Scientists at Johns Hopkins used a combination of molecular labeling and imaging techniques to create a three-dimensional map of the blood vessels in the mouse skull. Their approach also reveals niches where stem cell populations lurk, which could help researchers to understand how blood vessels and cells behave in various states of disease or injury. The researchers are interested in tissue engineering to replace lost bone, and their new methodology could allow them to understand how interventions to repair bone defects in the skull, such as biomaterial implants, are working. Understanding the distribution of specific cell types and blood vessels within our body is no easy task, and detailed maps are hard to come by. Part of the issue is the lack of transparency of our tissues, which do not easily permit detailed viewing. Dissecting tissues allows for some exploration, but damages the delicate structures we want to see, making it difficult to get an overall picture of the tissue of interest. “We need to see what’s happening inside the skull, including the relative locations of blood vessels and cells and how their organization changes during injury and over time,” said Warren Grayson, a researcher involved in the study, in a Hopkins press release. To achieve this, his team has developed a series of tissue processing, staining, and imaging steps that allow them to create impressive 3D maps of the vessels and cells within the skull. To begin, the team used immunofluorescent stains to label specific cell types and blood vessels with identifying tags. They then used a chemical that reduces the opacity of tissue, allowing light to penetrate through. The researchers describe the cleared tissue as resembling glass. Finally, they used a lightsheet microscope to obtain high-resolution images of the skull area, without bleaching out the fluorescent signals from the immunostains. “This tool helps us avoid deterioration of the fluorescent dye when tissues are exposed to light sources for a long time,” said Alexandra Rindone, another researcher involved in the study. The results are quite impressive, with the researchers using the map to identify niches within the skull that contain communities of stem cells. They may be able to target such regions to help with the healing process when repairing skull defects. These stem cell communities were previously unknown and reside near the transcortical canals within the skull. Source
