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Notes: Purified antibodies against type IV collagen and laminin were used to localize basement membranes by indirect immunofluorescence in various anatomical regions of normal and diseased human skin. The two proteins showed extensive codistribution. A continuous linear staining was found along the epidermal-dermal junction and around hair follicles, sebaceous gland acini and small capillaries. The same proteins also surrounded individual cells such as those found in vessels, hair erector muscles and subcutaneous tissue.Blister formation in bullous pemphigoid left type IV collagen and laminin on the floor of the blister, while the bullous pemphigoid antigen as detected by human autoantibodies was found on both sides of the blister. In solid basal cell carcinoma a strong staining was found around all tumour islands as well as focally within the cell clusters. This suggests that the tumour cells produce these basement membrane proteins but have lost, at least in part, control of polar deposition.

Notes: LyphoDermTM (XCELLentis, Belgium) is an end-sterilized, freeze-dried lysate from cultured allogeneic epidermal keratinocytes, formulated into a hydrophilic gel. Its efficacy and safety were evaluated, in combination with standard care (hydrocolloid dressing and compression therapy), in 194 patients suffering from hard-to-heal (lasting more than 6 weeks and not responding to conventional therapy) venous leg ulcers. Two control groups received standard care, with or without vehicle, respectively. Patients had a median age of 67.5 years and the majority were females (61%). The median duration of the ulcer was 43 weeks and in 39% of the subjects it had been present for more than 1 year. Thirty-eight percent of the patients in the standard care + LyphoDermTM group had complete ulcer healing within 24 weeks (primary end point) compared to 27% of patients in the standard care + vehicle pooled groups (P = 0.114) in the “as treated” intent-to-treat cohort (37% vs. 27% in the “as randomized intent-to-treat cohort; p = 0.137). In the subgroup of patients with enlarging ulcers, the difference between the two groups was significant (30% vs. 11%; p = 0.024 in the “as treated” intent-to-treat cohort and 31% vs. 9%; p = 0.005 in the “as randomized” intent-to-treat cohort). LyphoDermTM was well tolerated and safe, and no differences in the frequency of adverse events were noted between the treatment groups. Although the primary objective of the study was not achieved, the exploratory analysis carried out in patients with enlarging ulcers suggests that LyphoDermTM could offer a new prospect for the treatment of patients with venous ulcers that may prove to be a significant adjunct to the overall provision of care.

Notes: Gene therapy is becoming a reality, and it is a particularly attractive approach for wound healing, because the wound site is often exposed, the treatment and condition should be transient, and gene products such as growth factors and cytokines suffer from problems with bioavailability and stability. Among the techniques for gene delivery to the wound site, particle-mediated bombardment with a device called the gene gun has become an important developmental tool. This instrument has been used in numerous examples of wound gene therapy with growth factors or their receptors in the last decade. Among the advantages of particle-mediated bombardment are ease and speed of preparation of the delivery vehicle, the stability of the DNA preparation, the absence of (viral) antigens, the ability to target the projectiles to different tissue depths and areas, and the rapid shedding of both particles and DNA if they are targeted to the epidermis. Clinical application of the technology remains limited by the relatively low efficiency of the method, the potential tissue damage created by impact of the particles, and the coverage area. The gene gun can also be used to facilitate the discovery and validation of gene products as wound healing agents.

Notes: Various culture procedures, suitable to maintain the differentiated phenotype of various types of cells in vitro, have been devised during the past decade. These culture systems use macromolecular components extracted from extracellular matrixes or synthetic polymers which provide cells with a three-dimensional, spatially structured support. Substantial information has come from the use of collagen lattices. Many types of cells, of mesenchymal or other origin, are able to organize collagen fibrils in these models and to form a connective tissue—like structure. Prerequisites for this process are active function of the cytoskeleton and the expression of α2β1 integrin collagen receptors. Interaction of cells with such a matrix has profound effects on morphologic status, proliferation, cellular metabolism, and state of differentiation. The in vitro procedures reviewed and described in this article offer the possibility to combine different types of cells to approximate the in vivo environment and to investigate such physiologic processes as cell-cell and cell-matrix interactions which are otherwise not easily accessible. In pathologic conditions of the skin, these models have proven to be useful tools in investigating diseases relating to impaired recognition of extracellular matrix structures or alterations of cytoskeletal assembly. In pharmacologic and toxicologic studies, activity of drugs and potentially useful therapeutic substances has been evaluated. The use of cells and matrix components derived from skin has lead to refined systems which could be adapted and extended to other organs in an attempt to better understand pathophysiologic mechanisms.

Notes: Cellular responses to platelet-derived growth factor, which affects all phases of the wound healing process, are dependent on the interaction of the growth factor with its cell surface receptors. Recently, we have shown that the platelet-derived growth factor-receptor was not expressed in uninjured human skin. In acute human wounds healing by secondary intention, both platelet-derived growth factor-receptor subunits were coordinately expressed, whereas no expression was found after reepithelialization at day 47. Even though impaired wound healing may be due to uncoordinated expression or the failure to express platelet-derived growth factor-receptor subunits, little is known regarding their expression in chronic ulcers. We studied the localization of platelet-derived growth factor-receptor expression in chronic venous leg ulcers of 15 patients with a median age of 73 years. Cryostat sections of biopsy specimens were immunostained with the use of antibodies against the α- and the β-platelet-derived growth factor subunits. RNA was extracted from biopsy specimens and subjected to Northern blot analysis with the use of oligolabeled complementary DNA for the platelet-derived growth factor-receptor. Platelet-derived growth factor-receptor α- and β-subunit expression was found in fibroblast-like cells within the wound bed and in cells beneath the epidermis of the wound edge. Platelet-derived growth factor-receptor β-subunit expression was detected in endothelial cells of the vessels, in the granulation tissue, and the wound edge, whereas platelet-derived growth factor-receptor α-subunit was not expressed in endothelial cells of the uninjured skin. This finding suggests that the platelet-derived growth factor α-subunit may be involved in vessel formation during tissue repair. Both platelet-derived growth factor-receptor subunits were expressed at the messenger RNA level indicating that the synthesis is at least partly regulated at a pretranslational level. As the cellular responsiveness to growth factors depends on their specific receptors, our finding that both platelet-derived growth factor-receptor subunits are expressed in chronic venous ulcers substantiates the concept of therapeutic trials with recombinant platelet-derived growth factor.