Tag: stem cells

Technology Transfer

Johns Hopkins Researchers Develop a Road Map to Stem…

Johns Hopkins Researchers Develop a Road Map to Stem Cell Development

June 10, 2019

The following was originally published in Johns Hopkins Medicine’s Newsroom.

Johns Hopkins Medicine researchers report they have created a method of mapping how the central nervous system develops by tracking the genes expressed in cells. The technique, demonstrated in mouse retinas for this study, follows the activity of the genes used by individual cells during development, allowing researchers to identify patterns in unprecedented detail. This precise kind of road map, say the researchers, could be used to develop future regenerative treatments for blinding and other neurological diseases.

“This is far and away the most comprehensive map we have of cellular development in the central nervous system,” says Seth Blackshaw, Ph.D., professor of neuroscience at the Johns Hopkins University School of Medicine, and a faculty member of the Johns Hopkins Institute for Cell Engineering.

This branching structure, seen here as a still from a moving illustration, allows researchers to see the genes turned on and off over the lifetime of a cell. (Video from Loyal Goff. Work by Brian Clark, Genevieve Stein-O’Brien, Fion Shiau, Gabrielle Cannon, Emily Davis, Thomas Sherman, Fatemeh Rajaii, Rebecca James-Esposito, Richard Gronostajski, Elana Fertig, Loyal Goff, and Seth Blackshaw.)

“If we can harness this kind of map to teach stem cells to create a certain type of retinal cell, we could someday replace cells lost to macular degeneration and other blinding diseases,” says Genevieve Stein-O’Brien, Ph.D., a postdoctoral fellow at the Johns Hopkins University School of Medicine in the laboratory of Elana Fertig, Ph.D., associate professor of oncology at the Johns Hopkins University School of Medicine and a member of the Johns Hopkins Kimmel Cancer Center.

A description of the work is published May 22 in the online edition of the journal Neuron. The technology is available for license through Johns Hopkins Technology Ventures.

Blackshaw says the retina, a well-understood structure that contains many of the cell types found throughout the rest of the nervous system and as such, is an excellent example to use in studies of how the central nervous system develops.

Each of the many cell types that make up the retina are produced by neural progenitors — stem cell-like cells that have the capacity to develop into virtually any retinal cell type depending on what genes are switched on and off during development. The gene patterns needed to create each cell type occur along a strict timeline. Neurons, like the light-absorbing rods and cones in the retina, are produced by younger progenitors, while supportive glial cells are created by older progenitors.

To study the process in detail, and create the road map, the researchers first sequenced the DNA of individual mouse retinal cells at different developmental time points — from the first progenitors to cells of the adult retina.

The researchers then put this information into a machine learning computer program that Stein-O’Brien developed. The program was designed to rapidly compress the immense amount of genetic data, grouping like cells together to generate a map that let researchers visualize the development process. The computer program produced a branching structure that allowed researchers to see which cell types give rise to others, which are most closely related, and what genetic changes occur that lead to the over 100 cell types found in the mouse and human retina.

“The map provides a way for us to see the impact individual genes and gene networks have on the developing central nervous system,” says Blackshaw.

In a proof-of-principle experiment, the researchers next chose to take a close look at three of the genes: nuclear factor 1 (NFI) a, b and x, which are essential to helping a progenitor determine its age and what types of retinal cells it can produce.

The researchers genetically engineered mice to either express the three genes in abundance or not at all, and observed how their retinal cells’ life cycles changed by tracking which genes were turned “on” at any given time in the computer program. They found that cells expressing increased levels of the NFI genes acted older than they were and produced more of the corresponding cell types (glial cells) than normal retinal progenitors. In contrast, progenitors without the NFI genes continued to create earlier cell types, such as rod photoreceptors, and continued to divide, acting like young progenitor cells.

Ultimately, the researchers say, they hope to apply the mapping technique to other cell types to better understand which genes affect the development of diseases in the body’s other tissues. “If we know precisely how progenitors get from an uncommitted stem cell population to mature tissues, we can use this road map to redirect them to specifically follow other paths, says Loyal Goff, Ph.D., assistant professor of neuroscience at the Johns Hopkins University School of Medicine and a faculty member at the McKusick-Nathans Institute of Genetic Medicine at Johns Hopkins.

Other researchers involved in this study include Brian Clark, Fion Shiau, Gabrielle Cannon, Clayton Santiago, Thanh Hoang, Fatemeh Rajaii and Rebecca James-Esposito of the Johns Hopkins University School of Medicine; Emily Davis-Marcisak of the Sidney Kimmel Comprehensive Cancer Center and the Johns Hopkins University School of Medicine; Thomas Sherman of the Sidney Kimmel Comprehensive Cancer Center and Richard Gronostajski of the University at Buffalo.

Funding for this research was provided by the National Eye Institute (R01EY020560, U01EY027267, F32EY024201, K99EY027844, K08EY027093), the National Cancer Institute (R01XA177669, U01CA212007) the New York State Stem Cell Science awards (C026429, C03133, C30290GG), the Chan-Zuckerberg Initiative DAF (2018-183445, 2018-183444), the Johns Hopkins University Catalyst, the Johns Hopkins University Discovery Awards and the Johns Hopkins University School of Medicine Synergy Awards.

Technology Transfer

Stem Cells Make More ‘Cargo’ Packets to Carry Cellular…

Stem Cells Make More ‘Cargo’ Packets to Carry Cellular Aging Therapies

May 9, 2019

The following was originally published in Johns Hopkins Medicine’s Newsroom.

Johns Hopkins scientists report that adult cells reprogrammed to become primitive stem cells, called induced pluripotent stem cells (iPSCs), make tiny “cargo packets” able to deliver potentially restorative or repairing proteins, antibodies or other therapies to aged cells. They say the human iPSCs they studied produced much more of the packets, formally known as extracellular vesicles, than other kinds of adult stem cells commonly used for this purpose in research.

Extracellular vesicles are naturally abundant in many types of cells, which use the cargo-containing spheres to communicate with other cells. They are about one one-hundredth the diameter of a cell and can carry anything from fats and proteins to nucleic acids. When a cell releases an extracellular vesicle, other cells nearby slurp up the tiny packet and its contents, making it an attractive target for packaging treatments for diseased cells that are deteriorating or aging prematurely, researchers say.

An iPSC-derived extracellular vesicle (Courtesy of Vasiliki Machairaki)

The Johns Hopkins scientists have filed for a patent related to the technology described in this report, which is available for license through Johns Hopkins Technology Ventures.

To package a potential treatment in an extracellular vesicle, scientists typically use a cell called a mesenchymal stem cell, which is found among fat or bone marrow cells and gives rise to other fat and bone cells. Scientists genetically modify the stem cell to produce vesicles with the treatment-related cellular therapy — usually a protein.

But the Johns Hopkins scientists say that mesenchymal stem cells aren’t the best sources for extracellular vesicles. The cells don’t multiply as often as iPSCs, and more cells are necessary to produce larger quantities of extracellular vesicles needed for therapeutic use. In addition, mesenchymal cells grow best in a liquid called fetal bovine serum, which contains potentially treatment-contaminating extracellular vesicles that are difficult to distinguish and separate from extracellular vesicles derived from mesenchymal cells.

By contrast, the liquid used to store and feed human iPSCs in the laboratory, called Essential 8, is free of extracellular vesicles and animal proteins, and the Johns Hopkins scientists found the cells could produce 16 times more vesicles than mesenchymal stem cells.

“We wanted to show other scientists working on such potential therapies that human iPSCs can efficiently produce highly purified extracellular vesicles that could, one day, be used to treat aging-related diseases,” says Linzhao Cheng, Ph.D., a professor of medicine and oncology at the Johns Hopkins University School of Medicine and a member of the Johns Hopkins Kimmel Cancer Center.

Cheng and his colleague Vasiliki Machairaki, Ph.D., assistant professor of neurology at the Johns Hopkins University School of Medicine, postdoctoral fellow Senquan Liu and other colleagues reported results of their experiments online Feb. 27 in Stem Cells.

The scientists fed extracellular vesicles made from human iPSCs and mesenchymal cells to human cells that had been genetically engineered to model a disease called progeria, which ages cells far more quickly than normal cells. When the scientists broke open the human recipient cells and analyzed their proteins, they found a more than two-fold increase in the amounts of several antioxidant proteins, called peroxiredoxins, inside the cells fed with extracellular vesicles made from iPSCs, which prevented the aging-related damage to cells. Antioxidants help to prevent aging-related damage to cells.

Cheng says that iPSCs may one day be best suited for delivering treatments for cellular aging-related diseases such as progeria and ALS (or Lou Gehrig’s disease), but not other age-related diseases such as cancer because iPSCs are naturally designed for promoting cell growth and regeneration, a process that can fuel cancer.

Machairaki cautioned that a great deal of research is needed before vesicles are deemed effective and safe for human disease therapy. The research team, she says, is currently planning studies in animal models to figure out how to overcome the potential for the immune system to reject extracellular vesicles, for example.

Other scientists who contributed to the research include Hao Bai, Zheng Ding, Jiaxin Li and Kenneth W. Witwer from Johns Hopkins.

The research is funded by the Maryland Stem Cell Research Fund and National Institutes of Health (R56 AG-057430).

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