Abstract
Several in vitro models that mimic different aspects of local skin inflammation exist. The use of ex vivo human skin organ culture (HSOC) has been reported previously. However, comprehensive evaluation of the cytokine secretory capacity of the system and its kinetics has not been performed. Objective: the aim of the current study was to investigate the levels and secretion pattern of key cytokine from human skin tissue upon lipopolysaccharide (LPS) stimulation. HSOC maintained in an air–liquid interface was used. Epidermal and tissue viability was monitored by MTT and Lactate Dehydrogenase (LDH) activity assay, respectively. Cytokine levels were examined by ELISA and multiplex array. HSOCs were treated without or with three different LPS subtypes and the impact on IL-6 and IL-8 secretion was evaluated. The compounds enhanced the secreted levels of both cytokines. However, differences were observed in their efficacy and potency. Next, a kinetic multiplex analysis was performed on LPS-stimulated explants taken from three different donors to evaluate the cytokine secretion pattern during 0–72 h post-induction. The results revealed that the pro-inflammatory cytokines IL-6, IL-8, TNFα and IL-1β were up-regulated by LPS stimuli. IL-10, an anti-inflammatory cytokine, was also induced by LPS, but exhibited a different secretion pattern, peak time and maximal stimulation values. IL-1α and IL-15 showed donor-specific changes. Lastly, dexamethasone attenuated cytokine secretion in five independent repetitions, supporting the ability of the system to be used for drug screening. The collective results demonstrate that several cytokines can be used as valid inflammatory markers, regardless of changes in the secretion levels due to donor’s specific alterations.
Highlights
The skin is the largest organ of the human body, primarily acting to maintain homeostasis and protect the body from the deleterious action of the environment
Though inflammation is a common feature of several skin diseases including psoriasis, atopic dermatitis, seborrheic dermatitis and contact dermatitis [3], the characteristics of the cellular immune response and composition of cytokine profile vary among them [4]
The secretion profile and kinetics depend on both cell-specific expressions of inflammatory mediators and profile and kinetics depend on both cell-specific expressions of inflammatory mediators and the original signaling that caused their secretion
Summary
The skin is the largest organ of the human body, primarily acting to maintain homeostasis and protect the body from the deleterious action of the environment. Upon external or internal signals, skin-resident cells, such as Langerhans cells, keratinocytes, melanocytes, mast cells and macrophages, secrete small, hormone-like signal peptides called cytokines that act as local immune modulators or recruit additional immune cells [2]. Their action depends on the presence of specific membrane receptors found on the majority of cells. Three main in vitro platforms are available: cell cultures (e.g., keratinocyte, Langerhans cells, co-cultures, etc.), 3D reconstructed skin equivalent and human/porcine organ culture (ex vivo models). The objective of the current study was to monitor the secreted profile of 15 key cytokines in response to LPS and to assess the benefits and limitations of this experimental system
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