Tissue engineering is a hot field. Electrospinning is a promising technique for making a three dimensional scaffold for growing replacement tissue.
RICHMOND, Va. – Traditional heart bypass surgeries require using veins from the leg to replace damaged blood vessels. Using a nanotechnology developed by Virginia Commonwealth University researchers, doctors soon could be using artificial blood vessels grown in a laboratory to help save half a million lives every year.
The new technology produces a natural human blood vessel grown around a scaffold, or tube, made of collagen. Using a process called electrospinning, VCU scientists are making tubes as small as one millimeter in diameter. That’s more than four times smaller than the width of a drinking straw and six times smaller than the smallest commercially available vascular graft.
VCU Biomedical Engineer Gary L. Bowlin, Ph.D., said patients don’t always have enough spare veins for a heart bypass, and even when they do, complications and failures often result because they are not compatible. “So what’s really needed is a blood vessel you can pull off the shelf,” said Bowlin.
After the scaffold is spun, smooth muscle cells are “seeded” or placed on its surface in a laboratory. The cells grow and within three-to-six weeks the tissue-engineered blood vessel is ready to implant.
Unlike current synthetic plastic blood vessels, collagen is a natural component of the body, allowing cells to grow on its surface and avoid rejection. “The cells are in a happy environment and they’re just going to stay and think ‘I’m a blood vessel, I’m going to act like a blood vessel,’” said Bowlin.
The collagen scaffold is biodegradable and eventually is replaced by the body. Pre-made blood vessels could be made available to emergency rooms where every second counts. Other applications include pediatric surgery where implanted blood vessels must grow with the patient and diabetic patients who often lose blood vessels to vascular disease.
The same collagen electrospinning technology can also be used to regenerate or replace skin, bone, nerves, muscles and even repair spinal cord injuries, according to co-inventor Gary E. Wnek, Ph.D., a VCU chemical engineer. “Anything you want to repair can start from a scaffold. We’re very excited about the potential,” said Wnek.
Practical applications of the new technology could be commercially available within three years.
Through VCU, the researchers formed a company called NanoMatrix to produce and test their products. Within two to three years, NanoMatrix expects to have products on the market, Bowlin said.
His co-inventors are Gary E. Wnek, a chemical engineer interested in nerve repair, and David Simpson, an associate professor of anatomy and neurobiology, who is looking at hearts and skeletal muscles.
``We're trying to make corneas, cartilage, skin, bones, tendons,'' Bowlin said. ``The Holy Grail is to make a whole liver, a whole heart, but we have to take baby steps.''
NanoMatrix and VCU are pursuing US government funding thru the National Institute of Standards and Techology Advanced Technology Program. The NanoMatrix grant application summary provides an idea of their direction of development.
More than 1.4 million surgical procedures that require arterial prostheses are performed each year in the United States, approximately 500,000 of these are coronary artery bypass operations. Because there are no acceptable synthetic prostheses for small-diameter blood vessels, surgeons must harvest the patient's own blood vessels for the transplant. This procedure is time-consuming, prone to complications, and greatly increases the recovery time for the patient. It also limits the number of patients who are good candidates for the surgery, because there are only a few vessels in the body potentially available for transplantation. Attempts have been made for years to develop a viable synthetic or tissue-engineered prostheses for small blood vessels, but all have had high failure rates for one reason or another. To answer this need, NanoMatrix proposes a three-year project to design and fabricate three-dimensional (3D) "scaffolds" out of collagen, the body's natural structural material, that can be seeded with various types of cells to mimic natural, small-diameter blood vessels. Studies suggest that muscle cells, once implanted in the scaffold, will develop the function, shape, morphology, and cellular architecture of the "normal" vessel. In practice, natural blood vessels are difficult to mimic -- they are composed of three distinct layers of different types of cells and attempts to artificially create the blood-vessel tube have been frustrating. NanoMatrix's innovation is a novel "electrospinning" technology to produce nanofibers from collagen and other biological proteins, together with a special bioreactor to culture the implanted cells on this scaffold of collagen. Electrospinning has been used in the past to produce very fine fibers of polymers -- and even collagen -- but lacking precise, controlled orientation of the fibers. NanoMatrix will design and build an electrospinning device that incorporates computerized, multi-axis controls to build collagen scaffolds with the proper layering and orientation to mimic blood vessels. A novel cell culture bioreactor will maintain the constructs and prevent necrosis as the cells grow. Human endothelial cells, smooth muscle cells, and fibroblasts will be used in the inner, middle, and outer layers, respectively, of the vascular constructs. A key challenge will be to achieve the proper alignment, architecture, abundance of cell types, and behavior in each cell layer. The company will optimize the structure, mechanical properties, and biological efficacy of the vascular grafts and then conduct implantation studies. Virginia Commonwealth University (Richmond, Va.) will be subcontracted to conduct the tests. ATP support is necessary because the long history of previous failures to develop small artificial blood vessels discourages venture capital. If successfully developed and approved for clinical use, the new technology could replace all other vascular grafts, reduce coronary bypass surgical costs by 10 percent and other hospital costs as well, and improve productivity and quality of life for people who undergo vascular graft procedures. The technology platform also would be applicable to the engineering of skin, cartilage, bone, muscle, heart muscle, neural tissue, and other tissues.
In 1934, a process was patented by Formhals [1-3], wherein an experimental setup was outlined for the production of polymer filaments using electrostatic force. When used to spin fibers this way, the process is termed as electrospinning.
In the electrospinning process a high voltage is used to create an electrically charged jet of polymer solution or melt, which dries or solidifies to leave a polymer fiber [4, 5]. One electrode is placed into the spinning solution/melt and the other attached to a collector. Electric field is subjected to the end of a capillary tube that contains the polymer fluid held by its surface tension. This induces a charge on the surface of the liquid. Mutual charge repulsion causes a force directly opposite to the surface tension . As the intensity of the electric field is increased, the hemispherical surface of the fluid at the tip of the capillary tube elongates to form a conical shape known as the Taylor cone . With increasing field, a critical value is attained when the repulsive electrostatic force overcomes the surface tension and a charged jet of fluid is ejected from the tip of the Taylor cone. The discharged polymer solution jet undergoes a whipping process  wherein the solvent evaporates, leaving behind a charged polymer fiber, which lays itself randomly on a grounded collecting metal screen. In the case of the melt the discharged jet solidifies when it travels in the air and is collected on the grounded metal screen.
The collage scaffolding is biodegradable. Now, you might be asking "Sounds great, but where will we get the natural immunocompatible human collagen from?" Silk worms! Japanese researchers have genetically engineered silk worms to make human collagen.
The team, from Hiroshima University and other institutions, constructed a DNA sequence that encodes for the production of human Type III procollagen, a mini-chain that is a kind of precursor to the full collagen molecule, which is a long-chained polymer. This DNA was combined with other genetic material and then injected into silkworm embryos.
The resulting silkworms secreted procollagen along with silk proteins in forming their cocoons. The researchers reported in Nature Biotechnology that they had found it relatively simple to separate the procollagen from the silk.
Los Angeles, Dec. 23 –– Researchers at the Keck School of Medicine of the University of Southern California, along with colleagues from across the country, have for the first time genetically engineered mouse cells to produce a type of human collagen--type VII--that is missing in a family of inherited skin diseases called dystrophic epidermolysis bullosa. They also prompted the mouse cells to create the structural fibers that normally arise from type VII collagen. Their work was published in the December issue of Nature Genetics.
"This is the first demonstration of in vivo gene therapy where the genes have made a large extracellular molecular structure that you can actually see with a microscope," says David Woodley, M.D., professor and chief of dermatology at the Keck School and the principal investigator on this study. Scientists from Shriners Hospital for Children in Portland, Oregon, Northwestern University in Chicago, and Xgene Corporation in San Carlos, California, also participated in the study.
Woodley was helped by his previous efforts in the field: In 1992, he and some of his colleagues became the first team to clone the human gene for type VII collagen, which is one of the key components of the skin's extracellular matrix. Collagen makes up the tendrils and fibrils that provide a cushion for the skin's cells to rest upon; type VII collagen, in particular, is critical to the creation of the skin's so-called anchoring fibrils.
The goal of the USC researchers is to treat some human inherited skin diseases. They are studying the human type VII collagen gene in the mouse in preparation for the development of a gene therapy to treat the sufferers of these diseases. The mouse may not turn out to be a useful organism for the production of human collagen. Still, its an important result.
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