Biology subjects

Tzoulis, C.

Publications and source records attributed to Tzoulis, C..

11 recordsLinked to original sources

bioRxiv2026

Selective nigrosome vulnerability in Parkinson's disease revealed by histology-anchored ultra-high-field quantitative MRI

Selective vulnerability is a hallmark of Parkinson's disease (PD): post-mortem, dopamine-containing cell loss in the substantia nigra first and most severely affects calbindin-poor compartments termed nigrosomes. Testing this in vivo has been limited by circularity, as MRI studies have delineated nigrosomes from the same disease-related contrast they then measure. Here, we warp histology-derived, calbindin-validated probabilistic masks of nigrosomes 1-5 into a cohort-specific multimodal 7 Tesla quantitative MRI template, providing a degeneration-independent anatomical scaffold, in 28 patients with PD and 28 age- and sex-matched controls. In controls, nigrosomes differ from the surrounding nigral matrix in magnetic susceptibility and R1. PD shifts nigrosomal values upward relative to the matrix in susceptibility, R2*, and R1, concentrated in nigrosomes 1, 2, and 4, whereas whole-nigra analysis shows no group difference. Within the matrix, disease effects are largest caudally and emerge dorsally from the ventral tier, and susceptibility alterations in nigrosomes 1 and 4 track contralateral motor symptoms asymmetry. The multimodal pattern suggests that the detectable signal reflects predominantly secondary, iron-associated glial processes rather than neuronal loss itself. These findings recover the post-mortem spatial pattern of nigral vulnerability in vivo and establish a degeneration-independent framework for subregional nigral imaging in PD.

neuroscience↗

bioRxiv2026

Neuroinflammation and metabolic dysfunction in POLG-related mitochondrial epilepsy

Super-refractory status epilepticus is a common neurological manifestation of mitochondrial disease caused by bi-allelic pathogenic variants in POLG. Epilepsy in POLG-related disease typically presents with an explosive onset of status epilepticus, often from an occipital focus, and is associated with extensive neurodegeneration. The neuropathological mechanisms underlying POLG-related mitochondrial epilepsy remain poorly understood, however, neuroinflammation and glial dysfunction are hypothesised to play a significant role. In this study, we performed a neuropathological and proteomic investigation of post-mortem brain tissues from 12 patients with POLG-related mitochondrial epilepsy (age range: 3 - 28 years) and matched control cases. Given that the primary visual cortex is prominently involved in this epileptic disorder, occipital cortical tissues (Brodmann area 17) were compared to frontal cortical tissues (Brodmann area 9). Liquid chromatography-mass spectrometry (LC-MS/MS) analysis identified a distinct immunometabolic signature in the occipital cortex, and to a lesser extent in the frontal cortex, in POLG-related epilepsy. This was characterised by decreased abundance of mitochondrial proteins coupled to an increased expression of innate immune and inflammatory proteins, consistent with neuroinflammation. To validate these observations, we confirmed an increased density of cells immunoreactive for acute phase proteins (C-reactive protein, osteopontin and serpin A3), immune co-receptors (CD14 and HLA-DR), the inflammatory glycoprotein YKL40, the cytokine TNF-alpha, and mitochondrial translocator protein (TSPO). We also demonstrate a decreased expression of mitochondrial oxidative phosphorylation (OXPHOS) subunits within POLG patient microglia, indicative of mitochondrial dysfunction. Finally, we show enrichment of mitochondrial OXPHOS and interneuron proteins in the control primary visual cortex compared with the frontal cortex, which may underlie the selective regional vulnerability observed in POLG-related mitochondrial disease. Overall, these findings provide strong neuropathological evidence implicating neuroinflammation and glial dysfunction in POLG-related epilepsy.

neuroscience↗

bioRxiv2024

Mitochondrial complex I deficiency occurs in skeletal muscle of a subgroup of individuals with Parkinson's disease

Widespread neuronal complex I (CI) deficiency was recently reported to be a characteristic in a subgroup of individuals with idiopathic Parkinsons disease (PD). Here, we sought to determine whether a CI deficient subgroup could be discerned using clinically accessible muscle biopsy. Vastus lateralis needle biopsies were collected from 83 individuals with PD and 29 neurologically healthy controls and analyzed by immunohistochemistry for complexes I and IV, cytochrome c oxidase/succinate dehydrogenase (COX/SDH) histochemistry, and spectrophotometric activity assays of complexes I-IV. Mitochondrial DNA (mtDNA) copy number, deletions, and point variation were analyzed in single muscle fibers and bulk biopsy samples. PD muscle exhibited reduced CI activity at the group level, with 9% of cases falling below two standard deviations of the control group. This deficiency was not associated with mtDNA abnormalities. Our findings support the existence of a PD subpopulation characterized by CI pathology and suggest that stratification by extra-neural mitochondrial dysfunction may be informative for selecting individuals for clinical trials.

neuroscience↗

bioRxiv2023

Mitochondrial complex I deficiency stratifies idiopathic Parkinson's disease

Idiopathic Parkinsons disease (iPD) is believed to have a heterogeneous pathophysiology, but molecular disease subtypes have not been identified. Here, we show that iPD can be stratified according to the severity of neuronal respiratory complex I (CI) deficiency, and identify two emerging disease subtypes with distinct molecular and clinical profiles. The CI deficient (CI-PD) subtype accounts for approximately a fourth of all cases, and is characterized by anatomically widespread neuronal CI deficiency, a distinct cell type-specific gene expression profile, increased load of neuronal mtDNA deletions, and a predilection for non-tremor dominant motor phenotypes. In contrast, the non-CI deficient (nCI-PD) subtype exhibits no evidence of mitochondrial impairment outside the dopaminergic substantia nigra and has a predilection for a tremor dominant phenotype. These findings constitute a step towards resolving the biological heterogeneity of iPD with implications for both mechanistic understanding and treatment strategies.

neuroscience↗

bioRxiv2023

Hallmark molecular and pathological features of POLG disease are recapitulated in cerebral organoids

In our research, we developed a 3D brain organoid model to study POLG-related encephalopathy, a mitochondrial disease stemming from POLG gene mutations. We utilized induced pluripotent stem cells (iPSCs) derived from patients with these mutations to generate cortical organoids, which exhibited typical POLG disease features, such as altered morphology, neuronal loss, and mtDNA depletion. We also identified significant dysregulation in pathways crucial for neuronal development and function, alongside upregulated NOTCH and JAK-STAT signaling pathways. Metformin treatment ameliorated many of these abnormalities, except for the persistent affliction of inhibitory DA GLU neurons. This novel model effectively mirrors both the molecular and pathological attributes of POLG disease, providing a valuable tool for mechanistic understanding and therapeutic screening for POLG-related disorders and other conditions characterized by compromised neuronal mtDNA maintenance and complex I deficiency. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/558087v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1759de3org.highwire.dtl.DTLVardef@d0a3dborg.highwire.dtl.DTLVardef@1b58eaborg.highwire.dtl.DTLVardef@a986a8_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIWe have successfully developed cortical organoid model that represents POLG-related disease. C_LIO_LIThis model effectively replicates both histological and molecular signatures seen in the brains of patients. C_LIO_LIThe cortical organoid model displays a range of features common in POLG-related disease, including neurodegeneration, mtDNA depletion, and neuronal complex I deficiency. C_LIO_LIThe use of metformin supplementation in this model improved mitochondria protein and reduced cell death. C_LI

cell biology↗

bioRxiv2023

The NAD+ Precursor Nicotinamide Riboside Rescues Mitochondrial Defects and Neuronal Loss in iPSC derived Cortical Organoid of Alpers' Disease

Alpers syndrome is an early-onset neurodegenerative disorder usually caused by biallelic pathogenic variants in the gene encoding the catalytic subunit of polymerase-gamma (POLG), which is essential for mitochondrial DNA (mtDNA) replication. The disease is progressive, incurable, and inevitably it leads to death from drug-resistant status epilepticus. The neurological features of Alpers syndrome are intractable epilepsy and developmental regression, with no effective treatment; the underlying mechanisms are still elusive, partially due to lack of good experimental models. Here, we generated the patient-derived induced pluripotent stem cells from one Alpers patient carrying the compound heterozygous mutations of A467T (c.1399G>A) and P589L (c.1766C>T), and further differentiated them into cortical organoids and neural stem cells (NSCs) for mechanistic studies of neural dysfunction in Alpers syndrome. Patient cortical organoids exhibited a phenotype that faithfully replicated the molecular changes found in patient postmortem brain tissue, as evidenced by cortical neuronal loss and depletion of mtDNA and complex I (CI). Patient NSCs showed mitochondrial dysfunction leading to ROS overproduction and downregulation of the NADH pathway. More importantly, the NAD+ precursor nicotinamide riboside (NR) significantly ameliorated mitochondrial defects in patient brain organoids. Our findings demonstrate that the iPSC model and brain organoids are good in vitro models of Alpers disease; this first-in-its-kind stem cell platform for Alpers syndrome enables therapeutic exploration and has identified NR as a viable drug candidate for Alpers disease and, potentially, other mitochondrial diseases with similar causes.

neuroscience↗

bioRxiv2021

Distinct mitochondrial remodeling during early cardiomyocyte development in a human-based stem cell model

Given the considerable interest in using stem cells for modelling and treating disease, it is essential to understand what regulates self-renewal and differentiation. Remodeling of mitochondria and metabolism, with the shift from glycolysis to oxidative phosphorylation (OXPHOS), play a fundamental role in maintaining pluripotency and stem cell fate. It has been suggested that metabolic switch from glycolysis to OXPHOS is germ-layer specific as during early ectoderm commitment, glycolysis remains active while during the transition to mesoderm and endoderm lineages, it is downregulated. How mitochondria adapt during these metabolic changes and whether mitochondria remodeling is tissue specific remains unclear. Here we address the question of mitochondrial adaption by examining the differentiation of human pluripotent stem cells to cardiac progenitors and further to functional cardiomyocytes. Contrary to recent findings in neuronal differentiation, we found that mitochondrial content decreases continuously during mesoderm differentiation, despite clear mitochondrial remodeling giving increased mitochondrial activity and higher levels of ATP-linked respiration. Thus, our work both highlights similarities in mitochondrial remodeling during the transition from pluripotent to multipotent state in ectodermal and mesodermal lineages, while at the same time demonstrating cell-lineage-specific adaptions upon further differentiation. Our results improve understanding of how mitochondria remodeling and the metabolism interact during differentiation and show that it is erroneous to assume that increased OXPHOS activity during differentiation requires a simultaneous expansion of mitochondrial content. Summary statementWe found that mitochondrial content decreases continuously during mesoderm differentiation, despite clear mitochondrial remodeling giving increased mitochondrial activity and higher levels of ATP-linked respiration during mesoderm differentiation.

cell biology↗

bioRxiv2021

Ultra-deep whole genome bisulfite sequencing reveals a single methylation hotspot in human brain mitochondrial DNA

While DNA methylation is established as a major regulator of gene expression in the nucleus, the existence of mitochondrial DNA (mtDNA) methylation remains controversial. Here, we characterised the mtDNA methylation landscape in the prefrontal cortex of neurological healthy individuals (n=26) and patients with Parkinsons disease (n=27), using a combination of whole genome bisulfite sequencing (WGBS) and bisulfite-independent methods. Accurate mtDNA mapping from WGBS data required alignment to an mtDNA reference only, to avoid misalignment to nuclear mitochondrial pseudogenes. Once correctly aligned, WGBS data provided ultra-deep mtDNA coverage (16,723{+/-}7,711), and revealed overall very low levels of cytosine methylation. The highest methylation levels (5.49{+/-}0.97%) were found on CpG position m.545, located in the heavy-strand promoter 1 region. The m.545 methylation was validated using a combination of methylation-sensitive DNA digestion and quantitative PCR analysis. We detected no association between mtDNA methylation profile and Parkinsons disease. Interestingly, m.545 methylation correlated with the levels of mtDNA transcripts, suggesting a putative role in regulating mtDNA gene expression. In addition, we propose a robust framework for methylation analysis of mtDNA from WGBS data, which is less prone to false-positive findings due to misalignment of nuclear mitochondrial pseudogene sequences. Graphical abstract of the analyses and main findingsFresh-frozen brain tissue was obtained from the prefrontal cortex (Brodmann area 9) of 53 individuals, comprising 27 patients with idiopathic PD and 26 healthy controls. Tissue from the same samples was used in three different downstream analyses. WGBS was conducted on all 53 samples and the data were analysed using three different alignment strategies. Alignment against an mtDNA reference only was clearly superior as it gave the highest and most even depth of coverage. WGBS analysis revealed that mtDNA harbours very low levels of cytosine methylation, with the exception of the CpG position m.545 within the HSP1 region (lower right inset). The m.545 methylation was confirmed by bisulfite- and sequencing-independent methods, employing methylation-specific MspJI DNA digestion, followed by quantification with qPCR or fluorescent PCR and capillary electrophoresis. Finally, mtDNA transcript levels were determined by RT-qPCR and correlated to m.545 methylation levels, showing a positive association. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=196 SRC="FIGDIR/small/437685v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1d478e3org.highwire.dtl.DTLVardef@4cbf9forg.highwire.dtl.DTLVardef@b863faorg.highwire.dtl.DTLVardef@1abe6c5_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2020

Stem cell derived astrocytes with POLG mutations and mitochondrial dysfunction including abnormal NAD+ metabolism is toxic for neurons

The inability to reliably replicate mitochondrial DNA (mtDNA) by mitochondrial DNA polymerase gamma (POLG) leads to a subset of common mitochondrial diseases associated with neuronal death and depletion of neuronal mtDNA. Defining disease mechanisms remains difficult due to the limited access to human tissue. Astrocytes are highly abundant in the brain, playing a crucial role in the support and modulation of neuronal function. Astrocytes also respond to insults affecting the brain. Following damage to the center neural system, which can be hypoxia, inflammation or neurodegeneration, astrocytes become activated and lose their supportive role and gain toxic functions that induce rapid death of neurons and oligodendrocytes. The role of astrocyte reactivation and the consequences this has for neuronal homeostasis in mitochondrial diseases has not been explored. Here, using patient cells carrying POLG mutations, we generated iPSCs and then differentiated into astrocytes. We demonstrated that POLG-astrocytes exhibited both mitochondrial dysfunctions, including loss of mitochondrial membrane potential, energy failure, complex I and IV defects, disturbed NAD+/NADH metabolism, and mtDNA depletion. Further, POLG derived astrocytes presented an A1-like reactive phenotype with increased proliferation, invasion, upregulation of pathways involved in response to stimulus, immune system process, cell proliferation and cell killing. Under direct and indirect co-culture with neurons, POLG-astrocytes exhibited a toxic effect leading to the death of neurons. Our findings demonstrate that mitochondrial dysfunction caused by POLG mutations leads not only to intrinsic defects in energy metabolism affecting both neurons and astrocytes, but also to neurotoxic damage driven by astrocytes. Our studies provide a robust astroglia-neuronal interaction model for future investigation of mitochondrial involvement in neurogenesis and neurodegenerative diseases. Highlights{blacksquare} Patient-specific astrocytes harbouring a POLG mutation showed lower mitochondrial membrane potential and mtDNA depletion. {blacksquare}POLG-astrocytes generated elevated L-lactate as the end glycolytic product. {blacksquare}Patient-specific astrocytes with POLG mutations exhibited mitochondrial respiratory chain disruption accompanied with abnormal UCP2/SirT1/SirT3 mediated NAD+ metabolism. {blacksquare}Suppressed complex I and IV-driven respiration contributed to the pathological mechanisms in POLG-related disease. {blacksquare}POLG-astrocytes exhibit A1-reactive phenotype and neurotoxic potential. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=166 SRC="FIGDIR/small/423652v1_ufig1.gif" ALT="Figure 1"> View larger version (61K): org.highwire.dtl.DTLVardef@18995fdorg.highwire.dtl.DTLVardef@1d615d4org.highwire.dtl.DTLVardef@334aa8org.highwire.dtl.DTLVardef@16a84ba_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2019

Dysregulation of histone acetylation and decoupling from transcription in Parkinson disease

Parkinson disease (PD) is a complex neurodegenerative disorder of largely unknown etiology. While several genetic risk factors have been identified, the involvement of epigenetics in the pathophysiology of PD is mostly unaccounted for. We conducted a histone acetylome-wide association study in PD, using brain tissue from two independent cohorts of cases and controls. Immunoblotting revealed increased acetylation at several histone sites in PD, with the most prominent change observed for H3K27, a marker of active promoters and enhancers. Chromatin immunoprecipitation sequencing (ChIP-seq) further indicated that H3K27 hyperacetylation in the PD brain is a genome-wide phenomenon, with a strong predilection for genes implicated in the disease, including SNCA, PARK7, PRKN and MAPT. Integration of the ChIP-seq with transcriptomic data revealed that the correlation between promoter H3K27 acetylation and gene expression is attenuated in PD patients, suggesting that H3K27 acetylation may be decoupled from transcription in the PD brain. Our findings strongly suggest that dysregulation of histone acetylation plays an important role in the pathophysiology of PD and identify novel epigenetic signatures associated with the disease.

genomics↗

bioRxiv2019

Common gene expression signatures in Parkinson’s disease are driven by changes in cell composition

BackgroundThe etiology of Parkinsons disease (PD) is largely unknown. Genome-wide transcriptomic studies in bulk brain tissue have identified several molecular signatures associated with the disease. While these studies have the potential to shed light into the pathogenesis of PD, they are also limited by two major confounders: RNA post mortem degradation and heterogeneous cell type composition of bulk tissue samples. We performed RNA sequencing following ribosomal RNA depletion in the prefrontal cortex of 49 individuals from two independent case-control cohorts. Using cell-type specific markers, we estimated the cell-type composition for each sample and included this in our analysis models to compensate for the variation in cell-type proportions.\n\nResultsRibosomal RNA depletion results in substantially more even transcript coverage, compared to poly(A) capture, in post mortem tissue. Moreover, we show that cell-type composition is a major confounder of differential gene expression analysis in the PD brain. Correcting for cell-type proportions attenuates numerous transcriptomic signatures that have been previously associated with PD, including vesicle trafficking, synaptic transmission, immune and mitochondrial function. Conversely, pathways related to endoplasmic reticulum, lipid oxidation and unfolded protein response are strengthened and surface as the top differential gene expression signatures in the PD prefrontal cortex.\n\nConclusionsDifferential gene expression signatures in PD bulk brain tissue are significantly confounded by underlying differences in cell-type composition. Modeling cell-type heterogeneity is crucial in order to unveil transcriptomic signatures that represent regulatory changes in the PD brain and are, therefore, more likely to be associated with underlying disease mechanisms.

neuroscience↗