IL-1beta-induced Langerhans' cell migration and TNF-alpha production in human skin: regulation by lactoferrin

Abstract

In mice, the roles of cytokines in the initiation of epidermal Langerhans' cell (LC) migration are well documented; however, the mechanism of this response in humans is less well defined. The purpose of the present investigation was to examine the contribution of interleukin (IL)-1beta to human epidermal LC migration and to define further the mechanisms of this response. We demonstrate here that homologous recombinant IL-1beta administered intradermally to healthy human volunteers provides a stimulus for LC migration, with significant (P < 0.01) reductions in LC densities being observed at both 2 h and 4 h following treatment. At the later time-point of 4 h, injection of IL-1beta was also accompanied by activation of those LC remaining in the epidermis. Analysis of fluid aspirated from suction blisters formed at injection sites revealed significant (P < 0.01) tumour necrosis factor (TNF)-alpha production (2.99 +/- 1.18 pg TNF-alpha/mg protein; mean +/- s.d. of n = 10) in response to IL-1beta treatment compared with saline control injections (0.90 +/- 1.05 pg TNF-alpha/mg protein). Prior topical application of human recombinant lactoferrin (LF), an iron-binding protein found in exocrine secretions and skin, inhibited IL-1beta-mediated LC migration and also compromised the production of TNF-alpha protein as measured in suction blister fluids derived from each of the treatment sites. Taken together, these data demonstrate that IL-1beta is associated with both the stimulation of human epidermal LC migration and local TNF-alpha production. Topical treatment with LF compromises both these responses. These data suggest that topical LF may potentially represent a novel therapeutic in the treatment of skin inflammation where TNF-alpha is an important mediator.

Fig. 1

Modulation of human epidermal CD1a⁺ LC frequency by homologous recombinant IL-1β. Each volunteer received intradermal injections of IL-1β (10, 50 and 100 U) and a single control injection of normal saline alone (0 U). Biopsies for each volunteer were taken 2 h (a–d) or 4 h (e–g) later and epidermal sheets prepared. The frequency of CD1a⁺ LC was assessed following indirect immunofluorescence staining of epidermal sheets. Results are expressed as the mean ± s.d. number of cells/mm² derived from examination of 50 fields/sample. The percentage reduction in frequency of CD1a⁺ LC induced by 10, 50 and 100 U IL-1β, respectively, for each volunteer was: (a) 12·0, 25·0 and 27·9%; (b) 2·0, 15·5 and 14·1%; (c) 0·0, 10·1 and 13·0%; (d) 17·7, 23·4 and 24·7%; (e) 10·0, 26·9 and 19·5%; (f) 0·0, 14·6 and 15·4%; (g) 21·5, 26·2 and 38·5%.

Fig. 2

Morphological changes induced in human epidermal CD1a⁺ LC by IL-1β. Epidermal sheets were prepared from volunteer g (Fig. 1) 4 h following intradermal administration of (a) saline, or (b) 50 U IL-1β and stained by indirect immunofluorescence for CD1a expression. Scale bar: 150 µm.

Fig. 3

Inhibition by LF of IL-1β-induced changes in epidermal CD1a⁺ LC frequency. Volunteers (h–l) were exposed topically at two sites to LF (50 µl) and at a further two sites to an equivalent volume of aqueous cream alone. Two hours later, IL-1β was injected intradermally into paired sites (one pretreated with LF and one with cream) and biopsies taken 2 h later. CD1a⁺ LC densities were assessed following indirect immunofluorescence staining of epidermal sheets. Results are expressed as the mean ± s.d. number of cells/mm² derived from examination of 50 fields/sample. The percentage reduction in frequency of CD1a⁺ LC induced by IL-1β (at sites pretreated with cream) was: (h) 19·0%; (i) 17·6%; (j) 23·9%; (k) 37·6%; (l) 28·2%. In all cases, prior treatment with LF inhibited the IL-1β-induced response.

Fig. 4

Inhibition by LF of IL-1β-induced epidermal TNF-α production. A further 10 volunteers were exposed topically to LF or cream followed by intradermal injections of IL-1β or saline, as described in the legend to Fig. 3. Each symbol represents the results from one of the 10 volunteers. Immediately following injections, suction blister cups were applied to each of the four treated sites. Suction blister fluids were collected and analysed by ELISA for TNF-α content and total protein expression by modified Lowry [22]. Results are expressed as pg TNF-α/mg protein.