Mechanical properties of human patellar tendon at the hierarchical levels of tendon and fibril

Tendons are strong hierarchical structures, but how tensile forces are transmitted between different levels remains incompletely understood. Collagen fibrils are thought to be primary determinants of whole tendon properties, and therefore we hypothesized that the whole human patellar tendon and its distinct collagen fibrils would display similar mechanical properties. Human patellar tendons (n = 5) were mechanically tested in vivo by ultrasonography. Biopsies were obtained from each tendon, and individual collagen fibrils were dissected and tested mechanically by atomic force microscopy. The Young's modulus was 2.0 ± 0.5 GPa, and the toe region reached 3.3 ± 1.9% strain in whole patellar tendons. Based on dry cross-sectional area, the Young's modulus of isolated collagen fibrils was 2.8 ± 0.3 GPa, and the toe region reached 0.86 ± 0.08% strain. The measured fibril modulus was insufficient to account for the modulus of the tendon in vivo when fibril content in the tendon was accounted for. Thus, our original hypothesis was not supported, although the in vitro fibril modulus corresponded well with reported in vitro tendon values. This correspondence together with the fibril modulus not being greater than that of tendon supports that fibrillar rather than interfibrillar properties govern the subfailure tendon response, making the fibrillar level a meaningful target of intervention. The lower modulus found in vitro suggests a possible adverse effect of removing the tissue from its natural environment. In addition to the primary work comparing the two hierarchical levels, we also verified the existence of viscoelastic behavior in isolated human collagen fibrils.     

Affine versus non-affine fibril kinematics in collagen networks: theoretical studies of network behavior

The microstructure of tissues and tissue equivalents (TEs) plays a critical role in determining the mechanical properties thereof. One of the key challenges in constitutive modeling of TEs is incorporating the kinematics at both the macroscopic and the microscopic scale. Models of fibrous microstructure commonly assume fibrils to move homogeneously, that is affine with the macroscopic deformation. While intuitive for situations of fibril-matrix load transfer, the relevance of the affine assumption is less clear when primary load transfer is from fibril to fibril. The microstructure of TEs is a hydrated network of collagen fibrils, making its microstructural kinematics an open question. Numerical simulation of uniaxial extensile behavior in planar TE networks was performed with fibril kinematics dictated by the network model and by the affine model. The average fibril orientation evolved similarly with strain for both models. The individual fibril kinematics, however, were markedly different. There was no correlation between fibril strain and orientation in the network model, and fibril strains were contained by extensive reorientation. As a result, the macroscopic stress given by the network model was roughly threefold lower than the affine model. Also, the network model showed a toe region, where fibril reorientation precluded the development of significant fibril strain. We conclude that network fibril kinematics are not governed by affine principles, an important consideration in the understanding of tissue and TE mechanics, especially when load bearing is primarily by an interconnected fibril network.     

The molecular and fibrillar structure of collagen and its relationship to the mechanical properties of connective tissue

The conformation of type I collagen molecules has been refined using a linked-atom least-squares procedure in conjunction with high-quality X-ray diffraction data. In many tendons these molecules pack in crystalline arrays and a careful measurement of the positions of the Bragg reflections allows the unit cell to be determined with high precision. From a further analysis of the X-ray data it can be shown that the highly ordered overlap region of the collagen fibrils consists of a crystalline array of molecular segments inclined by a small angle with respect to the fibril axis. In contrast, the gap region is less well ordered and contains molecular segments that are likely to be inclined by a similar angle but in a different vertical plane to that found in the overlap region. The collagen molecule thus has a D-periodic crimp in addition to the macroscopic crimp observed visually in the collagen fibres of many connective tissues. The growth and development of collagen fibrils have been studied by electron microscopy for a diverse range of connective tissues and the general pattern of fibril growth has been established as a function of age. In particular, relationships between fibril size distribution, the content and composition of the glycosaminoglycans in the matrix and the mechanical role played by the fibrils in the tissue have been formulated and these now seem capable of explaining many new facets of connective tissue structure and function.     

A computational remodeling approach to predict the physiological architecture of the collagen fibril network in corneo-scleral shells

Organized collagen fibrils form complex networks that introduce strong anisotropic and highly nonlinear attributes into the constitutive response of human eye tissues. Physiological adaptation of the collagen network and the mechanical condition within biological tissues are complex and mutually dependent phenomena. In this contribution, a computational model is presented to investigate the interaction between the collagen fibril architecture and mechanical loading conditions in the corneo-scleral shell. The biomechanical properties of eye tissues are derived from the single crimped fibril at the micro-scale via the collagen network of distributed fibrils at the meso-scale to the incompressible and anisotropic soft tissue at the macro-scale. Biomechanically induced remodeling of the collagen network is captured on the meso-scale by allowing for a continuous re-orientation of preferred fibril orientations and a continuous adaptation of the fibril dispersion. The presented approach is applied to a numerical human eye model considering the cornea and sclera. The predicted fibril morphology correlates well with experimental observations from X-ray scattering data.     

Collagen fibril morphology and organization: implications for force transmission in ligament and tendon.

Connective tissue mechanical behavior is primarily determined by the composition and organization of collagen. In ligaments and tendons, type I collagen is the principal structural element of the extracellular matrix, which acts to transmit force between bones or bone and muscle, respectively. Therefore, characterization of collagen fibril morphology and organization in fetal and skeletally mature animals is essential to understanding how tissues develop and obtain their mechanical attributes. In this study, tendons and ligaments from fetal rat, bovine, and feline, and mature rat were examined with scanning electron microscopy. At early fetal developmental stages, collagen fibrils show fibril overlap and interweaving, apparent fibril ends, and numerous bifurcating/fusing fibrils. Late in fetal development, collagen fibril ends are still present and fibril bundles (fibers) are clearly visible. Examination of collagen fibrils from skeletally mature tissues, reveals highly organized regions but still include fibril interweaving, and regions that are more randomly organized. Fibril bifurcations/fusions are still present in mature tissues but are less numerous than in fetal tissue. To address the continuity of fibrils in mature tissues, fibrils were examined in individual micrographs and consecutive overlaid micrographs. Extensive microscopic analysis of mature tendons and ligaments detected no fibril ends. These data strongly suggest that fibrils in mature ligament and tendon are either continuous or functionally continuous. Based upon this information and published data, we conclude that force within these tissues is directly transferred through collagen fibrils and not through an interfibrillar coupling, such as a proteoglycan bridge.     

Effect of various nerve decompression procedures on the functions of distal limbs in streptozotocin-induced diabetic rats: further optimism in diabetic neuropathy

It is known that diabetic neuropathy is the result of endoneurial edema caused by various biochemical reactions triggered by hyperglycemia. This sequence of events can cause cessation of circulation at the perineurial level, or the tough layer, which is not resilient enough to spread intraneural pressure. Internal and external limiting structures create a double crush phenomenon to the nerve structure. Decompression of the nerve trunk at separate levels is one of the adjuncts to the overall treatment plan for diabetic neuropathy. In this study, the right sciatic nerves of 30 rats with streptozotocin-induced diabetes were used; three groups were created. In the control group, the sciatic nerves were explored and dissected only. In group II, tarsal tunnel release was performed and accompanied by epineurotomy of the sciatic nerve and its peroneal and tibial extensions. In group III, in addition to the procedures performed in group II, perineural sheaths, exposed through the epineurotomy sites at both the peroneal and tibial nerves, were incised for decompression of the fascicles. Improvement in diabetic neuropathy was evaluated by using footprint parameters. The last print length values, estimated according to the 38-month measurements, were 26.1 +/- 0.12 mm in the control group, 23.2 +/- 0.07 mm in group II, and 22.2 +/- 0.1 mm in group III. The toe spread and intermediate toe spread values of the groups were parallel to improvements in print lengths throughout the study. The best improvement was observed in the perineurotomy group. Finally, an electron microscopic study revealed variable degenerative changes in all groups, but they were milder in groups II and III. This experimental study reveals that adding internal decompression to external release doubled the effect in reducing derangement in the sciatic nerves of the rats and, in the authors' opinion, offers cause for further optimism in the treatment of diabetic neuropathy.     

Tensile mechanical properties of three-dimensional type I collagen extracellular matrices with varied microstructure

The importance and priority of specific micro-structural and mechanical design parameters must be established to effectively engineer scaffolds (biomaterials) that mimic the extracellular matrix (ECM) environment of cells and have clinical applications as tissue substitutes. In this study, three-dimensional (3-D) matrices were prepared from type I collagen, the predominant compositional and structural component of connective tissue ECMs, and structural-mechanical relationships were studied. Polymerization conditions, including collagen concentration (0.3-3 mg/mL) and pH (6-9), were varied to obtain matrices of collagen fibrils with different microstructures. Confocal reflection microscopy was used to assess specific micro-structural features (e.g., diameter and length) and organization of component fibrils in 3-D. Microstructural analyses revealed that changes in collagen concentration affected fibril density while maintaining a relatively constant fibril diameter. On the other hand, both fibril length and diameter were affected by the pH of the polymerization reaction. Mechanically, all matrices exhibited a similar stress-strain curve with identifiable "toe," "linear," and "failure" regions. However the linear modulus and failure stress increased with collagen concentration and were correlated with an increase in fibril density. Additionally, both the linear modulus and failure stress showed an increase with pH, which was related to an increasedfibril length and a decreasedfibril diameter. The tensile mechanical properties of the collagen matrices also showed strain rate dependence. Such fundamental information regarding the 3-D microstructural-mechanical properties of the ECM and its component molecules are important to our overall understanding of cell-ECM interactions (e.g., mechanotransduction) and the development of novel strategies for tissue repair and replacement.     

Stress related collagen ultrastructure in human aortic valves--implications for tissue engineering

Understanding the response of tissue structures to mechanical stress is crucial for optimization of mechanical conditioning protocols in the field of heart valve tissue engineering. In heart valve tissue, it is unclear to what extent mechanical loading affects the collagen fibril morphology. To determine if local stress affects the collagen fibril morphology, in terms of fibril diameter, its distribution, and the fibril density, this was investigated in adult native human aortic valve leaflets. Transmission electron microscopy images of collagen fibrils were analyzed at three locations: the commissures, the belly, and the fixed edge of the leaflets. Subsequently, the mechanical behavior of human aortic valves was used in a computational model to predict the stress distribution in the valve leaflet during the diastolic phase of the cardiac cycle. The local stresses at the three locations were related to the collagen fibril morphology. The fibril diameter and density varied significantly between the measured locations, and appeared inversely related. The average fibril diameter increased from the fixed edge, to the belly, and to the commissures of the leaflets, while fibril density decreased. Interestingly, these differences corresponded well with the level of stress at the locations. The presented data showed that large tissue stress is associated with greater average fibril diameter, lower fibril density, and wider fibril size distribution compared with low stress locations in the leaflets. The findings here provide insight in the effect of mechanical loading on the collagen ultrastructure, and are valuable to improve conditioning protocols for tissue engineering.     

The relation between collagen fibril kinematics and mechanical properties in the mitral valve anterior leaflet

We have recently demonstrated that the mitral valve anterior leaflet (MVAL) exhibited minimal hysteresis, no strain rate sensitivity, stress relaxation but not creep (Grashow et al., 2006, Ann Biomed Eng., 34(2), pp. 315-325; Grashow et al., 2006, Ann Biomed. Eng., 34(10), pp. 1509-1518). However, the underlying structural basis for this unique quasi-elastic mechanical behavior is presently unknown. As collagen is the major structural component of the MVAL, we investigated the relation between collagen fibril kinematics (rotation and stretch) and tissue-level mechanical properties in the MVAL under biaxial loading using small angle X-ray scattering. A novel device was developed and utilized to perform simultaneous measurements of tissue level forces and strain under a planar biaxial loading state. Collagen fibril D-period strain (epsilonD) and the fibrillar angular distribution were measured under equibiaxial tension, creep, and stress relaxation to a peak tension of 90 N/m. Results indicated that, under equibiaxial tension, collagen fibril straining did not initiate until the end of the nonlinear region of the tissue-level stress-strain curve. At higher tissue tension levels, epsilonD increased linearly with increasing tension. Changes in the angular distribution of the collagen fibrils mainly occurred in the tissue toe region. Using epsilonD, the tangent modulus of collagen fibrils was estimated to be 95.5+/-25.5 MPa, which was approximately 27 times higher than the tissue tensile tangent modulus of 3.58+/-1.83 MPa. In creep tests performed at 90 N/m equibiaxial tension for 60 min, both tissue strain and epsilonD remained constant with no observable changes over the test length. In contrast, in stress relaxation tests performed for 90 min epsilonD was found to rapidly decrease in the first 10 min followed by a slower decay rate for the remainder of the test. Using a single exponential model, the time constant for the reduction in collagen fibril strain was 8.3 min, which was smaller than the tissue-level stress relaxation time constants of 22.0 and 16.9 min in the circumferential and radial directions, respectively. Moreover, there was no change in the fibril angular distribution under both creep and stress relaxation over the test period. Our results suggest that (1) the MVAL collagen fibrils do not exhibit intrinsic viscoelastic behavior, (2) tissue relaxation results from the removal of stress from the fibrils, possibly by a slipping mechanism modulated by noncollagenous components (e.g. proteoglycans), and (3) the lack of creep but the occurrence of stress relaxation suggests a "load-locking" behavior under maintained loading conditions. These unique mechanical characteristics are likely necessary for normal valvular function.     

Relative orientation of collagen molecules within a fibril: a homology model for homo sapiens type I collagen

Type I collagen is an essential extracellular protein that plays an important structural role in tissues that require high tensile strength. However, owing to the molecule’s size, to date no experimental structural data are available for the Homo sapiens species. Therefore, there is a real need to develop a reliable homology model and a method to study the packing of the collagen molecules within the fibril. Through the use of the homology model and implementation of a novel simulation technique, we have ascertained the orientations of the collagen molecules within a fibril, which is currently below the resolution limit of experimental techniques. The longitudinal orientation of collagen molecules within a fibril has a significant effect on the mechanical and biological properties of the fibril, owing to the different amino acid side chains available at the interface between the molecules.     

Tip-mediated fusion involving unipolar collagen fibrils accounts for rapid fibril elongation, the occurrence of fibrillar branched networks in skin and the paucity of collagen fibril ends in vertebrates.

Collagen fibrils are the principal source of mechanical strength of connective tissues such as tendon, skin, cornea, cartilage and bone. The ability of these tissues to withstand tensile forces is directly attributable to the length and diameter of the fibrils, and to interactions between individual fibrils. Although electron microscopy studies have provided information on fibril diameters, little is known about the length of fibrils in tissue and how fibrils interact with each other. The question of fibril length has been difficult to address because fibril ends are rarely observed in cross-sections of tissue. The paucity of fibril ends, or tips, has led to controversy about how long individual fibrils might be and how the fibrils grow in length and diameter. This review describes recent discoveries that are relevant to these questions. We now know that vertebrate collagen fibrils are synthesised as short (1-3 microm) early fibrils that fuse end-to-end in young tissues to generate very long fibrils. The diameter of the final fibril is determined by the diameter of the collagen early fibrils. During a late stage of tissue assembly fibril tips fuse to fibril shafts to generate branched networks. Of direct relevance to fibril fusion is the fact that collagen fibrils can be unipolar or bipolar, depending on the orientation of collagen molecules in the fibril. Fusion relies on: (1) specific molecular interactions at the carboxyl terminal ends of unipolar collagen fibrils; and (2) the insulator function of small proteoglycans to shield the surfaces of fibrils from inappropriate fusion reactions. The fusion of tips to shafts to produce branched networks of collagen fibrils is an elegant mechanism to increase the mechanical strength of tissues and provides an explanation for the paucity of fibril tips in older tissue.     

Advanced Glycation End-Products Reduce Collagen Molecular Sliding to Affect Collagen Fibril Damage Mechanisms but Not Stiffness

Advanced glycation end-products (AGE) contribute to age-related connective tissue damage and functional deficit. The documented association between AGE formation on collagens and the correlated progressive stiffening of tissues has widely been presumed causative, despite the lack of mechanistic understanding. The present study investigates precisely how AGEs affect mechanical function of the collagen fibril – the supramolecular functional load-bearing unit within most tissues. We employed synchrotron small-angle X-ray scattering (SAXS) and carefully controlled mechanical testing after introducing AGEs in explants of rat-tail tendon using the metabolite methylglyoxal (MGO). Mass spectrometry and collagen fluorescence verified substantial formation of AGEs by the treatment. Associated mechanical changes of the tissue (increased stiffness and failure strength, decreased stress relaxation) were consistent with reports from the literature. SAXS analysis revealed clear changes in molecular deformation within MGO treated fibrils. Underlying the associated increase in tissue strength, we infer from the data that MGO modified collagen fibrils supported higher loads to failure by maintaining an intact quarter-staggered conformation to nearly twice the level of fibril strain in controls. This apparent increase in fibril failure resistance was characterized by reduced side-by-side sliding of collagen molecules within fibrils, reflecting lateral molecular interconnectivity by AGEs. Surprisingly, no change in maximum fibril modulus (2.5 GPa) accompanied the changes in fibril failure behavior, strongly contradicting the widespread assumption that tissue stiffening in ageing and diabetes is directly related to AGE increased fibril stiffness. We conclude that AGEs can alter physiologically relevant failure behavior of collagen fibrils, but that tissue level changes in stiffness likely occur at higher levels of tissue architecture.     

Type V collagen fibrils in mouse metanephroi

Type V collagen (Col V) molecule, a minor component of kidney connective tissues, was found in adult cornea, and has been considered as a regulatory fibril-forming collagen that emerges into type I collagen to trigger the initiation of Col I fiber assembly. Col V was also found in injured, wound healing tissues or placenta, and was considered as a dysfunctional extracellular matrix (ECM). Reconstituted Col V fibril was characterized as an ECM to detach cells in vitro, and our previous study showed that the reconstituted Col V fibril facilitated the migration of glomerular endothelial cells and induced ECM remodeling, whereas Col V molecules stabilized cells. These facts suggest that not only the structure but also the function of Col V fibril are different from Col V molecule. Recently, Col V molecule has been reported existing in various developing tissues such as bone and lung, but Col V fibril has not been reported yet. In this study, we firstly explored the existence of Col V fibril in metanephroi, and found it distributed in the immature kidney tissues whereas disappeared when the tissues reached mature. It is likely that Col V fibril may form a prototype of pericellular microenvironment and the transient existence of Col V fibril may play a role as the pioneering ECM during metanephric tissue morphogenesis.     

Structure of corneal scar tissue: an X-ray diffraction study.

Full-thickness corneal wounds (2 mm diameter) were produced in rabbits at the Schepens Eye Research Institute, Boston. These wounds were allowed to heal for periods ranging from 3 weeks to 21 months. The scar tissue was examined using low- and wide-angle x-ray diffraction from which average values were calculated for 1) the center-to-center collagen fibril spacing, 2) the fibril diameter, 3) the collagen axial periodicity D, and 4) the intermolecular spacing within the collagen fibrils. Selected samples were processed for transmission electron microscopy. The results showed that the average spacing between collagen fibrils within the healing tissue remained slightly elevated after 21 months and there was a small increase in the fibril diameter. The collagen D-periodicity was unchanged. There was a significant drop in the intermolecular spacing in the scar tissues up to 6 weeks, but thereafter the spacing returned to normal. The first-order equatorial reflection in the low-angle pattern was visible after 3 weeks and became sharper and more intense with time, suggesting that, as healing progressed, the number of nearest neighbor fibrils increased and the distribution of nearest neighbor spacings reduced. This corresponded to the fibrils becoming more ordered although, even after 21 months, normal packing was not achieved. Ultrastructural changes in collagen fibril density measured from electron micrographs were consistent with the increased order of fibril packing measured by x-ray diffraction. The results suggest that collagen molecules have a normal axial and lateral arrangement within the fibrils of scar tissue. The gradual reduction in the spread of interfibrillar spacings may be related to the progressive decrease in the light scattered from the tissue as the wound heals.     

Prediction of peak pressure from clinical and radiological measurements in patients with diabetes

Background

Various structural and functional factors of foot function have been associated with high local plantar pressures. The therapist focuses on these features which are thought to be responsible for plantar ulceration in patients with diabetes. Risk assessment of the diabetic foot would be made easier if locally elevated plantar pressure could be indicated with a minimum set of clinical measures.

Methods

Ninety three patients were evaluated through vascular, orthopaedic, neurological and radiological assessment. A pressure platform was used to quantify the barefoot peak pressure for six forefoot regions: big toe (BT) and metatarsals one (MT-1) to five (MT-5). Stepwise regression modelling was performed to determine which set of the clinical and radiological measures explained most variability in local barefoot plantar peak pressure in each of the six forefoot regions. Comprehensive models were computed with independent variables from the clinical and radiological measurements. The difference between the actual plantar pressure and the predicted value was examined through Bland-Altman analysis.

Results

Forefoot pressures were significant higher in patients with neuropathy, compared to patients without neuropathy for the whole forefoot, the MT-1 region and the MT-5 region (respectively 138 kPa, 173 kPa and 88 kPa higher: mean difference). The clinical models explained up to 39 percent of the variance in local peak pressures. Callus formation and toe deformity were identified as relevant clinical predictors for all forefoot regions. Regression models with radiological variables explained about 26 percent of the variance in local peak pressures. For most regions the combination of clinical and radiological variables resulted in a higher explained variance. The Bland and Altman analysis showed a major discrepancy between the predicted and the actual peak pressure values.

Conclusion

At best, clinical and radiological measurements could only explain about 34 percent of the variance in local barefoot peak pressure in this population of diabetic patients. The prediction models constructed with linear regression are not useful in clinical practice because of considerable underestimation of high plantar pressure values. Identification of elevated plantar pressure without equipment for quantification of plantar pressure is inadequate. The use of quantitative plantar pressure measurement for diabetic foot screening is therefore advocated.     

Type III collagen is a key regulator of the collagen fibrillar structure and biomechanics of articular cartilage and meniscus

Despite the fact that type III collagen is the second most abundant collagen type in the body, its contribution to the physiologic maintenance and repair of skeletal tissues remains poorly understood. This study queried the role of type III collagen in the structure and biomechanical functions of two structurally distinctive tissues in the knee joint, type II collagen-rich articular cartilage and type I collagen-dominated meniscus. Integrating outcomes from atomic force microscopy-based nanomechanical tests, collagen fibril nanostructural analysis, collagen cross-link analysis and histology, we elucidated the impact of type III collagen haplodeficiency on the morphology, nanostructure and biomechanical properties of articular cartilage and meniscus in Col3a1^+/− mice. Reduction of type III collagen leads to increased heterogeneity and mean thickness of collagen fibril diameter, as well as reduced modulus in both tissues, and these effects became more pronounced with skeletal maturation. These data suggest a crucial role of type III collagen in mediating fibril assembly and biomechanical functions of both articular cartilage and meniscus during post-natal growth. In articular cartilage, type III collagen has a marked contribution to the micromechanics of the pericellular matrix, indicating a potential role in mediating the early stage of type II collagen fibrillogenesis and chondrocyte mechanotransduction. In both tissues, reduction of type III collagen leads to decrease in tissue modulus despite the increase in collagen cross-linking. This suggests that the disruption of matrix structure due to type III collagen deficiency outweighs the stiffening of collagen fibrils by increased cross-linking, leading to a net negative impact on tissue modulus. Collectively, this study is the first to highlight the crucial structural role of type III collagen in both articular cartilage and meniscus extracellular matrices. We expect these results to expand our understanding of type III collagen across various tissue types, and to uncover critical molecular components of the microniche for regenerative strategies targeting articular cartilage and meniscus repair.     

Mechanical model for a collagen fibril pair in extracellular matrix

In this paper, we model the mechanics of a collagen pair in the connective tissue extracellular matrix that exists in abundance throughout animals, including the human body. This connective tissue comprises repeated units of two main structures, namely collagens as well as axial, parallel and regular anionic glycosaminoglycan between collagens. The collagen fibril can be modeled by Hooke’s law whereas anionic glycosaminoglycan behaves more like a rubber-band rod and as such can be better modeled by the worm-like chain model. While both computer simulations and continuum mechanics models have been investigated for the behavior of this connective tissue typically, authors either assume a simple form of the molecular potential energy or entirely ignore the microscopic structure of the connective tissue. Here, we apply basic physical methodologies and simple applied mathematical modeling techniques to describe the collagen pair quantitatively. We found that the growth of fibrils was intimately related to the maximum length of the anionic glycosaminoglycan and the relative displacement of two adjacent fibrils, which in return was closely related to the effectiveness of anionic glycosaminoglycan in transmitting forces between fibrils. These reveal the importance of the anionic glycosaminoglycan in maintaining the structural shape of the connective tissue extracellular matrix and eventually the shape modulus of human tissues. We also found that some macroscopic properties, like the maximum molecular energy and the breaking fraction of the collagen, were also related to the microscopic characteristics of the anionic glycosaminoglycan.     

Hyposensitivity of C-fiber Afferents at the Distal Extremities as an Indicator of Early Stages Diabetic Bladder Dysfunction in Type 2 Diabetic Women

Purpose

To investigate the relationship between distal symmetric peripheral neuropathy and early stages of autonomic bladder dysfunction in type 2 diabetic women.

Materials and Methods

A total of 137 diabetic women with minimal coexisting confounders of voiding dysfunction followed at a diabetes clinic were subject to the following evaluations: current perception threshold (CPT) tests on myelinated and unmyelinated nerves at the big toe for peroneal nerve and middle finger for median nerve, uroflowmetry, post-void residual urine volume, and overactive bladder (OAB) symptom score questionnaire. Patients presenting with voiding difficulty also underwent urodynamic studies and intravesical CPT tests.

Results

Based on the OAB symptom score and urodynamic studies, 19% of diabetic women had the OAB syndrome while 24.8% had unrecognized urodynamic bladder dysfunction (UBD). The OAB group had a significantly greater mean 5 Hz CPT test value at the big toe by comparison to those without OAB. When compared to diabetic women without UBD, those with UBD showed greater mean 5 Hz CPT test values at the middle finger and big toe. The diabetic women categorized as C-fiber hyposensitivity at the middle finger or big toe by using CPT test also had higher odds ratios of UBD. Among diabetic women with UBD, the 5 Hz CPT test values at the big toe and middle finger were significantly associated with intravesical 5 Hz CPT test values.

Conclusions

Using electrophysiological evidence, our study revealed that hyposensitivity of unmyelinated C fiber afferents at the distal extremities is an indicator of early stages diabetic bladder dysfunction in type 2 diabetic women. The C fiber dysfunction at the distal extremities seems concurrent with vesical C-fiber neuropathy and may be a sentinel for developing early diabetic bladder dysfunction among female patients.     

Deterministic material-based averaging theory model of collagen gel micromechanics

Mechanics of collagen gels, like that of many tissues, is governed by events occurring on a length scale much smaller than the functional scale of the material. To deal with the challenge of incorporating deterministic micromechanics into a continuous macroscopic model, we have developed an averaging-theory-based modeling framework for collagen gels. The averaging volume, which is constructed around each integration point in a macroscopic finite-element model, is assumed to experience boundary deformations homogeneous with the macroscopic deformation field, and a micromechanical problem is solved to determine the average stress at the integration point. A two-dimensional version was implemented with the microstructure modeled as a network of nonlinear springs, and 500 segments were found to be sufficient to achieve statistical homogeneity. The method was then used to simulate the experiments of Tower et al. (Ann. Biomed. Eng., 30, pp. 1221-1233) who performed uniaxial extension of prealigned collagen gels. The simulation captured many qualitative features of the experiments, including a toe region and the realignment of the fibril network during extension. Finally, the method was applied to an idealized wound model based on the characterization measurements of Bowes et al. (Wound Repair Regen., 7, pp. 179-186). The model consisted of a strongly aligned "wound" region surrounded by a less strongly aligned "healthy" region. The alignment of the fibrils in the wound region led to reduced axial strains, and the alignment of the fibrils in the healthy region, combined with the greater effective stiffness of the wound region, caused rotation of the wound region during uniaxial stretch. Although the microscopic model in this study was relatively crude, the multiscale framework is general and could be employed in conjunction with any microstructural model.     

Candidate cell and matrix interaction domains on the collagen fibril, the predominant protein of vertebrates

Type I collagen, the predominant protein of vertebrates, polymerizes with type III and V collagens and non-collagenous molecules into large cable-like fibrils, yet how the fibril interacts with cells and other binding partners remains poorly understood. To help reveal insights into the collagen structure-function relationship, a data base was assembled including hundreds of type I collagen ligand binding sites and mutations on a two-dimensional model of the fibril. Visual examination of the distribution of functional sites, and statistical analysis of mutation distributions on the fibril suggest it is organized into two domains. The "cell interaction domain" is proposed to regulate dynamic aspects of collagen biology, including integrin-mediated cell interactions and fibril remodeling. The "matrix interaction domain" may assume a structural role, mediating collagen cross-linking, proteoglycan interactions, and tissue mineralization. Molecular modeling was used to superimpose the positions of functional sites and mutations from the two-dimensional fibril map onto a three-dimensional x-ray diffraction structure of the collagen microfibril in situ, indicating the existence of domains in the native fibril. Sequence searches revealed that major fibril domain elements are conserved in type I collagens through evolution and in the type II/XI collagen fibril predominant in cartilage. Moreover, the fibril domain model provides potential insights into the genotype-phenotype relationship for several classes of human connective tissue diseases, mechanisms of integrin clustering by fibrils, the polarity of fibril assembly, heterotypic fibril function, and connective tissue pathology in diabetes and aging.