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Schwann cell autophagy, myelinophagy, initiates myelin clearance from injured nerves.

Gomez-Sanchez JA, Carty L, Iruarrizaga-Lejarreta M, Palomo-Irigoyen M, Varela-Rey M, Griffith M, Hantke J, Macias-Camara N, Azkargorta M, Aurrekoetxea I, De Juan VG, Jefferies HB, Aspichueta P, Elortza F, Aransay AM, Martínez-Chantar ML, Baas F, Mato JM, Mirsky R, Woodhoo A, Jessen KR - J. Cell Biol. (2015)

Bottom Line: Myelinophagy was positively regulated by the Schwann cell JNK/c-Jun pathway, a central regulator of the Schwann cell reprogramming induced by nerve injury.We also present evidence that myelinophagy is defective in the injured central nervous system.These results reveal an important role for inductive autophagy during Wallerian degeneration, and point to potential mechanistic targets for accelerating myelin clearance and improving demyelinating disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell and Developmental Biology, University College London, London WC1E 6BT, England, UK.

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Genetic inactivation of autophagy retards myelin degradation in vivo. (A) Western blot showing higher levels of the myelin proteins MPZ, MBP, and Periaxin in 5 d cut nerves from Atg7 cKO mice compared with WT controls. (B) Densitometric analysis of Western blots showing higher levels of myelin proteins in 5 and 7 d cut nerves from Atg7 cKO mice compared with WT controls. For each comparison, the value for cKO is normalized to that seen in WT. n = a minimum of three mice for the genotype/time point. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT). (C) Immunolabeling showing MPZ+ myelin inclusions in cultured WT Schwann cells and cells from Atg7 cKO nerves that often show bloated “cauliflower” morphology. The graph shows quantification of MPZ+ area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 60 cells analyzed per condition. **, P < 0.01 (Atg7 cKO relative to WT). (D) Electron micrographs showing several intact myelin sheath profiles in 5 d cut nerves from Atg7 cKO mice. The graph shows a quantification of the number of intact myelin sheaths. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 18 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (E) Teased fibers of 5 d cut nerves from WT and Atg7 cKO mice stained with FluoroMyelin red to show myelin. The graph shows a quantification of the myelin fluorescent area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 10 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (F) The lipid composition of whole sciatic nerves from 5 d cut WT and Atg7 cKO mice expressed as log2 fold change compared with uncut nerves. n = a minimum of four mice for each genotype. Data are presented as mean ± SEM (error bars). **, P < 0.01. (G) The lipid composition of purified myelin obtained from sciatic nerves from 5 d cut WT and Atg7 cKO mice, expressed as log2 fold change compared with uncut nerves. The individual lipid species detected by UPLC were grouped in distinct lipid classes as shown in the graph. “Membrane lipids” refers to all lipid species detected that are the major structural lipids in the eukaryotic membrane, including phosphatidylethanolamines, phosphatidylcholines, and phosphatidylinositols, and “storage lipids” include triacylglycerides and cholesteryl esters. See Table S2 for changes in levels of individual lipid species in WT and Atg7 cKO mice. n = 3 mice for each genotype. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT).
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fig4: Genetic inactivation of autophagy retards myelin degradation in vivo. (A) Western blot showing higher levels of the myelin proteins MPZ, MBP, and Periaxin in 5 d cut nerves from Atg7 cKO mice compared with WT controls. (B) Densitometric analysis of Western blots showing higher levels of myelin proteins in 5 and 7 d cut nerves from Atg7 cKO mice compared with WT controls. For each comparison, the value for cKO is normalized to that seen in WT. n = a minimum of three mice for the genotype/time point. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT). (C) Immunolabeling showing MPZ+ myelin inclusions in cultured WT Schwann cells and cells from Atg7 cKO nerves that often show bloated “cauliflower” morphology. The graph shows quantification of MPZ+ area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 60 cells analyzed per condition. **, P < 0.01 (Atg7 cKO relative to WT). (D) Electron micrographs showing several intact myelin sheath profiles in 5 d cut nerves from Atg7 cKO mice. The graph shows a quantification of the number of intact myelin sheaths. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 18 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (E) Teased fibers of 5 d cut nerves from WT and Atg7 cKO mice stained with FluoroMyelin red to show myelin. The graph shows a quantification of the myelin fluorescent area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 10 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (F) The lipid composition of whole sciatic nerves from 5 d cut WT and Atg7 cKO mice expressed as log2 fold change compared with uncut nerves. n = a minimum of four mice for each genotype. Data are presented as mean ± SEM (error bars). **, P < 0.01. (G) The lipid composition of purified myelin obtained from sciatic nerves from 5 d cut WT and Atg7 cKO mice, expressed as log2 fold change compared with uncut nerves. The individual lipid species detected by UPLC were grouped in distinct lipid classes as shown in the graph. “Membrane lipids” refers to all lipid species detected that are the major structural lipids in the eukaryotic membrane, including phosphatidylethanolamines, phosphatidylcholines, and phosphatidylinositols, and “storage lipids” include triacylglycerides and cholesteryl esters. See Table S2 for changes in levels of individual lipid species in WT and Atg7 cKO mice. n = 3 mice for each genotype. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT).

Mentions: Striking differences were found when myelin breakdown was compared in Atg7 cKO and WT nerves after injury. A significant reduction was seen in injury-induced breakdown of myelin proteins in transected nerves in vivo (Fig. 4, A and B), in cultured nerve segments (Fig. S4 D), and in Schwann cell cultures where Atg7 cKO cells typically retained bloated “cauliflower” morphology for many days due to the persistence of undigested myelin (Fig. 4 C). In transected Atg7 cKO nerves, there was also a striking preservation of noncollapsed myelin (Fig. 4 D), confirming previous results using the autophagy blocker 3-MA (Fig. 3 C).


Schwann cell autophagy, myelinophagy, initiates myelin clearance from injured nerves.

Gomez-Sanchez JA, Carty L, Iruarrizaga-Lejarreta M, Palomo-Irigoyen M, Varela-Rey M, Griffith M, Hantke J, Macias-Camara N, Azkargorta M, Aurrekoetxea I, De Juan VG, Jefferies HB, Aspichueta P, Elortza F, Aransay AM, Martínez-Chantar ML, Baas F, Mato JM, Mirsky R, Woodhoo A, Jessen KR - J. Cell Biol. (2015)

Genetic inactivation of autophagy retards myelin degradation in vivo. (A) Western blot showing higher levels of the myelin proteins MPZ, MBP, and Periaxin in 5 d cut nerves from Atg7 cKO mice compared with WT controls. (B) Densitometric analysis of Western blots showing higher levels of myelin proteins in 5 and 7 d cut nerves from Atg7 cKO mice compared with WT controls. For each comparison, the value for cKO is normalized to that seen in WT. n = a minimum of three mice for the genotype/time point. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT). (C) Immunolabeling showing MPZ+ myelin inclusions in cultured WT Schwann cells and cells from Atg7 cKO nerves that often show bloated “cauliflower” morphology. The graph shows quantification of MPZ+ area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 60 cells analyzed per condition. **, P < 0.01 (Atg7 cKO relative to WT). (D) Electron micrographs showing several intact myelin sheath profiles in 5 d cut nerves from Atg7 cKO mice. The graph shows a quantification of the number of intact myelin sheaths. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 18 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (E) Teased fibers of 5 d cut nerves from WT and Atg7 cKO mice stained with FluoroMyelin red to show myelin. The graph shows a quantification of the myelin fluorescent area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 10 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (F) The lipid composition of whole sciatic nerves from 5 d cut WT and Atg7 cKO mice expressed as log2 fold change compared with uncut nerves. n = a minimum of four mice for each genotype. Data are presented as mean ± SEM (error bars). **, P < 0.01. (G) The lipid composition of purified myelin obtained from sciatic nerves from 5 d cut WT and Atg7 cKO mice, expressed as log2 fold change compared with uncut nerves. The individual lipid species detected by UPLC were grouped in distinct lipid classes as shown in the graph. “Membrane lipids” refers to all lipid species detected that are the major structural lipids in the eukaryotic membrane, including phosphatidylethanolamines, phosphatidylcholines, and phosphatidylinositols, and “storage lipids” include triacylglycerides and cholesteryl esters. See Table S2 for changes in levels of individual lipid species in WT and Atg7 cKO mice. n = 3 mice for each genotype. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT).
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fig4: Genetic inactivation of autophagy retards myelin degradation in vivo. (A) Western blot showing higher levels of the myelin proteins MPZ, MBP, and Periaxin in 5 d cut nerves from Atg7 cKO mice compared with WT controls. (B) Densitometric analysis of Western blots showing higher levels of myelin proteins in 5 and 7 d cut nerves from Atg7 cKO mice compared with WT controls. For each comparison, the value for cKO is normalized to that seen in WT. n = a minimum of three mice for the genotype/time point. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT). (C) Immunolabeling showing MPZ+ myelin inclusions in cultured WT Schwann cells and cells from Atg7 cKO nerves that often show bloated “cauliflower” morphology. The graph shows quantification of MPZ+ area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 60 cells analyzed per condition. **, P < 0.01 (Atg7 cKO relative to WT). (D) Electron micrographs showing several intact myelin sheath profiles in 5 d cut nerves from Atg7 cKO mice. The graph shows a quantification of the number of intact myelin sheaths. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 18 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (E) Teased fibers of 5 d cut nerves from WT and Atg7 cKO mice stained with FluoroMyelin red to show myelin. The graph shows a quantification of the myelin fluorescent area. Data are presented as mean ± SEM (error bars) from three independent experiments with a minimum of 10 picture frames analyzed per condition/experiment. **, P < 0.01 (Atg7 cKO relative to WT). (F) The lipid composition of whole sciatic nerves from 5 d cut WT and Atg7 cKO mice expressed as log2 fold change compared with uncut nerves. n = a minimum of four mice for each genotype. Data are presented as mean ± SEM (error bars). **, P < 0.01. (G) The lipid composition of purified myelin obtained from sciatic nerves from 5 d cut WT and Atg7 cKO mice, expressed as log2 fold change compared with uncut nerves. The individual lipid species detected by UPLC were grouped in distinct lipid classes as shown in the graph. “Membrane lipids” refers to all lipid species detected that are the major structural lipids in the eukaryotic membrane, including phosphatidylethanolamines, phosphatidylcholines, and phosphatidylinositols, and “storage lipids” include triacylglycerides and cholesteryl esters. See Table S2 for changes in levels of individual lipid species in WT and Atg7 cKO mice. n = 3 mice for each genotype. Data are presented as mean ± SEM (error bars). *, P < 0.05; **, P < 0.01 (Atg7 cKO relative to WT).
Mentions: Striking differences were found when myelin breakdown was compared in Atg7 cKO and WT nerves after injury. A significant reduction was seen in injury-induced breakdown of myelin proteins in transected nerves in vivo (Fig. 4, A and B), in cultured nerve segments (Fig. S4 D), and in Schwann cell cultures where Atg7 cKO cells typically retained bloated “cauliflower” morphology for many days due to the persistence of undigested myelin (Fig. 4 C). In transected Atg7 cKO nerves, there was also a striking preservation of noncollapsed myelin (Fig. 4 D), confirming previous results using the autophagy blocker 3-MA (Fig. 3 C).

Bottom Line: Myelinophagy was positively regulated by the Schwann cell JNK/c-Jun pathway, a central regulator of the Schwann cell reprogramming induced by nerve injury.We also present evidence that myelinophagy is defective in the injured central nervous system.These results reveal an important role for inductive autophagy during Wallerian degeneration, and point to potential mechanistic targets for accelerating myelin clearance and improving demyelinating disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell and Developmental Biology, University College London, London WC1E 6BT, England, UK.

Show MeSH
Related in: MedlinePlus