Biology subjects

Venturini, L.

Publications and source records attributed to Venturini, L..

5 recordsLinked to original sources

bioRxiv2026

Direct 3D isolation in gelatin methacryloyl microbeads versus conventional 2D isolation and culture: a trade-off between mesenchymal stromal cell purity and niche-associated transcriptional profiles

Bone-marrow mesenchymal stromal cell (MSC) isolation typically couples plastic-adherence selection to planar expansion. Therefore, the recovered cells may reflect culture adaptation as much as native stromal biology. Direct isolation within a three-dimensional matrix could reduce planar selection, but its effects on cellular recovery, composition and MSC transcriptional profiles remain insufficiently resolved. We therefore developed a gelatin methacryloyl (GelMA) workflow progressing from bulk hydrogels to medium-surrounded GelMA microbeads (GMBs) and compared 3D-derived cells with conventional 2D cultures. Qualitative live/dead staining with calcein acetoxymethyl ester (CAM) and propidium iodide (PI) and 5-ethynyl-2'-deoxyuridine (EdU) imaging established cellular activity in both GelMA formats. In bulk GelMA cultures from four human donors, CD73-, CD90- and CD105-associated flow-cytometric readouts did not increase from day 7 to day 21. At day 21, the combined marker readout was numerically higher in donor-matched 2D cultures than in bulk 3D cultures, which retained heterogeneous marker-defined populations. GMBs enabled cell recovery and flow-cytometric characterization and motivated an exploratory one-donor single-cell RNA sequencing (scRNA-seq) analysis. The final dataset contained 37,784 cells across four samples and 23 annotated populations. MSC-focused re-embedding resolved 13,175 cells into two 3D and four 2D clusters. A pooled comparison identified 4,840 differentially expressed genes (DEGs, false discovery rate [FDR]<0.01, |log2FC|[≥]1), indicating a broad method-associated expression shift. The 3D C1 cluster combined progenitor-, extracellular-matrix-, adhesion-, cytokine-, hypoxia/glycolysis- and predicted-interaction-associated programmes, whereas 2D clusters showed mitochondrial, cytoskeletal/mechanoadaptive and proliferative programmes. Thus, 2D culture yielded a more strongly represented and compositionally purer MSC compartment, while 3D isolation contained rare MSC clusters with niche-associated transcriptional profiles within a heterogeneous marrow environment. These findings support a testable native-associated tendency for 3D, but donor replication, matched recovery and direct native-reference mapping are required before inferring greater in-vivo similarity or functional superiority.

cell biology↗

bioRxiv2026

Spatiotemporal atlas of pro-inflammatory (NF-kB) and anti-inflammatory (STAT6) signalling using reporter mice during mRNA vaccination

The immunization process unfolds through a precisely orchestrated sequence of innate and adaptive events across distinct anatomical sites. Although many mechanisms underlying vaccination are well described, most vaccines have been developed empirically, partly due to the lack of tools enabling rapid, organ-specific analysis of immune activation. To address this gap, we developed and validated a novel STAT6 reporter mouse enabling dynamic in vivo whole-body imaging and ex vivo analysis of STAT6-mediated anti-inflammatory signalling, and combined it with an established NF-{kappa}B reporter model to dissect immune activation induced by two LNP-encapsulated mRNA vaccines encoding the same antigen but differing in RNA chemistry (unmodified versus N{superscript 1}-methyl-pseudouridine (m{superscript 1}{Psi})-modified). This dual-reporter system enabled the creation of a spatiotemporal atlas of vaccine-induced signalling, revealing chemistry-dependent immune dynamics and identifying the liver as a predominant early hub for both NF-{kappa}B and STAT6 activity following systemic administration. Integration with antibody measurements demonstrated that early STAT6 activation followed by rapid signal resolution--rather than prolonged NF-{kappa}B-mediated inflammation--correlated with robust humoral responses, suggesting that monitoring NF-{kappa}B and STAT6 dynamics could provide predictive insight into vaccine immunogenicity. Together, these findings establish NF-{kappa}B and STAT6 reporter mice as rapid in vivo screening tools for the early assessment of vaccine immunogenicity and performance. By enabling dynamic, organ-resolved immune profiling, this approach paves the way for more rational, mechanism-driven design of mRNA vaccines and underscores the importance of further investigating the effects of vaccines on the liver, both as a primary LNP target and as an immunologically tolerogenic organ.

immunology↗

bioRxiv2017

Efficient and accurate detection of splice junctions from RNAseq with Portcullis

Next generation sequencing (NGS) technologies enable rapid and cheap genome-wide transcriptome analysis, providing vital information about gene structure, transcript expression and alternative splicing. Key to this is the the accurate identification of exon-exon junctions from RNA sequenced (RNA-seq) reads. A number of RNA-seq aligners capable of splitting reads across these splice junctions (SJs) have been developed, however, it has been shown that while they correctly identify most genuine SJs available in a given sample, they also often produce large numbers of incorrect SJs. Herein we describe the extent of this problem using popular RNA-seq mapping tools, and present a new method, called Portcullis, to rapidly filter false SJs junctions from spliced alignments produced by any RNA-seq mapper capable of creating SAM/BAM files. We show that Portcullis distinguishes between genuine and false positive junctions to a high-degree of accuracy across different species, samples, expression levels, error profiles and read lengths. Portcullis makes efficient use of memory and threading and, to our knowledge, is currently the only SJ prediction tool that reliably scales for use with large RNAseq datasets and large highly fragmented genomes, whilst delivering highly accurate SJs.\n\nAvailabilityPortcullis is available under the GPLv3 license at: http://maplesond.github.io/portcullis/\n\nContactdaniel.mapleson@earlham.ac.uk

bioinformatics↗

bioRxiv2017

Leveraging multiple transcriptome assembly methods for improved gene structure annotation

The performance of RNA-Seq aligners and assemblers varies greatly across different organisms and experiments, and often the optimal approach is not known beforehand. Here we show that the accuracy of transcript reconstruction can be boosted by combining multiple methods, and we present a novel algorithm to integrate multiple RNA-Seq assemblies into a coherent transcript annotation. Our algorithm can remove redundancies and select the best transcript models according to user-specified metrics, while solving common artefacts such as erroneous transcript chimerisms. We have implemented this method in an open-source Python3 and Cython program, Mikado, available at https://github.com/lucventurini/Mikado.

bioinformatics↗

bioRxiv2016

An improved assembly and annotation of the allohexaploid wheat genome identifies complete families of agronomic genes and provides genomic evidence for chromosomal translocations.

Advances in genome sequencing and assembly technologies are generating many high quality genome sequences, but assemblies of large, repeat-rich polyploid genomes, such as that of bread wheat, remain fragmented and incomplete. We have generated a new wheat whole-genome shotgun sequence assembly using a combination of optimised data types and an assembly algorithm designed to deal with large and complex genomes. The new assembly represents more than 78% of the genome with a scaffold N50 of 88.8kbp that has a high fidelity to the input data. Our new annotation combines strand-specific Illumina RNAseq and PacBio full-length cDNAs to identify 104,091 high confidence protein-coding genes and 10,156 non-coding RNA genes. We confirmed three known and identified one novel genome rearrangements. Our approach enables the rapid and scalable assembly of wheat genomes, the identification of structural variants, and the definition of complete gene models, all powerful resources for trait analysis and breeding of this key global crop. [Supplemental material is available for this article.]

genomics↗