Nanomedicine, Volume I: Basic Capabilities

© 1999 Robert A. Freitas Jr. All Rights Reserved.

Robert A. Freitas Jr., Nanomedicine, Volume I: Basic Capabilities, Landes Bioscience, Georgetown, TX, 1999 ECM Brachiation

Taking the standard reference male human body volume of 60,000 cm3 and subtracting ~16,000 cm3 (27%) for fluids, digestive contents, and expandables (Section 8.2.5), then subtracting another ~27,000 cm3 (45%) for the volume of all tissue cells (Section 8.5.1), leaves ~17,000 cm3 (28%) which comprises the intercellular tissue volume. This volume is thoroughly penetrated by the extracellular matrix or ECM, a fibrous scaffolding that helps organize cells into tissues. The ECM contains both protein and carbohydrate components. (Matrical water is not in the form of a dilute aqueous solution but is strongly influenced by the macromolecules that are present; diffusion of small molecules is much slower in matrix water than in bulk water.)

Nanorobots with at least two appendages can alternately grasp and release a succession of adjacent ECM elements, brachiating1621 "hand over hand" through tissue in a manner crudely analogous to a scuba diver pulling himself along an underwater rope mesh ladder. Similarly, fibroblasts migrating through tissue toward the site of a wound move toward a chemotactic distress gradient by extending lamellipodia toward the stimulus while their opposite poles remain firmly bound until released (haptotaxis). ECM fibrils strongly influence the direction of migration since cultured fibroblasts tend to align and migrate along discontinuities in substrata to which they are attached (contact guidance), and only along, but not across, fibronectin fibrils.1537

ECMs are composed of different collagen types, elastin, large glycoproteins (e.g., fibronectin, laminin, entactin, osteopontin), and proteoglycans that contain large glycosaminoglycan side chains (e.g., heparan sulfate, chondroitin sulfate, dermatan sulfate, keratan sulfate, hyaluronic acid).938,985,1511 While all ECMs share these components, the organization, form, and mechanical properties of ECMs vary widely in different tissues. For example, interstitial collagens (e.g., Types I & III) self-assemble into a three-dimensional lattice which binds fibronectin and proteoglycans. This type of ECM hydrogel forms the backbone of loose connective tissues such as dermis.985 In contrast, basement tissue membrane collagens (Types IV & V) assemble into planar arrays. When these collagenous sheets interact with fibronectin, laminin, and heparan sulfate proteoglycan, the result is a planar ECM scaffolding. The ability of tendons to resist tension, and of cartilage and bone to resist compression, similarly results from local differences in the composition and organization of the ECM.985

Tissues are dynamic structures that exhibit continual turnover of all molecular and cellular components. The ECM helps to maintain tissue pattern integrity, allowing cells that are lost due to injury or aging to be replaced in an organized fashion. For instance, when tissue cells are killed by freezing or poisoning, all of the cellular components may die and be removed, but the basement tissue membrane often remains intact. These residual scaffoldings ensure correct repositioning of cells (e.g., cell polarity) and proper restoration of different cell types to their correct locations (e.g., muscle cells in muscle basement tissue membrane, nerve cells in nerve sheaths, endothelium within vessels). Conversely, loss of ECM integrity during wound healing may cause permanent disorganization of tissue patterns, producing dermal scars.985 The ECM also plays a role in controlling cell function and morphology. Hepatocytes cultured upon fibronectin or laminin at low fiber surface density (1-50 ng/cm2) exhibit differentiation, but switch to growth at high fiber surface density (1000 ng/cm2).1501 Capillary-like tube formation by endothelial cells can be induced by purely mechanical means (simulating mechanical stresses in the natural ECM).1502 Cells cultured on substrates containing adhesive islands (mimicking spatial patterns of ECM adhesivity) change shape to match the shape of the islands.718

The mesh size of the fibrous components of the ECM varies widely according to tissue type, but a few generalizations and specific examples can be given. For instance, the matrical function of collagen and elastin is mostly structural, while the function of fibronectin and laminin is primarily adhesive. Yet collagens are also adhesive for cells and other macromolecules, and fibronectin provides the major matrical support in clot and early granulation tissue formation (the first stages of wound repair).985

Collagens are the major proteins of the ECM and comprise ~25% of total mammalian protein mass.396 The collagen family currently contains 19 members.971,1497,1498,3614 The collagen molecule is a long (~300 nm), thin (~1.5 nm) fiber, the center of which is a characteristic triple helix running ~95% of the length.938 Micrographs of endothelial cells cultured on a disorganized collagen-rich gel show that the cells quickly remodel the substrate into a network of collagen pads, cords, cables and bridges with grid sizes ranging from 100-1000 microns.1499 Photomicrographs of excised tissue samples show a gap of 4-12 microns between adjacent collagen fibrils in skin tissue and in tendons.938 Cartilage (which contains much collagen) has large aggregates of proteoglycans, with as many as 100 molecules attached to a single hyaluronic acid, giving a length of ~10 microns and a diameter of 500-600 nm; a tissue micrograph of fibrous cartilage in the disks between the vertebral bodies shows gaps of 10-30 microns between adjacent fibrils.938

Fibronectin is usually formed as a disulfide-linked dimer consisting of two 220-250 kilodalton peptide subunits, each containing binding sites for collagen, for heparin, for the clotting protein fibrin, and for several other ECM molecules, along with cell surface receptors.938 The main function of laminin (a 50 nm x 70 nm cross-shaped molecule938) is to mediate the binding of cells to Type IV collagen. Fibronectins or laminins may be present in ECM at low density (<1 ng/mm3) or high density (>5 ng/mm3),985 giving, from Eqn. 8.7, an estimated typical intermolecular spacing of Lgrid ~ 10 microns at low density and Lgrid ~ 4 microns at high density.

Although the matrix grid can be as narrow as 0.3-3 microns in special cases such as the ECM between the epidermis and the somites of the axolotl embryo, or between the highly flexible elastin fibers in the walls of the aorta,938 in most cases the gaps between ECM elements should be wide enough for micron-size nanorobots to slip through without much difficulty. Even quite large rigid objects should be able to negotiate the ECM. Experimentally, teflon particles up to 80 microns in diameter were injected extravascularly and migrated to distant locations around the body including lymph nodes, lungs, and kidneys.946

From Eqn. 9.75, an isolated cylindrical ECM element ~1 micron long and ~0.1 micron wide could be shoved aside at a lateral velocity of ~1 mm/sec in plasma with a force of ~5 pN, easily within the strength limits of medical nanorobots having the appropriate manipulatory and motive appendages. Chemosensor pads also will allow the nanorobot to identify fiber type and tissue type (having unique chemical signatures) as the device crawls or brachiates through the ECM forest. For example, only two kinds of carbohydrate are found attached to the hydroxylysines of collagen. Elastin lacks the amino acid methionine. The family of integrins binds fibronectin, and fibronectin has lots of binding sites. Proteoglycans typically have a long core protein to which are attached up to 100 very large glycosaminoglycan (GAG) chains, with each population of core proteins having many different carbohydrates, varying in size, charge, and even composition.938 Sensor pads can make a cell-type or tissue-type determination in as little as 2 millisec (Section, suggesting an upper limit on nanorobot ECM brachiation of nbrach ~ 500 Hz if chemopositional validation is required at every step.

What is the maximum safe nanorobot brachiation speed through the ECM? In theory, a dibrachial armswing of ~1 micron repeated at ~100 Hz allows an ECM transit speed up to 100 microns/sec through clear fluid lanes in the matrix. From Eqn. 9.73, the force required to pull a 1 micron spherical object through 310 K interstitial fluid (Table 9.4) at 100 microns/sec is ~2 pN, somewhat less than the ~10 pN likely to stimulate a mechanically-transduced cellular response (Sections and 15.5.4). (One study found that the force required to break individual integrin bonds to fibrinogen is ~2.1 pN;1508 another study found ~1000 cell adhesion proteoglycan molecules on each marine sponge cell surface, with pairs on adjacent surfaces having an experimentally measured adhesion of 40-400 pN per pair, depending upon Ca++ concentration.1248) Assuming a pair of 1-micron long, 0.1-micron thick cylindrical nanorobot arms and applying Eqn. 9.75, an additional ~0.5 pN is required to move each arm forward, giving a total power requirement of only ~0.0003 pW for brachiation at ~100 microns/sec. For comparison, a proposal to drive a millimeter-size "seed" through human brain parenchyma using external magnetic forces, while maintaining what was believed to be an adequate margin of safety, was tested experimentally at a net tissue transit speed of ~8 microns/sec.1256,1257

Natural movement of biological entities through ECM is much slower. The ECM provides a handhold-rich scaffold for fibroblast migration.359 Leukocyte and fibroblast amoeboid motion through extracellular tissue is typically ~0.050-0.7 microns/sec (Section, although white cell diapedesis through the blood vessel walls (~0.005 microns/sec; Section is much slower. Chain migration of neuronal precursor cells across a restricted pathway in the brain, through a complex parenchyma between two regions separated by several millimeters, occurs at ~0.008 microns/sec.947 The migratory speed of cultured smooth muscle cells taken from coronary and peripheral arterial walls is also only ~0.006-0.015 microns/sec.1503 Epithelial cells migrate over a wound surface in a sheet at ~0.035 microns/sec, while wound contraction proceeds at 0.007-0.009 microns/sec.359 Osteoclasts tunnel through old bone, remodeling it, advancing at a rate of only ~0.0006 microns/sec.531 However, inside cytomatrix-rich cells, membrane-bound vesicles undergo axonal transport at a typical speed ~2-4 microns/sec,938,939 pigment granules in chromatophores move at 2-10 microns/sec,938 and mitochondria (~micron-sized organelles) are shuttled around the cellular interior at up to ~10 microns/sec,453 so faster ECM transit speeds up to ~100 microns/sec are probably safe.


Last updated on 16 April 2004