Corneal collagen fibril structure in three dimensions: Structural insights into fibril assembly, mechanical properties, and tissue organization

Abstract

The ability of the cornea to transmit light while being mechanically resilient is directly attributable to the formation of an extracellular matrix containing orthogonal sheets of collagen fibrils. The detailed structure of the fibrils and how this structure underpins the mechanical properties and organization of the cornea is understood poorly. In this study, we used automated electron tomography to study the three-dimensional organization of molecules in corneal collagen fibrils. The reconstructions show that the collagen molecules in the 36-nm diameter collagen fibrils are organized into microfibrils (Ϸ4-nm diameter) that are tilted by Ϸ15°to the fibril long axis in a right-handed helix. An unexpected finding was that the microfibrils exhibit a constant-tilt angle independent of radial position within the fibril. This feature suggests that microfibrils in concentric layers are not always parallel to each other and cannot retain the same neighbors between layers. Analysis of the lateral structure shows that the microfibrils exhibit regions of order and disorder within the 67-nm axial repeat of collagen fibrils. Furthermore, the microfibrils are ordered at three specific regions of the axial repeat of collagen fibrils that correspond to the N-and C-telopeptides and the d-band of the gap zone. The reconstructions also show macromolecules binding to the fibril surface at sites that correspond precisely to where the microfibrils are most orderly.automated electron tomography ͉ microfibril ͉ filaments T he cornea is a highly specialized tissue that combines optical transparency with mechanical resilience. These properties are directly attributable to the formation of an extracellular matrix containing narrow-diameter (Ϸ36 nm) and heterotypic collagen fibrils that are spaced and organized uniformly into sheets of parallel fibrils. Each sheet is arranged orthogonal to its neighbor and to the path of light through the cornea. We reasoned that an understanding of the morphogenesis of the corneal stroma requires detailed knowledge of the structure of individual collagen fibrils.Corneal fibrils are D-periodic (axial periodicity of collagen fibrils, where D Ϸ67 nm), uniformly narrow (Ϸ30-35 nm in diameter), and indeterminate in length, particularly in older animals. The D-periodicity of the fibril arises from side-to-side associations of triple-helical collagen molecules that are Ϸ300 nm in length (i.e., the molecular length ϭ 4.4 ϫ D) and are staggered by D. The D-stagger of collagen molecules produces alternating regions of protein density in the fibril, which explains the characteristic gap and overlap appearance of fibrils negatively contrasted for transmission electron microscopy. Previous attempts to study the high-resolution structure of collagen fibrils have used primarily x-ray fiber diffraction of rat tail tendon (e.g., ref. 1 and references therein) and lamprey notochord (2) in which the fibrils are aligned and composed mostly of one genetic type of collagen. This method has yielded information about the triple...

In vivo comparison of jellyfish and bovine collagen sponges as prototype medical devices

“…This is encouraging, as bovine collagen is well known to be of low antigenicity and tolerated well, and it is incorporated into a wide range of medical applications and devices13, 33, 34 and means that the jellyfish collagen implants are showing suitability for the same medical uses.…”

Imaging the Nonlinear Susceptibility Tensor of Collagen by Nonlinear Optical Stokes Ellipsometry

“…Previous SEM measurements of rat tail tendons have found that individual collagen fibrils lie primarily along the fiber axis, but also exhibit regions of interweaving fibrils and regions exhibiting random fibril order (39,40). A study of fibril structures in cornea using electron tomography found that microfibril substructures adopted tilt angles of 15 with respect to the fibril axis (41). Constant tilt angles between 0 and 19 for collagen helices and/or fibrils within collagen fibers have been measured for several different collagen structures using a variety of techniques (20,33,38,41).…”

“…A study of fibril structures in cornea using electron tomography found that microfibril substructures adopted tilt angles of 15 with respect to the fibril axis (41). Constant tilt angles between 0 and 19 for collagen helices and/or fibrils within collagen fibers have been measured for several different collagen structures using a variety of techniques (20,33,38,41). This discrepancy indicates that the simple d-function distribution is not sufficient to compare the average experimental local-frame tensor elements to the molecular level ab initio calculations.…”

“…2, which describes the relative local-frame tensor elements of the structures (37). From the breadth of work on analysis of collagen fiber substructure (20,33,(38)(39)(40)(41), it is apparent that a distribution of tilt angles of collagen …”

An In Situ Study of Collagen Self-Assembly Processes

“…Therefore we have to consider the definition of the acidity constant (for acetic acid in this example; however, all equations can be generalized for any weak acid and strong base) Furthermore, we have to take into account the definition of the dissociation constant of water (3) Upon addition of NaOH these equilibrium conditions are affected. We need to ensure that the number of charges is conserved ([Na + ] = [NaOH]): (4) Equations (1) - (4) have to be coupled and solved in order to determine the pH before the stoichiometric point is reached as a function of the acid and base concentrations.…”