The finding is important, investigators say, because the cells rescued by this treatment represent potential targets for future therapy in human babies with DS. And it suggests that similar success for other DS-related disruptions of brain growth, such as occurs in the hippocampus, could lead to additional treatments - perhaps prenatally - that restore memory and the ability to orient oneself in space.
DS is caused by an extra chromosome 21, a condition called trisomy - a third copy of a chromosome in addition to the normal two copies. Children with DS have a variety of abnormalities, such as slowed growth, abnormal facial features and mental retardation. The brain is always small and has a greatly reduced number of neurons.
A report on the Hopkins work with trisomic mice, led by Roger H. Reeves, Ph.D., professor in the Department of Physiology and the McKusick-Nathans Institute for Genetic Medicine at Hopkins, appears in the January 24 issue of the Proceedings of the National Academy of Sciences (PNAS).
Reeves and his team used an animal model of DS called the Ts65Dn trisomic mouse to show that pre-nerve cells called granule cell precursors (GCP) fail to grow correctly in response to stimulation by a natural growth-triggering protein. This protein, called Sonic hedgehog (Shh), normally activates the so-called Hedgehog pathway of signals in these cells. These signals stimulate mitosis (cell division) and multiplication of the cells in the growing, newborn brain, according to the researchers.
The GCP originate near the surface of the cerebellum and migrate deeper into the brain to form the internal granule layer (IGL), the researchers note. Therefore, the team studied the growth of the cerebellum in Ts65Dn trisomic mice at seven time points -- beginning at birth - to determine when GCP abnormalities first occurred. The IGL was similar in both normal and Ts65Dn mice at birth, but was significantly reduced in the trisomic mice by day six after birth.
Furthermore, the researchers found that the reduced number of GCP in these mice compared to normal mice was not due to cell death; rather, there were 21 percent fewer GCP undergoing cell division in Ts65Dn mice. This suggested that stimulating these cells might restore normal numbers of GCP, according to Reeves.
The Hopkins team then showed in test-tube experiments that GCP from the brains of Ts65Dn mice had a significantly lower response to increasing concentrations of a potent form of Shh called ShhNp. That is, increasing concentrations of ShhNp triggered increasing rates of mitosis. Despite their lower response, trisomic cells did show a dose response with increasing ShhNp concentrations.
"The fact that trisomic GCP responded to stimulation of their Hedgehog pathway even in a reduced way is significant," says Reeves, the senior author of the PNAS paper. "It suggested that these cells could be stimulated to reach normal levels of cell division by artificially increasing their exposure to Hedgehog growth factor."
Based on this initial discovery, the team injected into newborn Ts65Dn mice a molecule that stimulates the Hedgehog pathway to trigger cell growth. Treatment of the trisomic mice with this molecule, called SAG 1.1, restored both the numbers of GCP and the number of GCP cells undergoing mitosis to levels seen in normal mice by six days after birth.
"The normal mouse cerebellum attains about a third of its adult size in the first week after birth," says Randall J. Roper, Ph.D. "This is the time during which SAG 1.1 treatment of Ts65Dn restored GCP populations and the rate of mitosis of those cells," he adds. "However, further research is needed to determine if it's possible to reverse the effects of trisomy in other parts of the DS mouse." Roper is a postdoctoral fellow in the laboratory of Reeves and a co-first author of the PNAS paper.
The other authors of the Hopkins paper include Drs. Laura L. Baxter, Nidhi G. Saran, Donna K. Klinedinst, and Philip A. Beachy. Baxter is a co-first author of this paper and is currently at the National Human Genome Research Institute of the National Institutes of Health (Bethesda, Md.).
This work was supported in part by the Public Health Service. P.A.B. is a Howard Hughes Medical Institute investigator.
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