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  • 1.
    Beltran-Pardo, Eliana
    et al.
    Pontificia Universidad Javeriana, Instituto de Genética Humana, Bogotá.
    Jönsson, Ingemar
    Kristianstad University, School of Education and Environment, Avdelningen för Naturvetenskap. Kristianstad University, Forskningsmiljön Man and Biosphere Health (MABH).
    Wojcik, Andrzej
    Department of Genetics, Microbiology and Toxicology, Stockholm University.
    Haghdoost, Siamak
    Department of Genetics, Microbiology and Toxicology, Stockholm University.
    Bermúdez-Cruz, Rosa María
    4Departamento de Genéticas y Biología Molecular, Centro de Investigación y Estudios Avanzados del Instituto Politécnico Nacional - CINVESTAV, Mexico City.
    Bernal Villegas, Jaime E.
    Pontificia Universidad Javeriana, Instituto de Genética Humana, Bogotá.
    Sequence analysis of the DNA-repair gene rad51 in the tardigrades Milnesium cf. tardigradum, Hypsibius dujardini and Macrobiotus cf. harmsworthi2013In: Journal of limnology, ISSN 1129-5767, E-ISSN 1723-8633, Vol. 72, no s1, p. 80-91Article in journal (Refereed)
    Abstract [en]

    Tardigrades are known for being resistant to extreme conditions, including tolerance to ionising and UV radiation in both the hydratedand the dehydrated state. It is known that these factors may cause damage to DNA. It has recently been shown that single and double DNAstrand breaks occur when tardigrades are maintained for a long time in the anhydrobiotic state. This may suggest that perhaps tardigrades rely on efficient DNA repair mechanisms. Among all proteins that comprise the DNA repair system, recombinases such as RecA or Rad51 have a very important function: DNA exchange activity. This enzyme is used in the homologous recombination and allows repair of thedamaged strand using homologous non-damaged strands as a template. In this study, Rad51 induction was evaluated by western blot in Milnesium cf. tardigradum, after exposure to gamma radiation. The Rad51 protein was highly induced by radiation, when compared to the control. The rad51 genes were searched in three tardigrades: Milnesium cf. tardigradum, Hypsibius dujardini and Macrobiotus cf. harmsworthi. The gene sequences were obtained by preparing and sequencing transcriptome libraries for H. dujardini and M. cf. harmsworthi and designing rad51 degenerate primers specific for M. cf. tardigradum. Comparison of Rad51 putative proteins from tardigrades with other organisms showed that they are highly similar to the corresponding sequence from the nematode Trichinella spiralis. A structure-based sequence alignment from tardigrades and other organisms revealed that putative Rad51 predicted proteins from tardigrades contain the expected motifs for these important recombinases. In a cladogram tree based on this alignment, tardigrades tend to cluster together suggesting that they have selective differences in these genes that make them diverge between species. Predicted Rad51 structures from tardigrades were also compared with crystalline structure of Rad51 in Saccharomyces cerevisiae. These results reveal that S. cerevisiae Rad51 structure is very similar to that of the three analysed tardigrades. On the other hand the predicted structure of Rad51 from M. cf. harmsworthi and H. dujardini are closer related to each other, than each of them to that of M. cf. tardigradum.

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  • 2.
    Borg, Johanna
    Kristianstad University, Faculty of Natural Sciences.
    Jämförelse av kemiinstrument och validering av referensintervall hos hund och katt2021Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    Clinical chemical analyzes has high clinical relevance. In serum/plasma, different parameters can be quantified. Parameters can be proteins, enzymes, ions, metals, lipids, and carbohydrates. With reference intervals, veterinarians can set diagnosis, follow treatment and development of the disease. Parameters are detected with different analysis principles/methods; colorimetry, immunoturbidimetry, enzymatic method and potentiometry. The animal hospital, AniCura, in Hässleholm accept dogs and cats. About 120 chemical analyzes are analyzed every day. AniCura purchased a new instrument, Indiko Plus, which will replace Cobas C111. Indiko Plus provide, more sample positions, 8 new analyzes, increased capacity, and facilitated use.

    The purpose of this study was to compare the instruments and produce a new reference interval which was compared to the reference interval provided by Thermo Fisher. To verify Indiko Plus, a precision and linearity study were conducted. Blood sampling of dogs and cats was performed by staff at AniCura. The comparison was made with patient samples and the reference intervals were obtained using samples from healthy animals.

    The comparison showed 9 of 13 analyzes had a statistically significant difference. The reason for this is probably due to the difference in reagents, the age of the instruments and the time between measurements. A new reference interval was developed and did not differ much from the Thermo Fisher interval. Further validation due to low population was recommended. The precision for Indiko Plus was approved. The linearity study shows not linear trend but was likely due to a bad pipette and should be redone.

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  • 3.
    Jönsson, Ingemar
    et al.
    Kristianstad University, School of Education and Environment, Avdelningen för Naturvetenskap. Kristianstad University, Forskningsmiljön Man and Biosphere Health (MABH).
    Beltran-Pardo, Eliana
    Pontificia Universidad Javeriana, Instituto de Genética Humana, Bogotá.
    Haghdoost, Siamak
    Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University.
    Wojcik, Andrzej
    Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University.
    Bermúdez-Cruz, Rosa María
    Centro de Investigación y Estudios Avanzados del Instituto Politécnico Nacional - CINVESTAV, Mexico City.
    Bernal Villegas, Jaime E.
    Pontificia Universidad Javeriana, Instituto de Genética Humana, Bogotá.
    Harms-Ringdahl, Mats
    Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University.
    Tolerance to gamma-irradiation in eggs of the tardigrade Richtersius coronifer depends on stage of development2013In: Journal of limnology, ISSN 1129-5767, E-ISSN 1723-8633, Vol. 72, no s1, p. 73-79Article in journal (Refereed)
    Abstract [en]

    Tardigrades are known as one of the most radiation tolerant animals on Earth, and several studies on tolerance in adult tardigrades have been published. In contrast, very few studies on radiation tolerance of embryonic stages have been reported. Here we report a study on tolerance to gamma irradiation in eggs of the eutardigrade Richtersius coronifer. Irradiation of eggs collected directly from a natural substrate (moss) showed a clear dose-response, with a steep decline in hatchability at doses up to 0.4 kGy followed by a relatively constant hatchability around 25% up to 2 kGy, and a decline to ca. 5% at 4 kGy above which no eggs hatched. Analysis of the time required for eggs to hatch after irradiation (residual development time) showed that hatching of eggs after exposure to high doses of gamma radiation was associated with short residual development time. Since short residual development time means that the egg was irradiated at a late developmental stage, this suggests that eggs were more tolerant to radiation late in development. This was also confirmed in another experiment in which stage of development at irradiation was controlled. No eggs irradiated at the early developmental stage hatched, and only one egg at middle stage hatched, while eggs irradiated in the late stage hatched at a rate indistinguishable from controls. This suggests that the eggs are more sensitive to radiation in the early stages of development, or that tolerance to radiation is acquired only late in development, shortly before the eggs hatch, hypotheses that are not mutually exclusive. Our study emphasizes the importance of considering specific cell cycle phases and developmental stages in studies of tolerance to radiation in tardigrades, and the potential importance of embryonic studies in revealing the mechanisms behind the radiation tolerance of tardigrades and other cryptobiotic animals.

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  • 4.
    Nasir, Irem
    et al.
    Lund University.
    Fatih, Warda
    Upper 2nd school, Klippan, Sweden.
    Svensson, Anja
    Upper 2nd school, Klippan, Sweden.
    Radu, Dennis
    Upper 2nd school, Klippan, Sweden.
    Linse, Sara
    Lund University.
    Cabaleiro-Lago, Celia
    Lund University.
    Lundqvist, Martin
    Lund University.
    High throughput screening method to explore protein interactions with nanoparticles2015In: PLOS ONE, E-ISSN 1932-6203, Vol. 10, no 8Article in journal (Refereed)
    Abstract [en]

    The interactions of biological macromolecules with nanoparticles underlie a wide variety of current and future applications in the fields of biotechnology, medicine and bioremediation. The same interactions are also responsible for mediating potential biohazards of nanomaterials. Some applications require that proteins adsorb to the nanomaterial and that the protein resists or undergoes structural rearrangements. This article presents a screening method for detecting nanoparticle-protein partners and conformational changes on time scales ranging from milliseconds to days. Mobile fluorophores are used as reporters to study the interaction between proteins and nanoparticles in a high-throughput manner in multi-well format. Furthermore, the screening method may reveal changes in colloidal stability of nanomaterials depending on the physicochemical conditions.

  • 5.
    Nasir, Irem
    et al.
    Lund University.
    Linse, Sara
    Lund University.
    Cabaleiro-Lago, Celia
    Lund University.
    Fluorescent filter-trap assay for amyloid fibril formation kinetics in complex solutions2015In: ACS Chemical Neuroscience, E-ISSN 1948-7193, Vol. 6, no 8, p. 1436-44Article in journal (Refereed)
    Abstract [en]

    Amyloid fibrils are the most distinct components of the plaques associated with various neurodegenerative diseases. Kinetic studies of amyloid fibril formation shed light on the microscopic mechanisms that underlie this process as well as the contributions of internal and external factors to the interplay between different mechanistic steps. Thioflavin T is a widely used noncovalent fluorescent probe for monitoring amyloid fibril formation; however, it may suffer from limitations due to the unspecific interactions between the dye and the additives. Here, we present the results of a filter-trap assay combined with the detection of fluorescently labeled amyloid β (Aβ) peptide. The filter-trap assay separates formed aggregates based on size, and the fluorescent label attached to Aβ allows for their detection. The times of half completion of the process (t1/2) obtained by the filter-trap assay are comparable to values from the ThT assay. High concentrations of human serum albumin (HSA) and carboxyl-modified polystyrene nanoparticles lead to an elevated ThT signal, masking a possible fibril formation event. The filter-trap assay allows fibril formation to be studied in the presence of those substances and shows that Aβ fibril formation is kinetically inhibited by HSA and that the amount of fibrils formed are reduced. In contrast, nanoparticles exhibit a dual-behavior governed by their concentration.

  • 6.
    Szczepankiewicz, Olga
    et al.
    Lund University.
    Cabaleiro-Lago, Celia
    Lund University.
    Tartaglia, Gian Gaetano
    England.
    Vendruscolo, Michele
    England.
    Hunter, Thérèse
    Malta.
    Hunter, Gary J
    Malta.
    Nilsson, Hanna
    Lund University.
    Thulin, Eva
    Lund University.
    Linse, Sara
    Lund University.
    Interactions in the native state of monellin, which play a protective role against aggregation2011In: Molecular Biosystems, ISSN 1742-206X, E-ISSN 1742-2051, Vol. 7, no 2, p. 521-32Article in journal (Refereed)
    Abstract [en]

    A series of recent studies have provided initial evidence about the role of specific intra-molecular interactions in maintaining proteins in their soluble state and in protecting them from aggregation. Here we show that the amino acid sequence of the protein monellin contains two aggregation-prone regions that are prevented from initiating aggregation by multiple non-covalent interactions that favor their burial within the folded state of the protein. By investigating the behavior of single-chain monellin and a series of five of its mutational variants using a variety of biochemical, biophysical and computational techniques, we found that weakening of the non-covalent interaction that stabilizes the native state of the protein leads to an enhanced aggregation propensity. The lag time for fibrillation was found to correlate with the apparent midpoint of thermal denaturation for the series of mutational variants, thus showing that a reduced thermal stability is associated with an increased aggregation tendency. We rationalize these findings by showing that the increase in the aggregation propensity upon mutation can be predicted in a quantitative manner through the increase in the exposure to solvent of the amyloidogenic regions of the sequence caused by the destabilization of the native state. Our findings, which are further discussed in terms of the structure of monellin and the perturbation by the amino acid substitutions of the contact surface between the two subdomains that compose the folded state of monellin, provide a detailed description of the specific intra-molecular interactions that prevent aggregation by stabilizing the native state of a protein.

  • 7.
    Sävblom, Charlotta
    et al.
    Lund University, Skåne University Hospital, Malmö.
    Halldén, Christer
    Kristianstad University, School of Education and Environment, Avdelningen för Naturvetenskap.
    Cronin, Angel M
    Memorial Sloan-Kettering Cancer Center, New York.
    Säll, Torbjörn
    Department of Cell and Organism Biology, Lund University, Lund.
    Savage, Caroline
    Memorial Sloan-Kettering Cancer Center, New York.
    Vertosick, Emily A
    Memorial Sloan-Kettering Cancer Center, New York.
    Klein, Robert J
    Memorial Sloan-Kettering Cancer Center, New York.
    Giwercman, Aleksander
    Memorial Sloan-Kettering Cancer Center, New York.
    Lilja, Hans
    Lund University, Skåne University Hospital, Malmö.
    Genetic variation in KLK2 and KLK3 is associated with concentrations of hK2 and PSA in serum and seminal plasma in young men2014In: Clinical Chemistry, ISSN 0009-9147, E-ISSN 1530-8561, Vol. 60, no 3, p. 490-499Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Genetic variants in KLK2 and KLK3 have been associated with increased serum concentrations of their encoded proteins, human kallikrein-related peptidase 2 (hK2) and prostate-specific antigen (PSA), and with prostate cancer in older men. Low PSA concentrations in seminal plasma (SP) have been associated with low sperm motility. To evaluate whether KLK2 and KLK3 genetic variants affect physiological prostatic secretion, we studied the association of SNPs with hK2 and PSA concentrations in SP and serum of young, healthy men.

    METHODS: Leukocyte DNA was extracted from 303 male military conscripts (median age 18.1 years). Nine SNPs across KLK2-KLK3 were genotyped. We measured PSA and hK2 in SP and serum using immunofluorometric assays. The association of genotype frequencies with hK2 and PSA concentrations was tested with the Kruskal-Wallis test.

    RESULTS: Four KLK2 SNPs (rs198972, rs198977, rs198978, and rs80050017) were strongly associated with hK2 concentrations in SP and serum, with individuals homozygous for the major alleles having 3- to 7-fold higher concentrations than the intermediate concentrations found in other homozygotes and heterozygotes (all P < 0.001). Three of these SNPs were significantly associated with percentage of free PSA (%fPSA) in serum (all P < 0.007). Three KLK3 SNPs showed associations with PSA in SP, and the rs1058205 SNP was associated with total PSA in serum (P = 0.001) and %fPSA (P = 0.015).

    CONCLUSIONS: Associations observed in young, healthy men between the SP and serum concentrations of hK2 and PSA and several genetic variants in KLK2 and KLK3 could be useful to refine models of PSA cutoff values in prostate cancer testing.

  • 8.
    Ter Schure, Arnout F.H.
    et al.
    Lund University.
    Larsson, Per
    Lund University.
    Merilä, Juha
    Finland.
    Jönsson, K. Ingemar
    Lund University.
    Latitudinal Fractionation of Polybrominated Diphenyl Ethers and Polychlorinated Biphenyls in Frogs (Rana temporaria)2002In: Environmental Science and Technology, ISSN 0013-936X, E-ISSN 1520-5851, Vol. 36, no 23, p. 5057-5061Article in journal (Refereed)
    Abstract [en]

    The flame retardant polybrominated diphenyl ethers (PBDEs) have become ubiquitous environmental pollutants. The environmental distribution of PBDEs is much less studied than that of the polychlorinated biphenyls (PCBs).To compare the environmental fate of the PCBs withtheir partial substitute, the PBDEs, common frogs (Ranatemporaria) were collected along a 1500-km-long latitudinal gradient of the Scandinavian Peninsula and their livers analyzed for PCBs and PBDEs. Mean concentrations of total PCBs and BDE47 ranged from 9200 to 92 900 and 30 to120 ng kg-1 fresh weight, respectively, whereas BDE99 was detected in less than 50% of the frogs. PCB concentrations were higher than that of the PBDEs, and the differences decreased in the northern latitudes. Moreover, the pollutant concentrations in frog livers were negative functions of latitude. The observed scatter and regression slopes imply several influencing factors (such as habitat, exposure route, uptake, metabolism, excretion, etc.) and indicate release events instead of the grasshopper effect. Biological variables such as gender, age, body size, and lipid content did not influence pollutant concentrations in the frog livers. The total PCB and BDE47 concentrations in frogs were highly correlated. Hence, their environmental fate is analogous and our results highlight the need to examine the potential role of xenobiotics on amphibian populations.

  • 9.
    Zorita, Saioa
    et al.
    Division of Analytical Chemistry, University of Lund.
    Hallgren, Pär
    Kristianstad University College, Department of Mathematics and Science.
    Mathiasson, Lennart
    Division of Analytical Chemistry, University of Lund.
    Steroid hormone determination in water using an environmentally friendly membrane based extraction technique2008In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1192, no 1, p. 1-8Article in journal (Refereed)
    Abstract [en]

    In this study, a method was developed for determination of steroid hormones (17beta-estradiol, estrone, 17alpha-ethynylestradiol) in tap and sewage water samples from Sweden. Sample preparation and analysis were performed by a hollow-fibre microporous membrane liquid-liquid extraction (HF-MMLLE) set-up combined with gas chromatography-mass spectrometry (GC-MS). In this approach, only the organic liquid in the lumen (10microL) of the hollow-fibre membrane was utilised for depleting extraction. Several parameters were studied, including: type of organic solvent, sample pH, salt and humic acid content. The optimised method allowed the determination of the analyte at the low ngL(-1) level in tap and sewage water. A linear plot gave correlation coefficients better than 0.995 and resulted in a method limit of detection of 1.6, 3 and 10ngL(-1) for 17beta-estradiol, estrone, and 17alpha-ethynylestradiol, respectively, in sewage water. Enrichment factors were over 1400 after derivatisation. The repeatabilities at 50 and 600ngL(-1) were better than 10% and 6%, respectively.

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