Multiplex cytokine analysis of dermal interstitial blister fluid defines local disease mechanisms in systemic sclerosis

Abstract

IntroductionClinical diversity in systemic sclerosis (SSc) reflects multifaceted pathogenesis and the effect of key growth factors or cytokines operating within a disease-specific microenvironment. Dermal interstitial fluid sampling offers the potential to examine local mechanisms and identify proteins expressed within lesional tissue. We used multiplex cytokine analysis to profile the inflammatory and immune activity in the lesions of SSc patients.MethodsDermal interstitial fluid sample from the involved forearm skin, and synchronous plasma samples were collected from SSc patients (n = 26, diffuse cutaneous SSc (DcSSc) n = 20, limited cutaneous SSc (LcSSc) n = 6), and healthy controls (HC) (n = 10) and profiled by Luminex® array for inflammatory cytokines, chemokines, and growth factors.ResultsLuminex® profiling of the dermal blister fluid showed increased inflammatory cytokines (median interleukin ( IL)-6 in SSc 39.78 pg/ml, HC 5.51 pg/ml, p = 0.01, median IL-15 in SSc 6.27 pg/ml, HC 4.38 pg/ml, p = 0.03), chemokines (monocyte chemotactic protein (MCP)-3 9.81 pg/ml in SSc, 7.18 pg/ml HC, p = 0.04), and profibrotic growth factors (platelet derived growth factor (PDGF)-AA 10.38 pg/ml versus 6.94 pg/ml in HC, p = 0.03). In general dermal fluid and plasma cytokine levels did not correlate, consistent with predominantly local production of these factors within the dermal lesions, rather than leakage from the serum. In hierarchical clustering and network analysis IL-6 emerged as a key central mediator.ConclusionsOur data confirm that an immuno-inflammatory environment and aberrant vascular repair are intimately linked to fibroblast activation in lesional skin in SSc. This non-invasive method could be used to profile disease activity in the clinic, and identifies key inflammatory or pro-fibrotic proteins that might be targeted therapeutically. Distinct subgroups of SSc may be defined that show innate or adaptive immune cytokine signatures.

Blockade of PDGF Receptors by Crenolanib Has Therapeutic Effect in Patient Fibroblasts and in Preclinical Models of Systemic Sclerosis

“…PDGF-A is overexpressed in SSc lesions (Yamakage et al, 1992) and increased expression of PDGFRα and PDGFRβ on spindle cells correlates with collagen deposition in SSc biopsies (Daoussis et al, 2012). Increased PDGF-A has also been reported in the dermal interstitial blister fluid of SSc patients (Clark et al, 2015). In addition, agonistic anti-PDGFRα autoantibodies have been detected in the serum of SSc patients (Moroncini et al, 2015).…”

Minimally-invasive Sampling of Interleukin-1α and Interleukin-1 Receptor Antagonist from the Skin: A Systematic Review of In vivo Studies in Humans

“…In the articles included in this review, the negative pressure was between 200 and 400 mmHg, the suction cups ranged in size from 5 to 15 mm and the time needed for the formation of the blisters was mostly 2-3 h. Warming was sometimes added to aid blister formation. For the quantification of the protein levels of IL-1α and/or IL-1RA, the majority used EIA/ ELISA (n = 10) (43-52) and the remaining used multiplex arrays (53)(54)(55)(56)(57). All studies but 2 analysed additional cytokines and chemokines, albeit many reported that not all could be detected (43-45, 51, 52, 54, 55).…”

Prospects for Using Tocilizumab in Systemic Sclerosis

“…Возникло предположение об ассоциации этих субтипов с ответом на иммуносупрессивную терапию [49]. Недавнее изучение интерстициальной жидкости из пораженной кожи боль-ных ССД с помощью мультиплексного анализа показало существенные различия в локальной концентрации воспа-лительных и профиброзных белков [50]. Это исследование подтвердило существование субпопуляций больных с раз-ным профилем доминирующих цитокинов, а также выяви-ло подгруппу больных с высокими уровнями ИЛ6 в коже (без повышения в циркуляции), что, по мнению авторов, может предполагать перспективность применения таргет-ной терапии против ИЛ6 в этой подгруппе больных.…”

“…Средний возраст больных составил 58 (46)(47)(48)(49)(50)(51)(52)(53)(54)(55)(56)(57)(58)(59)(60)(61) лет. Исходно 73% больных получали низкие дозы ГК (≤10 мг/сут в пересчете на преднизолон) и 57% -метотре-ксат.…”