Experiments point to new theory of skeletal development

Curious children and developmental biologists have long pondered the question: what makes a thumb a thumb and a pinkie a pinkie? The answer UW Medical School researchers have found may force scientists to revise their theories of how cells of the developing skeleton organize into exquisitely patterned tissue, from fingers to spines.

In various microsurgical experiments on the developing toes of chicken embryos still in the egg, the researchers discovered that the future uniqueness of each budding embryonic digit is determined by the webbed tissue located next to it. Until now, scientists considered this interdigital tissue, which disappears before birth, to be of little importance. It was interesting mainly because in most species it is eliminated by programmed cell death in order to free digits from one another.

“We were very surprised to find that developing cartilage, which eventually forms the skeleton, gets the information about what it will ultimately look like from surrounding soft tissue,” said John Fallon, UW Medical School Harland Winfield Mossman Professor of Anatomy. “The general thinking has been that embryonic cartilage, digits and limbs are self-differentiating systems that develop independently.”

Fallon and graduate student Randall Dahn report the find in the July 21 issue of Science.

In their experiments, they also found that specific levels of a substance called bone morphogenic protein (BMP), which is produced by interdigital web cells, play a crucial role in determining the distinct characteristics, or identity, of each digit.

The UW team set out to test the existing, decades-old theory of how a developing digit learns that it is destined to become, for example, a big toe. According to the theory, a gene called Sonic hedgehog somehow gives each precursor digit detailed instructions about its particular identity. The developing tissue follows through on the cue days later, after Sonic hedgehog is no longer produced, remembering exactly which digit it should become.

Dahn separated one precursor chicken digit into two halves by surgically inserting a piece of foil between them. By the existing theory, each half should contain the same positional information and develop into two digits of the same identity. To the researchers’ surprise, one half formed the digit it was supposed to form, but the other half transformed to the identity of the neighboring digit.

Translating to human anatomy terms, Dahn explained that by splitting the pointer finger in two, the half nearest the middle finger would develop into a pointer and the half nearest the thumb would develop into a thumb.

“This showed us that digital identity is not a fixed, inherent property under the direct influence of Sonic hedgehog in early development. Now we know that the identity of each finger is progressively determined as it develops,” he said.

The researchers then demonstrated that the interdigital tissue (ID) that lies between budding digits is responsible for instructing them as to what they will become. Specifically, each ID instructs the identity of the digit located anterior to it, towards the thumb or big toe.

Fallon and Dahn next investigated signalling molecules that IDs might use to send their messages to precursor digits. The researchers turned to the large family of BMPs because the proteins are involved in patterning many tissues of the body, and may contribute to ID cell death.

By manipulating interdigital BMP levels, the Wisconsin researchers transformed the identities of the neighboring digits. Taken together with reports from other scientists, this observation implies that the developing embryo uses BMPs to influence the way individual structures in a series-fingers, teeth, vertebrae-acquire their own discrete identity.

“It’s possible that the precise development of all skeletal elements in the body requires input from surrounding tissue,” said Dahn. “These new findings may eventually be clinically useful in providing novel insights into genetic diseases of the skeleton as well as the repair and replacement of cartilage and bone.”

Adds Fallon, “Another intriguing aspect of the work is that it may have implications for understanding how to bring about the regeneration of normally patterned human digits after traumatic amputation.”

Fallon’s teams have studied tissue patterning in the developing embryo for some 30 years, analyzing the way asymmetrical digits and limbs emerge and become fixed in newts, turtles, reptiles and humans.

The research was funded by a National Institute of Child Health and Human Development grant to Fallon and a National Science Foundation fellowship to Dahn.