Thickness Of Collagen Bundles Research Articles

Similar collagen distribution in full-thickness skin grafts in intraperitoneal and onlay positions, an experimental mice-study

PurposeAutologous full-thickness skin grafting (FTSG) has the potential to become an option in abdominal wall repair. An understanding of tissue remodelling in the extracellular matrix (ECM) is crucial as this interplay determines such parameters as tissue strength and flexibility. This cross-sectional preclinical laboratory study in mice provides information on the distribution of collagen types and matrix metalloproteinases (MMPs) in the ECM of FTSGs in the intraperitoneal and onlay positions compared with internal controls. The aim was to evaluate morphologic changes after tissue remodelling and repair in FTSGs applied in the two positions and to detect any adverse host response.MethodsECM components were evaluated as follows: qualitative examination of collagen bundle thickness using Picrosirius Red staining (collagen types I, III and IV); and evaluation of collagen types IV and V, as well as MMPs 1, 8 and 9 using immunohistochemical staining. Full-thickness grafts transplanted between female twin mice were examined as this best mimics autologous transplantation.ResultsAt 8 weeks, FTSGs in the intraperitoneal position did not show any noticeable differences in morphologic appearance to those in the onlay position. Both intraperitoneal and onlay FTSGs showed increases in the amount of thick collagen bundles compared to internal controls. No correlation was seen between distribution of MMPs 1, 8 or 9 and distribution of collagen types I, III, IV or V.ConclusionThis preclinical study shows that FTSGs in both intraperitoneal and onlay positions are possible application site options and, by extension, promising application site options for abdominal wall reinforcement in hernia surgery. Clinical studies in humans are required to confirm these findings.

Intense pulsed light versus cryotherapy in the treatment of hypertrophic scars: A clinical and histopathological study.

Different therapeutic modalities have been tried for hypertrophic scar treatment. To our knowledge, intense pulsed light (IPL) has not been previously evaluated in comparison with cryotherapy as a stand-alone treatment for hypertrophic scars. We aimed to evaluate the efficacy of IPL as a monotherapy for hypertrophic scar treatment as compared with cryotherapy both clinically and histopathologically. This study included 28 patients with hypertrophic scars. Patients were divided randomly and equally into two groups; group I patients received cryotherapy while group II patients received IPL. All patients received treatments for a total of six sessions or until resolution of the lesion whichever was nearer. The outcome was evaluated clinically and histopathologically. Scar height showed a significant decrease and scar color and pliability showed a significant improvement in group I. No significant changes were detected in group II except in scar pliability. Vancouver scar scale (VSS) mean decreased by -53.7% in group I versus -11.5% decrease in group II. Histopathologically, group I showed a significantly increased epidermal thickness and decreased dermal and collagen bundle thickness, while group II showed insignificant histopathological changes. Group I exhibited a statistically significant clinical and histopathological improvement compared to group II, yet with more complications. Cryotherapy is more effective than IPL in the treatment of hypertrophic scars both on clinical and histopathological level yet with more complications.

Ameliorative effects of paricalcitol against 1800-MHz mobile phone radiation-induced skin damage in rats

This study aimed to evaluate the protective role of paricalcitol, a vitamin D analog, against hazardous effects of 1800-MHz radiofrequency (RF) radiation on the skin of male Wistar albino rats exposed one h/day for 4 weeks. The maximum specific absorption rate was calculated as 0.0042 W/kg. Superoxide dismutase and catalase activity, collagen bundle thickness, breaking force, energy, and stress were decreased, while malondialdehyde level and mast cell number were increased. The exposure to 1800-MHz RF radiation caused a reduction in skin biomechanical quality through oxidative stress and mast cell activation. The administration of paricalcitol caused a significant improvement.

959: Molecular characterization of the post cesarean uterine scar

Cesarean section (CS) accounts for 31.9% of births in the United States and is associated with a 3.6-fold increase in maternal mortality, in part due to complications in future pregnancies such as uterine rupture or placenta accreta. CS technique is largely anecdotal because it is not known how the unique structure of the uterus nor CS surgical factors impact scar formation. The purpose of this study was to investigate the histologic structure and density of post-CS uterine scar. Lower uterine segment was prospectively biopsied at the time of repeat CS (n=4). Samples from a primary CS and premenopausal non-pregnant hysterectomy specimen were used for comparison. Data collected included demographic information and inter-conception interval (ICI). Biopsy specimens were formalin fixed, paraffin embedded, and sections were stained with Masson Trichome to differentiate collagen (blue) and myometrium (red/purple). Representative images were analyzed using ImageJ software. Collagen bundle thickness was quantified, and ANOVA was used to detect differences between samples. Tissue specimens were sent for 3- dimensional molecular imaging with MicroCT. Vivoquant software was used for segmentation analysis of MicroCT images. Differences in tissue density were compared using Chi-square, with significance at p<0.05. Collagen bundles were thicker and more disorganized in subjects undergoing repeat CS compared to unscarred pregnant and post-menopausal specimens (p < 0.001). The range of collagen bundle size was also wider in subjects with a shorter 29-34 month ICI compared to a longer 106-108 month ICI (2.37-23.84 mm vs 2.37-9.00 mm). MicroCT analysis revealed a heterogeneous distribution of scar density within RCS tissue samples (p < 0.001), with collagen density randomly distributed between serosal and endometrial surfaces. Post-CS scar structure is characterized by increased collagen bundle thickness compared to native uterine tissue. Scar structure is dynamic over time with more abundant and disorganized collagen bundles present earlier in the healing process, and more contracted scar over time. MicroCT analysis shows heterogeneity in scar density, even among samples of similar ICI, suggesting the importance of individual variation. Further study is needed to determine if molecular characteristics of uterine scar structure are associated with adverse pregnancy outcomes.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Comparative Study on Surface Geometric Properties of Decellularized and Glutaraldehyde fixed Bovine Pericardium and Possible Effects on Cell Seeding

Background:Collagen fibers arrangement and density play a pivotal role in cell migration and proliferation. Bovine Pericardium (BP) is a collagen – rich tissue which is currently used as a biological scaffold.Aim of the work:The aim of this study was to examine the effects of the decellulrazation processes on the surface architecture and geometry of the BP and consequently cell seeding and epitheliliazation onto it.Methods and Results:To achieve these goals bovine pericardium (BP) was decellularized with 1 % TritonX-100 and 0.1 % sodium dodecyl sulfate . Control group was only treated by PBS and post fixed in glutaraldehyed (GAD) 1%. The buccal cell suspensions were then overlaid on the serous side of the BP scaffolds at concentrations2×105 Cells/mL. 10 days following seeding the samples were fixed and SEM results were analyzed by Matlab software. The pore surfaces were measured 182.05±11.61μm2 and 132.44±12.35 μm2 in acellular and GAD BP respectively (P<0.05). The con surfaces were measured 93.54±13.41 μm2and 114.78±11.67 μm2 acellular and GAD PB respectively (P<0.05).The thickness of collagen bundles in decellularized BP and GAD BP (10 random fields) were obtained 19.5±6.3 μm and 16.8±.99 μm respectively (p<0.05).Conclusion: The results of acellular group showed more attached cells onto scaffold, epithelialization growth, and spreading on the scaffold. While in contrast GAD-fixed group only scattered cell clumping were seen. Our findings showed that topographical changes after decellularization could provide a suitable growth pool or microenvironment, that can influence cellular attachment and subsequently cell-ECM integration in biological scaffold.

The histomorphometric and histopathologic effect of methionine on the epidermis and dermis layers of skin in rat

The skin is the largest organ of the body and is composed of epidermis (the outermost layer of cells consisting mainly of keratinized, stratified, squamous epithelium) and dermis (a layer of connective tissue composed of cells and fibrils such as collagen bundles). The aim of this study was to assess the effect of excess methionine on the histomorphology and histomorphometry of collagen bundles in these layers. Eighty, 9–10-week old, male rats were randomly allocated into eight equal groups. Each group received one of the following treatments: L-methionine (50, 100 or 200 mg/kg B.W.), vitamin C (50 mg/kg B.W), combination of L-methionine (all 3 concentrations) and vitamin C (Met + Vit C), or normal saline as control (CT) for 20 consecutive days. After terminating the animals on day 21, skin samples were taken, and the epidermis, papillary, and reticular layers were evaluated histologically. The data obtained revealed that methionine does not increase collagen bundle thickness unless combined with vitamin C. We suggest that lack of crosslinking occurs when using methionine alone. The greatest increase in both epidermis and dermis occurred in the Met 100 + Vit C group. There was no significant difference between Vit C and control groups. Histopathology did not show any congestion or inflammatory cell infiltration in any treatment groups compared to the controls.

Tissue Inhibitor of Metalloproteinase-2 Suppresses Collagen Synthesis in Cultured Keloid Fibroblasts.

Background:Keloids are defined as a kind of dermal fibroproliferative disorder resulting from the accumulation of collagen. In the remodeling of extracellular matrix, the balance between matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) is as critical as the proper production of extracellular matrix. We investigate the role of TIMPs and MMPs in the pathogenesis of keloids and examine the therapeutic potential of TIMP-2.Methods:The expression of TIMPs and MMPs in most inflamed parts of cultured keloid fibroblasts (KFs) and peripheral normal skin fibroblasts (PNFs) in the same individuals and the reactivity of KFs to cyclic mechanical stretch were analyzed by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay (n = 7). To evaluate the effect of treating KFs with TIMP-2, collagen synthesis was investigated by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay, and microscopic analysis was used to examine the treatment effects of TIMP-2 on ex vivo cultures of keloid tissue (n = 6).Results:TIMP-2 was downregulated in cultured KFs compared with PNFs in the same individuals, and the reduction in TIMP-2 was exacerbated by cyclic mechanical stretch. Administration of TIMP-2 (200 or 300 ng/mL) significantly suppressed expression of Col1A2 and Col3A1 mRNA and collagen type I protein in KFs. TIMP-2 also significantly reduced the skin dermal and collagen bundle thickness in ex vivo cultures of keloid tissue.Conclusion:These results indicated that downregulation of TIMP-2 in KFs is a crucial event in the pathogenesis of keloids, and the TIMP-2 would be a promising candidate for the treatment of keloids.

Morphological and mechanical examination of the atrial 'intima'

To examine morphology and mechanical properties of the atrial 'intima', which we defined as the tissue interposed between atrial endocardium and myocardium, in patients without known cardiovascular disease. Post-mortem right and left atrial tissue was obtained from male infants (<1 year, n = 4), children (10-19 years, n = 4), and adults (58-69 years, n = 7). Using light microscopy and an ocular micrometer, atrial intimal (AIT) thickness was measured. Intimal collagen bundle thickness was measured using electron microscopy. Passive atrial wall stiffness was measured using a planar biaxial testing device. Among infants, left AIT (0.2 ± 0.2 mm) and right (0.2 ± 0.1 mm) AIT were not significantly different (P = 0.84). Among children, left AIT (0.6 ± 0.2 mm) was significantly greater than right (0.2 ± 0.1 mm) AIT (P = 0.03), and left AIT was marginally greater than in infants (P = 0.07). Among adults, with the exception of the appendage region, left AIT (1.0 ± 0.2 mm) was markedly greater than right AIT (0.3 ± 0.1 mm; P < 0.05), and left AIT was significantly greater than that in other age groups (P < 0.05). There were no differences in right AIT among age groups. Left intimal collagen bundle thickness was greater in adults (0.0512 ± 0.0056 µm) than infants (0.0432 ± 0.0071 µm) or children (0.0435 ± 0.0013 µm), and bundles were less organized. Wall stiffness was attributable primarily to the intima (1245 ± 132, vs. 260 ± 45 N/m(2) for the remaining atrial wall). The left atrial intima, but not the right, thickens with age, becomes more disorganized ultrastructurally, and is responsible for the majority of atrial wall stiffness.

Effects of Natural Beta-carotene and Commercially Available Combination of Beta-carotene, Vitamin E, Vitamin C and Minerals: A Prospective Comparative Study

Aim and objective: The purpose of this study was to determine effects of natural beta-carotene and revox in, reducing the oral symptoms of submucous fibrosis patients, thickness of collagen fibers in submucous fibrosis patients and to correlate the changes in epithelium and connective tissue. Materials and methods: The study population consisted of 50 patients with submucous fibrosis divided in two groups randomly. For patents in group A recived intralesional injection of 0.5 ml hyaluronidase 1500 IU and 05 ml hydrocortisone acetate 25 mg/ml were injected in buccal mucosa bilaterally in alternative week and natural beta-carotene capsule twice daily for 12 weeks along with ice cream stick mouth exercise. For group B patients intralesional injections and capsule revox were given twice daily for 12 weeks along with ice cream stick mouth exercise. Results: There is statistical significant improvement in mouth opening, reduction of burning sensation and decrease in thickness of collagen bundles were seen in all patients but there was early response in group A than group B Conclusion: Our preliminary findings suggest that both groups showed statistical significant results with early response in group A. The clinical comparison and effectiveness of natural beta-carotene and revox is not statistical significant.

Collagen bundle morphometry in skin and scar tissue: a novel distance mapping method provides superior measurements compared to Fourier analysis

Histopathological evaluations of fibrotic processes require the characterization of collagen morphology in terms of geometrical features such as bundle orientation thickness and spacing. However, there are currently no reliable and valid techniques of measuring bundle thickness and spacing. Hence, two objective methods quantifying the collagen bundle thickness and spacing were tested for their reliability and validity: Fourier first-order maximum analysis and Distance Mapping, with the latter constituting a newly developed morphometric technique. Histological slides were constructed and imaged from 50 scar and 50 healthy human skin biopsies and subsequently analyzed by two observers to determine the interobserver reliability via the intraclass correlation coefficient. An intraclass correlation coefficient larger than 0.7 is considered as representing good reliability. The interobserver reliability for the Fourier first-order maximum and for the Distance Mapping algorithms, respectively, showed an intraclass correlation coefficient above 0.72 and 0.89. Additionally, we performed an assessment of validity in the form of responsiveness, in particular, demonstrating medium to excellent results via a calculation of the effect size, highlighting that both methods are sensitive enough to measure a treatment effect in clinical practice. In summary, two reliable and valid measurement methods were demonstrated for collagen bundle morphometry for the first time. Due to its superior reliability and more useful measures (bundle thickness and bundle spacing), Distance Mapping emerges as the preferred and more practical method. Nevertheless, in the future, both methods can be used for reliable and valid collagen morphometry of skin and scars, whereas further applications evaluating the quantitative microscopy of other fibrotic processes are anticipated.

Human Skin Culture as an Ex Vivo Model for Assessing the Fibrotic Effects of Insulin-Like Growth Factor Binding Proteins

Systemic sclerosis (SSc) is a connective tissue disease of unknown etiology. A hallmark of SSc is fibrosis of the skin and internal organs. We recently demonstrated increased expression of IGFBP-3 and IGFBP-5 in primary cultures of fibroblasts from the skin of patients with SSc. In vitro, IGFBP-3 and IGFBP-5 induced a fibrotic phenotype and IGFBP-5 triggered dermal fibrosis in mice. To assess the ability of IGFBPs to trigger fibrosis, we used an ex vivo human skin organ culture model. Our findings demonstrate that IGFBP-3 and IGFBP-5, but not IGFBP-4, increase dermal and collagen bundle thickness in human skin explants, resulting in substantial dermal fibrosis and thickening. These fibrotic effects were sustained for at least two weeks. Our findings demonstrate that human skin ex vivo is an appropriate model to assess the effects of fibrosis-inducing factors such as IGFBPs, and for evaluating the efficacy of inhibitors/therapies to halt the progression of fibrosis and potentially reverse it.

Insulin‐like growth factor binding protein 5 induces skin fibrosis: A novel murine model for dermal fibrosis

To determine the role of insulin-like growth factor binding protein 5 (IGFBP-5) in the development of skin fibrosis in vivo, by examining the effect of overexpression of IGFBP-5 in mouse skin. Wild-type C57BL/6J mice were injected subcutaneously with replication-deficient serotype 5 adenovirus expressing human IGFBP-3 (Ad3), IGFBP-5 (Ad5), or no complementary DNA (cAd). Mice were killed 3, 8, or 22 days postinjection. The dermal thickness and dermal collagen bundle thickness in skin sections were measured. The deposition of collagen in the extracellular matrix (ECM) was quantified using the Sircol assay. Expression of proliferating cell nuclear antigen (PCNA) and fibronectin, as determined by immunohistochemical analysis, was used to evaluate fibroblast activation, and vimentin and alpha-smooth muscle actin (alpha-SMA) were used to evaluate the fibroblast phenotype. Adenovirally expressed IGFBP was detected in dermal fibroblasts, endothelial cells, epithelial cells, and muscle bundles in Ad3- and Ad5-injected mice. Increased collagen deposition, denser dermal connective tissue, and increased collagen bundle thickness were observed in IGFBP-5-overexpressing mice. Dermal thickness and collagen bundle thickness were significantly increased in Ad5-injected mice compared with cAd- and Ad3-injected mice. Treatment with Ad5 resulted in a dose-dependent increase in dermal and collagen bundle thickness. Increased deposition of collagen and fibronectin, increased numbers of PCNA-positive fibroblasts, as well as increased numbers of vimentin- and alpha-SMA-double-positive fibroblasts were detected in the dermis of IGFBP-5-overexpressing mouse skin. IGFBP-5 is a key mediator of fibrosis. IGFBP-5 mediates its profibrotic effects through fibroblast activation, increased ECM deposition, and myofibroblastic transformation of dermal fibroblasts. Overexpression of IGFBP-5 provides a novel model for studying the pathogenesis of skin fibrosis in systemic sclerosis.

Ultrasonic Backscatter and Attenuation (11–27 MHz) Variation with Collagen Fiber Distribution in Ex Vivo Human Dermis

This ex vivo study explores the relationship of ultrasonic attenuation and backscatter to dermal microarchitecture by comparing ultrasonic measurements of these parameters (11-27 MHz) to a microscopic analysis of three parameters describing the collagen distribution (mean thickness and spacing of collagen bundles along the insonification direction and the percent area occupied by collagen). Skin samples (N= 31) were obtained from patients undergoing breast or abdominal reduction surgery. Radio-frequency (rf) signals were acquired in a B-scan format using an ultrasound system developed for skin imaging (Ultrasons Technologies, Tours, France). Ultrasonic data were analyzed to calculate average integrated backscatter (IBS in dB) and frequency dependence of backscatter (n, dimensionless) of each specimen at depths centered approximately 370,620 and 880 microm beneath the skin surface. Average integrated attenuation coefficient (IA in dB.cm(-1)) and frequency dependence of attenuation coefficient (beta in dB.cm(-1).MHz(-1)) were estimated across the depth between 240 and 1,000 microm. The three collagen distribution parameters were estimated using digitized microcopic fields from matched regions of histological sections stained with hematoxylin-eosin-saffron. No significant correlation was identified between collagen distribution parameters and IA or beta. For the most superficial depth studied in abdominal skin, n was inversely correlated to collagen bundle thickness (r = -0.67,p = 0.002) and percent area (r = -0.65,p = 0.003). At the same depth, IBS was inversely correlated to percent area of collagen (r = -0.51,p = 0.03). The rather high collagen packing (48 to 82% area) measured in histological sections and the inverse relationship observed between IBS and percent area of collagen suggest that a packing factor should be included in models relating skin collagen distribution to ultrasound spectral parameters. A better understanding of the relationship between ultrasound parameters and the microarchitecture of the dermis should help to interpret changes in ultrasonic parameters observed during in vivo ultrasonic skin examinations.

Evaluation of histologic changes after microdermabrasion in a porcine model

Background There is a paucity of scientific information on the histologic changes associated with the clinical response to microdermabrasion. Objective In this study we sought to evaluate those changes in the skin after microdermabrasion, using a porcine model. Methods Using the left flank of an 8-week-old pig, we tattooed 4 areas with “A,” “B,” “C,” and “D.” Each of these areas was subdivided, in a gridlike fashion, from the dorsal to the ventral surface into 3 × 3-cm squares, which were numbered 1 through 8. Vacuum was initiated at 25 mm Hg the first week and increased by 5 mm Hg for each of 7 weeks to a high of 55 mm Hg in week 7. Biopsy specimens from each grid were obtained weekly. These samples were stained with hematoxylin and eosin and with the Verhoff/Van Geison (elastin) stain. Blinded histologic review of the biopsy specimens was performed by a dermatopathologist. Histometric analyses of the biopsy specimens were performed in which dermal thickness (millimeters), collagen-bundle thickness (microns), and epidermal thickness (microns) were all measured. Results Dermal thickness was increased by as much as 40% in the thinner skin and by 27% in the thicker skin. Similarly, the increase in collagen-bundle thickness was 22%, whereas the increase in the epidermal thickness was 9%. Conclusions The findings of this preliminary study appear to indicate that microdermabrasion produces real increases in dermal thickness, collagen-bundle thickness, and epidermal thickness. Histologically, the procedure produces changes similar to those seen after dermabrasion, laser resurfacing, and chemical peels.

The development of fibrocartilage in the elastic tendon of the chicken wing.

The elastic tendon of the chicken wing has five morphologically distinct regions. One of these regions is a distally located fibrocartilage from which fibrous connections extend to the capsule of the distal radius. In adult birds, this region shows the characteristics of a tendon-compressed fibrocartilage, with an accumulation of proteoglycans between thick collagen bundles arranged in a basket-weave formation. Here we study the development of this fibrocartilage in order to of compare it with other tendon fibrocartilages and try to identify the factors involved in fibrocartilage differentiation. This fibrocartilage initially developed by cell enlargement and accumulation of vimentin, with simultaneous deposition of proteoglycans in the extracellular matrix and an increase in the amount and thickness of collagen bundles. Elastic fibers were minor components associated with the collagen bundles. Cells could be classified into two main types. One was typically fibrocartilaginous and the other was fibroblast-like, the latter occurring in close association with the collagen bundles. These results establish the steps in the development of the elastic tendon fibrocartilage and provide a basis for future studies.

The fetlock tunnel syndrome: a macroscopic and microscopic study.

Chronic changes of several structures in around the fetlock tunnel can be a cause of the so-called fetlock tunnel syndrome (FTS) in the horse. Forty-nine annular ligaments (AL) from dead horses without a known history or clinical evidence of lameness and/or digital tendon sheath problems in these legs and 30 AL biopsies from horses suffering from FTS were studied macroscopically and microscopically. Macroscopically, the normal AL had a shiny white appearance, whereas the affected AL were often thicker and less white. Microscopically, the normal AL were about +/- 1 mm thick and were composed of undulating, parallel bundles of collagen. Small blood vessels with a diameter of 0.03-0.12 mm were found. The affected AL showed an increased thickness of collagen bundles, a changed direction of longitudinal axis of collagen bundles, and irregularly dispersed fibroblast nuclei. The number of blood vessels had increased, the external diameter of arteriolae could be up to 0.3 mm and arterial wall changes were observed. Possible relationships between the histological findings and the aetiology of the FTS are discussed.

Regional and age-related properties of human skin dermis relative to the thickness of collagen bundles

Regional and age-related properties of human skin dermis relative to the thickness of collagen bundles