Search results for: collagen like peptide

Authors: A. Bajek, J. Skopinska-Wisniewska, A. Rynkiewicz, A. Jundzill, M. Bodnar, A. Marszalek, T. Drewa

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Traumatic injury and age-related degenerative diseases associated with cartilage are major health problems worldwide. The articular cartilage is comprised of a relatively small number of cells, which have a relatively slow rate of turnover. Therefore, damaged articular cartilage has a limited capacity for self-repair. New clinical methods have been designed to achieve better repair of injured cartilage. However, there is no treatment that enables full restoration of it. The aim of this study was to evaluate how collagen gel with bone marrow mesenchymal stem cells (MSCs) and collagen gel alone will influence on the hip cartilage repair after injury. Collagen type I was isolated from rats’ tails and cross-linked with N-hydroxysuccinimide in 24-hour process. MSCs were isolated from rats’ bone marrow. The experiments were conducted according to the guidelines for animal experiments of Ethics Committee. Fifteen 8-week-old Wistar rats were used in this study. All animals received hip joint surgery with a total of 30 created cartilage defects. Then, animals were randomly divided into three groups and filled, respectively, with collagen gel (group 1), collagen gel cultured with MSCs (group II) or left untreated as a control (control group). Immunohistochemy and radiological evaluation was carried out 11 weeks post implantation. It has been proved that the surface of the matrix is non-toxic, and its porosity promotes cell adhesion and growth. However, the in vivo regeneration process was poor. We observed the low integration rate of biomaterial. Immunohistochemical evaluation of cartilage after 11 weeks of treatment showed low II and high X collagen expression in two tested groups in comparison to the control one, in which we observed the high II collagen expression. What is more, after radiological analysis, we observed the best regeneration process in control group. The biomaterial construct and mesenchymal stem cells, as well as the use of the biomaterial itself was not sufficient to regenerate the hip cartilage surfaces. These results suggest that the collagen gel based biomaterials, even with MSCs, are not satisfactory in repar of hip cartilage defect. However, additional evaluation is needed to confirm these results.

Keywords: collafen gel, MSCs, cartilage repair, hip cartilage

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Authors: Chin-Tsu Ma, Yi-Jhen Wu, Hsia Ying Cheng, Han Hsiang Huang, Shyh Ming Kuo

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The use of specific molecules including nutrients and pharmacological agents has been tried in modulation of stem cells differentiation (MSCs) to insulin producing cells. The aim of this study is to investigate the ability of nano collagen molecules (nutrient or scaffold) to enhance the MSCs differentiation into insulin-producing cells in combination with nicotinamide and exendin-4 (pharmacological agents) in vitro and in vivo. The results demonstrated that the cells exhibit morphologically islet-like clusters after treatment with nano collagen molecules, nicotinamide and exendin-4. MSCs extra treated with nano collagen molecules showed significant increases in Nkx6.1 and insulin mRNA expression at 14-d and 21-d culture compared with those merely treated with nicotinamide and exendin-4. Early 7-day elevation in PDX-1 mRNA expression was observed. Furthermore, the MSCs exposed to nano collagen molecules produced the highest secretion of insulin (p ＜ 0.05). Type-2 diabetes induced by high-fat diet and low dose of streptozotocin in rat model was built in this study. This rat exhibited higher food intake, water intake, lower glucose tolerance, lower-insulin tolerance, and higher HbA1C (significant increases, p ＜ 0.01) as compared with the normal rat that demonstrated the model of type-2 diabetes was successfully built. Biopsy examinations also showed that obvious destruction of islet. After injection of differentiated MSCs into the destructed pancreas of diabetes rat, more regenerated islet were observed at the rats that treated with nano collagen molecules and exhibited much lower HbA1C as compared with the normal rat and diabetes rat after 4 weeks (significant deceases, p ＜ 0.001). These results indicate that the culturing MSCs with nano collagen molecules, nicotinamide, and exendin-4 are beneficial for MSCs differentiation into islet-like cells. These nano collagen molecules may lead to alternations or up-regulation of gene expression and influence the differentiated outcomes induced by nicotinamide and exendin-4.

Keywords: nano collagen molecules, nicotinamide, MSCs, diabetes

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Authors: Shah Abbas

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Rheumatoid arthritis is a systemic, inflammatory autoimmune disease, affecting an overall 1% of the global population. Despite being tremendous efforts by scientists, early diagnosis of RA still has not been achieved. In the current study, a Graphene oxide (GO) based electrochemical sensor has been developed for early diagnosis of RA through Cyclic voltammetry. Chitosan (CHI), a CPnatural polymer has also been incorporated along with GO in order to enhance the biocompatibility and functionalization potential of the biosensor. CCPs are known antigens for Anti Citrullinated Peptide Antibodies (ACPAs) which can be detected in serum even 14 years before the appearance of symptoms, thus they are believed to be an ideal target for the early diagnosis of RA. This study has yielded some promising results regarding the binding and detection of ACPAs through changes in the electrochemical properties of biosensing material. The cyclic voltammogram of this biosensor reflects the binding of ACPAs to the biosensor surface, due to its shifts observed in the current flow (cathodic current) as compared to the when no ACPAs bind as it is absent in RA negative patients.

Keywords: rheumatoid arthritis, peptide sensor, graphene oxide, anti citrullinated peptide antibodies, cyclic voltammetry

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Authors: Bawa Inalegwu, Jacob A. Jato, Ovye Akyengo, John Akighir

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All humans will experience some type of wound in every lifetime. Most wounds heal quickly with little or no attention but, many people suffer from wounds that are complex and/or persistent therefore posing a burden. This study was designed to assess the efficacy of Mitrcarpus villous ointment against honey in diabetic rats. To achieve this, percentage wound closure and collagen assessments were used to express treatment efficacy. Results show that on day 21, rats treated with M. villosus ointment had the highest percentage closure (94.5%) while honey treated and non-treated recorded 90.0% and 83.3% respectively. Similarly, a significant difference (p < 0.05) was observed on day 21 in the total collagen deposited in wounds of diabetic rats (10.57 ± 0.7) and M. villous ointment treated wounds (11.77 ± 0.4) as compared with the non-treated diabetic rats. M. villosus ointment was efficacious in healing wounds in diabetic rats and heals wound faster than honey and may hold potential for wound healing in diabetes mellitus sufferers. However, the wound healing mechanism of this ointment

Keywords: collagen, diabetic rats, honey, Mitracarpus villosus, ointment, wound healing

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Authors: G. Wojcik, J. Gindlhuber, A. Syarif, K. Hoetzenecker, P. Bohm, P. Vesely, V. Biasin, G. Kwapiszewska

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Pulmonary fibrosis is a rare disease where uncontrolled wound healing processes damage the lung structure. Intensive changes within the extracellular matrix (ECM) and its interaction with fibroblasts have a major role in pulmonary fibrosis development. Among others, collagen is one of the main components of the ECM, and it is important for lung structure. In IPF, constant production of collagen by fibroblast, through TGFβ1-SMAD2/3 pathways, leads to an uncontrolled deposition of matrix and hence lung remodeling. Abnormal changes in lipid production, alterations in fatty acids (FAs) metabolism, enhanced oxidative stress, and lipid peroxidation in fibrotic lung and fibrotic fibroblasts have been reported; however, the interplay between the collagen and lipids is not yet established. One of the FAs influx regulators is Angiopoietin-like 4 (ANGPTL4), which inhibits lipoprotein lipase work, decreasing the availability of FAs. We hypothesized that altered lipid composition or availability could have the capability to influence the phenotype of different fibroblast populations in the lung and hence influence lung fibrosis. To prove our hypothesis, we aim to investigate lipids and their influence on human, animal, and in vitro levels. In the bleomycin model, treatment with the well-known metabolic drugs Rosiglitazone or Metformin significantly lower collagen production. Similar results were noticed in ANGPTL4 KO animals, where the KO of ANGPTL4 leads to an increase of FAs availability and lower collagen deposition after the bleomycin challenge. Currently, we study the treatment of different FAs on human lung para fibroblasts (hPF) isolated from donors. To understand the lipid composition, we are collecting human lung tissue from donors and pulmonary fibrosis patients for Liquid chromatography-mass spectrometry. In conclusion, our results suggest the lipid influence on collagen deposition during lung fibrosis, but further research needs to be conducted to understand the matter of this relationship.

Keywords: collagen, fibroblasts, lipidomics, lung, pulmonary fibrosis

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Authors: Wei-Ting Kuo, Feng-Huei Lin

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Epidermal growth factor receptor (EGFR) is often constitutively stimulated in cancer owing to the binding of ligands such as epidermal growth factor (EGF), so it is necessary to investigate the interaction between EGFR and its targeting biomolecules which were over ligands binding. This study would focus on the binding affinity and adhesion force of two targeting products anti-EGFR monoclonal antibody (mAb) and peptide A to EGFR comparing with EGF. Surface plasmon resonance (SPR) was used to obtain the equilibrium dissociation constant to evaluate the binding affinity. Atomic force microscopy (AFM) was performed to detect adhesion force. The result showed that binding affinity of mAb to EGFR was higher than that of EGF to EGFR, and peptide A to EGFR was lowest. The adhesion force between EGFR and mAb that was higher than EGF and peptide A to EGFR was lowest. From the studies, we could conclude that mAb had better adhesion force and binding affinity to EGFR than that of EGF and peptide A. SPR and AFM could confirm the interaction between receptor and targeting ligand easily and carefully. It provide a platform to screen ligands for receptor targeting and drug delivery.

Keywords: adhesion force, binding affinity, epidermal growth factor receptor, target molecule

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Authors: Ahmed Khalaf Reyad Raslan

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Cancer stem cells (CSCs) describe a class of pluripotent cancer cells that behave analogously to normal stem cells in their ability to differentiate into the spectrum of cell types observed in tumors. The de-differentiation processes, such as an epithelial-mesenchymal transition (EMT), are known to enhance cellular plasticity. Here, we demonstrate a new hypothesis to use hybrid superparamagnetic magnetite nanoparticles (Fe₃O₄)- graphene oxide functionalized surface with Collagen to target the cancer stem cell as an early detection tool for cancer. We think that with the use of magnetic resonance imaging (MRI) and the new hybrid system would be possible to track the cancer stem cells.

Keywords: hydrogel, alginate, reduced graphene oxide, collagen

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Authors: Chetna Dhand, Venkatesh Mayandi, Silvia Marrero Diaz, Roger W. Beuerman, Seeram Ramakrishna, Rajamani Lakshminarayanan

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Sturdy Insect Cuticle Sclerotization, Incredible Substrate independent Mussel’s bioadhesion, Tanning of Leather are some of catechol(amine)s mediated natural processes. Chemical contemplation spots toward a mechanism instigated with the formation of the quinone moieties from the respective catechol(amine)s, via oxidation, followed by the nucleophilic addition of the amino acids/proteins/peptides to this quinone leads to the development of highly strong, cross-linked and water-resistant proteinacious structures. Inspired with this remarkable catechol(amine)s chemistry towards amino acids/proteins/peptides, we attempted to design highly stable and water-resistant antifungal wound dressing mats with exceptional durability using collagen (protein), dopamine (catecholamine) and antifungal drugs (Amphotericin B and Caspofungin) as the key materials. Electrospinning technique has been used to fabricate desired nanofibrous mat including Collagen (COLL), COLL/Dopamine (COLL/DP) and calcium incorporated COLL/DP (COLL-DP-Ca2+). The prepared protein-based scaffolds have been studied for their microscopic investigations (SEM, TEM, and AFM), structural analysis (FT-IR), mechanical properties, water wettability characteristics and aqueous stability. Biocompatibility of these scaffolds has been analyzed for dermal fibroblast cells using MTS assay, Cell TrackerTM Green CMFDA and confocal imaging. Being the winner sample, COLL-DP-Ca2+ scaffold has been selected for incorporating two antifungal drugs namely Caspofungin (Peptide based) and Amphotericin B (Non-Peptide based). Antifungal efficiency of the designed mats has been evaluated for eight diverse fungal strains employing different microbial assays including disc diffusion, cell-viability assay, time kill kinetics etc. To confirm the durability of these mats, in term of their antifungal activity, drug leaching studies has been performed and monitored using disc diffusion assay each day. Ex-vivo fungal infection model has also been developed and utilized to validate the antifungal efficacy of the designed wound dressings. Results clearly reveal dopamine mediated crosslinking within COLL-antifungal scaffolds that leads to the generation of highly stable, mechanical tough, biocompatible wound dressings having the zone of inhabitation of ≥ 2 cm for almost all the investigated fungal strains. Leaching studies and Ex-vivo model has confirmed the durability of these wound dressing for more than 3 weeks and certified their suitability for commercialization. A model has also been proposed to enlighten the chemical mechanism involved for the development of these antifungal wound dressings with exceptional robustness.

Keywords: catecholamine chemistry, electrospinning technique, antifungals, wound dressings, collagen

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Authors: Sayanti Kar, Amitava Ghosh, Pritam Aitch, Gupinath Bhandari

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An extensive seasonal analysis of wastewater had been done from outfalls of river Ganga in Howrah, Hooghly, 24 PGS (N) District, West Bengal, India during 2017. The morphological parameters of Bitter gourd (Momordica charantia L.) were estimated under wastewater treatment. An approach to study the activity within the range of low molecular weight peptide 3-0.5 kDa were taken through its extraction and purification by ion exchange resin column, cation, and anion exchanger. HPLC analysis had been done for both in wastewater treated and untreated plants. The antioxidant activity by using DPPH and germination percentage in control and treated plants were also determined in relation to wastewater effect. The inhibition of growth and its parameters were maximum in pre-monsoon in comparing to post-monsoon and monsoon season. The study also helped to explore the effect of wastewater on the peptidome of Bitter gourd (Momordica charantia L.). Some of these low molecular weight peptide(s) (3-0.5 kDa) also inhibited during wastewater treatment. Expression of particular peptide(s) or absence of some peptide(s) in chromatogram indicated the adverse effects on plants which may be the indication of stressful condition. Pre monsoon waste water was found to create more impact than other two.

Keywords: bitter gourd (Momordica charantia l.), low molecular weight peptide, river ganga, waste water

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Authors: Palwinder Singh

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Beyond the conventional mode of working with anti-inflammatory agents through enzyme inhibition, herein, an alternate substrate of cyclooxygenase-2 was developed. Proline centered pentapeptide iso-conformational to arachidonic acid exhibited appreciable selectivity for COX-2 overcoming acetic acid and formalin induced pain in rats to almost 80% and was treated as a substrate by the enzyme. Remarkably, COX-2 metabolized the pentapeptide into small fragments consisting mainly of di- and tri-peptides that ensured the safe breakdown of the peptide under in-vivo conditions. The kinetic parameter Kcat/Km for COX-2 mediated metabolism of peptide 6.3 x 105 M-1 s-1 was quite similar to 9.5 x 105 M-1 s-1 for arachidonic acid. Evidenced by the dynamic molecular studies and the use of Y385F COX-2, it was observed that the breakage of the pentapeptide has probably taken place through H-bond activation of the peptide bond by the side chains of Y385 and S530.

Keywords: small peptides, anti-inflammatory agents, cyclooxygenase-2, unnatural substrates

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Authors: Amarjyoti Das Mahapatra, Umesh Kumar, Bhaskar Datta

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A pseudo peptide can mimic the biological or structural properties of natural peptides. They have become an increasing attention in medicinal chemistry because of their interesting advantages like more bioavailability and less biodegradation than compare to the physiologically active native peptides which increase their therapeutic applications. Many biologically active compounds contain urea as functional groups, and they have improved pharmacokinetic properties because of their bioavailability and metabolic stability. Recently we have reported a single-step synthesis of sulfonyl urea and sulfonyltriuret from sulfonyl chloride and sodium cyanate. But the yield of sulfonyltriuret was less around 40-60% because of the formation of other products like sulfonamide and sulfonylureas. In the present work, we mainly focused on the selective synthesis of sulfonyltriuret using tetrabutylammonium cyanate and sulfonyl chloride. More precisely, we are interested in the controlled synthesis of oligomeric urea mainly sulfonyltriuret as a new class of pseudo peptide and their application as protease modulators. The distinctive architecture of these molecules in the form of their pseudo-peptide backbone offers promise as a potential pharmacophore. The synthesized molecules have been screened on trypsin enzyme, and we observed that these molecules are the efficient modulator of trypsin enzyme.

Keywords: pseudo peptide, pharmacophore, sulfonyltriuret, trypsin

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Authors: Anna Trusek-Holownia, Andrzej Noworyta

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Using an enzyme of known specificity the hydrolysis of protein was carried out in a controlled manner. The aim was to obtain oligopeptides being the so-called active peptides or their direct precursors. An original way of expression of the protein hydrolysis kinetics was introduced. Peptide bonds contained in the protein were recognized as a diverse-quality substrate for hydrolysis by the applied protease. This assumption was positively verified taking as an example the hydrolysis of albumin by thermolysin. Peptide linkages for this system should be divided into at least four groups. One of them is a group of bonds non-hydrolyzable by this enzyme. These that are broken are hydrolyzed at a rate that differs even by tens of thousands of times. Designated kinetic constants were k'F = 10991.4 L/g.h, k'M = 14.83L/g.h, k'S about 10-1 L/g.h for fast, medium and slow bonds, respectively. Moreover, a procedure for unfolding of the protein, conducive to the improved susceptibility to enzymatic hydrolysis (approximately three-fold increase in the rate) was proposed.

Keywords: peptide bond hydrolysis, kinetics, enzyme specificity, biologically active peptides

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Authors: Thangavelu Muthukumar, Thotapalli Parvathaleswara Sastry

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Collagen is the most abundantly available connective tissue protein, which is being used as a biomaterial for various biomedical applications. Presently, fish wastes are disposed improperly which is causing serious environmental pollution resulting in offensive odour. Fish scales are promising source of Type I collagen. Medicinal plants have been used since time immemorial for treatment of various ailments of skin and dermatological disorders especially cuts, wounds, and burns. Developing biomaterials from the natural sources which are having wound healing properties within the search of a common man is the need of hour, particularly in developing and third world countries. With these objectives in view we have developed a wound dressing material containing fish scale collagen (FSC) incorporated with Macrotyloma uniflorum plant extract (PE). The wound dressing composite was characterized for its physiochemical properties using conventional methods. SEM image revealed that the composite has fibrous and porous surface which helps in transportation of oxygen as well as absorbing wound fluids. The biomaterial has shown 95% biocompatibility with required mechanical strength and has exhibited antimicrobial properties. This biomaterial has been used as a wound dressing material in experimental wounds of rats. The healing pattern was evaluated by macroscopic observations, panimetric studies, biochemical, histopathological observations. The results showed faster healing pattern in the wounds treated with CSPE compared to the other composites used in this study and untreated control. These experiments clearly suggest that CSPE can be used as wound/burn dressing materials.

Keywords: collagen, wound dressing, Macrotyloma uniflorum, burn dressing

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Authors: Rolivhuwa Bishop Ramagoma1*, Lynn Cairncross1, , Saartjie Roux1

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According to the world health organisation, colon cancer is among the most common cancers diagnosed in both men and women. Specifically, it is the second leading cause of cancer related deaths accounting for over 860 000 deaths worldwide in 2018. Currently, chemotherapy has become an essential component of most cancer treatments. Despite progress in cancer drug development over the previous years, traditional chemotherapeutic drugs still have low selectivity for targeting tumour tissues and are frequently constrained by dose-limiting toxicity. The creation of nanoscale delivery vehicles capable of directly directing treatment into cancer cells has recently caught the interest of researchers. Herein, the development of peptide-functionalized polyethylene glycol gold nanoparticles (Peptide-PEG-AuNPs) as a cellular probe and delivery agent is described, with the higher aim to develop a specific diagnostic prototype and assess their specificity not only against cell lines but primary human cells as well. Gold nanoparticles (AuNPs) were synthesized and stabilized through chemical conjugation. The synthesized AuNPs were characterized, stability in physiological solutions was assessed, their cytotoxicity against colon carcinoma and non-carcinoma skin fibroblasts was also studied. Furthermore, genetic effect through real-time polymerase chain reaction (RT-PCR), localization and uptake, peptide specificity were also determined. In this study, different peptide-AuNPs were found to have preferential toxicity at higher concentrations, as revealed by cell viability assays, however, all AuNPs presented immaculate stability for over 3 months following the method of synthesis. The final obtained peptide-PEG-AuNP conjugates showed good biocompatibility in the presence of high ionic solutions and biological media and good cellular uptake. Formulation of colon cancer specific targeting peptide was successful, additionally, the genes/pathways affected by the treatments were determined through RT-PCR. Primary cells study is still on going with promising results thus far.

Keywords: nanotechnology, cancer, diagnosis, therapeutics, gold nanoparticles.

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Authors: Supanan Chansap, Werachon Cheukamud, Pornanan Kueakhai, Narin Changklungmoa

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Fasciola species (Fasciola spp.) is caused fasciolosis in ruminants such as cattle, sheep, and buffalo. Fasciola gigantica (F.gigantica) commonly infects tropical regions. Fasciola hepatica (F.hepatica) in temperate regions. Liver fluke infection affects livestock economically, for example, reduced milk and meat production, weight loss, sterile animals. Currently, Triclabendazole is used to treat liver flukes. However, liver flukes have also been found to be resistant to drugs in countries. Therefore, vaccination is an attractive alternative to prevent liver fluke infection. Peptide vaccines are new vaccine technologies that mimic epitope antigens that trigger an immune response. An interesting antigen used in vaccine production is catepsin L, a family of proteins that play an important role in the life of the parasite in the host. This study aims to identify immunogenic regions of protein and construct a multi-epidetope vaccine using an immunoinformatic tool. Fasciola gigantica Cathepsin L1 (FgCatL1), Fasciola gigantica Cathepsin L1G (FgCatL1G), and Fasciola gigantica Cathepsin L1H (FgCatL1H) were predicted B-cell and Helper T lymphocytes (HTL) by Immune Epitope and Analysis Resource (IEDB) servers. Both B-cell and HTL epitopes aligned with cathepsin L of the host and Fasciola hepatica (F. hepatica). Epitope groups were selected from non-conserved regions and overlapping sequences with F. hepatica. All overlapping epitopes were linked with the GPGPG and KK linker. GPGPG linker was linked between B-cell epitope. KK linker was linked between HTL epitope and B-cell and HTL epitope. The antigenic scores of multi-epitope peptide vaccine was 0.7824. multi-epitope peptide vaccine was non-allergen, non-toxic, and good soluble. Multi-epitope peptide vaccine was predicted tertiary structure and refinement model by I-Tasser and GalaxyRefine server, respectively. The result of refine structure model was good quality that was generated by Ramachandran plot analysis. Discontinuous and linear B-cell epitopes were predicted by ElliPro server. Multi-epitope peptide vaccine model was two and seven of discontinuous and linear B-cell epitopes, respectively. Furthermore, multi-epitope peptide vaccine was docked with Toll-like receptor 2 (TLR-2). The lowest energy ranged from -901.3 kJ/mol. In summary, multi-epitope peptide vaccine was antigenicity and probably immune response. Therefore, multi-epitope peptide vaccine could be used to prevent F. gigantica infections in the future.

Keywords: fasciola gigantica, Immunoinformatic tools, multi-epitope, Vaccine

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Authors: Ratchada Sopakayang, Gerhard A. Holzapfel

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In this study, a new constitutive model is developed to describe the hyperelastic behavior of collagenous tissues with a parallel arrangement of collagen fibers such as ligaments and tendons. The model is formulated using a continuum approach incorporating the structural changes of the main tissue components: collagen fibers, proteoglycan-rich matrix and fiber-matrix interaction. The mechanical contribution of the interaction between the fibers and the matrix is simply expressed by a coupling term. The structural change of the collagen fibers is incorporated in the constitutive model to describe the activation of the fibers under tissue straining. Finally, the constitutive model can easily describe the stress-stretch nonlinearity which occurs when a ligament/tendon is axially stretched. This study shows that the interaction between the fibers and the matrix contributes to the mechanical tissue response. Therefore, the model may lead to a better understanding of the physiological mechanisms of ligaments and tendons under axial loading.

Keywords: constitutive model, fiber-matrix, hyperelasticity, interaction, ligament, tendon

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Authors: Saroj Poudel, Joshua Mancini, Douglas Pike, Jennifer Timm, Alexei Tyryshkin, Vikas Nanda, Paul Falkowski

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On the early Earth, protein-metal complexes likely harvested energy from a reduced environment. These complexes would have been precursors to the metabolic enzymes of ancient organisms. Hydrogenase is an essential enzyme in most anaerobic organisms for the reduction and oxidation of hydrogen in the environment and is likely one of the earliest evolved enzymes. To attempt to reinvent a precursor to modern hydrogenase, we computationally designed a short thirteen amino acid peptide that binds the often-required catalytic transition metal Nickel in hydrogenase. This simple complex can achieve hundreds of hydrogen evolution cycles using light energy in a broad range of temperature and pH. Biophysical and structural investigations strongly indicate the peptide forms a di-nickel active site analogous to Acetyl-CoA synthase, an ancient protein central to carbon reduction in the Wood-Ljungdahl pathway and capable of hydrogen evolution. This work demonstrates that prior to the complex evolution of multidomain enzymes, early peptide-metal complexes could have catalyzed energy transfer from the environment on the early Earth and enabled the evolution of modern metabolism

Keywords: hydrogenase, prebiotic enzyme, metalloenzyme, computational design

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Authors: Manh Tin Ho, Phorl Sophors, Ga Young Seo, Young Mee Kim, Youngho Lim, Moonjae Cho

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UV radiation in sunlight is one of the most potential factor induced skin ageing and photocarcinogenesis. UV may induce the melanin production and wrinkle formation. Recently, the natural secondary compounds have been reported that had the beneficial protective effects from UV light. In this study, we investigated the effects of two different compounds, glycitin and 38, on human dermal fibroblast. We first only treated the 38 on melanocyte cell to test the proliferation inhibition of 38 on this cell line. Then, we induced the combination of glycitin and 38 on human dermal fibroblast in 48h and investigate the proliferation, collagen production and the metalloproteinase family expression. The 38 alone could inhibit the proliferation of melanocyte which indicated the reduction of melanin production. The combination of glycitin and 38 truly increased the fibroblast proliferation and even they could recover the UV-induced and H2O2-induced damaged fibroblast proliferation. The co-treatment also promoted the collagen IV expression significantly and accelerated the total collagen secretion. In addition, metalloproteinase (MMPs) family such as MMP1, MMP2, MMP7 was down-regulated in transcriptional level. In conclusion, the combination of glycitin and 38 has induced the fibroblast proliferation even when it was damaged by UV exposure and H2O2, whereas augmented collagen production and inhibited the MMPs caused the wrinkle formation and decreased the melanocyte proliferation, suggested an potential UV-protective therapy.

Keywords: UV radiation, wrinkle, ageing, glycitin, dermal fibroblast

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Authors: M. H. A. D. de Silva, R. P. Hewawasam, M. A. G. Iresha

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Macrosomia is common in infants born to not only women diagnosed with gestational diabetes mellitus but also non-diabetic obese women. Maternal Body Mass Index (BMI) correlates with the incidence of large for gestational age infants. Obesity has reached epidemic levels in modern societies. During the past two decades, obesity in children and adolescents has risen significantly in Asian populations including Sri Lanka. There is increasing evidence to believe that infants who are born large for gestational age are more likely to be obese in childhood and adolescence and are at risk of cardiovascular and metabolic complications later in life. It is also established that Asians have lower skeletal muscle mass, low bone mineral content and excess body fat for a given BMI indicating a genetic predisposition in the occurrence of obesity. The objective of this study is to determine the effect of maternal weight, weight gain during pregnancy, c-peptide and insulin concentrations in the cord blood on the birth of appropriate for and large for gestational age infants in a tertiary care center in Southern Sri Lanka. Umbilical cord blood was collected from 90 newborns (Male 40, Female 50; gestational age 35-42 weeks) after double clamping the umbilical cord before separation of the placenta and the concentration of insulin and C-peptide were measured by ELISA technique. Anthropometric parameters of the newborn such as birth weight, length, ponderal index, occipital frontal, chest, hip and calf circumferences were measured, and characteristics of the mother were collected. The relationship between insulin, C-peptide and anthropometrics were assessed by Spearman correlation. The multiple logistic regression analysis examined influences of maternal weight, weight gain during pregnancy, C-peptide and insulin concentrations in cord blood as covariates on the birth of large for gestational age infants. A significant difference (P<0.001) was observed between the insulin levels of infants born large for gestational age (18.73 ± 0.52 µlU/ml) and appropriate for gestational age (13.08 ± 0.56 µlU/ml). Consistently, A significant decrease in concentration (41.68%, P<0.001) was observed between C-peptide levels of infants born large for gestational age and appropriate for gestational age. Cord blood insulin and C-peptide levels had a significant correlation with birth weight (r=0.35, P<0.05) of the newborn at delivery. Maternal weight and BMI which are indicators of maternal nutrition were proven to be directly correlated with birth weight and length. To our knowledge, this relationship was investigated for the first time in a Sri Lankan setting and was also evident in our results. This study confirmed the fact that insulin and C-peptide play a major role in regulating fetal growth. According to the results obtained in this study, we can suggest that the increased BMI of the mother has a direct influence on increased maternal insulin secretion, which may subsequently affect cord insulin and C-peptide levels and also birth weight of the infant.

Keywords: C-peptide, insulin, large for gestational age, maternal weight

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Authors: Esra Turker, Umit Hakan Yildiz, Ahu Arslan Yildiz

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The key of regenerative medicine is mimicking natural three dimensional (3D) microenvironment of tissues by utilizing appropriate biomaterials. In this study, a synthetic biodegradable polymer; poly (L-lactide-co-ε-caprolactone) (PLLCL) and a natural polymer; collagen was used to mimic the biochemical structure of the natural extracellular matrix (ECM), and by means of electrospinning technique the real physical structure of ECM has mimicked. PLLCL/Collagen biocomposite scaffolds enables cell attachment, proliferation and nutrient transport through fabrication of micro to nanometer scale nanofibers. Biocomposite materials are commonly preferred due to limitations of physical and biocompatible properties of natural and synthetic materials. Combination of both materials improves the strength, degradation and biocompatibility of scaffold. Literature studies have shown that collagen is mostly solved with heavy chemicals, which is not suitable for cell culturing. To overcome this problem, a new approach has been developed in this study where polyvinylpyrrolidone (PVP) is used as co-electrospinning agent. PVP is preferred due to its water solubility, so PLLCL/collagen biocomposite scaffold can be easily and rapidly produced. Hydrolytic and enzymatic biodegradation as well as mechanical strength of scaffolds were examined in vitro. Cell adhesion, proliferation and cell morphology characterization studies have been performed as well. Further, on-chip drug screening analysis has been performed over 3D tumor models. Overall, the developed biocomposite scaffold was used for 3D tumor model formation and obtained results confirmed that developed model could be used for drug screening studies to predict clinical efficacy of a drug.

Keywords: biomaterials, 3D cell culture, drug screening, electrospinning, lab-on-a-chip, tissue engineering

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Authors: Armando A. Rodríguez, Emilio Salceda, Anoland Garateix, André J. Zaharenko, Steve Peigneur, Omar López, Tirso Pons, Michael Richardson, Maylín Díaz, Yasnay Hernández, Ludger Ständker, Jan Tytgat, Enrique Soto

Abstract:

Acid-sensing ion channels are cation (Na+) channels activated by a pH drop. These proteins belong to the ENaC/degenerin superfamily of sodium channels. ASICs are involved in sensory perception, synaptic plasticity, learning, memory formation, cell migration and proliferation, nociception, and neurodegenerative disorders, among other processes; therefore those molecules that specifically target these channels are of growing pharmacological and biomedical interest. Sea anemones produce a large variety of ion channels peptide toxins; however, those acting on ligand-gated ion channels, such as Glu-gated, Ach-gated ion channels, and acid-sensing ion channels (ASICs), remain barely explored. The peptide PhcrTx1 is the first compound characterized from the sea anemone Phymanthus crucifer, and it constitutes a novel ASIC inhibitor. This peptide was purified by chromatographic techniques and pharmacologically characterized on acid-sensing ion channels of mammalian neurons using patch-clamp techniques. PhcrTx1 inhibited ASIC currents with an IC50 of 100 nM. Edman degradation yielded a sequence of 32 amino acids residues, with a molecular mass of 3477 Da by MALDI-TOF. No similarity to known sea anemone peptides was found in protein databases. The computational analysis of Cys-pattern and secondary structure arrangement suggested that this is a structurally ICK (Inhibitor Cystine Knot)-type peptide, a scaffold that had not been found in sea anemones but in other venomous organisms. These results show that PhcrTx1 represents the first member of a new structural group of sea anemones toxins acting on ASICs. Also, this peptide constitutes a novel template for the development of drugs against pathologies related to ASICs function.

Keywords: animal toxin, inhibitor cystine knot, ion channel, sea anemone

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Authors: F. Bellali, M. Kharroubi, Y. Rady, N. Bourhim

Abstract:

In Morocco, fish processing industry is an important source income for a large amount of by-products including skins, bones, heads, guts, and scales. Those underutilized resources particularly scales contain a large amount of proteins and calcium. Sardina plichardus scales from resulting from the transformation operation have the potential to be used as raw material for the collagen production. Taking into account this strong expectation of the regional fish industry, scales sardine upgrading is well justified. In addition, political and societal demands for sustainability and environment-friendly industrial production systems, coupled with the depletion of fish resources, drive this trend forward. Therefore, fish scale used as a potential source to isolate collagen has a wide large of applications in food, cosmetic, and biomedical industry. The main aim of this study is to isolate and characterize the acid solubilize collagen from sardine fish scale, Sardina pilchardus. Experimental design methodology was adopted in collagen processing for extracting optimization. The first stage of this work is to investigate the optimization conditions of the sardine scale deproteinization on using response surface methodology (RSM). The second part focus on the demineralization with HCl solution or EDTA. And the last one is to establish the optimum condition for the isolation of collagen from fish scale by solvent extraction. The advancement from lab scale to pilot scale is a critical stage in the technological development. In this study, the optimal condition for the deproteinization which was validated at laboratory scale was employed in the pilot scale procedure. The deproteinization of fish scale was then demonstrated on a pilot scale (2Kg scales, 20l NaOH), resulting in protein content (0,2mg/ml) and hydroxyproline content (2,11mg/l). These results indicated that the pilot-scale showed similar performances to those of lab-scale one.

Keywords: deproteinization, pilot scale, scale, sardine pilchardus

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Authors: Abu Salim Mustafa

Abstract:

Rv3873 is a relatively large size protein (371 amino acids in length) and its gene is located in the immunodominant genomic region of difference (RD)1 that is present in the genome of Mycobacterium tuberculosis but deleted from the genomes of all the vaccine strains of Bacillus Calmette Guerin (BCG) and most other mycobacteria. However, when tested for cellular immune responses using peripheral blood mononuclear cells from tuberculosis patients and BCG-vaccinated healthy subjects, this protein was found to be a major stimulator of cell mediated immune responses in both groups of subjects. In order to further identify the sequence of immunodominant epitopes and explore their Human Leukocyte Antigen (HLA)-restriction for epitope recognition, 24 peptides (25-mers overlapping with the neighboring peptides by 10 residues) covering the sequence of Rv3873 were synthesized chemically using fluorenylmethyloxycarbonyl chemistry and tested in cell mediated immune responses. The results of these experiments helped in the identification of an immunodominant peptide P9 that was recognized by people expressing varying HLA-DR types. Furthermore, it was also predicted to be a promiscuous binder with multiple epitopes for binding to HLA-DR, HLA-DP and HLA-DQ alleles of HLA-class II molecules that present antigens to T helper cells, and to HLA-class I molecules that present antigens to T cytotoxic cells. In addition, the evaluation of peptide P9 using an immunogenicity predictor server yielded a high score (0.94), which indicated a greater probability of this peptide to elicit a protective cellular immune response. In conclusion, P9, a peptide with multiple epitopes and ability to bind several HLA class I and class II molecules for presentation to cells of the cellular immune response, may be useful as a peptide-based vaccine against tuberculosis.

Keywords: mycobacterium tuberculosis, PPE68, peptides, vaccine

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Authors: Abu Salim Mustafa

Abstract:

The comparisons of mycobacterial genomes have identified several Mycobacterium tuberculosis-specific genomic regions that are absent in other mycobacteria and are known as regions of differences. Due to M. tuberculosis-specificity, the peptides encoded by these regions could be useful in the specific diagnosis of tuberculosis. To explore this possibility, overlapping synthetic peptides corresponding to 39 proteins predicted to be encoded by genes present in regions of differences were tested for antibody-reactivity with sera from tuberculosis patients and healthy subjects. The results identified four immunodominant peptides corresponding to four different proteins, with three of the peptides showing significantly stronger antibody reactivity and rate of positivity with sera from tuberculosis patients than healthy subjects. The fourth peptide was recognized equally well by the sera of tuberculosis patients as well as healthy subjects. Predication of antibody epitopes by bioinformatics analyses using ABCpred server predicted multiple linear epitopes in each peptide. Furthermore, peptide sequence analysis for sequence identity using BLAST suggested M. tuberculosis-specificity for the three peptides that had preferential reactivity with sera from tuberculosis patients, but the peptide with equal reactivity with sera of TB patients and healthy subjects showed significant identity with sequences present in nob-tuberculous mycobacteria. The three identified M. tuberculosis-specific immunodominant peptides may be useful in the serological diagnosis of tuberculosis.

Keywords: genomic regions of differences, Mycobacterium tuberculossis, peptides, serodiagnosis

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Authors: Hadi Rad

Abstract:

Unrestricted somatic stem cells (USSCs) loaded in nanofibrous PHBV scaffold can be used for skin regeneration when grafted into full-thickness skin defects of rats. Nanofibrous PHBV scaffolds were designed using electrospinning method and then, modified with the immobilized collagen via the plasma method. Afterward, the scaffolds were evaluated using scanning electron microscopy, physical and mechanical assays. In this study; nanofibrous PHBV scaffolds loaded with and without USSCs were grafted into the skin defects. The wounds were subsequently investigated at 21 days after grafting. Results of mechanical and physical analyses showed good resilience and compliance to movement as a skin graft. In animal models; all study groups excluding the control group exhibited the most pronounced effect on wound closure, with the statistically significant improvement in wound healing being seen on post-operative Day 21. Histological and immunostaining examinations of healed wounds from all groups, especially the groups treated with stem cells, showed a thin epidermis plus recovered skin appendages in the dermal layer. Thus, the graft of collagen-coated nanofibrous PHBV scaffold loaded with USSC showed better results during the healing process of skin defects in rat model.

Keywords: collagen, nanofibrous PHBV scaffold, unrestricted somatic stem cells, wound healing.

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Authors: Frida Carrillo Morales, Maria Maricela Carrasco Yépez, Saúl Rojas Hernández

Abstract:

Naegleria fowleri is the causative agent of primary amebic meningoencephalitis, this disease is acute and fulminant that affects humans. It has been reported that despite the existence of therapeutic options against this disease, its mortality rate is 97%. Therefore, the need arises to have vaccines that confer protection against this disease and, in addition to developing adjuvants to enhance the immune response. In this regard, in our work group, we obtained a peptide designed from the membrane protein MP2CL5 of Naegleria fowleri called Smp145 that was shown to be immunogenic; however, it would be of great importance to enhance its immunological response, being able to co-administer it with a non-toxic adjuvant. Therefore, the objective of this work was to carry out the bioinformatic design of a peptide of the Naegleria fowleri membrane protein MP2CL5 conjugated with a non-toxic modified adjuvant from the native A1 structure of Cholera Toxin. For which different bioinformatics tools were used to obtain a model with a modification in amino acid 61 of the A1 subunit of the CT (CTA1), to which the Smp145 peptide was added and both molecules were joined with a 13-glycine linker. As for the results obtained, the modification in CTA1 bound to the peptide produces a reduction in the toxicity of the molecule in in silico experiments, likewise, the prediction in the binding of Smp145 to the receptor of B cells suggests that the molecule is directed in specifically to the BCR receptor, decreasing its native enzymatic activity. The stereochemical evaluation showed that the generated model has a high number of adequately predicted residues. In the ERRAT test, the confidence with which it is possible to reject regions that exceed the error values was evaluated, in the generated model, a high score was obtained, which determines that the model has a good structural resolution. Therefore, the design of the conjugated peptide in this work will allow us to proceed with its chemical synthesis and subsequently be able to use it in the mouse meningitis protection model caused by N. fowleri.

Keywords: immunology, vaccines, pathogens, infectious disease

Procedia PDF Downloads 59

Authors: R. P. Hewawasam, M. H. A. D. de Silva, M. A. G. Iresha

Abstract:

Obesity is associated with an increased risk of developing insulin resistance. Insulin resistance often progresses to type-2 diabetes mellitus and is linked to a wide variety of other pathophysiological features including hypertension, hyperlipidemia, atherosclerosis (metabolic syndrome) and polycystic ovarian syndrome. Macrosomia is common in infants born to not only women with gestational diabetes mellitus but also non-diabetic obese women. During the past two decades, obesity in children and adolescents has risen significantly in Asian populations including Sri Lanka. There is increasing evidence to believe that infants who are born large for gestational age (LGA) are more likely to be obese in childhood. It is also established from previous studies that Asian populations have higher percentage body fat at a lower body mass index compared to Caucasians. High leptin levels in cord blood have been reported to correlate with fetal adiposity at birth. Previous studies have also shown that cord blood C-peptide and insulin levels are significantly and positively correlated with birth weight. Therefore, the objective of this preliminary study was to determine the relationship between parameters of fetal insulin resistance such as leptin, C-peptide and insulin and the birth weight of the newborn in a study population in Southern Sri Lanka. Umbilical cord blood was collected from 90 newborns and the concentration of insulin, leptin, and C-peptide were measured by ELISA technique. Birth weight, length, occipital frontal, chest, hip and calf circumferences of newborns were measured and characteristics of the mother such as age, height, weight before pregnancy and weight gain were collected. The relationship between insulin, leptin, C-peptide, and anthropometrics were assessed by Pearson’s correlation while the Mann-Whitney U test was used to assess the differences in cord blood leptin, C-peptide, and insulin levels between groups. A significant difference (p < 0.001) was observed between the insulin levels of infants born LGA (18.73 ± 0.64 µlU/ml) and AGA (13.08 ± 0.43 µlU/ml). Consistently, A significant increase in concentration (p < 0.001) was observed in C-peptide levels of infants born LGA (9.32 ± 0.77 ng/ml) compared to AGA (5.44 ± 0.19 ng/ml). Cord blood leptin concentration of LGA infants (12.67 ng/mL ± 1.62) was significantly higher (p < 0.001) compared to the AGA infants (7.10 ng/mL ± 0.97). Significant positive correlations (p < 0.05) were observed among cord leptin levels and the birth weight, pre-pregnancy maternal weight and BMI between the infants of AGA and LGA. Consistently, a significant positive correlation (p < 0.05) was observed between the birth weight and the C peptide concentration. Significantly high concentrations of leptin, C-peptide and insulin levels in the cord blood of LGA infants suggest that they may be involved in regulating fetal growth. Although previous studies suggest comparatively high levels of body fat in the Asian population, values obtained in this study are not significantly different from values previously reported from Caucasian populations. According to this preliminary study, maternal pre-pregnancy BMI and weight may contribute as significant indicators of cord blood parameters of insulin resistance and possibly the birth weight of the newborn.

Keywords: large for gestational age, leptin, C-peptide, insulin

Procedia PDF Downloads 134

Authors: Prajna Metta, Yanti Rusyanti, Nunung Rusminah, Bremmy Laksono

Abstract:

The decrease of neutrophil chemotaxis function may cause increased susceptibility to aggressive periodontitis (AP). Neutrophil chemotaxis is affected by formyl peptide receptor 1 (FPR1), which when activated will respond to bacterial chemotactic peptide formyl methionyl leusyl phenylalanine (FMLP). FPR1 protein value is decreased in response to a wide number of inflammatory stimuli in AP patients. This study was aimed to assess the alteration of FPR1 protein value in AP patients and if FPR1 protein value could be used as an indicator of neutrophil chemotaxis dysfunction in AP. This is a case control study with 20 AP patients and 20 control subjects. Three milliliters of peripheral blood were drawn and analyzed for FPR1 protein value with ELISA. The data were statistically analyzed with Mann-Whitney test (p>0,05). Results showed that the mean value of FPR1 protein value in AP group is 0,353 pg/mL (0,11 to 1,18 pg/mL) and the mean value of FPR1 protein value in control group is 0,296 pg/mL (0,05 to 0,88 pg/mL). P value 0,787 > 0,05 suggested that there is no significant difference of FPR1 protein value in both groups. The present study suggests that FPR1 protein value has no significance alteration in AP patients and could not be used as an indicator of neutrophil chemotaxis dysfunction.

Keywords: aggressive periodontitis, chemotaxis dysfunction, FPR1 protein value, neutrophil

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Authors: Alok Pal Jain, Santram Lodhi

Abstract:

Martynia annua Linn. (Martyniaccae) is traditionally used in inflammation and applied locally to tuberculosis glands of camel’s neck. The leaves used topically to bites of venomous insects and wounds of domestic animals. Chemical examination of Martynia annua leaves revealed the presence of glycosides, tannins, proteins, phenols and flavonoids. The present study was aimed to evaluate the anti-arthritic activity of methanolic extract of Martynia annua leaves. Methanolic extract of Martynia annua leaves was tested by using in vivo collagen-induced arthritis mouse model to investigate the anti-rheumatoid arthritis activity. In addition, antioxidant effect of methanolic extract was determined by the estimation of antioxidants level in joint tissues. The severity of arthritis was assessed by arthritis score and edema. Levels of cytokines TNF-α and IL-6, in the joint tissue homogenate were measured using ELISA. A high dose (250 mg/kg) of methanolic extract was significantly reduced the degree of inflammation in mice as compared with reference drug. Antioxidants level and malondialdehyde (MDA) in joint tissue homogenate found significantly (p < 0.05) higher. Methanolic extract at dose of 250 mg/kg modulated the cytokines production and suppressed the oxidative stress in the mice with collagen-induced arthritis. This study suggested that Martynia annua might be alternative herbal medicine for the management of rheumatoid arthritis.

Keywords: Martynia annua, collagen, rheumatoid arthritis, antioxidants

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Authors: Recep Kesli, Onur Turkyilmaz, Cengiz Demir

Abstract:

Objective: Autoimmune collagen diseases occur with the immune reactions against the body’s own cell or tissues which cause inflammation and damage the tissues and organs. In this study, it was aimed to compare seropositivity rates and patterns of the anti-nuclear antibodies (ANA) in the diagnosis of four different autoimmune collagen tissue diseases (Rheumatoid Arthritis-RA, Systemic Lupus Erythematous-SLE, Scleroderma-SSc and Sjogren Syndrome-SS) with each other. Methods: One hundred eighty-eight patients applied to different clinics in Afyon Kocatepe University ANS Practice and Research Hospital between 11.07.2014 and 14.07.2015 that thought the different collagen disease such as RA, SLE, SSc and SS have participated in the study retrospectively. All the data obtained from the patients participated in the study were evaluated according to the included criteria. The historical archives belonging to the patients have been screened, assessed in terms of ANA positivity. The obtained data was analysed by using the descriptive statistics; chi-squared, Fischer's exact test. The evaluations were performed by SPSS 20.0 version and p < 0.05 level was considered as significant. Results: Distribution rates of the totally one hundred eighty-eight patients according to the diagnosis were found as follows: 82 (43.6%) were RA, 38 (20.2%) were SLE, 22 (11.7%) were SSc, and 46 (24.5%) were SS. Distribution of ANA positivity rates according to the collagen tissue diseases were found as follows; for RA were 54 (65,9 %), for SLE were 36 (94,7 %), for SSc were 18 (81,8 %), and for SS were 43 (93,5 %). Rheumatoid arthritis should be evaluated and classified as a different class among all the other investigated three autoimmune illnesses. ANA positivity rates were found as differently higher (91.5 %) in the SLE, SSc, and SS, from the RA (65.9 %). Differences at ANA positivity rates for RA and the other three diseases were found as statistically significant (p=0.015). Conclusions: Systemic autoimmune illnesses show broad spectrum. ANA positivity was found as an important predictor marker in the diagnosis of the rheumatologic illnesses. ANA positivity should be evaluated as more valuable and sensitive a predictor diagnostic marker in the laboratory findings of the SLE, SSc, and SS according to RA.

Keywords: antinuclear antibody (ANA), rheumatoid arthritis, scleroderma, Sjogren syndrome, systemic lupus Erythemotosus

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