Abstract
ABSTRACT Introduction: Gliomas are characterized by rapid proliferation and aggressive invasion into normal surrounding brain tissue. In medical laboratories, the in vitro wound healing assay stands out as a simple, easy, inexpensive and affordable method to evaluate cell migration and proliferation. Objective: To standardize the in vitro wound healing assay using antimicrotubule drugs as positive controls. Methods: U87MG glioma cells were seeded at different densities and, after 24 h, the monolayer was scratched using different micropipette tip size to create a gap with no cells. The cells were then treated with colchicine and paclitaxel in culture medium with the presence or absence of fetal bovine serum. The wound was photographed with the aid of an inverted microscope and the wound area was measured using the Image J software. Results: Better defined edges scratches and monolayer with approximately 90% confluence were obtained at 1.5 and 2 x 105 cells/well density. The width and area of the scratch were, respectively, 948 pm/2.193221 mm2; 964 pm/2.266 mm2 and 1448 pm/3.221 mm2 to 10. 200 and 1000 pl micropipette tips. Colchicine inhibited wound closure by 12.6% or 3.4%, both in the presence or absence of serum; paclitaxel 2.4 and 6.7% respectively. Conclusion: Under standardized conditions, colchicine and paclitaxel proved to be efficient positive controls into the in vitro wound healing assay.
Highlights
Gliomas are characterized by rapid proliferation and aggressive invasion into normal surrounding brain tissue
A cell-free area is generated in a confluent cell monolayer and the wound closure rate and cell migration can be quantified by time-lapsing photography with an inverted microscope at some time intervals[1]
Cell migration is essential for many biological processes, such as tissue repair and regeneration; the aberrant regulation of this process drives the progression of many diseases, including cancer invasion and metastasis[2]
Summary
Gliomas are characterized by rapid proliferation and aggressive invasion into normal surrounding brain tissue. The in vitro wound healing assay, known as the “scratch assay” is a simple, versatile, and cost-effective method for studying cell migration. In this methodology, a cell-free area is generated in a confluent cell monolayer and the wound closure rate and cell migration can be quantified by time-lapsing photography with an inverted microscope at some time intervals[1]. Cell migration is essential for many biological processes, such as tissue repair and regeneration; the aberrant regulation of this process drives the progression of many diseases, including cancer invasion and metastasis[2] Cytoskeleton components, such as the actin and tubulin filaments, are crucial for cell migration. Drugs capable of reaching both actin filaments[5], such as the microtubule network, formed by tubulin[6], are capable to promoting deregulation of cell structure and disruption of proliferative processes
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