Collagen Fiber Bundles Research Articles

Characterizing refractive index and thickness of biological tissues using combined multiphoton microscopy and optical coherence tomography

We present a noninvasive method for characterizing the refractive index (RI) and thickness distribution in biological tissues using a combined multiphoton microscopy (MPM) and optical coherence tomography (OCT) system. Tissue layers are distinguished by the MPM and OCT images, and the RI and thickness of each layer are determined by analyzing the co-registered MPM and OCT images. The precision of this method is evaluated on four standard samples which are water, air, immersion oil and cover glass. Precision of within ~1% error compared to reference values is obtained. Biological tissue measurement is demonstrated on fish cornea. Three layers are detected, which are identified as the epithelium and stroma I and II of the cornea. The corresponding RI of each layer is measured to be ~1.446–1.448, 1.345–1.372, and 1.392–1.436, respectively. The difference of RI in the three layers correlates with the tissue compositions including cells in epithelium, large collagen fiber bundles in stroma I, and small collagen fibers in stroma II. The combined MPM/OCT technique is shown to be able to distinguish tissue layers through biochemically specific contrasts and measure RI and thickness of tissue layers at different depths.

Changes of the Structural and Biomechanical Properties of the Bovine Pericardium after the Removal of α-Gal Epitopes by Decellularization and α-Galactosidase Treatment

BackgroundBovine pericardium is one of the most widely used materials in bioprosthetic heart valves. Immunologic responses have been implicated as potential causes of limited durability of xenogenic valves. This study aimed to determine the effectiveness of decellularization and α-galactosidase (α-gal) to remove major xenoreactive antigens from xenogenic tissues.Materials and MethodsRecombinant Bacteroides thetaiotaomicron (B. thetaiotaomicron) α-gal or decellularization, or both were used to remove α-gal from bovine pericardium. It was confirmed by α-gal-bovine serum albumin-based enzyme-linked immunosorbent assay (ELISA), high-performance anion exchange chromatography, flow cytometry, 3,3'-diaminobenzidine-staining, and lectin-based ELISA. The mechanical properties of bovine pericardium after decellularization or α-gal treatment were investigated by tests of tensile-strength, permeability, and compliance. Collagen fiber rearrangement was also evaluated by a 20,000× transmission electron microscope (TEM).ResultsRecombinant B. thetaiotaomicron α-gal could effectively remove α-gal from bovine pericardium B. thetaiotaomicron (0.1 U/mL, pH 7.2) while recombinant human α-gal removed it recombinant human α-gal (10 U/mL, pH 5.0). There was no difference in the mechanical properties of fresh and recombinant α-gal-treated bovine pericardium. Furthermore, the TEM findings demonstrated that recombinant α-gal made no difference in the arrangement of collagen fiber bundles with decellularization.ConclusionRecombinant B. thetaiotaomicron α-gal effectively removed α-gal from bovine pericardium with a small amount under physiological conditions compared to human recombinant α-gal, which may alleviate the harmful xenoreactive immunologic responses of α-gal. Recombinant α-gal treatment had no adverse effects on the mechanical properties of bovine pericardium.

Investigation of the human bridging veins structure using optical microscopy

In this paper, we investigated the brain-sinus junction and especially the bridging veins linking these two organs. Two types of optical microscopy were used: conventional optical microscopy and digital microscopy. We used thin histological sections prepared from a human brain, and stained with Masson's trichrome, hemalun and orcein. Finally we observed the path of the bridging vein inside the brain-skull interface. At smaller scales, wavy collagen fiber bundles were found and characterized inside the vein walls. Taking into account the orientations of the different sections with reference to frontal planes, we found that the bridging vein has a very complex geometry, which increases the difficulty to determine fiber orientations in its walls. Nevertheless, we found that collagen fiber bundles are mainly circumferentially oriented in the superior sagittal sinus walls. In this paper, we were able to characterize precisely the path of the bridging vein from the brain to the sinus, with different magnifications.

Mathematical modeling of collagen turnover in biological tissue

We present a theoretical and computational model for collagen turnover in soft biological tissues. Driven by alterations in the mechanical environment, collagen fiber bundles may undergo important chronic changes, characterized primarily by alterations in collagen synthesis and degradation rates. In particular, hypertension triggers an increase in tropocollagen synthesis and a decrease in collagen degradation, which lead to the well-documented overall increase in collagen content. These changes are the result of a cascade of events, initiated mainly by the endothelial and smooth muscle cells. Here, we represent these events collectively in terms of two internal variables, the concentration of growth factor TGF-β and tissue inhibitors of metalloproteinases TIMP. The upregulation of TGF-β increases the collagen density. The upregulation of TIMP also increases the collagen density through decreasing matrix metalloproteinase MMP. We establish a mathematical theory for mechanically-induced collagen turnover and introduce a computational algorithm for its robust and efficient solution. We demonstrate that our model can accurately predict the experimentally observed collagen increase in response to hypertension reported in literature. Ultimately, the model can serve as a valuable tool to predict the chronic adaptation of collagen content to restore the homeostatic equilibrium state in vessels with arbitrary micro-structure and geometry.

Clinical and pathological analysis of six cases of eruptive microvenular hemangioma

Objective To analyze the clinical and histopathologic features of eruptive microvenular hemangioma (EMVH). Methods The clinical and histological findings in six patients with EMVH were retrospectively reviewed.Results The patients were young or middle-aged,with a median age of onset of 34 years.The ratio of males to females was 1 ∶ 1.Clinically,all the patients experienced a sudden development of widespread,asymptomatic,brunneus,flat-topped papules and plaques measuring from 0.1 to 2.0 cm in diameter.Histopathologic examination revealed a poorly circumscribed proliferation of small,irregularly branched,thin-walled blood vessels haphazardly arranged between collagen fiber bundles in the dermis.The lumina of the blood vessels were often collapsed.There was no cellular atypia or mitosis.A variable degree of sclerosis was noted in the surrounding stroma.Immunobistochemical study revealed that the endothelial cells stained positive for CD34,CD31,and factor Ⅷ-related antigen.Conclusions The diagnosis of EMVH is proposed,and awareness of this disease may benefit its early diagnosis and avoidance of unnecessary treatment. Key words: Eruptive microvenular hemangioma; Skin manifestations; Pathologic processes

Spontaneous fibrosarcoma in an experimental aged Lewis rat

This case report describes the results of the gross pathological, histological and immunohistological examinations of a fibrosarcoma that spontaneously developed in the left neck region of an aged male Lewis rat that participated in a long-term facial nerve regeneration study. A 10 mm long polylactic acid (PLA) tube was implanted into a 7 mm critical defect gap of the buccal branch of the left facial nerve of a male eight-week-old Lewis rat. Forty-two weeks after implantation, an approximately 8 cm × 9 cm × 2.5 cm rapidly growing mass with ulceration of the overlying skin was found in the left neck region. Gross examination of the tumour and the surrounding tissues before tumour excision revealed that the tumour had clear boundaries, and had not invaded any facial tissues, the facial nerve and the PLA nerve guide. Gross examination of the tumour's cut surface revealed that the tumour comprised numerous smooth nodules with a homogeneous white tan colour. Examination of the haematoxylin and eosin-stained sections of the tumour revealed that the tumour was predominantly composed of bundles of spindle-shaped atypical proliferating cells that were mixed with bundles of collagen fibres and arranged in a storiform pattern. Tumour emboli and skin invasion were also observed. Immunohistological examination revealed that the tumour cells were weakly positive for vimentin, but negative for keratin, α-smooth muscle actin, S-100 protein and CD34. From the results of these analyses, the final pathological diagnosis of this tumour was a fibrosarcoma.

Contribution Towards the Anatomy of the Esophageal Hiatus and its Relationship with the Presence of Bundles of Collagen Fibers in its Margins

El objetivo fue detectar la presencia de haces de fibras de colageno en los margenes del hiato esofagico y establecer si existe relacion de estos con la anatomia del hiato. Fueron utilizados 10 cadaveres de individuos adultos de sexo masculino, sin alteraciones anatomicas evidentes causadas por trauma, cirugia o enfermedad, sobre el hiato esofagico. Fue retirada y disecada una seccion anatomica que contendia el diafragma con el hiato esofagico y los tejidos adyacentes y luego fijada en formol 10%. Las medidas del perimetro del hiato esofagico se realizaron con ayuda de un caliper digital en los lados abdominal y toracico. Para el estudio estructural cada margen fue dividido en seis secciones y tenidos con Tricomico de Masson y Picrosirius-hematoxilina. En 8/10 cadaveres estudiados fueron encontrados haces de fibras colagenas, distribuidos en 13 margenes del hiato esofagico, 7 izquierdos y 6 derechos. Las fibras musculares que originaban el pilar derecho participaron en la formacion de ambos margenes del hiato esofagico en 60% de los cadaveres, mientras que en el 40%, las fibras del pilar izquierdo habian formado el lado intermedio del margen derecho. El margen derecho fue estadisticamente mas grueso que el izquierdo. No se encontro correlacion entre las medidas de los vertices de los angulos superior/inferior y la medida transversal del hiato esofagico. Las medidas entre los vertices de los angulos superior/inferior, respectivamente, con el tendon central y el ligamento arqueado mediano, presentaron valores toracicos que fueron casi el doble em relacion a los abdominales, y fueron estadisticamente significativos. Estas distancias eran mas pequenas en los cadaveres que tenian haces de fibras de colageno en las margenes del hiato esofagico. Los paquetes de fibras de colageno fueron encontrados en 65% de los 20 margenes del hiato del esofago. Los margenes del hiato esofagico fueron formados principalmente por fibras musculares originadas del pilar derecho del diafragma. Los datos anatomicos y morfometricos presentaron valores estadisticamente significativos en relacion con: espesor del lado derecho en relacion al izquierdo; distancia entre vertice del angulo superior y el tendon central y distancia entre el vertice del angulo inferior y el ligamento arqueado mediano.

Electroacupuncture Increases the Concentration and Organization of Collagen in a Tendon Healing Model in Rats

The aim of this study was to investigate the effect of electroacupuncture (EA) on the composition and organization of the extracellular matrix of the rat Achilles tendon after a partial transection during the proliferative phase of healing. Wistar rats were divided into three groups: rats that were not tenotomized (G1), tenotomized rats (G2), and rats that were tenotomized and submitted to EA (G3). EA was applied 15 days after injury at the ST36 and BL57 acupoints for 20 min, three times per week on alternate days for a total of six sessions. Biochemical analyses were performed using non-collagenous proteins, glycosaminoglycans, and hydroxyproline quantifications. An analysis of metalloproteinase-2 was carried out by zymography. The general organization of the extracellular matrix and the metachromasy of the tendons were analyzed under light microscopy. The organization of the bundles of collagen fibers was analyzed by birefringence analysis. The results showed that EA did not alter the concentration of non-collagenous proteins or glycosaminoglycans or the enzymatic activity of metalloproteinase-2 in the transected tendons. However, the concentration of hydroxyproline was significantly increased when these tendons were treated by EA. The analysis of birefringence showed a higher organization of collagen fibers in the group treated by EA. These results indicate, for the first time, that EA may offer therapeutic benefits for the treatment of tendon injuries by increasing the concentration of collagen and by inducing a better molecular organization of the collagen fibers, which may improve the mechanical strength of the tendon after injury.

Si and Ca Individually and Combinatorially Target Enhanced MC3T3-E1 Subclone 4 Early Osteogenic Marker Expression

This study tests the hypothesis that silicon and calcium ions combinatorially target gene expression during osteoblast differentiation. MC3T3-E1 subclone 4 osteoblast progenitors (transformed mouse calvarial osteoblasts) were exposed to Si(4+) (from Na(2)SiO(3)) and Ca(2+) (from CaCl(2):H(2)O) ion treatments both individually (0.4 mM each + control treatment) and combinatorially (0.4 mM Si(4+) + 0.4 mM Ca(2+) + control treatment) and compared to control treated (α-minimum essential medium, 10% fetal bovine serum, and 1% penicillin-streptomycin) cells. Cell proliferation studies showed no significant increase in cell density between treatments over 5 days of culture. Cellular differentiation studies involved addition of ascorbic acid (50 mg/L) for all treatments. Relative gene expression was determined for collagen type 1 (Col(I)α1/Col(I)α2), core-binding factor a (cbfa1/Runx2), and osteocalcin (OCN), which indicated osteoblast progenitor differentiation into a mineralizing phenotype. Increased Si(4+) or Ca(2+) ion treatments enhanced Col(I)α1, Col(I)α2, Runx2, and OCN expression, while increased Si(4+) + Ca(2+) ion treatments enhanced OCN expression. Moreover, it was found that a Si(4+)/Ca(2+) ratio of unity was optimal for maximal expression of OCN. Collagen fiber bundles were dense, elongated, and thick within extracellular matrices (ECM) exposed to Si(4+) and Si(4+) + Ca(2+) treatments, while collagen fiber bundles were sparse, short, and thin within Ca(2+) and control treated ECM. These results indicated that individual ions enhance multiple osteogenic gene expression, while combined ion treatments enhance individual gene expression. In addition, these results indicated that Si(4+) enhanced osteoblast gene expression and ECM formation at higher levels than Ca(2+). These results support the larger concept that ions (possibly released from bioactive glasses) could control bone formation by targeting osteoblast marker expression.

Structure and function of the median finfold in larval teleosts

This paper offers a structural and mechanical analysis of the median finfold in larval teleosts. The median finfold is strengthened by bundles of collagen fibres, known as actinotrichia. We demonstrate that these structures contribute to increase the mass of backward accelerated water during swimming. The amount, dimensions, orientation and growth of actinotrichia were measured at various locations along the finfold in several developmental stages of common carp (Cyprinus carpio) and zebrafish (Danio rerio). Actinotrichia morphology, using light microscopy (e.g. diameter, orientation) and electron microscopy (which revealed their anchoring at proximal and distal ends), correlated with expected lateral forces exerted on the water during swimming. An analytical model is proposed that predicts the extent of camber from the oblique arrangement of the actinotrichia and curvature of the body. Camber of the finfold during swimming was measured from high-speed video recordings and used to evaluate the model predictions. Based on structural requirements for swimming and strain limits for collagen, the model also predicts optimal orientations of actinotrichia. Experimental data confirm the predictions of the model.

Hemangiopericytoma in a cat: a case report

In dogs and men, hemangiopericytoma is a well-recognised neoplasm, thought to originate from vascular pericytes. In cats, however, hemangiopericytoma is an extremely rare finding. The present report describes the pathological features of a 7-cm-diameter, dense, white, multilobulated tumour-like growth on the thigh of a ten-year-old Persian cat. Histologically, the mass consisted of polygonal neoplastic cells, concentrically arranged around thin-walled endothelium-lined blood vessels occasionally forming typical staghorn-configurations. In reticulin-stained sections, a dense meshwork of argyrophilic collagen fibres was evident, surrounding the central vessels and separating individual tumour cells. Within the tumour tissue, there were areas of extensive necrosis and degeneration. In the periphery, the tumour displayed a moderate infiltrative growth into the adjacent musculature. Immunohistochemical analysis revealed strong diffuse cytoplasmatic immunoreactivity for vimentin, a focal immunoreactivity for S-100, and a weak interstitial staining for laminin, whereas neoplastic cells stained negative for cytokeratin, desmin, actin, calponin, von Willebrand factor, glial fibrillary acidic protein, neuron specific enolase, CD79a, MAC387, lysozyme and MHCII. Ultrastructurally, the neoplastic cells contained few intracytoplasmatic filaments, and were surrounded by interlacing bundles of intercellular long-spacing collagen fibres. Occasionally, desmosome-like intermediate junctions were observed between neighbouring tumour cells. On the basis of histomorphology, ultrastructure and immunohistochemical reactivity, the neoplasm was diagnosed as a hemangiopericytoma, representing the second reported case of this rare tumour entity in a cat.  

Enhancing collagen stability through nanostructures containing chromium(III) oxide

Stabilization of collagen for various applications employs chemicals such as aldehydes, metal ions, polyphenols, etc. Stability against enzymatic, thermal and mechanical degradation is required for a range of biomedical applications. The premise of this research is to explore the use of nanoparticles with suitable functionalization/encapsulation to crosslink with collagen, such that the three dimensional architecture had the desired stability. Collagen solution prepared as per standard protocols is treated with chromium(III) oxide nanoparticules encapsulated within a polymeric matrix (polystyrene-block-polyacrylic acid copolymer). Selectivity towards encapsulation was ensured by the reaction in dimethyl sulfoxide, where the PS groups popped out and encapsulated the Cr2O3. Subsequently when immersed in aqueous solution, PAA units popped up to react with functional groups of collagen. The interaction with collagen was monitored through techniques such as CD, FTIR, viscosity measurements, stress analysis. CD studies and FTIR showed no degradation of collagen. Thermal stability was enhanced upon interaction of nanostructures with collagen. Self-assembly of collagen was delayed but not inhibited, indicating a compete binding of the metal oxide encapsulated polymer to collagen. Metal oxide nanoparticles encapsulated within a polymeric matrix could provide thermal and mechanical stability to collagen. The formed fibrils of collagen could serve as ideal material for various smart applications such as slow/sustained drug release. The study is also relevant to the leather industry in that the nanostructures can diffuse through the highly networked collagen fibre bundles in skin matrix easily, thus overcoming the rate limiting step of diffusion.

Ultrastructural features of the myotendinous junction of the sternomastoid muscle in Wistar rats: From newborn to aging

The myotendinous junction (MTJ) is a major area for transmitting force from the skeletal muscle system and acts in joint position and stabilization. This study aimed to use transmission electron microscopy to describe the ultrastructural features of the MTJ of the sternomastoid muscle in Wistar rats from newborn to formation during adulthood and possible changes with aging. Ultrastructural features of the MTJ from the newborn group revealed pattern during development with interactions between muscle cells and extracellular matrix elements with thin folds in the sarcolemma and high cellular activity evidenced through numerous oval mitochondria groupings. The adult group had classical morphological features of the MTJ, with folds in the sarcolemma forming long projections called "finger-like processes" and sarcoplasmic invaginations. Sarcomeres were aligned in series, showing mitochondria near the Z line in groupings between collagen fiber bundles. The old group had altered "finger-like processes," thickened in both levels of sarcoplasmic invaginations and in central connections with the lateral junctions. We conclude that the MTJ undergoes intense activity from newborn to its formation during adulthood. With increasing age, changes to the MTJ were observed in the shapes of the invaginations and "finger-like processes" due to hypoactivity, potentially compromising force transmission and joint stability.

Structured three‐dimensional co‐culture of mesenchymal stem cells with meniscus cells promotes meniscal phenotype without hypertrophy

Menisci play a crucial role in weight distribution, load bearing, shock absorption, lubrication, and nutrition of articular cartilage within the knee joint. Damage to the meniscus typically does not heal spontaneously due to its partial avascular nature. Partial or complete meniscectomy is a common clinical treatment of the defective meniscus. However, this procedure ultimately leads to osteoarthritis due to increased mechanical stress to the articular cartilage. Meniscus tissue engineering offers a promising solution for partial or complete meniscus deficiency. Mesenchymal stem cells (MSC) have the potential to differentiate into meniscal fibrochondrocyte as well as deliver trophic effects to the differentiated cells. This study tested the feasibility of using MSC co-cultured with mature meniscal cells (MC) for meniscus tissue engineering. Structured cell pellets were created using MC and MSC at varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) and cultured with or without transforming growth factor-beta 3 supplemented chondrogenic media for 21 days. The meniscal and hypertrophic gene expression, gross appearance and structure of the pellets, meniscus extracellular matrix (ECM), histology and immunohistochemistry of proteoglycan and collagen were evaluated. Co-culture of MC with MSC at 75:25 demonstrated highest levels of collagen type I and glycosaminoglycans (GAG) production, as well as the lowest levels of hypertrophic genes, such as COL10A1 and MMP13. All co-culture conditions showed better meniscus ECM production and hypertrophic inhibition as compared to MSC culture alone. The collagen fiber bundles observed in the co-cultures are important to produce heterogenic ECM structure of meniscus. In conclusion, co-culturing MC and MSC is a feasible and efficient approach to engineer meniscus tissue with enhanced ECM production without hypertrophy.

Evaluation of socket healing in irradiated rats after diode laser exposure (histological and morphometric studies)

ObjectiveThe purpose of this study is to evaluate the effect of low-level laser therapy (LLLT) in enhancing bone repair in irradiated sockets of albino rats. DesignThirty male Swiss Albino rats ranging from 120 to 150g were used in this study. The animals were subjected 6 gray gamma radiations. Three days post irradiation, right and left mandibular first molars were extracted. The sockets of the left sides were irradiated by (GaAIAs) diode laser device immediately after extraction, while the sockets of the right side were not exposed to the laser and served as control. The rats were randomly assigned into three groups (10 rats each) according to the date of sacrifice, 3, 7 and 10days into groups I, II and III, respectively. The two sides of each mandible were separated. Each group was further subdivided into subgroups A and B (10 specimens each), where A represents the right side of the mandible and B represents the left side. The specimens were stained with hematoxylin and eosin, and Masson's trichrome. ResultsLLLT accelerated bone healing, while, radiotherapy induced delay of bone healing along the three experimental groups. This acceleration was assessed histologically by the presence of mature collagen fibre bundles and early new bone formation in the lased groups. Histomorphometric analysis revealed an increase in the area percentage of bone trabeculae in the lased sockets compared to the control ones in group II. This increase was statistically significant (p=0.0274). The increase in the area percentage of bone trabeculae between the lased and control sockets of group III was statistically insignificant (p=0.1903). ConclusionsIn a rat model application of LLLT with a GaAIAs diode laser device can enhance bone healing and mineralisation in sockets subjected to gamma radiation.

Regeneration of Periodontal Ligament for Apatite-coated Tooth-shaped Titanium Implants with and without Occlusion Using Rat Molar Model

Regeneration of periodontal ligament (PDL) is still a major issue in implant dentistry. In this study, we evaluated the effects of hydroxyapatite (HA) coating and occlusion on the regeneration of PDL around tooth-shaped titanium implants with root form. Three groups of tooth-shaped titanium implants, namely, non-HA-coated without occlusion (HA-/OCL-), HA-coated without occlusion (HA+/OCL-), and HA-coated with occlusion (HA+/OCL+), were immediately implanted into extracted tooth sockets with remaining PDL of rat molar model and the regeneration of PDL was examined histomorphometrically and histologically. Direct bony attachments were formed at the HA+/OCL- implant interface. In contrast, soft peri-implant tissue was distinctly seen between the HA+/OCL+ implant surface and surrounding alveolar bones. The shape of the load-time curve for the HA+/OCL+ group was similar to that for natural teeth. Peri-implant tissue of the HA+/OCL+ implants exhibited alkaline phosphatase with a distribution and activity similar to those of the PDL. Collagen fiber bundles were found functionally oriented in some regions of the peri-implant tissue. These findings indicate that the remaining PDL tissue around extracted sockets has the ability to regenerate bone and PDL-like tissues on HA-coated tooth-shaped implants. Occlusal loads to the HA-coated implants may induce regeneration of PDL-like tissue in the peri-implant tissue.

Can Marine Mammals Be a Reliable Source for the Manufacture of Prosthetic Heart Valves for Percutaneous Surgery?

Marine mammals experience unique physiological conditions when diving. Myocardial function is sustained despite a 90% reduction of the blood flow in the coronaries. Therefore, their heart valves and pericardium could serve as a unique source of tissue for the manufacture of prosthetic heart valves. The pericardium of a stillborn pup sea lion was investigated to determine its morphology using gross observation, scanning electron microscopy (SEM), light microscopy, and transmission electron microscopy (TEM). Depending upon the site of sampling, the structure of the pericardium varied significantly. The atrial sample was well structured with wavy bundles of collagen fibers. The thickness in the atrial sample was regular with a smooth serous surface. The fibrous side of the pericardium of the auricular sample was irregular and incorporated microcapillaries. Both the sternoperitoneal ligament and the phrenoperitoneal ligament section were irregular and incorporated various amounts of adipocytes. Because of the increased amount of adipocytes, the fibrils of the collagen fibers were also observed to be occasionally agglutinated. Practically, the harvesting of pericardium would have to be restricted to the atrial surface. The presence of adipocytes in the pericardium wall makes the selection of this tissue a poor choice compared to alternative existing tissue sources.

The Layered Structure of Coronary Adventitia under Mechanical Load

The mechanical loading-deformation relation of elastin and collagen fibril bundles is fundamental to understanding the microstructural properties of tissue. Here, we use multiphoton microscopy to obtain quantitative data of elastin and collagen fiber bundles under in situ loading of coronary adventitia. Simultaneous loading-imaging experiments on unstained fresh coronary adventitia allowed morphometric measurements of collagen and elastin fibril bundles and their individual deformation. Fiber data were analyzed at five different distension loading points (circumferential stretch ratio λθ = 1.0, 1.2, 1.4, 1.6, and 1.8) at a physiological axial stretch ratio of λaxial = 1.3. Four fiber geometrical parameters were used to quantify the fibers: orientation angle, waviness, width, and area fraction. The results show that elastin and collagen fibers in inner adventitia form concentric densely packed fiber sheets, and the fiber orientation angle, width, and area fraction vary transmurally. The extent of fiber deformation depends on the initial orientation angle at no-distension state (λθ = 1.0 and λaxial = 1.3). At higher distension loading, the orientation angle and waviness of fibers decrease linearly, but the width of collagen fiber is relatively constant at λθ = 1.0–1.4 and then decrease linearly for λθ ≥ 1.4. A decrease of the relative dispersion (SD/mean) of collagen fiber waviness suggests a heterogeneous mechanical response to loads. This study provides fundamental microstructural data for coronary artery biomechanics and we consider it seminal for structural models.

Collagen fibrillogenesis in the development of the annulus fibrosus of the intervertebral disc

The annulus fibrosus of the intervertebral disc is a complex, radial-ply connective tissue consisting of concentric lamellae of oriented collagen. Whilst much is known of the structure of the mature annulus, less is known of how its complex collagenous architecture becomes established; an understanding of which could inform future repair/regenerative strategies. Here, using a rat disc developmental series, we describe events in the establishment of the collagenous framework of the annulus at light and electron microscopic levels and examine the involvement of class I and II small leucine rich proteoglycans (SLRPs) in the matrix assembly process. We show that a period of sustained, ordered matrix deposition follows the initial cellular differentiation/orientation phase within the foetal disc. Fibrillar matrix is deposited from recesses within the plasma membrane into compartments of interstitial space within the outer annulus - the orientation of the secreted collagen reflecting the initial cellular orientation of the laminae. Medially, we demonstrate the development of a reinforcing 'cage' of collagen fibre bundles around the foetal nucleus pulpous. This derives from the fusion of collagen bundles between presumptive end-plate and inner annulus. By birth, the distinct collagenous architectures are established and the disc undergoes considerable enlargement to maturity. We show that fibromodulin plays a prominent role in foetal development of the annulus and its attachment to vertebral bodies. With the exception of keratocan, the other SLRPs appear associated more with cartilage development within the vertebral column, but all become more prominent within the disc during its growth and differentiation.

Observations on the ultrastructure of equatorial scleral collagen fibrils in sheep eyes

To investigate collagen fibrils of the equatorial sclera in relation to the age-related changes in eye size in sheep. Lambs and outbred ewes. Sheep eyes (three lamb and three from adult outbred ewes), presumed disease-free, were processed for transmission electron microscopy (TEM) immediately postmortem. Tissue blocks from the equatorial region were sectioned across fibril bundles orientated along the equator. Micrographs including at least 500 fibrils were projected at 22,000× magnification for measures of fibril diameters (FDs). Lamb eyes were smaller than those of adult ewes but equatorial scleral thickness was only marginally less at 0.232±0.013 vs. 0.254±0.012 mm (P value not significant). Scleral tissue was composed of compacted bundles of collagen fibers that tended to be rounder in outer compared to being flatter in inner regions. In typical (normal) appearing regions, FDs were distinctly larger (68-410 nm) in outer sclera compared to inner sclera (63-281 nm). Outer sclera FDs were bimodal averaging 192±58 nm, compared to unimodal distributions at inner locations averaging 156±48 nm (P<0.001). Some atypical regions, especially at outer-mid sclera locations, were also noted where the FD distribution was bimodal but also included numerous microfibrils (<50 nm diameter), with similar appearances being found for both lamb and adult ewe eyes. The equatorial sclera is a mixture of rounder versus flatter collagen fiber bundles, the former being more likely to be made up of a mixture of both smaller and larger fibrils, as compared to slightly smaller fibrils.