A protocol for the culture and isolation of murine synovial fibroblasts

Abstract

The culture of synovial fibroblasts (SFs) is one of the most effective tools for investigating the pathology and physiology of synovial tissues and should prove useful for identifying the importance of SFs in disease as well as for the development of novel therapeutic approaches for several chronic joint diseases, such as rheumatoid arthritis. However, thus far, a detailed protocol for the primary culture and isolation of murine SFs has not been established. Therefore, the present study describes an easy and convenient method for isolating and culturing SFs from C57BL/6 mice. This protocol can be divided into 4 stages: Isolation of synovial tissues, isolation of SFs, seeding of SFs for growth in culture and purity analysis of SFs using the four cell markers, vimentin, cluster of differentiation 90.2 (CD90.2; Thy-1.2), intracellular adhesion molecule 1 (CD54) and vascular cell adhesion molecule 1 (CD106). This method is efficient and a purified population of SFs can be obtained 10 days after the initiation of culture.

Standardized Xylocarpus moluccensis fruit fraction mitigates collagen-induced arthritis in mice by regulating immune response

“…CIA‐FLS was isolated following previously reported protocol with some modification . CIA mice were used to isolate FLS.…”

“…Collagen induced arthritis-fibroblast-like synoviocyte (CIA-FLS) isolation, proliferation and apoptosis assay CIA-FLS was isolated following previously reported protocol with some modification. [22] CIA mice were used to isolate FLS. Under aseptic conditions, synovial tissues were collected and digested with 2 mg/ml collagenase type IA at 37°C for 30 min.…”

Kruppel-Like Factor 4 Positively Regulates Autoimmune Arthritis in Mouse Models and Rheumatoid Arthritis in Patients via Modulating Cell Survival and Inflammation Factors of Fibroblast-Like Synoviocyte

“…OC17TNSI0155). Murine FLS were isolated from hip joint tissue from C57BL/6 mice as previously described ( 18 ).…”

BATF regulates collagen-induced arthritis by regulating T helper cell differentiation

“…FLS were isolated from WT or Batf knockout (KO) mice and cultured as described by Zhao et al [ 23 ]. FLS of passages 4–8 were used for further analysis.…”

“…CD4 + T cell were isolated from the spleens of WT and Batf −/− mice using an EasySep™ Mouse CD4 + T cell Isolation Kit (Stem Cell). The cells (2.5 × 10 5 ) were cultured for 120 h under Th cell-differentiating conditions [ 23 ]. Briefly, for in-vitro differentiation of CD4 + T cells into Th1 cells, isolated cells were cultured with anti-mouse CD3 (5 μg/ml), anti-mouse CD28 (5 μg/ml), IL-12 (20 ng/ml), IL-2 (20 ng/ml), and anti-IL-4 (10 μg/ml).…”