Biology subjects

Schmitz, A. E.

Publications and source records attributed to Schmitz, A. E..

3 recordsLinked to original sources

bioRxiv2025

An automated image analysis pipeline for wide-field optical redox imaging of patient-derived cancer organoids

Wide-field optical redox imaging provides a fast and accessible method to monitor metabolic changes in cells and has recently been developed for drug screening in patient-derived cancer organoids (PDCOs). However, manual analysis of wide-field optical redox images is inefficient and laborious for large-scale drug screens. Here, we developed an automated pipeline for PDCO segmentation, single-PDCO tracking, and background correction in autofluorescence images. This pipeline was tested on two imaging systems over a 3-day time-course with two drug doses to demonstrate generalizability across imaging systems. Segmentation was performed using a fine-tuned Cellpose model, which when compared to manual masks, achieved mean Dice scores >0.8 across systems, indicating high reproducibility. Automated single-PDCO tracking was compared to manual tracking and the accuracy of the tracking algorithm exceeded 94% by two metrics, recall and Jaccard index. For background correction, the automated pipeline uses the full field-of-view to reduce sampling bias. Compared to the manual analysis pipeline, the automated pipeline resolves single-PDCO responses with comparable sensitivity to drug treatment but with over 127x faster processing time. This novel automated image analysis pipeline improves throughput and robustness in PDCO image analysis, which increases the accessibility and scalability of wide-field optical redox imaging for PDCO drug screening.

bioengineering↗

bioRxiv2025

The role of amygdala calcitonin gene-related peptide receptors on the development of persistent bladder pain in mice.

Bladder pain significantly impacts millions worldwide, severely affecting their quality of life and posing a major clinical challenge. Understanding the mechanisms underlying persistent bladder pain is critical for developing better therapeutic strategies. In this study, we investigate the effects of cyclophosphamide (CYP)-induced persistent bladder sensitization to explore the lateralized contribution of amygdala calcitonin gene-related peptide receptors (CGRP-Rs) on pain-like changes in mice. We demonstrate that CYP induces hypersensitivity lasting up to 14 days post-injury (DPI) in the urinary bladder distention assay and up to 21 DPI when assessing abdominal mechanical sensitivity. Despite persistent pain-like changes, no significant bladder histological changes were observed. Based on previous findings that CGRP signaling from the parabrachial nucleus contributes to central amygdala (CeA) lateralization, we hypothesized that CGRP-Rs play a key role in driving visceral bladder pain-related hemispherical differences. We show that inhibiting CGRP-R activity with the antagonist CGRP8-37, in the right CeA attenuates bladder pain-like behavior, whereas left CeA inhibition sustains CYP-induced hypersensitivity. Electrophysiological recordings revealed increased firing frequency in CGRP-R positive cells in the right CeA 7 DPI. In vivo single photon calcium imaging demonstrated increased Ca transients in CGRP-R-positive cells in the right CeA, upon the presentation of a stimulus at 0 DPI, and overall at 2DPI, further confirming the pronociceptive role of CGRP-Rs in the right CeA. Taken together, these findings provide a crucial foundation for understanding pain-induced CeA lateralization and for further studies identifying how targeting CGRP signaling could provide bladder pain relief.

neuroscience↗

bioRxiv2024

Wide-field optical redox imaging with leading-edge detection for assessment of patient-derived cancer organoids

Patient-derived cancer organoids (PDCOs) are a valuable model to recapitulate human disease in culture with important implications for drug development. However, current methods for assessing PDCOs are limited. Label-free imaging methods are a promising tool to measure organoid level heterogeneity and rapidly screen drug response in PDCOs. The aim of this study was to assess and predict PDCO response to treatments based on mutational profiles using label-free wide-field optical redox imaging (WF ORI). WF ORI provides organoid-level measurements of treatment response without labels or additional reagents by measuring the autofluorescence intensity of the metabolic co-enzymes NAD(P)H and FAD. The optical redox ratio is defined as the fluorescence intensity of [NAD(P)H / NAD(P)H +FAD] which measures the oxidation-reduction state of PDCOs. We have implemented WF ORI and developed novel leading-edge analysis tools to maximize the sensitivity and reproducibility of treatment response measurements in colorectal PDCOs. Leading-edge analysis improves sensitivity to redox changes in treated PDCOs (G{Delta} = 1.462 vs G{Delta} = 1.233). Additionally, WF ORI resolves FOLFOX treatment effects across all PDCOs better than two-photon ORI, with [~]7X increase in effect size (G{Delta} = 1.462 vs G{Delta} = 0.189). WF ORI distinguishes metabolic differences based on driver mutations in CRC PDCOs identifying KRAS+PIK3CA double mutant PDCOs vs wildtype PDCOs with 80% accuracy and can identify treatment resistant mutations in mixed PDCO cultures (G{Delta} = 1.39). Overall, WF ORI enables rapid, sensitive, and reproducible measurements of treatment response and heterogeneity in colorectal PDCOs that will impact patient management, clinical trials, and preclinical drug development. Statement of SignificanceLabel-free wide-field optical redox imaging of patient-derived cancer organoids enables rapid, sensitive, and reproducible measurements of treatment response and heterogeneity that will impact patient management, clinical trials, and preclinical drug development.

bioengineering↗