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  • 401.
    Wang, Chao
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Klechikov, Alexey G.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gharibyan, Anna L.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Wärmländer, Sebastian K. T. S.
    Jarvet, Jüri
    Zhao, Lina
    Jia, Xueen
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Shankar, S. K.
    Olofsson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Brännström, Thomas
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Mu, Yuguang
    Gräslund, Astrid
    Morozova-Roche, Ludmilla A.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    The role of pro-inflammatory S100A9 in Alzheimer's disease amyloid-neuroinflammatory cascade2014Ingår i: Acta Neuropathologica, ISSN 0001-6322, E-ISSN 1432-0533, Vol. 127, nr 4, s. 507-522Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Pro-inflammatory S100A9 protein is increasingly recognized as an important contributor to inflammation-related neurodegeneration. Here, we provide insights into S100A9 specific mechanisms of action in Alzheimer's disease (AD). Due to its inherent amyloidogenicity S100A9 contributes to amyloid plaque formation together with A beta. In traumatic brain injury (TBI) S100A9 itself rapidly forms amyloid plaques, which were reactive with oligomer-specific antibodies, but not with A beta and amyloid fibrillar antibodies. They may serve as precursor-plaques for AD, implicating TBI as an AD risk factor. S100A9 was observed in some hippocampal and cortical neurons in TBI, AD and non-demented aging. In vitro S100A9 forms neurotoxic linear and annular amyloids resembling A beta protofilaments. S100A9 amyloid cytotoxicity and native S100A9 pro-inflammatory signaling can be mitigated by its co-aggregation with A beta, which results in a variety of micron-scale amyloid complexes. NMR and molecular docking demonstrated transient interactions between native S100A9 and A beta. Thus, abundantly present in AD brain pro-inflammatory S100A9, possessing also intrinsic amyloidogenic properties and ability to modulate A beta aggregation, can serve as a link between the AD amyloid and neuroinflammatory cascades and as a prospective therapeutic target.

  • 402.
    Wang, Xi
    et al.
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Ning, Yujie
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Zhang, Feng
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Yu, Fangfang
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Tan, Wuhong
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Lei, Yanxia
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Wu, Cuiyan
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Zheng, Jingjing
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Wang, Sen
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Yu, Hanjie
    National Engineering Research Center for Miniaturized Detection Systems, Northwest University, Xi’an, China.
    Li, Zheng
    National Engineering Research Center for Miniaturized Detection Systems, Northwest University, Xi’an, China.
    Lammi, Mikko
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Guo, Xiong
    School of Public Health, Xi’an Jiaotong University Health Science Center, Key Laboratory of Trace Elements and Endemic Diseases, Xi'an, China.
    Gene expression signature in endemic osteoarthritis by microarray analysis2015Ingår i: International Journal of Molecular Sciences, ISSN 1422-0067, E-ISSN 1422-0067, Vol. 16, nr 5, s. 11465-11481Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Kashin-Beck Disease (KBD) is an endemic osteochondropathy with an unknown pathogenesis. Diagnosis of KBD is effective only in advanced cases, which eliminates the possibility of early treatment and leads to an inevitable exacerbation of symptoms. Therefore, we aim to identify an accurate blood-based gene signature for the detection of KBD. Previously published gene expression profile data on cartilage and peripheral blood mononuclear cells (PBMCs) from adults with KBD were compared to select potential target genes. Microarray analysis was conducted to evaluate the expression of the target genes in a cohort of 100 KBD patients and 100 healthy controls. A gene expression signature was identified using a training set, which was subsequently validated using an independent test set with a minimum redundancy maximum relevance (mRMR) algorithm and support vector machine (SVM) algorithm. Fifty unique genes were differentially expressed between KBD patients and healthy controls. A 20-gene signature was identified that distinguished between KBD patients and controls with 90% accuracy, 85% sensitivity, and 95% specificity. This study identified a 20-gene signature that accurately distinguishes between patients with KBD and controls using peripheral blood samples. These results promote the further development of blood-based genetic biomarkers for detection of KBD.

  • 403.
    Weber, Elvira
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
    Finsterbusch, Katja
    Innate Immunity and Infection, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Lindquist, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
    Nair, Sharmila
    Innate Immunity and Infection, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Lienenklaus, Stefan
    Department of Molecular Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Gekara, Nelson O
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Janik, Dirk
    Institute of Pathology, Helmholtz Center Munich, Neuherberg, Germany.
    Weiss, Siegfried
    Department of Molecular Immunology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Kalinke, Ulrich
    Institute for Experimental Infection Research, TWINCORE, Hannover, Germany.
    Överby, Anna K
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
    Kröger, Andrea
    Innate Immunity and Infection, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Type I interferon protects mice from fatal neurotropic infection with Langat virus by systemic and local antiviral responses2014Ingår i: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 89, nr 21, s. 12202-12212Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Vector-borne flaviviruses, such as tick-borne encephalitis virus (TBEV), West Nile virus, and dengue virus, cause millions of infections in humans. TBEV causes a broad range of pathological symptoms, ranging from meningitis to severe encephalitis or even hemorrhagic fever, with high mortality. Despite the availability of an effective vaccine, the incidence of TBEV infections is increasing. Not much is known about the role of the innate immune system in the control of TBEV infections. Here, we show that the type I interferon (IFN) system is essential for protection against TBEV and Langat virus (LGTV) in mice. In the absence of a functional IFN system, mice rapidly develop neurological symptoms and succumb to LGTV and TBEV infections. Type I IFN system deficiency results in severe neuroinflammation in LGTV-infected mice, characterized by breakdown of the blood-brain barrier and infiltration of macrophages into the central nervous system (CNS). Using mice with tissue-specific IFN receptor deletions, we show that coordinated activation of the type I IFN system in peripheral tissues as well as in the CNS is indispensable for viral control and protection against virus induced inflammation and fatal encephalitis. IMPORTANCE: The type I interferon (IFN) system is important to control viral infections; however, the interactions between tick-borne encephalitis virus (TBEV) and the type I IFN system are poorly characterized. TBEV causes severe infections in humans that are characterized by fever and debilitating encephalitis, which can progress to chronic illness or death. No treatment options are available. An improved understanding of antiviral innate immune responses is pivotal for the development of effective therapeutics. We show that type I IFN, an effector molecule of the innate immune system, is responsible for the extended survival of TBEV and Langat virus (LGTV), an attenuated member of the TBE serogroup. IFN production and signaling appeared to be essential in two different phases during infection. The first phase is in the periphery, by reducing systemic LGTV replication and spreading into the central nervous system (CNS). In the second phase, the local IFN response in the CNS prevents virus-induced inflammation and the development of encephalitis.

  • 404. Wentzel, Christian
    et al.
    Rajcan-Separovic, Evica
    Ruivenkamp, Claudia A L
    Chantot-Bastaraud, Sandra
    Metay, Corinne
    Andrieux, Joris
    Annerén, Göran
    Gijsbers, Antoinet C J
    Druart, Luc
    Hyon, Capucine
    Portnoi, Marie-France
    Stattin, Eva-Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Medicinsk och klinisk genetik.
    Vincent-Delorme, Catherine
    Kant, Sarina G
    Steinraths, Michelle
    Marlin, Sandrine
    Giurgea, Irina
    Thuresson, Ann-Charlotte
    Genomic and clinical characteristics of six patients with partially overlapping interstitial deletions at 10p12p112011Ingår i: European Journal of Human Genetics, ISSN 1018-4813, E-ISSN 1476-5438, Vol. 19, nr 9, s. 959-964Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    With the clinical implementation of genomic microarrays, the detection of cryptic unbalanced rearrangements in patients with syndromic developmental delay has improved considerably. Here we report the molecular karyotyping and phenotypic description of six new unrelated patients with partially overlapping microdeletions at 10p12.31p11.21 ranging from 1.0 to 10.6 Mb. The smallest region of overlap is 306 kb, which includes WAC gene, known to be associated with microtubule function and to have a role in cell division. Another patient has previously been described with a 10 Mb deletion, partially overlapping with our six patients. All seven patients have developmental delay and a majority of the patients have abnormal behaviour and dysmorphic features, including bulbous nasal tip, deep set eyes, synophrys/thick eyebrows and full cheeks, whereas other features varied. All patients also displayed various visual impairments and six out of seven patients had cardiac malformations. Taken together with the previously reported patient, our study suggests that the detected deletions may represent a new contiguous gene syndrome caused by dosage-sensitive genes that predispose to developmental delay.

  • 405. Williams, Jessica S.
    et al.
    Smith, Dana J.
    Marjavaara, Lisette
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Lujan, Scott A.
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Kunkel, Thomas A.
    Topoisomerase 1-Mediated Removal of Ribonucleotides from Nascent Leading-Strand DNA2013Ingår i: Molecular Cell, ISSN 1097-2765, E-ISSN 1097-4164, Vol. 49, nr 5, s. 1010-1015Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    RNase H2-dependent ribonucleotide excision repair (RER) removes ribonucleotides incorporated during DNA replication. When RER is defective, ribonucleotides in the nascent leading strand of the yeast genome are associated with replication stress and genome instability. Here, we provide evidence that topoisomerase 1 (Top1) initiates an independent form of repair to remove ribonucleotides from genomic DNA. This Top1-dependent process activates the S phase checkpoint. Deleting TOP1 reverses this checkpoint activation and also relieves replication stress and genome instability in RER-defective cells. The results reveal an additional removal pathway for a very common lesion in DNA, and they imply that the "dirty" DNA ends created when Top1 incises ribonucleotides in DNA are responsible for the adverse consequences of ribonucleotides in RNase H2-defective cells.

  • 406.
    Winestrand, Sandra
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Gandla, Madhavi Latha
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hong, Feng
    Chen, Qi Zhi
    Jönsson, Leif J
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Oxalate decarboxylase of Trametes versicolor: biochemical characterization and performance in bleaching filtrates from the pulp and paper industry2012Ingår i: Journal of chemical technology and biotechnology (1986), ISSN 0268-2575, E-ISSN 1097-4660, Vol. 87, nr 11, s. 1600-1606Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    BACKGROUND: Oxalate decarboxylase (ODC) from acid-induced cultures of the white-rot fungus Trametes versicolor was purified and characterized with respect to its biochemical properties and the possibility to utilize the enzyme for treatment of process water with the intention to prevent problems with calcium-oxalate scaling in the pulp and paper industry. RESULTS: Purified T. versicolor ODC was identified by tandem mass spectrometry. As estimated by using SDS-PAGE, the molecular mass was 69 kDa, and 60 kDa after deglycosylation with N-glycosidase F. The pH optimum was 2.5 and the temperature optimum was 4045 degrees C. The effects of ten potential inhibitors in industrial filtrates were examined. The enzyme was sensitive to low concentrations (0.1 mmol L-1) of chlorite and sulfite. T. versicolor ODC exhibited activity in 16 filtrates collected from mechanical pulping and kraft pulping. It had higher activity than ODC from Aspergillus niger in all of the filtrates and higher activity than oxalate oxidase from barley in all filtrates except two. CONCLUSIONS: The investigation shows basic biochemical properties of T. versicolor ODC and indicates that the enzyme may be useful for treatment of industrial filtrates under acidic conditions. Copyright (c) 2012 Society of Chemical Industry

  • 407.
    Wixner, Jonas
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Obayashi, Konen
    Department of Diagnostic Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan.
    Ando, Yukio
    Department of Diagnostic Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan.
    Karling, Pontus
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Anan, Intissar
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Loss of gastric interstitial cells of Cajal in patients with hereditary transthyretin amyloidosis2013Ingår i: Amyloid: Journal of Protein Folding Disorders, ISSN 1350-6129, E-ISSN 1744-2818, Vol. 20, nr 2, s. 99-106Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background: Hereditary transthyretin (TTR) amyloidosis is a systemic neuropathic disorder caused by TTR gene mutations. Gastrointestinal complications are common and the underlying mechanisms remain unclear. The interstitial cells of Cajal (ICC) function as pacemaker cells in the gastrointestinal tract and are important for gastrointestinal motility. The aim of this study was to investigate the densities of gastric ICC and nerves in patients with TTR amyloidosis compared to non-amyloidosis controls.

    Methods: Antral wall autopsy specimens from 11 Japanese ATTR V30M patients and 10 controls were analyzed with immunohistochemistry and computerized analysis. Antibodies to c-Kit and TMEM16A were used to assess ICC and an antibody to PGP 9.5 was used to assess nervous tissue. The study was approved by a Japanese ethical committee.

    Results: The densities of c-Kit-immunoreactive (IR) ICC were significantly lower in the circular and longitudinal muscle layers of patients compared to controls (p = 0.004 for both). Equivalent results were found for TMEM 16A-IR ICC. There were no significant differences in PGP 9.5-IR cells in the circular or longitudinal muscle layers between patients and controls (p = 0.173 and 0.099, respectively).

    Conclusions: A loss of gastrointestinal ICC may be an important factor for the digestive disturbances in hereditary TTR amyloidosis.

  • 408.
    Wixner, Jonas
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin.
    Westermark, Per
    Ihse, Elisabet
    Pilebro, Björn
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin.
    Lundgren, Hans-Erik
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin.
    Anan, Intissar
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin.
    The Swedish open-label diflunisal trial (DFNS01) on hereditary transthyretin amyloidosis and the impact of amyloid fibril composition2019Ingår i: Amyloid: Journal of Protein Folding Disorders, ISSN 1350-6129, E-ISSN 1744-2818, Vol. 26, s. 39-40Artikel i tidskrift (Refereegranskat)
  • 409.
    Wu, Chen
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Wang, Sihan
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi. Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Xu, Caihua
    Tyler, Andreas
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Li, Xingru
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Andersson, Charlotta
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi. Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Oji, Yusuke
    Sugiyama, Haruo
    Chen, Yijiang
    Li, Aihong
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    WT1 Enhances Proliferation and Impedes Apoptosis in KRAS Mutant NSCLC via Targeting cMyc2015Ingår i: Cellular Physiology and Biochemistry, ISSN 1015-8987, E-ISSN 1421-9778, Vol. 35, nr 2, s. 647-662Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background: A novel link between oncogenic KRAS signalling and WT1 was recently identified. We sought to investigate the role of WT1 and KRAS in proliferation and apoptosis. Methods: KRAS mutations and WT1 (cMyc) expression were detected using Sanger sequencing and real-time PCR in 77 patients with non-small cell lung cancer (NSCLC). Overexpression and knockdown of WT1 were generated with plasmid and siRNA via transient transfection technology in H1299 and H1568 cells. MTT assay for detection of cell proliferation, and TUNEL assay amd proteomic profiler assay for apoptosis evaluation were carried out. Dual luciferase reporter assay and ChIP-PCR were performed to validate the effect of WT1 on the cMyc promoter. Results: KRAS mutations showed a negative impact on overall survival ( OS). High expressions of WT1 and cMyc were associated with poor OS in KRAS mutant subgroup. The potential mechanisms that WT1 promotes proliferation and impedes apoptosis through affecting multiple apoptosis-related regulators in KRAS mutant NSCLC cells were identified. WT1 could activate cMyc promoter directly in KRAS mutant cells. Conclusion: The results suggest that WT1 and c-MYC expression is important for survival in KRAS mutant tumors as opposed to KRAS wild-type tumors. For treatment of KRAS mutant NSCLC, targeting WT1 and cMyc may provide alternative therapeutic strategies.

  • 410.
    Wu, H. D.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
    Mei, Ya-Fang
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
    Replication-competent adenovirus 11p vector armed with ADP gene at E1 region significantly improved tumour-killing effect on metastatic prostate cells in vitro and in vivo2017Ingår i: Human Gene Therapy, ISSN 1043-0342, E-ISSN 1557-7422, Vol. 28, nr 12, s. A29-A29Artikel i tidskrift (Övrigt vetenskapligt)
  • 411.
    Wu, Junfang
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Domellöf, Magnus
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk vetenskap, Pediatrik.
    Zivkovic, Angela M.
    Larsson, Göran
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Öhman, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Klinisk neurovetenskap.
    Nording, Malin L.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    NMR-based metabolite profiling of human milk: A pilot study of methods for investigating compositional changes during lactation2016Ingår i: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 469, nr 3, s. 626-632Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Low-molecular-weight metabolites in human milk are gaining increasing interest in studies of infant nutrition. In the present study, the milk metabolome from a single mother was explored at different stages of lactation. Metabolites were extracted from sample aliquots using either methanol water (MeOH/H2O) extraction or ultrafiltration. Nuclear magnetic resonance (NMR) spectroscopy was used for metabolite identification and quantification, and multi- and univariate statistical data analyses were used to detect changes over time of lactation. Compared to MeOH/H2O extraction, ultrafiltration more efficiently reduced the interference from lipid and protein resonances, thereby enabling the identification and quantification of 36 metabolites. The human milk metabolomes at the early (9-24 days after delivery) and late (31-87 days after delivery) stages of lactation were distinctly different according to multi- and univariate statistics. The late lactation stage was characterized by significantly elevated concentrations of lactose, choline, alanine, glutamate, and glutamine, as well as by reduced levels of citrate, phosphocholine, glycerophosphocholine, and N-acetylglucosamine. Our results indicate that there are significant compositional changes of the human milk metabolome also in different phases of the matured lactation stage. These findings complement temporal studies on the colostrum and transitional metabolome in providing a better understanding of the nutritional variations received by an infant.

  • 412.
    Wu, Jungfang
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Gouveia-Figueira, Sandra
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Farmakologi.
    Domellöf, Magnus
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk vetenskap, Pediatrik.
    Zivkovic, Angela M
    Nording, Malin
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Oxylipins, endocannabinoids, and related compounds in human milk: levels and effects of storage conditions2016Ingår i: Prostaglandins & other lipid mediators, ISSN 1098-8823, E-ISSN 2212-196X, Vol. 122, s. 28-36Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The presence of fatty acid derived oxylipins, endocannabinoids and related compounds in human milk may be of importance to the infant. Presently, clinically relevant protocols for storing and handling human milk that minimize error and variability in oxylipin and endocannabinoid concentrations are lacking. In this study, we compared the individual and combined effects of the following storage conditions on the stability of these fatty acid metabolites in human milk: state (fresh or frozen), storage temperature (4 °C, -20 °C or -80 °C), and duration (1 day, 1 week or 3 months). Thirteen endocannabinoids and related compounds, as well as 37 oxylipins were analyzed simultaneously by liquid chromatography coupled to tandem mass spectrometry. Twelve endocannabinoids and related compounds (2–111 nM) and 31 oxylipins (1.2 pM–1242 nM) were detected, with highest levels being found for 2-arachidonoylglycerol and 17(R)-hydroxydocosahexaenoic acid, respectively. The concentrations of most endocannabinoid-related compounds and oxylipins were dependent on storage condition, and especially storage at 4 °C introduced significant variability. Our findings suggest that human milk samples should be analyzed immediately after, or within one day of collection (if stored at 4 °C). Storage at -80 °C is required for long-term preservation, and storage at -20 °C is acceptable for no more than one week. These findings provide a protocol for investigating the oxylipin and endocannabinoid metabolome in human milk, useful for future milk-related clinical studies.

  • 413. Wu, Xuanjun
    et al.
    McKay, Craig
    Pett, Christian
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Leibniz-Institut für Analytische Wissenschaften ISAS e.V., Dortmund, Germany.
    Yu, Jin
    Schorlemer, Manuel
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Leibniz-Institut für Analytische Wissenschaften ISAS e.V., Dortmund, Germany.
    Ramadan, Sherif
    Lang, Shuyao
    Behren, Sandra
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Leibniz-Institut für Analytische Wissenschaften ISAS e.V., Dortmund, Germany.
    Westerlind, Ulrika
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Leibniz-Institut für Analytische Wissenschaften ISAS e.V., Dortmund, Germany.
    Finn, M. G.
    Huang, Xuefei
    Synthesis and Immunological Evaluation of Disaccharide Bearing MUC-1 Glycopeptide Conjugates with Virus-like Particles2019Ingår i: ACS Chemical Biology, ISSN 1554-8929, E-ISSN 1554-8937, Vol. 14, nr 10, s. 2176-2184Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Mucin-1 (MUC1) is a highly attractive antigenic target for anticancer vaccines. Naturally existing MUC1 can contain multiple types of O-linked glycans, including the Thomsen–Friedenreich (Tf) antigen and the Sialyl Thomsen-nouveau (STn) antigen. In order to target these antigens as potential anticancer vaccines, MUC1 glycopeptides SAPDT*RPAP (T* is the glycosylation site) bearing the Tf and the STn antigen, respectively, have been synthesized. The bacteriophage Qβ carrier is a powerful carrier for antigen delivery. The conjugates of MUC1-Tf and -STn glycopeptides with Qβ were utilized to immunize immune-tolerant human MUC1 transgenic (MUC1.Tg) mice, which elicited superior levels of anti-MUC1 IgG antibodies with titers reaching over 2 million units. The IgG antibodies recognized a wide range of MUC1 glycopeptides bearing diverse glycans. Antibodies induced by Qβ-MUC1-Tf showed strongest binding, with MUC1-expressing melanoma B16-MUC1 cells, and effectively killed these cells in vitro. Vaccination with Qβ-MUC1-Tf first followed by tumor challenge in a lung metastasis model showed significant reductions of the number of tumor foci in the lungs of immunized mice as compared to those in control mice. This was the first time that a MUC1-Tf-based vaccine has shown in vivo efficacy in a tumor model. As such, Qβ-MUC1 glycopeptide conjugates have great potential as anticancer vaccines.

  • 414.
    Wuolikainen, Anna
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Jonsson, Pär
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Ahnlund, Maria
    Antti, Henrik
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Marklund, Stefan L.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Klinisk kemi.
    Moritz, Thomas
    Forsgren, Lars
    Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Klinisk neurovetenskap.
    Andersen, Peter M.
    Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Klinisk neurovetenskap.
    Trupp, Miles
    Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Klinisk neurovetenskap.
    Multi-platform mass spectrometry analysis of the CSF and plasma metabolomes of rigorously matched amyotrophic lateral sclerosis, Parkinson's disease and control subjects2016Ingår i: Molecular Biosystems, ISSN 1742-206X, E-ISSN 1742-2051, Vol. 12, nr 4, s. 1287-1298Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Amyotrophic lateral sclerosis (ALS) and Parkinson's disease (PD) are protein-aggregation diseases that lack clear molecular etiologies. Biomarkers could aid in diagnosis, prognosis, planning of care, drug target identification and stratification of patients into clinical trials. We sought to characterize shared and unique metabolite perturbations between ALS and PD and matched controls selected from patients with other diagnoses, including differential diagnoses to ALS or PD that visited our clinic for a lumbar puncture. Cerebrospinal fluid (CSF) and plasma from rigorously age-, sex- and sampling-date matched patients were analyzed on multiple platforms using gas chromatography (GC) and liquid chromatography (LC)-mass spectrometry (MS). We applied constrained randomization of run orders and orthogonal partial least squares projection to latent structure-effect projections (OPLS-EP) to capitalize upon the study design. The combined platforms identified 144 CSF and 196 plasma metabolites with diverse molecular properties. Creatine was found to be increased and creatinine decreased in CSF of ALS patients compared to matched controls. Glucose was increased in CSF of ALS patients and alpha-hydroxybutyrate was increased in CSF and plasma of ALS patients compared to matched controls. Leucine, isoleucine and ketoleucine were increased in CSF of both ALS and PD. Together, these studies, in conjunction with earlier studies, suggest alterations in energy utilization pathways and have identified and further validated perturbed metabolites to be used in panels of biomarkers for the diagnosis of ALS and PD.

  • 415. Yamamoto, Shouji
    et al.
    Mitobe, Jiro
    Ishikawa, Takahiko
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Ohnishi, Makoto
    Watanabe, Haruo
    Izumiya, Hidemasa
    Regulation of natural competence by the orphan two-component system sensor kinase ChiS involves a non-canonical transmembrane regulator in Vibrio cholerae2014Ingår i: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 91, nr 2, s. 326-347Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In Vibrio cholerae, 41 chitin-inducible genes, including the genes involved in natural competence for DNA uptake, are governed by the orphan two-component system (TCS) sensor kinase ChiS. However, the mechanism by which ChiS controls the expression of these genes is currently unknown. Here, we report the involvement of a novel transcription factor termed TfoS' in this process. TfoS is a transmembrane protein that contains a large periplasmic domain and a cytoplasmic AraC-type DNA-binding domain, but lacks TCS signature domains. Inactivation of tfoS abolished natural competence as well as transcription of the tfoR gene encoding a chitin-induced small RNA essential for competence gene expression. A TfoS fragment containing the DNA-binding domain specifically bound to and activated transcription from the tfoR promoter. Intracellular TfoS levels were unaffected by disruption of chiS and coexpression of TfoS and ChiS in Escherichia coli recovered transcription of the chromosomally integrated tfoR::lacZ gene, suggesting that TfoS is post-translationally modulated by ChiS during transcriptional activation; however, this regulation persisted when the canonical phosphorelay residues of ChiS were mutated. The results presented here suggest that ChiS operates a chitin-induced non-canonical signal transduction cascade through TfoS, leading to transcriptional activation of tfoR.

  • 416.
    Yanamandra, Kiran
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Studies of in vivo prostate amyloidosis and autoimmune responses towards amyloid structures in neurodegeneration2010Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    By using multidisciplinary analysis of CA inclusions in prostate glands of patients diagnosed with prostate cancer, we have revealed that their major components are the amyloid forms of S100A8 and S100A9 proteins associated with numerous inflammatory conditions and types of cancer. We have demonstrated that material closely resembling CA can be produced from S100A8/A9 in vitro and shows the characters of amyloids. This process is facilitated by calcium or zinc, both of which are abundant in ex vivo inclusions. These observations were supported by computational analysis of the S100A8/A9 calcium-dependent aggregation propensity profiles. We have found DNA and proteins from Escherichia coli in CA bodies, suggesting that their formation is likely to be associated with bacterial infection. CA inclusions were also accompanied by the activation of macrophages and by an increase in the concentration of S100A8/A9 in the surrounding tissues, indicating inflammatory reactions. These findings, taken together, suggest a link between bacterial infection, inflammation and amyloid deposition of pro-inflammatory proteins S100A8/A9 in the prostate gland, such that a self-perpetuating cycle can be triggered and may increase the risk of malignancy in the ageing prostate.

    We evaluated the autoimmune reactions to endocrine (insulin) and astrocytical (S100B) biomarkers in the blood sera of PD patients compared with healthy controls. Peripheral immune responses can be sensitive indicators of disease pathology. We found a statistically significant increase of the autoimmune responses to both antigens in patients compared with controls. Heterogeneity of the immune responses observed in patients may reflect the modulating effect of multiple variables associated with neurodegeneration and also changes in the basic mechanisms of individual autoimmune reactivity. We did not detect any pronounced immune reactions towards insulin amyloid fibrils and oligomers in patients, indicating that an amyloid-specific conformational epitope is not involved in immune recognition of this amyloid type. Immune reactions towards S100B and insulin may reflect the neurodegenerative brain damaging processes and impaired insulin homeostasis occurring in PD.

    Generated auto-antibodies towards the major amyloidogenic protein involved in PD Lewy bodies - a-synuclein and its amyloid oligomers and fibrils were measured in the blood sera of early and late PD patients and controls by using ELISA, Western blot and Biacore surface plasmon resonance analyses. We found significantly higher antibody levels towards monomeric a-synuclein in the blood sera of PD patients compared to controls, though the responses decreased with PD progression. There were no noticeable immune responses towards amyloid oligomers, but substantially increased levels of IgGs towards a-synuclein amyloid fibrils both in PD patients and controls, which subsided with the disease progression. Pooled IgGs from PD patients and controls interacted also with amyloid fibrils of Ab (1-40) and hen lysozyme, however the latter were recognized with lower affinity. This suggests that IgGs bind to amyloid conformational epitope, though displaying higher specificity towards human amyloid species associated with neurodegeneration. The findings suggest the protective role of autoimmunity in PD and therefore immune reactions towards PD major amyloid protein - a-synuclein can be used in treatment strategies and in diagnostics, especially in identifying early disease.

  • 417.
    Yang, Hairu
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hultmark, Dan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    JAK/STAT and insulin signaling in Drosophila muscles regulate cellular immune responses against parasitoid wasp infectionManuskript (preprint) (Övrigt vetenskapligt)
  • 418.
    Yang, Hairu
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hultmark, Dan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). BioMediTech, University of Tampere, Tampere, Finland.
    Tissue communication in a systemic immune response of Drosophila.2016Ingår i: Fly, ISSN 1933-6942, Vol. 10, nr 3, s. 115-122Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Several signaling pathways, including the JAK/STAT and Toll pathways, are known to activate blood cells (hemocytes) in Drosophila melanogaster larvae. They are believed to regulate the immune response against infections by parasitoid wasps, such as Leptopilina boulardi, but how these pathways control the hemocytes is not well understood. Here, we discuss the recent discovery that both muscles and fat body take an active part in this response. Parasitoid wasp infection induces Upd2 and Upd3 secretion from hemocytes, leading to JAK/STAT activation mainly in hemocytes and in skeletal muscles. JAK/STAT activation in muscles, but not in hemocytes, is required for an efficient encapsulation of wasp eggs. This suggests that Upd2 and Upd3 are important cytokines, coordinating different tissues for the cellular immune response in Drosophila. In the fat body, Toll signaling initiates a systemic response in which hemocytes are mobilized and activated hemocytes (lamellocytes) are generated. However, the contribution of Toll signaling to the defense against wasps is limited, probably because the wasps inject inhibitors that prevent the activation of the Toll pathway. In conclusion, parasite infection induces a systemic response in Drosophila larvae involving major organ systems and probably the physiology of the entire organism.

  • 419.
    Yang, Hairu
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Kronhamn, Jesper
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Ekstrom, Jens-Ola
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Korkut, Gul Gizem
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hultmark, Dan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    JAK/STAT signaling in Drosophila muscles controls the cellular immune response against parasitoid infection2015Ingår i: EMBO Reports, ISSN 1469-221X, E-ISSN 1469-3178, Vol. 16, nr 12, s. 1664-1672Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The role of JAK/STAT signaling in the cellular immune response of Drosophila is not well understood. Here, we show that parasitoid wasp infection activates JAK/STAT signaling in somatic muscles of the Drosophila larva, triggered by secretion of the cytokines Upd2 and Upd3 from circulating hemocytes. Deletion of upd2 or upd3, but not the related os (upd1) gene, reduced the cellular immune response, and suppression of the JAK/STAT pathway in muscle cells reduced the encapsulation of wasp eggs and the number of circulating lamellocyte effector cells. These results suggest that JAK/STAT signaling in muscles participates in a systemic immune defense against wasp infection.

  • 420.
    Ylärinne, Janne
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB), Histologi med cellbiologi.
    Production of neocartilage tissues using primary chondrocytes2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Hyaline cartilage is a highly specialized tissue, which plays an important role in the articulating joints of an individual. It provides the joints with a nearly frictionless, impact resisting surface to protect the ends of the articulating bones. Articular cartilage has a poor self-repair capacity and, therefore, it rarely heals back to normal after an injury. Overweight, injuries, overloading and genetic factors may initiate a degenerative disease of the joint called osteoarthritis.

    Osteoarthiritis is a major global public health issue. Currently, the most used treatment for large articular cartilage defects is joint replacement surgery. However, possibilities to replace this highly invasive operation with strategies based on tissue engineering are currently investigated. The idea of the tissue engineering is to optimize the use of the cells, biomaterials and culture conditions to regenerate a new functional tissue for the defect site.

    The goal of this thesis was to manufacture cartilage tissue in cell culture conditions in vitro. Bovine primary chondrocytes isolated from the femoral condyles were used in all the experiments for neocartilage production. The samples were collected for histology, gene expression level quantifications, and analyses of proteoglycan (PG) content and quality. The histological sections were stained for type II collagen and PGs, the quantitative RT-PCR was used to observe the relative expressions of aggrecan, Sox9, procollagen α2(I) and procollagen α1(II) genes. The PGs were quantified using a spectrophotometric method, and agarose gel electrophoresis was used to separate the PGs according to their size.

    In the two first studies, we optimized the culture conditions of in vitro scaffold-free culture technique to produce the native-type hyaline cartilage of a good quality. We found out that high glucose concentration and hypertonic medium at 20% oxygen tension promoted the best hyaline-like neocartilage tissue production. Glucosamine sulfate supplementation, low oxygen tension, 5 mM glucose concentration and a transient TGF-β3 supplementation were not beneficial for the neocartilage formation in the scaffold-free cell culture system.

    In the third study, we used these newly defined, optimized culture conditions to produce the neocartilage tissues in the HyStem™ and the HydroMatrix™ scaffold materials and we compared these tissues to the ones grown as scaffold-free control cultures. We noticed that there was no difference between the controls and the scaffolds, and occasionally the scaffold-free controls had produced better quality cartilage than the ones with the scaffolds. Overall, the neocartilage tissues were of good hyaline-like quality in the third study. Their extracellular matrix contents were close to the native cartilage, although the neotissues lacked the zonal organization typical to the normal articular cartilage. The tissues had the right components, but their ultrastructure differed from the native cartilage.

    In conclusion, we were able to optimize our in vitro neocartilage culture method further, and discovered a good combination of the culture conditions to produce hyaline-like cartilage of good quality. Surprisingly, the scaffold materials were not beneficial for the cartilage formation.

     

  • 421.
    Ylärinne, Janne
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Qu, Chengjuan
    Department of Applied Physics, University of Eastern Finland, Kuopio, Finland.
    Lammi, Mikko
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB). School of Public Health, Health Science Center of Xi’an Jiaotong University, Key Laboratory of Trace Elements and Endemic Diseases, National Health and Family Planning Commission, Xi’an, P. R. China.
    Comparison of the neocartilages generated in scaffolds and scaffold-free agarose gel supported primary chondrocyte culture: Generation of neocartilage in vitro Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Objective: The present study was conducted to compare the neocartilages generated in scaffolds and scaffold-free, agarose gel-supported primary chondrocyte cultures.

    Design: Six million bovine primary chondrocytes were embedded in HyStem™ or HydroMatrix™ scaffolds, or scaffold-free in chondrocyte culture medium, and then loaded to agarose gel supported culture wells. Neocartilages were grown in the presence of hypertonic high glucose DMEM medium in 37 °C incubator at 20% O2 and 5% CO2 for one, three or six weeks. By the end of culture periods, the formed tissues were analyzed by histological staining for proteoglycans (PGs) and type II collagen, gene expression measurements of chondrocyte-specific genes aggrecan, Sox9 and procollagen α1(II), and procollagen α2(I) were performed using quantitative RT-PCR, and analyses of PG contents and structure were conducted by spectrophotometric and agarose gel electrophoretic methods, respectively.

    Results: The neocartilage generated in scaffold-free cultures appeared slightly bigger in size than in HyStem™- or HydroMatrix™-containing scaffolds. Histology visualized that the PGs and type II collagen were abundantly present in both in scaffold-free and scaffold-containing tissues. The PG content gradually increased following the culture period. However, the mRNA expression levels of the cartilage-specific genes of aggrecan, procollagen α1(II) and Sox9 gradually decreased following culture period, while procollagen α2(I) levels increased.

    Conclusions: After six weeks cultivations, the PG concentrations in neocartilage tissues manufactured with HyStem™ or HydroMatrix™ scaffolds, and in scaffold-free agarose gel-supported cell cultures, were similar to native cartilage. No obvious benefits could be seen on the extracellular matrix assembly in HyStem™ or HydroMatrix™ scaffolds cultures.

  • 422.
    Ylärinne, Janne
    et al.
    Department of Applied Physics, University of Eastern Finland, Kuopio, Finland.
    Qu, Chengjuan
    Department of Applied Physics, University of Eastern Finland, Kuopio, Finland.
    Lammi, Mikko
    Department of Applied Physics, University of Eastern Finland, Kuopio, Finland.
    Hypertonic conditions enhance cartilage formation in scaffold-free primary chondrocyte cultures2014Ingår i: Cell and Tissue Research, ISSN 0302-766X, E-ISSN 1432-0878, ISSN 1432-0878, Vol. 358, nr 2, s. 541-550Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The potential of hypertonic conditions at in vivo levels to promote cartilage extracellular matrix accumulation in scaffold-free primary chondrocyte cultures was investigated. Six million bovine primary chondrocytes were cultured in transwell inserts in low glucose (LG), high glucose (HG), or hypertonic high glucose (HHG) DMEM supplemented with fetal bovine serum, antibiotics, and ascorbate under 5 % or 20 % O2 tension with and without transforming growth factor (TGF)-β3 for 6 weeks. Samples were collected for histological staining of proteoglycans (PGs) and type II collagen, analysis by quantitative reverse transcription plus the polymerase chain reaction (RT-PCR) of mRNA expression of aggrecan and procollagen α1 (II) and of Sox9 and procollagen α2 (I), and quantitation of PGs and PG separation in agarose gels. Cartilage tissues produced at 20 % O2 tension were larger than those formed at 5 % O2 tension. Compared with LG, the tissues grew to larger sizes in HG or HHG medium. Histological staining showed the strongest PG and type II collagen staining in cartilage generated in HG or HHG medium at 20 % O2 tension. Quantitative RT-PCR results indicated significantly higher expression of procollagen α1 (II) mRNA in cartilage generated in HHG medium at 20 % O2 tension compared with that in the other samples. TGF-β3 supplements in the culture medium provided no advantage for cartilage formation. Thus, HHG medium used at 20 % O2 tension is the most beneficial combination of the tested culture conditions for scaffold-free cartilage production in vitro and should improve cell culture for research into cartilage repair or tissue engineering.

  • 423.
    Ylärinne, Janne
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Qu, Chengjuan
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Lammi, Mikko
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB). Xi’an Jiaotong Univ., Xi'an, China.
    HyStemTM and HydroMatrixTM scaffolds for articular cartilage tissue engineering2016Ingår i: Osteoarthritis and Cartilage, ISSN 1063-4584, E-ISSN 1522-9653, Vol. 24, s. S466-S466Artikel i tidskrift (Övrigt vetenskapligt)
    Abstract
  • 424. Yousefzadeh, Matthew J
    et al.
    Wyatt, David W
    Takata, Kei-Ichi
    Mu, Yunxiang
    Hensley, Sean C
    Tomida, Junya
    Bylund, Göran O
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Doublié, Sylvie
    Johansson, Erik
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Ramsden, Dale A
    McBride, Kevin M
    Wood, Richard D
    Mechanism of suppression of chromosomal instability by DNA polymerase POLQ2014Ingår i: PLOS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 10, nr 10, s. e1004654-Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Although a defect in the DNA polymerase POLQ leads to ionizing radiation sensitivity in mammalian cells, the relevant enzymatic pathway has not been identified. Here we define the specific mechanism by which POLQ restricts harmful DNA instability. Our experiments show that Polq-null murine cells are selectively hypersensitive to DNA strand breaking agents, and that damage resistance requires the DNA polymerase activity of POLQ. Using a DNA break end joining assay in cells, we monitored repair of DNA ends with long 3' single-stranded overhangs. End joining events retaining much of the overhang were dependent on POLQ, and independent of Ku70. To analyze the repair function in more detail, we examined immunoglobulin class switch joining between DNA segments in antibody genes. POLQ participates in end joining of a DNA break during immunoglobulin class-switching, producing insertions of base pairs at the joins with homology to IgH switch-region sequences. Biochemical experiments with purified human POLQ protein revealed the mechanism generating the insertions during DNA end joining, relying on the unique ability of POLQ to extend DNA from minimally paired primers. DNA breaks at the IgH locus can sometimes join with breaks in Myc, creating a chromosome translocation. We found a marked increase in Myc/IgH translocations in Polq-defective mice, showing that POLQ suppresses genomic instability and genome rearrangements originating at DNA double-strand breaks. This work clearly defines a role and mechanism for mammalian POLQ in an alternative end joining pathway that suppresses the formation of chromosomal translocations. Our findings depart from the prevailing view that alternative end joining processes are generically translocation-prone.

  • 425.
    Zamotin, Vladimir
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gharibyan, Anna
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gibanova, NV
    Lavrikova, MA
    Dolgikh, DA
    Kirpichnikov, MP
    Kostanyan, IA
    Morozova-Roche, Ludmilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Cytotoxicity of albebetin oligomers depends on cross-beta-sheet formation2006Ingår i: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 580, nr 10, s. 2451-2457Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Prefibrillar cytotoxicity was suggested as a common amyloid characteristic. We showed two types of albebetin prefibrillar oligomers are formed during incubation at pH 7.3. Initial round-shaped oligomers consist of 10–15 molecules determined by atomic force microscopy, do not bind thioflavin-T and do not affect viability of granular neurons and SH-SY5Y cells. They are converted into ca. 30–40-mers possessing cross-β-sheet and reducing viability of neuronal cells. Neither monomers nor fibrils possess cytotoxicity. We suggest that oligomeric size is important for stabilising cross-β-sheet core critical for cytotoxicity. As albebetin was used as a carrier-protein for drug delivery, examination of amyloidogenicity is required prior polypeptide biomedical applications.

  • 426.
    Zlatkov, Nikola
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Regulatory mechanisms involved in pathoadaptation of extraintestinal pathogenic Escherichia coli2019Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Establishment of commensal bacteria within a new niche of their host usually promotes the transition from commensalism to pathogenicity. Extraintestinal Pathogenic Escherichia coli (ExPEC) represent different pathovars with biphasic lifestyle – they can reside in the gut as commensals or they can escape and cause diseases elsewhere in the human body. Depending on the disease they are linked to, ExPEC can be divided into Uropathogenic E. coli (UPEC), Newborn Meningitis-causing E. coli (NMEC) and Sepsis-associated E. coli (SEPEC).

    Pathoadaptive mutations linked to c-di-GMP signaling were investigated in the NMEC strain IHE3034 which lacks the main global stress regulator RpoS. We investigated the role of ycgG2 in the lifestyle of NMEC. Deletion of ycgG2, shown by us to encode an YcgG allozyme with c-di-GMP phosphodiesterase (PDE) activity, and the restored RpoS led to a decrease in the S-fimbriae, otherwise robustly produced in artificial urine, hinting that the urinary tract could serve as a habitat for NMEC. We showed that NMEC were capable of aerobic citrate utilization in the presence of a co-substrate - a property that normally does not exist in E. coli. Our data hint that this metabolic upgrade is enhanced by the lack, or reduced activity, of c-di-GMP PDEs. We also found that citrate utilization is a property of ExPEC, since we reconstituted it in E. coli UTI89 (a cystitis isolate) via inactivation of its RpoS, and since a set of pyelonephritis E. coli isolates use citrate aerobically in the presence of glucose. The main reason for this metabolic capability is the absence of RpoS which leads to the production of the citrate transporter CitT. Furthermore, we highlighted the deletion of the fec operon (required for the ferric citrate uptake) in a large group of different ExPEC strains and we showed that NMEC can use CitT for in vitro ferric citrate uptake dependent on YcgG2 as an alternative system.

    Another pathoadaptive mutation which influences the fitness of ExPEC is the clyA (cytolysin A) gene inactivation, resulting from different deletions in different ExPEC genomes. When we restored the clyA+ locus, the UPEC strain 536 displayed increased susceptibility to antimicrobial peptides and urea. We also showed that the ClyA expression in 536 was increased by the presence of the DNA-binding regulator SfaX and another stand-alone PDE similar to YcgG2, called SfaY. The results were further confirmed by ClyA downregulation in NMEC deficient in SfaY and SfaX.

    We also studied the role of sfaY - a gene coding for another stand-alone c-di-GMP PDE. The expression of sfaY is under the regulation of the main promoter of the horizontally acquired sfa gene cluster. The latter is responsible for the regulation and assembly of the virulence-associated S-fimbriae, via which ExPEC bacteria bind to sialylated receptors. We found that NMEC are competent for filamentation because of a c-di-GMP-dependent program under the control of a phase-variation event which selectively turns ‘ON’ the sfa promoter in a subpopulation of bacteria. When SfaY is present, c-di-GMP levels are reduced, thus inducing the SOS stress response via the canonical LexA-RecA pathway. The signaling resulted in an internal differentiation of the bacterial population into a subpopulation exhibiting a filamentous morphotype (bacteria with induced SOS stress response) and a subpopulation of small motile and non-motile bacteria. Hence, this molecular program could serve as a clue to explain the formation of the intracellular bacterial communities observed during urinary tract infection by UPEC.

  • 427.
    Zlatkov, Nikola
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Absence of Global Stress Regulation in Escherichia coli Promotes Pathoadaptation and Novel c-di-GMP-dependent Metabolic Capability2019Ingår i: Scientific Reports, ISSN 2045-2322, Vol. 9, artikel-id 2600Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    athoadaptive mutations linked to c-di-GMP signalling were investigated in neonatal meningitis-causing Escherichia coli (NMEC). The results indicated that NMEC strains deficient in RpoS (the global stress regulator) maintained remarkably low levels of c-di-GMP, a major bacterial sessility-motility switch. Deletion of ycgG2, shown here to encode a YcgG allozyme with c-di-GMP phosphodiesterase activity, and the restoration of RpoS led to a decrease in S-fimbriae, robustly produced in artificial urine, hinting that the urinary tract could serve as a habitat for NMEC. We showed that NMEC were skilled in aerobic citrate utilization in the presence of glucose, a property that normally does not exist in E. coli. Our data suggest that this metabolic novelty is a property of extraintestinal pathogenic E. coli since we reconstituted this ability in E. coli UTI89 (a cystitis isolate) via deactivation rpoS; additionally, a set of pyelonephritis E. coli isolates were shown here to aerobically use citrate in the presence of glucose. We found that the main reason for this metabolic capability is RpoS inactivation leading to the production of the citrate transporter CitT, exploited by NMEC for ferric citrate uptake dependent on YcgG2 (an allozyme with c-di-GMP phosphodiesterase activity).

  • 428.
    Zlatkov, Nikola
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    C-di-GMP-mediated Morphotypic Pathoadaptability of Neonatal Meningitis Escherichia coliManuskript (preprint) (Övrigt vetenskapligt)
  • 429.
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    NMR studies of protein dynamics and structure2010Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Enzymes are extraordinary molecules that can accelerate chemical reactions by several orders of magnitude. With recent advancements in structural biology together with classical enzymology the mechanism of many enzymes has become understood at the molecular level. During the last ten years significant efforts have been invested to understand the structure and dynamics of the actual catalyst (i. e. the enzyme). There has been a tremendous development in NMR spectroscopy (both hardware and pulse programs) that have enabled detailed studies of protein dynamics. In many cases there exists a strong coupling between enzyme dynamics and function. Here I have studied the conformational dynamics and thermodynamics of three model systems: adenylate kinase (Adk), Peroxiredoxin Q (PrxQ) and the structural protein S16. By developing a novel chemical shift-based method we show that Adk binds its two substrates AMP and ATP with an extraordinarily dynamic mechanism. For both substrate-saturated states the nucleotide-binding subdomains exchange between open and closed states, with the populations of these states being approximately equal. This finding contrasts with the traditional view of enzyme-substrate complexes as static low entropy states. We are also able to show that the individual subdomains in Adk fold and unfold in a non-cooperative manner. This finding is relevant from a functional perspective, since it allows a change in hydrogen bonding pattern upon substrate-binding without provoking global unfolding of the entire enzyme (as would be expected from a two-state folding mechanism). We also studied the structure and dynamics of the plant enzyme PrxQ in both reduced and oxidized states. Experimentally validated structural models were generated for both oxidation states. The reduced state displays unprecedented μs-ms conformational dynamics and we propose that this dynamics reflects local and functional unfolding of an α-helix in the active site. Finally, we solved the structure of S16 from Aquifex aeolicus and propose a model suggesting a link between thermostability and structure for a mesophilic and hyperthermophilic protein pair. A connection between the increased thermostability in the thermophilic S16 and residual structure in its unfolded state was discovered, persistent at high denaturant concentrations, thereby affecting the difference in heat capacity difference between the folded and unfolded state. In summary, we have contributed to the understanding of protein dynamics and to the coupling between dynamics and catalytic activity in enzymes.

  • 430.
    Ådén, Jörgen
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wallgren, Marcus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Storm, Patrik
    Weise, Christoph
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Christiansen, Alexander
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Schröder, Wolfgang
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Funk, Christiane
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wolf-Watz, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Arabidopsis thaliana peroxiredoxin Q is extraordinarily dynamic on the μs-ms timescaleManuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Peroxiredoxin Q (PrxQ) isolated from Arabidopsis thaliana belongs to a family of redox enzymes called peroxiredoxins, which are thioredoxin- or glutaredoxin dependent peroxidases acting to reduce peroxides and in particular hydrogen peroxide. PrxQ cycles between an active reduced state and an inactive oxidized state during its catalytic cycle. The catalytic mechanism involves a nucleophilic attack of the catalytic cysteine on hydrogen peroxide to generate a sulfonic acid intermediate with a concerted release of a water molecule. This intermediate is subsequently relaxed by the reaction of a second cysteine, denoted as the resolving cysteine, generating an intermolecular disulphide bond to expel a second water molecule into solution. PrxQ is finally recycled to the active state by a thioredoxin dependent reduction. Previous structural studies of PrxQ homologues have provided the structural basis for the switch between reduced and oxidized conformations. Here we have performed a detailed study of the structure and dynamics of PrxQ in both the oxidized and reduced state. Reliable and experimentally validated structural models of PrxQ in both oxidation states were generated using homology based modeling. Model-free analyses of NMR spin relaxation show that PrxQ is monomeric in both oxidation states. As evident from fast R2 relaxation rates the reduced form of PrxQ undergoes unprecedented dynamics on the slow μs-ms timescale. The ground state of the conformational dynamics is likely the stably folded reduced state as implied by circular dichroism spectroscopy. We speculate that the extensive dynamics is intimately related to the catalytic function of PrxQ.

  • 431. Önnerfjord, Patrik
    et al.
    Khabut, Areej
    Reinholt, Finn P
    Svensson, Olle
    Umeå universitet, Medicinska fakulteten, Institutionen för kirurgisk och perioperativ vetenskap, Ortopedi.
    Heinegård, Dick
    Quantitative proteomic analysis of eight cartilaginous tissues reveals characteristic differences as well as similarities between subgroups2012Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, nr 23, s. 18913-18924Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Human synovial joints display a characteristic anatomic distribution of arthritis, e.g. rheumatoid arthritis primarily affects the metacarpophalangeal and proximal finger joints, but rarely the distal finger joints, whereas osteoarthritis occurs in the distal and proximal finger joints. Pelvospondylitis has a selective localization to the spine and sacroiliac joints. Is this tropism due to differences between the cartilages at the molecular level? To substantiate this concept the present study provides a background detailed compositional analysis by relative quantification of extracellular matrix proteins in articular cartilages, meniscus, intervertebral disc, rib, and tracheal cartilages on samples from 5-6 different individuals using an optimized approach for proteomics. Tissue extraction followed by trypsin digestion and two-dimensional LC separations coupled to tandem mass spectrometry, relative quantification with isobaric labeling, iTRAQ (TM), was used to compare the relative abundance of about 150 proteins. There were clear differences in protein patterns between different kinds of cartilages. Matrilin-1 and epiphycan were specific for rib and trachea, whereas asporin was particularly abundant in the meniscus. Interestingly, lubricin was prominent in the intervertebral disc, especially in the nucleus pulposus. Fibromodulin and lumican showed distributions that were mirror images of one other. Analyses of the insoluble residues from guanidine extraction revealed that a fraction of several proteins remained unextracted, e.g. asporin, CILP, and COMP, indicating cross-linking. Distinct differences in protein patterns may relate to different tissue mechanical properties, and to the intriguing tropism in different patterns of joint pathology.

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