Fibroblast Cell Cultures Research Articles

Microbiological Assessment and Evaluation of Virus Yield in Germ-free Chicken Eggs

Background Increasing yield will have major implications for vaccine manufacturers. Internal bacterial contamination of specific pathogen-free (SPF) eggs results in batches of vaccines being rejected and may negatively impact vaccine yield. Objective The objectives of this study were to analyze Germ-Free (GF) eggs microbiologically and evaluate the yield of H1N1 when compared with SPF eggs from 2 different suppliers. Methods The contents of GF eggs were analyzed using three different methods (sterility test, total viable count, and bioluminescence). Eggs from each source were inoculated with H1N1 virus. Chick Embryo Fibroblast (CEF) cell cultures were generated from GF and SPF eggs from both suppliers and infected with H1N1. The supernatant was titrated for viral Haemagglutinin activity and virus infectivity while cellular RNA extracts were assayed for Interferon (α and β) gene expression by RT-qPCR using gene-specific primers. Results The analysis of GF eggs confirmed that the contents were sterile. The batch yield of GF eggs was ~1.2-fold and ~1.8-fold higher than SPF supplier 1 and SPF supplier 2 virus yields, respectively. Replication of H1N1 was 2-fold higher in CEF derived from GF eggs than CEF derived from SPF supplier 1 or SPF supplier 2. IFN-β antiviral response of GF-derived CEF cells was 2-fold lower than SPF supplier 1-derived and ~3-fold lower than SPF supplier 2-derived CEF cells. Conclusion The content of GF eggs was sterile. The yield of H1N1 was higher in GF eggs than in SPF eggs, and the higher yield was associated with a significant reduction in interferon Beta levels.

Engineering Zn/Fe Mixed Metal Oxides with Tunable Structural and Magnetic Properties for Magnetic Particle Imaging.

Engineering magnetic nanoparticles with tunable structural properties and magnetism is critical to develop desirable magnetic particle imaging (MPI) tracers for biomedical applications. Here we present a new superparamagnetic metal oxide nanoparticle with a controllable chemical composition and magnetism for imaging tumor xenografts in living mice. Superparamagnetic Zn/Fe mixed metal oxide (ZnFe-MMO) nanoparticles are fabricated via a facile one-pot co-precipitation method in water followed by thermal decomposition with tunable Zn/Fe ratios and at various calcination temperatures. This work, for the first time, presented LDH-derived metal oxides for an MPI application. The metal composition is tunable to present an optimized MPI performance. The analytical results demonstrate that ZnFe-MMO nanoparticles at the designed molar ratio of Zn/Fe = 2:1 after 650 °C calcination demonstrate a higher saturation magnetization (MS) value and optimal MPI signal than the samples presented with other conditions. The excellent biocompatibility of ZnFe-MMO is demonstrated in both breast cancer cells and fibroblast cell cultures. In vivo imaging of 4T1 tumor xenografts in mice using ZnFe-MMO as a tracer showed that the mean signal intensity is 1.27-fold higher than the commercial tracer VivoTrax at 72 h post-injection, indicating ZnFe-MMO's promise for prolonged MPI imaging applications.

Dermatological Nanotechnology: Gelatin films with O/W emulsions for skin lesion repair.

The development of films, scaffolds, hydrogels, and other innovations based on biopolymers for the treatment of skin injuries is on the rise. Therefore, it is important to focus on their functionality, influence on human use, and environmental impact. This work investigates the antimicrobial capacity of gelatin films that incorporate O/W emulsions encapsulating bactericidal and healing active ingredients (EA). Their biocompatibility was evaluated in vitro in human skin keratinocyte and murine fibroblast cell cultures, as well as in vivo using the zebrafish model. Finally, its potential to heal wounds was assessed through a keratinocyte cell migration assay. The EA films exhibited antimicrobial activity against Pseudomonas aeruginosa and Staphylococcus aureus. The films were biocompatible with monolayer cultures, without affecting cell viability, metabolic activity, or membrane integrity. The films did not exhibit general toxicological effects in zebrafish nor specific organ toxicity in the heart, liver, or brain. Further, the EA films promoted keratinocyte migration in the wound healing assay. In conclusion, the films could be used as a potential treatment for various types of skin injuries, being safe for both potential human application and the environment after use and disposal.

Cytotoxicity test of Bovine Gelatin- based hydrogel with Hibiscus rosa-sinensis L. leaf extract in topical drug development

Introduction: Herbal drugs have healing properties to prevent and treat various health conditions, including topical treatments, which contain therapeutic active ingredients like Hibiscus rosa-sinensis L. leaf extract combined with a carrier substance such as bovine gelatin hydrogel. Hibiscus rosa-sinensis L. extract also exhibits other pharmacological effects including antioxidant, antibacterial, antifungal, anti- inflammatory, antidiabetic, and anticancer properties. Bovine gelatin-based hydrogels have non-immunogenic properties derived from natural materials, with controlled drug release, as well as good biodegradability and biocompatibility, making them non-toxic. The combination of bovine gelatin hydrogel with Hibiscus rosa-sinensis L. leaf extract has been studied to develop a new topical drug. This research focuses on assessing the safety of this combination through cytotoxicity testing. Materials and Methods: This in vitro experimental study, utilized a post-test only control group design. The control group utilizes a bovine gelatin-based hydrogel, while the treatment group uses the same hydrogel combined with Hibiscus rosa-sinensis L. leaf extract on BHK-21 fibroblast cell cultures. Preclinical testing using the MTT assay served as a cytotoxicity test to validate the safety of this topical drug. Comparative study using a T-test with a significance level of p<0.05. Data analysis was carried out using Statistical Product and Service Solution (SPSS) Software for Mac version 29. Results: The cytotoxicity test results for bovine gelatin hydrogel and the combination with the extract of Hibiscus rosa-sinensis L. leaves were shown in the percentage of viable cells of 92.3% and 96.4%. The parametric T-test revealed no significant difference between the two groups (p = 0.754). Conclusions: The combination of bovine gelatin hydrogel with the extract of Hibiscus rosa-sinensis L. leaves in the development of herbal drugs are non-toxic

Heterophase CuO/Cu/Fe nanoparticle incorporated polyurethane coating for combating catheter-induced infection

To combat biofilm formation on the surface of medical devices, a new coatings based on polyurethane filled with heterophase nanoparticles CuO/Cu/Fe composition have been developed. Due to the synergistic antibacterial effect of the nanoparticle constituents, the obtained coatings demonstrated exceptional antibacterial activity. Thus the coating containing 5 % wt. or more of the CuO/Cu/Fe nanoparticles was at least 99.99 % against bacteria S. aureus, A. bumannii, K. pneumonia, P. aeruginosa. For this coating, the value of surface energy was maximum, indicating an increase in its hydrophobicity. The operational life of the catheter samples was more than 30 days, which exceeds the operational life of standard urinary catheters. Based on the results obtained, the application of the developed coating is a promising strategy for solving the catheter-associated urinary tract infection problem. Good biocompatibility was determined in vitro using 3T3 fibroblast cell culture, which is highly sensitive to toxic effects. Compared to other strategies of catheter surface modification, the use of polyurethane coating with nanoparticles of complex composition has a number of advantages: high antibacterial activity of nanoparticles due to the synergistic effect of constituents; biocompatibility of the coating provided by the use of biometals; long-term service life due to good adhesion of the coating to the standard silicon Foley catheter. Thus, the proposed coating containing nanoparticles effectively prevents bacterial colonization on catheters or other implants, thereby reducing the risk of postoperative catheter-induced infection.

Impact of various endodontic sealers on HPDLF Cell viability and apoptosis

This study aimed to investigate the cytotoxicity and apoptotic activity of different endodontic sealers: Sealapex, Apexit Plus, AH Plus, MTA-Fillapex and TotalFill BC Sealer in the culture of human periodontal ligament fibroblast (HPDLF) cells. The sealers were mixed, set for 24 h, and then covered with culture medium to obtain extracts, which were diluted to 1:0, 1:1, 1:2, 1:4, and 1:8. Simultaneously, HPDLF cells (1 × 104) were seeded in 96-well plates and incubated for 24 h at 37 °C 5% CO2 conditions. The cells were then exposed to 100 µL of diluted extract medium. Cytotoxicity was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to check cell viability, while apoptosis was assessed by TUNEL assay. Statistical analyses were performed using Kruskal-Wallis and Dunn post hoc tests with Mann Whitney U. In MTT and TUNEL assay cells were treated with sealers both 24 and 72 h. All materials showed higher toxicity at 72 h compared to 24 h. AH Plus exhibited the highest cytotoxicity, followed by MTA-Fillapex, Apexit Plus, Sealapex, while TotalFill BC Sealer had the lowest cytotoxicity. Consuquently, it was considered that TotalFill BC Sealer had the lowest cytotoxic potency when compared to other sealers, so it can be considered biocompatible.

Anti-inflammatory, cytotoxic and morphological impact of phycocyanin on ultraviolet radiation irradiated human fibroblast cells

Background Recently, the use of natural products as skin photoprotective agents has been in increasing demand. This study investigated the bioactivity of phycocyanin (PC) extracted from Spirulina sp. on human skin fibroblast cell line (CCD-966SK), specifically focusing on apoptosis, necrosis, anti-inflammatory effects, and enzymatic reactions. Methods The first step of this study was cyanobacterial cell culture and the extraction and purification of PC. After that, CCD-966SK cell line was cultivated under normal and UV irradiation. The bioassays included the cytotoxicity measurement, cell viability assay, morphology determination, tumour necrosis factor-α and Interleukin 6 release assays, enzyme activity for superoxide dismutase and glutathione peroxidase as well as malondialdehyde content and the cell-free extract of cyanobacterial stains were assessed. Results The cell viability results showed that as the concentration of the PC increased, the viability of CCD-966SK cell line was reduced, which suggested that the effect of PC on the growth of fibroblast cells was dose dependent. The morphological results indicated that presence of PC in the fibroblast cell culture medium led to a transformation in cell morphology from spindle-shaped to spherical. PC released anti-inflammatory IL-6 and TNF-a cytokines, indicating high inflammation resistance. Furthermore, the findings revealed that PC dramatically reduced the release of superoxide dismutase, glutathione peroxidase, and malondialdehyde from inflammatory cells, with the reduction being more apparent at increasing doses. Conclusions In conclusion, the results indicated that PC inhibit the CCD-966SK cell line by membrane destructor, which led to the increase the leakage of cell constituent and increase enzymes activities.

Development of Technology for the Synthesis of Nanocrystalline Cerium Oxide Under Production Conditions with the Best Regenerative Activity and Biocompatibility for Further Creation of Wound-Healing Agents.

Background/Objectives: The issue of effective wound healing remains highly relevant. The objective of the study is to develop an optimal method for the synthesis of nanosized cerium oxide powder obtained via the thermal decomposition of cerium carbonate precipitated from aqueous nitrate solution for the technical creation of new drugs in production conditions; the select modification of synthesis under different conditions based on the evaluation of the physicochemical characteristics of the obtained material and its biological activity, and an evaluation of the broad-spectrum effect on cells involved in the regeneration of skin structure as well as antimicrobial properties. Methods: Several modes of the industrial synthesis of cerium dioxide nanoparticles (NPs) were carried out. The synthesis stages and the chemical and physical parameters of the obtained NPs were described using transmission electron microscopy (TEM), X-ray diffraction, Raman spectroscopy, and mass spectrometry. The cell cultures of human fibroblasts and keratinocytes were cultured with different concentrations of different nanoceria variations, and the cytotoxicity and the metabolic and proliferative activity were investigated. An MTT test and cell counting were performed. The antimicrobial activity of CeO2 variations at a concentration of 0.1-0.0001 M against Pseudomonas aeruginosa was studied. Results: The purity of the synthesized nanoceria powders in all the batches was >99.99%. According to TEM data, the size of the NPs varied from 1 nm to 70 nm under different conditions and methodologies. The most optimal technology for the synthesis of the nanoceria with the maximum biological effect was selected. A method for obtaining the most bioactive NPs of optimal size (up to 10 nm) was proposed. The repeatability of the results of the proposed method of nanoceria synthesis in terms of particle size was confirmed. It was proven that the more structural defects on the surface of the CeO2 crystal lattice, the higher the efficiency of the NPs due to oxygen vacancies. The strain provided the best redox activity and antioxidant properties of the nanoceria, which was demonstrated by better regenerative potential on various cell lines. The beneficial effect of synthesized nanoceria on the proliferative and metabolic activity of the cell lines involved in skin regeneration (human fibroblasts, human keratinocytes) was demonstrated. The antimicrobial effect of synthesized nanoceria on the culture of the most-resistant-to-modern-antibiotics microorganism Pseudomonas aeruginosa was confirmed. The optimal concentrations of the nanoceria to achieve the maximum biological effect were determined (10-3 M). Conclusions: It was possible to develop a method for the industrial synthesis of nanoceria, which can be used to produce drugs and medical devices containing CeO2 NPs.

Arginine-Biofunctionalized Ternary Hydrogel Scaffolds of Carboxymethyl Cellulose-Chitosan-Polyvinyl Alcohol to Deliver Cell Therapy for Wound Healing.

Wound healing is important for skin after deep injuries or burns, which can lead to hospitalization, long-term morbidity, and mortality. In this field, tissue-engineered skin substitutes have therapy potential to assist in the treatment of acute and chronic skin wounds, where many requirements are still unmet. Hence, in this study, a novel type of biocompatible ternary polymer hybrid hydrogel scaffold was designed and produced through an entirely eco-friendly aqueous process composed of carboxymethyl cellulose, chitosan, and polyvinyl alcohol and chemically cross-linked by citric acid, forming three-dimensional (3D) matrices, which were biofunctionalized with L-arginine (L-Arg) to enhance cellular adhesion. They were applied as bilayer skin biomimetic substitutes based on human-derived cell cultures of fibroblasts and keratinocytes were seeded and grown into their 3D porous structures, producing cell-based bio-responsive hybrid hydrogel scaffolds to assist the wound healing process. The results demonstrated that hydrophilic hybrid cross-linked networks were formed via esterification reactions with the 3D porous microarchitecture promoted by foam templating and freeze-drying. These hybrids presented chemical stability, physicochemical properties, high moisture adsorption capacity, surface properties, and a highly interconnected 3D porous structure well suited for use as a skin substitute in wound healing. Additionally, the surface biofunctionalization of these 3D hydrogel scaffolds with L-arginine through amide bonds had significantly enhanced cellular attachment and proliferation of fibroblast and keratinocyte cultures. Hence, the in vivo results using Hairless mouse models (an immunocompromised strain) confirmed that these responsive bio-hybrid hydrogel scaffolds possess hemocompatibility, bioadhesion, biocompatibility, adhesiveness, biodegradability, and non-inflammatory behavior and are capable of assisting the skin wound healing process.

Molecular characterization and immunopathological investigation of Avian reticuloendotheliosis virus in breeder flocks in Egypt

BackgroundReticuloendotheliosis virus (REV) is an oncogenic immunosuppressive retrovirus that infects different kinds of avian species; posing significant economic losses to the poultry industry worldwide.MethodsIn Egypt, there is an unidentified disease associated with the runting-stunting syndrome with neoplasia, suspected to be REV, that has been continuously monitored in several breeder flocks. To diagnose and analyze REV by cell cultures, enzyme-linked immunosorbent assay (ELISA), histopathological investigation, the polymerase chain reaction (PCR) test, and sequencing analysis, 200 blood samples, and 50 tissue specimens were collected. The current study targets the occurrence and genetic characteristics of a viral neoplastic disease, resembling REV infection, circulating in breeder flocks from 2022 to 2023 in the Ismailia, El-Sharqia, and El-Dakahliya governorates.ResultHere, REV was isolated on chicken embryo fibroblast cell culture; exhibiting cell aggregation, rounding, and cell detachments. Collectively, only 70 serum samples were positive for anti‐REV antibodies with seroprevalence rates of 35% based on the ELISA test. The histopathological observation demonstrated lymphoreticular tumors in the liver, spleen, and other examined organs. The immunohistochemical staining method confirmed the REV-positive signals in all examined organs (liver, kidney, spleen, bursa, ovaries) except for the heart. The PCR assay of the LTR gene assessed 370 base pairs with only 5 positive samples with a percentage of 16.6%. Three positive samples were further sequenced and submitted to the Genbank under accession numbers (PP763709, PP763710, PP763711). Phylogenetic analysis of the REV-LTR gene showed that our three isolates (Sharquia-1-REV, Ismilia-2-REV, Mansoura-3-REV) are REV subtype III which predominantly circulated in breeders in Egypt. These three isolates are highest similar to American, Chinese, and Taiwanese REV reference strains, and other Egyptian strains with nucleotide identity percentages of 100%, 99%, and 99%; respectively, and on the amino acid identity level were with (99–100%), (98%, 99%), (99%, 100%); respectively.ConclusionsThis study established that REV infection was extensively distributed in the breeders and became one of the causes of the clinical outbreaks of tumors, raising awareness of REV as the causative agent of avian oncogenic disease in Egypt.

Innovative synthesis and molecular modeling of actinomycetes-derived silver nanoparticles for biomedical applications

The rising demand for innovative antimicrobial solutions has shifted focus towards silver nanoparticles (AgNPs), especially those produced through eco-friendly methods. This study introduces a novel approach utilizing actinomycetes strains—Streptomyces albus, Micromonospora maris, and Arthrobacter crystallopoietes—to biosynthesize AgNPs with remarkable antibacterial properties. Through molecular characterization, we identified unique features of these nanoparticles, and computational modeling suggested significant ion-ligand interactions with proteins 6REV and 3K07. Our research highlights the promise of these biogenically synthesized nanoparticles in advancing biomedical applications. Actinomycetes were sourced and screened for their ability to produce metallic nanoparticles, revealing that among 35 samples, only six showed this capability. Notably, Streptomyces albus strain smmdk14 (OR685674), Micromonospora maris strain smmdk13 (OR685672), and Arthrobacter crystallopoietes strain smmdk12 (OR685674) were identified as effective silver nanoparticle producers. The synthesized nanoparticles demonstrated potent antibacterial activity against common pathogens including E. coli, Pseudomonas aeruginosa, Klebsiella spp., Enterococcus faecalis, Staphylococcus aureus, and Acinetobacter spp. The data obtained from color change observation, UV–visible spectrophotometry, Zeta potential, FTIR spectroscopy, and transmission electron microscopy (TEM) characterized AgNPs potentiality. The nanoparticles were spherical, with sizes ranging from 6.46 nm to 24.7 nm. Optimization of production conditions, comparison of antimicrobial effects with antibiotics, evaluation of potential toxicity, and assessment of wound-healing capabilities were also conducted. The biosynthesized AgNPs exhibited superior antibacterial properties compared to traditional antibiotics and significantly accelerated wound healing by approximately 66.4 % in fibroblast cell cultures. Additionally, computational analysis predicted interactions between various metal ions and specific amino acid residues in proteins 6REV and 3K07. Overall, this study demonstrates the successful creation of AgNPs with notable antibacterial and wound-healing properties, underscoring their potential for medical applications.

Cromolyn sodium and masitinib combination inhibits fibroblast-myofibroblast transition and exerts additive cell-protective and antioxidant effects on a bleomycin-induced invitro fibrosis model.

Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal fibrotic lung disease. While recent studies have suggested the potential efficacy of tyrosine kinase inhibitors in managing IPF, masitinib, a clinically used tyrosine kinase inhibitor, has not yet been investigated for its efficacy in fibrotic lung diseases. In a previous study on an invitro neurodegenerative model, we demonstrated the synergistic antitoxic and antioxidant effects of masitinib combined with cromolyn sodium, an FDA-approved mast cell stabilizer. This study aims to investigate the anti-fibrotic and antioxidant effects of the masitinib-cromolyn sodium combination in an invitro model of pulmonary fibrosis. Fibroblast cell cultures treated with bleomycin and/or hydrogen peroxide (H2O2) were subjected to masitinib and/or cromolyn sodium, followed by assessments of cell viability, morphological and apoptotic nuclear changes, triple-immunofluorescence labeling, and total oxidant/antioxidant capacities, besides ratio of Bax and Bcl-2 mRNA expressions as an indication of apoptosis. The combined treatment of masitinib and cromolyn sodium effectively prevented the fibroblast myofibroblast transition, a hallmark of fibrosis, and significantly reduced bleomycin / H2O2-induced apoptosis and oxidative stress. This study is the first to demonstrate the additive anti-fibrotic, cell-protective, and antioxidant effects of the masitinib-cromolyn sodium combination in an invitro fibrosis model, suggesting its potential as an innovative therapeutic approach for pulmonary fibrosis. Combination therapy may be more advantageous in that both drugs could be administered in lower doses, exerting less side effects, and at the same time providing diverse mechanisms of action simultaneously.

Autologous Cultured Fibroblast Cell Therapy for Treatment of Stria Alba

Abstract Background There is growing evidence showing that human autologous cultured fibroblast transplantation is a safe alternative treatment to prevent and reduce the structural changes in aging skin. Previous data indicate that injections of autologous fibroblasts increase collagen formation, accompanied by a concomitant increase in thickness and density of dermal collagen. our study will highlight the technique of autologous cultured fibroblast and it is indication for treatment of stria alba and this wasn't done before. Objective To evaluate the efficacy and safety of autologous cultured fibroblast for improvement of stria alba. Patients and Methods The present study included 15 patients a with stria alba. The study was approved by the Research Ethical Committee, Faculty of Medicine, Ain Shams University and fulfilled all the ethical aspects required in human research. All patients received full information about description of the procedure of treatment, possible side effects, photo documentation and they all provided written consent. the patients were injected intradermally by autologous fibroblast suspension at 2 weeks intervals with total of 3 sessions. Results Before autologous fibroblast injection (baseline), the width of stria ranged from 4-17.4 mm (mean 8.18 ± 3.85). After treatment at 3rd month, stria ranged from 2.9 – 14.8 (mean 7.17 ± 3.26) and at the 6th month ranged from 2.9 – 14.8 (mean 7.16 ± 3.27). Comparison of width of stria before versus after injection showed non significant difference (p value 0.062)On contrast, the histopathological examination by H&E, Masson Trichome and Orcein stain a showed significant improvement in the form of increased epidermal thickness, better collagen arrangement and thickness and increased elastic fibers contents. Conclusion Autologous fibroblast injection in treatment of stria alba may have promising results in the future especially if combined with other tretment modalities.

Toll-like Receptor 9 Inhibition Mitigates Fibroproliferative Responses in Translational Models of Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease for which current treatment options only slow clinical progression. Previously, we identified a subset of patients with IPF with an accelerated disease course associated with fibroblast expression of Toll-Like Receptor 9 (TLR9) mediated by interactions with its ligand mitochondrial DNA (mtDNA). We aimed to show that TLR9 activation induces fibroproliferative responses that are abrogated by its antagonism by using two commercially-available indirect inhibitors and a proprietary, selective direct small molecule inhibitor. We employed two independent cohorts of patients with IPF, multiple in vitro fibroblast cell culture platforms, an in vivo mouse model, and an ex vivo human precision cut lung slices system to investigate the clinical and biologic significance of TLR9 in this disease. In two independent IPF cohorts, plasma mtDNA activates TLR9 in a manner associated with the expression of MCP-1, IL-6, TNFα, and IP-10 and worsened transplant-free survival. Our cell culture platform showed that TLR9 mediates fibroblast activation via TGFβ1 and stiff substrates, and that its antagonism, particularly direct inhibition, ameliorates this process, including production of these TLR9 associated pharmacodynamic endpoints. We further demonstrated that direct TLR9 inhibition mitigates these fibroproliferative responses in our in vivo and ex vivo models of pulmonary fibrosis. In this novel study, we found that direct TLR9 inhibition mitigates fibroproliferative responses in preclinical models of pulmonary fibrosis. Our work demonstrates the therapeutic potential of direct TLR9 antagonism in IPF and related fibrotic lung diseases.

Exploring the complexity of systemic sclerosis etiology by trio whole genome sequencing.

Systemic sclerosis (SSc) is a heterogeneous rare autoimmune fibrosing disorder affecting connective tissue. The etiology of systemic sclerosis is largely unknown and many genes have been suggested as susceptibility loci of modest impact by genome-wide association study (GWAS). Multiple factors can contribute to the pathological process of the disease, which makes it more difficult to identify possible disease-causing genetic alterations. In this study, we have applied whole genome sequencing (WGS) in 101 indexed family trios, supplemented with transcriptome sequencing on cultured fibroblast cells of four patients and five family controls where available. Single nucleotide variants (SNVs) and copy number variants (CNVs) were examined, with emphasis on de novo variants. We also performed enrichment test for rare variants in candidate genes previously proposed in association with systemic sclerosis. We identified 42 exonic and 34 ncRNA de novo SNV changes in 101 trios, from a total of over 6000 de novo variants genome wide. We observed higher than expected de novo variants in PRKXP1 gene. We also observed such phenomenon along with increased expression in patient group in NEK7 gene. Additionally, we also observed significant enrichment of rare variants in candidate genes in the patient cohort, further supporting the complexity/multi-factorial etiology of systemic sclerosis. Our findings identify new candidate genes including PRKXP1 and NEK7 for future studies in SSc. We observed rare variant enrichment in candidate genes previously proposed in association with SSc, which suggest more efforts should be pursued to further investigate possible pathogenetic mechanisms associated with those candidate genes.

Anti-inflammatory activities of phycoerythrin and phycocyanin on human fibroblast cells

Background and AimCyanobacteria are highly regarded for their nutritional value and the scientific community has shown great interest in studying their biological activity and bioactive compounds, especially pigments. They have a wide range of properties, making them potentially useful in various health applications, including dermatology. Given the growing demand for natural products and actives in the cosmetics and medical sectors, these microorganisms have emerged as a valuable source. Therefore, this study investigated the bioactivity of phycoerythrin (PE) and phycocyanin (PC) extracted from Nostoc sp. and Spirulina sp. on CCD-966SK cell lines, specifically focusing on apoptosis, necrosis, anti-inflammatory effects, and enzymatic reactions. MethodsThe study involved conducting various bioassays on a human skin fibroblast cell line (CCD-966SK) after performing cyanobacterial cell culture, pigment extraction, and fibroblast cell culture. The bioassays included the MTT test, TNF-α and IL-6 release assays, measurement of human collagen I and III, assessment of apoptosis and necrosis, and analysis of enzyme activity for superoxide dismutase (SOD) and glutathione peroxidase (GPx). ResultsThe cytotoxicity test results indicated that higher concentrations of PE and PC were associated with lower survival rates. The flow cytometry analysis confirmed a significant increase in necrotic cells and a decrease in living cells at high concentrations of both pigments. The largest percentages of collagen I for PC and PE were identified at 125 and 62.5 ppm, whereas the lowest quantities were reported at 1000 ppm for both pigments. PC and PE had the highest collagen II concentrations at 3.91 and 15.63 ppm, respectively. PC and PE had the lowest collagen II levels at 1000 ppm. PE and PC released anti-inflammatory IL-6 and TNF-a cytokines, indicating high inflammation resistance. Superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity and malondialdehyde (MDA) concentrations showed that both pigments were most active at 1000 ppm. ConclusionIn conclusion, both PC and PE pigments exhibited positive effects in enhancing necrotic, anti-inflammatory, and enzymatic activity against the human skin fibroblast cell line.

Cerium Dioxide-Dextran Nanocomposites in the Development of a Medical Product for Wound Healing: Physical, Chemical and Biomedical Characteristics.

the creation of a dextran coating on cerium oxide crystals using different ratios of cerium and dextran to synthesize nanocomposites, and the selection of the best nanocomposite to develop a nanodrug that accelerates quality wound healing with a new type of antimicrobial effect. Nanocomposites were synthesized using cerium nitrate and dextran polysaccharide (6000 Da) at four different initial ratios of Ce(NO3)3x6H2O to dextran (by weight)-1:0.5 (Ce0.5D); 1:1 (Ce1D); 1:2 (Ce2D); and 1:3 (Ce3D). A series of physicochemical experiments were performed to characterize the created nanocomposites: UV-spectroscopy; X-ray phase analysis; transmission electron microscopy; dynamic light scattering and IR-spectroscopy. The biomedical effects of nanocomposites were studied on human fibroblast cell culture with an evaluation of their effect on the metabolic and proliferative activity of cells using an MTT test and direct cell counting. Antimicrobial activity was studied by mass spectrometry using gas chromatography-mass spectrometry against E. coli after 24 h and 48 h of co-incubation. According to the physicochemical studies, nanocrystals less than 5 nm in size with diffraction peaks characteristic of cerium dioxide were identified in all synthesized nanocomposites. With increasing polysaccharide concentration, the particle size of cerium dioxide decreased, and the smallest nanoparticles (<2 nm) were in Ce2D and Ce3D composites. The results of cell experiments showed a high level of safety of dextran nanoceria, while the absence of cytotoxicity (100% cell survival rate) was established for Ce2D and C3D sols. At a nanoceria concentration of 10-2 M, the proliferative activity of fibroblasts was statistically significantly enhanced only when co-cultured with Ce2D, but decreased with Ce3D. The metabolic activity of fibroblasts after 72 h of co-cultivation with nano composites increased with increasing dextran concentration, and the highest level was registered in Ce3D; from the dextran group, differences were registered in Ce2D and Ce3D sols. As a result of the microbiological study, the best antimicrobial activity (bacteriostatic effect) was found for Ce0.5D and Ce2D, which significantly inhibited the multiplication of E. coli after 24 h by an average of 22-27%, and after 48 h, all nanocomposites suppressed the multiplication of E. coli by 58-77%, which was the most pronounced for Ce0.5D, Ce1D, and Ce2D. The necessary physical characteristics of nanoceria-dextran nanocomposites that provide the best wound healing biological effects were determined. Ce2D at a concentration of 10-3 M, which stimulates cell proliferation and metabolism up to 2.5 times and allows a reduction in the rate of microorganism multiplication by three to four times, was selected for subsequent nanodrug creation.

Electrofluidic control for textile-based cell culture: Identification of appropriate conditions required to integrate cell culture with electrofluidics.

Electric field-driven microfluidics, known as electrofluidics, is a novel attractive analytical tool when it is integrated with low-cost textile substrate. Textile-based electrofluidics, primarily explored on yarn substrates, is in its early stages, with few studies on 3D structures. Further, textile structures have rarely been used in cellular analysis as a low-cost alternative. Herein, we investigated novel 3D textile structures and develop optimal electrophoretic designs and conditions that are favourable for direct 3D cell culture integration, developing an integrated cell culture textile-based electrofluidic platform that was optimised to balance electrokinetic performance and cell viability requirements. Significantly, there were contrasting electrolyte compositional conditions that were required to satisfy cell viability and electrophoretic mobility requiring the development of and electrolyte that satisfied the minimum requirements of both these components within the one platform. Human dermal fibroblast cell cultures were successfully integrated with gelatine methacryloyl (GelMA) hydrogel-coated electrofluidic platform and studied under different electric fields using 5mM TRIS/HEPES/300mM glucose. Higher analyte mobility was observed on 2.5% GelMA-coated textile which also facilitated excellent cell attachment, viability and proliferation. Cell viability also increased by decreasing the magnitude and time duration of applied electric field with good cell viability at field of up to 20Vcm-1.

Glycopolymer Inhibitors of Galectin-3 Suppress the Markers of Tissue Remodeling in Pulmonary Hypertension.

Pulmonary hypertension is a cardiovascular disease with a low survival rate. The protein galectin-3 (Gal-3) binding β-galactosides of cellular glycoproteins plays an important role in the onset and development of this disease. Carbohydrate-based drugs that target Gal-3 represent a new therapeutic strategy in the treatment of pulmonary hypertension. Here, we present the synthesis of novel hydrophilic glycopolymer inhibitors of Gal-3 based on a polyoxazoline chain decorated with carbohydrate ligands. Biolayer interferometry revealed a high binding affinity of these glycopolymers to Gal-3 in the subnanomolar range. In the cell cultures of cardiac fibroblasts and pulmonary artery smooth muscle cells, the most potent glycopolymer 18 (Lac-high) caused a decrease in the expression of markers of tissue remodeling in pulmonary hypertension. The glycopolymers were shown to penetrate into the cells. In a biodistribution and pharmacokinetics study in rats, the glycopolymers accumulated in heart and lung tissues, which are most affected by pulmonary hypertension.

Co electrospinning -poly (vinyl alcohol)-chitosan/gelatin-poly (ϵ-caprolacton) nanofibers for diabetic wound-healing application

With the increasing prevalence of diabetes, the healing of diabetic wounds has become a significant challenge for both healthcare professionals and patients. Recognizing the urgent need for effective solutions, it is crucial to develop suitable scaffolds specifically tailored for diabetic wound healing. In line with this objective, we have developed novel hybrid nanofibrous scaffolds by combining polyvinyl alcohol/chitosan (PVA/CS) and gelatin/poly(ε-caprolactone) (Gel/PCL) polymers through a double-nozzle electrospinning technique. In this study, we investigated the influence of the Gel/PCL blend ratio on the properties of the resulting nanofibers. Three different hybrid scaffold structures were examined: Gel/PCL (80:20)-PVA/CS (80:20), Gel/PCL (50:50)-PVA/CS (80:20), and Gel/PVA (20:80)-PVA/CS (80:20). Our findings demonstrate that the electrospun nanofibers of PVA/CS (80:20)-Gel/PCL (80:20) exhibited optimal mechanical performance, with a contact angle of approximately 54° and a diameter of 183 nm. Considering the crucial role of inhibiting bacterial adhesion in the success of implanted materials, we evaluated the cytocompatibility of the hybrid electrospun nanofibers using mouse fibroblast cells (L-929 cells). The in vitro cytotoxicity results obtained from L-929 fibroblast cell culture on the hybrid scaffolds revealed enhanced cell proliferation and appropriate cell morphology on the PVA/CS (80:20)-Gel/PCL (80:20) sample, indicating its capability to support tissue cell integration. Based on the information obtained from this study, the fabricated hybrid scaffold holds great promise for diabetic ulcer healing. Its optimal mechanical properties, suitable contact angle, and favorable cytocompatibility highlight its potential as a valuable tool in the field of diabetic wound healing. The development of such hybrid scaffolds represents a significant step forward in addressing the challenges associated with diabetic wound care.