WHAT A TANGLED WEB:
Courtesy of Philip B. Messersmith
Fibroblasts are cellular workhorses of extracellular matrix production, spinning out the majority of collagens, the most abundant proteins in the animal kingdom.
Appreciation in biology can come slowly. Researchers once deemed as junk the parts of genes not represented in proteins; likewise, neuroglia were thought to be mere bystanders to neurons. So it is with the extracellular matrix (ECM), the "scaffolding" and "glue" that fill the spaces among cells. New ways of excavating the ECM reveal that it is much more.
An eclectic collection of molecules, the ECM was once relegated to the backwater of connective tissues in histology texts. But, ECM's status is shifting. Henry E. Young, an associate professor in anatomy at the Mercer University School of Medicine in Macon, Ga., calls it "a three-dimensional 'spider web' lattice-work-like structure within a fourth-dimensional time continuum." The various guises of ECM...
NOT JUST GLUE
Over a lifetime, the ECM oversees the intricate intercellular choreographies that shape the embryo, guide wound healing, and maintain adult organs. And timing is everything. Adult tissue expressing embryonic-ECM proteins could be in a dire situation. "They tend to be re-expressed in pathological conditions, such as wound healing or cancer," says Fred Bosman, professor of pathology at the University of Lausanne in Switzerland.
DNA-microarray technology is revealing the biochemical steps behind ECM actions. "Prior to the availability of such a powerful high-throughput technology, scientists would have to work on individual genes separately, by techniques such as Northern blotting," says Shu Chien, director of the Whitaker Institute of Biomedical Engineering at the University of California, San Diego.
Microarrays bearing suites of the 2,000-plus human ECM genes can monitor sweeping gene-expression changes as well as glimpse fleeting events. Chien's team, for example, profiled human aorta cells exposed to 24 hours of shear stress, modeling blood's rush from the heart. Genes controlling division and inflammation were down-regulated; those enhancing survival, angiogenesis, and vascular remodeling were upregulated.1
ECM recipes vary: A plant has cellulose, lignin, and pectin; a fungus has chitin, cellulose, and glucans; an animal has mostly collagens. The cellular slime mold
Michael Sarras, professor of anatomy and cell biology at the University of Kansas Medical Center, studies the simplest known animal with tissue layers, the hydra, whose epithelial bilayer sandwiches a matrix of sheetlike layers surrounding a fibrillar interior. The matrix forms during development when collagens from the outermost layer communicate with laminin from the innermost layer. Integrins, specialized cell-surface receptors, mediate the signaling. "With the development of defined tissue came the presence of an ECM, with structural similarities to all subsequent animal groups," says Sarras, whose team observes that new individuals arise from pellets of cells.
Hydra uses the same ECM components as other animals, but in characteristic ways. For example, the mature forms of fibrillar collagens in hydra are similar to those in people with Ehlers-Danlos ("stretchy skin") syndrome. "In hydra, flexibility is desirable because the organism continuously contracts and extends. An abnormality in [humans] is the normal condition in hydra," Sarras says.
THE ECM TOOLBOX
In the human body, armies of fibroblasts and contractile myofibroblasts churn out 20 types of collagen, which self-assemble into varied fibrils, networks, and beads and provide tensile strength. Other proteins, such as elastin and fibrillin, impart resilience.
From the collagen frameworks extend adhesive glycoproteins, such as the cross-shaped laminins and multipronged tenascins. Proteoglycans emanating from cell surfaces sport polysaccharide chains that bind enzymes, growth factors, cytokines, and matrix proteins. Matrix metalloproteinases dismantle parts of the ECM, a process crucial in tissue remodeling.
During development, as tissues fold into organs, ECM establishes characteristic architectures. In the heart, the mostly-collagen ECM coordinates the pulsating cells, whereas the liver's scaffolding connects the parenchyma tissue to the circulation, essential for regeneration. At least four collagens, five glycoproteins, and a half-dozen proteoglycans build the liver's framework.
ECM is crucial to kidney function. In the glomeruli, the tiny tangles of capillaries that are the initial filtering sectors for each of a human kidney's million nephrons, ECM composition constantly adjusts to bloodstream contents. "In the glomerulus, [the] endothelial, mesangial, and epithelial cells enable the ECM to maintain a filtration rate of water that is higher than in any other capillary bed. Unfortunately, our knowledge of this complexity, both in normal and disease states, is primitive," says H. David Humes, professor of internal medicine at the University of Michigan Medical Center in Ann Arbor. Model organisms could help to unravel that complexity; the cellular bilayer sandwiching ECM in the kidney's subunits is highly reminiscent of hydra, says Sarras.
THE ECM–CANCER LINK
The ECM-cancer connection was sown when English surgeon Stephen Paget evoked a "seed and soil" metaphor in 1889 to explain why primary tumors spread only to certain organs. In suggesting that the secondary sites beckon the errant cells, Paget was describing the stroma, the ECM that hugs epithelia, where many cancers arise.2 "If one could modify the ECM around tumor cells to resemble the ECM of a nonpropagating body location, then this might prevent tumor progression," says Stephen Yarwood, lecturer in the division of biochemistry and molecular biology at the University of Glasgow.
Courtesy of Donald Ferry
The sandwich-like layering of cells and fibers in the stretchy hydra provides a simple model for studying the extracellular matrix.
The stroma participates in cancer early on. "Invasion of epithelial cells necessitates cross-signaling between epithelial cells and their stromal neighbors. Unraveling those factors is a major research challenge," says Olivier De Wever, a researcher in the laboratory of experimental cancerology at Ghent University Hospital in Belgium. Then, under the influence of an altered stroma, cell adhesion ebbs, cancer cells pile up into tumors, and differentiation fades. Later, the stroma entices angiogenesis and clears pathways for metastasis.
In epithelia and elsewhere, DNA microarrays are illuminating the molecular interactions behind cancer cell activities. For example, researchers from the Barrow Neurological Institute in Phoenix ran microarrays on glioma cells to look for ECM proteins that promote cell motility. The profiles fulfilled expectations for migrating brain cancer cells: diminished factors that promoted adhesion, division, and apoptosis.3
A QUESTION OF BALANCE
Excess ECM drowns organs in collagen and other matrix proteins, first causing fibrosis as fibers accumulate, then sclerosis as the material hardens. Variations on these themes arise throughout the body.
In the kidneys, diabetes mellitus causes mesangial cells to overproduce fibronectin and underproduce collagenase, gumming up the filtration apparatus and setting the stage for renal failure. Extra ECM in blood-vessel walls causes atherosclerosis; in the heart, it causes arrhythmia. Fibrosis following a heart attack prevents rupture of the affected area, but eventually it hardens the tissue, impairing cardiac function. Widespread fibrosis in the liver precedes cirrhosis.
But fibrosis is physiological too, says Shukti Chakravarti, assistant professor of medicine and ophthalmology at the Johns Hopkins Medical Institute. She works with a mouse model of inflammatory bowel disease. "As soon as the cell senses danger it makes ECM molecules. We don't know if they are laid down as ECM, but the molecules play a role in cell behavior. ECM controls cell proliferation and apoptosis, too," she says.
Too little ECM also hampers health. In Schwartz-Jampel syndrome, deficient perlecan, a proteoglycan in basement membranes, causes a mask-like face. Normally, perlecan delivers the enzyme that degrades acetylcholine in neuromuscular junctions. Without it, facial muscles contract abnormally.
Appreciation of the ECM's role in pathology will likely flower further as stem-cell biology moves forward. For example, Ichiro Nishimura's group, at the Weintraub Center for Reconstructive Biotechnology at the UCLA School of Dentistry, is tracking gene-expression changes that accompany the specialization of stem cells from liposuction aspirates into bone cells. "We hope to establish the ECM microarray as an essential testing protocol for future stem cell-based therapeutics, such as tissue engineering," says Nishimura.
And so the intercellular substance formerly known as amorphous and inert has achieved a new status as a vital part of a multicellular organism's body. Bryon Petersen, assistant professor of pathology, immunology, and laboratory medicine at the University of Florida in Gainesville, works with liver stem cells and says that the mechanisms governing the communication between cell and scaffolding are varied and conjure many unanswered questions. "It is clear that the answer to these questions entails, to a large degree, the differential response of the ECM components, and the cells that produce these proteins, to various stimuli. The ECM is more than the mortar that holds the bricks together."