Enteric and hepatic glia-like cell reactivation after astrocyte-directed twinkle helicase loss

Mitochondrial brain diseases, such as Alpers-Huttenlocher syndrome and mitochondrial recessive ataxia syndrome, are complex and severe metabolic diseases with variety of symptoms that lack any treatment. Multiple different mutations in the proteins taking part in the mitochondrial DNA (mtDNA) replication, such as mutations in the Twinkle helicase gene, can lead to mtDNA depletion. This causes for example dysfunction of the oxidative phosphorylation complexes and problems in adenosine triphosphate (ATP) synthesis resulting in mitochondrial stress and disease. A mouse model has been established to study these diseases by knocking out the twinkle gene from astrocytes, glial cells of the brain, causing spongiotic encephalopathy, astrogliosis and ciliogenesis activation in the mouse brain. This model is created by using a glial fibrillary acid (GFAP) promoter, driving a Cre recombinase that together with LoxP sequences leads to a knockout of the twinkle gene in the cells expressing GFAP. In this thesis I studied whether the astrocytic twinkle gene knockout (TwKOastro) affects non-nervous system cells that express Cre-transgene. The main methods include histological stainings, both haematoxylin-eosin (HE) and immunofluorescence stainings on the mouse brain, intestine and liver. In addition to central nervous system astrocytes, also peripheric nervous system’s enteric glial cells and liver’s hepatic stellate cells express GFAP and mimic the functions of astrocytes by facilitating surrounding cells in their tasks. There were no changes in the gross pathology of either intestine or liver tissue that could be comparable to the brain manifestations. In the intestine, the GFAP immunofluorescence signal localized to the enteric glial cells as expected, but there were no differences between the TwKOastro mice and controls indicating reactivation of these cells. The finding is interesting, suggesting that the tolerance of enteric glia to Twinkle loss is high. GFAP signal in the tdTomato-GFAP (tandem-dimer tomato) mouse intestine localized similarly and confirmed these results. In the liver, the GFAP signal was spot-like and could not be used as a reliable marker to detect the hepatic stellate cells (HSCs) although some cell somas could be identified. In addition, another HSC marker, alpha smooth muscle actin, or the tdTomato protein did not bring any further results. We did not analyze mtDNA amount in situ, which would be a consequence of Twinkle loss, provided that this helicase acts as the replicative helicase in the intestine, as it does in the nervous system. However, these results provide a good baseline for future experiments that want to explore the possible triggers for these mitochondrial diseases and the effects of twinkle knockout in different tissues in this mouse model. Further research considering the roles of enteric glia subtypes as well as liver zonation and its metabolic differences would possibly generate interesting results.

Grantor dc:publisher

Helsingin yliopisto

Year dc:date.issued

2024

Author dc:creator

• Sarkola, Emmi

Contributors dc:contributor

• Helsingin yliopisto, Lääketieteellinen tiedekunta
• University of Helsinki, Faculty of Medicine
• Helsingfors universitet, Medicinska fakulteten

Language dc:language.iso

eng

Identifier URI

URN:NBN:fi:hulib-202501301272

OAI identifier oai:identifier

oai:helda.helsinki.fi:10138/591652