Kozicz, Tamas

Abstract Oxidative phosphorylation (OXPHOS) is a central function and a key indicator of mitochondrial fitness, yet studies in human tissue remain limited. Inclusion body myositis (IBM) is a progressive myopathy that lies at the intersection of aging, inflammation and mitochondrial dysfunction. We aimed to perform a comprehensive profiling of mitochondrial respiration in muscle tissue from patients with IBM. A wide battery of complementary approaches from RNA level to high-resolution respirometry on permeabilized muscle fibers was employed. The relationship between mitochondrial respiration, mitochondrial content, mitochondrial DNA (mtDNA) abnormalities and mitophagy was examined, along with the correlation with various clinical parameters to determine their clinical relevance. The study included a total of 67 patients with IBM and 45 controls. On high resolution respirometry of permeabilized muscle fibers, IBM samples exhibited reduced maximal mitochondrial respiration per tissue weight in State 3 (high substrates, high ADP) and uncoupled state with decreased coupling efficiency and higher leak control ratios. When adjusting for citrate synthase reflecting mitochondrial content, male patients had decreased State 3 intrinsic respiration, whereas female patients had greater intrinsic respiration under leak states. Complex I activity was decreased mainly in female patients, in whom complex II control ratio positively correlated with disease duration and severity. IBM was further associated with decreased RNA levels of all complexes, and lower protein expression of complex I, III, IV and V, likely related to the lower mtDNA content seen in IBM samples. Regarding the production of reactive oxygen species, IBM samples exhibited lower maximal H2O2 emission, accompanied by a higher total antioxidant capacity that positively correlated with disease duration in female patients. Lastly, correlation analyses suggested that impaired mitochondrial respiration, altered mitophagy, and reduced mtDNA content are interconnected in IBM and maybe of clinical significance. IBM is characterized by multifaceted, clinically relevant impairments in mitochondrial respiration. Future studies should further explore underlying pathomechanisms and the variation of mitochondrial respiration by disease stage.

Abstract Background Mono-allelic Dehydrodolichyl Diphosphate Synthase ( DHDDS) variants are associated with juvenile Parkinsonism, developmental delay and seizures. Symptoms are progressive, and various mechanisms, such as defective glycosylation, lysosomal dysfunction and cholesterol accumulation have been hypothesized to underlie disease symptoms. There is no treatment for DHDDS-related disease. Methods Patient-derived cortical forebrain organoids were created to elucidate disease mechanisms and evaluate potential treatments. In these neuronal models, glycosylation, lipidomics, proteomics, cholesterol/ganglioside accumulation, mitochondrial function and electrophysiological activity were assessed. Finally, we investigated the effects of nicotinamide mononucleotide (NMN), identified through a yeast-based drug screen, in neuronal cell models and in six patients in an off-label, N-of-1, observational series. Results DHDDS-patient derived organoids showed visual signs of degeneration after four months of culturing. This was accompanied by significant cholesterol accumulation in astrocytes, decreased mitochondrial respiration and loss of deep-layer neurons. In addition, we identified glycosylation abnormalities, showing for the first time that glycosylation in human tissue is affected by monoallelic DHDDS variants. Proteomic analysis revealed altered protein expression of proteins involved in lipid metabolism, cytoskeletal organization and neuronal development. We found that oral Nicotinamide Mononucleotide supplementation led to significant improvement in mitochondrial respiration and electrophysiological parameters in organoids, concurring with clinical improvements in all of the treated patients, particularly regarding their ataxia and tremor. Conclusion Our findings reveal a progressive phenotype in DHDDS-patient-derived brain organoids, with mitochondrial dysfunction and astrocyte-specific metabolic alterations contributing to disease pathology. Notably, NMN treatment led to clinical improvements in patients with heterozygous DHDDS variants, highlighting its potential as a therapeutic strategy.

ABSTRACT Polyprenal reductase is an enzyme encoded by the SRD5A3 gene, which is involved in the synthesis of dolichol from polyprenol. Dolichol serves as a carrier for glycan precursors or monosaccharides in N‐linked glycosylation. Pathogenic variants in SRD5A3 can result in a congenital disorder of glycosylation (CDG), SRD5A3‐CDG, which is inherited in an autosomal recessive manner. Most plasma proteins are glycosylated and changes in the glycosylation of several glycoproteins are associated with pathological consequences. Despite the critical role of SRD5A3 in glycosylation, the impact of its deficiency on the glycosylation of serum proteins remains largely unexplored. In this study, we used tandem mass tag‐based multiplexed quantitative approach to analyze serum N‐glycoproteomics and proteomics in SRD5A3‐CDG patients and controls. We quantified 2200 serum N‐glycopeptides from 359 N‐glycosites from 204 serum proteins. Extensive hypoglycosylation of serum proteins was observed in patients, with 245 of 291 altered glycopeptides decreased in SRD5A3‐CDG. Altered glycopeptides included those derived from haptoglobin, plasma serine protease inhibitor, alpha‐1‐B glycoprotein, alpha‐2‐macroglobulin, and ceruloplasmin. Some of these proteins have previously been reported to be associated with liver dysfunction, anemia, and coagulopathy, which could underlie similar clinical features observed in SRD5A3‐CDG patients. Overall, our study provides novel insights into alterations in the glycosylation status of specific serum proteins in SRD5A3‐CDG. Some of these alterations could be further pursued to develop glycopeptide‐based biomarkers as the current diagnosis of SRD5A3‐CDG by screening assays remains challenging. In addition, knowledge of altered glycoproteins could enhance our understanding of the disease spectrum and potentially unveil additional therapeutic avenues.

Background: ALG13-CDG is an X-linked N-linked glycosylation disorder caused by pathogenic variants in the glycosyltransferase ALG13, leading to severe neurological manifestations. Despite the clear CNS involvement, the impact of ALG13 dysfunction on human brain glycosylation and neurodevelopment remains unknown. We hypothesize that ALG13-CDG causes brain-specific hypoglycosylation that disrupts neurodevelopmental pathways and contributes directly to cortical network dysfunction. Methods: We generated iPSC-derived human cortical organoids (hCOs) from individuals with ALG13-CDG to define the impact of hypoglycosylation on cortical development and function. Electrophysiological activity was assessed using MEA recordings and integrated with multiomic profiling, including scRNA-seq, proteomics, glycoproteomics, N-glycan imaging, lipidomics, and metabolomics. X-inactivation status was evaluated in both iPSCs and hCOs. Results: ALG13-CDG hCOs showed reduced glycosylation of proteins involved in ECM organization, neuronal migration, lipid metabolism, calcium homeostasis, and neuronal excitability. These pathway disruptions were supported by proteomic and scRNA-seq data and included altered intercellular communication. Trajectory analyses revealed mistimed neuronal maturation with early inhibitory and delayed excitatory development, indicating an E/I imbalance. MEA recordings demonstrated early network hypoactivity with reduced firing rates, immature burst structure, and shortened axonal projections, while transcriptomic and proteomic signatures suggested emerging hyperexcitability. Altered lipid and GlcNAc metabolism, along with skewed X-inactivation, were also observed. Conclusions: Our study reveals that ALG13-CDG is a disorder of brain-specific hypoglycosylation that disrupts key neurodevelopmental pathways and destabilizes cortical network function. Through integrated multiomic and functional analyses, we identify early network hypoactivity, mistimed neuronal maturation, and evolving E/I imbalance that progresses to compensatory hyperexcitability, providing a mechanistic basis for seizure vulnerability. These findings redefine ALG13-CDG as disorders of cortical network instability, offering a new framework for targeted therapeutic intervention.

Abstract Mono‐allelic DHDDS variants are associated with seizures, intellectual disability, and movement disorders. The age of onset and progression rates of symptoms vary greatly among patients, spanning from infancy to late adulthood. Yet, the reasons behind this clinical variability and the underlying pathophysiological mechanisms of the disease have remained elusive. We investigated the age of onset and the progression of symptoms over time in 59 patients with heterozygous DHDDS variants, drawing from medical literature and incorporating five previously unreported cases from the FCDGC Natural History Study. Clinical symptoms typically emerged early in life. Ataxia, tremor, dystonia, and dyskinesia manifested slightly later in childhood. Global developmental delay usually presented as the initial symptom. We observed diverse rates of symptom accumulation over time: some patients exhibited the full spectrum of symptoms in early childhood, while others developed novel symptoms well into adulthood. Interestingly, neither the sex nor the underlying DHDDS variants correlated with the age of symptom onset or specific clinical symptoms. Additionally, we found that 19% of patients presented with autism spectrum disorder. This study offers insight into the age of symptom onset and the rate of symptom accumulation in patients with DHDDS variants. We found no correlation between the age of onset and progression of clinical symptoms with specific DHDDS variants or patient sex. Autism spectrum disorder is common in patients and warrants attention in clinical management.

Given its polygenic nature, there is a need for a personalized approach to schizophrenia. The aim of the study was to select laboratory biomarkers from blood, brain imaging, and clinical assessment, with an emphasis on patients’ self-report questionnaires. Metabolomics studies of serum samples from 51 patients and 45 healthy volunteers, based on the liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS/MS), led to the identification of 3 biochemical indicators (cortisol, glutamate, lactate) of schizophrenia. These metabolites were sequentially correlated with laboratory tests results, imaging results, and clinical assessment outcomes, including patient self-report outcomes. The hierarchical cluster analysis on the principal components (HCPC) was performed to identify the most homogeneous clinical groups. Significant correlations were noted between blood lactates and 11 clinical and 10 neuroimaging parameters. The increase in lactate and cortisol were significantly associated with a decrease in immunological parameters, especially with the level of reactive lymphocytes. The strongest correlations with the level of blood lactate and cortisol were demonstrated by brain glutamate, N-acetylaspartate and the concentrations of glutamate and glutamine, creatine and phosphocreatine in the prefrontal cortex. Metabolomics studies and the search for associations with brain parameters and self-reported outcomes may provide new diagnostic evidence to specific schizophrenia phenotypes.

Congenital disorders of glycosylation (CDG) and mitochondrial disorders are multisystem disorders with overlapping symptomatology. Pathogenic variants in the PMM2 gene lead to abnormal N-linked glycosylation. This disruption in glycosylation can induce endoplasmic reticulum stress, contributing to the disease pathology. Although impaired mitochondrial dysfunction has been reported in some CDG, cellular bioenergetics has never been evaluated in detail in PMM2-CDG. This prompted us to evaluate mitochondrial function and autophagy/mitophagy in vitro in PMM2 patient-derived fibroblast lines of differing genotypes from our natural history study. We found secondary mitochondrial dysfunction in PMM2-CDG. This dysfunction was evidenced by decreased mitochondrial maximal and ATP-linked respiration, as well as decreased complex I function of the mitochondrial electron transport chain. Our study also revealed altered autophagy in PMM2-CDG patient-derived fibroblast lines. This was marked by an increased abundance of the autophagosome marker LC3-II. Additionally, changes in the abundance and glycosylation of proteins in the autophagy and mitophagy pathways further indicated dysregulation of these cellular processes. Interestingly, serum sorbitol levels (a biomarker of disease severity) and the CDG severity score showed an inverse correlation with the abundance of the autophagosome marker LC3-II. This suggests that autophagy may act as a modulator of biochemical and clinical markers of disease severity in PMM2-CDG. Overall, our research sheds light on the complex interplay between glycosylation, mitochondrial function, and autophagy/mitophagy in PMM2-CDG. Manipulating mitochondrial dysfunction and alterations in autophagy/mitophagy pathways could offer therapeutic benefits when combined with existing treatments for PMM2-CDG.

The altered cerebral energy metabolism central to schizophrenia can be linked to lactate accumulation. Lactic acid is produced by gastrointestinal bacteria, among others, and readily crosses the blood–brain barrier, leading to the brain acidity. This study aimed to examine the association of the oral microbiota with the effects of acid stress induced by an increase of brain lactate in schizophrenia patients. The study included patients with a diagnosis of acute polyphasic psychotic disorder meeting criteria for schizophrenia at 3-month follow-up. Results: Individuals with a significantly higher total score on the Positive and Negative Syndrome Scale had statistically significantly lower lactate concentrations compared to those with a lower total score and higher brain lactate. We observed a positive correlation between Actinomyces and lactate levels in the anterior cingulate cap and a negative correlation between bacteria associated with lactate metabolism and some clinical assessment scales. Conclusions: Shifts in the oral microbiota in favour of lactate-utilising bacterial genera may represent a compensatory mechanism in response to increased lactate production in the brain. Assessment of neuronal function mediated by ALA-LAC-dependent NMDA regulatory mechanisms may, thus, support new therapies for schizophrenia, for which acidosis has become a differentiating feature of individuals with schizophrenia endophenotypes.

Ischemic stroke accounts for over 80% of all strokes and is one of the leading causes of mortality and permanent disability worldwide. Intravenous administration of recombinant tissue plasminogen activator (rt-PA) is an approved treatment strategy for acute ischemic stroke of large arteries within 4.5 h of onset, and mechanical thrombectomy can be used for large arteries occlusion up to 24 h after onset. Improving diagnostic work up for acute treatment, reducing onset-to-needle time and urgent radiological access angiographic CT images (angioCT) and Magnetic Resonance Imaging (MRI) are real problems for many healthcare systems, which limits the number of patients with good prognosis in real world compared to the results of randomized controlled trials. The applied endovascular procedures demonstrated high efficacy, but some cellular mechanisms, following reperfusion, are still unknown. Changes in the morphology and function of mitochondria associated with reperfusion and ischemia-reperfusion neuronal death are still understudied research fields. Moreover, future research is needed to elucidate the relationship between continuously refined imaging techniques and the variable structure or physical properties of the clot along with vascular permeability and the pleiotropism of ischemic reperfusion lesions in the penumbra, in order to define targeted preventive procedures promoting long-term health benefits.