Human plasma as a dermal scaffold for the generation of a completely autologous bioengineered skin Histological and immunohistochemical examination
10.3969/j.issn.1673-8225.2009.47.001
- VernacularTitle:人血浆作为真皮支架对自体皮肤增殖的影响:组织学和免疫组织化学试验
- Author:
Negri STEFANO
;
Federici GIADA
;
Farinato SARA
;
Fila CHIARA
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2009;13(47):9211-9216
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: Tissue engineering is a multidisciplinary area of research which aims to regenerate damaged tissues and organs in the human body, starting from the assumption that almost all animal tissues in the human body can be cultivated in a laboratory. The general principle is to isolate stem cells from a patient who requires a transplant and then the cells are cultured to grow and differentiate on a suitable support to produce the replacement tissue. On the one hand, it is necessary to find a suitable support (matrix or scaffold) on which cells can adhere and form stratified structures. On the other hand, the conditions allowing cells to proliferate and differentiate into the various types of tissues must be understood and reproduced. At present, the materials used as support in the treatment of ulcers and bums are manifold, such as collagen, hyaluronic acid, fibrin and PLGA. OBJECTIVE: To discuss the effectiveness of a new kind of biomaterial, autologous plasma, as scaffold for the growth, expansion and differentiation of autologous fibroblasts and keratinocytes.DESIGN: We have devised a method for dermal regeneration using autologous fibroblasts immersed in a matrix of human plasma so as to eliminate any problem linked to the safety of the sample and rejection from a patient. On this new dermis, keratinocytes have then been seeded.TIME AND SETTING: This work was performed at Cell Factory of C. Poma Hospital in Mantova in 2008. PARTICIPANT/MATERIALS: The human keratinocytes and fibroblasts used for this research were taken from a skin fragment after mastectomy in a 58-year-old patient. The experiment was approved by the independent ethics committee of C. Poma Hospital and the patient was requested to give informed consent.INTERVENTIONS/METHODS: Our studies were focussed on the isolation of human basal keratinocytes and fibroblasts from autologous biopsied skin samples and on their proliferation in culture flasks. Autologous plasma was subsequently used as scaffold for the formation of skin grafts whose morphofunctional and immunohistochemical characteristics are similar to those of normal skin.MAIN OUTCOME MEASURES: In order to evaluate the results, the samples of plasma were fixed in formalin, embedded in paraffin, stained with haematoxilyn-eosin and viewed through a microscope to count the number of cells in 40-fold 5 fields. Furthermore, all specimens underwent immunohistochemical examination for vimentin, keratin AE1 and AE3, collagen IV, P63 and Mib 1. RESULTS: The skin grafts obtained in the present study consist of few layers of normally-shaped keratinocytes on a plasma matrix with fibroblasts embedded inside. The formation of basement membrane shows that autologous plasma is a good scaffold for the growth, differentiation and proliferation of keratinocytes and fibroblasts and it also demonstrates that dermal and epidermal cells can be interlocked to reconstruct artificial grafts identical to normal skin.CONCLUSION: Plasma has given a remarkable performance as scaffold for the proliferation and differentiation of keratinocytes and fibroblasts. In particular, we have observed that fibroblast-enriched plasma can replace dermis temporarily, besides being a robust scaffold for the growth of keratinocytes. In addition, plasma is relatively cheap and easy to prepare. The use of autologous keratinocytes and fibroblasts, as well as that of autologous plasma, is essential to avoid any type of rejection. This kind of therapy is ideal for patients with chronic defects requiring continuous grafts.
