Changes to the Collagen Structure using Vibrational Spectroscopy and Chemometrics: A Comparison between Chemical and Sulfide-Free Leather Process

Chemical and physical changes take place when hides and skins are processed to leather that affect the quality and strength of the material. Understanding the structure at each leather-making stage is the basis of this study but also intend to improve the process through a biochemical approach, employing a proteolytic enzyme for processing leather more cleanly with reduced environmental impact. Raman and ATR-FTIR spectroscopy in conjunction with chemometrics was used to investigate each leather-making stage from fresh green cattle hide to dry crust leather. The changes in proteins, lipids, nucleic acids and other biomolecules with leather processing was measured and reported using three novel Raman ratiometric markers, 920/1476, 1345/1259 and 1605/1476 cm-1, to discriminate the structural changes in collagen of hide using standard chemical and enzymatic method. Amide I band was deconvoluted to investigate thecollagen secondary structures using curve fitting by Gaussians function. The results of Principal Component Analysis are well-corroborated with the ratiometric markers of structural changes.

Evaluation of Gene Polymorphism and Gingival Crevicular Fluid Levels of Matrix Metalloproteinase-3 in a Group of Turkish Periodontitis Patients

Introduction: Periodontitis is characterized by the destruction of tooth-supporting tissues. Matrix metalloproteinases (MMPs) play a significant part in the degradation of collagen structure. The gingival crevicular fluid (GCF) levels of MMPs increase with the progression of periodontal inflammation. Polymorphisms can be responsible for high expression of MMPs and can exacerbate the breakdown of collagen structure. This study aims to investigate the effect of MMP-3 -1171 5A/6A polymorphism and the GCF levels of MMP-3 in a group of Turkish periodontitis patients. Materials and Methods: Non-smoking, stage II grade A periodontitis (S II-Gr A) (n = 68) and stage II grade B periodontitis (S II-Gr C) (n = 64) patients were recruited. Healthy individuals (H) (n = 72) without signs of gingivitis or periodontitis served as the control. Venous blood was collected from participants to obtain DNA, and the polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) method was used to detect polymorphism. GCF samples were taken to assess MMP-3 levels using an enzyme-linked immunosorbent assay (ELISA). Results: The MMP-3 -1179 5A/6A distribution showed no significant difference between the groups (p > 0.05). However, the MMP-3 GCF levels of the S II-Gr C group were higher than those of both the S II-Gr A and H groups (p < 0.05), and elevated MMP-3 levels were detected in S II-Gr A compared to H (p < 0.05). Conclusion: The MMP-3 GCF levels showed an association with periodontal tissue destruction, although single nucleotide polymorphism was not associated with the S II-Gr C and S II-Gr A groups in the Turkish population.

Abstract P377: Collagen Remodeling Of Spontaneously Hypertensive Rats Undergoing Quinapril Treatment Measured By Three Dimensional Shape Analysis

Introduction: Myocardial collagen structure is a complex three dimensional (3D) hierarchy, organizing cardiomyocytes and providing structural integrity. Changes in collagen organization have a crucial role in cardiac remodeling. However, quantitative methods for the analysis of collagen structures are still lacking. Herein, we apply our collagen shape analysis framework to compare within-sheetlet collagen (both endomysial collagen surrounding myocytes and perimysial cords) against sheetlet-surface collagen (perimysial meshwork collagen deposited between sheetlets) in spontaneously hypertensive rats undergoing quinapril treatment. Hypothesis: The different functional properties of endomysial and perimysial meshwork collagen are linked to their structure and will have markedly different features quantifiable by 3D shape analysis. Methods: Rat myocardium was stained for collagen (picrosirius red), and the left ventricular free wall imaged using extended volume confocal microscopy (0.4 μm 3 voxel size). The 3D image volumes were segmented into within-sheetlet and sheetlet-surface regions. Shape analysis was performed using eigenanalysis of the inertia matrix to derive 3D morphological (elongation and flatness) parameters, characterizing the collagen shape in the two regions. Results: Shape analysis revealed marked differences between within-sheetlet and sheetlet-surface collagen organization. Long-term quinapril treatment resulted in notable differences in within-sheetlet collagen shape in rats at 14 months of age (mo) compared with 24 mo. At 14 mo, the within-sheetlet collagen is a mix of sheet-, ribbon-, and rod-like structures (elongation: 0.43, flatness: 0.05), but by 24 mo this has remodeled to predominately thin rod-like structures (elongation: 0.03, flatness: 0.35). The sheetlet-surface collagen consisted of thick sheet-like structures in both 14 and 24 mo rats (elongation: 0.83 vs 0.79, flatness: 0.13 vs 0.09). Conclusion: Shape analysis in 3D showed marked differences between within-sheetlet and sheetlet-surface collagen structure, as well as differences between early treatment and late treatment of diseased hearts.

A multiscale four-layer finite element model to predict the effects of collagen fibers on skin behavior under tension

Human skin is a complex multilayered multiscale material that exhibits nonlinear and anisotropic mechanical behavior. It has been reported that its macroscopic behavior in terms of progression of wrinkles induced by aging is strongly dependent on its microscopic composition in terms of collagen fibers in the dermis layer. In the present work, a multiscale four-layer 2D finite element model of the skin was developed and implemented in Matlab code. The focus here was to investigate the effects of dermal collagen on the macroscopic mechanical behavior of the skin. The skin was modeled by a continuum model composed of four layers: the Stratum Corneum, the epidermis, the dermis, and the hypodermis. The geometry of the different layers of the skin was represented in a 2D model with their respective thicknesses and material properties taken from literature data. The macroscopic behavior of the dermis was modeled with a nonlinear multiscale approach based on a multiscale elastic model of collagen structure going from cross-linked molecules to the collagen fiber, combined with a Mori-Tanaka homogenization scheme. The model includes the nonlinear elasticity of the collagen fiber density, the fiber radius, the undulation, and the fiber orientation. An axial tension was applied incrementally to the lateral surfaces of the skin model. A parametric study was performed in order to investigate the effect of the collagen constituents on the macroscopic skin mechanical behavior in terms of the predicted macroscopic stress-strain curve of the skin. The results of the FE computations under uniaxial tension showed that the different layers undergo different strains, leading to a difference in the transversal deformation at the top surface. In addition, the parametric study revealed a strong correlation between macroscopic skin elasticity and its collagen structure.

FORMATION OF LEATHER QUALITY INDICATORS OF DIFFERENT TYPES OF LEATHER RAW MATERIALS WITH THE USE OF MONTMORILONITE

The tanning process is important in stabilizing the collagen structure of the derma. Сhrome tanning is the traditional option. The basic chromium sulfate is used in the production of 80% of leather in use. A peculiarity of chromium tanning turns out to be an inefficient use of chromium compounds, since almost 40% of tannins remain in wastewater. Considering the disadvantages of using chromium compounds, scientific research on the replacement and use of rather eco-friendlier substances has been conducted for a long time. One of the directions of ecologization of leather manufacturing is the use of modified montmorillonite dispersions in various technological processes such as tanning, filling, dyeing, fat-liquoring, etc. Researches have been aimed at identifying the feasibility of making leather for the uppers of children’s shoes from different types of leather raw materials using modified dispersions of montmorillonite at the stage of tanning. According to the technology of making leather, cattle (medium heifer), goat and sheep skins were used for the upper of the shoes. The analysis of leather quality indicators from different types of raw materials obtained with the use of chromefree tanning. It is proved that hides from raw materials of goats and sheepskins in comparison with cattle are characterized by increased yield area by 3.5-7.2 %, thickness by 0.8-2.6 %, volume yield by 4.5-6.1 % and uniformity of quality indicators in different topographic areas. The level of indicators of physical and mechanical, hygienic properties and chemical composition of leather of different types of raw materials is almost identical within the requirements of regulatory documents. It is advisable to use for children’s shoes leather from raw goat and sheepskin given the peculiarities of the manufacture of children’s shoes, dimensional characteristics of parts, requirements for dimensional stability and shape conservation.

Mechanical Properties of Porcine and Fish Skin-Based Collagen and Conjugated Collagen Fibers

Collagen is a protein that is a major component of animal skins and tendons. It is used in various medical, cosmetic, and food products through extraction and purification. The fibrous products of purified collagen fibers extracted from raw mammal materials have relatively excellent mechanical properties and are used for high-end medical products. In this study, we examined collagen materials produced from porcine and fish skins, which are major sources of collagen raw materials. We examined a method for spinning collagen fibers from fish skin-based collagen and analyzed the physical properties of those collagen fibers. In addition, we examined the characteristics and advantages of conjugated fibers according to their porcine- and/or fish skin-based compositions. The spinnability and mechanical properties of these conjugated fibers were analyzed according to their compositions. The mechanical properties of collagen structure are determined by hydroxyproline content and can be manipulated by the composition of collagen in the conjugated fibers.

Impact of Porcine Pancreas Decellularization Conditions on the Quality of Obtained dECM

Due to the limited number of organ donors, 3D printing of organs is a promising technique. Tissue engineering is increasingly using xenogeneic material for this purpose. This study was aimed at assessing the safety of decellularized porcine pancreas, together with the analysis of the risk of an undesirable immune response. We tested eight variants of the decellularization process. We determined the following impacts: rinsing agents (PBS/NH3·H2O), temperature conditions (4 °C/24 °C), and the grinding method of native material (ground/cut). To assess the quality of the extracellular matrix after the completed decellularization process, analyses of the following were performed: DNA concentration, fat content, microscopic evaluation, proteolysis, material cytotoxicity, and most importantly, the Triton X-100 content. Our analyses showed that we obtained a product with an extremely low detergent content with negligible residual DNA content. The obtained results confirmed the performed histological and immuno-fluorescence staining. Moreover, the TEM microscopic analysis proved that the correct collagen structure was preserved after the decellularization process. Based on the obtained results, we chose the most favorable variant in terms of quality and biology. The method we chose is an effective and safe method that gives a chance for the development of transplant and regenerative medicine.