Wavy Collagen Bundles Research Articles

760 Melanin may mediate visible light induced phototoxicity in darker skin types

Visible light (VL), a major component of solar radiation, differentially induces pigmentation in skin of color, an effect likely related to inherent melanization. A chromophore, melanin absorbs light within the VL wavelength. We hypothesize that clinical and histologic responses to VL are determined by epidermal melanin content and distribution. Nuclear matrix metalloproteinase (MMP-1), a proapoptotic mediator of photodamage may play a central role in skin type specific responses to VL. Enrolled participants were grouped by skin type (light=6,dark=6). Photoprotected sites were exposed daily x4days to 480J/cm2 of VL at 200mW(FiberLite High Intensity Illuminator Series 180,Dolan Jenner Industries) for a total dose of 1920J/cm2as tolerated. Treated and control sites were biopsied 24h after the last exposure and stained with Masson’s Trichrome and Fontana-Masson (FM) to assess collagen and melanin deposition, respectively. MMP-1 deposition was detected by indirect immunofluorescence (IF). Dark skin types did not tolerate the full VL regimen with blistering occurring in all subjects at doses of 220-880J/cm2. We found epidermal MMP-1 expression and nuclear localization in keratinocytes in dark but not in light skin. Nuclear localization correlated with areas of highest melanin density per FM stain. While there was no significant difference in MMP expression of treated light skin from control, collagen stains showed decreased stain intensity and thickened, wavy collagen bundles in both light (mean fold change[FC]=1.46,SEM+0.11), and dark skin (FC=1.19,+0.05). Herein, we demonstrate the acute effects of high power VL exposure in light and dark skin. VL can cause degenerative changes in collagen similar to acute UV irradiation or thermal injury, occurring independent of skin melanization. While the role of melanin in photoprotection is well defined, given the pro-apoptotic function of nuclear MMPs, our findings suggest a potential mechanism by which melanin may mediate VL induced phototoxicity. These observations show a “light” and “dark” side to melanin that warrants further investigation.

Identification of Human Pathological Mitral Chordae Tendineae Using Polarization-sensitive Optical Coherence Tomography

Defects of the mitral valve complex imply heart malfunction. The chordae tendineae (CTs) are tendinous strands connecting the mitral and tricuspid valve leaflets to the papillary muscles. These CTs are composed of organized, wavy collagen bundles, making them a strongly birefringent material. Disorder of the collagen structure due to different diseases (rheumatic, degenerative) implies the loss or reduction of tissue birefringence able to be characterized with Polarization Sensitive Optical Coherence Tomography (PS-OCT). PS-OCT is used to discriminate healthy from diseased chords, as the latter must be excised and replaced in clinical conventional interventions. PS-OCT allows to quantify birefringence reduction in human CTs affected by degenerative and rheumatic pathologies. This tissue optical property is proposed as a diagnostic marker for the identification of degradation of tendinous chords to guide intraoperative mitral valve surgery.

The Red Kangaroo pericardium as a material source for the manufacture of percutaneous heart valves

The kangaroo pericardium might be considered to be a good candidate material for use in the manufacture of the leaflets of percutaneous heart valves based upon the unique lifestyle. The diet consists of herbs, forbs and strubs. The kangaroo pericardium holds an undulated structure of collagen. A Red Kangaroo was obtained after a traffic fatality and the pericardium was dissected. Four compasses were cut from four different sites: auricular (AUR), atrial (ATR), sternoperitoneal (SPL) and phrenopericardial (PPL). They were investigated by means of scanning electron microscopy, light microscopy and transmission electron microscopy. All the samples showed dense and wavy collagen bundles without vascularisation from both the epicardium and the parietal pericardium. The AUR and the ATR were 150±25μm thick whereas the SPL and the PPL were thinner at 120±20μm. The surface of the epicardium was smooth and glistening. The filaments of collagen were well individualized without any aggregation, but the banding was poorly defined and somewhat blurry. This detailed morphological analysis of the kangaroo pericardium illustrated a surface resistant to thrombosis and physical characteristics resistant to fatigue. The morphological characteristics of the kangaroo pericardium indicate that it represents an outstanding alternative to the current sources e.g., bovine and porcine. However, procurement of tissues from the wild raises supply and sanitary issues. Health concerns based upon sanitary uncertainty and reliability of supply of wild animals remain real problems.

Microstructural alterations owing to handling of bovine pericardium to manufacture bioprosthetic heart valves: A potential risk for cusp dehiscence

Cross-linking and anti-calcification of prosthetic heart valves have been continuously improved to prevent degeneration and calcification. However, non-calcific structural deteriorations such as cuspal dehiscences along the stent still require further analysis. Based upon the previous analysis of an explanted valve after 7years, a fresh commercial aortic valve was embedded in poly(methyl methacrylate) (PMMA) and cut into slices to ensure the detailed observation of the assembly and material structures. A pericardial patch embossed to provide the adequate shape of the cusps was investigated after paraffin embedding and appropriate staining. The microstructural damages that occurred during manufacturing process were identified and evaluated by light microscopy, polarized microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The wavy collagen bundles, the key structure of the pericardium patch, were damaged to a great extent at suture sites along the stent and in the compressed areas around the stent post. The fixation of the embossed pericardium patch along the plots of the stent aggravated the microstructural modifications. The damages mainly appeared as the elimination of collagen bundle waviness and delamination between the bundles. Considering the modes of failure of the explant, the damages to the collagen bundles may identify the vulnerable sites that play an important role in the cusp dehiscence of heart valve implants. Such information is important to the manufacturers. Recommendations to prevent in vivo cusp dehiscence can therefore be formulated.

Unusual Case of a Proptosed Eye: Isolated Right Maxillary Neurofibroma.

Neurofibroma is a slow growing benign tumour of the peripheral nerve sheath which is frequently associated with neurofibromatosis type 1 (Prakash et al., 2014). Isolated solitary occurrence of neurofibroma in the maxillary sinus is rare with only 29 reported cases in the literature. We present a rare case of a 70-year-old gentleman who was referred to ENT with a right maxillary sinus neurofibroma with extension into the right inferior orbit. He has significant proptosis, ptosis, and limitation in abduction of the right eye. He has a complicated past history of multiple neurofibromas which were completely excised. Craniofacial MRI shows a large mass filling the right maxillary antrum extending anteriorly into subcutaneous tissue towards nasal ala and posterolaterally into inferior temporal fossa and superiorly into orbit and cavernous sinus involvement. Biopsy of the right maxillary mass revealed cellular spindle cell tumour with wavy collagen bundles within myxoid stroma which is consistent with a neurofibroma. Patient's case was discussed in the skull-base MDT and he has been referred to a specialist center for surgical removal of the neurofibroma with reconstructive surgery. Despite the rarity of this disease, otorhinolaryngologist should consider a possibility of neurofibroma of the paranasal sinuses.

Capsular Contracture after Calf Augmentation with Silicone Implant Insertion.

Capsular Contracture after Calf Augmentation with Silicone Implant Insertion.

Helicase-Like Transcription Factor (Hltf) Regulates G2/M Transition, Wt1/Gata4/Hif-1a Cardiac Transcription Networks, and Collagen Biogenesis

HLTF/Hltf regulates transcription, remodels chromatin, and coordinates DNA damage repair. Hltf is expressed in mouse brain and heart during embryonic and postnatal development. Silencing Hltf is semilethal. Seventy-four percent of congenic C57BL/6J Hltf knockout mice died, 75% within 12-24 hours of birth. Previous studies in neonatal (6-8 hour postpartum) brain revealed silencing Hltf disrupted cell cycle progression, and attenuated DNA damage repair. An RNA-Seq snapshot of neonatal heart transcriptome showed 1,536 of 20,000 total transcripts were altered (p < 0.05) - 10 up- and 1,526 downregulated. Pathway enrichment analysis with MetaCore™ showed Hltf’s regulation of the G2/M transition (p=9.726E-15) of the cell cycle in heart is nearly identical to its role in brain. In addition, Brca1 and 12 members of the Brca1 associated genome surveillance complex are also downregulated. Activation of caspase 3 coincides with transcriptional repression of Bcl-2. Hltf loss caused downregulation of Wt1/Gata4/Hif-1a signaling cascades as well as Myh7b/miR499 transcription. Hltf-specific binding to promoters and/or regulatory regions of these genes was authenticated by ChIP-PCR. Hif-1a targets for prolyl (P4ha1, P4ha2) and lysyl (Plod2) collagen hydroxylation, PPIase enzymes (Ppid, Ppif, Ppil3) for collagen trimerization, and lysyl oxidase (Loxl2) for collagen-elastin crosslinking were downregulated. However, transcription of genes for collagens, fibronectin, Mmps and their inhibitors (Timps) was unaffected. The collective downregulation of genes whose protein products control collagen biogenesis caused disorganization of the interstitial and perivascular myocardial collagen fibrillar network as viewed with picrosirius red-staining, and authenticated with spectral imaging. Wavy collagen bundles in control hearts contrasted with collagen fibers that were thin, short and disorganized in Hltf null hearts. Collagen bundles in Hltf null hearts were tangled and fragmented. Thus, silencing Hltf during heart organogenesis compromised DNA double-strand break repair, and caused aberrant collagen biogenesis altering the structural network that transmits cardiomyocyte force into muscle contraction.

Editorial Comment: Pericardial traumatic injury in trans-catheter aortic valve implantation

Trans-catheter aortic valve implantation (TAVI) is a very promising new treatment for native aortic stenosis or failed bioprostheses [1]. Glutaraldehyde xenograft tissue is employed because of its pliability for folding during implantation and minimizing its size in order to reach its final position in the aortic root. In the CoreValve system, the stent is self-expanding, whereas in the Edwards Sapien, the valve has to be dilated by balloon inflation to approximate the stent to the aortic annulus without suturing, so as to prevent escape of the prosthesis and paravalvular leakage. In the latter procedure the pericardium is crushed against the stent. Since the operation is relatively easily accomplished with early success, the question arises of whether it might also be indicated in low-risk patients, thus avoiding sternotomy, general anaesthesia, cardio-pulmonary bypass and cardiac arrest. However, in contrast to the traditional surgical implant, TAVI requires cusp crimping and, in the case of the Edwards Sapien, balloon inflation. Both manoeuvres may be responsible for tissue injury. Previous observations by Zegdi et al. have shown evidence of cusp injury following percutaneous aortic valve deployment of the Edwards Sapien [2]. To confirm these preliminary findings, the same group made an experimental study, comparing the severity of traumatic cusp injury in home-made balloonvs self-expanding valved stents (VS); a non-crimped pericardium was used as a control [3]. According to the authors’ findings, all deployed valves were associated with microscopic lesions, in keeping with traumatic disruption, both in terms of transverse fractures and longitudinal cleavages. Transverse fractures were seen both in balloonand in self-expanding VS, whereas longitudinal cleavages were more frequently observed in balloon-expanded VS. Crushing and shearing of the pericardium during crimping and deployment are the most plausible causes. They may facilitate plasma and lipid insudation with tearing in the long term, thus affecting durability. These data support and expand the preliminary observations of the same authors [2]. An editorial comment by our group expressed some concern in interpreting these data, due to the limitations of technology used in pathological investigations [4]. However, the new data provided herein by Amahzoune et al. [3] seem to be convincing, even in the absence of macroscopic evidence of lacerations, commissural dehiscences or tears. They support the occurrence of some iatrogenic damage, particularly in balloon-expandable VS. However, in our opinion, the experiment should be repeated in a core lab employing special histological staining, such as picrosirius red, and scanning electron microscopy, to assess collagen waviness and to exclude the artifacts arising from manipulation at histology. If confirmed, these observations may cast a shadow on the safety of the TAVI procedure in terms of tissue integrity and biological compatibility, although crimping was proven not to influence propensity to cusp calcification in the long term [5]. Glutaraldehyde-fixed pericardium is the best available tissue because of its strength (thick fibrosa) and pliability (elasticity of wavy collagen bundles). If handled gently, as in the current secondand third-generation stented or stentless pericardial valve prostheses, it is not at risk of mechanical failure. Whether this is the case with TAVI—in which the pericardium is exposed to crimping or even crushing, with a potential traumatic injury— is a matter of concern. According to the findings by Amahzoune et al. [3], the following considerations may be put forward:

Lateral ligament injury of the ankle and associated articular cartilage degeneration in the talocrural joint: anatomic study using elderly cadavers

We investigated degenerative changes (DCs) of the articular cartilage using 33 cadaveric talocrural joints with anterior talofibular ligament injuries. The control group (93 normal ankles) were age- and sex-matched with the injury group. The injured ligament carried unusual wavy collagen bundles with hyalinization (or elongation) or rupture with scar. In both groups, we found talar-side dominance of DCs, multifocus occurrence, and a high incidence of mirror-image DCs in the medial parts of the joint. However, the mirrorimage lesion was multiple or large (or both) in the injury group in contrast to its restricted nature in the control. Thus, the averaged numbers of affected areas or sectors in the injury group were almost twice as large as in the controls. A significantly high incidence of DCs, especially of the mirrorimage lesion, was found in the lateral malleolar facet in the injury group. Although initial DC lesions might be at a medial site in the joint, as hypothesized in the control group, the lateral ligament injury seemed to increase the dominance of DCs on the lateral site. Consequently, early evaluation and treatment for lateral ligament insufficiency is more necessary in elderly patients than in younger patients to avoid widespread development of osteoarthritis.

Bulbus arteriosus of the antarctic teleosts. I. The white-blooded Chionodraco hamatus.

The bulbus arteriosus of teleost fish is a thick-walled chamber that extends between the single ventricle and the ventral aorta. The functional importance of the bulbus resides in the fact that it maintains a steady blood flow into the gill system through heart contraction. Despite of this, a thorough study of the structure of the bulbus in teleost fish is still lacking. We have undertaken a morphologic study of the bulbus arteriosus in the stenothermal teleosts of the Antarctic sea. The structural organization of the bulbus arteriosus of the icefish Chionodraco hamatus has been studied here by conventional light, scanning, and transmission electron microscopy. The inner surface of the bulbus shows a festooned appearance due to the presence of longitudinal, unbranched ridges that extend between the ventricle and the arterial trunk. The wall of the bulbus is divided into endocardial, subendocardial, middle, and external layers. Endocardial cells show a large number of moderately-dense bodies. The endocardium invaginates into the subendocardium forming solid epithelial cords that contain numerous secretory vacuoles. Cells in the subendocardium group into small domains, have some of the morphological characteristics of smooth muscle cells, and appear enmeshed in a three-dimensional network of matrix filaments. Cells in the middle layer are typical smooth muscle cells. They appear arranged into layers and are surrounded by a filamentous meshwork that excludes collagen fibers. Orientation of this meshwork occurs in the vicinity of the smooth muscle cells. Elastin fibers are never observed. The external layer is formed by wavy collagen bundles and fibroblast-like cells. This layer lacks blood vessels and nerve fibers. The endocardium and the endocardium-derived cords are secretory epithelia that may be involved in the formation ofmucins or glycosaminoglycans. These mucins may have a protecting effect on the endocardium. The subendocardium and the middle layer appear to be formed by the same cell type, smooth muscle, with a gradient of differentiation from the secretory (subendocardium) to the contractile (middle layer) phenotype. Despite the absence of elastin fibers, the filamentous matrix could maintain the elastic properties of the bulbus wall. Smooth muscle cells appear to be actively involved in bulbus wall dynamics. The restriction of collagen to the external layer suggests that it may control wall dilatation and bulbus compliance. When comparison was possible, structural differences between C. hamatus and temperate teleosts seemed to be not species-related, but of phenotypic adaptative significance. This is remarkable since Antarctic fishes have lived isolated in freezing waters for the last two million years.

Structural Changes of Collagen Components and Diminution of Nerves in Congenital Ureteropelvic Junction Obstruction

Purpose Three-dimensional arrangements of smooth muscle cells, collagenous component and peripheral nerves of congenital ureteropelvic junction (UPJ) obstruction were studied in order to clarify the pathogenetic mechanism of interaction among these neuro-myo-stromal components. Materials and Methods The UPJ and upper ureters were obtained from 14 patients with congenital hydronephrosis (7 intrinsic and 4 extrinsic obstruction) and 7 normal controls. Three-dimensional arrangement of each structural component was observed by scanning electron microscopy, and the nerve distribution was analyzed with immunohistochemistry for protein gene product 9.5. Results The UPJ of intrinsic obstruction had structural features as follows. Muscle fascicles were sparse and thin. Each muscle cell was thin in diameter. Intercellular spaces were six to seven times wider than controls. Collagen fibrillar sheaths of smooth muscle cells (pericellular collagen fibrils attached to the basement membrane) were interwoven to form a dense felt-like structure against thin lace-like sheaths in controls. Interstitial collagenous component showed dense and compact structure against loose network of wavy collagen bundles in controls. In the muscular layer, nerve distribution was decreased to about one-third of controls. In contrast, non-stenotic portion of intrinsic UPJ obstruction as well as materials from extrinsic UPJ obstruction showed no structural difference as compared with controls. Conclusions In the intrinsic obstruction, nerve fibers were depleted in the muscular layers in the ureteric walls, resulting in dysfunction and atrophy of muscle fibers and an increase of collagen fibers in the muscle layers with abnormal accumulation of intercellular and interstitial collagen. These changes may disrupt the mobility of UPJ and lead to both mechanical and functional obstruction.

Atrioventricular valves of the mouse: III. Collagenous skeleton and myotendinous junction

The leaflet tissue of the mouse atrioventricular (AV) valves contains a system of wavy collagen bundles that organize like tendons, orientate along lines of tension, and constitute an essential component of the valve tissue. The organization of these bundles is different in the two AV valves, reflecting differences in the anatomy of the entire valvular complex. Further insights into this kind of organization are needed to gain a complete understanding of the functional anatomy of the mouse AV valves. The endocardial covering of the mouse AV valves (from 21 days to 1 year of age) was eliminated by the sonication or the maceration method. This allowed us to study in situ the organization of the collagenous valve skeleton, as well as the structure of the myotendinous junction. The leaflets of the two AV valves are formed by a fibrous layer (on the ventricular side) and a spongy layer (on the atrial side). The fibrosa is formed by undulating collagen bundles that organize and orientate differently on the right and left sides. The spongiosa is formed, on both sides, by a loose network of thin collagen fibers with no apparent orientation. Myocardial cells in the papillary muscles of the tricuspid valve are elongated and show cone-shaped tips. Collagen fibers attach to the myocyte surface. Collagen struts and thin septa can also be recognized. On the other hand, the collagenous components of the mitral leaflets attach tangentially to the mitral papillary muscles. On the two sides, the myocytes appear to be ensheathed in a layer of collagenous tissue. The sheaths are formed by circularly arranged fibers and appear to be tightly interconnected. The differences in the collagenous organization between the two AV valves reflect differences in the gross anatomy of the valves. The attachment of collagen to the papillary myocytes in the tricuspid valve resembles that of a typical myotendinous junction. However, the collagen-muscle junction in the mitral valve is more similar to the structure of a pennate muscle. The collagen matrix of the heart has been divided into endomysial, perimysial, and epimysial components. The presence of sheaths housing individual myocytes and capillaries, struts, and thin septa, corresponds to the endomysium. The absence of perimysial septa, which aggregate myocytes into groups, is striking, but this may just be a species difference. The appropriateness of the term epimysium, as applied to the heart, is discussed.

Three-Dimensional Arrangement of Collagen and Elastin Fibers in the Human Urinary Bladder: A Scanning Electron Microscopic Study

To clarify the arrangements of collagen and elastin fibers of the urinary bladder, we examined 9 human (male, aged 42 to 72) urinary bladders by scanning electron microscopy with chemical digestion methods. The mucosal layer was divided into 3 portions according to the collagen arrangement: the superficial portion interwoven densely by collagen fibrils, the middle portion layered by flat bundles of collagen fibrils and the deep portion made of a loose network of twisted collagen bundles. In the muscular layer, the smooth muscle fascicles were firmly covered with collagen sheets, while each muscle cell in a fascicle was accommodated by a thin sheath of collagen fibrils. The serosal layer consists of wavy collagen bundles piled up in a sheet, which was intercalated by clusters of adipose cells. Elastic fibers were, on the other hand, sparse throughout the bladder wall, except for denser networks around the blood vessels and muscle fascicles and beneath the peritoneal mesothelium. The arrangements of these components were discussed in relation to the mechanical function and compliance of the urinary bladder.