Transcriptomic and proteomic profiles of fetal versus adult mesenchymal stromal cells and mesenchymal stromal cell-derived extracellular vesicles.

BACKGROUND: Mesenchymal stem/stromal cells (MSCs) can regenerate tissues through engraftment and differentiation but also via paracrine signalling via extracellular vesicles (EVs). Fetal-derived MSCs (fMSCs) have been shown, both in vitro and in animal studies, to be more efficient than adult MSC (aMSCs) in generating bone and muscle but the underlying reason for this difference has not yet been clearly elucidated. In this study, we aimed to systematically investigate the differences between fetal and adult MSCs and MSC-derived EVs at the phenotypic, RNA, and protein levels. METHODS: We carried out a detailed and comparative characterization of culture-expanded fetal liver derived MSCs (fMSCs) and adult bone marrow derived MSCs (aMSCs) phenotypically, and the MSCs and MSC-derived EVs were analysed using transcriptomics and proteomics approaches with RNA Sequencing and Mass Spectrometry. RESULTS: Fetal MSCs were smaller, exhibited increased proliferation and colony-forming capacity, delayed onset of senescence, and demonstrated superior osteoblast differentiation capability compared to their adult counterparts. Gene Ontology analysis revealed that fMSCs displayed upregulated gene sets such as "Positive regulation of stem cell populations", "Maintenance of stemness" and "Muscle cell development/contraction/Myogenesis" in comparison to aMSCs. Conversely, aMSCs displayed upregulated gene sets such as "Complement cascade", "Adipogenesis", "Extracellular matrix glycoproteins" and "Cellular metabolism", and on the protein level, "Epithelial cell differentiation" pathways. Signalling entropy analysis suggested that fMSCs exhibit higher signalling promiscuity and hence, higher potency than aMSCs. Gene ontology comparisons revealed that fetal MSC-derived EVs (fEVs) were enriched for "Collagen fibril organization", "Protein folding", and "Response to transforming growth factor beta" compared to adult MSC-derived EVs (aEVs), whereas no significant difference in protein expression in aEVs compared to fEVs could be detected. CONCLUSIONS: This study provides detailed and systematic insight into the differences between fMSCs and aMSCs, and MSC-derived EVs. The key finding across phenotypic, transcriptomic and proteomic levels is that fMSCs exhibit higher potency than aMSCs, meaning they are in a more undifferentiated state. Additionally, fMSCs and fMSC-derived EVs may possess greater bone forming capacity compared to aMSCs. Therefore, using fMSCs may lead to better treatment efficacy, especially in musculoskeletal diseases.

Novel LDLR Variant in Familial Hypercholesterolemia: NGS-Based Identification, In Silico Characterization, and Pharmacogenetic Insights.

BACKGROUND: Familial Hypercholesterolemia (FH) is a hereditary condition that causes a rise in blood cholesterol throughout a person's life. FH can result in myocardial infarction and even sudden death if not treated. FH is thought to be caused mainly by variants in the gene for the low-density lipoprotein receptor (LDLR). This study aimed to investigate the genetic variants in FH patients, verify their pathogenicity, and comprehend the relationships between genotype and phenotype. Also, review studies assessed the relationship between the LDLR null variants and the reaction to lipid-lowering therapy. METHODS: The study utilised high-throughput next-generation sequencing for genetic screening of FH-associated genes and capillary sequencing for cascade screening. Furthermore, bioinformatic analysis was employed to describe the pathogenic effects of the revealed novel variant on the structural features of the corresponding RNA molecule. RESULTS: We studied the clinical signs of hypercholesterolemia in a Saudi family with three generations of FH. We discovered a novel frameshift variant (c.666_670dup, p.(Asp224Alafs*43) in the LDLR and a known single nucleotide variant (c.9835A > G, p.(Ser3279Gly) in the APOB gene. It is thought that the LDLR variant causes a protein to be prematurely truncated, likely through nonsense-mediated protein decay. The LDLR variant is strongly predicted to be pathogenic in accordance with ACMG guidelines and co-segregated with the FH clinical characteristics of the family. This LDLR variant exhibited severe clinical FH phenotypes and was restricted to the LDLR protein's ligand-binding domain. According to computational functional characterization, this LDLR variant was predicted to change the free energy dynamics of the RNA molecule, thereby affecting its stability. This frameshift variant is thought to eliminate important functional domains in LDLR that are required for receptor recycling and LDL particle binding. We provide insight into how FH patients with a null variant in the LDLR gene respond to lipid-lowering therapy. CONCLUSIONS: The findings expand the range of FH variants and assist coronary artery disease preventive efforts by improving diagnosis, understanding the genotype-phenotype relationship, prognosis, and personalised therapy for patients with FH.

Carotid Plaque Phenotyping by Correlating Plaque Morphology from Computed Tomography Angiography with Transcriptional Profiling.

OBJECTIVE: Ischaemic strokes can be caused by unstable carotid atherosclerosis, but methods for identification of high risk lesions are lacking. Carotid plaque morphology imaging using software for visualisation of plaque components in computed tomography angiography (CTA) may improve assessment of plaque phenotype and stroke risk, but it is unknown if such analyses also reflect the biological processes related to lesion stability. Here, we investigated how carotid plaque morphology by image analysis of CTA is associated with biological processes assessed by transcriptomic analyses of corresponding carotid endarterectomies (CEAs). METHODS: Carotid plaque morphology was assessed in patients undergoing CEA for symptomatic or asymptomatic carotid stenosis consecutively enrolled between 2006 and 2015. Computer based analyses of pre-operative CTA was performed to define calcification, lipid rich necrotic core (LRNC), intraplaque haemorrhage (IPH), matrix (MATX), and plaque burden. Plaque morphology was correlated with molecular profiles obtained from microarrays of corresponding CEAs and models were built to assess the ability of plaque morphology to predict symptomatology. RESULTS: Carotid plaques (n = 93) from symptomatic patients (n = 61) had significantly higher plaque burden and LRNC compared with plaques from asymptomatic patients (n = 32). Lesions selected from the transcriptomic cohort (n = 40) with high LRNC, IPH, MATX, or plaque burden were characterised by molecular signatures coupled with inflammation and extracellular matrix degradation, typically linked with instability. In contrast, highly calcified plaques had a molecular signature signifying stability with enrichment of profibrotic pathways and repressed inflammation. In a cross validated prediction model for symptoms, plaque morphology by CTA alone was superior to the degree of stenosis. CONCLUSION: The study demonstrates that CTA image analysis for evaluation of carotid plaque morphology, also reflects prevalent biological processes relevant for assessment of plaque phenotype. The results support the use of CTA image analysis of plaque morphology for risk stratification and management of patients with carotid stenosis.