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Lasers in Medical Science

, Volume 34, Issue 3, pp 561–569 | Cite as

Visualize and quantify the structural alteration of the rat spinal cord injury based on multiphoton microscopy

  • Chenxi Liao
  • Xiaoqin ZhuEmail author
  • Linquan Zhou
  • Zhenyu WangEmail author
  • Wenge Liu
  • Jianxin Chen
Original Article
  • 101 Downloads

Abstract

The development of imaging technique to visualize and quantify the structural alteration of the spinal cord injury (SCI) may lead to better understanding and treatments of the injuries. In this work, multiphoton microscopy (MPM) based on two-photon excited fluorescence (TPEF) and second-harmonic generation (SHG) was tentatively applied to quantitatively visualize the cellular microstructures of SCI to demonstrate the feasibility and superiority of MPM in SCI imaging. High-contrast MPM images of normal and injured spinal cord tissue were obtained for comparison. Moreover, the changes of injured spinal cord were characterized by the quantitative analysis of the MPM images. These results showed that MPM combined with quantitative method has the ability to identify the characteristics of spinal cord injury including the changes in the contents of nerve fibers and extracellular matrix. With the advancement of MPM, we believe that this technique has great potential to provide the histological diagnosis for the monitoring and evaluation of SCI.

Keywords

Multiphoton microscopy (MPM) Two-photon excited fluorescence (TPEF) Second-harmonic generation (SHG) Spinal cord injury (SCI) 

Notes

Funding information

The work was supported by the Changjiang Scholars Program and the University Innovative Research Team (grant no. IRT_15R10), the National Natural Science Foundation of China (grant nos. 81671730), the Natural Science Foundation of Fujian Province (2015J01241), and a grant from the Education Bureau of Fujian Province (grant no. JA13060).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

The experimental procedures with rats were conducted according to the Guide for Care and Use of Laboratory Animals and approved by the Animal Care and Use Committee in Fujian Medical University.

Informed consent

It does not apply, since the study was developed with Sprague-Dawley rats.

References

  1. 1.
    Galli R, Uckermann O, Winterhalder MJ, Sitoci-Ficici KH, Geiger KD, Koch E, Schackert G, Zumbusch A, Steiner G, Kirsch M (2012) Vibrational spectroscopic imaging and multiphoton microscopy of spinal cord injury. Anal Chem 84:8707–8714CrossRefGoogle Scholar
  2. 2.
    Galtrey CM, Kwok JC, Carulli D, Rhodes KE, Fawcett JW (2008) Distribution and synthesis of extracellular matrix proteoglycans, hyaluronan, link proteins and tenascin-R in the rat spinal cord. Eur J Neurosci 27:1373–1390CrossRefGoogle Scholar
  3. 3.
    Wiese S, Faissner A (2015) The role of extracellular matrix in spinal cord development. Exp Neurol 274:90–99CrossRefGoogle Scholar
  4. 4.
    Gaudet AD, Popovich PG (2014) Extracellular matrix regulation of inflammation in the healthy and injured spinal cord. Exp Neurol 258:24–34CrossRefGoogle Scholar
  5. 5.
    Ellingson BM, Salamon N, Holly LT (2014) Imaging techniques in spinal cord injury. World Neurosurg 82:1351–1358CrossRefGoogle Scholar
  6. 6.
    Powers BE, Sellers DL, Lovelett EA, Cheung W, Aalami SP, Zapertov N, Maris DO, Horner PJ (2013) Remyelination reporter reveals prolonged refinement of spontaneously regenerated myelin. Proc Natl Acad Sci U S A 110:4075–4080CrossRefGoogle Scholar
  7. 7.
    Petersen JA, Wilm BJ, von Meyenburg J, Schubert M, Seifert B, Najafi Y, Dietz V, Kollias S (2012) Chronic cervical spinal cord injury: DTI correlates with clinical and electrophysiological measures. J Neurotrauma 29:1556–1566CrossRefGoogle Scholar
  8. 8.
    Wang M, Dai Y, Han Y, Haacke EM, Dai J, Shi D (2011) Susceptibility weighted imaging in detecting hemorrhage in acute cervical spinal cord injury. Magn Reson Imaging 29:365–373CrossRefGoogle Scholar
  9. 9.
    Kang CE, Clarkson R, Tator CH, Yeung IW, Shoichet MS (2010) Spinal cord blood flow and blood vessel permeability measured by dynamic computed tomography imaging in rats after localized delivery of fibroblast growth factor. J Neurotrauma 27:2041–2053CrossRefGoogle Scholar
  10. 10.
    Zoumi A, Yeh A, Tromberg BJ (2002) Imaging cells and extracellular matrix in vivo by using second-harmonic generation and two photon excited fluorescence. Proc Natl Acad Sci U S A 99:11014e9CrossRefGoogle Scholar
  11. 11.
    Uckermann O, Galli R, Beiermeister R, Sitoci-Ficici KH, Later R, Leipnitz E, Neuwirth A, Chavakis T, Koch E, Schackert G, Steiner G, Kirsch M (2015) Endogenous two-photon excited fluorescence provides label-free visualization of the inflammatory response in the rodent spinal cord. Biomed Res Int 2015:859084CrossRefGoogle Scholar
  12. 12.
    Israel JS, Esquibel CR, Dingle AM, Liu Y, Keikhosravi A, Pisaniello JA, Hesse MA, Brodnick SK, Novello J, Krugner-Higby L, Williams JC, Eliceiri KW, Poore SO (2017) Quantification of collagen organization after nerve repair. Plast Reconstr Surg Glob Open 5:e1586CrossRefGoogle Scholar
  13. 13.
    Noble LJ, Wrathall JR (1985) Spinal cord contusion in the rat: morphometric analyses of alterations in the spinal cord. Exp Neurol 88:135–149CrossRefGoogle Scholar
  14. 14.
    Zhu X, Tang Y, Chen J, Xiong S, Zhuo S (2013) Monitoring wound healing of elastic cartilage using multiphoton microscopy. Osteoarthr Cartilage 21:1799–1806CrossRefGoogle Scholar
  15. 15.
    Quinn KP, Georgakoudi I (2013) Rapid quantification of pixel-wise fiber orientation data in micrographs. J Biomed Opt 18:046003–046003CrossRefGoogle Scholar
  16. 16.
    Shapiro SS, Wilk MB (1965) An analysis of variance test for normality (complete samples). Biometrika 52:591–611CrossRefGoogle Scholar
  17. 17.
    Nam MH, Baek M, Lim J, Lee S, Yoon J, Kim J, Soh KS (2014) Discovery of a novel fibrous tissue in the spinal pia mater by polarized light microscopy. Connect Tissue Res 55:147–155CrossRefGoogle Scholar
  18. 18.
    Bignami A, Asher R, Perides G (1992) The extracellular matrix of rat spinal cord: a comparative study on the localization of hyaluronic acid, glial hyaluronate-binding protein, and chondroitin sulfate proteoglycan. Exp Neurol 117:90–93CrossRefGoogle Scholar
  19. 19.
    Li D, Zheng W, Zeng Y, Luo Y, Qu JY (2011) Two-photon excited hemoglobin fluorescence provides contrast mechanism for label-free imaging of microvasculature in vivo. Opt Lett 36:834–836CrossRefGoogle Scholar
  20. 20.
    Saytashev I, Glenn R, Murashova GA, Osseiran S, Spence D, Evans CL, Dantus M (2016) Multiphoton excited hemoglobin fluorescence and third harmonic generation for non-invasive microscopy of stored blood. Biomed Opt Express 7:3449–3460CrossRefGoogle Scholar
  21. 21.
    Liao CX, Wang ZY, Zhou Y, Zhou LQ, Zhu XQ, Liu WG, Chen JX (2017) Label-free identification of the microstructure of rat spinal cords based on nonlinear optical microscopy. J Microsc 267:143–149CrossRefGoogle Scholar
  22. 22.
    Waxman SG, Kocsis JD, Stys PK (1995) The axon: structure, function, and pathophysiology. Oxford University Press, OxfordCrossRefGoogle Scholar
  23. 23.
    Wang F, Bélanger E, Paquet ME, Côté DC, De Koninck Y (2016) Probing pain pathways with light. Neuroscience 338:248–271CrossRefGoogle Scholar
  24. 24.
    Johansen-Berg H, Behrens TE (2013) Diffusion MRI: from quantitative measurement to in vivo neuroanatomy. Academic press, San DiegoGoogle Scholar
  25. 25.
    Cheran S, Shanmuganathan K, Zhuo J, Mirvis SE, Aarabi B, Alexander MT, Gullapalli RP (2011) Correlation of MR diffusion tensor imaging parameters with ASIA motor scores in hemorrhagic and nonhemorrhagic acute spinal cord injury. J Neurotrauma 28:1881–1892CrossRefGoogle Scholar
  26. 26.
    Quencer RM, Bunge RP, Egnor M, Green BA, Puckett W, Naidich TP, Post MJD, Norenberg M (1992) Acute traumatic central cord syndrome: MRI-pathological correlations. Neuroradiology 34:85–94CrossRefGoogle Scholar
  27. 27.
    Douek P, Turner R, Pekar J, Patronas N, Le Bihan D (1991) MR color mapping of myelin fiber orientation. J Comput Assist Tomogr 15:923–929CrossRefGoogle Scholar
  28. 28.
    Li LM, Li JB, Zhu Y, Fan GY (2010) Soluble complement receptor type 1 inhibits complement system activation and improves motor function in acute spinal cord injury. Spinal Cord 48:105–111CrossRefGoogle Scholar
  29. 29.
    Dray C, Rougon G, Debarbieux F (2009) Quantitative analysis by in vivo imaging of the dynamics of vascular and axonal networks in injured mouse spinal cord. Proc Natl Acad Sci U S A 106:9459–9464CrossRefGoogle Scholar
  30. 30.
    Helmchen F, Denk W, Kerr JN (2013) Miniaturization of two-photon microscopy for imaging in freely moving animals. Cold Spring Harb Protoc 2013:904–913Google Scholar

Copyright information

© Springer-Verlag London Ltd., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, Fujian Provincial Key Laboratory of Photonics TechnologyFujian Normal UniversityFuzhouChina
  2. 2.Department of OrthopedicsAffiliated Union Hospital of Fujian Medical UniversityFuzhouPeople’s Republic of China

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