By David N. Leff

Worldwide, about 200 sufferers from far-advanced Parkinson's disease have received brain transplants of human fetal brain tissue in the past decade.

That tissue came from the substantia nigra, a region of the brain's mesencephalon that produces dopamine. This is the potent neurotransmitter that dwindles away in the aging brains of people with Parkinson's disease (PD). (See BioWorld Today, May 4, 1998, p. 1.)

The empirical rationale that prompted this draconian treatment was that the substantia nigra contains cells specialized in churning out dopamine, to replace that lost from diminishing dopaminergic neurons.

"What's been going on," observed neuroscientist Ron McKay at NINDS — the National Institute of Neurological Disorders and Stroke, "is that so far people have been engrafting tissue that is immediately obtained from the fetal nervous system. At the moment," he went on, "the difficulty with that current technology is that there is no easy access to large numbers of quality-controlled neural stem cells. We have shown that you can actually get these cells to grow in culture."

McKay, who heads the laboratory of molecular biology at NINDS, is senior author of a paper in the August 1998 issue of Nature Neuroscience. Its title is "Transplantation of expanded mesencephalic precursors leads to recovery in parkinsonian rats."

"Early on in mammalian development," he explained, "there's a cell called an embryonic stem cell. That's an all-purpose cell that people use when they want to do genetic engineering — homologous recombination — in mice.

"Each somatic tissue in the body," McKay pointed out, "is thought to have its own stem cell, which goes right to generating the major cell types of that tissue. The neural stem cell that we've identified is abundant in the fetal brain during development of the nervous system. It gives rise to the neurons, astrocytes and oligodendrocytes that are the brain's three major cell types."

He recalled, "The first demonstration that you could identify a cell that was a precursor to neurons was evidence from my group, published in 1988."

Expand, Differentiate, Aggregate, Transplant

As recounted in their present paper, McKay and his co-authors harvested rodent neural stem cells from the substantia nigra of 12-day-old rat embryos. That initial step started a three-phase process that ended two weeks later in adult rats with brains previously altered to mimic the motor disorder of PD.

Phase One consisted of expanding those few fetal cells 30-fold to a population that could restore enough dopamine to correct the rats' mindless, endless rotating around an imaginary axis — hallmark of their pseudo-PD.

To stimulate neural stem cell expansion, the team laced the culture dish with basic fibroblast growth factor (bFGF), a mighty mitogenic galvanizer of cell division. "If those stem cells aren't dividing," McKay pointed out, "they'll start differentiating too soon. And we actually used bFGF as a trick to differentiate when the timing became right."

Phase Two caused those thickly multiplying stem cells to quit growing, after six or eight days, and begin differentiating into neurons specialized in secreting dopamine.

"We got the cells," he recalled, "from the part of the brain that's going to give rise normally to these dopaminergic neurons, which normally express tyrosine hydroxylase. If the expanded cells remember where they came from," he added, "they'll differentiate into the class of neurons you'd expect."

Phase Three, aggregation, saw the newly hatching neurons removed from the tissue-culture plates to which their precursors had clung, and rotated in a tube inside the tissue culture incubator.

"That prevents the cells from attaching to the surface," McKay pointed out. "So what they do under those new culture conditions is attach to each other and differentiate. This forms these aggregating neurons into barely visible, free-floating spheres of a particular size that will fit into a needle. Then they can be grafted without further disruption into the rats' brains."

Each PD-mimicking rodent in this preclinical in vivo trial received injections of the dopamine-dedicated neurons into only one side of its brain. The opposite side served as a control. Eighty days after the transplants, three-quarters of the animals were able to overcome their slow spinning motion, showing that the transplant had worked.

From Rats To Folks In Two To Three Years

McKay foresees clinical trials of expanded, differentiated and aggregated human neural stem cells in the nearest feasible future. "Human cells," he observed, "appear to be very similar to rodent ones.

"I think there is no question," he stated, "that clinical trials are going to be pursued. It's just a question, really, of getting all the pieces in place, and going forward without delay. The delay is not so much that there's any particular experimental hurdle. It's mostly generating enough human stem cells and getting organized."

McKay estimates that such human studies can begin "within two to three years from now. It's certainly not clear when we will know if the transplant will work in people. My guess about that is between three and four years."

Two years ago, McKay and his coworkers showed that "there are cells in the adult nervous system with essentially the same properties as the fetal neural stem cells. That's been an important finding in the field, because in neurobiology the idea that the adult brain does not have the powers to regenerate is being radically changed now. One of the factors that's leading to this altered view is the finding that the adult brain has stem cells in it."

He continued: "The question remains: Why don't those neurons regenerate? It's not because they don't have the cells, which is what people have previously thought. The answer," he surmised, "is likely to come out of a deeper understanding of the basic biology involved, of the mechanisms that control the differentiation of neural stem cells."

McKay has submitted the work described in this Nature Neuroscience journal for possible patenting, "through the NIH system, in what they call an 'employee invention report.' They're looking at this at the moment," he said, "and it's my guess they might actually go forward. There are things in here that are potentially commercially useful.

"Besides any such commercial considerations," McKay concluded, "the ability to transplant populations of cultured human dopaminergic neurons into the brains of Parkinson's patients has a likely ethical benefit as well. It would minimize the controversial use of human fetal tissue." *