Biology subjects

Thierry, B.

Publications and source records attributed to Thierry, B..

5 recordsLinked to original sources

bioRxiv2024

3D printed microtissue cassettes enabling high throughput proton radiobiological assays

Uncertainties on proton relative biological effectiveness (RBE) across the spread out of Bragg peak (SOBP) (typically assumed to be 1.1) may lead to suboptimal treatment plan and unwarranted toxicity to organs-at-risk. Herein, we report a reliable analytical method to determine the proton RBE along the SOBP and distal fall-off region. The 3D microtissue cassette enables the high throughput assessment of biological assays including clonogenic assay and {gamma}-H2AX assay following a single proton irradiation. Clonogenic assay shows the RBE of 1.6 (10% cellular survival) which is consistent with the deter-mined RBE of 1.58 using the {gamma}-H2AX assay. Besides, we also show that the high spatial resolution of the cassette can distinguish the minute but significant foci changes (number, area) in response to small proton radiation dose fraction. The results validate the reliability of our setup in addressing critical proton radiobiological questions.

bioengineering↗

bioRxiv2024

A pumpless and tubeless microfluidic device enables extended in vitro development of Cryptosporidium parvum

The enteric parasite Cryptosporidium remains a treatment challenge for drinking water utilities globally due to its resistance to chlorine disinfection. However, the lack of an in vitro culture system for Cryptosporidium that is both cost-effective and reliable remains a key bottleneck in Cryptosporidium research. Here we report that the microfluidic culture of HCT-8 cells under fluid shear stress enables the extended development of Cryptosporidium parvum. Specifically, the growth of C. parvum in a user-friendly pumpless microfluidic device was assessed using immunofluorescence assays, scanning electron microscopy and quantitative PCR, which revealed that development peaked at six days post-infection but continued for ten days in total. Oocysts produced within the microfluidic device were infective to fresh HCT-8 monolayers, however these oocysts were only present at low levels. We anticipate that such microfluidic approaches will facilitate a wide range of in vitro studies on Cryptosporidium and may have the potential to be further developed as a routine infectivity assessment tool for the water industry.

microbiology↗

bioRxiv2022

Preclinical assessment of Fibroblast Activation Protein-specific MRI as an alternative to PSMA for image-guided focal therapy of prostate cancer

Accurate and precise delineation of gross tumour volumes remains a barrier to radiotherapy dose escalation and boost dosing in the treatment of solid tumours, such as prostate cancer. Magnetic resonance imaging of tumour molecular targets has the power to enable focal dose boosting, particularly when combined with technological advances such as MRI-LINAC. Fibroblast activation protein (FAP) is a transmembrane protein overexpressed in stromal components of >90% of epithelial carcinomas. Herein we compare targeted MRI of gold standard PSMA with FAP in the delineation of orthotopic tumours in a mouse model of prostate cancer. Control (no ligand), FAP and PSMA-targeting iron oxide nanoparticles were prepared with modification of an MRI agent (FerroTrace). Mice with orthotopic LNCaP tumours underwent T2-weighted 3D MRI 24 hours after intravenous injection of contrast agents. FAP and PSMA nanoparticles produced contrast enhancement on MRI when compared to control nanoparticles, which was most pronounced on the tumour periphery. FAP-targeted MRI increased the proportion of tumour contrast enhancing black pixels by 13.37% when compared to PSMA. Furthermore, analysis of changes in R2 values between healthy prostates and LNCaP tumours indicated an increase in contrast enhancing pixels in the tumour border of 15%, when targeting FAP, in contrast to PSMA This study demonstrates preclinical feasibility of PSMA and FAP-targeted MRI which can enable targeted image-guided focal therapy of localized prostate cancer.

bioengineering↗

bioRxiv2022

Magnetic Enrichment of Immuno-Specific Extracellular Vesicles for Mass Spectrometry Using Biofilm-Derived Iron Oxide Nanowires

Immuno-specific enrichment of extracellular vesicles (EVs) can provide important information into cellular pathways underpinning various pathologies and for non-invasive diagnostics, including mass spectrometry-based analyses. Herein, we report an optimised protocol for immuno-magnetic enrichment of specific EV subtypes and their subsequent processing with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Specifically, we conjugated placental alkaline phosphatase (PLAP) antibodies to magnetic iron oxide nanowires (NWs) derived from bacterial biofilms and demonstrated the utility of this approach by enriching placental specific EVs (containing PLAP) from cell culture media. We demonstrate efficient PLAP+ve EV enrichment for both NW-PLAP and Dynabeads-PLAP, with high PLAP protein recovery (83.7{+/-}8.9% and 83.2{+/-}5.9%, respectively), high particle-to-protein ratio (7.5{+/-}0.7x109 and 7.1{+/-}1.2x109, respectively), and low non-specific binding of non-target EVs (7{+/-}3.2% and 5.4{+/-}2.2%, respectively). Furthermore, our optimized EV enrichment and processing approach identified 2518 and 2545 protein groups with LC-MS/MS for NW-PLAP and Dynabead-PLAP, respectively, with excellent reproducibility (Pearson correlation 0.986 and 0.988). These findings demonstrate that naturally occurring iron oxide NWs have comparable performance to current gold standard immune-magnetic beads. The optimized immunospecific EV enrichment for LC-MS/MS method provides a low-cost and highly-scalable yet efficient, high-throughput approach for quality EV proteomic studies.

bioengineering↗

bioRxiv2021

Optical Cellular Micromotion: A New Paradigm to Measure Tumour Cells Invasion in 3D Tumour Environments

Measuring tumour cell invasiveness through three-dimensional (3D) tissues, particularly at the single cell level, can provide important mechanistic understanding and assist in identifying therapeutic targets of tumour invasion. However, current experimental approaches, including standard in vitro invasion assays, have limited physiological relevance and offer insufficient insight about the vast heterogeneity in tumour cell migration through tissues. To address these issues, here we report on the concept of optical cellular micromotion, where digital holographic microscopy (DHM) is used to map the optical thickness fluctuations at sub-micron scale within single cells. These fluctuations are driven by the dynamic movement of subcellular structures including the cytoskeleton and inherently associated with the biological processes involved in cell invasion within tissues. We experimentally demonstrate that the optical cellular micromotion correlates with tumour cells motility and invasiveness both at the population and single cell levels. In addition, the optical cellular micromotion significantly reduced upon treatment with migrastatic drugs that inhibit tumour cell invasion. These results demonstrate that micromotion measurements can rapidly and non-invasively determine the invasive behaviour of single tumour cells within tissues, yielding a new and powerful tool to assess the efficacy of approaches targeting tumour cell invasiveness. Significance StatementTumour cells invasion through tissues is a key hallmark of malignant tumour progression and its measurement is essential to unraveling biological processes and screening for new approaches targeting cell motility. To address the limitations of current approaches, we demonstrate that sub-micron scale mapping of the dynamic optical thickness fluctuations within single cells, referred to as optical cellular micromotion, correlates with their motility in ECM mimicking gel, both at the population and single cell levels. We anticipate that 3D optical micromotion measurement will provide a powerful new tool to address important biological questions and screen for new approaches targeting tumour cell invasiveness.

bioengineering↗