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Biology subjects

Bressan, M.

Publications and source records attributed to Bressan, M..

4 recordsLinked to original sources

A Combined Computational Fluid Dynamics Modeling and Geometric Morphometrics Methods Approach to Quantifying Hemodynamic and Anatomical Features of Embryonic Chick Heart Anatomies Reconstructed from Light Sheet Fluorescence Microscopy Imaging

Although congenital heart defects occur in approximately 1% of newborns in the US annually, their pathogenesis remains largely unknown. Less than a third of congenital heart defects are traced a known genetic or environmental cause. It has been demonstrated that hemodynamic forces such as wall shear stress are critical for heart development. However, measuring these hemodynamic factors in vivo is infeasible due to physical limitations, such as the small size and constant motion of the embryonic heart. An alternative approach is to recapitulate the hemodynamic environment by simulating blood flow and calculating the resulting hemodynamic forces through computational fluid dynamics modeling. We use computational fluid dynamics modeling to quantify hemodynamics in a cohort of cell-accurate embryonic chick heart anatomies reconstructed using light sheet fluorescent microscopy. Additionally, we perform a quantitative analysis on geometric features using geometric morphometric methods. Together, the high-resolution but accessible imaging technique of light sheet fluorescence microscopy to reconstruct the anatomies paired with computational fluid dynamics modeling and geometric morphometrics methods produces a fast and accessible pipeline for quantitative hemodynamic and anatomical analysis in embryonic heart development.

developmental biology↗

GPT-4 based AI agents - the new expert system for detection of antimicrobial resistance mechanisms?

BackgroundEUCAST recommends a two-step process for beta-lactamases in Gram-negative bacteria. Screening with minimal inhibitory concentrations (MICs) or inhibition zone diameters for potential extended spectrum beta-lactamase (ESBL), plasmid-mediated AmpC beta-lactamase, or carbapenemase production is followed by confirmatory tests. GPT-4 and its newly released customized GPT-agent may support the initial EUCAST-screening process. We aimed to validate a customized GPT-agent to identify potential resistance mechanisms. MethodsWe used 225 Gram-negative isolates. Based on phenotypic resistances against beta-lactam antibiotics, we formed four categories: "none", "ESBL", "AmpC", or "carbapenemase". We included 862 phenotypic categories. Next, we customized a GPT-agent with EUCAST-guidelines, expert rules, and EUCAST-breakpoint table (v13.1). We compared routine diagnostic outputs (reference) to (i) EUCAST-GPT-expert, (ii) medical microbiologists, and (iii) GPT-4 without customization. We determined performance as sensitivities and specificities to flag suspect resistance mechanisms. ResultsThree human readers showed concordance in 814/862 (94.4%) phenotypic categories and used in median eight words (IQR 4-11) for reasoning. Median sensitivity and specificity for ESBL, AmpC, and carbapenemase were 98%/99.1%, 96.8%/97.1%, and 95.5%/98.5%, respectively. Three independent prompting rounds of the GPT-agent showed concordance in 706/862 (81.9%) categories but used in median 158 words (IQR 140-174) for reasoning. Median sensitivity and specificity for ESBL, AmpC, and carbapenemase prediction were 95.4%/69.23%, 96.9%/86.3%, and 100%/98.8%, respectively. In the non-customized GPT-4, 169/862 (19.6%) categories could be interpreted. Of these 137/169 (81.1%) categories agreed with routine diagnostic. The non-customized GPT-4 used in median 85 words (IQR 72-105) for reasoning. ConclusionHuman experts showed higher concordance and shorter argumentations compared to GPT-agents. Human experts showed comparable median sensitivities and higher specificities compared to GPT-agents. GPT-agents showed more unspecific flagging of ESBL and AmpC, potentially, resulting in additional testing, diagnostic delays, and higher costs. GPT-4 and GPT-agents are not IVDR/FDA-approved, but validation of LLMs is critical and datasets for benchmarking are needed.

microbiology↗

A Hedgehog-FGF signaling axis patterns anterior mesoderm during gastrulation

The application of single cell technologies to early development holds promise for resolving complex developmental phenotypes. Here we define a novel role for Hedgehog (Hh) signaling for the formation of anterior mesoderm lineages during gastrulation. Single-cell transcriptome analysis of Hh-deficient mesoderm revealed selective deficits in anterior mesoderm populations that later translate to physical defects to anterior embryonic structures including the first pharyngeal arch, heart, and anterior somites. We found that Hh-dependent anterior mesoderm defects were cell non-autonomous to Hh-signal reception. Transcriptional profiling of Hh-deficient mesoderm during gastrulation revealed disruptions to both transcriptional patterning of the mesoderm and a key FGF signaling pathway for mesoderm migration. FGF4 protein application was able to restore cellular migration during gastrulation that was decreased by Hh pathway antagonism. These findings implicate that primitive streak-mediated regulation of anterior mesoderm patterning is controlled by a multicomponent signaling hierarchy activated by Hh signaling and executed by FGF signal transduction.\n\nSIGNIFICANCE STATEMENTHow signaling events during gastrulation pattern the mesoderm is a fascinating developmental process. Although Hedgehog signaling has been implicated in early mesoderm development, its mechanistic role has not been described. We applied single cell sequencing to describe mesodermal defects in Hedgehog pathway mutants--revealing selective defects in anterior mesoderm populations. Transcriptional profiling of gastrulating Hedgehog mutants indicated that several pathways essential for primitive streak function, including FGF, required Hh signaling. Blocking Hedgehog signaling abrogated cell migration during gastrulation, which could be mitigated by addition of FGF4 ligand. This work uncovers a novel Hedgehog to FGF signaling event and describes a unique mechanism by which signals from the node impact to anterior mesoderm formation through the modulation of primitive streak function.

developmental biology↗

Optimized Cas9 expression systems for highly efficient Arabidopsis genome editing facilitate isolation of complex alleles in a single generation

Genetic resources for the model plant Arabidopsis comprise mutant lines defective in almost any single gene in reference accession Columbia. However, gene redundancy and/or close linkage often render it extremely laborious or even impossible to isolate a desired line lacking a specific function or set of genes from segregating populations. Therefore, we here evaluated strategies and efficiencies for the inactivation of multiple genes by Cas9-based nucleases and multiplexing. In first attempts, we succeeded in isolating a mutant line carrying a 70 kb deletion, which occurred at a frequency of ~1.6% in the T2 generation, through PCR-based screening of numerous individuals. However, we failed to isolate a line lacking Lhcb1 genes, which are present in five copies organized at two loci in the Arabidopsis genome. To improve efficiency of our Cas9-based nuclease system, regulatory sequences controlling Cas9 expression levels and timing were systematically compared. Indeed, use of DD45 and RPS5a promoters improved efficiency of our genome editing system by approximately 25-30-fold in comparison to the previous ubiquitin promoter. Using an optimized genome editing system with RPS5a promoter-driven Cas9, putatively quintuple mutant lines lacking detectable amounts of Lhcb1 protein represented approximately 30% of T1 transformants. These results show how improved genome editing systems facilitate the isolation of complex mutant alleles, previously considered impossible to generate, at high frequency even in a single (T1) generation.

plant biology↗