Biology subjects

Bury, A.

Publications and source records attributed to Bury, A..

4 recordsLinked to original sources

bioRxiv2026

Complement Dysregulation During the Early Phases of Synucleinopathy

Parkinsons disease (PD) is characterized by progressive degeneration of nigrostriatal dopamine neurons and synucleinopathy, which is the accumulation of aggregated -synuclein (-syn). Increasing evidence implicates -syn-associated neuroinflammation as a contributor to PD pathogenesis; however, immune mechanisms linking synucleinopathy to neurodegeneration remain incompletely defined. Activation of the complement cascade occurs in PD and other neurodegenerative disorders, but most studies report complement activation after overt neurodegeneration, making it difficult to conclude if complement is directly activated by pathological -syn or secondarily following neurodegeneration. We used the rat -syn preformed fibril (PFF) mode, in vitro complement assays and human postmortem PD tissue to test whether pathological -syn directly activates complement prior to overt neurodegeneration. The -syn PFF model exhibits a protracted pathological time course and distinct temporal separation between peak -syn aggregation and nigrostriatal degeneration; thus we quantified complement expression, activation, and regulation during the aggregation phase. Synucleinopathy induced complement activation prior to nigrostriatal degeneration, including upregulation of components of both the classical (C1qa, C1r, C4b) and alternative (Cfd, Cfb) pathways, the anaphylatoxin (C3aR, C5aR) and phagocytic (CR3) complement receptors, and activation of complement C3. During early synucleinopathy, microglia upregulated C3 which significantly correlated with synucleinopathy burden across several brain regions, including the substantia nigra pars compacta (SNc) and cortex. Concurrently, complement regulatory proteins, including CD55, CD59, neuronal pentraxin-1 (Nptx1), and the neuronal pentraxin receptor were downregulated in the synucleinopathy-affected SNc. Importantly, increased levels of C1q and iC3b along with downregulation of CD55 and NPTX1 were also observed in human postmortem PD SNc, supporting the translational relevance of our findings. Mechanistically, we demonstrate that aggregated, but not monomeric, -syn directly binds C1q and activates the complement cascade in a C1q-dpendent manner. These data provide the first in vivo evidence that synucleinopathy triggers complement activation and dysregulation prior to neurodegeneration.

neuroscience↗

bioRxiv2023

Nanobiopsy investigation of the subcellular mtDNA heteroplasmy in human tissues

Mitochondrial function is critical to continued cellular vitality and is an important contributor to a growing number of human diseases. Mitochondrial dysfunction is typically heterogeneous, mediated through the clonal expansion of mitochondrial DNA (mtDNA) variants in a subset of cells in a given tissue. To date, our understanding of the dynamics of clonal expansion of mtDNA variants has been technically limited to the single cell-level. Here, we report the use of nanobiopsy for subcellular sampling from human tissue, combined with next-generation sequencing to assess subcellular mtDNA mutation load in human tissue from mitochondrial disease patients. The ability to map mitochondrial mutation loads within individual cells of diseased tissue samples will further our understanding of mitochondrial genetic diseases.

genomics↗

bioRxiv2023

Incorporation of TRDV segments into TCR alpha chains

In human tumor models we tried to identify and clone the TCR of tumor-reactive T cells enriched in mixed lymphocyte tumor-cell cultures (MLTC). In a particular MLTC, we identified a predominant TCR beta chain using the Beta Mark Vbeta Kit, but a corresponding alpha chain could not be amplified via RT-PCR using TRAV-specific forward primers. Therefore, we applied 5RACE to obtain the TCR alpha chain sequences. The 5RACE product revealed an alpha chain that encompassed 89bp of the TRDV1 5UTR, followed by the TRDV1 coding sequence joined in frame to TRAJ24. The ORF reaching from the TRDV1 start codon to the TRAC segment was intact, suggesting a functional TCR. To analyze this MLTC population in greater depth we conducted 10X VDJ sequencing. CellRanger identified the beta chain known from the Beta Mark analysis, but no corresponding alpha chain in the filtered results. The corresponding TRDV-containing TCR alpha chain could, however, be detected in the "all_contig_annotations" files. In a separate project, we performed TCR sequencing of tumor-infiltrating lymphocytes (TILs) in a murine tumor model. Also here, a predominant clonotype contained a TCR alpha chain joining Trdv2-2 in frame to Traj49. Transfection of both TCR cDNAs resulted in cell surface localization of TCR and CD3 as validated by FACS. Tumor recognition of the human, TRDV1-containing TCR could be demonstrated by IFNgamma ELISpot whereas the murine TCR did not recognize a tumor-derived cell line. TRDV-containing alpha chains have been reported in the literature for two HLA I-restricted TCRs against HIV peptides (Ueno et al, Eur J Immunol, 2003). To determine whether such TDRV-containing TCRs are unique events or whether Vdelta segments are commonly incorporated into TCR alpha chains, we queried the NCBI Sequence Read Archive (SRA) for 10X VDJ data and analyzed 21 human and 23 murine datasets. We found that especially TRDV1, Trdv1 and to some extent Trdv2-2 are more commonly incorporated into TCR alpha chains than some TRAV genes, making the TRDV segments a relevant contribution to TCR alpha diversity. For apparently solitary beta chains in 10X VDJ datasets, we suggest to scrutinize the "all_contig_annotations" files as these may contain an accompanying, TRDV-containing alpha chain.

immunology↗

bioRxiv2021

A subcellular cookie cutter for spatial genomics in human tissue

Intracellular heterogeneity contributes significantly to cellular physiology and, in a number of debilitating diseases, cellular pathophysiology. This is greatly influenced by distinct organelle populations and to understand the aetiology of disease it is important to have tools able to isolate and differentially analyse organelles from precise location within tissues. Here we report the development of a subcellular biopsy technology that facilitates the isolation of organelles, such as mitochondria, from human tissue. We compared the subcellular biopsy technology to laser capture microdissection (LCM) that is the state of art technique for the isolation of cells from their surrounding tissues. We demonstrate an operational limit of (>20m) for LCM and then, for the first time in human tissue, show that subcellular biopsy can be used to isolate mitochondria beyond this limit.

genomics↗