TGF-β1–induced phospholamban expression alters esophageal smooth muscle cell contraction in patients with eosinophilic esophagitis

Abstract

Background Eosinophilic esophagitis (EoE) is a chronic antigen mediated disease characterized by esophageal eosinophilia, remodeling, and fibrosis. TGFβ1 is a central regulator of EoE remodeling and increases esophageal smooth muscle cell contraction. Objective In this study, we aimed to understand the molecular mechanisms by which TGFβ1 could induce esophageal smooth muscle cell contraction. Methods We used primary human esophageal smooth muscle (ESM) cells and EoE myofibroblasts (EMF) to assess the mechanisms of TGFβ1-induced contraction. We analyzed the expression, phosphorylation, and function of phospholamban (PLN), a sarcoendoplasmic reticulum regulatory protein which was induced by TGFβ1. Expression of PLN, phospho-PLN, and its regulatory pathway was analyzed in the esophageal smooth muscle of EoE and control biopsies. Gene silencing in EoE EMFs was utilized to understand the role of PLN in contraction. Results TGFβ1 induced and phosphorylated PLN in primary human ESM and EoE EMFs. PLN and phospho-PLN were elevated in EoE as compared to control subject smooth muscle in vivo. PLN inhibition significantly diminished TGFβ1 induced EoE EMF contraction. PLN expression and ESM/EMF contraction depended on TGFβ receptor I signals. Conclusion We describe a previously unrecognized mechanism for esophageal smooth muscle cell contraction that depends on TGFβ1, its receptors, and PLN. Since PLN is elevated in EoE smooth muscle and PLN silencing diminishes contraction, we provide a novel potential mechanistic framework and therapeutic target for esophageal smooth muscle dysfunction in EoE.

GSDMB induces an asthma phenotype characterized by increased airway responsiveness and remodeling without lung inflammation

“…Our studies of epithelial cells transfected with GSDMB in vitro, as well as our studies of hGSDMB Zp3-Cre mice in vivo, identified that GSDMB induced high levels of expression of 5-LO and TGF-β1, mediators associated with AHR and smooth muscle remodeling. The 5-LO products can directly induce smooth muscle contraction with rapid onset kinetics, whereas TGF-β1 can directly increase smooth muscle contractility with slow onset kinetics (15). Previous studies have demonstrated that TGF-β1 and leukotrienes can produce synergistic effects to increase smooth muscle contraction, collagen synthesis, and epithelial cell proliferation (16).…”

Eosinophilic Esophagitis

“…Inhibiting PLN gene expression and signals through TGF-b receptor I both significantly decrease esophageal smooth muscle contraction in response to TGF-b1. 59 In the context of innate immunity, recent studies have shown increased messenger RNA for CXCL16, CD1d, and va23, consistent with increased numbers of invariant natural killer T (iNKT) cells. Studies in murine models show that iNKT are necessary for EoE.…”

“…15 Mast cell and eosinophils both make TGF-b1, which can directly alter the contraction of collagen gels impregnated with esophageal smooth muscle cells. 17,59 Recently, this has been demonstrated to rely on the expression of phospholamban (PLN), a sarcoplasmic reticulum protein that regulates calcium flux. 59 Although biopsies from control subjects do not express PLN, EoE esophageal smooth muscle expresses PLN.…”

“…17,59 Recently, this has been demonstrated to rely on the expression of phospholamban (PLN), a sarcoplasmic reticulum protein that regulates calcium flux. 59 Although biopsies from control subjects do not express PLN, EoE esophageal smooth muscle expresses PLN. Inhibiting PLN gene expression and signals through TGF-b receptor I both significantly decrease esophageal smooth muscle contraction in response to TGF-b1.…”

Eosinophilic Esophagitis

“…40 In vitro exposure of esophageal fibroblasts and smooth-muscle cells to transforming growth factor β (TGF-β) leads to smooth-muscle contraction and fibrosis. [41][42][43] …”