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Investigating ultrastructural morphology in MIRAGE syndrome-derived fibroblasts using transmission electron
Federica Buonocore1, Monika Balys2, Glenn Anderson2
1Genetics and Genomic Medicine Research and Teaching Department, UCL Great Ormond Street Institute of Child Health, University College London, London, UK.
Background:
Heterozygous de novo variants in the gene SAMD9 cause the complex multisystem disorder, MIRAGE syndrome. Patients are characterised by myelodysplasia, infections, growth restriction, adrenal insufficiency, gonadal dysfunction and enteropathies. Pathogenic variants in SAMD9 are gain-of-function and enhance its role as a growth repressor, leading to growth restriction of many tissues. Two studies have reported changes in skin fibroblasts derived from MIRAGE patients, more specifically identifying enlarged endosomes. We have also previously shown subtle changes in endosome size in patients' fibroblasts compared to controls. However, these variations in endosomes were not as marked as those described in the literature.
Methods:
We have performed an observational study using transmission electron microscopy (TEM) in a larger number of cells derived from three patients' fibroblasts to assess ultrastructure morphology compared to control images.
Results:
Consistent changes were observed in cell organelles in all patient samples. In particular, increased endosomal activity was detected, characterised by augmented pinocytosis and vesicle budding, increased endosome number, as well as by large lysosomes and endosomes. Endoplasmic reticulum was also prominent. Mitochondria appeared enlarged in selected cells, possibly due to cellular stress. Cell nuclei did not display major differences compared to controls.
Conclusions:
TEM is a powerful tool to investigate morphological features of tissues and cell organelles, although TEM data could be affected by sample preparation methodology, therefore potentially explaining the variability between independent studies, and its analysis can be dependent on the experience of the researcher. The increased endosomal activity we have observed in patients' fibroblasts could indicate that SAMD9 regulates endocytosis of receptors, acting as an endosome fusion facilitator, or in lysosomal activation. However, the precise mechanism(s) by which SAMD9 regulates cell growth is still not fully understood, and further studies are needed to elucidate its pathogenic pathway and develop therapeutic approaches to support patients.
Insights
Transmission electron microscopy revealed consistent cellular changes in MIRAGE syndrome fibroblasts, notably increased endosomal activity and enlarged organelles. These findings shed light on SAMD9 gene variants
Area of Science:
- Cell Biology
- Genetics
- Pathology
Background:
- MIRAGE syndrome is a complex multisystem disorder caused by heterozygous de novo variants in the SAMD9 gene.
- Pathogenic SAMD9 variants are gain-of-function, enhancing its growth-repressor role and causing tissue growth restriction.
- Previous studies noted subtle endosome size changes in MIRAGE patient fibroblasts, but findings were not consistently marked.
Purpose of the Study:
- To investigate ultrastructure morphology of fibroblasts from MIRAGE syndrome patients using transmission electron microscopy (TEM).
- To compare cellular organelle morphology in patient-derived fibroblasts with control samples.
- To further characterize the cellular basis of MIRAGE syndrome and the role of SAMD9.
Main Methods:
- Observational study utilizing transmission electron microscopy (TEM).
- Analysis of fibroblast ultrastructure morphology from three MIRAGE syndrome patients.
- Comparison of patient cell images with control fibroblast images.
Main Results:
- Consistent ultrastructural changes observed in all patient fibroblast samples.
- Significant increase in endosomal activity, including augmented pinocytosis, vesicle budding, and increased endosome/lysosome size.
- Prominent endoplasmic reticulum and enlarged mitochondria in selected cells; no major differences in cell nuclei.
Conclusions:
- TEM is a valuable tool for ultrastructural analysis, though methodology and researcher experience can influence results.
- Increased endosomal activity in MIRAGE fibroblasts suggests SAMD9 may regulate endocytosis or lysosomal pathways.
- Further research is required to fully understand SAMD9's growth regulation mechanisms and develop therapeutic strategies.
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