Biology subjects

Filatova, A.

Publications and source records attributed to Filatova, A..

3 recordsLinked to original sources

bioRxiv2025

Chromatin remodelling subunit SMARCB1 is implicated in dendrite development and complex brain functions

SMARCB1 encodes a core component of the BAF chromatin remodelling complex and pathogenic variants in this gene are associated with neurodevelopmental disorders such as Coffin-Siris syndrome. The relationship between altered SMARCB1 protein products and severe functional brain changes in Coffin-Siris syndrome remains largely unknown. We performed cellular, molecular, and behavioural analyses of a Coffin-Siris syndrome mouse model with a heterozygous nervous system-specific Smarcb1 mutation. In addition, we evaluated general cognitive abilities, as well as cognitive and behavioural functioning, in individuals with SMARCB1-related Coffin-Siris syndrome. Smarcb1 mutant mice exhibited deficits in fine motor coordination and balance, as well as impaired spatial learning and memory. Furthermore, these mice showed anxiety-like behaviours and agitation when exposed to novel environments. The detected behavioural abnormalities could indicate impaired decision-making, which results in impaired risk assessment. Comparable cognitive and behavioural deviations were identified in individuals with Coffin-Siris syndrome and SMARCB1 pathogenic variants. Our analysis of the Smarcb1 mouse model revealed structural alterations in the brain, including decreased dendritic length and complexity of dendritic trees. These alterations may explain the observed functional impairments. Notably, our finding of reduced Wasl transcripts in mutant Purkinje cell nuclei suggests that dysregulation of actin polymerization may be involved in the discovered dendritic defects. Taken together, we demonstrate a link between the chromatin remodelling complex component SMARCB1, complex brain functions, neuronal structure, and a key regulator of actin branching.

neuroscience↗

bioRxiv2025

Self-organized vascularized human liver spheroids: Serum-free culture conditions and use as tissue building blocks

Engineering vascularized human liver tissue for in vitro models and in vivo applications remains a major challenge. Here, we describe a scalable approach to generate human liver spheroids with self-organized, lumen-containing vascular networks and demonstrate their use as building blocks for fabricating vascularized tissue layers. Spheroids were formed from HepaRG liver cells, human umbilical vein endothelial cells (HUVECs), and adipose tissue-derived mesenchymal stem cells (MSCs). The inclusion of MSCs prevented spatial segregation of hepatic and endothelial compartments and enabled endothelial network formation. We present two media that are suitable for culturing these spheroids: a serum-reduced medium and a defined serum-free medium containing GibcoTM KnockOut serum replacement. These media supported the long-term maintenance of hepatocytes in a metabolically active, relatively mature state, as well as the persistence of endothelial networks. Endothelial cell identity and organization were confirmed by VE-cadherin and ICAM-2 immunostaining and by transmission electron microscopy, which revealed adherens junctions and luminal morphologies consistent with a capillary-like organization. Spheroid-derived HUVECs established anastomoses with external endothelial channels in microfluidic devices. Moreover, endothelial sprouts emerging from the spheroids formed inter-spheroid connections within permissive hydrogels (fibrin or collagen-methylcellulose), a process that depended on the inter-spheroid distance. Finally, we demonstrate the fabrication of planar tissue layers with vascularly interconnected spheroids. Together, we identify key conditions, including cellular ratios, medium formulations, biomaterials, and spatial design criteria that enable the generation and assembly of vascularized liver spheroids as scalable tissue building blocks for tissue engineering applications. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=187 SRC="FIGDIR/small/684548v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@1b8c8a2org.highwire.dtl.DTLVardef@7c0ed2org.highwire.dtl.DTLVardef@b10886org.highwire.dtl.DTLVardef@a6ab4f_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2024

Revision of splicing variants in the DMD gene

BackgroundPathogenic variants in the dystrophin (DMD) gene lead to X-linked recessive Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). Nucleotide variants that affect splicing are a known cause of hereditary diseases. However, their representation in the public genomic variation databases is limited due to the low accuracy of their interpretation, especially if they are located within exons. The analysis of splicing variants in the DMD gene is essential both for understanding the underlying molecular mechanisms of the dystrophinopathies pathogenesis and selecting suitable therapies for patients. ResultsUsing deep in silico mutagenesis of the entire DMD gene sequence and subsequent SpliceAI splicing predictions, we identified 7,948 DMD single nucleotide variants that could potentially affect splicing, 863 of them were located in exons. Next, we analyzed over 1,300 disease-associated DMD SNVs previously reported in the literature (373 exonic and 956 intronic) and intersected them with SpliceAI predictions. We predicted that [~]95% of the intronic and [~]10% of the exonic reported variants could actually affect splicing. Interestingly, the majority (75%) of patient-derived intronic variants were located in the AG-GT terminal dinucleotides of the introns, while these positions accounted for only 13% of all intronic variants predicted in silico. Of the 97 potentially spliceogenic exonic variants previously reported in patients with dystrophinopathy, we selected 38 for experimental validation. For this, we developed and tested a minigene expression system encompassing 27 DMD exons. The results showed that 35 (19 missense, 9 synonymous, and 7 nonsense) of the 38 DMD exonic variants tested actually disrupted splicing. We compared the observed consequences of splicing changes between variants leading to severe Duchenne and milder Becker muscular dystrophy and showed a significant difference in their distribution. This finding provides extended insights into relations between molecular consequences of splicing variants and the clinical features. ConclusionsOur comprehensive bioinformatics analysis, combined with experimental validation, improves the interpretation of splicing variants in the DMD gene. The new insights into the molecular mechanisms of pathogenicity of exonic single nucleotide variants contribute to a better understanding of the clinical features observed in patients with Duchenne and Becker muscular dystrophy.

genetics↗