In a new research report, scientists at Johns Hopkins Medicine say they have developed a technology that allows them to capture a 3D molecular “snapshot” of hundreds of hair follicles as they develop and then reconstruct the fourth dimension — time — to effectively create a stop-motion animation of how this organ forms.  

The National Institutes of Health-funded research was published online July 1 and will appear in the Sept. 3 print issue of Cell. 

Hair follicles are the smallest, most numerous organs in our body but they form in similar ways as the other organs, the scientists say. This advancement thus paves the way for understanding how organ development may go awry for people with certain congenital conditions.   

“Our four-dimensional (4D) map of the hair follicle from mice serves as a model system for understanding broad-stroke fundamentals of how organs develop,” says Reza Kalhor, Ph.D., associate professor of biomedical engineering at the Johns Hopkins University School of Medicine, who led the recent study. 

By analyzing the 4D map, the researchers were able to identify distinct phases in this organ’s formation — a complex choreography of thousands of cells. First, the precursor cells organize themselves in space to establish the organ’s spatial axis perpendicular to skin surface. Next, these precursors differentiate into the many cell types needed to form hair follicles. Finally, these new cell types grow and morph into a mature follicle, preparing to produce the hair strand.  

The research further compares the hair follicles of normal mice to hairless mice lacking a gene, Foxn1, which is critical for hair growth. This comparison may help scientists understand how medical conditions that cause hair loss develop, which may lead to new ways to prevent and treat hair loss in the future, says Luis Garza, M.D., Ph.D., a professor of dermatology at the Johns Hopkins University School of Medicine and co-author of the paper.   

Scientists have long sought out a way to visualize how organs develop over time. Doing so may lead to a better understanding of how certain inherited conditions may affect organ development, or capture how and when tumors develop, and could lead to earlier diagnoses and treatments, Kalhor says. 

“This problem remains complicated because organs are generated by millions of cells in very complex, coordinated processes,” says Soichiro Asami, the first author of the paper and a Ph.D. candidate in Kalhor’s lab. “We developed a technology that tackles the problem of visualizing this, and applied it to hair follicles, because there are hundreds of them spanning each stage of organ development.”  

Biomedical engineers Kalhor and Asami partnered with Garza, who frequently studies Foxn1 mice as a model for understanding how hair growth disorders develop and to find new ways to treat them. Garza provided key context and background on hair follicle biology and tissue samples from normal and hairless mice in this recent study.  

The researchers developed a new molecular imaging tool, 3D DNase-Enhanced Expression Profiling (3DEEP), that allows them to analyze pieces of tissue large enough to capture hair follicle organs in entirety in skin samples of normal mice and bald mice. 3DEEP removes genomic DNA from the skin samples, which can interfere with chemical reactions used to visualize fragile pieces of messenger RNA essential for understanding organ development. From there, the researchers labeled the positions of millions of RNA molecules in the sample, providing a precise, 3D spatial map of gene expression within the tissue. 

Then, the scientists classified the cell types in these complex organs, calculated the molecular age of each hair follicle and lined up the hair follicles from youngest to oldest, turning frozen 3D snapshots of the skin of normal and hairless mice into 3D stop-motion animation. Jean Fan, Ph.D., assistant professor of biomedical engineering at Johns Hopkins, created an online interface for exploring and interacting with this animation.  

“These tools show us how a hair follicle grows from a tiny thickening of the skin to a  deep, matured structure,” says Garza. “It is a window into organogenesis.” 

Comparing the maps of hair follicle samples from normal and hairless mice, the scientists say they determined that hair follicles from hairless mice experienced delayed development as they formed. Notably, the cells from hairless mice follicles were able to proliferate, or divide and grow, at a higher rate, but had a reduced ability to mature and take on specific roles at the right time, destabilizing the delicate choreography of the organ’s development, Kalhor says. 

“The Foxn1 mutation led to a breakdown in cellular communication and timing, causing the hair follicle organs to structurally collapse before hair had the chance to form,” says Asami. 

In the future, Garza says this technique of visualizing hair follicle development has the potential to help researchers understand the cause of certain hair loss conditions in people.  

“Eventually if we can apply this technology to people, then we can find out a tremendous amount more from each patient and help individually treat them,” Garza says.  

In addition to Asami, Fan, Garza and Kalhor, another Johns Hopkins scientist, Chenshuo Yin, contributed to this paper. 

Funding for this research was provided by the National Institutes of Health (R01HG012357, U01HL156056), the Simons Foundation and the David & Lucile Packard Foundation.