CERTL reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer’s disease

Simone M. Crivelli; Qian Luo; Jo A.A. Stevens; Caterina Giovagnoni; Daan van Kruining; Gerard Bode; Sandra den Hoedt; Barbara Hobo; Anna Lena Scheithauer; Jochen Walter; Monique T. Mulder; Christopher Exley; Matthew Mold; Michelle M. Mielke; Helga E. De Vries; Kristiaan Wouters; Daniel L.A. van den Hove; Dusan Berkes; María Dolores Ledesma; Joost Verhaagen; Mario Losen; Erhard Bieberich; Pilar Martinez-Martinez

doi:10.1186/s13195-021-00780-0

CERT_L reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer’s disease

Simone M. Crivelli, Qian Luo, Jo A.A. Stevens, Caterina Giovagnoni, Daan van Kruining, Gerard Bode, Sandra den Hoedt, Barbara Hobo, Anna Lena Scheithauer, Jochen Walter, Monique T. Mulder, Christopher Exley, Matthew Mold, Michelle M. Mielke, Helga E. De Vries, Kristiaan Wouters, Daniel L.A. van den Hove, Dusan Berkes, María Dolores Ledesma, Joost VerhaagenMario Losen, Erhard Bieberich, Pilar Martinez-Martinez^*

^*Corresponding author for this work

Research output: Contribution to Journal › Article › Academic › peer-review

Abstract

Background: Dysregulation of ceramide and sphingomyelin levels have been suggested to contribute to the pathogenesis of Alzheimer’s disease (AD). Ceramide transfer proteins (CERTs) are ceramide carriers which are crucial for ceramide and sphingomyelin balance in cells. Extracellular forms of CERTs co-localize with amyloid-β (Aβ) plaques in AD brains. To date, the significance of these observations for the pathophysiology of AD remains uncertain. Methods: A plasmid expressing CERT_L, the long isoform of CERTs, was used to study the interaction of CERT_L with amyloid precursor protein (APP) by co-immunoprecipitation and immunofluorescence in HEK cells. The recombinant CERT_L protein was employed to study interaction of CERT_L with amyloid-β (Aβ), Aβ aggregation process in presence of CERT_L, and the resulting changes in Aβ toxicity in neuroblastoma cells. CERT_L was overexpressed in neurons by adeno-associated virus (AAV) in a mouse model of familial AD (5xFAD). Ten weeks after transduction, animals were challenged with behavior tests for memory, anxiety, and locomotion. At week 12, brains were investigated for sphingolipid levels by mass spectrometry, plaques, and neuroinflammation by immunohistochemistry, gene expression, and/or immunoassay. Results: Here, we report that CERT_L binds to APP, modifies Aβ aggregation, and reduces Aβ neurotoxicity in vitro. Furthermore, we show that intracortical injection of AAV, mediating the expression of CERT_L, decreases levels of ceramide d18:1/16:0 and increases sphingomyelin levels in the brain of male 5xFAD mice. CERT_L in vivo over-expression has a mild effect on animal locomotion, decreases Aβ formation, and modulates microglia by decreasing their pro-inflammatory phenotype. Conclusion: Our results demonstrate a crucial role of CERT_L in regulating ceramide levels in the brain, in amyloid plaque formation and neuroinflammation, thereby opening research avenues for therapeutic targets of AD and other neurodegenerative diseases.

Original language	English
Article number	45
Journal	Alzheimer's Research and Therapy
Volume	13
Issue number	1
DOIs	https://doi.org/10.1186/s13195-021-00780-0
Publication status	Published - Dec 2021

Bibliographical note

Funding Information:
This work was supported by grants to NMdW, SdH, MTM, JW, AR, PMM, JV, and HEV from ZonMw Memorabel program (projectnr: 733050105). PMM is also supported by the International Foundation for Alzheimer Research (ISAO) (projectnr: 14545). SMC received a travel grant support from Alzheimer Nederlands foundation (AN) to spend a month in the laboratory of EB, University of Kentucky, Lexington KY, USA. Aspects of this work were supported by the grants NIH R01AG034389, R01NS095215, and R56AG064234; VA I01BX003643 to EB. MMM was supported by R01AG049704.

Funding Information:
We thank Prof. S. Kugler Department of Neurology, University of Gottingen, for the kind gift of adeno-associated virus plasmid. We thank Geertjan van Zonneveld for the professional approach and dedication to complete the illustrations of this research paper.

Publisher Copyright:
© 2021, The Author(s).

Copyright:
Copyright 2021 Elsevier B.V., All rights reserved.

Funding

This work was supported by grants to NMdW, SdH, MTM, JW, AR, PMM, JV, and HEV from ZonMw Memorabel program (projectnr: 733050105). PMM is also supported by the International Foundation for Alzheimer Research (ISAO) (projectnr: 14545). SMC received a travel grant support from Alzheimer Nederlands foundation (AN) to spend a month in the laboratory of EB, University of Kentucky, Lexington KY, USA. Aspects of this work were supported by the grants NIH R01AG034389, R01NS095215, and R56AG064234; VA I01BX003643 to EB. MMM was supported by R01AG049704. We thank Prof. S. Kugler Department of Neurology, University of Gottingen, for the kind gift of adeno-associated virus plasmid. We thank Geertjan van Zonneveld for the professional approach and dedication to complete the illustrations of this research paper.

Funders	Funder number
International Foundation for Alzheimer Research
University of Gottingen
ZonMw Memorabel program	733050105
National Institutes of Health	R01AG049704, R01AG034389, VA I01BX003643, R56AG064234
National Institute of Neurological Disorders and Stroke	R01NS095215
University of Kentucky
Internationale Stichting Alzheimer Onderzoek	14545
Alzheimer Nederland

Keywords

5xFAD
Adeno-associated virus (AAV)
Alzheimer’s disease (AD)
Amyloid-β plaques
Ceramide
Ceramide transporter protein (CERT)
Microglia
Neuroinflammation
Sphingomyelin

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Access to Document

10.1186/s13195-021-00780-0

Persistent URL (handle)

Persistent link

Cite this

Crivelli, S. M., Luo, Q., Stevens, J. A. A., Giovagnoni, C., van Kruining, D., Bode, G., den Hoedt, S., Hobo, B., Scheithauer, A. L., Walter, J., Mulder, M. T., Exley, C., Mold, M., Mielke, M. M., De Vries, H. E., Wouters, K., van den Hove, D. L. A., Berkes, D., Ledesma, M. D., ... Martinez-Martinez, P. (2021). CERT_L reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer’s disease. Alzheimer's Research and Therapy, 13(1), Article 45. https://doi.org/10.1186/s13195-021-00780-0

@article{4861577e37b244d3a5fe3bb6f591f79f,

title = "CERTL reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer{\textquoteright}s disease",

abstract = "Background: Dysregulation of ceramide and sphingomyelin levels have been suggested to contribute to the pathogenesis of Alzheimer{\textquoteright}s disease (AD). Ceramide transfer proteins (CERTs) are ceramide carriers which are crucial for ceramide and sphingomyelin balance in cells. Extracellular forms of CERTs co-localize with amyloid-β (Aβ) plaques in AD brains. To date, the significance of these observations for the pathophysiology of AD remains uncertain. Methods: A plasmid expressing CERTL, the long isoform of CERTs, was used to study the interaction of CERTL with amyloid precursor protein (APP) by co-immunoprecipitation and immunofluorescence in HEK cells. The recombinant CERTL protein was employed to study interaction of CERTL with amyloid-β (Aβ), Aβ aggregation process in presence of CERTL, and the resulting changes in Aβ toxicity in neuroblastoma cells. CERTL was overexpressed in neurons by adeno-associated virus (AAV) in a mouse model of familial AD (5xFAD). Ten weeks after transduction, animals were challenged with behavior tests for memory, anxiety, and locomotion. At week 12, brains were investigated for sphingolipid levels by mass spectrometry, plaques, and neuroinflammation by immunohistochemistry, gene expression, and/or immunoassay. Results: Here, we report that CERTL binds to APP, modifies Aβ aggregation, and reduces Aβ neurotoxicity in vitro. Furthermore, we show that intracortical injection of AAV, mediating the expression of CERTL, decreases levels of ceramide d18:1/16:0 and increases sphingomyelin levels in the brain of male 5xFAD mice. CERTL in vivo over-expression has a mild effect on animal locomotion, decreases Aβ formation, and modulates microglia by decreasing their pro-inflammatory phenotype. Conclusion: Our results demonstrate a crucial role of CERTL in regulating ceramide levels in the brain, in amyloid plaque formation and neuroinflammation, thereby opening research avenues for therapeutic targets of AD and other neurodegenerative diseases.",

keywords = "5xFAD, Adeno-associated virus (AAV), Alzheimer{\textquoteright}s disease (AD), Amyloid-β plaques, Ceramide, Ceramide transporter protein (CERT), Microglia, Neuroinflammation, Sphingomyelin",

author = "Crivelli, {Simone M.} and Qian Luo and Stevens, {Jo A.A.} and Caterina Giovagnoni and {van Kruining}, Daan and Gerard Bode and {den Hoedt}, Sandra and Barbara Hobo and Scheithauer, {Anna Lena} and Jochen Walter and Mulder, {Monique T.} and Christopher Exley and Matthew Mold and Mielke, {Michelle M.} and {De Vries}, {Helga E.} and Kristiaan Wouters and {van den Hove}, {Daniel L.A.} and Dusan Berkes and Ledesma, {Mar{\'i}a Dolores} and Joost Verhaagen and Mario Losen and Erhard Bieberich and Pilar Martinez-Martinez",

note = "Funding Information: This work was supported by grants to NMdW, SdH, MTM, JW, AR, PMM, JV, and HEV from ZonMw Memorabel program (projectnr: 733050105). PMM is also supported by the International Foundation for Alzheimer Research (ISAO) (projectnr: 14545). SMC received a travel grant support from Alzheimer Nederlands foundation (AN) to spend a month in the laboratory of EB, University of Kentucky, Lexington KY, USA. Aspects of this work were supported by the grants NIH R01AG034389, R01NS095215, and R56AG064234; VA I01BX003643 to EB. MMM was supported by R01AG049704. Funding Information: We thank Prof. S. Kugler Department of Neurology, University of Gottingen, for the kind gift of adeno-associated virus plasmid. We thank Geertjan van Zonneveld for the professional approach and dedication to complete the illustrations of this research paper. Publisher Copyright: {\textcopyright} 2021, The Author(s). Copyright: Copyright 2021 Elsevier B.V., All rights reserved.",

year = "2021",

month = dec,

doi = "10.1186/s13195-021-00780-0",

language = "English",

volume = "13",

journal = "Alzheimer's Research and Therapy",

issn = "1758-9193",

publisher = "BioMed Central",

number = "1",

}

Crivelli, SM, Luo, Q, Stevens, JAA, Giovagnoni, C, van Kruining, D, Bode, G, den Hoedt, S, Hobo, B, Scheithauer, AL, Walter, J, Mulder, MT, Exley, C, Mold, M, Mielke, MM, De Vries, HE, Wouters, K, van den Hove, DLA, Berkes, D, Ledesma, MD, Verhaagen, J, Losen, M, Bieberich, E & Martinez-Martinez, P 2021, 'CERT_L reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer’s disease', Alzheimer's Research and Therapy, vol. 13, no. 1, 45. https://doi.org/10.1186/s13195-021-00780-0

TY - JOUR

T1 - CERTL reduces C16 ceramide, amyloid-β levels, and inflammation in a model of Alzheimer’s disease

AU - Crivelli, Simone M.

AU - Luo, Qian

AU - Stevens, Jo A.A.

AU - Giovagnoni, Caterina

AU - van Kruining, Daan

AU - Bode, Gerard

AU - den Hoedt, Sandra

AU - Hobo, Barbara

AU - Scheithauer, Anna Lena

AU - Walter, Jochen

AU - Mulder, Monique T.

AU - Exley, Christopher

AU - Mold, Matthew

AU - Mielke, Michelle M.

AU - De Vries, Helga E.

AU - Wouters, Kristiaan

AU - van den Hove, Daniel L.A.

AU - Berkes, Dusan

AU - Ledesma, María Dolores

AU - Verhaagen, Joost

AU - Losen, Mario

AU - Bieberich, Erhard

AU - Martinez-Martinez, Pilar

N1 - Funding Information: This work was supported by grants to NMdW, SdH, MTM, JW, AR, PMM, JV, and HEV from ZonMw Memorabel program (projectnr: 733050105). PMM is also supported by the International Foundation for Alzheimer Research (ISAO) (projectnr: 14545). SMC received a travel grant support from Alzheimer Nederlands foundation (AN) to spend a month in the laboratory of EB, University of Kentucky, Lexington KY, USA. Aspects of this work were supported by the grants NIH R01AG034389, R01NS095215, and R56AG064234; VA I01BX003643 to EB. MMM was supported by R01AG049704. Funding Information: We thank Prof. S. Kugler Department of Neurology, University of Gottingen, for the kind gift of adeno-associated virus plasmid. We thank Geertjan van Zonneveld for the professional approach and dedication to complete the illustrations of this research paper. Publisher Copyright: © 2021, The Author(s). Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/12

Y1 - 2021/12

N2 - Background: Dysregulation of ceramide and sphingomyelin levels have been suggested to contribute to the pathogenesis of Alzheimer’s disease (AD). Ceramide transfer proteins (CERTs) are ceramide carriers which are crucial for ceramide and sphingomyelin balance in cells. Extracellular forms of CERTs co-localize with amyloid-β (Aβ) plaques in AD brains. To date, the significance of these observations for the pathophysiology of AD remains uncertain. Methods: A plasmid expressing CERTL, the long isoform of CERTs, was used to study the interaction of CERTL with amyloid precursor protein (APP) by co-immunoprecipitation and immunofluorescence in HEK cells. The recombinant CERTL protein was employed to study interaction of CERTL with amyloid-β (Aβ), Aβ aggregation process in presence of CERTL, and the resulting changes in Aβ toxicity in neuroblastoma cells. CERTL was overexpressed in neurons by adeno-associated virus (AAV) in a mouse model of familial AD (5xFAD). Ten weeks after transduction, animals were challenged with behavior tests for memory, anxiety, and locomotion. At week 12, brains were investigated for sphingolipid levels by mass spectrometry, plaques, and neuroinflammation by immunohistochemistry, gene expression, and/or immunoassay. Results: Here, we report that CERTL binds to APP, modifies Aβ aggregation, and reduces Aβ neurotoxicity in vitro. Furthermore, we show that intracortical injection of AAV, mediating the expression of CERTL, decreases levels of ceramide d18:1/16:0 and increases sphingomyelin levels in the brain of male 5xFAD mice. CERTL in vivo over-expression has a mild effect on animal locomotion, decreases Aβ formation, and modulates microglia by decreasing their pro-inflammatory phenotype. Conclusion: Our results demonstrate a crucial role of CERTL in regulating ceramide levels in the brain, in amyloid plaque formation and neuroinflammation, thereby opening research avenues for therapeutic targets of AD and other neurodegenerative diseases.

AB - Background: Dysregulation of ceramide and sphingomyelin levels have been suggested to contribute to the pathogenesis of Alzheimer’s disease (AD). Ceramide transfer proteins (CERTs) are ceramide carriers which are crucial for ceramide and sphingomyelin balance in cells. Extracellular forms of CERTs co-localize with amyloid-β (Aβ) plaques in AD brains. To date, the significance of these observations for the pathophysiology of AD remains uncertain. Methods: A plasmid expressing CERTL, the long isoform of CERTs, was used to study the interaction of CERTL with amyloid precursor protein (APP) by co-immunoprecipitation and immunofluorescence in HEK cells. The recombinant CERTL protein was employed to study interaction of CERTL with amyloid-β (Aβ), Aβ aggregation process in presence of CERTL, and the resulting changes in Aβ toxicity in neuroblastoma cells. CERTL was overexpressed in neurons by adeno-associated virus (AAV) in a mouse model of familial AD (5xFAD). Ten weeks after transduction, animals were challenged with behavior tests for memory, anxiety, and locomotion. At week 12, brains were investigated for sphingolipid levels by mass spectrometry, plaques, and neuroinflammation by immunohistochemistry, gene expression, and/or immunoassay. Results: Here, we report that CERTL binds to APP, modifies Aβ aggregation, and reduces Aβ neurotoxicity in vitro. Furthermore, we show that intracortical injection of AAV, mediating the expression of CERTL, decreases levels of ceramide d18:1/16:0 and increases sphingomyelin levels in the brain of male 5xFAD mice. CERTL in vivo over-expression has a mild effect on animal locomotion, decreases Aβ formation, and modulates microglia by decreasing their pro-inflammatory phenotype. Conclusion: Our results demonstrate a crucial role of CERTL in regulating ceramide levels in the brain, in amyloid plaque formation and neuroinflammation, thereby opening research avenues for therapeutic targets of AD and other neurodegenerative diseases.

KW - 5xFAD

KW - Adeno-associated virus (AAV)

KW - Alzheimer’s disease (AD)

KW - Amyloid-β plaques

KW - Ceramide

KW - Ceramide transporter protein (CERT)

KW - Microglia

KW - Neuroinflammation

KW - Sphingomyelin

UR - http://www.scopus.com/inward/record.url?scp=85101175394&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85101175394&partnerID=8YFLogxK

U2 - 10.1186/s13195-021-00780-0

DO - 10.1186/s13195-021-00780-0

M3 - Article

C2 - 33597019

AN - SCOPUS:85101175394

SN - 1758-9193

VL - 13

JO - Alzheimer's Research and Therapy

JF - Alzheimer's Research and Therapy

IS - 1

M1 - 45

ER -