Mechanism of mast cell adhesion to human tenocytes in vitro

Abstract

Mast cells and fibroblasts are two key players involved in many fibrotic and degenerative disorders. In the present study we examined the nature of binding interactions between human mast cells and tendon fibroblasts (tenocytes). In the mast cellfibroblast co-culture model, mast cells were shown to spontaneously bind to tenocytes, in a process that was partially mediated by a5b1 integrin receptors. The same receptors on mast cells significantly mediated binding of these cells to tissue culture plates in the presence of tenocyte-conditioned media; the tenocyte-derived fibronectin in the media was shown to also play a major role in these binding activities. Upon binding to tenocytes or tissue culture plates, mast cells acquired an elongated phenotype, which was dependent on a5b1 integrin and tenocyte fibronectin. Additionally, tenocyte-derived fibronectin significantly enhanced mRNA expression of the adhesion molecule, THY1, by mast cells. Our data suggests that a5b1 integrin mediates binding of mast cells to human tenocyte and to tenocyte-derived ECM proteins, in particular fibronectin. Decreased collagen production and increased MMP activities have been detected in tendon biopsies from patients with tendinopathy.3 To date, only a handful of studies have investigated the possible contribution of mast cells in tendinopathy. Mast cells have previously been reported to directly communicate with fibroblasts via gap junction intercellular communications, 4,5 and the SCF/c-kit receptor interaction has been shown to mediate adhesion of mouse mast cells to 3T3 fibroblasts. 6 It is very likely that due to heterogeneity in fibroblast and mast cell populations, different receptors are involved in the process of adhesion, and understanding the exact nature of these specific cell-cell interactions could help delineate these connections.In this study, we utilized an in vitro cell culture system using primary human tendon fibroblasts (tenocytes) and an established human mast cell line (HMC-1) 7 to identify adhesion receptors that mediate binding of human mast cells to tenocytes and their ECM proteins. Mast cells avidly bound to tenocytes upon co-culture, and the adhesion of mast cells to tissue culture plates in response to culture with tenocytederived conditioned media was primarily mediated by a5b1 integrin. Upon binding, mast cells acquired an elongated morphology and enhanced their expression of the adhesion receptor, THY1 (cluster of differentiation 90/CD90), events that were significantly dependent on a5b1 and fibronectin mediated interactions. Our data provides a mechanism by which human mast cells adhere to human tendon fibroblasts and its ECM components. MATERIAL AND METHODS Cell CultureTendon fibroblasts (tenocytes) were isolated from human hamstring tendon. Ethics approval was obtained from the review board at the University of British Columbia.3 Samples of healthy hamstring tendons from living male and female patients undergoing anterior cruciate ligament reconstruction were trimmed to remove fat and muscl...

Tendon Immune Regeneration: Insights on the Synergetic Role of Stem and Immune Cells during Tendon Regeneration

“…Firstly, tenocytes exposed to mast cell line conditioned media presented a higher viability rate compared to the control sample during the in vitro studies [116]. Additionally, the obtained results showed that the contact with mast cells conditioned media stimulated human tenocytes to increase the levels of COX-2, which resulted in an increase in the PGE 2 levels [117]. This upregulation of PGE 2 resulted in a decrease of the COL1A1 mRNA levels, thus affecting the biomechanical properties of the tendon [116,118].…”

MRGPRX2-mediated mast cell activation by substance P from overloaded human tenocytes induces inflammatory and degenerative responses in tendons

“…We next examined the gene expression of factors in stretched human tenocytes that could activate MCs. Previous data indicate that human tenocytes are capable of producing fibronectin and substance proteins 14 , 19 , which are factors known to activate mast cells 20 , 21 . The qPCR analysis revealed elevated expression of SP and fibronectin in stretched tenocytes at the mRNA level (Fig.…”

“…Tenocytes are capable of producing a number of mast cell chemoattractants such as prostaglandins, chemokines, TGF-β isoforms and stem cell factor (SCF). We previously showed that cyclic stretching of human tenocytes 19 , 38 , 39 increases the expression of Cox- 2 and the activity of TGF-β 12 , 40 . In addition to autocrine effects of the factors on tenocytes, they can stimulate mast cell migration and degranulation such as SP 21 .…”

“…Fibronectin plays a crucial role in regulating mast cell adhesion to tenocytes through integrin receptors, ultimately leading to mast cell degranulation and increased cytokine release 19 . We conducted an adhesion assay to compare MCs' adhesion to fibronectin and collagen.…”