Versican is upregulated in circulating monocytes in patients with systemic sclerosis and amplifies a CCL2-mediated pathogenic loop

Abstract

IntroductionAltered phenotypes of circulating monocytes of patients with systemic sclerosis (SSc) have been reported, but the role of these alterations in the pathogenesis of SSc remains unclear. This study was undertaken to identify molecules that are preferentially expressed by SSc monocytes, and to investigate the roles of these molecules in the pathogenic process of SSc.MethodsWe analyzed circulating CD14+ monocytes isolated from 36 patients with SSc and 32 healthy control subjects. The monocytes' gene expression profiles were assessed by Oligo GEArray® (SABiosciences, Frederic, MA, USA) and semiquantitative or quantitative PCR; their protein expression was evaluated in culture supernatants of unstimulated monocytes by immunoblotting or ELISA, and by immunocytostaining. Monocyte chemoattractant activity of CCL2 was assessed in a TransWell® system (Corning Incorporated, Corning, NY, USA) in the presence or absence of chondroitin sulfate (CS).ResultsA step-wise approach to profiling gene expression identified that versican and CCL2 were upregulated in SSc monocytes. Subsequent analysis of proteins expressed in monocyte culture supernatants confirmed enhanced production of versican and CCL2 in SSc monocytes compared with control monocytes. CCL2 bound to CS chains of versican and colocalized with versican in the monocytes' Golgi apparatus. Finally, CCL2 had a greater ability to mediate monocyte migration when bound to CS chains, because this binding provided efficient formation of CCL2 gradients and protection from protease attack.ConclusionCirculating monocytes with elevated versican and CCL2 levels may contribute to the fibrotic process in a subset of SSc patients by amplifying a positive feedback loop consisting of versican, CCL2, and the influx of monocytes.

Versican and the control of inflammation

“…While accumulation of versican during the inflammatory response can originate in the stromal connective tissue, a number of past as well as more recent studies have identified versican as a gene which is upregulated in monocytes in a number of pro-inflammatory states, such as myocardial infarction (Toeda et al, 2005), coronary stenosis (Wingrove et al, 2008), autoimmunity (Masuda et al, 2013; Olsen et al, 2004; Shou et al, 2006), and in response to pro-inflammatory stimulants such as lipopolysaccharide (LPS) (Lang et al, 2002) and hypoxia (Asplund et al, 2011; Asplund et al, 2009). In addition, the versican gene is differentially expressed in M1 macrophages, as opposed to M2 macrophages, as they differentiate from monocytes (Asplund et al, 2011; Martinez et al, 2006).…”

“…Such changes raise the possibility that the myeloid cells themselves are creating their own microenvironment to support the myeloid phenotype as part of the inflammatory response. For example, versican expression is elevated in CD14 positive monocytes taken from patients with systemic sclerosis (Masuda et al, 2013) and this elevated expression is accompanied by elevated expression of CCL2 (Monocyte Chemoattractant Protein 1). Furthermore, versican protects CCL2 from degradation and this interaction promotes monocyte migration (Masuda et al, 2013).…”

“…For example, versican expression is elevated in CD14 positive monocytes taken from patients with systemic sclerosis (Masuda et al, 2013) and this elevated expression is accompanied by elevated expression of CCL2 (Monocyte Chemoattractant Protein 1). Furthermore, versican protects CCL2 from degradation and this interaction promotes monocyte migration (Masuda et al, 2013). Such results confirm earlier studies that show CCL2 binds to versican and impacts inflammation in a model of neuronal inflammation hyperalgesia (Bogen et al, 2009).…”

Versican Deficiency Significantly Reduces Lung Inflammatory Response Induced by Polyinosine-Polycytidylic Acid Stimulation

“…Monocyte Chemotaxis Assay-As adapted from Masuda et al (77), the bottom wells of a 24-well transwell assay system (8-m pore size; Greiner Bio-One) were coated overnight with 200 l of PBS alone or containing various concentrations of CS (Sigma; CSA, C8529), purified bovine versican, or HA. In some experiments, the upper membrane surface of the transwell insert was also coated with versican.…”

Advanced in vitro systems for efficacy and toxicity testing in nanomedicine

“…In other products, 3D scaffolds fabricated from biodegradable materials with welldefined pores size can be used in studies focused on stem cells, tumour models and -important from a biodegradable perspective tissue engineering. In a more well-established example, Corning ® , Transwell ® (Corning, Tewksbury, MA, USA) permeable supports have become a standard method for culturing cells by permitting cells to uptake and secrete molecules on both its basal and apical surfaces (105,106). Insphero GravityPLUS ™ (Insphero, Schlieren, Switzerland) can be used for scalable microspheroid assembly applicable in toxicology and oncology applications (107,108).…”