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Project description:Cystogenesis is a morphological consequence of numerous genetic diseases of the epithelium. In the kidney, the pathogenic mechanisms underlying the program of altered cell and tubule morphology are obscured by secondary effects of cyst expansion. Here, we developed a new 3D tubuloid system to isolate the rapid changes in protein localization and gene expression that correlate with altered cell and tubule morphology during cyst initiation. Mouse renal tubule fragments were pulsed with a cell differentiation cocktail including glial-derived neurotrophic factor (GDNF) to yield collecting duct-like tubuloid structures with appropriate polarity, primary cilia, and gene expression. Using the 3D tubuloid model with an inducible Pkd2 knockout system allowed the tracking of morphological, protein, and genetic changes during cyst formation. Within hours of inactivation of Pkd2 and loss of polycystin-2, we observed significant progression in tubuloid to cyst morphology that correlated with 35 differentially expressed genes, many related to cell junctions, matrix interactions, and cell morphology previously implicated in cystogenesis.This article has an associated First Person interview with the first author of the paper.
| S-EPMC7375472 | biostudies-literature
Project description:We report a significant change in epithelial differentiation pathways and principal cell specific transcription factors in tubuloids with a GDNF pulse, when compared to control tubuloid cultures.
2020-06-08 | GSE151949 | GEO
Project description:We report a profile of 35 genes that are differentially expressed in Pkd2- tubuloids compared to controls.
2020-06-08 | GSE151950 | GEO
Project description:GDNF drives rapid tubule morphogenesis in novel 3D in vitro model for ADPKD
| PRJNA637871 | ENA
Project description:GDNF drives rapid tubule morphogenesis in novel 3D in vitro model for ADPKD [GDNF RNAseq]
| PRJNA637873 | ENA
Project description:GDNF drives rapid tubule morphogenesis in novel 3D in vitro model for ADPKD [Pkd2- RNAseq]
| PRJNA637874 | ENA
Project description:Angiogenesis is fundamentally required for the initialization, development and metastatic spread of cancer. A rapidly expanding number of new experimental, chemical modulators of endothelial cell function have been described for the therapeutic inhibition of angiogenesis in cancer. Despite this expansion, there has been very limited parallel growth of in vitro angiogenesis models or experimental tools. Here we present the Responsive Angiogenic Implanted Network (RAIN)-Droplet model and novel angiogenesis assay using an endothelial cell culture model of microvascular endothelial cells encapsulated in a spontaneously self-assembling, toroidal hydrogel droplet uniquely yielding discrete, pre-formed, angiogenic networks that may be embedded in 3D matrices. On embedding, radial growth of capillary-like sprouts and cell invasion was observed. The sprouts formed not only as outgrowths from endothelial cells on the surface of the droplets, but also, uniquely, from the pre-formed network structures within the droplet. We demonstrate proof of principle for the utility of the model showing significant inhibition of sprout formation (P<0.001) in the presence of bevacizumab, an anti-angiogenic antibody. Using the RAIN-Droplet assay, we also demonstrate a novel dose-dependent pro-angiogenic function for the characteristically anti-angiogenic multi-kinase inhibitor sorafenib. Exposure of endothelial cells in 3D culture to low, non-lethal doses (<1 ?M) of sorafenib after initiation of sprouting resulted in the formation of significantly (P<0.05) more endothelial sprouts compared with controls over a 48-h period. Higher doses of sorafenib (5 ?M) resulted in a significant (P<0.05) reduction of sprouting over the same time period. The RAIN-Droplet model is a highly versatile and simply constructed 3D focal sprouting approach well suited for the study of vascular morphogenesis and for preclinical testing of drugs. Furthermore, the RAIN-Droplet model has facilitated the discovery of a novel pro-angiogenic capacity for sorafenib, which may impact the clinical application and dosing regimen of that drug.
| S-EPMC4043634 | biostudies-literature
Project description:Cultured proximal tubule epithelial cells (PTECs) under physiological flow in a 3D channel embedded within an engineered extracellular matrix (ECM) that is colocalized with an adjacent channel lined with human glomerular endothelial cells (HGECs) to mimic a peritubular capillary. After a period of maturation under continuous flow, both cell types were harvested for RNAseq analyses. Our results revealed that PTECs’ transcriptional profile is highly dependent on both the matrix on which these cells are cultured and the flow conditions. By contrast, endothelial cells exhibit greater phenotypic plasticity and are affected by matrix, flow, and co-culture conditions.
2022-03-20 | GSE198822 | GEO
Project description:Clinical trials and animal studies on the gut microbiota are often limited by the difficult access to the gut, restricted possibility of in vivo monitoring, and ethical issues. An easily accessible and monitorable in vitro model of the gut microbiota represents a valid tool for a wider comprehension of the mechanisms by which microbes interact with the host and with each other. Herein, we present a novel and reliable system for culturing the human gut microbiota in vitro. An electrospun gelatin structure was biofabricated as scaffold for microbial growth. The efficiency of this structure in supporting microbial proliferation and biofilm formation was initially assessed for five microbes commonly inhabiting the human gut. The human fecal microbiota was then cultured on the scaffolds and microbial biofilms monitored by confocal laser and scanning electron microscopy and quantified over time. Metagenomic analyses and Real-Time qPCRs were performed to evaluate the stability of the cultured microbiota in terms of qualitative and quantitative composition. Our results reveal the three-dimensionality of the scaffold-adhered microbial consortia that maintain the bacterial biodiversity and richness found in the original sample. These findings demonstrate the validity of the developed electrospun gelatin-based system for in vitro culturing the human gut microbiota.
| S-EPMC7725811 | biostudies-literature
Project description:Human in vitro-manufactured tissue and organ models can serve as powerful enabling tools for the exploration of fundamental questions regarding cell, matrix, and developmental biology in addition to the study of drug delivery dynamics and kinetics. To date, the development of a human model of the renal proximal tubule (PT) has been hindered by the lack of an appropriate cell source and scaffolds that allow epithelial monolayer formation and maintenance. Using extracellular matrices or matrix proteins, an in vivo-mimicking environment can be created that allows epithelial cells to exhibit their typical phenotype and functionality. Here, we describe an in vitro-engineered PT model. We isolated highly proliferative cells from cadaveric human kidneys (human kidney-derived cells [hKDCs]), which express markers that are associated with renal progenitor cells. Seeded on small intestinal submucosa (SIS), hKDCs formed a confluent monolayer and displayed the typical phenotype of PT epithelial cells. PT markers, including N-cadherin, were detected throughout the hKDC culture on the SIS, whereas markers of later tubule segments were weak (E-cadherin) or not (aquaporin-2) expressed. Basement membrane and microvilli formation demonstrated a strong polarization. We conclude that the combination of hKDCs and SIS is a suitable cell-scaffold composite to mimic the human PT in vitro.
| S-EPMC4074761 | biostudies-literature