Cilia: From Mechanisms to Disease–Part A
- 1st Edition, Volume 175 - March 21, 2023
- Editors: Lorenzo Galluzzi, Jose Manuel Bravo-San Pedro
- Language: English
- Hardback ISBN:9 7 8 - 0 - 4 4 3 - 1 8 5 8 6 - 1
- eBook ISBN:9 7 8 - 0 - 4 4 3 - 1 8 5 8 7 - 8
Cilia: From Mechanisms to Disease, Part A, Volume 175 in the Methods in Cell Biology series, offers a range of techniques and protocols that can be used to study aspects of this… Read more
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Request a sales quoteCilia: From Mechanisms to Disease, Part A, Volume 175 in the Methods in Cell Biology series, offers a range of techniques and protocols that can be used to study aspects of this interesting cellular organelle both in vitro and in vivo. Sections in this new release include Protocols to induce and study ciliogenesis, Flow Cytometry-Based Approach for the study of primary Cilia, Microscopic observation of human airway ciliary movement using wheat germ agglutinin, Time-lapse imaging of primary cilia behavior with physiological expression of fluorescent ciliary proteins, Evaluation of ciliary-GPCR dynamics using a validated organotypic brain slice culture method, and much more.
Other sections cover Studying the morphology, composition and function of the photoreceptor primary cilium in zebrafish, Visualizing Multiciliated Cells in the Zebrafish, Isolation of Ciliary Ectosomes and Analysis of Peptide-mediated Chemotaxis in Chlamydomonas, Using Paramecium as a Model for Ciliopathies, Using organoids to study cilia, Using in vivo cerebellar electroporation to study neuronal cell proliferation and differentiation in a Joubert syndrome mouse model, and more.
- Offers a detailed overview of the protocols used to study cilia structure and various aspects of ciliary function
- Provides an approach to the study of some diseases related to ciliary dysfunction, also known as ciliopathies
- Written in an accessible style by renowned experts in the field
Students and entry-level scientists who are for the first time approaching the study of cilia, as well as experienced researchers.
- Cover image
- Title page
- Table of Contents
- Copyright
- Contributors
- Cilia: From Mechanisms to Disease - Part A
- Acknowledgments
- References
- Chapter 1: Protocols to induce and study ciliogenesis
- Abstract
- 1: Introduction
- 2: Material
- 3: Treatments to induce ciliogenesis in vitro
- 4: Cilia visualization
- 5: Quantification and statistical analysis
- 6: Conclusion remarks
- 7: Notes
- Acknowledgments
- References
- Chapter 2: A flow cytometry-based approach for the study of primary cilia
- Abstract
- 1: Introduction
- 2: Tissue dissection and preparation
- 3: Deciliation
- 4: Staining
- 5: FACS analysis
- 6: Cilia sorting and treatment for subsequent proteomic analysis
- 7: Troubleshooting
- 8: Concluding remarks
- References
- Chapter 3: Microscopic observation of human airway ciliary movement using wheat germ agglutinin
- Abstract
- 1: Introduction
- 2: Preparation of tracheal sample
- 3: Observation of ciliary beating
- 4: Analysis of the orientation of ciliary beating
- 5: Analysis of ciliary beating frequency
- 6: Concluding remarks
- 7: Notes
- References
- Chapter 4: Time-lapse imaging of primary cilium behavior with physiological expression of fluorescent ciliary proteins
- Abstract
- 1: Introduction
- 2: Generation of knock-in cells
- 3: Cell culture
- 4: Image acquisition
- 5: Image processing
- 6: Data quantification
- 7: Concluding remarks
- 8: Notes
- Acknowledgments
- References
- Chapter 5: Evaluation of ciliary-GPCR dynamics using a validated organotypic brain slice culture method
- Abstract
- 1: Introduction
- 2: Slice cultures
- 3: Evaluation of ciliary dynamics
- 4: Concluding remarks
- Acknowledgment
- References
- Chapter 6: High-efficient CRISPR/Cas9-mediated gene targeting to establish cell models of ciliopathies
- Abstract
- 1: Introduction
- 2: Construction of plasmids
- 3: Cell culture
- 4: Plasmids transfection and drug screening
- 5: Genotyping of drug-resistant clones
- 6: Validation of established clones by immunofluorescence microscopy
- 7: Concluding remarks
- Acknowledgments
- References
- Chapter 7: Studying the morphology, composition and function of the photoreceptor primary cilium in zebrafish
- Abstract
- 1: Introduction
- 2: Basic buffers and solutions used in multiple protocols
- 3: Histological analysis of retinal lamination and photoreceptor integrity on plastic-embedded sections and cryosections with immunofluorescence
- 4: Ultrastructural visualization of photoreceptors using transmission electron microscopy and correlative light and electron microscopy
- 5: Electrophysiological assessment of retinal function
- 6: Molecular examination of isolated photoreceptor outer segments
- 7: Conclusion
- References
- Chapter 8: Visualizing multiciliated cells in the zebrafish
- Abstract
- 1: Introduction
- 2: Zebrafish models of ciliopathic disease
- 3: Development of MCCs in the zebrafish pronephros
- 4: Methods to assess MCC development
- 5: Conclusion
- Acknowledgments
- References
- Chapter 9: Isolation of ciliary ectosomes and analysis of amidated peptide-mediated chemotaxis in Chlamydomonas
- Abstract
- 1: Introduction
- 2: Isolation of ciliary ectosomes from mating gametes
- 3: Microfluidic assay to assess chemotaxis in C. reinhardtii
- 4: Concluding remarks
- Acknowledgment
- References
- Chapter 10: Methods for Paramecium tetraurelia ciliary membrane protein identification and function
- Abstract
- 1: Introduction
- 2: Triton X-114-phase separation for the analysis of membrane proteins by mass spectrometry
- 3: Analysis of detergent resistant membranes (DRMs) using sucrose and OptiPrep™ density gradient ultracentrifugation
- 4: Epitope tagging, expression of proteins, and immunofluorescence
- 5: RNA interference (RNAi)
- 6: Semi-quantitative reverse transcriptase PCR, SEM, and TEM
- 7: Genetic crosses
- 8: Electrophysiology to measure membrane potential (Vm)
- 9: Conclusions
- Acknowledgment
- References
- Chapter 11: Using mammary organoids to study cilia
- Abstract
- 1: Introduction
- 2: Buffers
- 3: Mammary stem cell isolation
- 4: Mammary organoid culture and long-term maintenance
- 5: Immunofluorescence staining of primary cilia
- 6: Concluding remarks
- 7: Notes
- Acknowledgments
- References
- Chapter 12: Using in vivo cerebellar electroporation to study neuronal cell proliferation and differentiation in a Joubert syndrome mouse model
- Abstract
- 1: Introduction
- 2: In vivo cerebellar electroporation
- 3: Immunostaining markers for studying neuronal differentiation
- 4: Edu incorporation and cell-cycle analysis
- 5: Concluding remarks
- Acknowledgments
- References
- No. of pages: 268
- Language: English
- Edition: 1
- Volume: 175
- Published: March 21, 2023
- Imprint: Academic Press
- Hardback ISBN: 9780443185861
- eBook ISBN: 9780443185878
LG
Lorenzo Galluzzi
Lorenzo Galluzzi is Assistant Professor of Cell Biology in Radiation Oncology at the Department of Radiation Oncology of the Weill Cornell Medical College, Honorary Assistant Professor Adjunct with the Department of Dermatology of the Yale School of Medicine, Honorary Associate Professor with the Faculty of Medicine of the University of Paris, and Faculty Member with the Graduate School of Biomedical Sciences and Biotechnology of the University of Ferrara, the Graduate School of Pharmacological Sciences of the University of Padova, and the Graduate School of Network Oncology and Precision Medicine of the University of Rome “La Sapienza”. Moreover, he is Associate Director of the European Academy for Tumor Immunology and Founding Member of the European Research Institute for Integrated Cellular Pathology.
Galluzzi is best known for major experimental and conceptual contributions to the fields of cell death, autophagy, tumor metabolism and tumor immunology. He has published over 450 articles in international peer-reviewed journals and is the Editor-in-Chief of four journals:
OncoImmunology (which he co-founded in 2011), International Review of Cell and Molecular Biology, Methods in Cell biology, and Molecular and Cellular Oncology (which he co-founded in 2013). Additionally, he serves as Founding Editor for Microbial Cell and Cell Stress, and Associate Editor for Cell Death and Disease, Pharmacological Research and iScience.
Affiliations and expertise
Assistant Professor of Cell Biology in Radiation Oncology, Department of Radiation Oncology, Weill Cornell Medical College, NY, USAJB
Jose Manuel Bravo-San Pedro
Jose Manuel Bravo-San Pedro is currently a researcher at the Department of Physiology of the Complutense University of Madrid thanks to a Ramon y Cajal contract grant. He got his Ph.D. in biochemistry, cellular biology and genetics from the University of Extremadura (Caceres, Spain) in 2011, and he did a post-doctoral stage in the laboratory of Prof. Guido Kroemer. His main research interests have always been linked to autophagy, addressing this cellular process associated with neurodegenerative diseases or cancer and recently obesity and specifically related to problems in the correct functioning of the cilium. He is co-inventor of two patents and co-author of 110 publications indexed in PubMed in prestigious international journals, with h-index 45 and 23768 cites (Dec 2022).
Affiliations and expertise
Departamento de Fisiología, Facultad de Medicina, Universidad Complutense de Madrid, Madrid, SpainRead Cilia: From Mechanisms to Disease–Part A on ScienceDirect