PMC10343430_01
PMC10343430_02
(A) Brain magnetic resonance imaging (MRI) shows abnormal signals in the bilateral periventricular anterior white matter and the anterior part of the corpus callosum. (B) Axial T2-weighted MRI shows abnormal signals in the frontal white matter and in the pyramidal tract at the internal capsule. There was no clear pattern of cortical atrophy, and the hippocampal volume was normal. (C) The diffusion-weighted image (DWI) shows patchy areas of restricted diffusion in the abnormal white matter. (D) Visuospatial ability performance in the Minimental Status Examination (MMSE) test showing visuospatial disintegration (no intersection, loss of some sides and angles); (E) Clock Drawing Test (CDT) with perseverative drawing; (F) Writing a sentence in the MMSE test showing dysgraphia.
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F1
The brain magnetic resonance imaging (MRI) showed abnormal signals in the bilateral periventricular anterior white matter and the anterior part of the corpus callosum (Figures 1A-C). 18-F FDG PET/MRI showed frontal leukoencephalopathy with hypometabolism at the fronto-cerebellar cortex and caudate nucleus. The performance in cognitive tests and neuropsychiatric evaluation reports are shown in Supplementary Table 1 and Figures 1D-F.
Anteverted uterus, with regular and poorly defined contours, with approximate dimensions of 4.6 x 2.6 x 2.6 cm in its longitudinal, transverse and anteroposterior axes, respectively.
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F2
The study revealed a diminished size uterus confirming the uterus infantilis (Figure 2).
(A) Sanger sequencing of the AARS2 in gDNA from the control, mother, and patient confirming the presence of both variants: NM_020745:c.1036C>A (p.P346T) and NM_020745:c.2864G>A (p.W955*) in the patient. The p. c.2864G>A * was identified in the mother. (B) Schematic representation of a wild type, and the alternative and constitutive splicing isoforms caused by the c.1036C>A variant. (C) Sanger sequencing showing the pathogenic alternative splicing caused by the c.1036C>A variant. Three PCR products were sequenced from a unique PCR band amplifying the AARS2 transcript. The corresponding sequences are shown in the AARS2 affected" section of the figure: the first corresponds to the allele with the c.2864G>A, p.W955* variant that appears normal in the shown area. The second read showed a deletion of seven exon nucleotides that remove the c.1036C>A and causes a change in the reading frame with a premature stop codon as a predicted consequence (p.P346Wfs*18). The third read corresponds to the c.1036C>A constitutively spliced variant translating as a P346T aminoacid substitution without a change in the reading frame.
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F3
The variant c.2864G>A (p.W955*) was also identified in the patient's healthy mother (Figure 3A). The schematic representation and the results are shown in Figures 3B, C.
Amino acid alignment of the AARS2 homologous proteins. The amino acid p.P346 is not conserved; only Ptroglodytes and M. Mulatta have proline in this position.
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F4
Amino acid alignment of the AARS2 homologous gene is shown in Figure 4.
Muscle biopsy images. (A) Hematoxylin-Eosin Stain (H-E). Panoramic cross-sectional view of muscle with myopathic and angulated fascicles (marked with white arrow), difference in shape and size of the fiber, intra- and inter-fascicular connective tissue. In (B), close-up of image (A) with the presence of the hypertrophic fiber (white asterisk), surrounded by angulated fibers (black asterisk). In (C, D) images (Gomori's Modified Trichrome Stain (T-G)) the same findings are observed as in (A). In (E) panoramic view with the technique of Reduced Nicotinamide Adenine Dinucleotide-Tetrazolium Reductase (NADH-TR) with type I fibers of darker color and type II fibers of lighter color. In (F) close-up of the previous image, where a hypertrophic fiber is observed, surrounded by some angulated fibers. In (G), view of a transverse section with cytochrome C oxidase (COX) showing pale (thin black arrow) and ghost (thick black arrow) fibers. In (H) Cytochrome C Oxidase/Succinate Dehydrogenase (COX/SDH) double-labeling method we observe two blue fibers.
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F5
Muscle morphological studies revealed myopathic and neuronopathic changes with loss of COX activity in some fibers and blue fibers presence in Cytochrome C Oxidase/Succinate Dehydrogenase (COX/SDH) double-labeling method which suggests COX deficiency (Figure 5).