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. 2011 Oct 26;44(5):191-9.
doi: 10.1267/ahc.11014. Epub 2011 Aug 10.

Open Wound Healing In Vivo: Monitoring Binding and Presence of Adhesion/Growth-Regulatory Galectins in Rat Skin during the Course of Complete Re-Epithelialization

Peter Gál  1 , Tomáš Vasilenko, Martina Kostelníková, Ján Jakubco, Ivan Kovác, František Sabol, Sabine André, Herbert Kaltner, Hans-Joachim Gabius, Karel Smetana Jr
Affiliations

Open Wound Healing In Vivo: Monitoring Binding and Presence of Adhesion/Growth-Regulatory Galectins in Rat Skin during the Course of Complete Re-Epithelialization

Peter Gál et al. Acta Histochem Cytochem. .

Abstract

Galectins are a family of carbohydrate-binding proteins that modulate inflammation and immunity. This functional versatility prompted us to perform a histochemical study of their occurrence during wound healing using rat skin as an in vivo model. Wound healing is a dynamic process that exhibits three basic phases: inflammation, proliferation, and maturation. In this study antibodies against keratins-10 and -14, wide-spectrum cytokeratin, vimentin, and fibronectin, and non-cross-reactive antibodies to galectins-1, -2, and -3 were applied to frozen sections of skin specimens two days (inflammatory phase), seven days (proliferation phase), and twenty-one days (maturation phase) after wounding. The presence of binding sites for galectins-1, -2, -3, and -7 as a measure for assessing changes in reactivity was determined using labeled proteins as probes. Our study detected a series of alterations in galectin parameters during the different phases of wound healing. Presence of galectin-1, for example, increased during the early phase of healing, whereas galectin-3 rapidly decreased in newly formed granulation tissue. In addition, nuclear reactivity of epidermal cells for galectin-2 occurred seven days post-trauma. The dynamic regulation of galectins during re-epithelialization intimates a role of these proteins in skin wound healing, most notably for galectin-1 increasing during the early phases and galectin-3 then slightly increasing during later phases of healing. Such changes may identify a potential target for the development of novel drugs to aid in wound repair and patients' care.

Keywords: differentiation; lectin; migration; proliferation; repair.

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Figures

Fig. 1
Fig. 1
H+E: Hematoxylin and eosin staining of skin wounds at three different stages of the healing process, starting at day 2: (2d): presence of tissue necrosis (S), formation of the demarcation line beneath the scab consisting mainly of polymorphonuclear leucocytes (see insert); seven-days healing wound (7d): migration of epidermal (E) cells over the wound, forming of the granulation tissue (GT) rich on fibroblasts and high-caliber vessels (see insert); 21-days healing wound (21d): completed epidermis regeneration, well-formed granulation tissue with decreased number of vessels and fibroblast (see insert) establishing into the scar. K+Vim: wide-spectrum cytokeratin+vimentin double-staining immunohistochemistry of healing skin wounds at the same time points: 2d: migration of epidermal cells beneath the scab; 7d: formation of the granulation tissue rich on vimentin-positive cells; 21d: completed epidermis regeneration, well-formed granulation tissue with decreased number of vimentin-positive cells. Fibronectin: simple staining immunohistochemical localization in the course of healing of skin wounds: 2d: wounds with low-level expression of fibronectin near the wound edge; 7d: granulation tissue rich on fibronectin; 21d: low-level presence of fibronectin in the developing scar. For orientation solid/dotted/broken lines are given separating distinct regions referred by the following abbreviations: E, epidermis; D, dermis; GT, granulation tissue; S, scab; in detail, a dotted line sets epidermis apart from dermis and/or granulation tissue; the broken line distinguishes dermis from granulation tissue and the solid line scab/necrosis from vital tissue.
Fig. 2
Fig. 2
Illustration of immunohistochemical galectin detection and localization of accessible binding sites (BS) for labeled galectins in the epidermis and in the dermis/granulation tissue during healing. Comparison between control data (first vertical panel) and specimens at each studied time point during healing (2d: second vertical panel; 7d: third vertical panel; 21d: fourth vertical panel) is thus made possible for each marker along each horizontal panel. In detail, the following assignments of type of probe and time point are given. First panel (Gal-1): strong signal intensity for galectin-1 two days after injury in both epidermis and dermis near the wound edge, decreasing over time to minimal presence in the dermis at day 21; second panel (Gal-1-BS): low level of galectin-1 reactivity in uninjured skin and wounds two and 21 days post wounding, increased reactivity to Gal-1-BS in the granulation tissue; third panel (Gal-2): galectin-2 detection in the epidermis, absence in granulation tissue; fourth panel (Gal-2-BS): galectin-2 reactivity in uninjured skin and wounds localized to epidermis, low-level presence of binding sites in the dermis, insert—galectin-2 nuclear reactivity in the epidermis (dashed line marks the nuclei of keratinocytes); fifth panel (Gal-3): presence in the suprabasal epidermal layer and dermis, low-level signal intensity in the granulation tissue; sixth panel Gal-3-BS: present in the suprabasal epidermal layer and in the surrounding dermis, low abundance presence in the scar forming. For orientation solid/dotted/broken lines are given separating distinct regions referred by the following abbreviations: E, epidermis; D, dermis; GT, granulation tissue; S, scab; in detail, a dotted line sets epidermis apart from dermis and/or granulation tissue; the broken line distinguishes dermis from granulation tissue and the solid line scab/necrosis from vital tissue.
Fig. 3
Fig. 3
Computation of the staining data on semi-quantitative scale for the tested galectins and respective binding sites in epidermis and in the dermis/granulation tissue at the three given time points during the healing process (top row) with respect to wound closure. Arrow indicates the moment of wounding.
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