Development of an automated 3D high content cell screening platform for organoid phenotyping.

The use of organoid models in biomedical research has grown substantially since their inception. As they gain popularity among scientists seeking more complex and biologically relevant systems, there is a direct need to expand and clarify potential uses of such systems in diverse experimental contexts. Herein we outline a high-content screening (HCS) platform that allows researchers to screen drugs or other compounds against three-dimensional (3D) cell culture systems in a multi-well format (384-well). Furthermore, we compare the quality of robotic liquid handling with manual pipetting and characterize and contrast the phenotypic effects detected by confocal imaging and biochemical assays in response to drug treatment. We show that robotic liquid handling is more consistent and amendable to high throughput experimental designs when compared to manual pipetting due to improved precision and automated randomization capabilities. We also show that image-based techniques are more sensitive to detecting phenotypic changes within organoid cultures than traditional biochemical assays that evaluate cell viability, supporting their integration into organoid screening workflows. Finally, we highlight the enhanced capabilities of confocal imaging in this organoid screening platform as they relate to discerning organoid drug responses in single-well co-cultures of organoids derived from primary human biopsies and patient-derived xenograft (PDX) models. Altogether, this platform enables automated, imaging-based HCS of 3D cellular models in a non-destructive manner, opening the path to complementary analysis through integrated downstream methods.

Effect of Filtered Blood Return on Outcomes of Pulmonary Aspiration Thrombectomy.

OBJECTIVES: To assess the effects of filtered blood reinfusion (FBR) on procedural outcomes of aspiration thrombectomy for pulmonary embolism (PE). MATERIALS AND METHODS: A total of 171 patients who underwent aspiration thrombectomy for intermediate-high-risk or high-risk PE between December 2018 and September 2022 were included, 84 of whom underwent thrombectomy with FBR and 87 without. Demographic data, vital signs, laboratory values, procedural details, pulmonary arterial pressures, transfusion needs, length of hospital stay, and procedure-related adverse events were recorded. RESULTS: The groups did not differ at baseline, other than the FBR cohort having a higher percentage of women. There was no significant difference in postprocedural vital signs or pulmonary arterial pressure. Mean fluoroscopy time and volume of contrast medium used were lower in the FBR cohort. The drop in hemoglobin level was lower in the FBR group at both 12 (FBR, -1.065; No FBR, -1.742; P > .001) and 24 hours (FBR, -1.526; No FBR, -2.380; P > .001) after procedure; accordingly, fewer patients required transfusions in the FBR cohort (FBR, 8 (9.5%); No FBR, 20 (23.0%); P = .016). There was no difference in the number or severity of adverse events or duration of intensive care unit or hospital admission. CONCLUSIONS: FBR use during aspiration pulmonary thrombectomy reduced blood loss and transfusion requirements but had no significant effect on procedural success or adverse event rates.

Single-cell analysis of endometriosis reveals a coordinated transcriptional programme driving immunotolerance and angiogenesis across eutopic and ectopic tissues.

Endometriosis is characterized by the growth of endometrial-like tissue outside the uterus. It affects many women during their reproductive age, causing years of pelvic pain and potential infertility. Its pathophysiology remains largely unknown, which limits early diagnosis and treatment. We characterized peritoneal and ovarian lesions at single-cell transcriptome resolution and compared them to matched eutopic endometrium, unaffected endometrium and organoids derived from these tissues, generating data on over 122,000 cells across 14 individuals. We spatially localized many of the cell types using imaging mass cytometry. We identify a perivascular mural cell specific to the peritoneal lesions, with dual roles in angiogenesis promotion and immune cell trafficking. We define an immunotolerant peritoneal niche, fundamental differences in eutopic endometrium and between lesion microenvironments and an unreported progenitor-like epithelial cell subpopulation. Altogether, this study provides a holistic view of the endometriosis microenvironment that represents a comprehensive cell atlas of the disease in individuals undergoing hormonal treatment, providing essential information for future therapeutics and diagnostics.