Stem Cell Basics
What are embryonic stem cells?
A. What stages of early embryonic development are important for generating embryonic stem cells?
Embryonic stem cells, as their name suggests, are derived from embryos. Most embryonic stem cells are derived from embryos that develop from eggs that have been fertilized in vitro—in an in vitro fertilization clinic—and then donated for research purposes with informed consent of the donors. They are not derived from eggs fertilized in a woman's body. The embryos from which human embryonic stem cells are derived are typically four or five days old and are a hollow microscopic ball of cells called the blastocyst. The blastocyst includes three structures: the trophoblast, which is the layer of cells that surrounds the blastocoel, a hollow cavity inside the blastocyst; and the inner cell mass, which is a group of cells at one end of the blastocoel that develop into the embryo proper.
B. How are embryonic stem cells grown in the laboratory?
Growing cells in the laboratory is known as cell culture. Human embryonic stem cells are isolated by transferring the inner cell mass into a plastic laboratory culture dish that contains a nutrient broth known as culture medium. The cells divide and spread over the surface of the dish. The inner surface of the culture dish is typically coated with mouse embryonic skin cells that have been treated so they will not divide. This coating layer of cells is called a feeder layer. The mouse cells in the bottom of the culture dish provide the inner cell mass cells a sticky surface to which they can attach. Also, the feeder cells release nutrients into the culture medium. Researchers have devised ways to grow embryonic stem cells without mouse feeder cells. This is a significant scientific advance because of the risk that viruses or other macromolecules in the mouse cells may be transmitted to the human cells.
The process of generating an embryonic stem cell line is somewhat inefficient, so lines are not produced each time an inner cell mass is placed into a culture dish. However, if the plated inner cell mass cells survive, divide and multiply enough to crowd the dish, they are removed gently and plated into several fresh culture dishes. The process of re-plating or subculturing the cells is repeated many times and for many months. Each cycle of subculturing the cells is referred to as a passage. Once the cell line is established, the original cells yield millions of embryonic stem cells. Embryonic stem cells that have proliferated in cell culture for six or more months without differentiating, are pluripotent, and appear genetically normal are referred to as an embryonic stem cell line. At any stage in the process, batches of cells can be frozen and shipped to other laboratories for further culture and experimentation.
C. What laboratory tests are used to identify embryonic stem cells?
At various points during the process of generating embryonic stem cell lines, scientists test the cells to see whether they exhibit the fundamental properties that make them embryonic stem cells. This process is called characterization.
Scientists who study human embryonic stem cells have not yet agreed on a standard battery of tests that measure the cells' fundamental properties. However, laboratories that grow human embryonic stem cell lines use several kinds of tests, including:
D. How are embryonic stem cells stimulated to differentiate?
Figure 1. Directed differentiation of mouse embryonic stem cells.
As long as the embryonic stem cells in culture are grown under appropriate conditions, they can remain undifferentiated (unspecialized). But if cells are allowed to clump together to form embryoid bodies, they begin to differentiate spontaneously. They can form muscle cells, nerve cells, and many other cell types. Although spontaneous differentiation is a good indication that a culture of embryonic stem cells is healthy, it is not an efficient way to produce cultures of specific cell types.
So, to generate cultures of specific types of differentiated cells—heart muscle cells, blood cells, or nerve cells, for example—scientists try to control the differentiation of embryonic stem cells. They change the chemical composition of the culture medium, alter the surface of the culture dish, or modify the cells by inserting specific genes. Through years of experimentation, scientists have established some basic protocols or "recipes" for the directed differentiation of embryonic stem cells into some specific cell types (Figure 1).
If scientists can reliably direct the differentiation of embryonic stem cells into specific cell types, they may be able to use the resulting, differentiated cells to treat certain diseases in the future. Diseases that might be treated by transplanting cells generated from human embryonic stem cells include Parkinson's disease, diabetes, traumatic spinal cord injury, Duchenne's muscular dystrophy, heart disease, and vision and hearing loss.