Simple wound exudate collection method identifies bioactive cytokines and chemokines in (arterio) venous ulcers

Abstract

A major challenge for clinicians treating (arterio) venous leg ulcers is to decide between standard therapy and advanced interventions. Here, we developed a simple method to collect human material representative of the ulcer wound bed, which can be used to identify biomarkers for prognostic test development. Superficial surgical debridement was performed using a small vidal curette during the weekly visit to the outpatient clinic. Moist, easily removable debridement material essentially blood free (including necrotic and nonviable slough) was collected from the surface of the ulcer. The amount ranged from 5.5 mg to 78 mg material per ulcer. Seventeen cytokines, chemokines, and growth factors were extracted and analyzed by enzyme-linked immunosorbent assay (concentration range: 0.0005-78 ng/mg total protein). Notably, CXCL8 was by far the most abundant protein present. Inflammatory mediators were more abundant than anti-inflammatory mediators (e.g., interleukin (IL)-10 and transforming growth factor-β1). Bioactivity assays showed chronic wound extracts to be capable of stimulating fibroblast migration in a chemokine-dependent manner and also capable of stimulating healthy cells within skin substitutes to secrete wound healing mediators (CCL2, CXCL1, CXCL8, IL-6) in an IL-1α dependent manner. Collection of debridement tissue enables investigation of the ulcer environment in an easy noninvasive manner that may be suitable for prognostic test development.

Differential Response of Human Adipose Tissue-Derived Mesenchymal Stem Cells, Dermal Fibroblasts, and Keratinocytes to Burn Wound Exudates: Potential Role of Skin-Specific Chemokine CCL27

“…21 This receptor is expressed in many cells, including keratinocytes, dermal fibroblasts, and ASC. 3,24 Since we previously detected this chemokine also in chronic wound exudates, 10 it is most probable that it is related to skin trauma in general. However, the function of this skin-specific chemokine during wound healing is still unknown.…”

“…3 In addition, we have shown that both ASC and dermal fibroblasts migrate predominantly toward chemokine CCL5, 3 which is present in the wound fluid of chronic cutaneous wounds. 10 However, it is still unclear how the migrated ASC and dermal fibroblasts respond to wound-healing mediators that are present in the wound bed. Generally, skin substitutes (SS) contain dermal fibroblasts and keratinocytes.…”

Comparison of advanced therapy medicinal product gingiva and skin substitutes and their in vitro wound healing potentials

“…Furthermore, elevated levels of CXCL8 have been described to contribute to delayed wound healing, as CXCL8 was increased in non‐healing human thermal wounds compared with healing wounds. High amounts of CXCL8 and IL‐6 were detected in wound extracts of non‐healing ulcers (Kroeze et al, 2012b). However, it has also been described that tumour necrosis factor‐α stimulates oral keratinocytes to produce more CXCL8 and IL‐6 than skin keratinocytes (Li, Farthing, Ireland, & Thornhill, 1996a; Li, Ireland, Farthing, & Thornhill, 1996b).…”

The Effect of PRF on Serum Starved Human Dermal Fibroblast

“…Chronic ulcers are difficult to heal spontaneously due to their characteristics of low cellular proliferation rates, 1-3 low cellular migration capacities, [3][4][5] followed by cellular senescence of fibroblasts on their wound bed and edges. 3,6 All of these may be caused by lack of growth factor supplies 7,8 as they are indicated by the decrease of GF-gene expressions among diabetic patients who tend to have chronic ulcers, 9 the entrapment of GFs by fibrin that spill out when capillaries leak due to high venous pressure in patients with venous leg ulcers, 7,10,11 and GFs degradation by protease contained in bio-film on the surface of chronic ulcer.…”