Limits...
P2X7 purinoceptors contribute to the death of Schwann cells transplanted into the spinal cord.

Luo J, Lee S, Wu D, Yeh J, Ellamushi H, Wheeler AP, Warnes G, Zhang Y, Bo X - Cell Death Dis (2013)

Bottom Line: Furthermore, ATP did not cause death of SCs isolated from P2X7R-knockout mice.All these results suggest that P2X7R is responsible for ATP-induced SC death in vitro.These results indicate that targeting P2X7R on SCs could be a potential strategy to improve the survival of transplanted cells.

View Article: PubMed Central - PubMed

Affiliation: 1] Centre for Neuroscience and Trauma, Blizard Institute, Queen Mary University of London, London E1 2AT, UK [2] Department of Physiology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.

ABSTRACT
The potential to use Schwann cells (SCs) in neural repair for patients suffering from neurotrauma and neurodegenerative diseases is well recognized. However, significant cell death after transplantation hinders the clinical translation of SC-based therapies. Various factors may contribute to the death of transplanted cells. It is known that prolonged activation of P2X7 purinoceptors (P2X7R) can lead to death of certain types of cells. In this study, we show that rat SCs express P2X7R and exposure of cultured SCs to high concentrations of ATP (3-5 mM) or a P2X7R agonist, 2'(3')-O-(4-benzoylbenzoyl)ATP (BzATP) induced significant cell death rapidly. High concentrations of ATP and BzATP increased ethidium uptake by SCs, indicating increased membrane permeability to large molecules, a typical feature of prolonged P2X7R activation. SC death, as well as ethidium uptake, induced by ATP was blocked by an irreversible P2X7R antagonist oxidized ATP (oxATP) or a reversible P2X7R antagonist A438079. oxATP also significantly inhibits the increase of intracellular free calcium induced by minimolar ATP concentrations. Furthermore, ATP did not cause death of SCs isolated from P2X7R-knockout mice. All these results suggest that P2X7R is responsible for ATP-induced SC death in vitro. When rat SCs were treated with oxATP before transplantation into uninjured rat spinal cord, 35% more SCs survived than untreated SCs 1 week after transplantation. Moreover, 58% more SCs isolated from P2X7R-knockout mice survived after being transplanted into rat spinal cord than SCs from wild-type mice. This further confirms that P2X7R is involved in the death of transplanted SCs. These results indicate that targeting P2X7R on SCs could be a potential strategy to improve the survival of transplanted cells. As many other types of cells, including neural stem cells, also express P2X7R, deactivating P2X7R may improve the survival of other types of transplanted cells.

Show MeSH

Related in: MedlinePlus

P2X7R-deficient SCs are resistant to ATP-induced cell death and survive better after transplantation. (a) Flow cytometry apoptosis assay showing that 5 mM ATP induced significant death of SCs from wild-type (WT) mice, whereas SCs from P2X7R-knockout (KO) mice did not show obvious cell death. ***P<0.001, Student's t-test, n=4. (b) Photomicrograph showing the surviving GFP-expressing mouse SCs from WT or P2X7R KO mouse 1 week after transplantation into rat spinal cords. (c) Quantification of the areas occupied by GFP/SCs from WT or P2X7R KO mice transplanted into the spinal cords of five rats (data from the same animal are linked by colored lines)
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3824653&req=5

fig7: P2X7R-deficient SCs are resistant to ATP-induced cell death and survive better after transplantation. (a) Flow cytometry apoptosis assay showing that 5 mM ATP induced significant death of SCs from wild-type (WT) mice, whereas SCs from P2X7R-knockout (KO) mice did not show obvious cell death. ***P<0.001, Student's t-test, n=4. (b) Photomicrograph showing the surviving GFP-expressing mouse SCs from WT or P2X7R KO mouse 1 week after transplantation into rat spinal cords. (c) Quantification of the areas occupied by GFP/SCs from WT or P2X7R KO mice transplanted into the spinal cords of five rats (data from the same animal are linked by colored lines)

Mentions: To test whether SCs deficient of P2X7R can survive better after transplantation, we isolated SCs from C57Bl/6J wild-type and P2X7R-knockout mice, and then transduced them with GFP-expressing adenovirus, as mouse SCs are not susceptible to lentiviral transduction. The same numbers of cells (100 000) from wild-type or P2X7R-knockout mice were transplanted into either side of dorsal columns at the thoracic eight level of the spinal cord of adult rats (n=5). Animals were injected with ciclosporin daily after surgery to suppress immune rejections. One week later, animals were killed and the areas occupied by GFP+ SCs in the spinal cord sections (Figure 7b) were measured using ImageJ. It was found that 54.8±8.8% more SCs from P2X7R-knockout mice survived compared with those from wild-type mice (Figure 7c, P<0.01, paired Student's t-test), which indicates that P2X7R knockout can promote the survival of transplanted SCs.


P2X7 purinoceptors contribute to the death of Schwann cells transplanted into the spinal cord.

Luo J, Lee S, Wu D, Yeh J, Ellamushi H, Wheeler AP, Warnes G, Zhang Y, Bo X - Cell Death Dis (2013)

P2X7R-deficient SCs are resistant to ATP-induced cell death and survive better after transplantation. (a) Flow cytometry apoptosis assay showing that 5 mM ATP induced significant death of SCs from wild-type (WT) mice, whereas SCs from P2X7R-knockout (KO) mice did not show obvious cell death. ***P<0.001, Student's t-test, n=4. (b) Photomicrograph showing the surviving GFP-expressing mouse SCs from WT or P2X7R KO mouse 1 week after transplantation into rat spinal cords. (c) Quantification of the areas occupied by GFP/SCs from WT or P2X7R KO mice transplanted into the spinal cords of five rats (data from the same animal are linked by colored lines)
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824653&req=5

fig7: P2X7R-deficient SCs are resistant to ATP-induced cell death and survive better after transplantation. (a) Flow cytometry apoptosis assay showing that 5 mM ATP induced significant death of SCs from wild-type (WT) mice, whereas SCs from P2X7R-knockout (KO) mice did not show obvious cell death. ***P<0.001, Student's t-test, n=4. (b) Photomicrograph showing the surviving GFP-expressing mouse SCs from WT or P2X7R KO mouse 1 week after transplantation into rat spinal cords. (c) Quantification of the areas occupied by GFP/SCs from WT or P2X7R KO mice transplanted into the spinal cords of five rats (data from the same animal are linked by colored lines)
Mentions: To test whether SCs deficient of P2X7R can survive better after transplantation, we isolated SCs from C57Bl/6J wild-type and P2X7R-knockout mice, and then transduced them with GFP-expressing adenovirus, as mouse SCs are not susceptible to lentiviral transduction. The same numbers of cells (100 000) from wild-type or P2X7R-knockout mice were transplanted into either side of dorsal columns at the thoracic eight level of the spinal cord of adult rats (n=5). Animals were injected with ciclosporin daily after surgery to suppress immune rejections. One week later, animals were killed and the areas occupied by GFP+ SCs in the spinal cord sections (Figure 7b) were measured using ImageJ. It was found that 54.8±8.8% more SCs from P2X7R-knockout mice survived compared with those from wild-type mice (Figure 7c, P<0.01, paired Student's t-test), which indicates that P2X7R knockout can promote the survival of transplanted SCs.

Bottom Line: Furthermore, ATP did not cause death of SCs isolated from P2X7R-knockout mice.All these results suggest that P2X7R is responsible for ATP-induced SC death in vitro.These results indicate that targeting P2X7R on SCs could be a potential strategy to improve the survival of transplanted cells.

View Article: PubMed Central - PubMed

Affiliation: 1] Centre for Neuroscience and Trauma, Blizard Institute, Queen Mary University of London, London E1 2AT, UK [2] Department of Physiology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.

ABSTRACT
The potential to use Schwann cells (SCs) in neural repair for patients suffering from neurotrauma and neurodegenerative diseases is well recognized. However, significant cell death after transplantation hinders the clinical translation of SC-based therapies. Various factors may contribute to the death of transplanted cells. It is known that prolonged activation of P2X7 purinoceptors (P2X7R) can lead to death of certain types of cells. In this study, we show that rat SCs express P2X7R and exposure of cultured SCs to high concentrations of ATP (3-5 mM) or a P2X7R agonist, 2'(3')-O-(4-benzoylbenzoyl)ATP (BzATP) induced significant cell death rapidly. High concentrations of ATP and BzATP increased ethidium uptake by SCs, indicating increased membrane permeability to large molecules, a typical feature of prolonged P2X7R activation. SC death, as well as ethidium uptake, induced by ATP was blocked by an irreversible P2X7R antagonist oxidized ATP (oxATP) or a reversible P2X7R antagonist A438079. oxATP also significantly inhibits the increase of intracellular free calcium induced by minimolar ATP concentrations. Furthermore, ATP did not cause death of SCs isolated from P2X7R-knockout mice. All these results suggest that P2X7R is responsible for ATP-induced SC death in vitro. When rat SCs were treated with oxATP before transplantation into uninjured rat spinal cord, 35% more SCs survived than untreated SCs 1 week after transplantation. Moreover, 58% more SCs isolated from P2X7R-knockout mice survived after being transplanted into rat spinal cord than SCs from wild-type mice. This further confirms that P2X7R is involved in the death of transplanted SCs. These results indicate that targeting P2X7R on SCs could be a potential strategy to improve the survival of transplanted cells. As many other types of cells, including neural stem cells, also express P2X7R, deactivating P2X7R may improve the survival of other types of transplanted cells.

Show MeSH
Related in: MedlinePlus