ABSTRACT
Maternal obesity and hyperglycemia are linked to an elevated risk for obesity, diabetes, and steatotic liver disease in the adult offspring. To establish and validate a noninvasive workflow for perinatal metabolic phenotyping, fixed neonates of common mouse strains were analyzed postmortem via magnetic resonance imaging (MRI)/magnetic resonance spectroscopy (MRS) to assess liver volume and hepatic lipid (HL) content. The key advantage of nondestructive MRI/MRS analysis is the possibility of further tissue analyses, such as immunohistochemistry, RNA extraction, and even proteomics, maximizing the data that can be gained per individual and therefore facilitating comprehensive correlation analyses. This study employed an MRI and 1H-MRS workflow to measure liver volume and HL content in 65 paraformaldehyde-fixed murine neonates at 11.7 T. Liver volume was obtained using semiautomatic segmentation of MRI acquired by a RARE sequence with 0.5-mm slice thickness. HL content was measured by a STEAM sequence, applied with and without water suppression. T1 and T2 relaxation times of lipids and water were measured for respective correction of signal intensity. The HL content, given as CH2/(CH2 + H2O), was calculated, and the intrasession repeatability of the method was tested. The established workflow yielded robust results with a variation of ~3% in repeated measurements for HL content determination. HL content measurements were further validated by correlation analysis with biochemically assessed triglyceride contents (R2 = 0.795) that were measured in littermates. In addition, image quality also allowed quantification of subcutaneous adipose tissue and stomach diameter. The highest HL content was measured in C57Bl/6N (4.2%) and the largest liver volume and stomach diameter in CBA (53.1 mm3 and 6.73 mm) and NMRI (51.4 mm3 and 5.96 mm) neonates, which also had the most subcutaneous adipose tissue. The observed effects were independent of sex and litter size. In conclusion, we have successfully tested and validated a robust MRI/MRS workflow that allows assessment of morphology and HL content and further enables paraformaldehyde-fixed tissue-compatible subsequent analyses in murine neonates.
ABSTRACT
PURPOSE: Rothmund-Thomson syndrome (RTS) is characterized by poikiloderma, sparse hair, small stature, skeletal defects, cancer, and cataracts, resembling features of premature aging. RECQL4 and ANAPC1 are the 2 known disease genes associated with RTS in >70% of cases. We describe RTS-like features in 5 individuals with biallelic variants in CRIPT (OMIM 615789). METHODS: Two newly identified and 4 published individuals with CRIPT variants were systematically compared with those with RTS using clinical data, computational analysis of photographs, histologic analysis of skin, and cellular studies on fibroblasts. RESULTS: All CRIPT individuals fulfilled the diagnostic criteria for RTS and additionally had neurodevelopmental delay and seizures. Using computational gestalt analysis, CRIPT individuals showed greatest facial similarity with individuals with RTS. Skin biopsies revealed a high expression of senescence markers (p53/p16/p21) and the senescence-associated ß-galactosidase activity was elevated in CRIPT-deficient fibroblasts. RECQL4- and CRIPT-deficient fibroblasts showed an unremarkable mitotic progression and unremarkable number of mitotic errors and no or only mild sensitivity to genotoxic stress by ionizing radiation, mitomycin C, hydroxyurea, etoposide, and potassium bromate. CONCLUSION: CRIPT causes an RTS-like syndrome associated with neurodevelopmental delay and epilepsy. At the cellular level, RECQL4- and CRIPT-deficient cells display increased senescence, suggesting shared molecular mechanisms leading to the clinical phenotypes.