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Keratinocyte growth factor and interleukin beta 1, 6, 8, 10, 12 and tumor necrosis factor alpha in media culture of dermal fibroblasts of patients with burns

Grant number: 11/24092-6
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): March 01, 2012
Effective date (End): February 28, 2014
Field of knowledge:Health Sciences - Medicine
Principal Investigator:Alfredo Gragnani Filho
Grantee:Anthony Gueratto Klepp
Home Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil

Abstract

Title: keratinocyte growth factor and inflammatory markers in cultured dermal fibroblasts of patients with burns. Student: Anthony Gueratto Klepp Level: Scientific Initiation - Early project Advisor: Doctor Alfredo Grangnani Filho INTRODUCTION: Intense Inflammatory responses are triggered in burns that affect a large body surface area. Cytokines are the primary mediators of the inflammatory response to thermal injury by promoting communication between different cell types. It is expected that the cytokines IL-1, IL-6, IL-8, IL-10, IL-12 and TNF-± and KGF are changed in patients with burns affecting more than 25% of the body, which are large burned parts. It is estimated that by the fact that the production of cytokines and KGF are two known mechanisms of tissue repair Keratinocytes are recognized as a source of proinflammatory cytokines that stimulates migration of inflammatory cells and may have systemic effects on the immune system, influencing the proliferation and differentiation processes of keratinocytes and fibroblasts, and ultimately affecting the production of other cytokines by keratinocytes and fibroblasts. Dermal fibroblasts also release cytokines and growth factors with autocrine and paracrine actions that induce the secretion of collagen and fibroblast proliferation, and affect the growth and differentiation of keratinocytes, especially through the secretion of the KGF. In response, the keratinocytes produce IL-1 that stimulate fibroblasts to produce KGF and so close a double paracrine loop with the primary function of promoting the ideal environment for the formation of the epidermis and facilitate wound healing. The study and the highest level of evidence in relation to these inflammatory mediators are crucial for understanding the evolution of the burn patient, because these mediators are closely related to the level of inflammation that the patient is developing. Compare these parameters between large and small burns is critical, by the fact that these mediators can alter in any way the reepitelization of superficial injuries as well as the clinical course resulting in alterations in healing on the esthetical and functional result of scars produced. The results may determine changes in the therapeutic protocol used in the treatment of burns units. The lack of knowledge supported by adequate evidence, has determined protocols based on personal experiences, reflecting high levels of morbidity and mortality OBJECTIVE Estimate the level of keratinocyte growth factor (KGF) and IL-1², IL-6, IL-8, IL-10, IL-12,tissue necrosis factor alpha (TNF-a) in the media of cultured dermal fibroblasts of burn patients. METHODS Procedure (s) operation s) Obtaining skin samples necessary for the development of research will be performed through standard surgical procedure used for the treatment of burns, burn care unit of the DCP-UNIFESP-HU-HSP. Culture of Human Dermal Fibroblasts Isolation and culture of human dermal fibroblasts are made from fragments of normalskin from discarded surgical procedures performed in the burn care unit-DCP-UNIFESP-HU-HSP. Sample preparation Concentrations of IL-1ß, IL-6, IL-8, IL-10, IL-12p70 and tumor necrosis factor (TNF-±)will be analyzed in the culture medium of cells from patients by Cytometric Bead Array Human Inflammation Kit (CBA , BD Biosciences, USA) according to the manufacturer's instructions. Keratinocyte growth factor KGF have their dosage in the supernatant of primary human fibroblast cultures by the method of immunoenzyme assay (ELISA) using KGF