Biology subjects

Pierson, W.

Publications and source records attributed to Pierson, W..

4 recordsLinked to original sources

bioRxiv2025

Single cell transcriptomics reveals enrichment of aggregation-prone alpha-synuclein isoforms across synucleinopathies

Alpha-synuclein (-Syn) is the primary component of Lewy bodies, the pathological hallmark of neurodegenerative synucleinopathies, including Parkinsons disease (PD) and dementia with Lewy bodies (DLB). Dysregulated expression of its encoding gene, SNCA, has been identified in association with both PD and DLB in short-read sequencing studies. However, such studies do not capture variation in transcript isoforms expressed. Here we combine for the first time SNCA-targeted long-read multiplexed arrays isoform sequencing (MAS-Iso-seq) with unbiased short-read single nucleus (sn) RNA-seq for simultaneous characterization of the SNCA transcript isoform landscape and mapping of isoform expression to specific cell types and subtypes in PD, DLB, and control sample cortical tissues. This approach enabled discovery of numerous SNCA transcript isoforms displaying novel splicing patterns and incorporating novel exons. We further identified an abundant class of transcript isoforms encoding a previously unreported -Syn protein variant (-Syn-115) and displaying increased proportional detection in excitatory neurons of PD and DLB tissues in comparison to controls. The proportion of these isoforms was found to be especially high within several specific glutamatergic neuron subtypes. In-depth characterization of the predicted structural and biochemical properties of -Syn-115 using an in silico modeling approach revealed a greater aggregative affinity compared with canonical -Syn-140, suggesting the potential for increased cytosolic -Syn-115 abundance to induce aggregation between heterogeneous -Syn isoforms, potentially driving fibril formation and disease progression. Together, our findings provide new insights into the molecular mechanisms underlying -Syn involvement in multiple synucleinopathies and have translational implications for the development of new precision medicine strategies to combat these diseases, indicating the potential for treatments targeting both specific transcript and protein isoforms, as well as disease-driving cell subtypes.

neuroscience↗

bioRxiv2023

Sustained liver HBsAg loss and clonal T and B cell expansion upon therapeutic DNA vaccination require low HBsAg levels

Background & AimsSuppression of HBV DNA, inhibition of HBsAg production and therapeutic vaccination to reverse HBV-specific T-cell exhaustion in chronic HBV patients are likely required to achieve functional cure. In the AAV-HBV mouse model, therapeutic vaccination can be effective in clearing HBsAg when hepatitis B surface (HBsAg) levels are low. The factor(s) required for mounting an effective immune control of HBV infection are unclear. Using a single-cell approach, we investigated the liver immune environment in the context of different levels of HBsAg as well as upon sustained HBsAg loss through treatment with an HBV specific GalNAc-siRNA followed by therapeutic vaccination. MethodsC57BL/6 mice were transduced with a range of rAAV-HBV DNA to express different HBsAg levels. Mice were treated with GalNAc-siRNA targeting HBV transcripts to lower the HBsAg levels and then vaccinated 4 times with a DNA vaccine encoding HBV Core, Pol and Surface. We used single-cell RNA-sequencing on homogenised liver resident cells, paired with single-cell V(D)J receptor sequencing to understand the changes in the liver immune microenvironment. ResultsTreatment with GalNAc-HBV siRNA followed by therapeutic vaccination, achieved a sustained HBsAg loss in all mice. This was accompanied by an induction of CD4 follicular helper T-cell responses, polyclonal activation of CD8 T-cells in the liver and clonal expansion of plasma cells that were responsible for antibody production. ConclusionsThis study provides novel insight into the immune changes in the liver at the single-cell level, highlighting the correlation between the induced reduction in HBsAg levels and the clonal expansion of CD4 follicular helper T-cells, CD8 cytotoxic T-cells, plasma cells, and ISG-producing neutrophils in the liver upon HBV siRNA and subsequent therapeutic vaccine treatment. Lay SummaryChronic hepatitis B infection is characterized by a complex interplay between immune responses and viral replication in the liver. To achieve functional cure a combination of different treatments is likely required. In this study single-cell approach was used to understand the liver microenvironment in the context of different HBsAg levels followed by therapeutic vaccination in AAV-HBV mouse model and to identify key factors required to achieve functional cure. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/556204v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@5776a4org.highwire.dtl.DTLVardef@2e1b9forg.highwire.dtl.DTLVardef@1179a2eorg.highwire.dtl.DTLVardef@8b801d_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIAAV-HBV transduced mice sequentially treated with GalNAc-siRNA and therapeutic vaccine showed sustained HBsAg loss. C_LIO_LIThe sustained HBsAg loss correlates with increased proportion and clonal expansion of CD4 follicular helper T-cells, CD8 cytotoxic T-cells, plasma cells, and ISG producing neutrophils in the liver. C_LIO_LIBaseline levels of HBsAg are important to determine outcome of therapeutic vaccination in mice. C_LI

immunology↗

bioRxiv2023

AAV-HBV mouse model replicates immune exhaustion patterns of chronic HBV patients at single-cell level

Background and AimsUnresolved hepatitis B virus (HBV) infection leads to a progressive state of immune exhaustion that impairs resolution of infection, leading to chronic infection (CHB). The immune-competent AAV-HBV mouse is a common HBV preclinical immune competent model, though a comprehensive characterization of the liver immune microenvironment and its translatability to human infection is still lacking. We investigated the intrahepatic immune profile of the AAV-HBV mouse model at a single-cell level and compared with data from CHB patients in immune tolerant (IT) and immune active (IA) clinical stages. MethodsImmune exhaustion was profiled through an iterative subclustering approach for cell-typing analyses of single-cell RNA-sequencing data in CHB donors and compared to the AAV-HBV mouse model 24-weeks post-transduction to assess its translatability. This was validated using an exhaustion flow cytometry panel at 40 weeks post-transduction. ResultsUsing single-cell RNA-sequencing, CD8 pre-exhausted T-cells with self-renewing capacity (TCF7+), and terminally exhausted CD8 T-cells (TCF7-) were detected in the AAV-HBV model. These terminally exhausted CD8 T-cells (expressing Pdcd1, Tox, Lag3, Tigit) were significantly enriched versus control mice and independently identified through flow cytometry. Importantly, comparison to CHB human data showed a similar exhausted CD8 T-cell population in IT and IA donors, but not in healthy individuals. ConclusionsLong term high titer AAV-HBV mouse liver transduction led to T-cell exhaustion, as evidenced by expression of classical immune checkpoint markers at mRNA and protein levels. In both IT and IA donors, a similar CD8 exhausted T-cell population was identified, with increased frequency observed in IA donors. These data support the use of the AAV-HBV mouse model to study T-cell exhaustion in HBV infection and the effect of immune-based therapeutic interventions. Lay SummaryThe AAV-HBV mouse model is used as a research tool to study hepatitis B infection. In this study we evaluated the translation value from mouse to human with regards to T-cell exhaustion. HighlightsO_LIAAV-HBV mice transduced with a high titer vector showed presence of CD8 exhausted T-cells after 24 weeks. C_LIO_LIHigh titer transduced mice, but not lower titer show increased expression of LAG-3, TOX, TIM-3 and TIGIT in CD8 T-cells. PD-1 was increased in CD8 T-cells, independent of HBV transduction titer. C_LIO_LIA similar exhausted CD8 T-cell population could be found in chronic HBV donors, but not in healthy individuals. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=75 SRC="FIGDIR/small/552328v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@17db34dorg.highwire.dtl.DTLVardef@186a0b1org.highwire.dtl.DTLVardef@19fd39borg.highwire.dtl.DTLVardef@1fd9f64_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2019

Follicular regulatory T cells can access the germinal centre independently of CXCR5

The germinal centre (GC) response is critical for generating high-affinity humoral immunity and immunological memory, which forms the basis of successful immunisation. Control of the GC response is thought to require follicular regulatory T (Tfr) cells, a subset of suppressive Foxp3+ Treg cells located within GCs. Relatively little is known about the exact role of Tfr cells within the GC, and the mechanism/s through which they exert their suppressive function. A unique feature of Tfr cells is their reported CXCR5-dependent localisation to the GC. Here we show that the lack of CXCR5 on Foxp3+ regulatory T cells resulted in a reduced frequency, but not an absence of, GC-localised Tfr cells. This demonstrates that additional, CXCR5-independent mechanisms facilitate Treg cell homing to the GC.

immunology↗