This study investigates a method for transnasal transplantation of induced pluripotent stem cell-derived human microglia (iPSMG) in mice. The approach aims to bypass the blood-brain barrier and enhance the delivery of therapeutic cells to the brain.
Take an anesthetized mouse that has been fed an inhibitor-containing diet.
In the brain tissue, this inhibitor selectively eliminates microglial cells.
Apply a hyaluronidase solution to both nostrils.
Place the mouse briefly in the supine position to prevent drainage of the solution.
Hyaluronidase breaks down the extracellular matrix of the nasal mucosa, loosening the cells and enhancing tissue penetration.
Administer induced pluripotent stem cell-derived human microglia or iPSMG cell suspension into one nostril.
Place the mouse in the supine position. Then, administer the suspension into the other nostril.
Repeat the administration several times.
The iPSMG cells penetrate the nasal mucosa and follow the olfactory pathway to reach the brain, thereby bypassing the blood-brain barrier.
Allow the mouse to recover.
Anesthetize the mouse and administer transplantation media intranasally at regular intervals.
Growth factors in the media support the growth and survival of transplanted iPSMG cells in the mouse brain.
To prepare mice for transnasal transplantation, feed the male mice with a diet containing PLX for seven days. At the end of the seventh day, seize the PLX diet and feed the mice with a normal diet. One hour before transnasal transplantation of iPSMG, administer 2.5 microliters of hyaluronidase in PBS to each nostril twice, using a 10 microliter pipette tip to increase the permeability to the nasal mucosa.
Place the mice in the supine position after the hyaluronidase application. Administer 2.5 microliters of hyaluronidase 10 minutes before the transnasal transplantation of iPSMG. Apply 2.5 microliters of cell suspension into one nostril of the mouse using a 10 microliter pipette tip. Place the mouse in the supine position for five minutes before administration of cell suspension to the other nostril.
Administer the cell suspension four times, applying a total volume of 20 microliters per animal. 48 hours after the cessation of PLX feeding, repeat the administration of hyaluronidase and cell suspension on the same mice. For administration of cytokines, apply 2.5 microliters of the transplantation medium into one nostril of the mouse using a 10 microliter pipette tip.
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