure of lamellar stack astrocytes. Glia, 59, 1075–1083.
71.
CraneJ. M., BennettJ. L., and VerkmanA. S. (2009). Live cell analysis of aquaporin-4 m1/m23 interactions and regulated orthogonal array assembly in glial cells. J. Biol. Chem., 284, 35850–35860.
72.
SorboJ. G., MoeS. E., OttersenO. P., and HolenT. (2008). The molecular composition of square arrays. Biochemistry, 47, 2631–2637.
73.
RateladeJ. and VerkmanA. S. (2012). Neuromyelitis optica: aquaporin-4 based pathogenesis mechanisms and new therapies. Int. J. Biochem. Cell Biol., 44, 1519–1530.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
