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Sökning: WFRF:(Bergström Lind Sara)

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1.
  • Hernroth, Bodil, 1951-, et al. (författare)
  • Impact of ocean acidification on antimicrobial activity in gills of the blue mussel (Mytilus edulis)
  • 2016
  • Ingår i: Fish and Shellfish Immunology. - : Elsevier. - 1050-4648 .- 1095-9947. ; 55, s. 452-459
  • Tidskriftsartikel (refereegranskat)abstract
    • Here, we aimed to investigate potential effects of ocean acidification on antimicrobial peptide (AMP) activity in the gills of Mytilus edulis, as gills are directly facing seawater and the changing pH (predicted to be reduced from ∼8.1 to ∼7.7 by 2100). The AMP activity of gill and haemocyte extracts was compared at pH 6.0, 7.7 and 8.1, with a radial diffusion assay against Escherichia coli. The activity of the gill extracts was not affected by pH, while it was significantly reduced with increasing pH in the haemocyte extracts. Gill extracts were also tested against different species of Vibrio (V. parahaemolyticus, V. tubiashii, V. splendidus, V. alginolyticus) at pH 7.7 and 8.1. The metabolic activity of the bacteria decreased by ∼65–90%, depending on species of bacteria, but was, as in the radial diffusion assay, not affected by pH. The results indicated that AMPs from gills are efficient in a broad pH-range. However, when mussels were pre-exposed for pH 7.7 for four month the gill extracts presented significantly lower inhibit of bacterial growth. A full in-depth proteome investigation of gill extracts, using LC-Orbitrap MS/MS technique, showed that among previously described AMPs from haemocytes of Mytilus, myticin A was found up-regulated in response to lipopolysaccharide, 3 h post injection. Sporadic occurrence of other immune related peptides/proteins also pointed to a rapid response (0.5–3 h p.i.). Altogether, our results indicate that the gills of blue mussels constitute an important first line defence adapted to act at the pH of seawater. The antimicrobial activity of the gills is however modulated when mussels are under the pressure of ocean acidification, which may give future advantages for invading pathogens.
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2.
  • Kassa, Eszter, et al. (författare)
  • Evaluation of affinity-purification coupled to mass spectrometry approaches for capture of short linear motif-based interactions
  • 2024
  • Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
    • Low affinity and transient protein-protein interactions, such as short linear motif (SLiM)-based interactions, require dedicated experimental tools for discovery and validation. Here, we evaluated and compared biotinylated peptide pulldown and protein interaction screen on peptide matrix (PRISMA) coupled to mass-spectrometry (MS) using a set of peptides containing interaction motifs. Eight different peptide sequences that engage in interactions with three distinct protein domains (KEAP1 Kelch, MDM2 SWIB, and TSG101 UEV) with a wide range of affinities were tested. We found that peptide pulldown can be an effective approach for SLiM validation, however, parameters such as protein abundance and competitive interactions can prevent the capture of known interactors. The use of tandem peptide repeats improved the capture and preservation of some interactions. When testing PRISMA, it failed to provide comparable results for a model peptide that successfully pulled down a known interactor using biotinylated peptide pulldown. Overall, in our hands, we find that albeit more laborious, biotin-peptide pulldown was more successful in terms of validation of known interactions. Our results highlight that the tested affinity-capture MS-based methods for validation of SLiM-based interactions from cell lysates are suboptimal, and we identified parameters for consideration for method development.
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3.
  • Kassa, Eszter, et al. (författare)
  • Evaluation of affinity-purification coupled to mass spectrometry approaches for capture of short linear motif-based interactions
  • 2023
  • Ingår i: Analytical Biochemistry. - : Elsevier. - 0003-2697 .- 1096-0309. ; 663
  • Tidskriftsartikel (refereegranskat)abstract
    • Low affinity and transient protein-protein interactions, such as short linear motif (SLiM)-based interactions, require dedicated experimental tools for discovery and validation. Here, we evaluated and compared biotinylated peptide pulldown and protein interaction screen on peptide matrix (PRISMA) coupled to massspectrometry (MS) using a set of peptides containing interaction motifs. Eight different peptide sequences that engage in interactions with three distinct protein domains (KEAP1 Kelch, MDM2 SWIB, and TSG101 UEV) with a wide range of affinities were tested. We found that peptide pulldown can be an effective approach for SLiM validation, however, parameters such as protein abundance and competitive interactions can prevent the capture of known interactors. The use of tandem peptide repeats improved the capture and preservation of some interactions. When testing PRISMA, it failed to provide comparable results for model peptides that successfully pulled down known interactors using biotinylated peptide pulldown. Overall, in our hands, we find that albeit more laborious, biotin-peptide pulldown was more successful in terms of validation of known interactions. Our results highlight that the tested affinity-capture MS-based methods for validation of SLiM-based interactions from cell lysates are suboptimal, and we identified parameters for consideration for method development.
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4.
  • Valarcher, Jean-Francois, et al. (författare)
  • Single-Shot Vaccines against Bovine Respiratory Syncytial Virus (BRSV) : Comparative Evaluation of Long-Term Protection after Immunization in the Presence of BRSV-Specific Maternal Antibodies
  • 2021
  • Ingår i: Vaccines. - : MDPI. - 2076-393X. ; 9:3
  • Tidskriftsartikel (refereegranskat)abstract
    • The induction of long-lasting clinical and virological protection is needed for a successful vaccination program against the bovine respiratory syncytial virus (BRSV). In this study, calves with BRSV-specific maternally derived antibodies were vaccinated once, either with (i) a BRSV pre-fusion protein (PreF) and Montanide(TM) ISA61 VG (ISA61, n = 6), (ii) BRSV lacking the SH gene (Delta SHrBRSV, n = 6), (iii) a commercial vaccine (CV, n = 6), or were injected with ISA61 alone (n = 6). All calves were challenged with BRSV 92 days later and were euthanized 13 days post-infection. Based on clinical, pathological, and proteomic data, all vaccines appeared safe. Compared to the controls, PreF induced the most significant clinical and virological protection post-challenge, followed by Delta SHrBRSV and CV, whereas the protection of PreF-vaccinated calves was correlated with BRSV-specific serum immunoglobulin (Ig)G antibody responses 84 days post-vaccination, and the IgG antibody titers of Delta SHrBRSV- and CV-vaccinated calves did not differ from the controls on this day. Nevertheless, strong anamnestic BRSV- and PreF-specific IgG responses occurred in calves vaccinated with either of the vaccines, following a BRSV challenge. In conclusion, PreF and Delta SHrBRSV are two efficient one-shot candidate vaccines. By inducing a protection for at least three months, they could potentially improve the control of BRSV in calves.
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5.
  • Artemenko, Konstantin A., et al. (författare)
  • Optimization of immunoaffinity enrichment and detection : toward a comprehensive characterization of the phosphotyrosine proteome of K562 cells by liquid chromatography-mass spectrometry
  • 2011
  • Ingår i: The Analyst. - : Royal Society of Chemistry (RSC). - 0003-2654 .- 1364-5528. ; 136:9, s. 1971-1978
  • Tidskriftsartikel (refereegranskat)abstract
    • Phosphorylation of protein tyrosine residues regulates many cell functions and has also been proved to be involved in oncogenesis. Thus, the identification of the phosphotyrosine (pTyr) proteome of cells is a very important task. Since tyrosine phosphorylation represents only around 1% of the total human phosphoproteome, the study of pTyr proteins is rather challenging. Here we report the optimization study of the phosphotyrosine proteome using K562 cells as a model system. A substantial segment of the phosphotyrosine proteome of K562 cells was characterized by immunoaffinity enrichment with 4G10 and PYKD1 antibodies followed by LC-MS/MS analysis. 480 non-redundant pTyr peptides corresponding to 342 pTyr proteins were found. 141 pTyr peptides were not described elsewhere. The mass spectrometry approach involving high-resolving FTMS analysis of precursor ions and subsequent detection of CID fragments in a linear ion trap was considered as optimal. For detection of low abundant pTyr peptides pooling of individual immunoaffinity enrichments for one LC-MS/MS analysis was crucial. The enrichment properties of the monoclonal PYKD1 antibody were presented for the first time, also in comparison to the 4G10 antibody. PYKD1 was found to be more effective for protein enrichment (1.2 and 5% efficiency at peptide and protein level correspondingly), while 4G10 showed better results when peptide enrichment was performed (15% efficiency versus 3.6% at protein level). Substantially different subsets of the phosphoproteome were enriched by these antibodies. This finding together with previous studies demonstrates that comprehensive pTyr proteome characterization by immunoprecipitation requires multiple antibodies to be used for the affinity enrichment.
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6.
  • Bergström Lind, Sara, et al. (författare)
  • A strategy for identification of protein tyrosine phosphorylation
  • 2012
  • Ingår i: Methods. - : Elsevier BV. - 1046-2023 .- 1095-9130. ; 56:2, s. 275-283
  • Tidskriftsartikel (refereegranskat)abstract
    • To develop methods for studying phosphorylation of protein tyrosine residues is an important task since this protein modification regulates many cellular functions and often is involved in oncogenesis. An optimal protocol includes enrichment of tyrosine phosphorylated (pTyr) peptides or proteins, followed by a high resolving analytical method for identification of the enriched components. In this Methods paper, we describe a working strategy on how immunoaffinity enrichments, using anti-pTyr antibodies, combined with mass spectrometric (MS) analysis can be used to study the pTyr proteome. We describe in detail how our procedure was used to characterize the pTyr proteome of K562 leukemia cells. Important questions concerning the use of different anti-pTyr antibodies, enrichments performed at the peptide and/or the protein level, pooling of enrichments and requirements for the MS characterization are discussed.
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7.
  • Bergström Lind, Sara, et al. (författare)
  • Immunoaffinity Enrichments Followed by Mass Spectrometric Detection for Studying Global Protein Tyrosine Phosphorylation
  • 2008
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 7:7, s. 2897-2910
  • Tidskriftsartikel (refereegranskat)abstract
    • Phosphorylation of protein tyrosine residues regulates important cell functions and is, when dysregulated, often crucially involved in oncogenesis. It is therefore important to develop and evaluate methods for identifying and studying tyrosine phosphorylated (P-Tyr) proteins. P-Tyr proteins are present at very low concentrations within cells, requiring highly selective enrichment methods to be detected. In this study, we applied immunoaffinity as enrichment step for P-Tyr proteins. Five selected anti-phosphotyrosine antibodies (monoclonal antibodies 4G10, PY100, PYKD1, 13F9 and one polyclonal antiserum) were evaluated with respect to their capability to enrich P-Tyr proteins from cell extracts of the K562 leukemia cell line. The enrichment resulted in the detection of a group of proteins that potentially were tyrosine-phosphorylated (putative P-Tyr proteins). High accuracy identification of actual P-Tyr sites were performed using a highly selective and sensitive liquid chromatography Fourier transform mass spectrometer (LC-FTMS) setup with complementary collision activated dissociation (CAD) and electron capture dissociation (ECD) fragmentations. 4G10 and PY100 antibodies recognized the greatest number of putative P-Tyr proteins in initial screening experiments and were therefore further evaluated and compared in immunoaffinity enrichment of both P-Tyr proteins and peptides. Using the 4G10 antibody for enrichment of proteins, we identified 459 putative P-Tyr proteins by MS. Out of these proteins, 12 were directly verified as P-Tyr proteins by MS analysis of the actual site. Using the PY100 antibody for enrichment of peptides, we detected 67 P-Tyr peptides (sites) and 89 putative P-Tyr proteins. Generally, enrichment at the peptide level made it difficult to reliably determine the identity of the proteins. In contrast, protein identification following immunoaffinity enrichment at the protein level gave greater sequence coverage and thus a higher confidence in the protein identification. By combining all available information, 40 proteins were identified as true P-Tyr proteins from the K562 cell line. In conclusion, this study showed that a combination of immunoaffinity enrichment using multiple antibodies of both intact and digested proteins in parallel experiments is required for best possible coverage of all possible P-Tyr proteins in a sample.
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8.
  • Bergström Lind, Sara, et al. (författare)
  • The phosphoproteome of the adenovirus type 2 virion
  • 2012
  • Ingår i: Virology. - : Elsevier BV. - 0042-6822 .- 1096-0341. ; 433:1, s. 253-261
  • Tidskriftsartikel (refereegranskat)abstract
    • We have used a proteomics approach to identify sites of phosphorylation in the structural proteins of the Adenovirus type 2 particle. This protein modification might play an important role during infection. Peptides from highly purified virus were enriched for phosphorylations and analyzed by liquid chromatography-high-resolving mass spectrometry. Phosphorylations were identified in 11 structural peptides and 29 non-redundant phosphorylation sites were unambiguously assigned to specific amino acid. An unexpected result was the finding of phosphotyrosine in two of the viral polypeptides. The most highly phosphorylated protein was pIIIa with 12 identified phosphorylation sites. An identified preference for proline or leucine residue flanking the phosphorylation sites downstream suggests that cellular kinases are involved in many of the phosphorylations. Structural modeling showed that one site in the hexon is located on the outer side of the virus and could be of importance for the virus when attaching and entering cells.
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9.
  • Bergström Lind, Sara, et al. (författare)
  • Toward a comprehensive characterization of the phosphotyrosine proteome
  • 2011
  • Ingår i: Cellular Signalling. - : Elsevier BV. - 0898-6568 .- 1873-3913. ; 23:8, s. 1387-1395
  • Tidskriftsartikel (refereegranskat)abstract
    • Tyrosine phosphorylation (pTyr) regulates important cell functions and plays a key role in carcinogenesis. The purpose of this study was to perform a comprehensive study of the phosphotyrosine proteome. Immunoaffinity enriched pTyr proteins and peptides from K562 leukemia cells were analyzed with high-resolving liquid chromatography mass spectrometry. Two different antibodies selective for the pTyr modification were used in repeated enrichments to identify as many pTyr peptides as possible. Stringent verification of putative pTyr sites was performed to assure high reliability in the subsequent biological interpretation of the data. Identified pTyr proteins were subjected to pathway analysis by using different analytical tools. In total, 294 pTyr peptides belonging to 217 pTyr proteins were identified, 15 of which had not previously been reported to be modified by pTyr. The pTyr proteins were clustered in six major groups based on the biological functions "cellular signaling", "cell motility and shape", "cell cycle process", "transport", "RNA processing" and "protein processing". The pTyr proteins were mainly positioned in the following cellular compartments: cytoplasm, cytoskeleton, nucleus and ribonucleoprotein complexes. An interesting finding was that many proteins were related to RNA processing and were found to be heterogeneous nuclear ribonucleoproteins. Also, more than half of the novel pTyr proteins were localized to the nucleus, of which three (PBX2, TEAD1 and DIDO1) were classified as transcription factors and two (CENPC1 and MAD2L1) are associated with cell division control.
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10.
  • Chu, Jiangtao, 1982-, et al. (författare)
  • Influence of surface modification and static pressure on microdialysis protein extraction efficiency
  • 2015
  • Ingår i: Biomedical microdevices (Print). - Springer : Springer Science and Business Media LLC. - 1387-2176 .- 1572-8781. ; 17:5
  • Tidskriftsartikel (refereegranskat)abstract
    • There is growing interest in using microdialysis (MD) for monitoring larger and more complexmolecules such as neuropeptides and proteins. This promotes the use of MD membranes withmolecular weight cut off (MWCO) of 100 kDa or above. The hydrodynamic property of themembrane goes to ultrafiltration or beyond, making the MD catheters more sensitive to pressure.In the meantime, despite the large pore size, studies have shown that membrane biofouling stilllead to unstable catheter performance. The objective is to study in vitro how 500 kDa dextranand Poloxamer 407 surface modification affect the fluid recovery (FR) and extraction efficiency(EE) of 100 kDa MWCO MD catheters. A pressure chamber was designed to facilitate the tests,using as MD sample a protein standard with similar concentrations as in human cerebral spinalfluid, comparing native and Poloxamer 407 modified MD catheters. The collected dialysatefractions were examined for FR and protein EE, employing Dot-it Spot-it Protein Assay for totalprotein EE and targeted mass spectrometry (MS) for EE of individual proteins and peptides. TheFR results suggested that the surface modified catheters were less sensitive to the pressure andprovide higher precision, and provided a FR closer to 100%. The surface modification did notshow a significant effect on the protein EE. The average total protein EE of surface modifiedcatheters was slightly higher than that of the native ones. The MS EE data of individual proteinsshowed a clear trend of complex response in EE with pressure.
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11.
  • Elfineh, Lioudmila, et al. (författare)
  • Tyrosine phosphorylation profiling via in situ proximity ligation assay
  • 2014
  • Ingår i: BMC Cancer. - : Springer Science and Business Media LLC. - 1471-2407. ; 14, s. 435-
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Tyrosine phosphorylation (pTyr) is an important cancer relevant posttranslational modification since it regulates protein activity and cellular localization. By controlling cell growth and differentiation it plays an important role in tumor development. This paper describes a novel approach for detection and visualization of a panel of pTyr proteins in tumors using in situ proximity ligation assay. Methods: K562 leukemia cells were treated with tyrosine kinase and/or phosphatase inhibitors to induce differences in pTyr levels and mimic cells with different malignant properties. Cells were then probed with one antibody against the pTyr modification and another probe against the detected protein, resulting in a detectable fluorescent signal once the probes were in proximity. Results: Total and protein specific pTyr levels on ABL, SHC, ERK2 and PI3K proteins were detected and samples of control and treated cells were distinguished at the pTyr level using this novel approach. Promising results were also detected for formalin fixed and paraffin embedded cells in the micro array format. Conclusions: This application of in situ proximity ligation assay is valuable in order to study the pTyr modification of a panel of proteins in large data sets to validate mass spectrometric data and to be combined with tissue microarrays. The approach offers new opportunities to reveal the pTyr signatures in cells of different malignant properties that can be used as biomarker of disease in the future.
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12.
  • Eriksson, Anna, et al. (författare)
  • On-target titanium dioxide-based enrichment for characterization of phosphorylations in the Adenovirus pIIIa protein
  • 2013
  • Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673 .- 1873-3778. ; 1317:SI, s. 105-109
  • Tidskriftsartikel (refereegranskat)abstract
    • A recently developed titanium dioxide (TiO2) based on-target method for phosphopeptide enrichment and matrix assisted laser desorption-ionization mass spectrometry (MALDI MS) analysis was used to investigate phosphorylations in the Adenovirus type 2 structural protein pIIIa. Lysates of purified virus particles were separated on 1-D SDS-PAGE and the band for the pIIIa protein was excised for tryptic digestion into peptides that were enriched with the on-target method. The enrichment provided by the method clearly improved the detectability of phosphorylated peptides and the results show for the first time evidence for multi-phosphorylated peptides in pIIIa. Moreover, three novel phosphorylations were identified in the protein sequence, even though the precise positions could not be determined. These results illustrate the potential of the method for the characterization of novel phosphoproteomes in biological samples of medical relevance.
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13.
  • Gromova, Arina, et al. (författare)
  • Identification of the adenovirus type 2 C-168 protein
  • 2017
  • Ingår i: Virus Research. - : Elsevier BV. - 0168-1702 .- 1872-7492. ; 238, s. 110-113
  • Tidskriftsartikel (refereegranskat)abstract
    • A hitherto predicted but undetected protein, C-168, in adenovirus type 2 (Ad2) has been identified using mass spectrometry (MS) based proteomics. The gene of this 17.7 kDa protein is located on the forward strand in the major late transcription unit between base pairs 9294 and 9797. A tryptic peptide, derived from the C-terminal part of the protein, was identified with high amino acid sequence coverage. A candidate splice site for the corresponding mRNA is also presented. The protein sequence is unusual with repeats of serine, glycine and arginine. A bioinformatics prediction of protein function and localization is presented.
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14.
  • Hemdan, Tammer, et al. (författare)
  • Stathmin-1 is a promising prognostic factor and potential therapeutic target in urinary bladder cancer
  • Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
    • Aim: The oncoprotein 18/stathmin 1 (STMN1), involved in cell cycle progression and cell migration, has been reported to be expressed in several types of cancer, and is associated with clinical outcome in e.g. breast and liver cancer. The aims in this study were to investigate the clinical significance of STMN1 and to examine if STMN1 might be a possible therapeutic target in urinary bladder cancer.Experimental design: Immunohistochemical analyses of STMN1 protein expression were performed in a wide-range tissue microarray (115 Ta-, 115 T1-, 112 T2-4-tumors) and in a metastatic primary tumor/matched metastasis-material (90 patients). In the T24 cell line, the effect of STMN1 on cell proliferation was evaluated by inhibiting the cellular expression of STMN using STMN1-siRNA.Results: Patients with T1- or muscle-invasive disease exhibiting high expression of the STMN1 protein had a poorer overall survival (OS) and disease specific survival (DSS). In a multivariate analysis adjusting for stage, age and gender the results were for T2-T4 patients: OS (HR=1.77 95% CI 1.02-3.07; p=0.04) and DSS (HR=2.04 95% CI 1.13-3.68; p=0.02); for T1-4 patients: DSS (HR=1.83 95% CI 1.09-3.08; p=0.02). In the metastatic bladder cancer material, the majority of the patients with one metastasis (69%) and with several matched metastases (70%) were STMN1-positive in both the primary tumor and the matched metastases. Moreover, the ability of the urinary bladder cancer cell line to grow was significantly reduced after 72 hours (p<0.0001) when transfecting the cells with a siRNA targeting STMN1.Conclusion: Our results suggest that STMN1 protein-expression has a potential both as a prognostic marker and a novel treatment target in urinary bladder cancer.
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15.
  • Heras, Gabriel, et al. (författare)
  • Muscle RING-finger protein-1 (MuRF1) functions and cellular localization are regulated by SUMO1 post-translational modification
  • 2019
  • Ingår i: Journal of Molecular Cell Biology. - : Oxford University Press (OUP). - 1674-2788 .- 1759-4685. ; 11:5, s. 356-370
  • Tidskriftsartikel (refereegranskat)abstract
    • The muscle RING-finger protein-1 (MuRF1) is an E3 ubiquitin ligase expressed in skeletal and cardiac muscle tissues and it plays important roles in muscle remodeling. Upregulation of MuRF1 gene transcription participates in skeletal muscle atrophy, on contrary downregulation of protein expression leads to cardiac hypertrophy. MuRF1 gene point mutations have been found to generate protein aggregate myopathies defined as muscle disorder characterized by protein accumulation in muscle fibers. We have discovered that MuRF1 turned out to be also a target for a new post-translational modification arbitrated by conjugation of SUMO1 and it is mediated by the SUMO ligases E2 UBC9 and the E3 PIASγ/4. SUMOylation takes place at lysine 238 localized at the second coiled-coil protein domain that is required for efficient substrate interaction for polyubiquitination. We provided evidence that SUMOylation is essential for MuRF1 nuclear translocation and its mitochondria accumulation is enhanced in hyperglycemic conditions delivering a stabilization of the overall SUMOylated proteins in cultured myocytes. Thus, our findings add this SUMO1 post-translational modification as a new concept to understand muscle disorders related to the defect in MuRF1 activity.
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16.
  • Hernroth, Bodil, 1951, et al. (författare)
  • Impact of ocean acidification on antimicrobial activity in gills of the blue mussel (Mytilus edulis)
  • 2016
  • Ingår i: Fish and Shellfish Immunology. - : Elsevier BV. - 1050-4648 .- 1095-9947. ; 55, s. 452-459
  • Tidskriftsartikel (refereegranskat)abstract
    • Here, we aimed to investigate potential effects of ocean acidification on antimicrobial peptide (AMP) activity in the gills of Mytilus edulis, as gills are directly facing seawater and the changing pH (predicted to be reduced from ∼8.1 to ∼7.7 by 2100). The AMP activity of gill and haemocyte extracts was compared at pH 6.0, 7.7 and 8.1, with a radial diffusion assay against Escherichia coli. The activity of the gill extracts was not affected by pH, while it was significantly reduced with increasing pH in the haemocyte extracts. Gill extracts were also tested against different species of Vibrio (V. parahaemolyticus, V. tubiashii, V. splendidus, V. alginolyticus) at pH 7.7 and 8.1. The metabolic activity of the bacteria decreased by ∼65–90%, depending on species of bacteria, but was, as in the radial diffusion assay, not affected by pH. The results indicated that AMPs from gills are efficient in a broad pH-range. However, when mussels were pre-exposed for pH 7.7 for four month the gill extracts presented significantly lower inhibit of bacterial growth. A full in-depth proteome investigation of gill extracts, using LC-Orbitrap MS/MS technique, showed that among previously described AMPs from haemocytes of Mytilus, myticin A was found up-regulated in response to lipopolysaccharide, 3 h post injection. Sporadic occurrence of other immune related peptides/proteins also pointed to a rapid response (0.5–3 h p.i.). Altogether, our results indicate that the gills of blue mussels constitute an important first line defence adapted to act at the pH of seawater. The antimicrobial activity of the gills is however modulated when mussels are under the pressure of ocean acidification, which may give future advantages for invading pathogens.
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17.
  • Holfeld, Aleš, et al. (författare)
  • Parallel Proteomic Workflow for Mass Spectrometric Analysis of Tissue Samples Preserved by Different Methods
  • 2018
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 90:9, s. 5841-5849
  • Tidskriftsartikel (refereegranskat)abstract
    • Formalin-fixed and paraffin-embedded (FFPE) and optimal cutting temperature (OCT)-embedded and frozen tissue specimens in biobanks are highly valuable in clinical studies but proteomic and post-translational modification (PTM) studies using mass spectrometry (MS) have been limited due to structural arrangement of proteins and contaminations from embedding material. This study aims to develop a parallel proteomic workflow for FFPE and OCT/frozen samples that allows for large-scale, quick, reproducible, qualitative, and quantitative high-resolution MS analysis. The optimized protocol gives details on removal of embedding material, protein extraction, and multienzyme digestion using filter-aided sample preparation method. The method was evaluated by investigating the protein expression levels in nonmuscle-invasive and muscle-invasive bladder cancer samples in two cohorts and MS spectra were carefully reviewed for contaminations. More than 2000 and 3000 proteins in FFPE and OCT/frozen samples, respectively, were identified, and samples could be clustered in different tumor stages based on their protein expression. Furthermore, more than 250 and 400 phosphopeptides could be identified from specific patient samples of FFPE and OCT/frozen, respectively, using titanium dioxide enrichment. The paper presents unique data describing the similarities and differences observed in FFPE and OCT/frozen samples and shows the feasibility to detect proteins and site-specific phosphorylation even after long-term storage of clinical samples.
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18.
  • Hörnaeus, Katarina, et al. (författare)
  • Mass spectrometry data from a quantitative analysis of protein expression in gills of immuno-challenged blue mussels (Mytilus edulis)
  • 2016
  • Ingår i: Data in Brief. - Elsevier : Elsevier BV. - 2352-3409. ; 8, s. 470-473
  • Tidskriftsartikel (refereegranskat)abstract
    • Here, we provide the dataset associated with our research article on the potential effects of ocean acidification on antimicrobial peptide (AMP) activity in the gills of Mytilus edulis, "Impact of ocean acidification on antimicrobial activity in gills of the blue mussel (Mytilus edulis)" [1]. Blue mussels were stimulated with lipopolysaccharides and samples were collected at different time points post injection. Protein extracts were prepared from the gills, digested using trypsin and a full in-depth proteome investigation was performed using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Protein identification and quantification was performed using the MaxQuant 1.5.1.2 software, "MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification" [2].
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19.
  • Karady, Julia, et al. (författare)
  • Coronary Plaque in People With HIV vs Non-HIV Asymptomatic Community and Symptomatic Higher-Risk Populations
  • 2024
  • Ingår i: JACC: Advances. - 2772-963X. ; 3:6
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: People with HIV (PWH) have a high burden of coronary plaques; however, the comparison to people without known HIV (PwoH) needs clarification. Objectives: The purpose of this study was to determine coronary plaque burden/phenotype in PWH vs PwoH. Methods: Nonstatin using participants from 3 contemporary populations without known coronary plaques with coronary CT were compared: the REPRIEVE (Randomized Trial to Prevent Vascular Events in HIV) studying PWH without cardiovascular symptoms at low-to-moderate risk (n = 755); the SCAPIS (Swedish Cardiopulmonary Bioimage Study) of asymptomatic community PwoH at low-to-intermediate cardiovascular risk (n = 23,558); and the PROMISE (Prospective Multicenter Imaging Study for Evaluation of Chest Pain) of stable chest pain PwoH (n = 2,291). The coronary plaque prevalence on coronary CT was compared, and comparisons were stratified by 10-year atherosclerotic cardiovascular disease (ASCVD) risk, age, and coronary artery calcium (CAC) presence. Results: Compared to SCAPIS and PROMISE PwoH, REPRIEVE PWH were younger (50.8 ± 5.8 vs 57.3 ± 4.3 and 60.0 ± 8.0 years; P < 0.001) and had lower ASCVD risk (5.0% ± 3.2% vs 6.0% ± 5.3% and 13.5% ± 11.0%; P < 0.001). More PWH had plaque compared to the asymptomatic cohort (48.5% vs 40.3%; P < 0.001). When stratified by ASCVD risk, PWH had more plaque compared to SCAPIS and a similar prevalence of plaque compared to PROMISE. CAC = 0 was more prevalent in PWH (REPRIEVE 65.2%; SCAPIS 61.6%; PROMISE 49.6%); among CAC = 0, plaque was more prevalent in PWH compared to the PwoH cohorts (REPRIEVE 20.8%; SCAPIS 5.4%; PROMISE 12.3%, P < 0.001). Conclusions: Asymptomatic PWH in REPRIEVE had more plaque than asymptomatic PwoH in SCAPIS but had similar prevalence to a higher-risk stable chest pain cohort in PROMISE. In PWH, CAC = 0 does not reliably exclude plaque.
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20.
  • Karady, Julia, et al. (författare)
  • Coronary plaque in people with HIV vs non-HIV asymptomatic community and symptomatic higher-risk populations
  • 2024
  • Ingår i: JACC: Advances. - : Elsevier. - 2772-963X. ; 3:6
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: People with HIV (PWH) have a high burden of coronary plaques; however, the comparison to people without known HIV (PwoH) needs clarification.Objectives: The purpose of this study was to determine coronary plaque burden/phenotype in PWH vs PwoH.Methods: Nonstatin using participants from 3 contemporary populations without known coronary plaques with coronary CT were compared: the REPRIEVE (Randomized Trial to Prevent Vascular Events in HIV) studying PWH without cardiovascular symptoms at low-to-moderate risk (n = 755); the SCAPIS (Swedish Cardiopulmonary Bioimage Study) of asymptomatic community PwoH at low-to-intermediate cardiovascular risk (n = 23,558); and the PROMISE (Prospective Multicenter Imaging Study for Evaluation of Chest Pain) of stable chest pain PwoH (n = 2,291). The coronary plaque prevalence on coronary CT was compared, and comparisons were stratified by 10-year atherosclerotic cardiovascular disease (ASCVD) risk, age, and coronary artery calcium (CAC) presence.Results: Compared to SCAPIS and PROMISE PwoH, REPRIEVE PWH were younger (50.8 ± 5.8 vs 57.3 ± 4.3 and 60.0 ± 8.0 years; P < 0.001) and had lower ASCVD risk (5.0% ± 3.2% vs 6.0% ± 5.3% and 13.5% ± 11.0%; P < 0.001). More PWH had plaque compared to the asymptomatic cohort (48.5% vs 40.3%; P < 0.001). When stratified by ASCVD risk, PWH had more plaque compared to SCAPIS and a similar prevalence of plaque compared to PROMISE. CAC = 0 was more prevalent in PWH (REPRIEVE 65.2%; SCAPIS 61.6%; PROMISE 49.6%); among CAC = 0, plaque was more prevalent in PWH compared to the PwoH cohorts (REPRIEVE 20.8%; SCAPIS 5.4%; PROMISE 12.3%, P < 0.001).Conclusions: Asymptomatic PWH in REPRIEVE had more plaque than asymptomatic PwoH in SCAPIS but had similar prevalence to a higher-risk stable chest pain cohort in PROMISE. In PWH, CAC = 0 does not reliably exclude plaque.
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21.
  • Khandagale, Avinash, et al. (författare)
  • Plasma-derived extracellular vesicles from myocardial infarction patients inhibits tumor necrosis factor-alpha induced cardiac cell death
  • 2021
  • Ingår i: Current research in translational medicine. - : Elsevier. - 2452-3186. ; 70:2
  • Tidskriftsartikel (refereegranskat)abstract
    • RATIONALE: Extracellular vesicles (EVs) derived exogenously from pluripotent stem cells or endogenously from healthy human serum exert cardioprotective effects after injury. However role of endogenous EVs from myocardial infarction (MI) patients not well understood in this settings.METHODS AND RESULTS: The EVs from plasma of MI patients with preserved or reduced left ventricular ejection fraction (LVEF) and healthy controls (HC) were purified and characterized by flow cytometry, mass spectrometry (MS) and transmission electron microscopy (TEM). HCM and human cardiac microvascular endothelial cells (hCMVECs), under individual culture or co-culture, were used to study functional effects of EVs upon TNFα stimulation. These effects of EVs on HCM and hCMVECs were observed using cell death assays, western blots and confocal microscopy. Higher concentrations of platelet-, leukocyte-, endothelial- and erythrocyte-derived EVs were found in MI patients, both with preserved and reduced LVEF, compared to HC, and MS data on MI EVs proteome displayed alteration in several proteins. MI EVs protected HCM and hCMVECs against staurosporine-induced apoptosis. Furthermore, MI EVs were observed to abrogate TNFα-triggered HCM and hCMVECs death under both individually cultured and co-cultured conditions. MI EVs failed to inhibit TNFα induced hCMVECs and HCM activation when cultured individually, however co-cultured hCMVECs with HCM supported MI EVs capacity to attenuate TNFα induced cells activation. MI CD41+ EVs but not HC EVs were found to be internalized by HCM directly or migrated through hCMVECs to HCM. MI EVs indirectly restores TNFα mediated drop in mitochondrial membrane potential.CONCLUSIONS: Endogenous EVs from MI patients, regardless of severity of the MI exert cardioprotective potential upon TNFα-induced cell death. Patient-derived EVs needs to be further explored to elucidate their potential cardioprotective role during MI.
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22.
  • Kjellander, Marcus, et al. (författare)
  • A flow-through nanoporous alumina trypsin bioreactor for mass spectrometry peptide fingerprinting
  • 2018
  • Ingår i: Journal of Proteomics. - : Elsevier. - 1874-3919 .- 1876-7737. ; 172, s. 165-172
  • Tidskriftsartikel (refereegranskat)abstract
    • Mass spectrometry-based proteomics benefits from efficient digestion of protein samples. In this study, trypsin was immobilized on nanoporous anodized alumina membranes to create an enzyme reactor suitable for peptide mass fingerprinting. The membranes were derivatized with 3-aminopropyltriethoxysilane and the amino groups were activated with carbonyldiimidazole to allow coupling of porcine trypsin via c-amino groups. The function was assessed using the artificial substrate Na-Benzoyl-L-arginine 4-nitroanilide hydrochloride, bovine ribonuclease A and a human plasma sample. A 10-membrane flow-through reactor was used for fragmentation and MS analysis after a single pass of substrate both by collection of product and subsequent off-line analysis, and by coupling on-line to the instrument. The peptide pattem allowed correct identification of the single target protein in both cases, and of > 70 plasma proteins in single pass mode followed by LC-MS analysis. The reactor retained 76% of the initial activity after 14 days of storage and repeated use at room temperature. Significance: This manuscript describes the design of a stable enzyme reactor that allows efficient and fast digestion with negligible leakage of enzyme and enzyme fragments. The high stability facilitates the use in an online-setup with MS detection since it allows the processing of multiple samples within an extended period of time without replacement.
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23.
  • Källsten, Malin, et al. (författare)
  • A comparative study of phosphopeptide-selective techniques for a sub-proteome of a complex biological sample.
  • 2016
  • Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 408:9, s. 2347-2356
  • Tidskriftsartikel (refereegranskat)abstract
    • Phosphorylation of proteins is important for controlling cellular signaling and cell cycle regulatory events. The process is reversible and phosphoproteins normally constitute a minor part of the global proteome in a cell. Thus, sample preparation techniques tailored for phosphoproteome studies are continuously invented and evaluated. This paper aims at evaluating the performances of the most popular techniques for phospho-enrichments in sub-proteome analysis, such as viral proteomes expressed in human cells during infection. A two-species sample of Adenovirus type 2 infected human cells was used, and in-solution digestion, strong cation exchange (SCX), and electrostatic repulsion hydrophilic interaction chromatography (ERLIC) fractionation, and subsequent enrichment by TiO2, were compared with SDS-PAGE fractionation and in-gel digestion. Evaluation was focused on phosphopeptide detection in the sub-proteome. The results showed that the SCX+TiO2 or ERLIC+TiO2 combinations had the highest enrichment efficiencies, but SDS-PAGE fractionation and in-gel digestion resulted in the highest number of identified proteins and phosphopeptides. Furthermore, the study demonstrates the usefulness of applying as many orthogonal techniques as possible in deep phosphoproteome analysis, since the overlap between approaches was low.
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24.
  • Källsten, Malin, et al. (författare)
  • Qualitative analysis of antibody-drug conjugates (ADCs) : an experimental comparison of analytical techniques of cysteine-linked ADCs.
  • 2018
  • Ingår i: The Analyst. - : Royal Society of Chemistry (RSC). - 0003-2654 .- 1364-5528. ; 143:22, s. 5487-5496
  • Tidskriftsartikel (refereegranskat)abstract
    • Antibody-drug conjugates (ADCs) are an emerging type of biotherapeutics that utilize multiple tissue-specific antibodies combined with a range of linker designs to enable the transportation and selective release of cytotoxic drugs in close proximity to tumours. Consisting of antibodies conjugated to small drug molecules through a variety of linkers, ADCs are chemically complex analytes. Here we present a unique experimental comparison of four techniques for ADC analysis: hydrophobic interaction chromatography (HIC-UV/Vis), reversed phase liquid chromatography mass spectrometry (RPLC-MS), using either a QToF or an Orbitrap analyser, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Four different ADCs consisting of Trastuzumab, monomethyl auristatin E (MMAE) and a peptidic linker moiety differing in their respective stoichiometric ratios in regard to drug-to-antibody ratio (DAR) were used for the comparison. We found that the determined DAR from all techniques was comparable, while the accuracy of the molecular weights for the conjugated light and heavy chain differed more extensively. This indicates that the choice of a mass analyser is more crucial for determining the accurate weights of the light and heavy chains than to evaluate the DAR of a given batch. However, ambiguous DAR assignment in HIC-UV/Vis or bias for either the light or heavy chain fragments in the mass spectrometry-based techniques can influence the obtained average DAR value and the use of complementary techniques is advisable. Out of the four techniques evaluated, HIC-UV/Vis and MALDI required less time to obtain an average DAR value and would therefore be good for initial screenings in the early stages of the discovery phase of new ADCs.
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25.
  • Lind, Sara Bergström, et al. (författare)
  • Detection of tyrosine phosphorylated proteins by combination of immunoaffinity enrichment, two-dimensional difference gel electrophoresis and fluorescent Western blotting
  • 2010
  • Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 401:4, s. 581-585
  • Tidskriftsartikel (refereegranskat)abstract
    • We describe fluorescence-based 2-D gel electrophoresis methods for visualization of low abundant cancer relevant tyrosine phosphorylated (pTyr) proteins The methods investigated were fluorescent Western blotting and two-dimensional difference gel electrophoresis (2-D DIGE) for detection of non-enriched and immunoaffinity enriched pTyr protein patterns The same anti-phosphotyrosine specific antibody 4G10 was used for both approaches The results from fluorescent Western blotting of total proteins and from enriched CyDye DIGE pre-labeled pTyr proteins showed similar down regulation of phosphorylation upon treating of cells from a cancer model system (K562 chronic myeloid leukemia cells) with imatinib This treatment introduced a known perturbation of phosphorylation that enabled testing of these new approaches to analyze variations in tyrosine phosphorylation levels Enrichment of pTyr proteins was found highly advantageous for the outcome Out of a simplified 2 D DIGE experiment of immunoaffinity enriched control and treated pTyr proteins differential analysis as well as protein identification by mass spectrometry (MS) was possible.
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26.
  • Lind, Sara Bergström, et al. (författare)
  • Post translational modifications in adenovirus type 2
  • 2013
  • Ingår i: Virology. - : Elsevier BV. - 0042-6822 .- 1096-0341. ; 447:1-2, s. 104-111
  • Tidskriftsartikel (refereegranskat)abstract
    • We have combined 2-D SOS-PAGE with liquid chromatography-high resolving mass spectrometry (LC-MS) to explore the proteome of the adenovirus type 2 (Ad2) at the level of post translational modifications (PTMs). The experimental design included in-solution digestion, followed by titanium dioxide enrichment, as well as in-gel digestion of polypeptides after separation of Ad2 capsid proteins by 1-D and 2-D SOS-PAGE. All samples were analyzed using LC-MS with subsequent manual verification of PTM positions. The results revealed new phosphorylation sites that can explain the observed trains of protein spots observed for the pIII, pIIIa and ply proteins. The pin protein was found to be the most highly modified protein with now 18 verified sites of phosphorylation, three sites of nitrated tyrosine and one sulfated tyrosine. Nitrated tyrosines were also identified in pII. Lysine acetylations were detected in pII and pVI. The findings make the Ad2 virion much more complex than hitherto believed. 
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27.
  • Lind, Sara Bergström, et al. (författare)
  • Proteome Analysis of Adenovirus Using Mass Spectrometry
  • 2014
  • Ingår i: Adenovirus: Methods and Protocols. - : Springer Science+Business Media B.V.. ; , s. 25-44
  • Bokkapitel (refereegranskat)abstract
    • Analysis of proteins and their posttranslational modifications is important for understanding different biological events. For analysis of viral proteomes, an optimal protocol includes production of a highly purified virus that can be investigated with a high-resolving analytical method. In this Methods in Molecular Biology paper we describe a working strategy for how structural proteins in the Adenovirus particle can be studied using liquid chromatography–high-resolving mass spectrometry. This method provides information on the chemical composition of the virus particle. Further, knowledge about amino acids carrying modifications that could be essential for any part of the virus life cycle is collected. We describe in detail alternatives available for preparation of virus for proteome analysis as well as choice of mass spectrometric instrumentation suitable for this kind of analysis.
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28.
  • Lindén, Mårten, et al. (författare)
  • Galectin-1, a potential therapeutic target, in primary tumors and metastases of urinary bladder carcinomas
  • 2013
  • Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
    • Urinary bladder cancer would gain from new protein biomarkers due to the heterogeneity of disease. The beta-galactoside-binding protein (GAL1) is one such candidate and in present study its prognostic value and expression at protein level in metastatic bladder cancer-disease, have been evaluated. The protein expression of GAL1 was investigated by immunohistochemistry in two tumor cohorts, one with primary tumors of different stage and grade (n=344) and another with primary tumors matched with metastases (n=90). The expression in the actual cancer cells as well as in stroma and blood vessels were considered since the presence of GAL1 in different tissue compartments has shown cancer relevance. The cellular expression increased with increased tumor stage and grade (p<0.001). For the majority of the patients, cells from both primary tumor and metastasis showed a positive immunoreactivity for GAL1 (91% (n=64) for primary tumors with single metastasis (n=70) and 100% (n=20) for primary tumors with multiple metastasis (n=20). Further, strong immunoreactivity in T1 tumor cells correlated with lower risk of recurrence (p<0.05). Both tumors and metastasis exhibited strong stromal-GAL1 staining that could not be correlated with clinical parameters. The expression in vessels showed that T1 tumors surrounded by GAL1 negative blood vessels had a higher risk of progression (p<0.0001) into muscle invasive T2-4 stages. The results show that GAL1 is an important bladder cancer-protein from several aspects. Further, GAL1 is a promising therapeutic target in bladder cancer due to the general expression in advanced disease.
  •  
29.
  • Lindén, Mårten, et al. (författare)
  • Proteomic analysis of urinary biomarker candidates for nonmuscle invasive bladder cancer
  • 2012
  • Ingår i: Proteomics. - : Wiley. - 1615-9853 .- 1615-9861. ; 12:1, s. 135-144
  • Tidskriftsartikel (refereegranskat)abstract
    • Nonmuscle invasive tumors of the bladder often recur and thereby bladder cancer patients need regular re-examinations which are invasive, unpleasant, and expensive. A noninvasive and less expensive method, e.g. a urine dipstick test, for monitoring recurrence would thus be advantageous. In this study, the complementary techniques mass spectrometry (MS) and Western blotting (WB)/dot blot (DB) were used to screen the urine samples from bladder cancer patients. High resolving MS was used to analyze and quantify the urinary proteome and 29 proteins had a significantly higher abundance (p<0.05) in bladder cancer samples compared with control urine samples. The increased abundance found in urine from bladder cancer patients compared with controls was confirmed with Western blot for four selected proteins; fibrinogen β chain precursor, apolipoprotein E, α-1-antitrypsin, and leucine-rich α-2-glycoprotein 1. Dot blot analysis of an independent urine sample set pointed out fibrinogen β chain and α-1-antitrypsin as most interesting biomarkers having sensitivity and specificity values in the range of 66-85%. Exploring the Human Protein Atlas (HPA) also revealed that bladder cancer tumors are the likely source of these proteins. They have the potential of being useful in diagnosis, monitoring of recurrence and thus may improve the treatment of bladder tumors, especially nonmuscle invasive tumors.
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30.
  • Lindén, Mårten, 1976- (författare)
  • Proteomic Analysis of Urinary Bladder Cancer : Aiming for Novel Biomarkers
  • 2013
  • Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
    • Urinary bladder cancer is a heterogeneous disease appearing in different forms, e.g. non-muscle invasive and muscle invasive. For all variants, the expression of proteins is interesting to analyze for diagnostic, predictive, prognostic and drug targeting purposes, since it reflects the altered gene expression causing the cancer. Since urothelial cells of the bladder are in direct contact with urine it is likely that this body fluid contains cancer-related proteins. In Paper I, unbiased analysis of proteins in urine from urinary bladder cancer patients and controls, using label-free quantification by mass spectrometry, was applied and four interesting proteins APOE, FGB, LRG and SERPINA1 were selected and further analyzed with western and dot blot. In Paper II, two more proteins, POLR1E and TOP2A, were validated as relevant proteins in bladder cancer urine. In Paper III and IV, the proteins GAL1 and STMN1 were investigated for their prognostic and therapeutic target potential in bladder cancer. In Paper II, III and IV, the expression of seven of the proteins were analyzed on tissue microarrays representing tumour tissue from 360 patients with different tumour stages. For the proteins identified by the urine screening approach, their protein expressions were confirmed in bladder cancer tissue. The expression level in tissue of five of the proteins, APOE, FGB, POLR1E (Paper II), GAL1 (Paper III) and STMN1 (Paper IV), increased with tumour stage, showing diagnostic relevance and three of the proteins, SERPINA1 (Paper II), STMN1 (Paper IV) and GAL1 (Paper III) had prognostic potential in urinary bladder cancer. In addition, GAL1 and STMN1 were demonstrated to be highly expressed in metastatic disease and inhibition of STMN1 reduced cell growth (Paper III and IV), indicating that these proteins are promising drug targets in urinary bladder cancer. In conclusion, the approach of this thesis has generated several candidate protein biomarkers in urine and tissue, validated with independent methods, which have the potential to improve the care for bladder cancer patients.
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31.
  • Lindén, Mårten, et al. (författare)
  • Tumour expression of bladder cancer-associated urinary proteins
  • 2013
  • Ingår i: BJU International. - 1464-4096 .- 1464-410X. ; 112:3, s. 407-415
  • Tidskriftsartikel (refereegranskat)abstract
    • WHAT'S KNOWN ON THE SUBJECT? AND WHAT DOES THE STUDY ADD?:The current basis for diagnosis and prognosis in urinary bladder cancer is based on the pathologists' assessment of a biopsy of the tumour. Urinary biomarkers are preferable as they can be non-invasively sampled. Urinary cytology is the only test with widespread use but is hampered by poor reproducibility and low sensitivity.By studying the protein expression in bladder tumour tissue samples of proteins previously found in elevated levels in the urine of patients with bladder cancer, we have been able to show that these proteins originate from the tumour. The immunoreactivity of three of the investigated proteins increased with higher stage. Also a serine peptidase inhibitor was found to be predictive of progression from non-muscle-invasive to muscle-invasive tumours.OBJECTIVES:To analyse the expression of five bladder cancer-associated urinary proteins and investigate if expression is related to the malignant phenotype of the tumour.To explore the possible prognostic value of these proteins.PATIENTS AND METHODS:Urine samples, 16 from patients with bladder cancer and 26 from controls, were used in Western Blotting experiments.Tissue microarrays with bladder tissue from 344 patients diagnosed with bladder cancer between 1984 and 2005 was used in immunohistochemistry experiments.The proteins apolipoprotein E (APOE), fibrinogen β chain precursor (FGB), leucine-rich α2-glycoprotein (LRG1), polymerase (RNA) I polypeptide E (POLR1E), α1-antitrypsin (SERPINA1) and topoisomerase 2A (TOP2A) were probed with antibodies validated by the Human Protein Atlas.RESULTS:Increased expressions of APOE, FGB and POLR1E were correlated with increased tumour stage (P < 0.001).Expression of SERPINA1 in Ta and T1 tumours was found to increase the risk of tumour progression (hazard ratio 2.57, 95% confidence interval 1.13-5.87; P = 0.025)CONCLUSIONS: All proteins previously detected in urine from patients with bladder cancer were also expressed in bladder cancer tissue.The expression of APOE, FGB and POLR1E increased with stage and they are potential diagnostic markers.SERPINA1 was identified as a prognostic marker candidate.
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32.
  • Ma'ayeh, Showgy Y., et al. (författare)
  • Characterization of the Giardia intestinalis secretome during interaction with human intestinal epithelial cells : The impact on host cells
  • 2017
  • Ingår i: PLoS Neglected Tropical Diseases. - : Public Library of Science (PLoS). - 1935-2727 .- 1935-2735. ; 11:12
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND:Giardia intestinalis is a non-invasive protozoan parasite that causes giardiasis in humans, the most common form of parasite-induced diarrhea. Disease mechanisms are not completely defined and very few virulence factors are known.METHODOLOGY:To identify putative virulence factors and elucidate mechanistic pathways leading to disease, we have used proteomics to identify the major excretory-secretory products (ESPs) when Giardia trophozoites of WB and GS isolates (assemblages A and B, respectively) interact with intestinal epithelial cells (IECs) in vitro.FINDINGS:The main parts of the IEC and parasite secretomes are constitutively released proteins, the majority of which are associated with metabolism but several proteins are released in response to their interaction (87 and 41 WB and GS proteins, respectively, 76 and 45 human proteins in response to the respective isolates). In parasitized IECs, the secretome profile indicated effects on the cell actin cytoskeleton and the induction of immune responses whereas that of Giardia showed anti-oxidation, proteolysis (protease-associated) and induction of encystation responses. The Giardia secretome also contained immunodominant and glycosylated proteins as well as new candidate virulence factors and assemblage-specific differences were identified. A minor part of Giardia ESPs had signal peptides (29% for both isolates) and extracellular vesicles were detected in the ESPs fractions, suggesting alternative secretory pathways. Microscopic analyses showed ESPs binding to IECs and partial internalization. Parasite ESPs reduced ERK1/2 and P38 phosphorylation and NF-κB nuclear translocation. Giardia ESPs altered gene expression in IECs, with a transcriptional profile indicating recruitment of immune cells via chemokines, disturbances in glucose homeostasis, cholesterol and lipid metabolism, cell cycle and induction of apoptosis.CONCLUSIONS:This is the first study identifying Giardia ESPs and evaluating their effects on IECs. It highlights the importance of host and parasite ESPs during interactions and reveals the intricate cellular responses that can explain disease mechanisms and attenuated inflammatory responses during giardiasis.
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33.
  • Namuduri, Arvind Venkat, et al. (författare)
  • A Proteomic Approach to Identify Alterations in the Small Ubiquitin-like Modifier (SUMO) Network during Controlled Mechanical Ventilation in Rat Diaphragm Muscle
  • 2017
  • Ingår i: Molecular & Cellular Proteomics. - : AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC. - 1535-9476 .- 1535-9484. ; 16:6, s. 1081-1097
  • Tidskriftsartikel (refereegranskat)abstract
    • The small ubiquitin-like modifier (SUMO) is as a regulator of many cellular functions by reversible conjugation to a broad number of substrates. Under endogenous or exogenous perturbations, the SUMO network becomes a fine sensor of stress conditions by alterations in the expression level of SUMO enzymes and consequently changing the status of SUMOylated proteins. The diaphragm is the major inspiratory muscle, which is continuously active under physiological conditions, but its structure and function is severely affected when passively displaced for long extents during mechanical ventilation (MV). An iatrogenic condition called Ventilator-Induced Diaphragm Dysfunction (VIDD) is a major cause of failure to wean patients from ventilator support but the molecular mechanisms underlying this dysfunction are not fully understood. Using a unique experimental Intensive Care Unit (ICU) rat model allowing long-term MV, diaphragm muscles were collected in rats control and exposed to controlled MV (CMV) for durations varying between 1 and 10 days. Endogenous SUMOylated diaphragm proteins were identified by mass spectrometry and validated with in vitro SUMOylation systems. Contractile, calcium regulator and mitochondrial proteins were of specific interest due to their putative involvement in VIDD. Differences were observed in the abundance of SUMOylated proteins between glycolytic and oxidative muscle fibers in control animals and high levels of SUMOylated proteins were present in all fibers during CMV. Finally, previously reported VIDD biomarkers and therapeutic targets were also identified in our datasets which may play an important role in response to muscle weakness seen in ICU patients. Data are available via ProteomeXchange with identifier PXD006085.
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34.
  • Sayols-Baixeras, Sergi, et al. (författare)
  • Streptococcus Species Abundance in the Gut Is Linked to Subclinical Coronary Atherosclerosis in 8973 Participants From the SCAPIS Cohort
  • 2023
  • Ingår i: Circulation. - : Lippincott Williams & Wilkins. - 0009-7322 .- 1524-4539. ; 148:6, s. 459-472
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Gut microbiota have been implicated in atherosclerotic disease, but their relation with subclinical coronary atherosclerosis is unclear. This study aimed to identify associations between the gut microbiome and computed tomography-based measures of coronary atherosclerosis and to explore relevant clinical correlates.Methods: We conducted a cross-sectional study of 8973 participants (50 to 65 years of age) without overt atherosclerotic disease from the population-based SCAPIS (Swedish Cardiopulmonary Bioimage Study). Coronary atherosclerosis was measured using coronary artery calcium score and coronary computed tomography angiography. Gut microbiota species abundance and functional potential were assessed with shotgun metagenomics sequencing of stool, and associations with coronary atherosclerosis were evaluated with multivariable regression models adjusted for cardiovascular risk factors. Associated species were evaluated for association with inflammatory markers, metabolites, and corresponding species in saliva.Results: The mean age of the study sample was 57.4 years, and 53.7% were female. Coronary artery calcification was detected in 40.3%, and 5.4% had at least 1 stenosis with >50% occlusion. Sixty-four species were associated with coronary artery calcium score independent of cardiovascular risk factors, with the strongest associations observed for Streptococcus anginosus and Streptococcus oralis subsp oralis (P<1×10-5). Associations were largely similar across coronary computed tomography angiography-based measurements. Out of the 64 species, 19 species, including streptococci and other species commonly found in the oral cavity, were associated with high-sensitivity C-reactive protein plasma concentrations, and 16 with neutrophil counts. Gut microbial species that are commonly found in the oral cavity were negatively associated with plasma indole propionate and positively associated with plasma secondary bile acids and imidazole propionate. Five species, including 3 streptococci, correlated with the same species in saliva and were associated with worse dental health in the Malmö Offspring Dental Study. Microbial functional potential of dissimilatory nitrate reduction, anaerobic fatty acid β-oxidation, and amino acid degradation were associated with coronary artery calcium score.Conclusions: This study provides evidence of an association of a gut microbiota composition characterized by increased abundance of Streptococcus spp and other species commonly found in the oral cavity with coronary atherosclerosis and systemic inflammation markers. Further longitudinal and experimental studies are warranted to explore the potential implications of a bacterial component in atherogenesis.
  •  
35.
  • Sayols-Baixeras, Sergi, et al. (författare)
  • Streptococcus Species Abundance in the Gut Is Linked to Subclinical Coronary Atherosclerosis in 8973 Participants From the SCAPIS Cohort
  • 2023
  • Ingår i: Circulation. - : American Heart Association. - 0009-7322 .- 1524-4539. ; 148:6, s. 459-472
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND: Gut microbiota have been implicated in atherosclerotic disease, but their relation with subclinical coronary atherosclerosis is unclear. This study aimed to identify associations between the gut microbiome and computed tomography–based measures of coronary atherosclerosis and to explore relevant clinical correlates.METHODS: We conducted a cross-sectional study of 8973 participants (50 to 65 years of age) without overt atherosclerotic disease from the population-based SCAPIS (Swedish Cardiopulmonary Bioimage Study). Coronary atherosclerosis was measured using coronary artery calcium score and coronary computed tomography angiography. Gut microbiota species abundance and functional potential were assessed with shotgun metagenomics sequencing of fecal samples, and associations with coronary atherosclerosis were evaluated with multivariable regression models adjusted for cardiovascular risk factors. Associated species were evaluated for association with inflammatory markers, metabolites, and corresponding species in saliva.RESULTS: The mean age of the study sample was 57.4 years, and 53.7% were female. Coronary artery calcification was detected in 40.3%, and 5.4% had at least 1 stenosis with >50% occlusion. Sixty-four species were associated with coronary artery calcium score independent of cardiovascular risk factors, with the strongest associations observed for Streptococcus anginosus and Streptococcus oralis subsp oralis (P<1×10–5). Associations were largely similar across coronary computed tomography angiography–based measurements. Out of the 64 species, 19 species, including streptococci and other species commonly found in the oral cavity, were associated with high-sensitivity C-reactive protein plasma concentrations, and 16 with neutrophil counts. Gut microbial species that are commonly found in the oral cavity were negatively associated with plasma indole propionate and positively associated with plasma secondary bile acids and imidazole propionate. Five species, including 3 streptococci, correlated with the same species in saliva and were associated with worse dental health in the Malmö Offspring Dental Study. Microbial functional potential of dissimilatory nitrate reduction, anaerobic fatty acid β-oxidation, and amino acid degradation were associated with coronary artery calcium score.CONCLUSIONS: This study provides evidence of an association of a gut microbiota composition characterized by increased abundance of Streptococcus spp and other species commonly found in the oral cavity with coronary atherosclerosis and systemic inflammation markers. Further longitudinal and experimental studies are warranted to explore the potential implications of a bacterial component in atherogenesis.
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36.
  • Tóth, Gábor, et al. (författare)
  • Interleukin-13 Treatment of Living Lung Tissue Model Alters the Metabolome and Proteome : A Nano-DESI MS Metabolomics and Shotgun Proteomics Study
  • 2024
  • Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 25:9
  • Tidskriftsartikel (refereegranskat)abstract
    • Asthma is a chronic respiratory disease with one of the largest numbers of cases in the world; thus, constant investigation and technical development are needed to unravel the underlying biochemical mechanisms. In this study, we aimed to develop a nano-DESI MS method for the in vivo characterization of the cellular metabolome. Using air-liquid interface (ALI) cell layers, we studied the role of Interleukin-13 (IL-13) on differentiated lung epithelial cells acting as a lung tissue model. We demonstrate the feasibility of nano-DESI MS for the in vivo monitoring of basal-apical molecular transport, and the subsequent endogenous metabolic response, for the first time. Conserving the integrity of the ALI lung-cell layer enabled us to perform temporally resolved metabolomic characterization followed by "bottom-up" proteomics on the same population of cells. Metabolic remodeling was observed upon histamine and corticosteroid treatment of the IL-13-exposed lung cell monolayers, in correlation with alterations in the proteomic profile. This proof of principle study demonstrates the utility of in vivo nano-DESI MS for characterizing ALI tissue layers, and the new markers identified in our study provide a good starting point for future, larger-scale studies.
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37.
  •  
38.
  • Undin, Torgny, 1975- (författare)
  • Characterization of Molecular Adsorption Using Liquid Chromatography and Mass Spectrometry
  • 2015
  • Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
    • Molecular adsorption is a key feature in several disciplines of chemistry, covering as diverse fields as chromatographic separation to biomaterial development. This thesis aims at developing methods and techniques for the characterization of molecular adsorption at the liquid-solid interface. Two different experimental models were used, small molecular interaction characterization using liquid chromatography and complex protein adsorption on polymeric materials possible for biological sampling. Holistic approaches, where both detailed molecular interactions and identifications of trends, could improve the fundamental understanding of adsorption systems, were invariably part of the scientific process.The characterization of small molecular interactions on liquid chromatography stationary phases via adsorption isotherm determination used combined data from physical phase parameters i.e. carbon loading, linear-, and nonlinear-characterization methods. These experiments were conducted on high performance liquid chromatography systems, using both ordinary reversed phase stationary phases, and hybrid phases. The expansion of the improved elusion by characteristic point (ECP) for adsorption isotherm determination, led to that previous impossible isotherm types, having inflexion points, now could be determined by the method. It also reduced errors in isotherm parameters due to the elimination of inaccurate determined retention times where the mobile phase concentration was zero.The characterization of protein adsorption where performed in an unbiased way. Adsorbed proteins on different surfaces were identified using mass spectrometry (MS) and data dependent acquisition or a targeted method. Prior MS, an improved on surface enzymatic digestion (oSED) method was used to enable identification and quantitation of adsorbed protein originating from ventricular cerebrospinal fluid (vCSF). oSED was found to be able to experimentally determine large variations in protein adsorption characteristics between native and coated polycarbonate surfaces in contact with vCSF. The method was also confirmed being mechanistic in favor of enzymatic digestion of the proteins adsorbed on a surface, rather than a prior desorption into solution before digestion.An improvement of the overall understanding of adsorption systems was not only achieved with the oSED method as a promising tool for characterization of protein adsorption on arbitrary surfaces, but also the use of linear and nonlinear approaches in stationary phase characterization that strengthened drawn conclusions.
  •  
39.
  • Undin, Torgny, 1975-, et al. (författare)
  • Mass Spectrometric Determination of the Effect of Surface Deactivation on Membranes Used for In-Situ Sampling of Cerebrospinal Fluid (CSF)
  • 2018
  • Ingår i: Separations. - : MDPI AG. - 2297-8739. ; 5:2
  • Tidskriftsartikel (refereegranskat)abstract
    • In this paper, a strategy for structured monitoring of surface modifications to control protein adsorption to membrane structures is presented. The already established on-surface enzymatic digestion (oSED) method combined with nano-liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis was employed for the analysis of proteins in ventricular cerebrospinal fluid (vCSF) from neurointensive care patients. Protein adsorption was studied by in-situ sampling in a temporally resolved manner on both immobilized native and Pluronic-deactivated membranes. Deactivation was significantly reducing the protein adsorption but it also induced novel selective properties of the surface. The proposed versatile strategy will facilitate protein-biomaterial, protein-polymer, protein-protein interaction studies in the future.
  •  
40.
  • Undin, Torgny, 1975-, et al. (författare)
  • Mechanistic investigation of the on surface enzymatic digestion (oSED) protein adsorption detection method using targeted mass spectrometry
  • 2016
  • Ingår i: The Analyst. - : Royal Society of Chemistry (RSC). - 0003-2654 .- 1364-5528. ; 141:5, s. 1714-1720
  • Tidskriftsartikel (refereegranskat)abstract
    • This study describes our efforts to study some of the mechanistic aspects of the earlier established onsurface enzymatic digestion (oSED) method. In a multitude of application areas, it has become important to be able to fully characterize and understand selective protein adsorption to biomaterial surfaces for various applications, including biomedicine (implants), nanotechnology (microchip surfaces and sensors) and materials sciences. Herein, the investigation of the mechanistic aspects was based on microdialysis catheter tubes that were flushed with controlled protein solutions mimicking the extracellular fluid of the brain. The protein adsorption properties were monitored using high-resolution liquid chromatography tandem mass spectrometry (LC-MS/MS) with a targeted method. The temporally resolved results show that most proteins stay adsorbed onto the surface during the entire digestion process and are only cut away piece by piece, whereas smaller proteins and peptides seem to desorb rather easily from the surface. This information will simplify the interpretation of data generated using the oSED method and can also be used for the characterization of the physicochemical properties controlling the adsorption of individual proteins to specific surfaces.
  •  
41.
  • Undin, Torgny, et al. (författare)
  • MS for investigation of time-dependent protein adsorption on surfaces in complex biological samples
  • 2015
  • Ingår i: Future Science OA. - : Future Science Ltd. - 2056-5623. ; 1:4
  • Tidskriftsartikel (refereegranskat)abstract
    • Aim: This study aims at developing a nondestructive way for investigating proteinadsorption on surfaces such as biomaterials using mass spectrometry. Methods: Ventricular cerebrospinal fluid in contact with poly carbonate membranes were usedas adsorption templates and on-surface enzymatic digestion was applied to desorbproteins and cleave them into peptides. Mass spectrometric analysis provided bothprotein identification and determination of protein specific adsorption behavior. Results: In general, the adsorption increased with incubation time but also proteinspecifictime-resolved adsorption patterns from the complex protein solutionwere discovered. Conclusion: The method developed is a promising tool for thecharacterization of biofouling, which sometimes causes rejection and encapsulationof implants and can be used as complement to other surface analytical techniques.One problem associated with artificial materials in the body is that proteins in thebody interact with the surface, which sometimes causes rejection of the implant.In this study, a method for investigating the time-dependent protein adsorptionon a surface originating from a complex biological protein solution was developed.Compared with other surface analyses, this method can identify what proteins thatadsorbs on the surface. In addition, determination of protein-specific adsorptionbehavior in relation to incubation was possible
  •  
42.
  • Valdes, Alberto, et al. (författare)
  • Mass Spectrometry-Based Analysis of Time-Resolved Proteome Quantification
  • 2020
  • Ingår i: Proteomics. - : Wiley. - 1615-9853 .- 1615-9861. ; 20:9
  • Forskningsöversikt (refereegranskat)abstract
    • The aspect of time is essential in biological processes and thus it is important to be able to monitor signaling molecules through time. Proteins are key players in cellular signaling and they respond to many stimuli and change their expression in many time-dependent processes. Mass spectrometry (MS) is an important tool for studying proteins, including their posttranslational modifications and their interaction partners-both in qualitative and quantitative ways. In order to distinguish the different trends over time, proteins, modification sites, and interacting proteins must be compared between different time points, and therefore relative quantification is preferred. In this review, the progress and challenges for MS-based analysis of time-resolved proteome dynamics are discussed. Further, aspects on model systems, technologies, sampling frequencies, and presentation of the dynamic data are discussed.
  •  
43.
  • Valdés, Alberto, et al. (författare)
  • Proteomic comparison between different tissue preservation methods for identification of promising biomarkers of urothelial bladder cancer
  • 2021
  • Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Samples in biobanks are generally preserved by formalin-fixation and paraffin-embedding (FFPE) and/or optimal cutting temperature compound (OCT)-embedding and subsequently frozen. Mass spectrometry (MS)-based analysis of these samples is now available via developed protocols, however, the differences in results with respect to preservation methods needs further investigation. Here we use bladder urothelial carcinoma tissue of two different tumor stages (Ta/T1-non-muscle invasive bladder cancer (NMIBC), and T2/T3-muscle invasive bladder cancer (MIBC)) which, upon sampling, were divided and preserved by FFPE and OCT. Samples were parallel processed from the two methods and proteins were analyzed with label-free quantitative MS. Over 700 and 1200 proteins were quantified in FFPE and OCT samples, respectively. Multivariate analysis indicates that the preservation method is the main source of variation, but also tumors of different stages could be differentiated. Proteins involved in mitochondrial function were overrepresented in OCT data but missing in the FFPE data, indicating that these proteins are not well preserved by FFPE. Concordant results for proteins such as HMGCS2 (uniquely quantified in Ta/T1 tumors), and LGALS1, ANXA5 and plastin (upregulated in T2/T3 tumors) were observed in both FFPE and OCT data, which supports the use of MS technology for biobank samples and encourages the further evaluation of these proteins as biomarkers.
  •  
44.
  • Valdés, Alberto, et al. (författare)
  • Time-resolved proteomics of adenovirus infected cells
  • 2018
  • Ingår i: PLOS ONE. - : PUBLIC LIBRARY SCIENCE. - 1932-6203. ; 13:9
  • Tidskriftsartikel (refereegranskat)abstract
    • Viral infections cause large problems in the world and deeper understanding of the disease mechanisms is needed. Here we present an analytical strategy to investigate the host cell protein changes during human adenovirus type 2 (HAdV-C2 or Ad2) infection of lung fibro-blasts by stable isotope labelling of amino acids in cell culture (SILAC) and nanoLC-MS/MS. This work focuses on early phase of infection (6 and 12 h post-infection (hpi)) but the data is combined with previously published late phase (24 and 36 hpi) proteomics data to produce a time series covering the complete infection. As many as 2169 proteins were quantitatively monitored from 6 to 36 hpi, while some proteins were time-specific. After applying different filter criteria, 2027 and 2150 proteins were quantified at 6 and 12 hpi and among them, 431 and 544 were significantly altered at the two time points. Pathway analysis showed that the De novo purine and pyrimidine biosynthesis, Glycolysis and Cytoskeletal regulation by Rho GTPase pathways were activated early during infection while inactivation of the Integrin signalling pathway started between 6 and 12 hpi. Moreover, upstream regulator analysis predicted MYC to be activated with time of infection and protein and RNA data for genes controlled by this transcription factor showed good correlation, which validated the use of protein data for this prediction. Among the identified phosphorylation sites, a group related to glycolysis and cytoskeletal reorganization were up-regulated during infection. The results show specific aspects on how the host cell proteins, the final products in the genetic information flow, are influenced by Ad2 infection, which would be overlooked if only knowledge derived from mRNA data is considered.
  •  
45.
  • Westergren Jakobsson, Amanda, et al. (författare)
  • The Human Adenovirus 2 Transcriptome : An Amazing Complexity of Alternatively Spliced mRNAs
  • 2021
  • Ingår i: Journal of Virology. - : American Society for Microbiology. - 0022-538X .- 1098-5514. ; 95:4
  • Tidskriftsartikel (refereegranskat)abstract
    • We have used the Nanopore long-read sequencing platform to demonstrate how amazingly complex the human adenovirus type 2 (Ad2) transcriptome is with a flexible splicing machinery producing a range of novel mRNAs both from the early and late transcription units. In total we report more than 900 alternatively spliced mRNAs produced from the Ad2 transcriptome whereof more than 850 are novel mRNAs. A surprising finding was that more than 50% of all E1A transcripts extended upstream of the previously defined transcriptional start site. The novel start sites mapped close to the inverted terminal repeat (ITR) and within the E1A enhancer region. We speculate that novel promoters or enhancer driven transcription, so-called eRNA transcription, is responsible for producing these novel mRNAs. Their existence was verified by a peptide in the Ad2 proteome that was unique for the E1A ITR mRNA. Although we show a high complexity of alternative splicing from most early and late regions, the E3 region was by far the most complex when expressed at late times of infection. More than 400 alternatively spliced mRNAs were observed in this region alone. These mRNAs included extended L4 mRNAs containing E3 and L5 sequences and readthrough mRNAs combining E3 and L5 sequences. Our findings demonstrate that the virus has a remarkable capacity to produce novel exon combinations, which will offer the virus an evolutionary advantage to change the gene expression repertoire and protein production in an evolving environment.IMPORTANCE Work in the adenovirus system led to the groundbreaking discovery of RNA splicing and alternative RNA splicing in 1977. These mechanisms are essential in mammalian evolution by increasing the coding capacity of a genome. Here, we have used a long-read sequencing technology to characterize the complexity of human adenovirus pre-mRNA splicing in detail. It is mindboggling that the viral genome, which only houses around 36,000 bp, not being much larger than a single cellular gene, generates more than 900 alternatively spliced mRNAs. Recently, adenoviruses have been used as the backbone in several promising SARS-CoV-2 vaccines. Further improvement of adenovirus-based vaccines demands that the virus can be tamed into an innocent carrier of foreign genes. This requires a full understanding of the components that govern adenovirus replication and gene expression.
  •  
46.
  • Zhao, Hongxing, et al. (författare)
  • Distinct temporal changes in host cell lncRNA expression during the course of an adenovirus infection
  • 2016
  • Ingår i: Virology. - : Elsevier BV. - 0042-6822 .- 1096-0341. ; 492, s. 242-250
  • Tidskriftsartikel (refereegranskat)abstract
    • The deregulation of cellular long non-coding RNA (lncRNA) expression during a human adenovirus infection was studied by deep sequencing. Expression of lncRNAs increased substantially following the progression of the infection. Among 645 significantly expressed lncRNAs, the expression of 398 was changed more than 2-fold. More than 80% of them were up-regulated and 80% of them were detected during the late phase. Eased on the genomic locations of the deregulated lncRNAs in relation to known mRNAs and miRNAs, they were predicted to be involved in growth, structure, apoptosis and wound healing in the early phase, cell proliferation in the intermediate phase and protein synthesis, modification and transport in the late phase. The most significant functions of cellular RNA-binding proteins, previously shown to interact with the deregulated lncRNAs identified here, are involved in RNA splicing, nuclear export and translation events. We hypothesize that adenoviruses exploit the lncRNA network to optimize their reproduction.
  •  
47.
  • Zhao, Hongxing, et al. (författare)
  • Posttranscriptional regulation in adenovirus infected cells
  • 2017
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 16:2, s. 872-888
  • Tidskriftsartikel (refereegranskat)abstract
    • A deeper understanding of how viruses reprogramtheir hosts for production of progeny is needed to combatinfections. Most knowledge on the regulation of cellular geneexpression during adenovirus infection is derived from mRNAstudies. Here, we investigated the changes in protein expressionduring the late phase of adenovirus type 2 (Ad2) infection of theIMR-90 cell line by stable isotope labeling in cell culture withsubsequent liquid chromatography−high resolution tandemmass spectrometric analysis. Two biological replicates of samplescollected at 24 and 36 h post-infection (hpi) were investigated using swapped labeling. In total, 2648 and 2394 proteins werequantified at 24 and 36 hpi, respectively. Among them, 659 and 645 were deregulated >1.6-fold at the two time points. Theprotein expression was compared with RNA expression using cDNA sequencing data. The correlation was surprisingly low(r = 0.3), and several examples of posttranscriptional regulation were observed; e.g., proteins related to carbohydrate metabolismwere up-regulated at the protein level but unchanged at the RNA level, whereas histone proteins were down-regulated at theprotein level but up-regulated at the RNA level. The deregulation of cellular gene expression by adenovirus is mediated atmultiple levels and more complex than hitherto believed.
  •  
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