Scientists at Johns Hopkins Medicine used glowing chemicals and other techniques to create a 3D map of the blood vessels and self-renewing “stem” cells that line and penetrate a mouse’s skull. The map provides precise locations of blood vessels and stem cells that scientists could eventually use to repair wounds and generate new bone and tissue in the skull.
We have to see what is going on inside the skull, including the relative locations of blood vessels and cells and how their organization changes during injury and over time. “
Warren Grayson, Ph.D., Professor of Biomedical Engineering and Director of the Craniofacial and Orthopedic Tissue Engineering Laboratory, Johns Hopkins University School of Medicine
His lab focuses on developing biomaterials and transplanting stem cells into the skull to recreate the missing bone tissue.
Other scientists have provided maps of small portions of blood vessels and stem cells in the skulls of mice. “However, a larger image of the skull gives us a better understanding of the entire vascular system and the distribution of different types of stem cells,” says Alexandra Rindone, graduate student at Johns Hopkins University and the Faculty of medicine and first author of the article.
The new map, published on October 28 in Nature Communication, is a 3D view of the top of a mouse skull – its cranial bone, or calvaria – which is made up of four connected skull bones.
To create the map, which includes hundreds of thousands of cells, Johns Hopkins researchers used four key techniques to locate the vessels and cells.
First, they used immunofluorescence to label molecules on the surface of a variety of blood vessels and stem cells with a fluorescent or bright chemical.
Next, scientists use a chemical compound that helps light enter the skull without being scattered – a method called optical tissue cleaning. “It makes the skull appear like glass,” says Rindone.
To take the 3D image, the scientists used an optical microscope, a device that takes images of large sections of tissue at high resolution and at high speed, but minimizes photobleaching. “This tool helps us prevent deterioration of the fluorescent dye when tissues are exposed to light sources for a long time,” says Rindone.
Finally, they used computer software to identify and segment the 3D cellular structures of the skull and recreate the spatial coordinates and volumes of the structures. “This shows us the prevalence of stem and bone cells and their orientation in the skull,” says Rindone.
The map revealed previously unknown niches in the skull where stem cells reside, especially near structures called transcortical channels, which are small channels that penetrate the bone of the skull and connect the outer walls of the skull to the central cavities. containing bone marrow.
Johns Hopkins scientists are working to adapt the method of creating 3D maps to image the human skull, which is difficult due to the large size of the human skull and the way light passes through it. Still, the method could be used to create 3D maps of cell types in bone and other human tissues.
Rindone, AN, et al. (2021) Quantitative 3D imaging of the cranial microvascular environment at unicellular resolution. Natural communications. doi.org/10.1038/s41467-021-26455-w.