ABSTRACT: Transcriptomic fingerprints of C. elegans exposed to superparamagnetic iron oxide nanoparticles coated with a monolayer of bovine serum albumin (BSA-SPIONs)
Project description:Transcriptomic fingerprints of C. elegans exposed to citrate coated superparamagnetic iron oxide nanoparticles (C-SPIONs) and to superparamagnetic iron oxide nanoparticles coated with a monolayer of bovine serum albumin (BSA-SPIONs)
Project description:Superparamagnetic Iron Oxide Nanoparticles (SPIONs) are currently being investigated for a range of biomedical applications. Their use have been related with different cytotoxic mechanisms including the generation of oxidative stress and the induction of metal detoxification pathways, among others. Different NP coatings are being explored, among them albumin which has been applied in some drugs delivery systems. We have investigated the molecular mechanisms responsive to in-house fabricated SPIONs coated with bovine serum albumin (BSA-SPIONs) in the nematode C. elegans to compare in vivo findings with previous in vitro studies. BSA-SPIONs (500 µg/ml) affected the transcriptional response of glycan metabolic pathways related to innate immune response, xenobiotics degradation, and triggered a lysosomal response, indicating a relevant biological role of this cellular compartment in the response to this nanoparticle treatment in C. elegans. Remarkably, key biological functions such as apoptosis or protein processing were not affected with significance despite the high dose of exposure.
Project description:Bloodstream trypanosomes are sensitive to iron starvation, and upregulate ESAG6/7 transferrin receptor (TfR) mRNA and protein levels when placed in low transferrin or iron-depleted media, and adjust TfR expression to compensate for reduced endocytosis. Both observations suggest a specific transcriptional-level response resulting from an iron-sensing mechanism. The transcriptomes of BSFs cultured in different proportions of fetal bovine serum (FBS) were analysed. <br><br>part 1: 4 biological replicates of SMB cells grown with 10% FBS (normal conditions) and 4 replicates of SMB cells grown with 30% FBS, as well as dye swaps, were used. <br><br>part 2: 3 biological replicates of SMB cells grown with 10% FBS (normal conditions) and 3 replicates of SMB cells grown without FBS (but supplemented with 5 mg/ml BSA to simulate physiological concentrations of albumin in media with 10% FBS), as well as dye swaps were used. <br><br>part 3: 3 biological replicates of SMB cells grown with 10% FBS (normal conditions) and 3 replicates of SMB cells grown without FBS (but supplemented with 5 mg/ml BSA and 0.3 mg/ml bovine holo-transferrin to simulate physiological concentrations of albumin and transferrin in media with 10% FBS), as well as dye swaps were used.
Project description:Due to the characteristics of multi-walled carbon nanotubes (MWCNTs) to aggregate and bind non-specifically, attempts have been made to inhibit their aggregation and increase their dispersion stability for biomedical applications by coating with bovine serum albumin (BSA). However, BSA concentration has not been studied in detail in terms of cellular response. To investigate the surface proteins formed in 10% fetal bovine serum cell culture media depending on the BSA concentration used to stabilize MWCNTs and to correlate the cellular responses with BSA concentration, we performed nanoflow liquid chromatography–electrospray ionization–tandem mass spectrometry analysis to quantify proteins of the protein corona (PC) and of A549 cells. Results showed that increasing the concentration of pre-incubated BSA changed the composition ratio of the proteins of the PCs adsorbed on the MWCNTs and also decreased the cellular uptake of A549 cells treated with BSA-coated MWCNTs. In the proteomic analysis, proteins related to the ribosome, ribosome biogenesis in eukaryotes, and oxidative phosphorylation pathways were downregulated in the low concentration BSA-treated cell groups, while proteins related to the mRNA surveillance pathway were downregulated in the high-BSA-concentration cell groups. Moreover, circular dichroism and Fourier-transform infrared spectroscopy results showed that the α-helix structure, a secondary structure of the adsorbed proteins, increased as a function of BSA concentration. Taken together, BSA concentration affects the adsorption and secondary structure of PCs on MWCNTs, which in turns affect their intracellular behavior. Our results may provide insight for the design of modification methods to increase the biocompatibility of MWCNTs.
Project description:To test our peak detection and label-free quantitation algorithm, practical applications of AB3D for LC-MS data sets were evaluated using a standard peptide mixture consisting of bovine serum albumin (BSA) peptides with different concentrations, peptides derived from four standard proteins (data set 1), and complex peptide samples derived from HeLa cells and BSA (data set 2).
Project description:Lactoperoxidase (LPO) is a component of milk and other external secretions. Recently, we found antiinflammatory effect of orally administered LPO in mouse model of influenza virus-induced pneumonia. In order to study influence of bovine LPO on the digestive tract as its possible target organ, we performed comprehensive gene expression analysis of the mouse small intestine using GeneChip microarray to compare the effect of LPO with that of water or bovine serum albumin (BSA). We observed that 3 h after single ingestion of LPO the levels of 78 gene expression are upregulated and those of 9 gene expressions are downregulated. Within most changed genes, 5 upregulated and 1 downregulated genes such as FK506 binding protein 5 (FKBP5) and serum/glucocorticoid regulated kinase (SGK) are known to be transcriptionally modulated by glucocorticoid. This result suggests that ingested LPO modulates gene expressions in the small intestine in a glucocorticoid-like manner and this activity may link to its systemic antiinflammatory effects. Experiment Overall Design: Mice were orally administered 200 microL of water (n=3), bovine serum albumin (BSA, 2.5 g/kg body weight, n=3), or bovine lactoperoxidase (LPO, 2.5 g/kg body weight, n=3). Three hours after administration, comprehensive gene expressions of jejunum detached payer's patch were analysed for comparison of LPO administration with water or BSA administration.
Project description:We determined the composition of the plasma protein corona on silica-coated versus dextran-coated iron oxide nanoparticles using mass spectrometry-based proteomics approaches. Gene ontology (GO) enrichment analysis and Ingenuity Pathway Analysis (IPA) revealed distinct protein corona compositions for the two different SPIONs. Relaxivity of silica-coated SPIONs was modulated by the presence of a protein corona. Moreover, the viability of primary human monocyte-derived macrophages was influenced by the protein corona on silica-coated, but not dextran-coated SPIONs, and the protein corona promoted cellular uptake of silica-coated SPIONs, but did not affect internalization of dextran-coated SPIONs.
Project description:Whole fetal livers were collected from mouse fetuses at embryonic day 14.5 (E14.5), and single-cell suspensions were prepared by successive passage through 18-, 21 and 23-gauge needles. Fetal liver cells were maintained in Dulbecco modified Eagle medium (DMEM; Invitrogen) supplemented with 10% fetal bovine serum (FBS; Invitrogen), 100 U/ml penicillin, 100µg/ml streptomycin, and 50ng/ml recombinant human thrombopoietin (TPO; Peprotech). After 5 days of culture, megakaryocytes were purified using a discontinuous bovine serum albumin gradient (BSA, Sigma–Aldrich; 3%, 1.5%, and 0%). Total RNA was isolated with Tri–Reagent (MRC) following manufacturer’s instructions, and its quality was assessed with ND–1000 Nanodrop (Peqlab) and on a 1.5% agarose gel.
