Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily

Thomas Bitterli; David Schmid; Ladina Ettinger; Olga Krupkova; Frances C. Bach; Marianna A. Tryfonidou; Björn P. Meij; Antonio Pozzi; Frank Steffen; Karin Wuertz‐Kozak; Lucas A. Smolders

doi:10.1002/jsp2.1292

Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily

Thomas Bitterli, David Schmid, Ladina Ettinger, Olga Krupkova, Frances C. Bach, Marianna A. Tryfonidou, Björn P. Meij, Antonio Pozzi, Frank Steffen, Karin Wuertz‐Kozak, Lucas A. Smolders^*

^*Corresponding author for this work

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

Abstract

Background
The regulation of inflammatory mediators in the degenerating intervertebral disc (IVD) and corresponding ligamentum flavum (LF) is a topic of emerging interest. The study aimed to investigate the expression of a broad array of inflammatory mediators in the degenerated LF and IVD using a dog model of spontaneous degenerative disc disease (DDD) to determine potential treatment targets.

Methods
LF and IVD tissues were collected from 22 normal dogs (Pfirrmann grades I and II) and 18 dogs affected by DDD (Pfirrmann grades III and IV). A qPCR gene array was used to investigate the expression of 80 inflammatory genes for LF and IVD tissues, whereafter targets of interest were investigated in additional tissue samples using qPCR, western blot (WB), and immunohistochemistry.

Results
Tumor necrosis factor superfamily (TNFSF) signaling was identified as a regulated pathway in DDD, based on the significant regulation (n-fold ± SD) of various TNFSF members in the degenerated IVD, including nerve growth factor (NGF; −8 ± 10), CD40LG (464 ± 442), CD70 (341 ± 336), TNFSF Ligand 10 (9 ± 8), and RANKL/TNFSF Ligand 11 (85 ± 74). In contrast, TNFSF genes were not significantly affected in the degenerated LF compared to the control LF. Protein expression of NGF (WB) was significantly upregulated in both the degenerated LF (4.4 ± 0.5) and IVD (11.3 ± 5.6) compared to the control group. RANKL immunopositivity was significantly upregulated in advanced stages of degeneration (Thompson grades IV and V) in the nucleus pulposus and annulus fibrosus of the IVD, but not in the LF.

Conclusions
DDD involves a significant upregulation of various TNFSF members, with tissue-specific expression profiles in LF and IVD tissues. The differential involvement of TNFSF members within multiple spinal tissues from the same individual provides new insights into the inflammatory processes involved in DDD and may provide a basis to formulate hypotheses for the determination of potential treatment targets.

Original language	English
Article number	e1292
Pages (from-to)	1-19
Number of pages	19
Journal	JOR Spine
Volume	7
Issue number	1
DOIs	https://doi.org/10.1002/jsp2.1292
Publication status	Published - Mar 2024

Bibliographical note

Publisher Copyright:
© 2023 The Authors. JOR Spine published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.

Funding

This work was financially supported by Forschungskredit Postdoc, University of Zurich, Switzerland (Grant number FK14-053) and OPO Stiftung, Zurich, Switzerland, 2016. We would like to acknowledge the work of the Musculoskeletal Research Unit (MRSU; Salim Darwiche, Aymone Lenisa) for conducting the immunohistochemistry experiments and the Functional Genomics Center Zurich (FGCZ) of ETH Zurich and the University of Zurich for their support in conducting the genomics experiments. This work was financially supported by Forschungskredit Postdoc, University of Zurich, Switzerland (Grant number FK14‐053) and OPO Stiftung, Zurich, Switzerland, 2016. We would like to acknowledge the work of the Musculoskeletal Research Unit (MRSU; Salim Darwiche, Aymone Lenisa) for conducting the immunohistochemistry experiments and the Functional Genomics Center Zurich (FGCZ) of ETH Zurich and the University of Zurich for their support in conducting the genomics experiments.

Funders	Funder number
Musculoskeletal Research Unit
OPO-Stiftung
Eidgenössische Technische Hochschule Zürich
Universität Zürich	FK14‐053
Functional Genomics Center Zurich

Keywords

RANKL
back pain
degenerative disc disease
dog
inflammation
nerve growth factor
tumornecrosis factor superfamily

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JOR Spine - 2023 - Bitterli - Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogsFinal published version, 35.7 MBLicence: CC BY

Cite this

Bitterli, T., Schmid, D., Ettinger, L., Krupkova, O., Bach, F. C., Tryfonidou, M. A., Meij, B. P., Pozzi, A., Steffen, F., Wuertz‐Kozak, K., & Smolders, L. A. (2024). Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily. JOR Spine, 7(1), 1-19. Article e1292. https://doi.org/10.1002/jsp2.1292

@article{074d1b7fb41f474eaf1d59fa6ed7c837,

title = "Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily",

abstract = "BackgroundThe regulation of inflammatory mediators in the degenerating intervertebral disc (IVD) and corresponding ligamentum flavum (LF) is a topic of emerging interest. The study aimed to investigate the expression of a broad array of inflammatory mediators in the degenerated LF and IVD using a dog model of spontaneous degenerative disc disease (DDD) to determine potential treatment targets.MethodsLF and IVD tissues were collected from 22 normal dogs (Pfirrmann grades I and II) and 18 dogs affected by DDD (Pfirrmann grades III and IV). A qPCR gene array was used to investigate the expression of 80 inflammatory genes for LF and IVD tissues, whereafter targets of interest were investigated in additional tissue samples using qPCR, western blot (WB), and immunohistochemistry.ResultsTumor necrosis factor superfamily (TNFSF) signaling was identified as a regulated pathway in DDD, based on the significant regulation (n-fold ± SD) of various TNFSF members in the degenerated IVD, including nerve growth factor (NGF; −8 ± 10), CD40LG (464 ± 442), CD70 (341 ± 336), TNFSF Ligand 10 (9 ± 8), and RANKL/TNFSF Ligand 11 (85 ± 74). In contrast, TNFSF genes were not significantly affected in the degenerated LF compared to the control LF. Protein expression of NGF (WB) was significantly upregulated in both the degenerated LF (4.4 ± 0.5) and IVD (11.3 ± 5.6) compared to the control group. RANKL immunopositivity was significantly upregulated in advanced stages of degeneration (Thompson grades IV and V) in the nucleus pulposus and annulus fibrosus of the IVD, but not in the LF.ConclusionsDDD involves a significant upregulation of various TNFSF members, with tissue-specific expression profiles in LF and IVD tissues. The differential involvement of TNFSF members within multiple spinal tissues from the same individual provides new insights into the inflammatory processes involved in DDD and may provide a basis to formulate hypotheses for the determination of potential treatment targets.",

keywords = "RANKL, back pain, degenerative disc disease, dog, inflammation, nerve growth factor, tumornecrosis factor superfamily",

author = "Thomas Bitterli and David Schmid and Ladina Ettinger and Olga Krupkova and Bach, {Frances C.} and Tryfonidou, {Marianna A.} and Meij, {Bj{\"o}rn P.} and Antonio Pozzi and Frank Steffen and Karin Wuertz‐Kozak and Smolders, {Lucas A.}",

note = "Publisher Copyright: {\textcopyright} 2023 The Authors. JOR Spine published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.",

year = "2024",

month = mar,

doi = "10.1002/jsp2.1292",

language = "English",

volume = "7",

pages = "1--19",

journal = "JOR Spine",

issn = "2572-1143",

publisher = "Wiley-Blackwell Publishing Ltd",

number = "1",

}

Bitterli, T, Schmid, D, Ettinger, L, Krupkova, O, Bach, FC , Tryfonidou, MA , Meij, BP, Pozzi, A, Steffen, F, Wuertz‐Kozak, K & Smolders, LA 2024, 'Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily', JOR Spine, vol. 7, no. 1, e1292, pp. 1-19. https://doi.org/10.1002/jsp2.1292

TY - JOUR

T1 - Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily

AU - Bitterli, Thomas

AU - Schmid, David

AU - Ettinger, Ladina

AU - Krupkova, Olga

AU - Bach, Frances C.

AU - Tryfonidou, Marianna A.

AU - Meij, Björn P.

AU - Pozzi, Antonio

AU - Steffen, Frank

AU - Wuertz‐Kozak, Karin

AU - Smolders, Lucas A.

PY - 2024/3

Y1 - 2024/3

N2 - BackgroundThe regulation of inflammatory mediators in the degenerating intervertebral disc (IVD) and corresponding ligamentum flavum (LF) is a topic of emerging interest. The study aimed to investigate the expression of a broad array of inflammatory mediators in the degenerated LF and IVD using a dog model of spontaneous degenerative disc disease (DDD) to determine potential treatment targets.MethodsLF and IVD tissues were collected from 22 normal dogs (Pfirrmann grades I and II) and 18 dogs affected by DDD (Pfirrmann grades III and IV). A qPCR gene array was used to investigate the expression of 80 inflammatory genes for LF and IVD tissues, whereafter targets of interest were investigated in additional tissue samples using qPCR, western blot (WB), and immunohistochemistry.ResultsTumor necrosis factor superfamily (TNFSF) signaling was identified as a regulated pathway in DDD, based on the significant regulation (n-fold ± SD) of various TNFSF members in the degenerated IVD, including nerve growth factor (NGF; −8 ± 10), CD40LG (464 ± 442), CD70 (341 ± 336), TNFSF Ligand 10 (9 ± 8), and RANKL/TNFSF Ligand 11 (85 ± 74). In contrast, TNFSF genes were not significantly affected in the degenerated LF compared to the control LF. Protein expression of NGF (WB) was significantly upregulated in both the degenerated LF (4.4 ± 0.5) and IVD (11.3 ± 5.6) compared to the control group. RANKL immunopositivity was significantly upregulated in advanced stages of degeneration (Thompson grades IV and V) in the nucleus pulposus and annulus fibrosus of the IVD, but not in the LF.ConclusionsDDD involves a significant upregulation of various TNFSF members, with tissue-specific expression profiles in LF and IVD tissues. The differential involvement of TNFSF members within multiple spinal tissues from the same individual provides new insights into the inflammatory processes involved in DDD and may provide a basis to formulate hypotheses for the determination of potential treatment targets.

AB - BackgroundThe regulation of inflammatory mediators in the degenerating intervertebral disc (IVD) and corresponding ligamentum flavum (LF) is a topic of emerging interest. The study aimed to investigate the expression of a broad array of inflammatory mediators in the degenerated LF and IVD using a dog model of spontaneous degenerative disc disease (DDD) to determine potential treatment targets.MethodsLF and IVD tissues were collected from 22 normal dogs (Pfirrmann grades I and II) and 18 dogs affected by DDD (Pfirrmann grades III and IV). A qPCR gene array was used to investigate the expression of 80 inflammatory genes for LF and IVD tissues, whereafter targets of interest were investigated in additional tissue samples using qPCR, western blot (WB), and immunohistochemistry.ResultsTumor necrosis factor superfamily (TNFSF) signaling was identified as a regulated pathway in DDD, based on the significant regulation (n-fold ± SD) of various TNFSF members in the degenerated IVD, including nerve growth factor (NGF; −8 ± 10), CD40LG (464 ± 442), CD70 (341 ± 336), TNFSF Ligand 10 (9 ± 8), and RANKL/TNFSF Ligand 11 (85 ± 74). In contrast, TNFSF genes were not significantly affected in the degenerated LF compared to the control LF. Protein expression of NGF (WB) was significantly upregulated in both the degenerated LF (4.4 ± 0.5) and IVD (11.3 ± 5.6) compared to the control group. RANKL immunopositivity was significantly upregulated in advanced stages of degeneration (Thompson grades IV and V) in the nucleus pulposus and annulus fibrosus of the IVD, but not in the LF.ConclusionsDDD involves a significant upregulation of various TNFSF members, with tissue-specific expression profiles in LF and IVD tissues. The differential involvement of TNFSF members within multiple spinal tissues from the same individual provides new insights into the inflammatory processes involved in DDD and may provide a basis to formulate hypotheses for the determination of potential treatment targets.

KW - RANKL

KW - back pain

KW - degenerative disc disease

KW - dog

KW - inflammation

KW - nerve growth factor

KW - tumornecrosis factor superfamily

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U2 - 10.1002/jsp2.1292

DO - 10.1002/jsp2.1292

M3 - Article

SN - 2572-1143

VL - 7

SP - 1

EP - 19

JO - JOR Spine

JF - JOR Spine

IS - 1

M1 - e1292

ER -

Targeted screening of inflammatory mediators in spontaneous degenerative disc disease in dogs reveals an upregulation of the tumor necrosis superfamily

Abstract

Bibliographical note

Funding

Keywords

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