Biology subjects

Chiappini, C.

Publications and source records attributed to Chiappini, C..

8 recordsLinked to original sources

bioRxiv2026

Immune reprogramming of 3D tumor models via optoporation-mediatedtargeted gene delivery to macrophages

The dynamics of the tumor microenvironment (TME) are a key determinant of cancer progression and therapeutic resistance through complex interactions between tumor, stromal and immune cell populations. Among these, tumor-associated macrophages (TAMs) play a central role in promoting tumor growth and immune suppression. However, the specific contributions of TAMs remain poorly understood due to the lack of tools enabling selective genetic manipulation in three-dimensional (3D) tumor models. Here, we present a gold nanoparticle-assisted optoporation approach that enables spatially selective plasmid-based gene delivery to TAMs within intact heterocellular 3D pancreatic ductal adenocarcinoma (PDAC) spheroids, thereby modulating the TME. In two-dimensional (2D) TAM cultures, conventional transfection of IRF5- and IKBKB-encoding plasmids validated their capacity to induce TAM repolarization, as evidenced by activation of interferon signaling. Extending this approach to 3D PDAC spheroids, nanoparticle-assisted optoporation achieved selective transfection of TAMs with IRF5- and IKBKB-encoding plasmids by transiently generating nanoscale membrane pores in illuminated cells. TAMs transfection elicited a robust interferon response, marked by transcriptional upregulation of IFNA, IFNB1, and CXCL10, and increased protein levels of IFNB1, IFNL1, and CXCL13, together with downregulation of pro-tumorigenic markers CEACAM5, IL19, and IL32. These coordinated changes indicate a shift towards an anti-tumorigenic TME. By enabling minimally invasive, TAM-specific gene delivery in complex multicellular 3D spheroids, this strategy allows precise modulation of the TME and opens new avenues for modeling its dynamics in cancer progression and therapeutic response.

bioengineering↗

bioRxiv2025

Morphomolecular Longitudinal Endoscopy of Carcinogenesis

Current diagnostic approaches for head and neck cancer rely primarily on white-light endoscopy and tissue biopsy, providing only static, episodic assessments that fail to capture dynamic changes in tumour biology over time. This absence of continuous, detailed molecular and microscopic structural information makes early identification of dysplasia or cancer difficult and can delay critical treatment. Here, we introduce a unified, label-free endoscopic platform that combines Raman spectroscopy (RS) and optical coherence tomography (OCT) within a miniaturised, forward-viewing fibre-optic probe. This integration enables simultaneous video-rate OCT imaging and subsecond RS acquisition linking morphological and molecular tissue trajectories in vivo. We developed an explainable AI-based fusion framework that learns joint temporal representations of biochemical and structural features to classify tissue states while highlighting the most influential predictors. In a longitudinal 4NQO-induced murine model of carcinogenesis, the system tracked epithelial transformation in real-time from hyperplasia through dysplasia to neoplasia. By capturing temporally resolved morphomolecular trajectories in vivo, this platform establishes a new paradigm for early cancer detection and real-time surveillance, supporting timely and individualised clinical decision-making.

bioengineering↗

bioRxiv2024

NoButter: An R package for reducing transcript dis-persion in CosMx Spatial Molecular Imaging Data

MotivationAdvances in spatial transcriptomics technologies at single-cell resolution have high-lighted the need for innovative quality assessment approaches and improved analytical tools. Imaging-based spatial transcriptomics technologies, such as the CosMx Spatial Molecular Imager (SMI), provide the location and abundance of transcripts through multifocal imaging. Optical sections (or Z-slices) form a Z-stack that represents the tissue depth. Transcript dispersion can be observed across these Z-slice and introduce considerable levels of technical noise to the data that can negatively impact downstream analysis. Package FunctionalityNoButter is an R package designed to evaluate transcript dispersion in CosMx SMI spatial transcriptomics data. Using the raw data, the transcript distribution is assessed for each Z-slice of a Z-stack across multiple fields of views (FOVs). To systematically identify transcript dispersion, the percentage of transcripts located outside cell boundaries is calculated. Z-slices exhibiting high levels of transcript dispersion can be excluded, while high-confidence transcripts are preserved. Usage ScenarioTo demonstrate the functionalities of NoButter, spatial transcriptomics data was generated using the CosMx SMI for lymph node tissue, a lung sample, and two triple-negative breast cancers (TNBCs). Use cases illustrate substantial transcript dispersion in optical planes closer to the glass slide. In these Z-slices, on average, an additional 10% of the transcripts were discarded using NoButter. Cleaning such Z-slices with high dispersion rates reduces technical noise and improves the overall quality of the spatial transcriptomics data. AvailabilityThe package can be accessed at https://github.com/cancerbioinformatics/NoButter.

bioinformatics↗

bioRxiv2023

Opto-lipidomics of tissues

Lipid metabolism and signalling play pivotal functions in biology and disease development. Despite this, there is currently no optical technique available that can directly visualise the lipidome in tissues. In this study, we introduce opto-lipidomics, a new approach to optical molecular tissue imaging. We expand the capability of vibrational Raman spectroscopy to identify individual lipids in complex tissue matrices through correlation with desorption electrospray ionisation (DESI) - mass spectrometry imaging in an integrated instrument. A computational pipeline of inter-modality regression analysis is established to extract lipidomic information from optical vibrational spectra. Opto-lipidomic imaging of transient cerebral ischemia-reperfusion injury in a murine model of ischemic stroke demonstrates the visualisation and identification of lipids in disease with unprecedented molecular specificity using light. Furthermore, we deploy opto-lipidomics in a handheld fiber-optic Raman probe and demonstrate real-time classification of bulk brain tissues based on specific lipid abundances. Opto-lipidomics opens a host of opportunities to study lipid biomarkers for diagnostics, prognostics, and novel therapeutic targets.

bioengineering↗

bioRxiv2023

Nondestructive Spatial Lipidomics for Glioma Classification

Mapping the molecular composition of tissues using spatial biology provides high-content information for molecular diagnostics. However, spatial biology approaches require invasive procedures to collect samples and destroy the investigated tissue, limiting the extent of analysis, particularly for highly functional tissues such as those of the brain. To address these limitations, we developed a workflow to harvest biomolecules from brain tissues using nanoneedles and characterise the distribution of lipids using desorption electrospray ionization mass spectrometry imaging. The nanoneedles preserved the original tissue while harvesting a reliable molecular profile and retaining the original lipid distribution for mouse and human brain samples, accurately outlining the morphology of key regions within the brain and tumour lesions. The deep neural network analysis of a cohort containing 23 human glioma biopsies showed that nanoneedle samples maintain the molecular signatures required to accurately classify disease state. Thus, nanoneedles provide a route for tissue-preserving spatial lipidomic and molecular diagnostics.

bioengineering↗

bioRxiv2022

Nanoneedles for targeted siRNA silencing of p16 in the Human Corneal Endothelium

Nanoneedles can target nucleic acid transfection to primary cells at tissue interfaces with high efficiency and minimal perturbation. The corneal endothelium is an ideal target for nanoneedle-mediated RNAi aimed at enhancing its proliferative capacity, necessary for tissue regeneration. Here we develop a strategy for siRNA nanoninjection of the human corneal endothelium. We show that nanoneedles can deliver p16-targeting siRNA to primary human corneal endothelial cells in vitro without toxicity. The nanoinjection of siRNA induces p16 silencing and increases cell proliferation, as monitored by ki67 expression. Furthermore, siRNA nanoinjection targeting the human corneal endothelium is non-toxic ex vivo and silences p16 in transfected cells. These data indicate that nanoinjection can support targeted RNAi therapy for the treatment of endothelial corneal dysfunction.

bioengineering↗

bioRxiv2021

Axonal length determines distinct homeostatic phenotypes in human iPSC derived motor neurons on a bioengineered platform

Stem cell-based experimental platforms for neuroscience can effectively model key mechanistic aspects of human development and disease. However, conventional culture systems often overlook the engineering constraints that cells face in vivo. This is particularly relevant for neurons covering long range connections such as spinal motor neurons (MNs). The axons of these neurons extend up to 1m in length and require a complex interplay of mechanisms to maintain cellular homeostasis. It follows that shorter axons in conventional cultures may not faithfully capture important aspects of their longer counterparts. Here we directly address this issue by establishing a bioengineered platform to assemble arrays of human axons ranging from micrometers to centimeters, permitting systematic investigation of the effects of length on human axonal biology for the first time. With this approach, we reveal a link between length and metabolism in human MNs in vitro, where axons above a "threshold" size induce specific molecular adaptations in cytoskeleton composition, functional properties, local translation and mitochondrial homeostasis. Our findings specifically demonstrate the existence of a length-dependent mechanism that switches homeostatic processes within human MNs in order to sustain long axons. Our findings have critical implications for in vitro modelling of several neurodegenerative disorders and reinforce the importance of modelling cell shape and biophysical constraints with fidelity and precision in vitro.

neuroscience↗

bioRxiv2020

Epicardial slices: a 3D organotypic model for the study of epicardial activation and differentiation

The epicardium constitutes an untapped reservoir for cardiac regeneration. Upon myocardial injury, the adult epicardium re-activates the embryonic program, leading to epithelial-to-mesenchymal transition, migration and differentiation. Despite some successes in harnessing the epicardial therapeutic potential by thymosin {beta}4 (T{beta}4) pre-treatment, further translational advancements are hampered by the paucity of representative experimental models. Here we apply innovative protocols to obtain living 3D organotypic slices from porcine hearts, encompassing the epicardial/myocardial interface. In culture, our slices preserve the in vivo architecture and functionality, presenting a continuous epicardium overlaying a healthy and connected myocardium. Upon T{beta}4 treatment of the slices, the epicardial cells become activated upregulating embryonic and EMT genes and invading the myocardium where they differentiate towards the mesenchymal lineage. Our 3D organotypic model enables to investigate the reparative potential of the adult epicardium, offering a new tool to explore ex vivo the complex 3D interactions occurring within the native heart environment.

bioengineering↗