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Controlled delivery of a neurotransmitter–agonist conjugate for functional recovery after severe spinal cord injury

Nature Nanotechnology volume 18, pages 1230–1240 (2023)Cite this article

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Abstract

Despite considerable unmet medical needs, effective pharmacological treatments that promote functional recovery after spinal cord injury remain limited. Although multiple pathological events are implicated in spinal cord injuries, the development of a microinvasive pharmacological approach that simultaneously targets the different mechanisms involved in spinal cord injury remains a formidable challenge. Here we report the development of a microinvasive nanodrug delivery system that consists of amphiphilic copolymers responsive to reactive oxygen species and an encapsulated neurotransmitter-conjugated KCC2 agonist. Upon intravenous administration, the nanodrugs enter the injured spinal cord due to a disruption in the blood–spinal cord barrier and disassembly due to damage-triggered reactive oxygen species. The nanodrugs exhibit dual functions in the injured spinal cord: scavenging accumulated reactive oxygen species in the lesion, thereby protecting spared tissues, and facilitating the integration of spared circuits into the host spinal cord through targeted modulation of inhibitory neurons. This microinvasive treatment leads to notable functional recovery in rats with contusive spinal cord injury.

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Fig. 1: Design and characterization of smart nanodrugs.
Fig. 2: Smart nanodrugs accumulate at the injury site in the spinal cord and selectively target specific neurons.
Fig. 3: The ROS-responsive nanodrug delivery system delivers small molecules into the spinal cord even after BSCB restoration.
Fig. 4: ROS Nano treatment protects spared tissues/axons from secondary injury.
Fig. 5: GABA Nano treatment improves the recovery of hind-limb locomotor function in rats with severe contusive SCI.
Fig. 6: GABA Nano treatment rebalances neuronal activity and activates rescued residual spinal cord circuits to perform functions.

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Data availability

The data to support the findings of this study are included in the paper, and further data are available from the corresponding author. Source data are provided with this paper.

References

  1. David, S. & Kroner, A. Repertoire of microglial and macrophage responses after spinal cord injury. Nat. Rev. Neurosci. 12, 388–399 (2011).

    Article  CAS  Google Scholar 

  2. Block, M. L., Zecca, L. & Hong, J. S. Microglia-mediated neurotoxicity: uncovering the molecular mechanisms. Nat. Rev. Neurosci. 8, 57–69 (2007).

    Article  CAS  Google Scholar 

  3. Ulndreaj, A., Badner, A. & Fehlings, M. G. Promising neuroprotective strategies for traumatic spinal cord injury with a focus on the differential effects among anatomical levels of injury. F1000Research 6, 1907 (2017).

    Article  Google Scholar 

  4. Li, L. et al. A MnO2 nanoparticle-dotted hydrogel promotes spinal cord repair via regulating reactive oxygen species microenvironment and synergizing with mesenchymal stem cells. ACS Nano 13, 14283–14293 (2019).

    Article  CAS  Google Scholar 

  5. Zhang, N. et al. A 3D fiber-hydrogel based non-viral gene delivery platform reveals that microRNAs promote axon regeneration and enhance functional recovery following spinal cord injury. Adv. Sci. 8, e2100805 (2021).

    Article  Google Scholar 

  6. Chen, B. et al. Reactivation of dormant relay pathways in injured spinal cord by KCC2 manipulations. Cell 174, 521–535.e13 (2018).

    Article  CAS  Google Scholar 

  7. Wilson, J. M., Blagovechtchenski, E. & Brownstone, R. M. Genetically defined inhibitory neurons in the mouse spinal cord dorsal horn: a possible source of rhythmic inhibition of motoneurons during fictive locomotion. J. Neurosci. 30, 1137–1148 (2010).

    Article  CAS  Google Scholar 

  8. Haring, M. et al. Neuronal atlas of the dorsal horn defines its architecture and links sensory input to transcriptional cell types. Nat. Neurosci. 21, 869–880 (2018).

    Article  Google Scholar 

  9. Brommer, B. et al. Improving hindlimb locomotor function by non-invasive AAV-mediated manipulations of propriospinal neurons in mice with complete spinal cord injury. Nat. Commun. 12, 781 (2021).

    Article  CAS  Google Scholar 

  10. Courtine, G. & Sofroniew, M. V. Spinal cord repair: advances in biology and technology. Nat. Med. 25, 898–908 (2019).

    Article  CAS  Google Scholar 

  11. Ramirez-Jarquin, U. N., Lazo-Gomez, R., Tovar, Y. R. L. B. & Tapia, R. Spinal inhibitory circuits and their role in motor neuron degeneration. Neuropharmacology 82, 101–107 (2014).

    Article  CAS  Google Scholar 

  12. Matsuya, R., Ushiyama, J. & Ushiba, J. Inhibitory interneuron circuits at cortical and spinal levels are associated with individual differences in corticomuscular coherence during isometric voluntary contraction. Sci. Rep. 7, 44417 (2017).

    Article  CAS  Google Scholar 

  13. Ramirez-Jarquin, U. N. & Tapia, R. Excitatory and inhibitory neuronal circuits in the spinal cord and their role in the control of motor neuron function and degeneration. ACS Chem. Neurosci. 9, 211–216 (2018).

    Article  CAS  Google Scholar 

  14. Rivera, C. et al. The K+/Cl– co-transporter KCC2 renders GABA hyperpolarizing during neuronal maturation. Nature 397, 251–255 (1999).

    Article  CAS  Google Scholar 

  15. Boulenguez, P. et al. Down-regulation of the potassium-chloride cotransporter KCC2 contributes to spasticity after spinal cord injury. Nat. Med. 16, 302–307 (2010).

    Article  CAS  Google Scholar 

  16. Gagnon, M. et al. Chloride extrusion enhancers as novel therapeutics for neurological diseases. Nat. Med. 19, 1524–1528 (2013).

    Article  CAS  Google Scholar 

  17. Reinig, S., Driever, W. & Arrenberg, A. B. The descending diencephalic dopamine system is tuned to sensory stimuli. Curr. Biol. 27, 318–333 (2017).

    Article  CAS  Google Scholar 

  18. Li, Y. et al. Pericytes impair capillary blood flow and motor function after chronic spinal cord injury. Nat. Med. 23, 733–741 (2017).

    Article  CAS  Google Scholar 

  19. Sharples, S. A. et al. A dynamic role for dopamine receptors in the control of mammalian spinal networks. Sci. Rep. 10, 16429 (2020).

    Article  CAS  Google Scholar 

  20. Grillner, S. & Jessell, T. M. Measured motion: searching for simplicity in spinal locomotor networks. Curr. Opin. Neurobiol. 19, 572–586 (2009).

    Article  CAS  Google Scholar 

  21. Li, W. C. & Moult, P. R. The control of locomotor frequency by excitation and inhibition. J. Neurosci. 32, 6220–6230 (2012).

    Article  CAS  Google Scholar 

  22. Kiehn, O. Decoding the organization of spinal circuits that control locomotion. Nat. Rev. Neurosci. 17, 224–238 (2016).

    Article  CAS  Google Scholar 

  23. Jiang, X. C. et al. Neural stem cells transfected with reactive oxygen species–responsive polyplexes for effective treatment of ischemic stroke. Adv. Mater. 31, e1807591 (2019).

    Article  Google Scholar 

  24. Liu, P. et al. Biomimetic dendrimer–peptide conjugates for early multi-target therapy of Alzheimer’s disease by inflammatory microenvironment modulation. Adv. Mater. 33, e2100746 (2021).

    Article  Google Scholar 

  25. Lu, Y. et al. Microenvironment remodeling micelles for Alzheimer’s disease therapy by early modulation of activated microglia. Adv. Sci. 6, 1801586 (2019).

    Article  Google Scholar 

  26. Xu, W. et al. Increased production of reactive oxygen species contributes to motor neuron death in a compression mouse model of spinal cord injury. Spinal Cord 43, 204–213 (2005).

    Article  CAS  Google Scholar 

  27. Zhang, M. et al. Oxidation and temperature dual responsive polymers based on phenylboronic acid and N-isopropylacrylamide motifs. Polym. Chem. 7, 1494–1504 (2016).

    Article  CAS  Google Scholar 

  28. Lin, L. et al. Nanodrug with ROS and pH dual-sensitivity ameliorates liver fibrosis via multicellular regulation. Adv. Sci. 7, 1903138 (2020).

    Article  CAS  Google Scholar 

  29. Zhang, D., Fan, Y., Chen, H., Trepout, S. & Li, M. H. CO2-activated reversible transition between polymersomes and micelles with AIE fluorescence. Angew. Chem. Int. Ed. 58, 10260–10265 (2019).

    Article  CAS  Google Scholar 

  30. Suk, J. S., Xu, Q., Kim, N., Hanes, J. & Ensign, L. M. PEGylation as a strategy for improving nanoparticle-based drug and gene delivery. Adv. Drug Deliv. Rev. 99, 28–51 (2016).

    Article  CAS  Google Scholar 

  31. Hu, J. et al. Long circulating polymeric nanoparticles for gene/drug delivery. Curr. Drug Metab. 19, 723–738 (2018).

    Article  CAS  Google Scholar 

  32. Zhang, Z. et al. Circulatory disturbance of rat spinal cord induced by occluding ligation of the dorsal spinal vein. Acta Neuropathol. 102, 335–338 (2001).

    Article  CAS  Google Scholar 

  33. Farrar, M. J., Rubin, J. D., Diago, D. M. & Schaffer, C. B. Characterization of blood flow in the mouse dorsal spinal venous system before and after dorsal spinal vein occlusion. J. Cereb. Blood Flow. Metab. 35, 667–675 (2015).

    Article  Google Scholar 

  34. Bartanusz, V., Jezova, D., Alajajian, B. & Digicaylioglu, M. The blood–spinal cord barrier: morphology and clinical implications. Ann. Neurol. 70, 194–206 (2011).

    Article  Google Scholar 

  35. Jin, L. Y. et al. Blood–spinal cord barrier in spinal cord injury: a review. J. Neurotrauma 38, 1203–1224 (2021).

    Article  Google Scholar 

  36. Zrzavy, T. et al. Acute and non-resolving inflammation associate with oxidative injury after human spinal cord injury. Brain 144, 144–161 (2021).

    Article  Google Scholar 

  37. Cooney, S. J., Zhao, Y. & Byrnes, K. R. Characterization of the expression and inflammatory activity of NADPH oxidase after spinal cord injury. Free Radic. Res. 48, 929–939 (2014).

    Article  CAS  Google Scholar 

  38. Bakh, N. A. et al. Glucose-responsive insulin by molecular and physical design. Nat. Chem. 9, 937–943 (2017).

    Article  CAS  Google Scholar 

  39. Chou, D. H. et al. Glucose-responsive insulin activity by covalent modification with aliphatic phenylboronic acid conjugates. Proc. Natl Acad. Sci. USA 112, 2401–2406 (2015).

    Article  CAS  Google Scholar 

  40. Ahuja, C. S. et al. Traumatic spinal cord injury. Nat. Rev. Dis. Prim. 3, 17018 (2017).

    Article  Google Scholar 

  41. Li, X. et al. The effect of a nanofiber-hydrogel composite on neural tissue repair and regeneration in the contused spinal cord. Biomaterials 245, 119978 (2020).

    Article  CAS  Google Scholar 

  42. Schucht, P., Raineteau, O., Schwab, M. E. & Fouad, K. Anatomical correlates of locomotor recovery following dorsal and ventral lesions of the rat spinal cord. Exp. Neurol. 176, 143–153 (2002).

    Article  CAS  Google Scholar 

  43. Qiao, Y. et al. Spinal dopaminergic mechanisms regulating the micturition reflex in male rats with complete spinal cord injury. J. Neurotrauma 38, 803–817 (2021).

    Article  Google Scholar 

  44. Shi, Y. et al. Effective repair of traumatically injured spinal cord by nanoscale block copolymer micelles. Nat. Nanotechnol. 5, 80–87 (2010).

    Article  CAS  Google Scholar 

  45. Ye, J. et al. Rationally designed, self-assembling, multifunctional hydrogel depot repairs severe spinal cord injury. Adv. Health. Mater. 10, e2100242 (2021).

    Article  Google Scholar 

  46. Watson, C. et al. in The Spinal Cord Ch 15 (Academic Press, 2008).

  47. Hong, L. T. A. et al. An injectable hydrogel enhances tissue repair after spinal cord injury by promoting extracellular matrix remodeling. Nat. Commun. 8, 533 (2017).

    Article  Google Scholar 

  48. Basso, D. M., Beattie, M. S. & Bresnahan, J. C. Graded histological and locomotor outcomes after spinal cord contusion using the NYU weight-drop device versus transection. Exp. Neurol. 139, 244–256 (1996).

    Article  CAS  Google Scholar 

  49. Wenger, N. et al. Spatiotemporal neuromodulation therapies engaging muscle synergies improve motor control after spinal cord injury. Nat. Med. 22, 138–145 (2016).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We acknowledge the excellent technical staff at the Imaging Facility, Core Facility of Zhejiang University School of Medicine and Center of Cryo-Electron Microscopy of Zhejiang University for their assistance with confocal laser scanning microscopy and TEM. This study was supported by the Scientific and Technological Innovation 2030 Program of China, major projects (2021ZD0200408 to X.W.); the National Natural Science Foundation of China (81971866 to X.W.); the Science Fund for Distinguished Young Scholars of Zhejiang Province (LR20H090002 to X.W.); the Leading Innovative and Entrepreneur Team Introduction Program of Zhejiang (2019R01007 to X.W.); the Fundamental Research Funds for the Central Universities (K20210195 to X.W.); and the National Major Project of Research and Development (2017YFA0104701 to B.Y.).

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Author notes

  1. These authors contributed equally: Yanming Zuo, Jingjia Ye.

Authors and Affiliations

  1. Department of Neurobiology and Department of Rehabilitation Medicine, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, P. R. China

    Yanming Zuo, Jingjia Ye, Zeinab Abdelrahman, Zuobin Chen & Xuhua Wang

  2. Liangzhu Laboratory, MOE Frontier Science Center for Brain Science and Brain-machine Integration, State Key Laboratory of Brain-machine Intelligence, Zhejiang University, Hangzhou, China

    Yanming Zuo, Jingjia Ye, Wanxiong Cai, Binjie Guo, Xiangfeng Chen, Lingmin Lin, Shuang Jin, Hanyu Zheng, Ao Fang, Xingran Qian, Zeinab Abdelrahman, Zhiping Wang & Xuhua Wang

  3. NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University, Hangzhou, China

    Yanming Zuo, Jingjia Ye, Wanxiong Cai, Binjie Guo, Xiangfeng Chen, Lingmin Lin, Shuang Jin, Hanyu Zheng, Ao Fang, Xingran Qian, Zeinab Abdelrahman, Zhiping Wang & Xuhua Wang

  4. School of Chemistry and Molecular Engineering, Frontiers Science Center for Materiobiology and Dynamic Chemistry, East China University of Science & Technology, Shanghai, P. R. China

    Zhipeng Zhang

  5. Key Laboratory of Neuroregeneration of Jiangsu and Ministry of Education, Nantong University, Nantong, P. R. China

    Bin Yu & Xiaosong Gu

  6. Co-innovation Center of Neuroregeneration, Nantong University, Nantong, P. R. China

    Xuhua Wang

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Contributions

X.W. and Y.Z. conceptualized and designed the study. Y.Z., J.Y., W.C., X.C., L.L., B.G., S.J., H.Z., A.F., X.Q. and X.W. conducted the experiments. Y.Z., J.Y. and W.C. collected the data. Y.Z., J.Y., W.C., B.G., X.G., Z.W., Z.C., Z.Z., B.Y. and X.W. analysed and interpreted the data. Y.Z., X.W. and J.Y. draughted the paper. All authors critically revised the manuscript and approved the final version for submission.

Corresponding author

Correspondence to Xuhua Wang.

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Competing interests

Zhejiang University has filed a patent application related to this work, with X.W., Y.Z., J.Y., W.C., X.C., L.L. and B.G. listed as inventors. X.W. is a scientific cofounder of WeQure AI Ltd. All the other authors declare no competing interests.

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Extended data

Extended Data Fig. 1 Synthesis and characterization of amphiphilic polymers and nanoparticles.

a, The synthetic route of a representative amphiphilic polymer. b, c, Representative 1H-NMR spectra of POEGMA30 and POEGMA30-BAA2. d, e, Properties of nanoparticles with different compositions of amphiphilic polymers. f-h, Representative TEM images of ROS Nano, DOPA Nano and GABA Nano. Scale bar, 100 nm. Independent experiments were repeated 3 times with similar results.

Source data

Extended Data Fig. 2 ROS Nano treatment protects spare propriospinal axons from secondary injury.

a, Schematic diagram of the experimental design. b, c, Representative images of cross sections at certain sites stained by RFP (representing propriospinal axon, red) and GFAP (green) in PBS- or ROS Nano-treated rats. Scale bar, 500 μm d, Quantitative analysis of propriospinal axons in rats treated with 9 weeks of ROS Nano or PBS. Data are shown as the mean ± SEM. Two-tailed paired t tests were used for comparisons between two groups. n = 3 rats for each group. p−4 = 0.0025, p−2 < 0.001, p0 = 0.0155, p2 = 0.004, p4 = 0.0220, * and ** indicate p < 0.05 and p < 0.01, respectively.

Source data

Extended Data Fig. 3 ROS Nano treatment relieved excessive inflammation and reduced apoptosis in the injured spinal cord.

a, Schematic diagram of the experimental design. b, Principal component analysis (PCA) of cytokine/chemokine expression in the spinal cord from 3 donors with different treatments. The effect of various ligand stimulations is largely described by PC1. c, Hierarchical clustering of the expression of cytokines/chemokines in the spinal cord from 3 donors with different treatments. The differential expression level is represented as the log2-fold change (log2[FC]) of normalized expression counts. Different treatments are arrayed by row and cytokine/chemokine by column. d, Volcano plot of cytokines/chemokines between the ROS Nano and PBS groups. Thresholds: −Log10 adjusted p value < 0.05, Log2[FC] >1. All data is presented in supplementary table. e, Quantitative analyses of MCP-1, GRO/KC and MIP-1α expression in the spinal cord after different treatments. n = 3 rats for each group. One-way ANOVA with Tukey’s post hoc test was used for comparisons among multiple groups. Two-tailed unpaired t tests were used for comparisons between two groups. ROS vs PBS (MCP-1, p = 0.00335), (GRO/KC, p = 0.00328) and (MIP-1α, p = 0.0113). * indicates p < 0.05 and ** indicates p < 0.001. Data are shown as the mean ± SEM. f,g, Representative immunoblots and quantitative analyses of TGF-β(p = 0.0419), BCL-2 (p = 0.0439) and BAX (p = 0.0271) expression in the injury site 1 week after PBS or ROS Nano (10 mg/kg) treatment. n = 3 rats for each group. Two-tailed unpaired t tests were used for comparisons between two groups. * indicates p < 0.05. Data are shown as the mean ± SEM. h, Representative images of transverse sections at the epicenter of an injury stained with iNOS (red)/IBA1 (green) or NeuN (red)/GFAP (green) after 4 weeks of treatment.

Source data

Extended Data Fig. 4 The preserved spinal cord circuits mediate limited hindlimb locomotor functional recovery.

a, Weekly BBB scores of the experimental rats treated with PBS or ROS Nano. Data are shown as the mean ± SEM. Two-tailed paired t tests were used for comparisons between two groups. n = 10 rats for each group. b, A simplified stick mode shows a one-step cycle of one intact hindlimb while the rat freely walked. The model also shows the quantification of the hip, knee and ankle angles and iliac crest height amplitudes during the cycle. c, Representative color-coded stick views of kinematic hindlimb movement of intact, PBS-treated or ROS Nano-treated rats. d, Representative curves of hip, knee and ankle angles during the one-step cycle. e, Representative EMG of the TA and GS muscles of rats with different treatments. Gray bars, stance; white bars, swing. Independent experiments were repeated 3 times with similar results.

Source data

Extended Data Fig. 5 Detailed statistical analysis of the hindlimb movement of rats with different treatments.

a, Quantification of the average maximal iliac crest height among groups. b, Quantification of the iliac crest height amplitude among rats with different treatments. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and comparisons within groups (#) for the data shown in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) <0.05. c, Quantification of the average maximal toe height among groups. d, Quantification of the average toe height amplitude among rats with different treatments. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and comparisons within groups (#) for the data in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) < 0.05. e, Quantification of the average hip oscillation among rats with different treatments. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and comparisons within groups (#) for the data in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) < 0.05. f, Quantification of the average knee angle oscillation among rats with different treatments. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and for comparisons within groups (#) for the data shown in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) < 0.05. g, Quantification of the average ankle angle oscillation among rats with different treatments. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and for comparisons within groups (#) for the data in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) < 0.05. h-i, Poincaré statistical analysis of the EMG signal amplitude rhythm of TA and GS muscles in the DOPA Nano and GABA Nano groups. One-way ANOVA with Tukey’s post hoc test was performed for comparisons among multiple groups (*) and for comparisons within groups (#) for the data shown in the violin plot. ***p (or ###p) < 0.001, **p (or ##p) < 0.01, *p (or #p) < 0.05. The complete stride cycle of each rat was recorded three times. n = 12 animals for intact group, n = 5 for PBS group, n = 6 for DOPA Nano group and ROS Nano group, n = 7 for GABA Nano group. The violin plot center indicates the median in all planes. Violin range covers 97.5th and 2.5th percentiles; extending whiskers show data distribution and probability density. Violin areas remain constant. Boxplot centerlines signify medians; boxes show first and third quartiles (Q1, Q3); whiskers extend from Q1 - 1.5xIQR to Q3 + 1.5xIQR; outliers lie outside whiskers. ANOVA for hip angle oscillation: Total: F = 19.098377, p = 5.200688*10−07,GABA-DOPA, p = 2.939199*10−04,PBS-DOPA, p = 1.637802*10−02, ROS-GABA, p = 7.304814*10−06, PBS-GABA, p = 6.557397*10−08 intact-GABA, p = 8.846887*10−04 ANOVA for knee angle oscillation Total: F = 8.73049, p = 0.000077, ROS-GABA, p = 0.034070, PBS-GABA p = 0.009822 intact-ROS p = 0.001111 intact-PBS p = 0.000313, ANOVA for ankle angle oscillation Total: F = 19.336293 p = 4.550813*10−08, GABA-DOPA p = 4.891140*10−03 ROS-DOPA P = 2.718774*10−02, PBS -DOPA p = 1.332916*10−03, ROS-GABA p = 1.476894*10−06, PBS -GABA p = 6.437209*10−08,intact-GABA p = 1.882720*10−05, intact- PBS P = 6.527743*10−03, ANOVA for Max Height of Crest Total: F = 38.22068, p = 1.439885 *10−11,intact- PBS p = 2.925660 *10−12, intact-ROS p = 1.030447 *10−07,intact-DOPA p = 1.880252 *10−06,intact-GABA p = 1.456296 *10−07, PBS-ROS p = 8.492501 *10−03, PBS-DOPA p = 9.491884 *10−03, PBS-GABA P = 2.243848 *10−03 ANOVA for maximal toe height Total: F = 3.735206 p = 0.013587. ANOVA for Toe Amplitude Total: F = 9.266889 p = 0.000048, intact- PBS p = 0.014311, intact-ROS p = 0.000182, intact-GABA p = 0.000631. ANOVA for crest high amplitude Total: F = 0.632153, p = 0.643303.

Source data

Extended Data Fig. 6 C-Fos expression in T8 and L2 spinal cord sections with different treatments.

a, Representative images of transverse sections of the T8 spinal cords of injured rats after 9 weeks of treatment with PBS, ROS Nano, DOPA Nano or GABA Nano, stained with c-Fos and NeuN. Scale bar, 500 μm. b, Representative images of transverse sections of the L2 spinal cords of injured rats after 9 weeks of treatment with PBS, ROS Nano, DOPA Nano or GABA Nano, stained with c-Fos and NeuN. Scale bar, 500 μm. T8 represents the 7th thoracic vertebral level, and L2 represents the second Lumbala’s vertebral level.

Supplementary information

Supplementary Information

Supplementary Figs. 1–11, Discussion and note.

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Supplementary Video 1

Vicon system observation of the movement of the rat hind-limb under different treatments.

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Zuo, Y., Ye, J., Cai, W. et al. Controlled delivery of a neurotransmitter–agonist conjugate for functional recovery after severe spinal cord injury. Nat. Nanotechnol. 18, 1230–1240 (2023). https://doi.org/10.1038/s41565-023-01416-0

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