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Influence of extracellular matrix components on the expression of integrins and regeneration of adult retinal ganglion cells

Vecino, Elena; Heller, Janosch P; Veiga-Crespo, Patricia LU ; Martin, Keith R and Fawcett, James W (2015) In PLoS ONE 10(5). p.0125250-0125250
Abstract

PURPOSE: Retinal ganglion cells (RGCs) are exposed to injury in a variety of optic nerve diseases including glaucoma. However, not all cells respond in the same way to damage and the capacity of individual RGCs to survive or regenerate is variable. In order to elucidate factors that may be important for RGC survival and regeneration we have focussed on the extracellular matrix (ECM) and RGC integrin expression. Our specific questions were: (1) Do adult RGCs express particular sets of integrins in vitro and in vivo? (2) Can the nature of the ECM influence the expression of different integrins? (3) Can the nature of the ECM affect the survival of the cells and the length or branching complexity of their neurites?

METHODS: Primary... (More)

PURPOSE: Retinal ganglion cells (RGCs) are exposed to injury in a variety of optic nerve diseases including glaucoma. However, not all cells respond in the same way to damage and the capacity of individual RGCs to survive or regenerate is variable. In order to elucidate factors that may be important for RGC survival and regeneration we have focussed on the extracellular matrix (ECM) and RGC integrin expression. Our specific questions were: (1) Do adult RGCs express particular sets of integrins in vitro and in vivo? (2) Can the nature of the ECM influence the expression of different integrins? (3) Can the nature of the ECM affect the survival of the cells and the length or branching complexity of their neurites?

METHODS: Primary RGC cultures from adult rat retina were placed on glass coverslips treated with different substrates: Poly-L-Lysine (PL), or PL plus laminin (L), collagen I (CI), collagen IV (CIV) or fibronectin (F). After 10 days in culture, we performed double immunostaining with an antibody against βIII-Tubulin to identify the RGCs, and antibodies against the integrin subunits: αV, α1, α3, α5, β1 or β3. The number of adhering and surviving cells, the number and length of the neurites and the expression of the integrin subunits on the different substrates were analysed.

RESULTS: PL and L were associated with the greatest survival of RGCs while CI provided the least favourable conditions. The type of substrate affected the number and length of neurites. L stimulated the longest growth. We found at least three different types of RGCs in terms of their capacity to regenerate and extend neurites. The different combinations of integrins expressed by the cells growing on different substrata suggest that RGCs expressed predominantly α1β1 or α3β1 on L, α1β1 on CI and CIV, and α5β3 on F. The activity of the integrins was demonstrated by the phosphorylation of focal adhesion kinase (FAK).

CONCLUSIONS: Adult rat RGCs can survive and grow in the presence of different ECM tested. Further studies should be done to elucidate the different molecular characteristics of the RGCs subtypes in order to understand the possible different sensitivity of different RGCs to damage in diseases like glaucoma in which not all RGCs die at the same time.

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publication status
published
keywords
Animals, Antigens, CD29, Cell Survival, Extracellular Matrix, Female, Immunohistochemistry, Integrin alpha1, Integrin alpha5, Integrin beta3, Integrins, Rats, Rats, Sprague-Dawley, Retinal Ganglion Cells, Journal Article, Research Support, Non-U.S. Gov't
in
PLoS ONE
volume
10
issue
5
pages
0125250 - 0125250
publisher
Public Library of Science
external identifiers
  • scopus:84957454361
ISSN
1932-6203
DOI
10.1371/journal.pone.0125250
language
English
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no
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d081bbf6-2ecb-4081-8e85-03c1a8825652
date added to LUP
2016-12-13 14:50:01
date last changed
2017-05-28 04:55:39
@article{d081bbf6-2ecb-4081-8e85-03c1a8825652,
  abstract     = {<p>PURPOSE: Retinal ganglion cells (RGCs) are exposed to injury in a variety of optic nerve diseases including glaucoma. However, not all cells respond in the same way to damage and the capacity of individual RGCs to survive or regenerate is variable. In order to elucidate factors that may be important for RGC survival and regeneration we have focussed on the extracellular matrix (ECM) and RGC integrin expression. Our specific questions were: (1) Do adult RGCs express particular sets of integrins in vitro and in vivo? (2) Can the nature of the ECM influence the expression of different integrins? (3) Can the nature of the ECM affect the survival of the cells and the length or branching complexity of their neurites?</p><p>METHODS: Primary RGC cultures from adult rat retina were placed on glass coverslips treated with different substrates: Poly-L-Lysine (PL), or PL plus laminin (L), collagen I (CI), collagen IV (CIV) or fibronectin (F). After 10 days in culture, we performed double immunostaining with an antibody against βIII-Tubulin to identify the RGCs, and antibodies against the integrin subunits: αV, α1, α3, α5, β1 or β3. The number of adhering and surviving cells, the number and length of the neurites and the expression of the integrin subunits on the different substrates were analysed.</p><p>RESULTS: PL and L were associated with the greatest survival of RGCs while CI provided the least favourable conditions. The type of substrate affected the number and length of neurites. L stimulated the longest growth. We found at least three different types of RGCs in terms of their capacity to regenerate and extend neurites. The different combinations of integrins expressed by the cells growing on different substrata suggest that RGCs expressed predominantly α1β1 or α3β1 on L, α1β1 on CI and CIV, and α5β3 on F. The activity of the integrins was demonstrated by the phosphorylation of focal adhesion kinase (FAK).</p><p>CONCLUSIONS: Adult rat RGCs can survive and grow in the presence of different ECM tested. Further studies should be done to elucidate the different molecular characteristics of the RGCs subtypes in order to understand the possible different sensitivity of different RGCs to damage in diseases like glaucoma in which not all RGCs die at the same time.</p>},
  author       = {Vecino, Elena and Heller, Janosch P and Veiga-Crespo, Patricia and Martin, Keith R and Fawcett, James W},
  issn         = {1932-6203},
  keyword      = {Animals,Antigens, CD29,Cell Survival,Extracellular Matrix,Female,Immunohistochemistry,Integrin alpha1,Integrin alpha5,Integrin beta3,Integrins,Rats,Rats, Sprague-Dawley,Retinal Ganglion Cells,Journal Article,Research Support, Non-U.S. Gov't},
  language     = {eng},
  number       = {5},
  pages        = {0125250--0125250},
  publisher    = {Public Library of Science},
  series       = {PLoS ONE},
  title        = {Influence of extracellular matrix components on the expression of integrins and regeneration of adult retinal ganglion cells},
  url          = {http://dx.doi.org/10.1371/journal.pone.0125250},
  volume       = {10},
  year         = {2015},
}