Main description:

This second edition volume expands on the previous edition with updated methodologies and practical tips to overcome obstacles associated with experimentation pertaining to chemistry, biology, physiology, pathology, medical and dental sciences, and pharmacology of CCN proteins. The chapters in this book cover topics such as CCN4 immunofluorescence for tissue microarray; utilizing public molecular biological databases for CCN family research; the effects of CCN4 on pancreatic beta cell proliferation; gene expression analysis of CCNs; novel cell biological assays for measuring bone remodeling activities of CCN proteins; and the function of CCN2 in tubular epithelium cells with a focus on renal fibrogenesis. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Cutting-edge and comprehensive, CCN Proteins: Methods and Protocols, Second Edition is a valuable resource for experienced CCN researchers looking for new approaches and novice researchers just starting out in the field of CCN research.

Contents:

1. CCN Proteins - Cellular Communication Network Factors: Expanding their Repertoire toward a New Concept

Masaharu Takigawa

Part I Basic Methods and Protocols

2. Optimal CCN4 Immunofluorescence for Tissue Microarray

Pierre-Olivier Gaudreau, Isabelle Cousineau, and John Stagg

3. Cellular Fluorescence Imaging for the Evaluation of Bioactivity of CCN Family Proteins

Harumi Kawaki, Satoshi Kubota, and Masaharu Takigawa

4. Imaging of Molecular Interaction between CCN Protein and Its Binding Partners: An In Situ Proximity Ligation Assay of Interaction Between CCN2 and Rab14 in Chondrocytes

Mitsuhiro Hoshijima, Eriko Aoyama, Hiroshi Kamioka, and Masaharu Takigawa

5. Western Blot Protocols for Analysis of CCN Proteins and Fragments in Exosomes, Vesicle-free Fractions, and Cells

Kisho Ono, Yuka Okusha, Manh Tien Tran, Koki Umemori, and Takanori Eguchi

6. Comprehensive Method for Exosome Isolation and Proteome Analysis for Detection of CCN Factors in/on Exosomes

Takanori Eguchi, Yuka Okusha, Yanyin Lu, Kisho Ono, Eman A. Taha, and Shiro Fukuoka

7. Evaluation of the Molecular Interaction between CCN Protein and Its Binding Partners: A Solid-Phase Binding Assay and Surface Plasmon Resonance

Eriko Aoyama and Masaharu Takigawa

8. Protocols for Screening Peptides Binding to CCN Family Proteins and their Extended Utility

Satoshi Kubota, Harumi Kawaki, and Masaharu Takigawa

9. Transfection, Spinfection, Exofection, and Luciferase Assays for Analysis of CCN Genes Expression Mechanism

Takanori Eguchi, Yayin Lu, Eman A. Taha, and Yuka Okusha

10. Analyses of the Post-Transcriptional Regulation of CCN Genes: Approach to Multiple Steps of CCN2 Gene Expression

Seiji Kondo, Satoshi Kubota, and Masaharu Takigawa

11. Protocol for CRISPR/Cas Genome Editing for Investigating Cell Communication Network

Yuka Okusha and Takanori Eguchi

12. Utilizing Public Molecular Biological Databases for CCN Family Research

Satoshi Kubota

Part II Functional Analysis

13. Evaluation of the Effects of CCN4 on Pancreatic Beta Cell Proliferation

Rebeca Fernandez-Ruiz and Rosa Gasa

14. Analysis of CCN Functions in Liver Regeneration After Partial Hepatectomy

Naiyuan Cheng, Ki-Hyun Kim, and Lester F. Lau

15. The Evaluation of Meniscus Regenerative Effects of LIPUS-Induced CCN Proteins: Induction by LIPUS of CCN2 and Meniscus-Related Genes in Cultured Meniscus Cells and Meniscus Tissues

Yusuke Kamatsuki, Eriko Aoyama, Takayuki Furumatsu, Toshifumi Ozaki, and Masaharu Takigawa

16. Analysis of Chemotactic Property of CCN2/CTGF in Intramembranous Osteogenesis

Nobuo Takeshita and Teruko Takano-Yamamoto

17. Novel Cell Biological Assays for Measuring Bone Remodeling Activities of CCN Proteins

Takashi Nishida, Satoshi Kubota, and Masaharu Takigawa

18. Imaging Analysis of Neurotrophic Effects by CCN2 Protein in Neuronal Precursor Cells Derived from Human Induced Pluripotent Stem Cells

Masaki Shoji and Takashi Kuzuhara

19. Gene Expression Analysis of CCNs Along with Odontoblastic Differentiation In Vivo

Kazumi Kawata and Satoshi Kubota

20.Angiogenesis Assays for the Analysis of CCN Proteins

Tsuyoshi Shimo, Mari Shimatani, Akihiko Tanimura, and Masaharu Takigawa

Part III Pathophysiology

21. Using the Bleomycin-Induced Model of Fibrosis to Study the Contribution of CCN Proteins to Scleroderma Fibrosis

Alexander Peidl, John Nguyen, Pratyusha Chitturi, Bruce L. Riser, and Andrew Leask

22. Atomic Force Microscopy-Based Measurements of Retinal Microvessel Stiffness in Mice with Endothelial-Specific Deletion of CCN1

Brahim Chaqour, Maria B. Grant, Lester F. Lau, Biran Wang, Mateusz M. Urbanski, and Carmen V. Melendez-Vasquez

23. Immunohistochemical Analysis of CCN2 in Experimental Fracture Healing Models

Tsuyoshi Shimo, Hiroaki Takebe, Saki Fujii, and Akihiro Hosoya

24. Mouse Models of Tumor Bone Metastasis and Invasion for Studying CCN Proteins

Tsuyoshi Shimo, Tatsuo Okui, Naohiro Horie, Kenji Yokozeki, Masaharu Takigawa, and Akira Sasaki

25. Regulation of CCN2 and Its Bioactivity by Advanced Glycation End Products

Xiaoyu Wang, Danqing Min, and Stephen M. Twigg

26. Analysis of CCN4/WISP1 Effects on Joint Tissues Using Gain and Loss of Function Approaches

Martijn H.J. van den Bosch and Esmeralda N. Blaney Davidson

27. Generation of Conditional KO Mice of CCN2 and Its Function in the Kidney

Hideki Yokoi, Naohiro Toda, and Masashi Mukoyama

28. Analysis of the Function of CCN2 in Tubular Epithelium Cells with a Focus on Renal Fibrogenesis

Hiroaki Amano, Tsutomu Inoue, Takeru Kusano, and Hirokazu Okada