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(Investigative Ophthalmology and Visual Science. 2000;41:601-609.)
© 2000 by The Association for Research in Vision and Ophthalmology, Inc.

Human Platelet Suspension Stimulates Porcine Retinal Glial Proliferation and Migration In Vitro

Laurent Castelnovo1, Christine Dosquet2, Alain Gaudric3, José Sahel1 and David Hicks1

1 From the Laboratoire de Physiopathologie Rétinienne, Clinique Ophthalmologique, INSERM–Université Louis Pasteur E9918, Centre Hospitalier Régional Universitaire, Strasbourg Cedex, France; the 2 Laboratoire des Cytokines, Etablissement de Transfusion Sanguine de l’AP-HP, Hôpital St. Louis, 1 Avenue Claude Vellefaux, 75475 Paris, France; and the 3 Service d’ Ophthalmologie, Hôpital Lariboisière, 75010 Paris, France.

PURPOSE. To characterize the cellular and molecular mechanisms underlying the efficacy of autologous platelet suspension adjuvant therapy in the treatment of macular hole.

METHODS. Platelet suspensions were prepared from whole blood samples obtained from informed volunteers. For proliferation assays, platelet suspensions or purified growth factors were added to semi-confluent cultures of porcine retinal glial cells for 24 hours, followed by [3H]thymidine for 15 hours, after which time cells were washed, solubilized, and counted for uptake of radioactive tracer. For cell migration assays, confluent glial cultures were scrape wounded and maintained in the presence or absence of platelet suspension or identified platelet constituents. Cell migration into the denuded area was scored as a function of time. In certain cases, specific pharmacologic inhibitors of growth factor action were added at the same time as platelet adjuvant or growth factors.

RESULTS. Platelet suspension adjuvant induced strong mitogenic and chemotactic responses in cultured glia, in a dose-dependent manner. Maximal incorporation of thymidine was two- to threefold that of control levels, with an ED50 ~5 x 106 platelets/ml, and migration was enhanced up to 80-fold after 48 hours. Platelet suspension-induced proliferation was completely blocked by addition of 25 µM genistein, a tyrosine kinase receptor inhibitor. However, the same concentration only partially blocked the cell migration response. Addition of any single growth factor or protein identified from ELISA analysis, or a combination of all factors, did not significantly stimulate proliferation or cell migration.

CONCLUSIONS. Human platelet suspensions exert both proliferative and chemotactic influences on retinal glial cells in vitro, suggesting that the same responses may occur in platelet-induced macular hole repair in humans. Growth factors or proteins that have been identified within the suspensions do not mimic these responses in vitro, implying that additional currently unidentified trophic activities are also present.






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