Protocol for rapid obtention and fractionation of anaerobic bacterial conditioned media to study calcium signaling in enteroendocrine cells.

STAR Protoc

Division of Biology and Biological Engineering, California Institute of Technology, 1200 E. California Boulevard, Pasadena, CA, USA. Electronic address:

Published: September 2022

Gut microbiota influences neurodevelopment, behavior and contributes to neurodegenerative disorders. One possible mechanism is the direct modulation of calcium (Ca) signaling and protein homeostasis in enteroendocrine cells (EECs), a component of the gut epithelium. Here, we present a protocol to isolate fractions of conditioned media (CM) from the anaerobic bacteria and the utilization of this CM to monitor Ca fluctuation in EECs by imaging. This protocol can be adapted and applied to various bacterial cultures and cell types. For complete details on the use and execution of this protocol, please refer to Amorim Neto et al. (2022).

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9214868PMC
http://dx.doi.org/10.1016/j.xpro.2022.101486DOI Listing

Publication Analysis

Top Keywords

conditioned media
8
calcium signaling
8
enteroendocrine cells
8
protocol
4
protocol rapid
4
rapid obtention
4
obtention fractionation
4
fractionation anaerobic
4
anaerobic bacterial
4
bacterial conditioned
4

Similar Publications

Background: Bacteria in physiological environments can generate mineralizing biofilms, which are associated with diseases like periodontitis or kidney stones. Modelling complex environments presents a challenge for the study of mineralization in biofilms. Here, we developed an experimental setup which could be applied to study the fundamental principles behind biofilm mineralization on rigid substrates, using a model organism and in a tailored bioreactor that mimics a humid environment.

View Article and Find Full Text PDF

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous group of immature myeloid cells playing a critical role in immune suppression. In vitro-generated MDSCs are a convenient tool to study the properties of tumour-associated MDSCs. Here, we compared six protocols for in vitro generation of functional mouse MDSCs from bone marrow progenitors.

View Article and Find Full Text PDF

Using a murine osteomyelitis model, we recently demonstrated that and mutants generated in the USA300 strain LAC are attenuated to a greater extent than an isogenic mutant and that this can be attributed to a significant extent to the increased production of extracellular proteases in both mutants. Based on this, we used a mass-based proteomics approach to compare the proteomes of LAC, its isogenic , , and mutants, and isogenic derivatives of all four of these strains unable to produce the extracellular proteases aureolysin, SspA, SspB, ScpA, or SplA-F. This allowed us to identify proteins that were present in reduced amounts in , and / mutants owing to the increased production of extracellular proteases.

View Article and Find Full Text PDF

Cutaneous T-cell lymphoma (CTCL) is a rare T-cell malignancy characterized by inflamed and painful rash-like skin lesions that may affect large portions of the body's surface. Patients experience recurrent infections due to a compromised skin barrier and generalized immunodeficiency resulting from a dominant Th2 immune phenotype of CTCL cells. Given the role of the unfolded protein response (UPR) in normal and malignant T-cell development, we investigated the impact of UPR-inducing drugs on the viability, transcriptional networks, and Th2 phenotype of CTCL.

View Article and Find Full Text PDF

Microvessel co-transplantation improves poor remuscularization by hiPSC-cardiomyocytes in a complex disease model of myocardial infarction and type 2 diabetes.

Stem Cell Reports

January 2025

Toronto General Hospital Research Institute, University Health Network, 101 College St., Toronto, ON M5G 1L7, Canada; Institute of Biomedical Engineering, University of Toronto, Toronto, ON, Canada; Laboratory of Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada; Heart & Stroke/Richard Lewar Centre of Excellence, University of Toronto, Toronto, ON, Canada; Ajmera Transplant Center, University Health Network, Toronto, ON, Canada. Electronic address:

People with type 2 diabetes (T2D) are at a higher risk for myocardial infarction (MI) than age-matched healthy individuals. Here, we studied cell-based cardiac regeneration post MI in T2D rats modeling the co-morbid conditions in patients with MI. We recapitulated the T2D hallmarks and clinical aspects of diabetic cardiomyopathy using high-fat diet and streptozotocin in athymic rats, which were then subjected to MI and intramyocardial implantation of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with or without rat adipose-derived microvessels (MVs).

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!