Engineers design bridges that span five-mile gulfs, and stand up to the twisting forces of the wind and the pounding of thousands of cars and 30-ton trucks, day in and day out. Engineers create industrial processes that pump out a myriad of materials each day. Engineers fabricate supercomputers that can make more than a trillion calculations each second.

But all these feats seem modest next to the challenges that the University of Michigan's David Mooney and other biomedical engineers are taking on. Melding the approaches and techniques of chemical engineers with the knowledge developed by cellular and molecular biologists over the last twenty years, Mooney is trying to create replacement body parts grown from human cells.

In the not-so-distant future, young mothers with leaky heart valves, children with defective livers, grandmothers who have lost breasts to cancer, or grandfathers with clogged heart arteries may be able to choose "grow-your-own" replacement tissues to make their bodies whole again.

Tissue problems-organs that fail, blood vessels that clog, and other body parts that are cancer-riddled, ripped, and worn-account for half our staggering $1 trillion in medical costs each year. Helping solve these problems in less costly ways, says Mooney, an assistant professor of dentistry and chemical engineering, is his basic motivation for doing this work.

A tissue is more than a big lump of cells. It has a specific organization, with certain types of cells located in particular patterns, all held together by an extracellular matrix. This matrix is composed of several types of molecules, generated by the cells. It provides a structure for the tissues, as well as biochemical needs. Tissues have specific functions, and to carry these out these jobs cells communicate with one another and with other parts of the body as necessary. Thus, the recipe for making a tissue is more complicated than tossing a mix of cells into a lab dish, incubating for an appropriate time, and then removing a well-formed tissue.

Instead, biomedical engineers are trying to provide the right structure, cues, and environment in the lab to grow tissues that mimic natural ones. Mooney combines biodegradable artificial materials and living cells to recreate body tissues.

Someday these replacement tissues could help alleviate severe shortages of organ transplants. "It provides the advantages of an organ tissue transplant with the availability of a prosthesis," Mooney explains. In addition, if replacement organs could be engineered from the patient's own cells, the recipient would be spared the need for drugs that prevent rejection.

And to get around the long-term problems that develop with artificial materials implanted in the body, such as the silicon breast implants that have sparked controversy, Mooney is working with biodegradable materials that in time would leave no trace of "foreign" substances in the person.

To engineer these tissues, Mooney creates a three-dimensional structure of biodegradable material that is "seeded" with tissue cells. In one research project underway, he loads the scaffold with body cells that normally make up smooth muscle tissue. The research team hopes the cells will proliferate and fill in the structure, forming a new tissue that functions like the smooth muscle found in heart valves, blood vessels, and the intestines. Mooney's research is composed of numerous steps in understanding how to make this happen and why it happens.

He uses the scaffolding material as a template to recreate the structure of the tissue. Mooney wants the scaffold to "dissolve," just as absorbable stitches do, so that in the end what remains is a totally biological material.

The scaffolding constituents are synthetic polymers that breakdown in the presence of water. The biodegradable material comes in many forms: fiber mats, tubes, or sponges. Mooney has coated the some filaments of the fiber mat with the other materials to give the mat more stability and flexibility. He has shown that cells will proliferate on a flat sheet of the polymer. Now he and graduate students Byung-Soo Kim, Martin Peters, and Leatrese Harris, are fashioning the scaffolding into three-dimensional structures.

The research team loads the structures with cells from rat muscle tissue, incubates the device under the right temperature, and keeps it bathed with the proper levels of oxygen and nutrients. The cells multiply on the scaffold to fill in the spaces and create the extracellular matrix around themselves.

So that the tissue created is stable but flexible, the researchers must pace three things: cellular proliferation, extracellular matrix formation, and the scaffolding degradation. If the scaffolding degrades before the tissue is well enough developed, then the tissue will collapse. However, if the scaffolding degrades too slowly and the cells multiply too rapidly, exuding too much extracellular matrix too soon, then the tissue will be "locked in," unable to respond to environmental signals.

At intervals, the laboratory team analyzes the tissue they have created to determine what kinds of cells proliferate, what cells are adjacent to one another, what sort of matrix molecules they put around themselves, and other factors. The researchers compare their "engineered" smooth muscle tissue to regular body tissue.

"It has become more and more clear, you can't take the observations from a few cultured cells and apply it to human conditions," Mooney says. A three-dimensional culture is a much better model system. "This will help us to design new materials that give the right of kind of signals to cells."

Mooney's laboratory is investigating how cells interact with the artificial materials, and how interactions regulate gene expression and tissue development. More than a framework for holding cells, the natural extracellular matrix transmits information to cells, which guides cell activity such as growth. Mooney's team is trying to get the artificial scaffolding to "stand in" for the extracellular matrix as these engineered tissues develop, and thus to regulate gene expression and tissue generation.

"We don't just want to observe what happens but to control the process to do specific things. We study it quantitatively. How quickly does it come together? How quickly does it degrade?" explains Mooney. "Engineering is about materials processing and synthesis, and studying in a quantitative way how the components interact."

Cell Mechanics

Tissues are constantly exposed to mechanical forces. Blood vessels undergo twisting and stretching as the body moves, and experience the constant pressure of blood passing through the vessel. Mooney suspects these forces are critical to the formation of a functional tissue.

To understand how mechanical forces may relate to tissue development, Mooney's research group is growing tissues that have various patterns of mechanical forces applied to them, as well as growing a comparison version of the three-dimensional culture under static conditions. They compare what happens at the level of the cell.

"We don't want to treat the cell like a black box. We want to know how mechanical forces are sensed outside the cell, and how that signal message goes inside the cell. We want to understand what happens inside the cell."

Mooney's research team, including graduate students Andy Putnam and Jim Cunningham, is studying changes inside the cells themselves to understand how mechanical forces regulate assembly and organization of tissues. For example, when you twist your arm, that strains the blood vessels in your arm. The strain may help signal the cells to grow. Just how that mechanical signal gets to the cell interior where growth is controlled is a matter of hypothesis.

One leading idea is that the strain is transmitted via the cytoskeleton-a spider-like system of tiny tubes that radiate throughout the interior of the cell that provides shape and structure. The strain is first transmitted to the extracellular matrix. The matrix is in contact with the cell walls where receptors are in contact with associated molecules on the other side of the cell membrane. This is a possible pathway for passing the mechanical force on to the cytoskeleton.

"We know the force we are applying outside the cell. We know some of the chemical pathways inside the cell that indicate growth. The cytoskeleton is a mechanical, structural pathway that physically integrates the system."

Mooney's hypothesis is that this microtubule network is a means of transmitting the mechanical forces from outside the cell into the cell nucleus where the "gene machinery" that directs the cell growth is activated.

Multiple Perspectives Add Insight

This research focus on the cytoskeleton also raised a seeming paradox on how the cell regulates the cytoskeleton formation. One of Mooney's studies provides an example of how having both biologists and engineers working on a question can lead to greater insight than either might learn alone.

Mooney applied an engineering analysis to a standard biological study of regulation of cell activities to resolve the paradox.

The molecule, tubulin, that makes up the microtubule is found in two forms in the cell: some of it is linked together like LegoŠ blocks to form the cytoskeleton. In addition, there is a reservoir of unlinked "blocks" in the cytoplasm of the cell.

A biological study of tubulin synthesis in the cell revealed that the cell maintained a constant level of tubulin protein. When the cell thinks it needs more protein, the gene machinery goes into action, and the DNA blueprint for tubulin is transcribed into messenger RNA, which serves as a template for protein formation. When biologists added more tubulin protein to the cell, there was a reduction in the messenger RNA. From this, the scientists concluded that the cell aimed to keep the supply of protein constant, and made more only when the supply fell.

Another analysis of the cell's activities, however, came up with a contradictory result. The microtubules and the pool of "free blocks" are in an equilibrium. When energy is added to the system through mechanical stress, the tubulin in the microtubules is in a high energy state (since the forces can only transmit through the microtubule structure) while the individual tubulin molecules exist in a state of low energy.

If force is applied to the microtubules, which adds energy to the system, the tubulin system needs to establish a new equilibrium. The microstructure would have to transfer energy to the "pool" by depolymerizing-coming apart. This would boost the number of free "blocks" in the pool.

Yet having a larger pool of blocks violates the automatic regulation by the gene machinery, which keeps the amount of protein constant. Also, it implies that the microtubules would come apart just when the cell needed more structure.

About two years ago, Mooney published the results of a study that explained the contradiction. He made a mass-balance study of the cell system, which is a standard chemical engineering analysis, he explains. His research showed that the rate of degradation of the protein in the reservoir made the difference. The cell is constantly making and remaking the blocks. When mechanical stress was present, the cell slowed the rate of turnover. Thus, an individual block "hung around" longer in the cell, rather than being degraded. That provided the necessary pool of free blocks to prevent the microtubules from coming apart as would have been necessary to preserve the energy balance. At the same time it was consistent with the gene autoregulation system. "There were actually two points of regulation in the cell," Mooney explains. His result added more understanding of the regulation of the cytoskeleton than the biological analysis alone had provided.

In these studies, cells were directly manipulated to induce changes inside the cell. Mooney is now conducting these analyses using a "more natural" context, subjecting cells to mechanical stress that the cell would experience through its connections to the extracellular matrix.

The Right Stuff

The extracellular matrix seems to play an important role in keeping a tissue functioning as it should. In a study using liver cells, Mooney is investigating how the matrix is involved in regulating the jobs that tissues do.

Efforts are underway at various institutions to grow replacement livers or to place liver cells in other tissues where they might take on normal liver functions. These are attempts to provide options for the many people who suffer liver failure each year. Aside from transplant, there is little available to compensate for liver failure. In the these engineered approaches, researchers have been able to keep the cells alive and get some proliferation, but cells soon quit working like liver cells.

Mooney believes they quit functioning because the cells require certain hormones and growth factors as well as contact with extracellular matrix. His theory is that the necessary factors to keeping cells alive such as insulin may be available throughout the body, but the contact with the extracellular matrix provides the specific local regulation that keeps the liver cells doing their liver-cell work.

Mooney and graduate student Jon Rowley are testing precisely which things are needed to keep the liver cells functioning, and they are developing a device to deliver these factors to liver cells grown in replacement organs or transplanted to other places. The researchers are creating microbeads loaded with the necessary factors. The beads could be infused among the liver cells in the artificial organs where they would gradually dissolve, releasing the chemicals over time to "bathe" the liver cells with the nutrients and hormones needed to maintain liver-cell functions.

Mooney has also studied in molecular biology laboratories to learn about genetic and physiologic controls on cell growth and tissue organization.

Still, the quantity of detailed information needed to engineer a tissue seems daunting. "No one person can have all the expertise needed," Mooney says, and he is busy establishing collaborations with colleagues in the dental school and elsewhere at the university.

In a collaboration with University of North Carolina researchers, he has used the template scaffoldings for growing tissue for breast implants. Women who have undergone mastectomies may someday "grow their own" implants rather than rely on artificial ones. A company called Reprogenesis is now conducting animal trials of that technology. The breast implants may be available to patients within a decade.

The potential for treating body breakdowns with engineered parts is a melding of engineering and biology that is possible because of the molecular biology developed over the last 10 to 20 years, Mooney explains. "We are building on basic advances in biology, and applying that to specific problems. We take this information from cellular and molecular biology and actually use it to make tissues."