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  Glial and vascular contributions to neurodegenerative diseases
Single cell sequencing-based exploration
of microglia subpopulations in accelerated aging mice
Robert-Josep Subirana-Slotos1, Sebastian Kunz2, Matthias Klein2, Tobias Bopp2, Kristina Endres1
1Department of Psychiatry and Psychotherapy, University Medical Center, Mainz, Germany,
2Institute for Immunology, University Medical Center, Johannes Gutenberg-Universität, Mainz, Germany
 Normal aging is defined as the accumulation of changes over time. An accelerated aging process can lead to defects and even diseases such as dementia. Worldwide, 10.9% of people over the age of 65 suffer from dementia, primarily due to Alzheimer's disease (AD). While familial AD is clearly evoked by mutations in a certain number of genes, the cause of sporadic AD is still enigmatic [1]. However, aging is a predominant risk factor and inflammation caused by chronic stimulation of the immune system has been shown to play a pivotal role in AD. Especially, distinct microglia subpopulations may be crucial in pathogenesis, as they can have beneficial but also detrimental effects regarding AD.
Senescence Accelerated Mouse-Prone 8 (SAMP8) [2] mice were subjected to various motor tests and visually monitored to assess an age-related frailty index. 3-month-old SAMP8 mice showed only a limited amount of decline as compared to the control, Senescence Accelerated Mouse-Restrained 1 (SAMR1). Older, 10-month-old mice revealed severe motor deterioration and low vitality. Henceforth, frontal brain of young and old SAMP8 and SAMR1 mice was subjected to single-cell sequencing.
We successfully applied the Adult Brain Dissociation Kit (Miltenyi Biotec) to dissociate parenchymal tissue and removed contaminants such as myelin debris and red blood cells while maintaining cell viability above 87% using a trehalose treatment modified from [3]. The high viability enabled successful generation of cDNA libraries from approximately 10,000 cells per brain. This allowed an accurate resolution of even smaller cell subpopulations.
Ultimately, this study will provide insight in age-related changes in cellular composition of the murine brain and will indicate if accelerated aging parallels AD in terms of microglial and other cell subpopulations.
time-dependent molecular signatures in the hippocampus that may be related to epileptogenesis. Principal component analysis showed that inflammatory signatures of acute (d1), latent (d3-7) and chronic (d14- 28) timepoints were distinct without a return to homeostatic conditions at 28 days post-SE, indicative of a chronic inflammatory state. The acute signature was enriched in pro-inflammatory immune and type II IFN signaling genes while the latent signature showed a loss of neuronal genes, which coincided with increased and sustained expression of neurodegeneration-associated genes, including Grn, C3, Igf1, CD74 and NLRP3 inflammasome. Rats that experienced SE exhibited enlarged lateral ventricles at 14-28 days post-SE consistent with progressive severe neuronal loss. Moreover, microglial and astrocyte activation increased over time and persisted up to 28 days post-SE.
Conclusion: Together, these data provide evidence of commonalities between delayed neuroinflammation following SE and neurode- generative diseases such as Alzheimer’s disease. This study also identified complement C3 as a potential mediator of neurodegeneration following SE that warrants further investigation.
Evidence of cerebellar TDP-43 loss of function in FTLD-TDP
Sarah Pickles1, Tania Gendron1, Yuka Koike1, Mei Yue1, Yuping Song1, Jennifer Kachergus1, J Shi1, Michael DeTure1, E. Aubrey Thompson1, Björn Oskarsson1, Neill Graff-Radford1, Bradley Boeve2, Ronald Petersen2, Zbigniew Wszolek1, Keith Josephs2, Dennis Dickson1, Leonard Petrucelli1, Casey Cook1,
Mercedes Prudencio1
1Mayo Clinic Florida, Jacksonville, USA , 2Mayo Clinic Rochester, Rochester, USA
Hippocampal transcriptomics reveal time- dependent neurodegeneration-associated immune signatures related to epileptogenesis Claudia Espinosa-Garcia1,2, Asheebo Rojas1, Hailian Xiao2, Young Lin2, Aditya Natu2, Alejandra Valdivia3, Raymond Dingledine1, Srikant Rangaraju2
1Department of Pharmacology and Chemical Biology, Emory University School of Medicine, Atlanta, USA, 2Department of Neurology, Emory University School of Medicine, Atlanta, USA, 3Department of Medicine, Emory University School of Medicine, Atlanta, USA
 Background: More than 50 million people worldwide suffer from epilepsy–a chronic neurological disease characterized by recurrent unprovoked seizures. Seizures result in neuroinflammation, a key factor contributing to epileptogenesis. We hypothesize that a time course study of hippocampal transcriptome profiles might identify new targets that can be useful in the prevention and treatment of epilepsy. Materials and Methods: We used the pilocarpine-induced status epilepticus (SE) model to recapitulate characteristics of human temporal lobe epilepsy. Cohorts of adult male rats (n=4) were euthanized at 1, 3, 7, 14 and 28 days post-SE, these timepoints correspond to the acute, latent, and chronic phases of epileptogenesis. Hippocampal transcriptome profiles were assessed using the nCounter Neuroinflammation Panel (NanoString). Genes exhibiting log2(fold change) > 1 or log2 (fold change) < -1 with adjusted p-value < 0.05 were identified as significant. Results and discussion: Transcriptomic profiling identified distinct
Background: Neuronal and glial inclusions containing TDP-43, accompanied by TDP-43 nuclear depletion, are a pathological hallmark of Frontotemporal lobar degeneration with TDP-43 pathology (FTLD- TDP), a neurodegenerative disease primarily affecting the frontal and/or temporal cortices. [1]. The cerebellum has historically been underappreciated in FTLD-TDP given the absence of TDP-43 inclusions and significant neurodegeneration in this region. However, a growing body of evidence suggests that the cerebellum contributes to biochemical, cognitive, and behavioral changes in FTLD-TDP. Previous studies discovered that TDP-43 suppresses the inclusion of cryptic exons in numerous transcripts [2], including STMN2 [3, 4]. A truncated variant of STMN2 (tSTMN2), generated by the aberrant inclusion of a stop-codon containing cryptic exon, is detected in ALS/FTLD cases with TDP-43 proteinopathy [3-5]. Intriguingly, tSTMN2 was also detected in the cerebellum of some ALS/FTLD cases [5], suggesting a loss of TDP-43 splicing activity in this region.
Materials and methods: To evaluate cerebellar TDP-43 expression and function in FTLD-TDP, we analyzed TDP-43 protein levels and the splicing of STMN2, in 95 FTLD-TDP cases and 25 non-neurological disease controls.
Results: Truncated STMN2 transcripts were elevated in the cerebellum of FTLD-TDP cases. Soluble TDP-43 was decreased in the cerebellum of FTLD-TDP cases but a concomitant increase in insoluble TDP-43 was not seen. Additionally, lower cerebellar TDP-43 associated with a younger age at disease onset.
Discussion: We provide evidence of TDP-43 loss of function in the cerebellum of FTLD-TDP cases, highlighting that the consequences of TDP-43 dysfunction may be more extensive than previously thought. Higher levels of TDP-43 in the cerebellum associated with an older age at disease onset, suggesting that maintaining TDP-43 levels in the cerebellum may be protective.
Conclusions: Taken together, our findings suggest that a loss of TDP- 43 in the cerebellum is implicated in FTLD pathogenesis, supporting further investigation into this underappreciated brain region.
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