Platelets Regulates Cell Viability and VEGF-A mRNA Expression in HaCaT Cell Line
Juliana Marques
Programa de Pós-Graduação Multicêntrico em Ciências Fisiológicas, Instituto de Biodiversidade e Sustentabilidade NUPEM, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Av. São José do Barreto, 764, Bairro Barreto, Cidade Macaé, CEP 27965-045, Brazil and Laboratório Integrado de Ciências Morfofuncionais, Instituto de Biodiversidade e Sustentabilidade NUPEM, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Av. São José do Barreto, 764, Bairro Barreto, Cidade Macaé, CEP 27965-045, Brazil.
Jackson de Souza-Menezes
*
Programa de Pós-Graduação Multicêntrico em Ciências Fisiológicas, Instituto de Biodiversidade e Sustentabilidade NUPEM, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Av. São José do Barreto, 764, Bairro Barreto, Cidade Macaé, CEP 27965-045, Brazil and Laboratório Integrado de Ciências Morfofuncionais, Instituto de Biodiversidade e Sustentabilidade NUPEM, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Av. São José do Barreto, 764, Bairro Barreto, Cidade Macaé, CEP 27965-045, Brazil.
*Author to whom correspondence should be addressed.
Abstract
Platelet Rich Plasma (PRP) is an autologous technique that uses centrifuged whole blood to concentrate platelets in plasma. The regenerative effect of PRP is attributed to platelets due to the release of growth factors involved in healing. This study sought to promote the isolation of platelets from PRP (PI-PRP) to identify the role of platelets in the modulation of cell surviving and VEGF-A mRNA expression. The coculture protocol with PI-PRP/ keratinocyte cell line HaCaT was established. Cellular viability by MTT, membrane integrity by trypan blue, cell and cytoskeletal cell morphology by DAPI and phalloidin staining and RNA extraction, for subsequently, qRT-PCR VEGF-A, were performed. The MTT test showed higher cell viability in PI-PRP group than CTRL. The trypan blue test showed no difference between CTRL and PI-PRP groups. Fluorescence microscopy analysis showed no changes in cellular morphology of the nucleus and distinct alterations in cytoskeletal between groups. In the qRT-PCR the VEGF-A expression was higher in PI-PRP group compared to CTRL. The centrifugation proved to be effective for platelet enrichment. This protocol demonstrated efficiency in studying interaction between platelet and cell lineage.
Keywords: PRP, keratinocyte, platelets, VEGF-A