The Skin Reservoir Model: A Tool for Evaluating Microdialysis Sampling of Large Biomarkers from Human Skin.

Microdialysis is a well-established technique for sampling of small molecules from the human skin, but larger molecules are more difficult to recover. Consequently, sampling feasibility must be evaluated before microdialysis is used in vivo. This report presents a tool for estimating the recovery of large biomarkers from human skin by microdialysis, using previously frozen human skin specimens as reservoirs for biomarker reference solutions. Recovery of the following 17 biomarkers was assessed: CCL27/CTACK, CXCL1/GROα, CXCL7/NAP-2, CXCL10/IP-10, EGF, GM-CSF, IFN-γ, IL-1α, IL-6, IL-8, IL-17, IL-22, IL-23, MIF, TNF-α, TSLP and VEGF. The relative skin recoveries of 13/17 biomarkers were successfully determined in the range 4.0-18.4%. Sampling in the skin reservoir model was not associated with probe leakage, as fluid recovery was stable, at between 80% and 110%. Furthermore, the skin reservoir model enabled studies and optimization of different parameters known to affect biomarker recovery, including flow rate and perfusate composition.

GSK2646264, a spleen tyrosine kinase inhibitor, attenuates the release of histamine in ex vivo human skin.

BACKGROUND AND PURPOSE: Chronic spontaneous urticaria presents as a heterogeneous syndrome characterised by wheals, angioedema, or both for greater than 6 weeks. Spleen tyrosine kinase mediates allergen-induced mast cell degranulation via the IgE signalling pathway, a central component of wheal formation and inflammation. In this study, we investigated the effects of perfused or topically administered GSK2646264 on IgE-mediated histamine release from mast cells in an ex vivo human skin model. EXPERIMENTAL APPROACH: Using a novel SkiP device, ex vivo human skin from mastectomy surgeries was challenged with anti-IgE, complement 5a (C5a), and buffer to induce histamine release from skin mast cells. Histamine was collected via microdialysis fibres and measured fluorometrically. GSK2646264 was delivered via perfusion either using microdialysis fibres or topically in a cream. Drug concentrations in the skin were measured by LC-MS, and a pharmacokinetic/ pharmacodynamic (PK/PD) relationship developed. KEY RESULTS: Perfused GSK2646264 significantly inhibited anti-IgE (but not C5a)-induced histamine release in a concentration-dependent manner. The 0.5, 1, and 3% cream delivered GSK2646264 to the dermis above the IC90 and dose-dependently attenuated anti-IgE-induced histamine release. CONCLUSIONS AND IMPLICATIONS: GSK2646264 administered topically or direct to the dermis blocked histamine release from in situ skin mast cells. A PK/PD relationship curve suggests that dermal concentrations above 6.8 µM should lead to approximately 90% inhibition of histamine release from skin mast cells following activation of the Fc fragment of IgE receptor 1a, implicating a potential use for the compound in skin mast cell diseases such as urticaria.

Is a positive intracutaneous test induced by penicillin mediated by histamine? A cutaneous microdialysis study in penicillin-allergic patients.

BACKGROUND: Diagnostic workup of penicillin allergy comprises skin testing with penicillins, and patients are deemed allergic if skin test is positive. However, the literature suggests that skin test-positive patients may be challenge-negative, indicating that the skin test may be falsely positive. OBJECTIVE: To investigate real-time histamine release from a positive intracutaneous test induced by penicillin in patients with positive and negative challenges to penicillin. METHODS: Skin microdialysis was performed in 21 penicillin-allergic patients with positive skin test, 13 non-allergic volunteers serving as negative controls, and 7 grass pollen-allergic patients serving as positive controls. Histamine was measured by microdialysis after skin test with penicillin/grass/NaCl. Penicillin challenge was subsequently performed in 12 of the patients. RESULTS: Only 10/21 patients (47.6%) were skin test positive at microdialysis. During microdialysis 13 single intracutaneous tests were positive and histamine was detected in 4/13 occurring in four challenge positive patients. Thirteen/21 patients (61.9%) were deemed allergic to penicillin; eight had positive skin test. Two patients with positive skin test were challenge negative. In grass pollen allergic patients, 7/7 had a positive intracutaneous test to grass and all released histamine in the wheals. All 13 negative controls had negative intracutaneous test to penicillin and no histamine release. CONCLUSION: Histamine was only detected in the minority of positive intracutaneous tests with penicillin in penicillin-allergic patients. Other mediators may be involved.

Measuring histamine and cytokine release from basophils and mast cells.

Basophils and mast cells are known for their capability to release both preformed and newly synthesized inflammatory mediators. In this chapter we describe how to stimulate and detect histamine released from basophils in whole blood, purified basophils, in vitro cultured mast cells, and in situ skin mast cells. We also give an example of an activation protocol for basophil and mast cell cytokine release and discuss approaches for cytokine detection.

A positive serum basophil histamine release assay is a marker for ciclosporin-responsiveness in patients with chronic spontaneous urticaria.

The electronic records of 398 patients with chronic spontaneous urticaria (CSU) who had had a serum basophil histamine release assay (BHRA) performed as a marker of functional autoantibodies were audited. The BHRA was positive in 105 patients (26.4%). Fifty eight were treated with ciclosporin because they were H1 anti-histamine unresponsive. CSU patients with a positive BHRA were more likely to respond clinically (P<0.001) and to have raised thyroid autoantibodies (P<0.02) than those with a negative BHRA. The BHRA offers a useful predictive biomarker for a good response of H1 antihistamine-unresponsive CSU patients to ciclosporin.