Identification of Spatio-Temporal Systems Using Multi-resolution Wavelet Models

A novel modelling structure for identifying spatio-temporal systems is proposed based on multi-resolution wavelet decompositions. Two classes of spatio-temporal systems, coupled map lattices and cellular automata, are considered and the new modelling approach is applied to identify these systems. Several examples are provided to demonstrate the applicability and effectiveness of the new identification procedure

Broadening the reach and investigating the potential of prime editors through fully viral gene-deleted adenoviral vector delivery

Prime editing is a recent precision genome editing modality whose versatility offers the prospect for a wide range of applications, including the development of targeted genetic therapies. Yet, an outstanding bottleneck for its optimization and use concerns the difficulty in delivering large prime editing complexes into cells. Here, we demonstrate that packaging prime editing constructs in adenoviral capsids overcomes this constrain resulting in robust genome editing in both transformed and non-transformed human cells with up to 90% efficiencies. Using this cell cycle-independent delivery platform, we found a direct correlation between prime editing activity and cellular replication and disclose that the proportions between accurate prime editing events and unwanted byproducts can be influenced by the target-cell context. Hence, adenovector particles permit the efficacious delivery and testing of prime editing reagents in human cells independently of their transformation and replication statuses. The herein integrated gene delivery and gene editing technologies are expected to aid investigating the potential and limitations of prime editing in numerous experimental settings and, eventually, in ex vivo or in vivo therapeutic contexts.Molecular Epidemiolog

Circulating microRNAs highly correlate to expression of cartilage genes potentially reflecting OA susceptibility: towards identification of applicable early OA biomarkers

Objective: To identify and validate circulating micro RNAs (miRNAs) that mark gene expression changes in articular cartilage early in osteoarthritis (OA) pathophysiology process. Methods: Within the ongoing RAAK study, human preserved OA cartilage and plasma (N = 22 paired samples) was collected for RNA sequencing (respectively mRNA and miRNA). Spearman correlation was determined for 114 cartilage genes consistently and significantly differentially expressed early in osteoarthritis and 384 plasma miRNAs. Subsequently, the minimal number of circulating miRNAs serving to discriminate between progressors and non-progressors was assessed by regression analysis and area under receiver operating curves (AUC) was calculated with progression data and plasma miRNA sequencing from the GARP study (N = 71). Results: We identified strong correlations (rho >= |0.7|) among expression levels of 34 unique plasma miRNAs and 21 genes, including 4 genes that correlated with multiple miRNAs. The strongest correlation was between let-7d-5p and EGFLAM (rho = -0.75, P = 6.9 x 10(-5)). Regression analysis of the 34 miRNAs resulted in a set of 7 miRNAs that, when applied to the GARP study, demonstrated clinically relevant predictive value with AUC > 0.8 for OA progression over 2 years and near-clinical value for progression over 5 years- (AUC = 0.8). Conclusions: We show that plasma miRNAs levels reflect gene expression levels in cartilage and can be exploited to represent ongoing pathophysiological processes in articular cartilage. We advocate that identified signature of 7 plasma miRNAs can contribute to direct further studies toward early biomarkers predictive for progression of osteoarthritis over 2 and 5 years.Molecular Epidemiolog

HbA(1c) is associated with altered expression in blood of cell cycle- and immune response-related genes

Therapeutic cell differentiatio

Radiofrequency ablation versus hepatic resection for hepatocellular carcinoma within the Milan criteria – A comparative study

AbstractBackgroundTo compare the results of radiofrequency ablation (RFA) with hepatic resection in the treatment of hepatocellular carcinoma (HCC) within the Milan criteria.MethodsA nonrandomized comparative study was performed with 111 consecutive patients who underwent laparoscopic RFA (n = 31) or curative hepatic resection (n = 80) for HCC within Milan criteria.ResultsProcedure related complications were less often and severe after RFA than resection (3.2% vs. 25%). There was no significant difference in hospital mortality (0% vs. 3.8%). Hospital stay was significantly shorter in the RFA group than in the resection group (mean, 3.8 vs. 6.8 days). The 1-, 3-, and 5-year disease-free survival rates for the RFA group and the resection group were 76%, 40%, 40% and 76%, 60%, 60%, respectively. Disease-free survival was significantly lower in the RFA group than in the resection group. The corresponding 1-, 3-, and 5-year overall survival rates for the RFA group and the resection group were 100%, 92%, 84%, and 92%, 75%, 71%, respectively. The overall survival for RFA and resection were not significantly different.ConclusionsOur result showed comparable overall survival between RFA and surgery, although RFA was associated with a significantly higher tumor recurrence rate. RFA had the advantages over surgical resection in being less invasive and having lower morbidity

Single-cell transcriptomics analysis of human small antral follicles

Stem cells & developmental biolog

Elucidating mechano-pathology of osteoarthritis: transcriptome-wide differences in mechanically stressed aged human cartilage explants

Background Failing of intrinsic chondrocyte repair after mechanical stress is known as one of the most important initiators of osteoarthritis. Nonetheless, insight into these early mechano-pathophysiological processes in age-related human articular cartilage is still lacking. Such insights are needed to advance clinical development. To highlight important molecular processes of osteoarthritis mechano-pathology, the transcriptome-wide changes following injurious mechanical stress on human aged osteochondral explants were characterized. Methods Following mechanical stress at a strain of 65% (65%MS) on human osteochondral explants (n(65%MS) = 14 versus n(control) = 14), RNA sequencing was performed. Differential expression analysis between control and 65%MS was performed to determine mechanical stress-specific changes. Enrichment for pathways and protein-protein interactions was analyzed with Enrichr and STRING. Results We identified 156 genes significantly differentially expressed between control and 65%MS human osteochondral explants. Of note, IGFBP5 (FC = 6.01; FDR = 7.81 x 10(-3)) and MMP13 (FC = 5.19; FDR = 4.84 x 10(-2)) were the highest upregulated genes, while IGFBP6 (FC = 0.19; FDR = 3.07 x 10(-4)) was the most downregulated gene. Protein-protein interactions were significantly higher than expected by chance (P = 1.44 x 10(-15) with connections between 116 out of 156 genes). Pathway analysis showed, among others, enrichment for cellular senescence, insulin-like growth factor (IGF) I and II binding, and focal adhesion. Conclusions Our results faithfully represent transcriptomic wide consequences of mechanical stress in human aged articular cartilage with MMP13, IGF binding proteins, and cellular senescence as the most notable results. Acquired knowledge on the as such identified initial, osteoarthritis-related, detrimental responses of chondrocytes may eventually contribute to the development of effective disease-modifying osteoarthritis treatments.Molecular Epidemiolog

CACTUS: integrating clonal architecture with genomic clustering and transcriptome profiling of single tumor cells

Background: Drawing genotype-to-phenotype maps in tumors is of paramount importance for understanding tumor heterogeneity. Assignment of single cells to their tumor clones of origin can be approached by matching the genotypes of the clones to the mutations found in RNA sequencing of the cells. The confidence of the cell-to-clone mapping can be increased by accounting for additional measurements. Follicular lymphoma, a malignancy of mature B cells that continuously acquire mutations in parallel in the exome and in B cell receptor loci, presents a unique opportunity to join exome-derived mutations with B cell receptor sequences as independent sources of evidence for clonal evolution.Methods: Here, we propose CACTUS, a probabilistic model that leverages the information from an independent genomic clustering of cells and exploits the scarce single cell RNA sequencing data to map single cells to given imperfect genotypes of tumor clones.Results: We apply CACTUS to two follicular lymphoma patient samples, integrating three measurements: whole exome, single-cell RNA, and B cell receptor sequencing. CACTUS outperforms a predecessor model by confidently assigning cells and B cell receptor-based clusters to the tumor clones.Conclusions: The integration of independent measurements increases model certainty and is the key to improving model performance in the challenging task of charting the genotype-to-phenotype maps in tumors. CACTUS opens the avenue to study the functional implications of tumor heterogeneity, and origins of resistance to targeted therapies. CACTUS is written in R and source code, along with all supporting files, are available on GitHub (https://github.com/LUMC/CACTUS).Development and application of statistical models for medical scientific researc

Characterizing the secretome of licensed hiPSC-derived MSCs

Although mesenchymal stromal cells (MSCs) from primary tissues have been successfully applied in the clinic, their expansion capabilities are limited and results are variable. MSCs derived from human-induced pluripotent stem cells (hiMSCs) are expected to overcome these limitations and serve as a reproducible and sustainable cell source. We have explored characteristics and therapeutic potential of hiMSCs in comparison to hBMSCs. RNA sequencing confirmed high resemblance, with average Pearson correlation of 0.88 and Jaccard similarity index of 0.99, and similar to hBMSCs the hiMSCs released extracellular vesicles with in vitro immunomodulatory properties. Potency assay with TNF alpha and IFN gamma demonstrated an increase in well-known immunomodulatory genes such as IDO1, CXCL8/IL8, and HLA-DRA which was also highlighted by enhanced secretion in the media. Notably, expression of 125 genes increased more than 1000-fold. These genes were predicted to be regulated by NFKB signaling, known to play a central role in immune response. Altogether, our data qualify hiMSCs as a promising source for cell therapy and/or cell-based therapeutic products. Additionally, the herewith generated database will add to our understanding of the mode of action of regenerative cell-based therapies and could be used to identify relevant potency markers.Molecular Epidemiolog

Efficient and scalable generation of primordial germ cells in 2D culture using basement membrane extract overlay

Current methods to generate human primordial germ cell-like cells (hPGCLCs) from human pluripotent stem cells (hPSCs) can be inefficient, and it is challenging to generate sufficient hPGCLCs to optimize in vitro gametogenesis. We present a differentiation method that uses diluted basement membrane extract (BMEx) and low BMP4 concentration to efficiently induce hPGCLC differentiation in scalable 2D cell culture. We show that BMEx overlay potentiated BMP/SMAD signaling, induced lumenogenesis, and increased expression of key hPGCLC-progenitor markers such as TFAP2A and EOMES. hPGCLCs that were generated using the BMEx overlay method were able to upregulate more mature germ cell markers, such as DAZL and DDX4, in human fetal ovary reconstitution culture. These findings highlight the importance of BMEx during hPGCLC differentiation and demonstrate the potential of the BMEx overlay method to interrogate the formation of PGCs and amnion in humans, as well as to investigate the next steps to achieve in vitro gametogenesis.Stem cells & developmental biolog