Biology subjects

Hinttala, R.

Publications and source records attributed to Hinttala, R..

7 recordsLinked to original sources

bioRxiv2026

A novel method for obtaining epileptic brain tissue for omic analyses using electrodes from a clinical stereoelectroencephalography study

Abstract Purpose: Developing a valid and practical method for obtaining brain tissue samples for transcriptomic and proteomic analyses using tissue adhered to stereoelectroencephalography (SEEG) electrodes, to enable investigation of the molecular mechanisms underlying chronic epilepsy. Method: Brain tissue samples adhered to SEEG electrodes were collected from six patients with drug-resistant epilepsy and preprocessed in a hospital environment after electrode removal. RNA extraction was initiated immediately, and proteomics samples were snap-frozen until subsequent analysis. The samples were categorized into three groups according to their electrophysiological profile: epileptogenic zone, propagation zone, and least-involved zone. The omic findings between these zones and anatomical brain areas were compared. Results: High-quality RNA and protein samples were obtained from tissue adhered to SEEG electrodes. Neuron- and brain-specific gene expression patterns and proteins were identified. Signs of activation of inflammatory mechanisms were most pronounced in the epileptogenic zone. Transcriptomic and proteomic findings demonstrated concordance. Conclusion: SEEG electrodes are a useful source for obtaining brain tissue for molecular characterization of chronic epilepsy. This method will enable the identification of shared and distinct molecular mechanisms in patients with varying etiologies of epilepsy.

neuroscience↗

bioRxiv2025

Stereospecific GPG acylation by CLN8 drives BMP biosynthesis and its loss leads to Batten disease

Loss-of-function mutations in the endoplasmic reticulum membrane protein CLN8 cause Batten disease, a neurodegenerative lysosomal storage disorder1. Together with the lysosomal enzyme CLN5, CLN8 mediates the biosynthesis of bis(monoacylglycero)phosphate (BMP), a phospholipid essential for lysosomal function and distinguished by its unique S,S stereochemistry2,3. However, the role of CLN8 in BMP synthesis has remained unclear. Here we establish that CLN8 is a glycerophosphoglycerol (GPG) acyltransferase that catalyses the stereospecific acylation of S,S-GPG to produce S,S-lysophosphatidylglycerol (LPG), the CLN5 substrate in BMP synthesis. Using cryo-electron microscopy, we resolve structures of the CLN8 homodimer in apo and substrate-bound states at 2.7 [A] resolution, revealing the active site architecture and a ping-pong acyl transfer mechanism. Batten disease-causing missense mutations impair CLN8 enzymatic activity in vitro and reduce BMP levels in a Cln8R24G knock- in mouse, whereas the Cln8mnd mouse frameshift mutation causes complete loss of BMP in vivo. Exogenous S,S-LPG, but not the R,S stereoisomer, restored BMP synthesis in CLN8- deficient cells and mice, and improved neurological phenotypes in cln8 mutant zebrafish. Together, these findings define the enzymatic function of CLN8, elucidate the biochemical basis of CLN8 Batten disease, and establish a proof-of-concept for treating it through stereospecific BMP precursor supplementation.

biochemistry↗

bioRxiv2025

Novel Cln8 p.R24G mouse line replicates major clinical features of Northern epilepsy

RationaleNorthern epilepsy belongs to a group of genetically diverse lysosomal storage diseases, the neuronal ceroid lipofuscinoses (NCLs). A characteristic feature of NCL pathology is the accumulation of autofluorescent ceroid lipofuscin in the central nervous system. Northern epilepsy is a late-infantile-onset disease. Patients develop normally until 5-10 years old, when they first present with general tonic-clonic seizures, followed by progressive cognitive impairment and a decline in motor skills. Northern epilepsy is caused by a missense variant in CLN8, causing a p.R24G amino acid substitution. CLN8 deficiency has been studied traditionally using motor neuron degeneration (mnd) mice, which carry a spontaneous frame shift variant in the murine orthologue Cln8, which is not known to exist in humans. MethodsWe have generated the first Cln8 p.R24G mouse model (Cln8R24G) using CRISPR/Cas9. Phenotyping analysis of the mice was conducted using behavioral and histopathological studies focusing on the brain and retina. ResultsAt birth, Cln8R24G KI mice were viable and asymptomatic. As the mice aged, a progressive accumulation of autofluorescent ceroid lipofuscin containing the mitochondrial ATP synthase subunit C was evident in different brain regions and retinal layers. Health monitoring revealed that mutant mice developed progressive but mild motor symptoms around 7 months of age. Spontaneous epileptic seizures, like those observed in Northern epilepsy patients, were detected and recorded. An increase in FosB staining intensity, reflecting neuronal hyperactivity, was observed in hippocampal CA1-CA3 pyramidal and dentate granule cells and correlated well with the intensity of seizure activity. Neuroinflammation was evident at 4 months and increased dramatically with age, mainly in the thalamic VPN/VPL nuclei and moderately in the cortex. Neurodegeneration was most prominent in the VPN/VPL thalamic nuclei. ConclusionsWe have generated the Cln8R24G mouse model that genocopies, for the first time, the pathogenic CLN8 variant present in patients with Northern epilepsy. These mice phenocopy major clinical features of the human disease, including mild motor impairment and, unlike the mnd mice, spontaneous generalized tonic-clonic seizures. We hypothesize that our mouse model will open new possibilities for developing and testing targeted treatment options for Northern epilepsy and, more broadly, for early-onset neurodegenerative disorders associated with epilepsy.

neuroscience↗

bioRxiv2024

GLE1 dysfunction compromises cellular homeostasis, spatial organization, and peripheral axon branching

The GLE1 protein is an enigmatic factor of RNA processing, associated with multiple developmental disorders including lethal congenital contracture syndrome 1 (LCCS1). Using in vivo genetic engineering to study disturbed GLE1 functions under physiological conditions we demonstrate that inactivation of Gle1 impedes cellular function and organization and causes pre-gastrulation lethality due to defects in adhesion and lineage specification. In contrast, the knock-in mice genocopying LCCS1-associated GLE1FinMajor variant (Gle1PFQ/PFQ) survive prenatal period but die suddenly at mid-adulthood. Gle1PFQ/PFQ mice present irregular count and distribution of spinal motor neurons and impaired development of neural crest-derived tissues as demonstrated by defects in their sympathetic innervation of heart ventricles, paravertebral sympathetic ganglia volume, and adrenal chromaffin cell counts. Unlike previously reported for yeast and HeLa cells, analysis of molecular consequences of GLE1FinMajor variant identified normal poly(A)+ RNA distribution in Gle1PFQ/PFQ cells, which however were impaired in RNA and protein synthesis and simultaneously showed typical signs of cellular senescence. Gle1PFQ/PFQ also induced disturbed stress responses with significant changes in G3BP1-positive stress granule count. Our results show necessity of GLE1 functions for life and indicate that LCCS1 etiology is resultant of pathogenic GLE1FinMajor variant impinging differentiation of neural crest derivatives and leading to complex multiorgan defects. HighlightsO_LITotal inactivation of GLE1 results in disorganization of blastocyst inner cell mass and early embryonic lethality. C_LIO_LIThe Gle1 knock-in (KI) mice, which genocopy the human GLE1FinMajor variant causative for lethal congenital contracture syndrome 1 (LCCS1), die suddenly in mid-adulthood. C_LIO_LINormal poly(A)+ RNA distribution was observed in Gle1 KI cells, but decreased number of G3BP1-positive stress granules were detected in response to stress. C_LIO_LIAbnormal sympathetic innervation of heart ventricles was detected in Gle1 KI mice. C_LIO_LINeural crest-derived tissues represent a new target of GLE1FinMajor and GLE1-related disorders. C_LI

developmental biology↗

bioRxiv2024

FINCA disease mouse model exhibits altered behaviour and immune response

Fibrosis, neurodegeneration and cerebral angiomatosis (FINCA) is a childhood-onset multi-organ neurodevelopmental disorder associated with multi-organ manifestations and recurrent infections. The disease is caused by variants in NHLRC2 initiating a cascade of unknown pathological events. Previously, we have demonstrated that despite the significant decrease at the molecular level, the compound heterozygosity of knock out and p.Asp148Tyr alleles in NHLRC2 does not lead to a severe phenotype in mice. Here, we analysed the behavioural and immunological phenotype of the FINCA mice and studied the molecular pathways affected by p.Asp148Tyr in NHLRC2 using mouse and human-derived cell culture models. The FINCA mice displayed a mild hyperactivity and deficient early immune response when challenged with LPS leading to altered cytokine responses, including IFN{gamma}, IL-12, and TNF. By comparing gene expression and putative interaction partners affected by p.Asp148Tyr, we identified Rho GTPase signalling as the common pathway. Altogether, these results establish a multi-dimensional impact of the p.Asp148Tyr variant in NHLRC2. Knowledge of the molecular pathways affected by NHLRC2 and the natural course of FINCA disease progression are instrumental for the development of effective therapeutics. Summary statementFINCA is a paediatric neurodevelopmental and multi-organ disorder caused by variants in NHLRC2. Here, mild hyperactivity in connection with altered early immune response is described in the FINCA mouse model.

neuroscience↗

bioRxiv2023

ciRS-7-miR7 regulate ischemia induced neuronal death via glutamatergic signaling

Brain functionality relies on finely tuned regulation of gene expression by networks of non-coding RNAs (ncRNAs) such as the one composed by the circular RNA ciRS-7 (also known as CDR1as), the microRNA miR-7 and the long non-coding RNA Cyrano. Here we describe ischemia induced alterations in the ncRNA network both in vitro and in vivo and in transgenic mice lacking ciRS-7 or miR-7. Our data show that cortical neurons downregulate ciRS-7 and Cyrano and upregulate miR-7 expression upon ischemic insults. Mice lacking ciRS-7 show reduced lesion size and motor impairment, whilst the absence of miR-7 alone leads to an increase in the ischemia induced neuronal death. Moreover, miR-7 levels in pyramidal excitatory neurons regulate dendrite morphology and glutamatergic signaling suggesting a potential molecular link to the in vivo phenotype. Our data reveal that ciRS-7 and miR-7 contribute to the outcome of ischemic stroke and shed new light into the pathophysiological roles of intracellular networks of non-coding RNAs in the brain.

molecular biology↗

bioRxiv2021

Analysis of human brain tissue derived from DBS surgery

BackgroundTranscriptomic and proteomic profiling of human brain tissue is hindered by availability of fresh samples from living patients. Postmortem samples usually represent the advanced disease stage of the patient. Furthermore, the postmortem interval affects the observed transcriptomic and proteomic profiles. Therefore, access to fresh brain tissue samples from living patients is valuable resource to obtain information on metabolically intact tissue. The implantation of deep brain stimulation (DBS) electrodes into the human brain is a neurosurgical treatment for, e.g., movement disorders. Here, we describe an improved approach to collect brain tissue from surgical instruments used in implantation of DBS device for transcriptomics and proteomics analyses. MethodsSamples were extracted from guide tubes and recording electrodes used in routine DBS implantation procedure that was carried out to treat patients with Parkinsons disease, genetic dystonia and tremor. RNA sequencing was carried out to tissue extracted from the recording microelectrodes and liquid chromatography-mass spectrometry was carried out to analyze tissue from guide tubes. To assess the performance of the current approach, obtained datasets were compared with previously published datasets representing brain tissue. ResultsIn RNA sequencing, altogether 32,034 transcripts representing unique Ensembl gene identifiers were detected from eight samples representing both hemispheres of four patients. By using liquid chromatography-mass spectrometry, we identified 734 unique proteins from 31 samples collected from 14 patients. Comparison with previously published brain derived data indicated that both of our datasets reflected the expected brain tissue specific features. The datasets are available via BioStudies database (accession number S-BSST667). ConclusionsSurgical instruments used in DBS installation retain enough brain material for protein and gene expression studies. Analysis of the datasets indicated that hemisphere-specific expression data can be obtained from individual patients without any sample pooling and without any modifications to the standard surgical protocol. Comparison with previously published datasets obtained with similar approach proved the robustness and reproducibility of the current improved protocol. This approach overcomes the issues that arise from using postmortem tissue, such as effect of postmortem interval, on proteomic and transcriptomic landscape of the brain and can be used for studying molecular aspects of DBS-treatable diseases.

neuroscience↗