@article{e738dc3b600b4ff886f5341715264aee,
title = "Rewired glycosylation activity promotes scarless regeneration and functional recovery in spiny mice after complete spinal cord transection",
abstract = "Regeneration of adult mammalian central nervous system (CNS) axons is abortive, resulting in inability to recover function after CNS lesion, including spinal cord injury (SCI). Here, we show that the spiny mouse (Acomys) is an exception to other mammals, being capable of spontaneous and fast restoration of function after severe SCI, re-establishing hind limb coordination. Remarkably, Acomys assembles a scarless pro-regenerative tissue at the injury site, providing a unique structural continuity of the initial spinal cord geometry. The Acomys SCI site shows robust axon regeneration of multiple tracts, synapse formation, and electrophysiological signal propagation. Transcriptomic analysis of the spinal cord following transcriptome reconstruction revealed that Acomys rewires glycosylation biosynthetic pathways, culminating in a specific pro-regenerative proteoglycan signature at SCI site. Our work uncovers that a glycosylation switch is critical for axon regeneration after SCI and identifies β3gnt7, a crucial enzyme of keratan sulfate biosynthesis, as an enhancer of axon growth.",
keywords = "acetylglucosaminyltransferase, Acomys cahirinus, axon regeneration, fibrotic scar, glial scar, glycosaminoglycan, glycosylation, proteoglycan, spinal cord injury, spiny mouse",
author = "Joana Nogueira-Rodrigues and Leite, {S{\'e}rgio C.} and Rita Pinto-Costa and Sousa, {Sara C.} and Luz, {Liliana L.} and Sintra, {Maria A.} and Raquel Oliveira and Monteiro, {Ana C.} and Pinheiro, {Gon{\c c}alo G.} and Marta Vitorino and Silva, {Joana A.} and S{\'o}nia Sim{\~a}o and Fernandes, {Vitor E.} and Jan Provazn{\'i}k and Vladimir Benes and Cruz, {C{\'e}lia D.} and Safronov, {Boris V.} and Ana Magalh{\~a}es and Reis, {Celso A.} and Jorge Vieira and Vieira, {Cristina P.} and Gustavo Tisc{\'o}rnia and Ara{\'u}jo, {In{\^e}s M.} and Sousa, {M{\'o}nica M.}",
note = "Funding Information: We thank the i3S Scientific Platforms (Animal, Histology and Electron Microscopy, Advanced Light Microscopy and BioSciences Screening Facilities, part of the national infrastructure Portuguese Platform of Bioimaging [PPBI]; PPBI-POCI-01-0145-FEDER-022122). This project was supported by Santa Casa da Miseric{\'o}rdia de Lisboa ( MC-39-2019 ), Wings for Life ( WFL-PT-21/20 ), FCT ( UID/BIM/04773/2013/CBMR , PTDC/BIA-ANM/0697/2014 , PTDC/MED-ONC/28489/2017 ). J.N.R. and S.C.S. are FCT fellows ( SFRH/BD/131565/2017 ; SFRH/BD/136760/2018 ). Funding Information: High-throughput analysis of the extracellular proteome in rats has supported the importance of inflammation and ECM composition in the response mounted upon SCI (Didangelos et al., 2016). Studies comparing non-regenerative and regenerative non-mammalian species after SCI, including the axolotl (Tica and Didangelos, 2018) and zebrafish (Tsata et al., 2021) have further pinpointed inflammation and ECM components as critical factors in interspecies regenerative differences. Similar to Acomys, in zebrafish, the deposition of a growth-supporting ECM deprived of growth inhibitory matrix molecules is crucial for axon regeneration after SCI (Tsata et al., 2021). Furthermore, in zebrafish, a regenerative permissive milieu is enabled by the recruitment of pdgfrb+ myoseptal and perivascular cells, in a PDGFR signaling-dependent manner (Tsata et al., 2021). In Acomys, we unveiled the glycosylation gene regulatory network as a key spinal cord-specific pathway that differentiates the Acomys and Mus lesion environment. In fact, injured Acomys exhibits a specific signature of ECM glycosaminoglycans at the SCI site. Although previous studies reported the potential role of KSPG as an inhibitor of axon regeneration (Imagama et al., 2011; Ishikawa et al., 2015; Jones and Tuszynski, 2002), here, we show that in fact KSPG glycosaminoglycan chains are increased in the pro-regenerative environment of the Acomys lesion site. Moreover, we identify ?3gnt7, a crucial enzyme in KSPG biosynthesis, as a potent enhancer of axon growth, which may be explored in the future design of therapies to enhance axon regeneration after SCI.Uninjured Mus (n=5) and Acomys (n=4), and injured Mus (n=3) and Acomys (n=10) were terminally anaesthetized with an i.p. injection of 50 mg/kg of pentobarbital at 8 WPI. Spinal cord from the upper cervical to lower lumbar levels was quickly removed, cleaned from the meninges and nerve roots, and kept in artificial cerebrospinal fluid (aCSF) containing: 115 mM NaCl (Sigma, S9888), 3 mM KCl (Sigma, 7447-40-7), 2.2 mM CaCl2 (Sigma, 10035-04-8), 1 mM MgCl2 (Sigma, M2393), 1 mM NaH2PO4 (Sigma, 10049-21-5), 25 mM NaHCO3 (Sigma, 144-55-8) and 11 mM glucose (Sigma, G8270) (bubbled with 95% O2 and 5% CO2). The spinal cord was transferred to the recording chamber perfused with oxygenated aCSF and allowed to recover for at least 45 min. All recordings were done at a temperature of 22?24?C. Suction electrodes fabricated from thick-walled glass (BioMedical Instruments, Germany) were used for both stimulation and recording. To study CAP conduction in descending motor tracts, the lateral funiculus of each side of the spinal cords (right/left) was stimulated at a location 8-10 mm rostral to the lesion site (Stim. A; Figure 2L) using the Isolated Pulse Stimulator (2100, A-M Systems, RRID:SCR_016677). Two recording electrodes were positioned 4-5 mm rostrally (Rec. B; Figure 2L) and 4-5 mm caudally to the lesion (Rec. C; Figure 2L), to compare CAP amplitudes before and after crossing the lesion site, thus revealing functional fibers crossing the scar. The recording electrode Rec.B allowed to control the viability of the spinal cord preparation and the proper propagation of CAPs. Each suction electrode had its own reference electrode. Spinal cords were stimulated with a 50 ?s pulse of 200 ?A at a 3s interval. For recording, we used the differential AC amplifier (1700, A?M Systems), in which the low cut-off filter was set 0.1 Hz. The signal was online low-pass filtered at 10 kHz and sampled using the A/D converter of the EPC9 amplifier (HEKA, Lambrecht, Germany). Of note, only injured Acomys that recovered weight support and presented locomotor recovery (BMS > 5) were used. Quantification of CAP conduction was performed by measuring the amplitude for each animal with the average of at least 10 traces.We thank the i3S Scientific Platforms (Animal, Histology and Electron Microscopy, Advanced Light Microscopy and BioSciences Screening Facilities, part of the national infrastructure Portuguese Platform of Bioimaging [PPBI]; PPBI-POCI-01-0145-FEDER-022122). This project was supported by Santa Casa da Miseric?rdia de Lisboa (MC-39-2019), Wings for Life (WFL-PT-21/20), FCT (UID/BIM/04773/2013/CBMR, PTDC/BIA-ANM/0697/2014, PTDC/MED-ONC/28489/2017). J.N.R. and S.C.S. are FCT fellows (SFRH/BD/131565/2017; SFRH/BD/136760/2018). M.M.S. coordinated the study; G.T. and I.M.A. coordinated research at University of Algarve; M.M.S. J.N.-R. C.D.C. B.V.S. A.M. and C.R. designed and analyzed experiments; J.N.R. S.C.L. R.P.-C. S.C.S. L.L.L. M.A.S. R.O. and A.C.M. performed experiments and quantifications; G.G.P. M.V. J.A.S. S.S. and V.E.F. supported preliminary experiments in Acomys; V.B. and J.P. performed RNA-seq experiments; C.P.V. and J.V. performed RNA-seq analyzes and their interpretation; M.M.S. and J.N.R. wrote the paper with input from all authors. The authors declare no competing interests. Publisher Copyright: {\textcopyright} 2021 Elsevier Inc.",
year = "2022",
month = feb,
day = "28",
doi = "10.1016/j.devcel.2021.12.008",
language = "English",
volume = "57",
pages = "440--450.e7",
journal = "Developmental Cell",
issn = "1534-5807",
publisher = "Cell Press",
number = "4",
}