BioMIPs: molecularly imprinted silk fibroin nanoparticles to recognize the iron regulating hormone hepcidin.
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ABSTRACT:
SUBMITTER: Bossi AM
PROVIDER: S-EPMC8782820 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Nanosized biomimetics prepared by the strategy of molecular imprinting, that is, the stamping of recognition sites by means of a template-assisted synthesis, are demonstrating potential as plastic antibodies in medicine, proving effective for cell imaging and targeted therapies. Most molecularly imprinted nanoparticles (MIP-NPs) are currently made of soft matter, such as polyacrylamide and derivatives. Yet, MIP-NPs biocompatibility is crucial for their effective translation into clinical uses. Here, we propose the original idea to synthesize fully biocompatible molecularly imprinted nanoparticles starting from the natural polymer silk fibroin (MIP SF-NPs), which is nontoxic and highly biocompatible. The conditions to produce MIP SF-NPs of different sizes (dmean ∼ 50 nm; dmean ∼ 100 nm) were set using the response surface method. The stamping of a single, high affinity (KD = 57 × 10-9 M), and selective recognition site per silk fibroin nanoparticle was demonstrated, together with the confirmation of nontoxicity. Additionally, MIP SF-NPs were used to decorate silk microfibers and silk nanofibers, providing a general means to add entailed biofunctionalities to materials.
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Project description:Quantification and characterisation of canine hepcidin by LC-MS
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Project description:Cartilage defects frequently occur around the knee joint yet cartilage has limited self-repair abilities. Hydrogel scaffolds have excellent potential for use in tissue engineering. Therefore, the aim of the present study was to assess the ability of silk fibroin (SF) hydrogel scaffolds incorporated with chitosan (CS) nanoparticles (NPs) to repair knee joint cartilage defects. In the present study, composite systems of CS NPs incorporated with transforming growth factor-β1 (TGF-β1; TGF-β1@CS) and SF incorporated with bone morphogenetic protein-2 (BMP-2; TGF-β1@CS/BMP-2@SF) were developed and characterized with respect to their size distribution, zeta potential, morphology, and release of TGF-β1 and BMP-2. Bone marrow stromal cells (BMSCs) were co-cultured with TGF-β1@CS/BMP-2@SF extracts to assess chondrogenesis in vitro using a cell counting kit-8 assay, which was followed by in vivo evaluations in a rabbit model of knee joint cartilage defects. The constructed TGF-β1@CS/BMP-2@SF composite system was successfully characterized and showed favorable biocompatibility. In the presence of TGF-β1@CS/BMP-2@SF extracts, BMSCs exhibited normal cell morphology and enhanced chondrogenic ability both in vitro and in vivo, as evidenced by the promotion of cell viability and the alleviation of cartilage defects. Thus, the TGF-β1@CS/BMP-2@SF hydrogel developed in the present study promoted chondrogenic ability of BMSCs both in vivo and in vitro by releasing TGF-β1 and BMP-2, thereby offering a novel therapeutic strategy for repairing articular cartilage defects in knee joints.
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Project description:Hepcidins are an evolutionarily conserved class of liver-expressed peptide, from which the twenty-five amino acid hormone, hepcidin-25 (herein hepcidin), has gained significant notoriety as the master regulator of iron homeostasis in mammals. Hepcidin maintains iron homeostasis by controlling the dietary absorption of iron and the mechanisms of recycling cellular iron stores. With the physiological significance of this hormone well established, it has emerged as an informative biomarker. In a comparison of the genome, transcriptome and peptidome of Canis lupis familiaris, we reveal the size of the hepcidin peptide in the canine, previous reports of which were contradictory to the evolutionary conservation predicted by genome annotation. Here, measurement of the peptide by mass spectrometry, following isolation from greyhound blood serum, revealed an amino acid sequence and peptide mass, differing from all accounts to date, yet demonstrating perfect sequence identity to that of the greater Canidae lineage of the Carnivora. Importantly, in the greyhound, the measured hepcidin peptide showed a similar temporal pattern to total serum iron, consistent with our understanding of hepcidin regulating iron homeostasis, in agreement with human diagnostics, and providing added translational evidence of the measured peptide being the iron regulatory hormone of the Canidae.
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Project description:In recent years, silk fibroin nanoparticles (SFNs) have been consolidated as drug delivery systems (DDSs) with multiple applications in personalized medicine. The design of a simple, inexpensive, and scalable preparation method is an objective pursued by many research groups. When the objective is to produce nanoparticles suitable for biomedical uses, their sterility is essential. To achieve sufficient control of all the crucial stages in the process and knowledge of their implications for the final characteristics of the nanoparticles, the present work focused on the final stage of sterilization. In this work, the sterilization of SFNs was studied by comparing the effect of different available treatments on the characteristics of the nanoparticles. Two different sterilization methods, gamma irradiation and autoclaving, were tested, and optimal conditions were identified to achieve the sterilization of SFNs by gamma irradiation. The minimum irradiation dose to achieve sterilization of the nanoparticle suspension without changes in the nanoparticle size, polydispersity, or Z-potential was determined to be 5 kiloGrays (kGy). These simple and safe methods were successfully implemented for the sterilization of SFNs in aqueous suspension and facilitate the application of these nanoparticles in medicine.
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Project description:BackgroundAbnormal expression of the liver peptide hormone hepcidin, a key regulator of iron homeostasis, contributes to the pathogenesis of anemia in conditions such as inflammatory bowel disease (IBD). Since little is known about the mechanisms that control hepcidin expression during states of intestinal inflammation, we sought to shed light on this issue using mouse models.Methodology/principal findingsHepcidin expression was evaluated in two types of intestinal inflammation caused by innate immune activation-dextran sulfate sodium (DSS)-induced colitis in wild-type mice and the spontaneous colitis occurring in T-bet/Rag2-deficient (TRUC) mice. The role of tumor necrosis factor (TNF) ? was investigated by in vivo neutralization, and by treatment of a hepatocyte cell line, as well as mice, with the recombinant cytokine. Expression and activation of Smad1, a positive regulator of hepcidin transcription, were assessed during colitis and following administration or neutralization of TNF?. Hepcidin expression progressively decreased with time during DSS colitis, correlating with changes in systemic iron distribution. TNF? inhibited hepcidin expression in cultured hepatocytes and non-colitic mice, while TNF? neutralization during DSS colitis increased it. Similar results were obtained in TRUC mice. These effects involved a TNF?-dependent decrease in Smad1 protein but not mRNA.Conclusions/significanceTNF? inhibits hepcidin expression in two distinct types of innate colitis, with down-regulation of Smad1 protein playing an important role in this process. This inhibitory effect of TNF? may be superseded by other factors in the context of T cell-mediated colitis given that in the latter form of intestinal inflammation hepcidin is usually up-regulated.
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