Connective Tissue Proteins Research Articles

Comprehensive proteome analysis of nasal lavage samples after controlled exposure to welding nanoparticles shows an induced acute phase and a nuclear receptor, LXR/RXR, activation that influence the status of the extracellular matrix

Background Epidemiological studies have shown that many welders experience respiratory symptoms. During the welding process a large number of airborne nanosized particles are generated, which might be inhaled and deposited in the respiratory tract. Knowledge of the underlying mechanisms behind observed symptoms is still partly lacking, although inflammation is suggested to play a central role. The aim of this study was to investigate the effects of welding fume particle exposure on the proteome expression level in welders suffering from respiratory symptoms, and changes in protein mediators in nasal lavage samples were analyzed. Such mediators will be helpful to clarify the pathomechanisms behind welding fume particle-induced effects.MethodsIn an exposure chamber, 11 welders with work-related symptoms in the lower airways during the last month were exposed to mild-steel welding fume particles (1 mg/m3) and to filtered air, respectively, in a double-blind manner. Nasal lavage samples were collected before, immediately after, and the day after exposure. The proteins in the nasal lavage were analyzed with two different mass spectrometry approaches, label-free discovery shotgun LC–MS/MS and a targeted selected reaction monitoring LC–MS/MS analyzing 130 proteins and four in vivo peptide degradation products.ResultsThe analysis revealed 30 significantly changed proteins that were associated with two main pathways; activation of acute phase response signaling and activation of LXR/RXR, which is a nuclear receptor family involved in lipid signaling. Connective tissue proteins and proteins controlling the degradation of such tissues, including two different matrix metalloprotease proteins, MMP8 and MMP9, were among the significantly changed enzymes and were identified as important key players in the pathways.ConclusionExposure to mild-steel welding fume particles causes measurable changes on the proteome level in nasal lavage matrix in exposed welders, although no clinical symptoms were manifested. The results suggested that the exposure causes an immediate effect on the proteome level involving acute phase proteins and mediators regulating lipid signaling. Proteases involved in maintaining the balance between the formation and degradation of extracellular matrix proteins are important key proteins in the induced effects.

Type I Collagen from Jellyfish Catostylus mosaicus for Biomaterial Applications.

Collagen is the predominant protein in animal connective tissues and is widely used in tissue regeneration and other industrial applications. Marine organisms have gained interest as alternative, nonmammalian collagen sources for biomaterial applications because of potential medical and economic advantages. In this work, we present physicochemical and biofunctionality studies of acid solubilized collagen (ASC) from jellyfish Catostylus mosaicus (JASC), harvested from the Persian Gulf, compared with ASC from rat tail tendon (RASC), the industry-standard collagen used for biomedical research. From the protein subunit (alpha chain) pattern of JASC, we identified it as a type I collagen, and extensive molecular spectroscopic analyses showed similar triple helical molecular signatures for JASC and RASC. Atomic force microscopy of fibrillized JASC showed clear fibril reassembly upon pH neutralization though with different temperature and concentration dependence compared with RASC. Molecular (natively folded, nonfibrillized) JASC was shown to functionalize rigid substrates and promote MC3T3 preosteoblast cell attachment and proliferation better than RASC over 6 days. On blended collagen-agarose scaffolds, both RASC and JASC fibrils supported cell attachment and proliferation, and scaffolds with RASC fibrils showed more cell growth after 6 days compared with those scaffolds with JASC fibrils. These results demonstrate the potential for this new type I collagen as a possible alternative to mammalian type I collagen for biomaterial applications.

Live Imaging of Type I Collagen Assembly Dynamics in Osteoblasts Stably Expressing GFP and mCherry-Tagged Collagen Constructs.

Type I collagen is the most abundant extracellular matrix protein in bone and other connective tissues and plays key roles in normal and pathological bone formation as well as in connective tissue disorders and fibrosis. Although much is known about the collagen biosynthetic pathway and its regulatory steps, the mechanisms by which it is assembled extracellularly are less clear. We have generated GFPtpz and mCherry-tagged collagen fusion constructs for live imaging of type I collagen assembly by replacing the α2(I)-procollagen N-terminal propeptide with GFPtpz or mCherry. These novel imaging probes were stably transfected into MLO-A5 osteoblast-like cells and fibronectin-null mouse embryonic fibroblasts (FN-null-MEFs) and used for imaging type I collagen assembly dynamics and its dependence on fibronectin. Both fusion proteins co-precipitated with α1(I)-collagen and remained intracellular without ascorbate but were assembled into α1(I) collagen-containing extracellular fibrils in the presence of ascorbate. Immunogold-EM confirmed their ultrastuctural localization in banded collagen fibrils. Live cell imaging in stably transfected MLO-A5 cells revealed the highly dynamic nature of collagen assembly and showed that during assembly the fibril networks are continually stretched and contracted due to the underlying cell motion. We also observed that cell-generated forces can physically reshape the collagen fibrils. Using co-cultures of mCherry- and GFPtpz-collagen expressing cells, we show that multiple cells contribute collagen to form collagen fiber bundles. Immuno-EM further showed that individual collagen fibrils can receive contributions of collagen from more than one cell. Live cell imaging in FN-null-MEFs expressing GFPtpz-collagen showed that collagen assembly was both dependent upon and dynamically integrated with fibronectin assembly. These GFP-collagen fusion constructs provide a powerful tool for imaging collagen in living cells and have revealed novel and fundamental insights into the dynamic mechanisms for the extracellular assembly of collagen. © 2018 American Society for Bone and Mineral Research.

Physicochemical changes of pepsin-solubilized and insoluble collagen in jumbo squid (Dosidicus gigas) muscle after cooking process

ABSTRACTCollagen is the major connective tissue (CT) protein and one of the main constituents of the jumbo squid (Dosidicus gigas). Therefore, physicochemical changes of pepsin-solubilized collagen (PSC) and insoluble collagen (IC) were studied after cooking (100°C/30 min) of muscle (mantle, fins, and arms). Different pyridinoline (Pyr) contents (the major cross-linking molecule in collagen fibers) were found in the fresh muscle of the three anatomical regions. After the cooking process, a decrease from 10 to 30% in the thermal resistance of collagen was observed, depending on the anatomical region and fraction evaluated. Furthermore, the electrophoretic profile, Fourier transform infrared (FTIR) spectroscopy, and the amino-acid profile revealed that structural changes occurred in the two different collagen fractions caused by the thermal process, and the changes were greater in the mantle. Under the conditions applied in this study, collagen fractions from the squid arms showed more stability during the cooking process due to the high cross-linking degree of their fibers.

Decorin, a growth hormone-regulated protein in humans.

Growth hormone (GH) stimulates connective tissue and muscle growth, an effect that is potentiated by testosterone. Decorin, a myokine and a connective tissue protein, stimulates connective tissue accretion and muscle hypertrophy. Whether GH and testosterone regulate decorin in humans is not known. To determine whether decorin is stimulated by GH and testosterone. Randomized, placebo-controlled, double-blind study. 96 recreationally trained athletes (63 men, 33 women) received 8 weeks of treatment followed by a 6-week washout period. Men received placebo, GH (2 mg/day), testosterone (250 mg/week) or combination. Women received either placebo or GH (2 mg/day). Serum decorin concentration. GH treatment significantly increased mean serum decorin concentration by 12.7 ± 4.2%; P < 0.01. There was a gender difference in the decorin response to GH, with greater increase in men than in women (∆ 16.5 ± 5.3%; P < 0.05 compared to ∆ 9.4 ± 6.5%; P = 0.16). Testosterone did not significantly change serum decorin. Combined GH and testosterone treatment increased mean decorin concentration by 19.5 ± 3.7% (P < 0.05), a change not significantly different from GH alone. GH significantly increases circulating decorin, an effect greater in men than in women. Decorin is not affected by testosterone. We conclude that GH positively regulates decorin in humans in a gender-dimorphic manner.

Existence of life-time stable proteins in mature rats—Dating of proteins’ age by repeated short-term exposure to labeled amino acids throughout age

In vivo turnover rates of proteins covering the processes of protein synthesis and breakdown rates have been measured in many tissues and protein pools using various techniques. Connective tissue and collagen protein turnover is of specific interest since existing results are rather diverging. The aim of this study is to investigate whether we can verify the presence of protein pools within the same tissue with very distinct turnover rates over the life-span of rats with special focus on connective tissue. Male and female Lewis rats (n = 35) were injected with five different isotopically labeled amino acids tracers. The tracers were injected during fetal development (Day -10 to -2), after birth (Day 5–9), at weaning (Day 25–32) at puberty (Day 54–58) and at adulthood (Day 447–445). Subgroups of rats were euthanized three days after every injection period, at different time point between injection periods and lastly at day 472. Tissue (liver, muscle, eye lens and patellar tendon) and blood samples were collected after euthanization. The enrichment of the labeled amino acids in the tissue or blood samples was measured using GC-MS-MS. In muscle and liver we demonstrated a rapid decrease of tracer enrichments throughout the rat’s life, indicating that myofibrillar and cytoskeleton proteins have a high turnover. In contrast, the connective tissue protein in the eye lens and patellar tendon of the mature rat showed detainment of tracer enrichment injected during fetal development and first living days, indicating very slow turnover. The data support the hypothesis that some proteins synthesized during the early development and growth still exist much later in life of animals and hence has a very slow turnover rate.

Evaluation of Therapeutic Tissue Crosslinking (TXL) for Myopia Using Second Harmonic Generation Signal Microscopy in Rabbit Sclera.

PurposeSecond harmonic generation signals (SHG) are emitted preferentially from collagenous tissue structures and have been used to evaluate photochemically-induced (CXL) crosslinking changes in the cornea. Since therapeutic tissue crosslinking (TXL) using sodium hydroxymethylglycinate (SMG) of the sclera is a potential treatment for high myopia, we explored the use of SHG microscopy to evaluate the effects.MethodsSingle sub-Tenon's (sT) injections (400 μL) using SMG (40–400 mM) were made at the equatorial 12 o'clock position of the right eye of cadaveric rabbit heads (n = 16 pairs). After 3.5 hours, confocal microscopy (CM) was performed using 860 nm two-photon excitation and 400 to 450 nm emission. Pixel density and fiber bundle “waviness” analyses were performed on the images. Crosslinking effects were confirmed using thermal denaturation (Tm) temperature. Comparison experiments with riboflavin photochemical crosslinking were done.ResultsTherapeutic tissue crosslinking localization studies indicated that crosslinking changes occurred at the site of injection and in adjacent sectors. Second harmonic generation signals revealed large fibrous collagenous bundled structures that displayed various degrees of waviness. Histogram analysis showed a nearly 6-fold signal increase in 400 mM SMG over 40 mM. This corresponded to a ΔTm = 13°C for 400 mM versus ΔTm = 4°C for 40 mM. Waviness analysis indicated increased fiber straightening as a result of SMG CXL.ConclusionsSecond harmonic generation signal intensity and fiber bundle waviness is altered by scleral tissue crosslinking using SMG. These changes provide insights into the macromolecular changes that are induced by therapeutic crosslinking technology and may provide a method to evaluate connective tissue protein changes induced by scleral crosslinking therapies.

Broadband diffuse optical characterization of elastin for biomedical applications

Elastin is a key structural protein of dynamic connective tissues widely found in the extracellular matrix of skin, arteries, lungs and ligaments. It is responsible for a range of diseases related to aging of biological tissues. The optical characterization of elastin can open new opportunities for its investigation in biomedical studies. In this work, we present the absorption spectra of elastin using a broadband (550–1350nm) diffuse optical spectrometer. Distortions caused by fluorescence and finite bandwidth of the laser source on estimated absorption were effectively accounted for in measurements and data analysis and compensated. A comprehensive summary and comparison between collagen and elastin is presented, highlighting distinct features for its accurate quantification in biological applications.

ОСОБЛИВОСТІ ОБМІНУ ВУГЛЕВОДНО-БІЛКОВИХ КОМПОНЕНТІВ СПОЛУЧНОЇ ТКАНИНИ ПАРОДОНТА ТА ЇХ ЦИТОКІНОВОЇ РЕГУЛЯЦІЇ У ХВОРИХ НА ВИРАЗКОВУ ХВОРОБУ ДВАНАДЦЯТИПАЛОЇ КИШКИ ЗАЛЕЖНО ВІД ВИДУ НЕЗНІМНИХ ЗУБНИХ ПРОТЕЗІВ

Мета роботи – встановити вплив незнімних металевих, металокерамічних та керамічних протезів на стан білково-вуглеводних компонентів сполучної тканини пародонта та стан їх цитокінової регуляції у крові хворих на виразкову хворобу (ВХ) дванадцятипалої кишки (ДПК).Матеріали і методи. Обстежено 147 хворих на ВХ ДПК у фазі рубцювання виразки, у тому числі 43 особи із дефектами зубного ряду без зубних протезів (1 група), 35 осіб із незнімними металевими зубними протезами (2 група), 42 – із незнімними металокерамічними протезами (3 група), 27 – із незнімними керамічними протезами (4 група). Використовували клінічні, біохімічні та імуноферментні методи дослідження.Результати досліджень. У хворих на ВХ ДПК без протезів зубного ряду виявлено істотні зміни метаболізму компонентів позаклітинного матриксу (ПКМ) пародонта: вірогідне зниження інтенсивності синтезу колагену, глікозаміногліканів, глікопротеїнів, фібронектину на тлі підвищення інтенсивності процесів колагенолізу та деградації фукоглікопротеїнів. Наявність металовмісних незнімних зубних протезів призвела до істотного поглиблення даного дисбалансу внаслідок підвищення активності колагеназ та еластази ПКМ, гіперпродукції прозапальних цитокінів (TNF-α, IL-1) та недостатності вмісту в крові факторів росту (TGF-β1) анаболічної дії, які у сильній взаємозалежності корелюють із інтенсивністю реакцій ана- та катаболізму колагену, еластину, вмістом у крові глікозаміногліканів, фукоглікопротеїнів, фібронектину.Висновки. Наявність металовмісних незнімних зубних протезів у хворих на ВХ ДПК сприяє зниженню синтезу колагену та підвищенню колагенолізу внаслідок зростання активності колагеназ та еластази ПКМ, що супроводжується вірогідно вищою гіперпродукцією прозапальних цитокінів (TNF-α, IL-1) на тлі недостатності вмісту в крові факторів росту (TGF-β1), які істотно перевищують показники хворих на ВХ без зубних протезів та за наявності незнімних керамічних протезів.

Dietary nitrite supplementation attenuates cardiac remodeling in l-NAME-induced hypertensive rats

Loss of nitric oxide (NO) bioavailability underlies the development of hypertensive heart disease. We investigated the effects of dietary nitrite on NG-nitro-l-arginine methyl ester (l-NAME)-induced hypertension. Sprague-Dawley rats were divided into five groups: an untreated control group, an l-NAME-treated group, and three other l-NAME-treated groups supplemented with 10 mg/L or 100 mg/L of nitrite or 100 mg/L of captopril in drinking water. After the 8-week experimental period, mean arterial blood pressure was measured, followed by sampling of blood and heart tissue for assessment of nitrite/nitrate levels in the plasma and heart, the plasma level of angiotensin II (AT II), and the heart transcriptional levels of AT II type 1 receptor (AT1R), transforming growth factor-β1 (TGF-β1), and connective tissue proteins such as type 1 collagen and fibronectin. Heart tissue was analyzed by histopathological morphometry, including assessments of ventricular and coronary vascular hypertrophy and fibrosis, as well as immunohistochemistry analyses of myocardial expression of AT1R. l-NAME treatment reduced the plasma nitrate level and led to the development of hypertension, with increased plasma levels of AT II and increased heart transcriptional levels of AT1R and TGF-β1-mediated connective tissue proteins, showing myocardial and coronary arteriolar hypertrophy and fibrosis. However, dietary nitrite supplementation inhibited TGF-β1-mediated cardiac remodeling by suppressing AT II and AT1R. These results suggest that dietary nitrite levels achievable via a daily high-vegetable diet could improve hypertensive heart disease by inhibiting AT II-AT1R-mediated cardiac remodeling.

Gelatin- and starch-based hydrogels. Part B: In vitro mesenchymal stem cell behavior on the hydrogels

Tissue regeneration often occurs only to a limited extent. By providing a three-dimensional matrix serving as a surrogate extracellular matrix that promotes adult stem cell adhesion, proliferation and differentiation, scaffold-guided tissue regeneration aims at overcoming this limitation. In this study, we applied hydrogels made from crosslinkable gelatin, the hydrolyzed form of collagen, and functionalized starch which were characterized in depth and optimized as described in Van Nieuwenhove et al., 2016. “Gelatin- and Starch-Based Hydrogels. Part A: Hydrogel Development, Characterization and Coating”, Carbohydrate Polymers 152:129–39. Collagen is the main structural protein in animal connective tissue and the most abundant protein in mammals. Starch is a carbohydrate consisting of a mixture of amylose and amylopectin. Hydrogels were developed with varying chemical composition (ratio of starch to gelatin applied) and different degrees of methacrylation of the applied gelatin phase. The hydrogels used exhibited no adverse effect on viability of the stem cells cultured on them. Moreover, initial cell adhesion did not differ significantly between them, while the strongest proliferation was observed on the hydrogel with the highest degree of cross-linking. On the least crosslinked and thus most flexible hydrogels, the highest degree of adipogenic differentiation was found, while osteogenic differentiation was the strongest on the most rigid, starch-blended hydrogels. Hydrogel coating with extracellular matrix compounds aggrecan or fibronectin prior to cell seeding exhibited no significant effects. Thus, gelatin-based hydrogels can be optimized regarding maximum promotion of either adipogenic or osteogenic stem cell differentiation in vitro, which makes them promising candidates for in vivo evaluation in clinical studies aiming at either soft or hard tissue regeneration.

The Impact of Replacing Backfat With Microcrystalline Cellulose Gel on Physico-Chemical and Sensory Quality of Frankfurter

The aim of this study was to evaluate the effect of different concentrations of dietary fiber Vitacel MCG 0018 Gel (consisting of microcrystalline cellulose - MCC and two different qualities of carboxymethyl cellulose - CMC) as back fat substitute on the physico-chemical and sensory quality of frankfurter. Five production series were evaluated to examine the back fat replacement: the control group (C) - a sample with just the addition of back fat (100% fat - 0% Vitacel MCG 0018 Gel); 75% fat and 25% gel (E1); 50% fat and 50% gel (E2); 25% fat and 75% gel (E3); 0% fat and 100% gel (E4). In our research, the moisture content increased (59.69% - 71.07%) and the fat content decreased (23.48% - 9.99%) with increasing of back fat replacement by Vitacel MCG 0018 gel. The protein content was higher than 10% (10.18% - 10.35%) and relative content of connective tissue proteins was lower than 20% (6.33 – 17.43) in all examined samples which is in accordance with the criteria of the Serbian Regulation.Sensory evaluation of color, smell and taste and texture and juiciness are less assessed in frankfurters where the fat is replaced by the Vitacel MCG 0018 gel in the proportion of more than half. In general, it can be concluded that the replacement of up to 50% back fat with cellulose fiber gives sausages almost unchanged properties, while the addition of larger fiber amount leads to decrease of sausage quality.

Some parameters of nutritional quality of meat obtained from Mangalitsa and Landrace pig breeds

The aim of this study was to provide a comparative analysis of chemical and fatty acid composition, as well as of the connective tissue proteins in pigs of different genotypes, Mangalitsa and Landrace. Both pig genotypes were fed with the same feed of standard composition and quality. At the end of the fattening period, in total 24 pigs of both genotypes were slaughtered. Based on the analysis of the chemical composition we came to the conclusion that the protein content in both genotypes was similar. Moisture and ash content in the Landrace pig genotype differed significantly (P &lt; 0.01) compared with genotype of Mangalitsa breed. Statistically significant differences (P &lt; 0.01) were established in the fat content, which was 7.95 g/100 g, in pigs of Mangalitsa breed and 1.59 g/100 g in the Landrace pigs breed. Content of hydroxyproline, non-proteinogenic amino acids, in meat of Landrace was significantly higher (P &lt; 0.01) compared to the content in the Mangalitsa breed. The same tendency was observed with regard to the connective tissue protein content, as well as with the relative connective tissue protein content. The fatty acid composition of the meat indicated that the most common saturated fatty acid (SFA) in both tested breeds was palmitic fatty acid (C16), whose content was significantly higher in Landrace (P &lt; 0.01) compared with its content in Mangalitsa breed. In addition, the share of stearic acid (C18) was significantly higher (P &lt; 0.01) in Landrace compared to Mangalitsa pig breed, what significantly contributed to the increase of the SFA share in Landrace compared to Mangalitsa breed. The most common monounsaturated fatty acid in both pig breeds was the oleic fatty acid (C18:1), whose share was significantly higher in Mangalitsa compared to the Landrace breed (P &lt; 0.01). Out of the polyunsaturated fatty acids (PUFA), linoleic fatty acid (C18:2) was the most predominant in both pig breeds, with no statistically significant differences (P &gt; 0.05). The content of PUFA was not statistically significantly different between the tested breeds, as well as the content of n-3 and n-6 fatty acids, which caused no statistically significant differences in the n-6/n-3 PUFA ratio. Ratio of the unsaturated fatty acids, i.e., of the sum of MUFA and PUFA, and of the saturated fatty acids was significantly higher in Mangalitsa compared to Landrace breed (1.86 vs. 1.4), and the same was observed when it comes to the relationship MUFA/SFA (1.51 in Mangalitsa vs. 1.08 in Landrace breed) and MUFA/PUFA (4.35 vs. 3.38).

Visualisation of Collagen in fixed skeletal muscle tissue using fluorescently tagged Collagen binding protein CNA35

Detection and visualisation of Collagen structure are important to understand the relationship between mechanical behaviour and microstructure in skeletal muscle since Collagen is the main structural protein in animal connective tissues, and is primarily responsible for their passive load-bearing properties.In the current study, the direct detection and visualization of Collagen using fluorescently tagged CNA35 binding protein (fused to EGFP or tdTomato) is reported for the first time on fixed skeletal muscle tissue. This Technical Note also establishes a working protocol by examining tissue preparation, dilution factor, exposure time etc. for sensitivity and specificity. Penetration of the binding protein into intact mature skeletal muscle was found to be very limited, but detection works well on tissue sections with higher sensitivity on wax embedded sections compared to frozen sections. CNA35 fused to tdTomato has a higher sensitivity than CNA35 fused to EGFP but both show specific detection. Best results were obtained with 15μm wax embedded sections, with blocking of non-specific binding in 1% BSA and antigen retrieval in Sodium Citrate. There was a play-off between dilution of the binding protein and time of incubation but both CNA35-tdTomato and CNA35-EGFP worked well with approximately 100μg/ml of purified protein with overnight incubation, while CNA35-tdTomato could be utilized at 5 fold less concentration.This approach can be applied to study the relationship between skeletal muscle micro-structure and to observe mechanical response to applied deformation. It can be used more broadly to detect Collagen in a variety of fixed tissues, useful for structure-functions studies, constitutive modelling, tissue engineering and assessment of muscle tissue pathologies.

Contraction mode and whey protein intake affect the synthesis rate of intramuscular connective tissue.

In this study we investigated the impact of whey protein hydrolysate and maltodextrin (WPH) intake on intramuscular connective tissue (IMCT) protein fractional synthesis rate (FSR) after maximal shortening and lengthening contractions. Twenty young men were randomized to receive either WPH or maltodextrin [carbohydrate (CHO)] immediately after completion of unilateral shortening and lengthening knee extensions. Ring-13 C6 -phenylalanine was infused, and muscle biopsies were obtained. IMCT protein FSR was measured at 1-5, as well as 1-3 and 3-5 hours after contractions and nutrient intake. During the 1-3-hour recovery, lengthening contractions resulted in a higher FSR than shortening contractions (P < 0.01), independent of supplementation type and, during the 3-5-hour recovery, WPH had a higher FSR than CHO (P < 0.05), independent of prior contraction mode. The later appearance of a stimulating effect of WPH on the IMCT FSR after strenuous muscle contractions lends support to its ability to promote recovery of the muscle connective tissue matrix after exercise. Muscle Nerve 55: 128-130, 2017.

Biomedical Significance of Alpha1-Antitrypsin: An Overview

Alpha1-antitrypsin (α1-AT) is a natural, endogenous inhibitor of serine proteases with preferential activity towards neutrophil elastase (NE), the enzyme that possess the ability to cleave important connective tissue protein, elastin. The normal circulating concentrations of α1-AT is thus very important to maintain the elasticity of tissues. Severely diminished levels of α1-AT are associated with development of various disease conditions, in particular disorders of lung. This review highlights the biological role of α1-AT and its involvement in diseases. New therapeutic approaches involving α1-AT are also discussed. Keywords: Alpha1-antitrypsin, Endogenous inhibitor, Neutrophil elastase, Serine protease, Serpin.

Chemical, Color, Texture and Sensory Properties of Čajna Kobasica, a Dry Fermented Sausage

The aim of this study was to conduct a comprehensive examination of quality parameters of dry fermented sausage (Čajna kobasica) from six different producers from the market in Vojvodina. The investigated parameters were chemical composition (moisture, total protein, relative content of connective tissue proteins RCCTP, free fat, ash, NaCl and nitrite content), pH value, aw value, instrumental color parameters (lightness L*, redness a* and yellowness b*), instrumental texture parameters (hardness, springiness, cohesiveness and chewiness) and sensory properties (external appearance and/or condition of the packaging, appearance and composition of cut surface, color and stability of color, odour and taste and texture and/ or juiciness). It could be concluded that out of six examined brands of dry fermented sausage (Čajna kobasica), two brands did not meet the requirements prescribed under the Regulations, because of a higher level of RCCTP. The lowest L* value was recorded in brand 4 (44.33). The brand 4 had the highest hardness and chewiness (22883.33g and 3857.58g, respectively), and the lowest springiness and cohesiveness (0.42 and 0.40, respectively). Parameters related to sensory properties showed significant variability (p &lt; 0.05). Overall sensory quality of tested samples ranged from 3.53 (brand 1) to 4.18 (brand 4).

Physicochemical Changes of Connective Tissue Proteins in Jumbo Squid (Dosidicus gigas) Muscle During Ice Storage

Connective tissue (CT) proteins in jumbo squid (Dosidicus gigas) play an important role because they are the responsible for the union between various cells; hence, there is a close relationship between their functions and muscle firmness during ice storage. In this study, the thermal resistance and solubility of the CT extracted from the fins, mantle and arms of jumbo squid during ice storage (20 days) was evaluated. The CT was fractionated based on solubility [NaCl-soluble (SSCT) and insoluble (ICT)]. The solubility of the CT was affected during ice storage. An increase in the thermal resistance of the SSCT after 10 days with a subsequent decrease was found in the mantle. Furthermore, the ICT was thermally more resistant than the SSCT in all the anatomical regions. Finally, the electrophoretic profile revealed that structural changes occurred, causing changes in its solubility, due to an increase in the thermal resistance. Practical Applications Jumbo squid (Dosidicus gigas) is an important seafood resource in Mexico, which is commercialized primarily iced and fresh-frozen. Its muscle connective tissue (CT) has different physicochemical characteristics that differ from other seafood products; it is thermally more resistant and might be responsible for the textural changes during ice storage. Moreover, specific studies about the behavior of CT proteins in such organism may result in improved post-catch management of jumbo squid muscle, focused on the processing of the species in the food industry or anywhere that CT proteins are used as raw material. The main objective of this research is to investigate solubility changes and thermal resistance of CT proteins of iced jumbo squid mantle.

Cloning, expression and antioxidant activity of a novel collagen from Pelodiscus sinensis.

Collagen is the main structural protein of various connective tissues in animals and naturally plays an important role within the body. It is increasingly used within certain areas, such as medicine, citology and cosmetology. The soft-shelled turtle (Pelodiscus sinensis) is a commercially important aquatic species rich in collagen. In this study, a novel collagen gene fragment of 756bp, which encodes 252 deduced amino acid residues, including 25 conserved Gly-X-Y motifs, was cloned from a soft-shelled turtle. Recombinant soft-shelled turtle collagen (rSTC) was stably expressed in Escherichia coli Rosetta and purified by His GraviTrap affinity columns. The antioxidant activities of rSTC were measured using hydroxyl and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals. The results showed that rSTC quenched the free radicals in a dose-dependent manner. The hydroxyl radical scavenging activity (HRSA) of rSTC was 98.9% at a concentration of 3mg/mL. At a concentration of 5mg/mL, rSTC exhibited a DPPH radical scavenging activity of 32.7%. At the tested concentrations, rSTC exhibited higher HRSA and lower DPPH radical scavenging activity.

The leucine-rich repeat protein PRELP binds fibroblast cell-surface proteoglycans and enhances focal adhesion formation.

PRELP (proline/arginine-rich end leucine-rich repeat protein) is a member of the leucine-rich repeat (LRR) family of extracellular matrix proteins in connective tissue. In contrast with other members of the family, the N-terminal domain of PRELP has a high content of proline and positively charged amino acids. This domain has previously been shown to bind chondrocytes and to inhibit osteoclast differentiation. In the present study, we show that PRELP mediates cell adhesion by binding to cell-surface glycosaminoglycans (GAGs). Thus, rat skin fibroblasts (RSFs) bound to full-length PRELP and to the N-terminal part of PRELP alone, but not to truncated PRELP lacking the positively charged N-terminal region. Cell attachment to PRELP was inhibited by addition of soluble heparin or heparan sulfate (HS), by blocking sulfation of the fibroblasts or by treating the cells with a combination of chondroitinase and heparinase. Using affinity chromatography, we identified syndecan-1, syndecan-4 and glypican-1 as cell-surface proteoglycans (PGs) binding to the N-terminal part of PRELP. Finally, we show that the N-terminal domain of PRELP in combination with the integrin-binding domain of fibronectin, but neither of the fragments alone, induced fibroblast focal adhesion formation. These findings provide support for a role of the N-terminal region of PRELP as an important regulator of cell adhesion and behaviour, which may be of importance in pathological conditions.