Quantifying lymphocyte vacuolization serves as a measure of CLN3 disease severity
Kuper, Willemijn F E; Oostendorp, Marlies; van den Broek, Brigitte T A; van Veghel, Karin; Nonkes, Lourens J P; Nieuwenhuis, Edward E S; Fuchs, Sabine A; Veenendaal, Tineke; Klumperman, Judith; Huisman, Albert; Nierkens, Stefan; van Hasselt, Peter M
(2020) JIMD Reports, volume 54, issue 1, pp. 87 - 97
(Article)
Abstract
Background: The CLN3 disease spectrum ranges from a childhood-onset neurodegenerative disorder to a retina-only disease. Given the lack of metabolic disease severity markers, it may be difficult to provide adequate counseling, particularly when novel genetic variants are identified. In this study, we assessed whether lymphocyte vacuolization, a well-known yet poorly
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explored characteristic of CLN3 disease, could serve as a measure of disease severity. Methods: Peripheral blood obtained from healthy controls and CLN3 disease patients was used to assess lymphocyte vacuolization by (a) calculating the degree of vacuolization using light microscopy and (b) quantifying expression of lysosomal-associated membrane protein 1 (LAMP-1), using flow cytometry in lymphocyte subsets as well as a qualitative analysis using electron microscopy and ImageStream analysis. Results: Quantifying lymphocyte vacuolization allowed to differentiate between CLN3 disease phenotypes (P =.0001). On immunofluorescence, classical CLN3 disease lymphocytes exhibited abundant vacuole-shaped LAMP-1 expression, suggesting the use of LAMP-1 as a proxy for lymphocyte vacuolization. Using flow cytometry in lymphocyte subsets, quantifying intracellular LAMP-1 expression additionally allowed to differentiate between infection and storage and to differentiate between CLN3 phenotypes even more in-depth revealing that intracellular LAMP-1 expression was most pronounced in T-cells of classical-protracted CLN3 disease while it was most pronounced in B-cells of “retina-only” CLN3 disease. Conclusion: Lymphocyte vacuolization serves as a proxy for CLN3 disease severity. Quantifying vacuolization may help interpretation of novel genetic variants and provide an individualized readout for upcoming therapies.
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Keywords: CLN3 disease, ImageStream, flow cytometry, lymphocyte vacuolization, lysosomal membrane‐associated protein‐1 (LAMP‐1), neuronal ceroid lipofuscinosis (NCL), lysosomal membrane-associated protein-1 (LAMP-1), Biochemistry, Genetics and Molecular Biology (miscellaneous), Internal Medicine, Endocrinology, Diabetes and Metabolism, Journal Article
ISSN: 2192-8304
Publisher: Springer Berlin
Note: Funding Information: The authors thank Evertine Heek, Amelia Lacna, and Jeroen van Velzen for their technical support. The present study is not industry sponsored. W. F. E. K. receives unrestricted financial support from the Beat Batten Foundation, the Friends of the Wilhelmina Children's Hospital Foundation, and the Bartiméus Foundation. B. T. A. B. receives unrestricted financial support from the Sylvia Toth Charity Foundation. The authors confirm independence from the sponsors; the content of this article has not been influenced by the sponsors. Funding Information: The authors thank Evertine Heek, Amelia Lacna, and Jeroen van Velzen for their technical support. The present study is not industry sponsored. W. F. E. K. receives unrestricted financial support from the Beat Batten Foundation, the Friends of the Wilhelmina Children's Hospital Foundation, and the Bartim?us Foundation. B. T. A. B. receives unrestricted financial support from the Sylvia Toth Charity Foundation. The authors confirm independence from the sponsors; the content of this article has not been influenced by the sponsors. Publisher Copyright: © 2020 The Authors. Journal of Inherited Metabolic Disease published by John Wiley & Sons Ltd on behalf of SSIEM.
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