Lucie Germain

INFLUENCE OF INITIAL COLLAGEN AND CELLULAR CONCENTRATIONS ON THE FINAL SURFACE AREA OF DERMAL AND SKIN EQUIVALENTS: A BOX-BEHNKEN ANALYSIS

SummaryOur laboratory has been involved in finding optimal conditions for producing dermal and skin equivalents. As an original approach, a Box-Behnken experimental design was used to study the effects of the initial collagen and fibroblast concentrations and the initial gel thickness on the contraction of dermal and skin equivalents. The final surface area of dermal equivalent varied significantly with the initial concentration of collagen and fibroblast, whereas the initial thickness of gel had no appreciable effect on the contraction of the dermal equivalent. When keratinocytes were grown on these dermal equivalents they produced a very severe contraction, to an extent that all skin equivalents had a similar final surface area. This severe contraction was independent of collagen and fibroblast concentrations. Models for the prediction of the final percentage contraction of dermal and skin equivalents as a function of the initial concentration of collagen, the logarithm of fibroblast concentration, and the initial gel thickness were obtained and analyzed. Keratinocytes grown at the lowest seeding density did not contract the equivalents. However, histologic analysis has shown an incomplete coverage by these cells of the equivalents. The extensive contraction of the skin equivalent presenting adequate morphology is a major drawback toward its clinical utilization for burn wound coverage.

What's new in human wound-healing myofibroblasts?

During wound healing and fibrocontractive diseases, clinical and experimental investigations have shown that fibroblastic cells acquire some morphological and biochemical features similar to those of smooth muscle cells [33]. These modified fibroblasts, called myofibroblasts, express de novo α-SM actin temporarily during wound healing and permanently in fibrotic situations, such as hypertrophic scars or fibromatosis. Myofibroblasts are thought to be involved in contraction and have been observed in practically all fibrotic conditions involving retraction and reorganization of connective tissues [32].

CHAPTER 50 – TENDONS AND LIGAMENTS

response is usually divided into three phases, more for convenience as the phases merge imperceptibly into eachAlthough tendons and ligaments generally respond similarly to other and represent a con-tinuum rather than discreteinjury, there are anatomic and regional differences in the speed stages. The firstphase, occurring in the firstweek after disruption of the tendon fibers,is characterized by inflammation.There is edema and infiltrationof a variety of cell types attracted to the region by inflammatorymediators. Platelets and mast cells release histamine, a potent agent promoting vasodilation and increasing blood vessel permeability. Serotonin, bradykinin, leukotri-enes, and prostaglandins act together to recruit polymorphonuclear leukocytes and lymphocytes from the circulation. Growth factors released by platelets include platelet-derived growth factor (PDGF), transforming growth factor-β (TGF-β), and epidermal growth factor (EGF). Macrophages are present withinue06a 24 hours, phagocytosing tissue debris andreleasing numerous inflammatorymediators growthfactors, including basic fibroblastgrowth factor (bFGF), transforming growth factor-α (TGF-α), TGF-β, and PDGF. These growth factors are chemotactic for fibroblastsand other cells and generally act to stimu-late matrix synthesis. Angiogenic factors such as bFGF and vascular endothelial growth factor (VEGF) stimulate capillary ingrowth into the fibrousclot. Toward the end of the inflammatoryphase, which may last several days, fibroblastsbecome the predominant cell type. The second phase, generally lasting up to 6 weeks after injury, is character-ized by cell proliferation and new matrix synthesis by fibroblasts.This new matrix is different in both quality and quantity from the normal matrix. Described as granulation tissue, it consists of a disorganized matrix with an elevated cell density that fillsthe tissue defect. The third phase, occurring from 3 to 6 weeks after injury and lasting for at least a year, is a prolonged period of remodeling and maturation in which the matrix components and tissue cellularity revert gradually toward normal. During this stage many cells in the scar are contractile myofibroblasts,which are specialized cells important for the organiza-tion of the wound tissue.

What is new in mechanical properties of tissue-engineered organs.

Tissue engineering is a promising new field based on expertise in cell biology, medicine and mechanical engineering. It raises exciting hopes of producing autologous tissue substitutes to replace altered organs. This challenge involves highly specialized technology in order to provide the proper shape to the tissue and promote the maintenance of its native physiological properties. Primary cell populations may lose some of their functional and morphological properties in vitro in the absence of a proper environment. In order to maintain cell integrity, a three-dimensional matrix that mimics the in vivo environment as closely as possible was developed, according to the type of tissue produced [1, 5, 18, 26, 27, 29, 34, 35].