Mathematical modelling of anisotropy in fibrous connective tissue

We present two modelling frameworks for studying dynamic anistropy in connective tissue, motivated by the problem of fibre alignment in wound healing. The first model is a system of partial differential equations operating on a macroscopic scale. We show that a model consisting of a single extracellular matrix material aligned by fibroblasts via flux and stress exhibits behaviour that is incompatible with experimental observations. We extend the model to two matrix types and show that the results of this extended model are robust and consistent with experiment. The second model represents cells as discrete objects in a continuum of ECM. We show that this model predicts patterns of alignment on macroscopic length scales that are lost in a continuum model of the cell population.     

Physical behavior of gelatin gels: a simple model for connective tissue

The swelling behavior of gelatin gels when immersed in buffer or in osmotically active solutions of dextran 500 has been studied, and the results can be described by means of the Flory-Rehner theory of the thermodynamic properties of three-dimensional network structures. The density of crosslinkages formed in the gel is found to be related to the concentration of gelatin at which gelation occurs. Values of rigidity moduli are estimated from the density of crosslinkages by use of the statistical thermodynamic theory of rubber elasticity. These values are in excellent agreement with direct measurements.     

Relationship between connective tissue cells and fibronectin in a sequential model of experimental hepatic fibrosis

The cellular and non-cellular components of fibrous septa formed at early and late stages in a sequential model of experimental hepatic fibrosis have been investigated using ultrastructural and immunocytochemical techniques. In the early septa, cells with intermediate features between lobular Ito cells and active fibroblasts were formed. These cells frequently displayed subplasmalemmal microfilaments (myofibroblast-like cells). Macrophages were also present. Scanty typical fibroblasts were present in the late septa. This cellular recruitment might be related to an extracellular glycoprotein-fibronectin-which is at present under investigation as a chemotactic factor for fibroblasts. Strong positivity for fibronectin in early septa and its sharp decrease in late septa seems to support this view. Fibroblasts and/or macrophages are the likely source of fibronectin synthesis.     

Application of a new probabilistic model for recognizing complex patterns in glycans

MOTIVATION: The study of carbohydrate sugar chains, or glycans, has been one of slow progress mainly due to the difficulty in establishing standard methods for analyzing their structures and biosynthesis. Glycans are generally tree structures that are more complex than linear DNA or protein sequences, and evidence shows that patterns in glycans may be present that spread across siblings and into further regions that are not limited by the edges in the actual tree structure itself. Current models were not able to capture such patterns. RESULTS: We have applied a new probabilistic model, called probabilistic sibling-dependent tree Markov model (PSTMM), which is able to inherently capture such complex patterns of glycans. Not only is the ability to capture such patterns important in itself, but this also implies that PSTMM is capable of performing multiple tree structure alignments efficiently. We prove through experimentation on actual glycan data that this new model is extremely useful for gaining insight into the hidden, complex patterns of glycans, which are so crucial for the development and functioning of higher level organisms. Furthermore, we also show that this model can be additionally utilized as an innovative approach to multiple tree alignment, which has not been applied to glycan chains before. This extension on the usage of PSTMM may be a major step forward for not only the structural analysis of glycans, but it may consequently prove useful for discovering clues into their function.     

Collagen-polysaccharide gel as a model of intercellular substance of the connective tissue]

Gel samples forming at 37 degrees C in the solutions containing tropocollagen and various polysaccharides were examined by electron microscopy. Contracting gel clots formed in the solutions containing chondroitin sulfate, proteoglycine from the tracheal cartilage, gum arabic. Electron microscopy showed such clots to be permeated with collagen fibrillae with transverse striations and a period of 640 A. An association between the density of the forming gel and the nature of the polysaccharide component is discussed. Gel forming in the solutions containing tropocollagen and various polysaccharides is regarded as a model of the connective tissue intercellular substance.     

Homologous peptide of connective tissue growth factor ameliorates epithelial to mesenchymal transition of tubular epithelial cells

The hallmark of failing renal transplants is tubular atrophy and interstitial fibrosis. The cytokine connective tissue growth factor (CTGF or CCN2) plays an important role in epithelial-mesenchymal transition (EMT) of tubular epithelial cells (TECs). A unique domain within CTGF (IRTPKISKPIKFELSG) which binds to its potential receptor integrin alpha v beta3 has been identified. This study was carried out to further characterize a synthetic hexadeca-peptide (P2) homologous to this domain and to determine its effect on CTGF-mediated solid phase cell adhesion, EMT induction and fibrogenesis in rat renal NRK-52E cells. Results showed that both P2 and recombinant CTGF bound to NRK-52E cells. Unlike CTGF, P2 had little effect on EMT induction including cytoskeleton remodeling and expression of alpha-smooth muscle actin (alpha-SMA) and E-cadherin, nor did it have effect on fibrogenic induction including alternation of extracellular matrix (ECM) proteins, collagen type I and IV at gene and protein levels. All data showed that P2 bound preferably on the surface of NRK-52E cells and inhibited the effect of CTGF on EMT induction and cell fibrogenesis, probably by occupying the binding sites of CTGF within its potential receptors. Therefore, P2 may be used as a potential anti-fibrotic agent.     

Stretching of the back improves gait, mechanical sensitivity and connective tissue inflammation in a rodent model

The role played by nonspecialized connective tissues in chronic non-specific low back pain is not well understood. In a recent ultrasound study, human subjects with chronic low back pain had altered connective tissue structure compared to human subjects without low back pain, suggesting the presence of inflammation and/or fibrosis in the low back pain subjects. Mechanical input in the form of static tissue stretch has been shown in vitro and in vivo to have anti-inflammatory and anti-fibrotic effects. To better understand the pathophysiology of lumbar nonspecialized connective tissue as well as potential mechanisms underlying therapeutic effects of tissue stretch, we developed a carrageenan-induced inflammation model in the low back of a rodent. Induction of inflammation in the lumbar connective tissues resulted in altered gait, increased mechanical sensitivity of the tissues of the low back, and local macrophage infiltration. Mechanical input was then applied to this model as in vivo tissue stretch for 10 minutes twice a day for 12 days. In vivo tissue stretch mitigated the inflammation-induced changes leading to restored stride length and intrastep distance, decreased mechanical sensitivity of the back and reduced macrophage expression in the nonspecialized connective tissues of the low back. This study highlights the need for further investigation into the contribution of connective tissue to low back pain and the need for a better understanding of how interventions involving mechanical stretch could provide maximal therapeutic benefit. This tissue stretch research is relevant to body-based treatments such as yoga or massage, and to some stretch techniques used with physical therapy.     

Application of a microstructural constitutive model of the pulmonary artery to patient-specific studies: validation and effect of orthotropy

We applied a statistical mechanics based microstructural model of pulmonary artery mechanics, developed from our previous studies of rats with pulmonary arterial hypertension (PAH), to patient-specific clinical studies of children with PAH. Our previous animal studies provoked the hypothesis that increased cross-linking density of the molecular chains may be one biological remodeling mechanism by which the PA stiffens in PAH. This study appears to further confirm this hypothesis since varying molecular cross-linking density in the model allows us to simulate the changes in the P-D loops between normotensive and hypertensive conditions reasonably well. The model was combined with patient-specific three-dimensional vascular anatomy to obtain detailed information on the topography of stresses and strains within the proximal branches of the pulmonary vasculature. The effect of orthotropy on stressstrain within the main and branch PAs obtained from a patient was explored. This initial study also puts forward important questions that need to be considered before combining the microstructural model with complex patient-specific vascular geometries.     

Application of experimental data to a new model of tissue oxygen transport

A recently published equation for oxygen uptake in cells (Hills, 1970) is corrected and applied to data gathered under appropriate experimental conditions. The result is found to support the hypothesis of facilitated intracellular oxygen transport.     

Finite element model of subsynovial connective tissue deformation due to tendon excursion in the human carpal tunnel

Carpal tunnel syndrome (CTS) is a nerve entrapment disease, which has been extensively studied by the engineering and medical community. Although the direct cause is unknown, in vivo and in vitro medical research has shown that tendon excursion creates microtears in the subsynovial connective tissue (SSCT) surrounding the tendon in the carpal tunnel. One proposed mechanism for the SSCT injury is shearing, which is believed to cause fibrosis of the SSCT. Few studies have reported quantitative observations of SSCT response to mechanical loading. Our proposed model is a 2-D section that consists of an FDS tendon, interstitial SSCT and adjacent stationary tendons. We believe that developing this model will allow the most complete quantitative observations of SSCT response to mechanical loading reported thus far. Boundary conditions were applied to the FEA model to simulate single finger flexion. A velocity was applied to the FDS tendon in the model to match loading conditions of the documented cadaver wrist kinematics studies. The cadaveric and FEA displacement results were compared to investigate the magnitude of stiffness required for the SSCT section of the model. The relative motions between the model and cadavers matched more closely than the absolute displacements. Since cadaveric models do not allow identification of the SSCT layers, an FEA model will help determine the displacement and stress experienced by each SSCT layer. Thus, we believe this conceptual model is a first step in understanding how the SSCT layers are recruited during tendon excursion.     

Ultrastructural changes in the connective tissue of the gastric region during the metamorphosis of Xenopus laevis

Development of the gastric connective tissue of Xenopus laevis during metamorphosis was investigated by electron microscopy. Throughout the larval period to stage 60, the layer of connective tissue underlying the gastric epithelium consists of immature fibroblasts surrounded by a sparse extracellular matrix. At the beginning of the transition from the larval to the adult epithelial form, at about stage 60, extensive changes occur in the connective tissue. The number of cells suddenly increses and different cell types appear. Numerous contacts between epithelial and connective tissue cells are established through random gaps in the thickened basal lamina. During stages 62–63, just after the beginning of the morphogenesis of adult-type glands, the basal lamina lining the glandular epithelium becomes thinner, and the number of contacts decreases rapidly except near the tips of the glands. After the glandular cells begin to produce zymogen granules at stage 64, contacts become rare. From stage 63, when the muscularis mucosae develops, until the completion of metamorphosis, the connective tissue consists mainly of typical fibroblasts. Outside the muscularis mucosae, the fibroblasts of the lamina propria are aligned in parallel with the curvature of the glands. These observations indicate that developmental changes in the connective tissue are closely related spatiotemporally to those of the epithelial transition from larval to adult form during metamorphic climax. Although some changes are similar to those in the intestine (Ishizuya-Oka and Shimozawa, '87b), others are specific to the gastric region, which suggests that connective tissue may have a role in organ-specific differentiation of the gastric epithelium.     

Activation barriers to structural transition determine deposition rates of Alzheimer's disease a beta amyloid

Brain amyloid composed of the approximately 40-amino-acid human beta-amyloid peptide A beta is integral to Alzheimer's disease pathology. To probe the importance of a conformational transition in Abeta during amyloid growth, we synthesized and examined the solution conformation and amyloid deposition activity of A beta congeners designed to have similar solution structures but to vary substantially in their barriers to conformational transition. Although all these peptides adopt similar solution conformations, a covalently restricted Abeta congener designed to have a very high barrier to conformational rearrangement was inactive, while a peptide designed to have a reduced barrier to conformational transition displayed an enhanced deposition rate relative to wild-type A beta. The hyperactive peptide, which is linked to a heritable A beta amyloidosis characterized by massive amyloid deposition at an early age, displayed a reduced activation barrier to deposition consistent with a larger difference in activation entropy than in activation enthalpy relative to wild-type A beta. These results suggest that in Alzheimer's disease, as in the prion diseases, a conformational transition in the depositing peptide is essential for the conversion of soluble monomer to insoluble amyloid, and alterations in the activation barrier to this transition affect amyloidogenicity and directly contribute to human disease.     

Acceptance of a connective tissue equivalent for grafting and capsuloligamentary reconstruction

A strip of a connective tissue equivalent prepared by using the patient's fibroblasts cultivated in vitro and calf skin collagen, was used for capsuloligamentary reconstruction of the knee. The study of the humoral and cell-mediated immune responses indicated that there was no cell-mediated immune reaction and only a transient humoral reaction 2 months after implantation. This first assay in human surgery gave a good functional result, and an immunological response was no longer observed 5 months after grafting.