Brain transcriptome analysis of a CLN2 mouse model as a function of disease progression

Abstract

Background Neuronal ceroid lipofuscinoses, (NCLs or Batten disease) are a group of inherited, early onset, fatal neurodegenerative diseases associated with mutations in 13 genes. All forms of the disease are characterized by lysosomal accumulation of fluorescent storage material, as well as profound neurodegeneration, but the relationship of the various genes’ function to a single biological process is not obvious. In this study, we used a well-characterized mouse model of classical late infantile NCL (cLINCL) in which the tripeptidyl peptidase 1 (Tpp1) gene is disrupted by gene targeting, resulting in loss of detectable TPP1 activity and leading to progressive neurological phenotypes including ataxia, increased motor deficiency, and early death. Methods In order to identify genes and pathways that may contribute to progression of the neurodegenerative process, we analyzed forebrain/midbrain and cerebellar transcriptional differences at 1, 2, 3 and 4 months of age in control and TPP1-deficient mice by global RNA-sequencing. Results Progressive neurodegenerative inflammatory responses involving microglia, astrocytes and endothelial cells were observed, accompanied by activation of leukocyte extravasation signals and upregulation of nitric oxide production and reactive oxygen species. Several astrocytic (i.e., Gfap, C4b, Osmr, Serpina3n) and microglial (i.e., Ctss, Itgb2, Itgax, Lyz2) genes were identified as strong markers for assessing disease progression as they showed increased levels of expression in vivo over time. Furthermore, transient increased expression of choroid plexus genes was observed at 2 months in the lateral and fourth ventricle, highlighting an early role for the choroid plexus and cerebrospinal fluid in the disease pathology. Based on these gene expression changes, we concluded that neuroinflammation starts, for the most part, after 2 months in the Tpp1−/− brain and that activation of microglia and astrocytes occur more rapidly in cerebellum than in the rest of the brain; confirming increased severity of inflammation in this region. Conclusions These findings have led to a better understanding of cLINCL pathological onset and progression, which may aid in development of future therapeutic treatments for this disease.

Glial Dysfunction and Its Contribution to the Pathogenesis of the Neuronal Ceroid Lipofuscinoses

“…The secretion of cytokines and chemokines is of paramount importance for both astrocytes and microglia to exert pro- and anti-inflammatory effects on the process of neurodegeneration ( 93 ). The progressive elevation of multiple cytokines and chemokines has been confirmed by whole transcriptomics and/or proteomics in the forebrains and cerebella of Tpp 1 −/− mice ( 75 , 90 ) and forebrains and spinal cords of Ppt 1 −/− mice ( 68 , 94 , 95 ). Such evidence for the region- and subtype-specific nature of neuroinflammatory changes in CLN1 and CLN2 diseases correlates with the previously shown region- and subtype-specific immunoreactivity of astrogliosis and microglial activation markers.…”

“…However, caution is needed in using the current A1/A2 classifications to interpret pathological roles of astrocytes, because such a binary A1/A2 paradigm may be an oversimplification of potentially more wide-ranging and heterogeneous states of astrogliosis ( 74 ). Indeed, the recent RNA sequencing data of Tpp 1 −/− mice have shown changes in the expression of a restricted subset of A1- or A2-specific genes, which does not match the typical A1/A2 classification ( 75 ). A lack of clear A1/A2 signature has also been reported in other diseases including Huntington disease ( 76 ), highlighting that astrocyte heterogeneity may convolute A1/A2 boundaries.…”

“…DAM further display upregulation of genes involved in lysosomal, phagocytic, and lipid metabolism pathways such as Apoe, Ctsd, Lpl, Tyrobp , and Trem2 , which perhaps makes the DAM classification particularly pertinent to LSDs. RNA sequencing data has revealed the existence of both TREM2-independent and TREM2-dependent DAM in Tpp 1 −/− mice, suggesting the pro-inflammatory and neurotoxic role of activated microglia in CLN2 disease ( 75 , 90 ). However, the pathological role of DAM still remains debatable; several recent studies have shown neuroprotective effects of TREM2-dependent DAM in mouse models of AD and GRN haploinsufficiency-causing frontotemporal lobar degeneration (GRN-FTLD) ( 91 , 92 ), suggesting the pathological contribution of DAM may well be disease-dependent.…”

“…Since phagocytosis requires focal exocytosis of lysosomes ( 104 ), it is plausible to speculate that lysosomal dysfunction due to NCL protein deficiency could also impair phagocytosis in these cells. While there have been several pieces of evidence from RNA sequencing or proteomics analysis suggesting altered phagocytosis in the brains of Ppt 1 −/− and Tpp 1 −/− mice ( 75 , 94 ), microglial-specific alteration of phagocytosis is yet to be elucidated. A better understanding of the nature of such dysfunctional phagocytosis by microglia and its contribution to NCL pathogenesis may therefore inform us of new therapeutic targets.…”

Paeonol Ameliorates Chronic Itch and Spinal Astrocytic Activation via CXCR3 in an Experimental Dry Skin Model in Mice

“…AEW induces persistent upregulation of Tlr4 mRNA in GFAPexpressing astrocytes and TLR4-dependent astrogliosis (GFAP upregulation) in the spinal dorsal horn (Tsuda, 2018). In addition, reactive astrogliosis consists of a rapid, but quickly weakened, induction of gene expression after injury, including Cp (Ryan et al, 2018;Wu et al, 2018), Vim (Qian et al, 2015;Smith et al, 2018;Adolf et al, 2019) and Serpina3n (Zamanian et al, 2012;Domowicz et al, 2021;Ji et al, 2021). In our study, their expression responds to AEW stimulation increased significantly.…”