A three-dimensional culture model and application of glioma stem cells in vitro

Abstract

The invention belongs to the field of cell biology, and in particular relates to an in vitro three-dimensional culture model for studying vascular mimicry of glioma stem cells and its application. The preparation of the in vitro three-dimensional culture model is carried out according to the following steps: the three-dimensional collagen scaffold is soaked in DMEM medium for 6-24 hours and then taken out, and the redundant DMEM adhered to the three-dimensional collagen scaffold is removed to obtain a pretreated three-dimensional collagen scaffold. Treat the three-dimensional collagen scaffold and place it in a cell culture plate, plant the glioma stem cells into the pretreated three-dimensional collagen scaffold, let it stand at 37°C for 2-6 hours, add endothelial medium, and culture at 37°C for 2-4 days , which is an in vitro three-dimensional culture model for studying vascular mimicry of glioma stem cells. All operations of this in vitro three-dimensional culture model are carried out at room temperature, and various stainings can be performed in situ, including immunohistochemistry and immunofluorescence staining; electron microscope detection is possible; RT‑PCR and Western Blot detection can be performed directly after digestion .

Description

technical field [0001] The invention belongs to the field of cell biology, and in particular relates to an in vitro three-dimensional culture model for studying vascular mimicry of glioma stem cells and its application. Background technique [0002] The growth of tumors and the formation of distant lesions depend on blood supply. According to the classic theory of angiogenesis, when a solid tumor grows to 1-2mm 3 When tumors are vaccinated, they need to induce the formation of new blood vessels to obtain blood supply, otherwise the tumor will stop growing. With the deepening of the research on tumor biology, in recent years, people have also discovered that tumor cells can deform themselves and form a lumen around them, in a way that is independent of endothelial cell-dependent vascular mimicry (VM). Construct tumor microcirculation. Since Maniotis et al. reported the existence of a new tumor microcirculation model, vascular mimicry (VM), in malignant melanoma in 1999, peo...

AI Technical Summary

Problems solved by technology

However, Matrigel glue is essentially a kind of Matrigel, which only provides a matrix for cell culture, not a real three-dimensional culture system.

In addition, due to the reasons of Matrigel itself, the following disadvantages were found during the research process: (1) Matrigel is jelly-like at room temperature, and the operation must be carried out in a low temperature environment below 4°C, which will cause damage to cells; (2) Because some cell phenotypes of cells in two-dimensional culture are different from those in three-dimensional culture, we want to find a cell culture model that can perform in situ detection when studying VM, but Matrigel glue is not easy to dissolve and the background is easy to color and other reasons, it cannot be tested in situ in this culture model

At present, Matrigel glue is only used to verify whether cells can form tube-like structures under certain treatment conditions, which largely limits the research progress of in vitro vascular mimicry

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