Biology subjects

Bickle, M.

Publications and source records attributed to Bickle, M..

7 recordsLinked to original sources

bioRxiv2025

An integrated platform for high-throughput phenospace learning of 3D multilineage organoid systems

Complex multilineage organoid systems lack quantitative phenotyping methods preserving spatial architecture at high throughput. Current approaches compromise biological complexity, spatial resolution, or robust homogeneous multilineage assembly. We establish an integrated experimental-computational platform for high-throughput spatial phenotyping of multilineage organoids through developing a modular tumoroid culture system incorporating pancreatic ductal adenocarcinoma (PDAC) cells and cancer-associated fibroblasts (CAFs) in 384-well format with multiplexed whole-mount imaging. We developed Phenocoder, a machine learning framework combining conditional variational autoencoders with spatial graph analysis to extract multiscale organoid features. Rigorous validation demonstrates robust performance in PDAC tumoroids. The platform identifies pathway modulators that disrupt the fibrotic microenvironment and discovers stroma-dependent vulnerabilities, undetectable in monocultures. Extending to immuno-competent tumoroids, we assess fibrosis modulators in combination with T cell bispecific antibodies, identifying treatments that enhance immune cell proliferation and infiltration inducing cancer cell death, validated in patient-derived explants. This platform establishes a generalizable framework for multilineage organoid phenotyping.

cancer biology↗

bioRxiv2025

Modeling host-microbe interactions in immunocompetent engineered human gut tissues

The intestinal mucosal barrier contains microbial organisms within the lumen while preserving the ability to absorb nutrients. Dietary, microbial, and other exposures shaped human barrier evolution and continue to impact disease susceptibility. Here, we established engineered barrier models of the human small intestine and colon composed of a multilineage epithelium, mucus layer, accessible microbial compartment and autologous tissue-resident immune cells. The epithelium has crypt- and villus-like topological domains, with stem cells differentiating into absorptive and secretory lineages with region-specific identities. Secreted mucins accumulate apically, forming a dense mucus layer separating the epithelium from colonizing commensal and pathogenic bacteria. Intestinal memory T cells integrate into and interact with the epithelium. We use the engineered intestinal tissues to identify an epithelial gene regulatory network underlying response to Salmonella Typhimurium infection, and uncover epithelial-immune-pathogen crosstalk coordinating cytokine release and epithelial damage. Overall, this work allows for the modular integration of epithelial, microbial, and immune compartments providing a versatile system for studying human intestinal physiology and pathologies.

bioengineering↗

bioRxiv2024

A genome-wide RNA interference screening reveals protectiveness of SNX5 knockdown in a Parkinson`s disease cell model

BackgroundAlpha-synuclein is a major player in the pathophysiology of a group of diseases called synucleinopathies, which include Parkinsons disease, dementia with Lewy bodies, and multiple system atrophy. To date, there is no disease-modifying therapy available for these synucleinopathies. Furthermore, the intracellular mechanisms by which alpha-synuclein confers toxicity are not yet fully understood. Therefore, it is of utmost importance to investigate the pathophysiology of alpha-synuclein-induced toxicity in order to identify novel molecular targets for the development of disease-modifying therapies. MethodsIn the present study, we performed the first genome-wide siRNA modifier screening in a human postmitotic neuronal cell model using alpha-synuclein-induced toxicity as read-out. In a multi-step approach, we identified several genes, whose knockdown protected from alpha-synuclein-induced toxicity. The main hit was further validated by different methods, including immunofluorescence microscopy, qPCR, and Western blot. ResultsThe highest protection was achieved by knockdown of SNX5, which encodes the SNX5 protein, a component of the retromer complex. We confirmed the protective efficacy of SNX5 knockdown with an independent siRNA system. SNX5 protein is part of SNX-BAR heterodimers, which are part of the retromer complex. We found that extracellular and overexpressed intracellular alpha-synuclein led to fragmentation of the trans-Golgi network, which was prevented by SNX5 knockdown by confining alpha-synuclein in early endosomes. ConclusionIn summary, our data suggest that SNX5 plays an important role in trafficking and toxicity of alpha-synuclein. Therefore, SNX5 appears to be a possible target for therapeutic interventions in synucleinopathies.

neuroscience↗

bioRxiv2021

Quantitative intracellular retention of delivered RNAs through optimized cell fixation and immuno-staining

Detection of nucleic acids within sub-cellular compartments is key to understanding their function. Determining the intracellular distribution of nucleic acids requires quantitative retention and estimation of their association with different organelles by immunofluorescence microscopy. This is important also for the delivery of nucleic acid therapeutics which depends on endocytic uptake and endosomal escape. However, the current methods fail to preserve the majority of exogenously delivered nucleic acids in the cytoplasm. To solve this problem, by monitoring Cy5-labeled mRNA delivered to primary human adipocytes via lipid nanoparticles (LNP), we optimized cell fixation, permeabilization and immuno-staining of a number of organelle markers, achieving quantitative retention of mRNA and allowing visualization of levels which escape detection using conventional procedures. Additionally, we demonstrated the protocol to be effective on exogenously delivered siRNA, miRNA, as well as endogenous miRNA. Our protocol is compatible with RNA probes of single molecule fluorescence in-situ hybridization (smFISH) and molecular beacon, thus demonstrating that it is broadly applicable to study a variety of nucleic acids.

cell biology↗

bioRxiv2021

Sequential in vivo labeling of insulin secretory granule pools in INS-SNAP transgenic pigs

{beta}-cells produce, store and secrete insulin upon elevated blood glucose levels. Insulin secretion is a highly regulated process. The probability for insulin secretory granules to undergo fusion with the plasma membrane or being degraded is correlated with their age. However, the molecular features and stimuli connected to this behavior have not yet been fully understood. Furthermore, our understanding of {beta}-cell function is mostly derived from studies of ex vivo isolated islets and/or rodent models. To overcome this translational gap and study insulin secretory granule turnover in vivo, we have generated a transgenic pig model with the SNAP-tag fused to insulin. We demonstrate the correct targeting and processing of the tagged insulin and normal glycemic control of the pig model. Furthermore, we show specific single- and dual-color granular labeling of in vivo labeled pig pancreas. This model may provide unprecedented insights into the in vivo insulin secretory granule behavior in an animal close to humans.

cell biology↗

bioRxiv2020

Endosomal escape of delivered mRNA from endosomal recycling tubules visualized at the nanoscale

Delivery of exogenous mRNA using lipid nanoparticles (LNP) is a promising strategy for therapeutics. However, a bottleneck remains the poor understanding of the parameters that correlate with endosomal escape vs. cytotoxicity. To address this problem, we compared the endosomal distribution of six LNP-mRNA formulations of diverse chemical composition and efficacy, similar to those employed in mRNA-based vaccines, in primary human adipocytes, fibroblasts and HeLa cells. Surprisingly, we found that total uptake is not a sufficient predictor of delivery and different LNP vary considerably in endosomal distributions. Prolonged uptake impaired endosomal acidification, a sign of cytotoxicity, and caused mRNA to accumulate in compartments defective in cargo transport and unproductive for delivery. In contrast, early endocytic/recycling compartments have the highest probability for mRNA escape. By super-resolution microscopy we could resolve single LNP-mRNA within sub-endosomal compartments and capture events of mRNA escape from endosomal recycling tubules. Our results change the view of the mechanisms of endosomal escape and define quantitative parameters to guide the development of mRNA formulations towards higher efficacy and lower cytotoxicity.

bioengineering↗

bioRxiv2019

Small molecules for modulating protein driven liquid-liquid phase separation in treating neurodegenerative disease

Neurodegenerave diseases such as amyotrophic lateral sclerosis (ALS) are oten associated with mutaons in proteins that are associated with stress granules. Stress granules are condensates formed by liquid-liquid phase separaon which, when aberrant, can lead to altered condensaon behaviours and disease phenotypes. Here, we identified lipoamide, a small molecule which specifically prevents cytoplasmic condensaon of stress granule proteins. Thermal proteome profiling showed that lipoamide preferentially stabilises intrinsically disordered domain-containing proteins. These include SRSF1 and SFPQ, stress granule proteins necessary for lipoamide activity. The redox state of SFPQ correlates with its condensate-dissolving behaviour, in concordance with the importance of the dithiolane ring for lipoamide activity. In animals, lipoamide ameliorates aging-associated aggregaon of a stress granule reporter, improves neuronal morphology, and recovers motor defects caused by expression of ALS-associated FUS and TDP-43 mutants. In conclusion, lipoamide is a well-tolerated small molecule modulator of stress granule condensaon and dissection of its molecular mechanism identified a cellular pathway for redox regulaon of stress granule formaon.

cell biology↗