Issues with the Specificity of Immunological Reagents for NLRP3: Implications for Age-related Macular Degeneration

Cassandra Kosmidou, Nikolaos E. Efstathiou, Mien V. Hoang, Shoji Notomi, Eleni K. Konstantinou, Masayuki Hirano, Kosuke Takahashi, Daniel E. Maidana, Pavlina Tsoka, Lucy Young, Evangelos S. Gragoudas, Timothy W. Olsen, Yuki Morizane, Joan W. Miller, Demetrios G. Vavvas

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.

Original languageEnglish
Article number461
JournalScientific Reports
Volume8
Issue number1
DOIs
Publication statusPublished - Dec 1 2018

Fingerprint

Inflammasomes
Macular Degeneration
Cryopyrin-Associated Periodic Syndromes
Induced Pluripotent Stem Cells
Western World
Antibody Specificity
National Institutes of Health (U.S.)
Nervous System
Transfection
Limit of Detection
Anti-Idiotypic Antibodies
Up-Regulation
Guidelines
RNA
Cell Line
Skin

ASJC Scopus subject areas

  • General

Cite this

Kosmidou, C., Efstathiou, N. E., Hoang, M. V., Notomi, S., Konstantinou, E. K., Hirano, M., ... Vavvas, D. G. (2018). Issues with the Specificity of Immunological Reagents for NLRP3: Implications for Age-related Macular Degeneration. Scientific Reports, 8(1), [461]. https://doi.org/10.1038/s41598-017-17634-1

Issues with the Specificity of Immunological Reagents for NLRP3 : Implications for Age-related Macular Degeneration. / Kosmidou, Cassandra; Efstathiou, Nikolaos E.; Hoang, Mien V.; Notomi, Shoji; Konstantinou, Eleni K.; Hirano, Masayuki; Takahashi, Kosuke; Maidana, Daniel E.; Tsoka, Pavlina; Young, Lucy; Gragoudas, Evangelos S.; Olsen, Timothy W.; Morizane, Yuki; Miller, Joan W.; Vavvas, Demetrios G.

In: Scientific Reports, Vol. 8, No. 1, 461, 01.12.2018.

Research output: Contribution to journalArticle

Kosmidou, C, Efstathiou, NE, Hoang, MV, Notomi, S, Konstantinou, EK, Hirano, M, Takahashi, K, Maidana, DE, Tsoka, P, Young, L, Gragoudas, ES, Olsen, TW, Morizane, Y, Miller, JW & Vavvas, DG 2018, 'Issues with the Specificity of Immunological Reagents for NLRP3: Implications for Age-related Macular Degeneration', Scientific Reports, vol. 8, no. 1, 461. https://doi.org/10.1038/s41598-017-17634-1
Kosmidou, Cassandra ; Efstathiou, Nikolaos E. ; Hoang, Mien V. ; Notomi, Shoji ; Konstantinou, Eleni K. ; Hirano, Masayuki ; Takahashi, Kosuke ; Maidana, Daniel E. ; Tsoka, Pavlina ; Young, Lucy ; Gragoudas, Evangelos S. ; Olsen, Timothy W. ; Morizane, Yuki ; Miller, Joan W. ; Vavvas, Demetrios G. / Issues with the Specificity of Immunological Reagents for NLRP3 : Implications for Age-related Macular Degeneration. In: Scientific Reports. 2018 ; Vol. 8, No. 1.
@article{a0d5b1c41e8645a5a36e0abb4f437eab,
title = "Issues with the Specificity of Immunological Reagents for NLRP3: Implications for Age-related Macular Degeneration",
abstract = "Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.",
author = "Cassandra Kosmidou and Efstathiou, {Nikolaos E.} and Hoang, {Mien V.} and Shoji Notomi and Konstantinou, {Eleni K.} and Masayuki Hirano and Kosuke Takahashi and Maidana, {Daniel E.} and Pavlina Tsoka and Lucy Young and Gragoudas, {Evangelos S.} and Olsen, {Timothy W.} and Yuki Morizane and Miller, {Joan W.} and Vavvas, {Demetrios G.}",
year = "2018",
month = "12",
day = "1",
doi = "10.1038/s41598-017-17634-1",
language = "English",
volume = "8",
journal = "Scientific Reports",
issn = "2045-2322",
publisher = "Nature Publishing Group",
number = "1",

}

TY - JOUR

T1 - Issues with the Specificity of Immunological Reagents for NLRP3

T2 - Implications for Age-related Macular Degeneration

AU - Kosmidou, Cassandra

AU - Efstathiou, Nikolaos E.

AU - Hoang, Mien V.

AU - Notomi, Shoji

AU - Konstantinou, Eleni K.

AU - Hirano, Masayuki

AU - Takahashi, Kosuke

AU - Maidana, Daniel E.

AU - Tsoka, Pavlina

AU - Young, Lucy

AU - Gragoudas, Evangelos S.

AU - Olsen, Timothy W.

AU - Morizane, Yuki

AU - Miller, Joan W.

AU - Vavvas, Demetrios G.

PY - 2018/12/1

Y1 - 2018/12/1

N2 - Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.

AB - Contradictory data have been presented regarding the implication of the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome in age-related macular degeneration (AMD), the leading cause of vision loss in the Western world. Recognizing that antibody specificity may explain this discrepancy and in line with recent National Institutes of Health (NIH) guidelines requiring authentication of key biological resources, the specificity of anti-NLRP3 antibodies was assessed to elucidate whether non-immune RPE cells express NLRP3. Using validated resources, NLRP3 was not detected in human primary or human established RPE cell lines under multiple inflammasome-priming conditions, including purported NLRP3 stimuli in RPE such as DICER1 deletion and Alu RNA transfection. Furthermore, NLRP3 was below detection limits in ex vivo macular RPE from AMD patients, as well as in human induced pluripotent stem cell (hiPSC)-derived RPE from patients with overactive NLRP3 syndrome (Chronic infantile neurologic cutaneous and articulate, CINCA syndrome). Evidence presented in this study provides new data regarding the interpretation of published results reporting NLRP3 expression and upregulation in RPE and addresses the role that this inflammasome plays in AMD pathogenesis.

UR - http://www.scopus.com/inward/record.url?scp=85040513596&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85040513596&partnerID=8YFLogxK

U2 - 10.1038/s41598-017-17634-1

DO - 10.1038/s41598-017-17634-1

M3 - Article

C2 - 29323137

AN - SCOPUS:85040513596

VL - 8

JO - Scientific Reports

JF - Scientific Reports

SN - 2045-2322

IS - 1

M1 - 461

ER -