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| 5. |
- Eriksson, Oskar, et al.
(författare)
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Neutrophil CD64 expression - comparison of two different flow cytometry protocols on EPICs MCL and the Leuko64 (TM) assay on a Celldyn Sapphire haematology analyser
- 2015
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Ingår i: Scandinavian Journal of Clinical and Laboratory Investigation. - : Informa UK Limited. - 0036-5513 .- 1502-7686. ; 75:5, s. 428-433
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Tidskriftsartikel (refereegranskat)abstract
- Objective. To evaluate the Trillium Diagnostics Leuko64 (TM) assay on Abbott Celldyn Sapphire haematology analyser compared to two flow cytometry protocols on Beckman Coulter EPICS MCL flow cytometer. Materials and methods. CD64 expression on neutrophils was determined by two flow cytometry protocols and by a commercial assay on an automatic haematology analyser. The inclusion of study subjects was based on elevated procalcitonin (PCT) values, identifying patients where a systemic infection was suspected. Healthy blood donors were used as a reference group. Results. Statistically significant correlations between the Trillium Diagnostics Leuko64 (TM) assay and the flow cytometry methods were found when measuring neutrophil CD64 expression. Conclusions. The good correlation between a reference method and an automated haematology analyser method for CD64 expression on neutrophils supports introduction of the latter assay for routine use as an independent biomarker of bacterial infection and inflammation.
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| 6. |
- Fadeel, Bengt, et al.
(författare)
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Kostmann disease and other forms of severe congenital neutropenia
- 2021
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Ingår i: Acta Paediatrica. - : John Wiley & Sons. - 0803-5253 .- 1651-2227. ; 110:11, s. 2912-2920
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Tidskriftsartikel (refereegranskat)abstract
- Congenital neutropenia with autosomal recessive inheritance was first described by the Swedish paediatrician Rolf Kostmann who coined the term ‘infantile genetic agranulocytosis’. The condition is now commonly referred to as Kostmann disease. These patients display a maturation arrest of the myelopoiesis in the bone marrow and reduced neutrophil numbers and suffer from recurrent, often life-threatening infections. The molecular mechanism underlying congenital neutropenia has been intensively investigated, and mutations in genes that impinge on programmed cell death have been identified. The present review provides an overview of these studies.
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| 7. |
- Garwicz, Daniel
(författare)
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Biosynthesis, Processing, and Sorting of Human Neutrophil Granule Proteins. Focus on Cathepsin G, Proteinase 3, and Azurocidin
- 2000
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The neutrophil granulocyte contains a number of multifunctional proteins stored in different subsets of granules. The azurophil granules, synthesized early during differentiation, harbour leukocyte elastase, cathepsin G, proteinase 3, and azurocidin; the members of the serprocidin family. In order to study the biosynthesis, processing, and intracellular sorting of the proteins, human cDNA encoding the latter three proteins, in their preproform, was stably transfected to two hematopoietic rodent cell lines (RBL-1 and 32Dcl3). To investigate the importance of distinct protein motifs, mutations were made in the cDNAs prior to transfection. Cathepsin G lacking functional glycosylation site was normally activated and sorted to granules. Similarly, cathepsin G, lacking both functional catalytic site and amino-terminal propeptide, was stably transfected to both cell lines. However, preterm activation of catalytically active cathepsin G lead to impaired cell survival. Proteinase 3 was expressed and stored in a form that allowed specific recognition by human cANCA-sera from patients with Wegener's granulomatosis. The amino-terminal heptapropeptide of azurocidin was shown to be removed in a stepwise manner involving at least two different enzymes. All serprocidins acquired high-mannose oligosaccharides that were converted into complex forms. Finally, the biosynthetic windows of several neutrophil granule proteins, including myeloperoxidase, lysozyme, cathepsin D, and all four serprocidins, were characterized in an experimental in vitro model for differentiation of human bone marrow progenitor cells into neutrophils.
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- Garwicz, Daniel, et al.
(författare)
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Early recognition of reverse pseudohyperkalemia in heparin plasma samples during leukemic hyperleukocytosis can prevent iatrogenic hypokalemia
- 2012
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Ingår i: Clinical Biochemistry. - : Elsevier BV. - 0009-9120 .- 1873-2933. ; 45:18, s. 1700-1702
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Tidskriftsartikel (refereegranskat)abstract
- Objectives: To investigate the cause of apparent hyperkalemia in leukemic heparin plasma. Design and methods: Lithium heparin plasma and serum samples from a patient with chronic lymphocytic leukemia (CLL) with hyperleukocytosis were transported by either a pneumatic tube system or manual transport and analyzed either immediately or after 4 h. Results: Pneumatic tube transported samples resulted in higher plasma potassium levels than manually transported samples. Serum potassium was lower than plasma potassium, confirming the suspicion of "reverse" pseudohyperkalemia. Letting the pneumatic tube transported samples stand on the bench for 4 h before centrifugation surprisingly resulted in decreased or unchanged plasma potassium. Conclusions: The reverse pseudohyperkalemia in heparin plasma samples from a CLL patient was caused by pneumatic tube transport. Our results suggest extracellular leakage of potassium, followed by active transport of potassium into intact leukemic cells. This is the first Swedish case of reverse pseudohyperkalemia in a CLL patient, where clinical suspicion of false hyperkalemia and awareness of the phenomenon lead to a rapid laboratory diagnosis. The demonstration of reverse pseudohyperkalemia prevented potentially dangerous medical interventions, such as potassium lowering treatment.
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| 11. |
- Garwicz, Daniel, et al.
(författare)
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Farmakogenetisk analys kan avslöja risk för statinbiverkningar : [Pharmacogenetic analysis can predict adverse effects of statins]
- 2013
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Ingår i: Läkartidningen. - 0023-7205 .- 1652-7518. ; 110:19-20, s. 951-952
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Tidskriftsartikel (refereegranskat)abstract
- Mer än var tionde vuxen i Sverige behandlas med statiner. Muskelsvaghet, -trötthet och -värk är kända biverkningar. I sällsynta fall ses rabdomyolys, som kan leda till akut njursvikt och någon gång dödsfall. Statiners kemiska egenskaper och serumkoncentration påverkar risken för allvarliga biverkningar. Serumkoncentrationen beror på dos och på patientens förmåga att omsätta läkemedlet.Akademiska sjukhuset har som första svenska sjukhus infört analys av en genetisk variant (SLCO1B1*5) som kan förutsäga ökad risk för sällsynta, allvarliga muskelbiverkningar vid statinbehandling.
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| 14. |
- Garwicz, Daniel, et al.
(författare)
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Normal levels of constitutive and death receptor-mediated apoptosis of peripheral blood neutrophils from patients with chronic idiopathic neutropenia.
- 2007
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Ingår i: Clinical Immunology. - : Elsevier BV. - 1521-6616 .- 1521-7035. ; 122:3, s. 349-355
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Tidskriftsartikel (refereegranskat)abstract
- To investigate the role of neutrophil apoptosis in the pathogenesis of chronic neutropenia, we examined constitutive and death receptor-mediated apoptosis ex vivo of peripheral blood neutrophils obtained from six chronic idiopathic neutropenia (CIN) patients and six healthy adult blood donors. Apoptosis was quantified based on phosphatidylserine externalization and caspase-3 activation in freshly isolated neutrophils or after overnight cultivation of neutrophils in the absence or presence of pro- or anti-apoptotic factors, including the pan-caspase inhibitor, zVAD-fmk. Neutrophils from CIN patients receiving treatment with granulocyte colony-stimulating factor appeared to be more prone to constitutive apoptosis than cells from untreated patients; however, further investigations in larger cohorts of patients are needed to validate these pilot studies. Overall, the level of neutrophil apoptosis was similar in patient and control groups, thus supporting the notion that the underlying defect in these neutropenia patients lies elsewhere, such as in the bone marrow microenvironment.
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- Gullberg, U, et al.
(författare)
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Biosynthesis, processing and sorting of neutrophil proteins : insight into neutrophil granule development
- 1997
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Ingår i: European Journal of Haematology. - : Wiley. - 0902-4441 .- 1600-0609. ; 58:3, s. 137-153
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil granulocytes are specialized phagocytic cells that carry a collection of granules for regulated secretion, each with distinct constituents. The granules can be classified as azurophil (primary), developed first, followed in time by specific (secondary) granules gelatinase granules, and secretory vesicles. Stage- and tissue-specific transcription factors govern the successive expression of genes for granule proteins to allow storage of the gene products in these organelle categories whose packaging is separated in time. Many of the granule proteins, in particular those of the heterogeneous lysosome-like azurophil granules, are subject to extensive post-translational proteolytic processing into mature proteins, most commonly as a post-sorting event. A selective aggregation of proteins destined for storage in granules, as discussed in this review, would facilitate their retention and eliminate a need for distinct sorting motifs on each granule protein. Aggregation of granule proteins, that are often cationic, would be assisted by the anionic serglycin proteoglycans present in neutrophils. The antibacterial granule proteins can serve as models for antibiotics and some of them possess a potentially useful therapeutic ability to bind and neutralize endotoxin. Because aberrant expression of transcription factors regulating the synthesis of granule proteins is often found in leukemia, the clarification of mechanisms regulating the timed expression of granule proteins will shed light on the maturation block in myeloid leukemias.
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| 23. |
- Melin, Malin, et al.
(författare)
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Assignment of the gene locus for severe congenital neutropenia to chromosome 1q22 in the original Kostmann family from Northern Sweden
- 2007
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 353:3, s. 571-575
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Tidskriftsartikel (refereegranskat)abstract
- Autosomal recessive severe congenital neutropenia (SCN) or Kostmann syndrome is characterised by reduced neutrophil counts and subsequent recurrent bacterial infections. The disease was originally described in a large consanguineous pedigree from Northern Sweden. A genome-wide autozygosity scan was initiated on samples from four individuals in the original pedigree using high density single nucleotide polymorphism (SNP) genotyping arrays in order to map the disease locus. Thirty candidate regions were identified and the ascertainment of samples from two additional patients confirmed a single haplotype with significant association to the disorder (p < 0.01) on chromosome 1q22. One affected individual from the original Kostmann pedigree was confirmed as a phenocopy. The minimal haplotype shared by affected individuals spans a candidate region of 1.2 Mb, containing several potential candidate genes.
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| 24. |
- Rögnvaldsson, Thorsteinn, et al.
(författare)
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Bioinformatic approaches for modeling the substrate specificity of HIV-1 protease : an overview
- 2007
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Ingår i: Expert Review of Molecular Diagnostics. - : Informa UK Limited. - 1473-7159 .- 1744-8352. ; 7:4, s. 435-451
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Tidskriftsartikel (refereegranskat)abstract
- HIV-1 protease has a broad and complex substrate specificity, which hitherto has escaped a simple comprehensive definition. This, and the relatively high mutation rate of the retroviral protease, makes it challenging to design effective protease inhibitors. Several attempts have been made during the last two decades to elucidate the enigmatic cleavage specificity of HIV-1 protease and to predict cleavage of novel substrates using bioinformatic analysis methods. This review describes the methods that have been utilized to date to address this important problem and the results achieved. The data sets used are also reviewed and important aspects of these are highlighted.
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| 25. |
- Rögnvaldsson, Thorsteinn, 1963-, et al.
(författare)
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How to find simple and accurate rules for viral protease cleavage specificities
- 2009
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Ingår i: BMC Bioinformatics. - London : BioMed Central Ltd.. - 1471-2105. ; 10, s. 149-156
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:Proteases of human pathogens are becoming increasingly important drug targets, hence it is necessary to understand their substrate specificity and to interpret this knowledge in practically useful ways. New methods are being developed that produce large amounts of cleavage information for individual proteases and some have been applied to extract cleavage rules from data. However, the hitherto proposed methods for extracting rules have been neither easy to understand nor very accurate. To be practically useful, cleavage rules should be accurate, compact, and expressed in an easily understandable way.RESULTS:A new method is presented for producing cleavage rules for viral proteases with seemingly complex cleavage profiles. The method is based on orthogonal search-based rule extraction (OSRE) combined with spectral clustering. It is demonstrated on substrate data sets for human immunodeficiency virus type 1 (HIV-1) protease and hepatitis C (HCV) NS3/4A protease, showing excellent prediction performance for both HIV-1 cleavage and HCV NS3/4A cleavage, agreeing with observed HCV genotype differences. New cleavage rules (consensus sequences) are suggested for HIV-1 and HCV NS3/4A cleavages. The practical usability of the method is also demonstrated by using it to predict the location of an internal cleavage site in the HCV NS3 protease and to correct the location of a previously reported internal cleavage site in the HCV NS3 protease. The method is fast to converge and yields accurate rules, on par with previous results for HIV-1 protease and better than previous state-of-the-art for HCV NS3/4A protease. Moreover, the rules are fewer and simpler than previously obtained with rule extraction methods.CONCLUSION: A rule extraction methodology by searching for multivariate low-order predicates yields results that significantly outperform existing rule bases on out-of-sample data, but are more transparent to expert users. The approach yields rules that are easy to use and useful for interpreting experimental data.
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| 26. |
- Rögnvaldsson, Thorsteinn, 1963-, et al.
(författare)
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State of the art prediction of HIV-1 protease cleavage sites
- 2015
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Ingår i: Bioinformatics. - Oxford : Oxford University Press. - 1367-4803 .- 1367-4811. ; 31:8, s. 1204-1210
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Tidskriftsartikel (refereegranskat)abstract
- Motivation: Understanding the substrate specificity of HIV-1 protease is important when designing effective HIV-1 protease inhibitors. Furthermore, characterizing and predicting the cleavage profile of HIV-1 protease is essential to generate and test hypotheses of how HIV-1 affects proteins of the human host. Currently available tools for predicting cleavage by HIV-1 protease can be improved.Results: The linear support vector machine with orthogonal encod-ing is shown to be the best predictor for HIV-1 protease cleavage. It is considerably better than current publicly available predictor ser-vices. It is also found that schemes using physicochemical proper-ties do not improve over the standard orthogonal encoding scheme. Some issues with the currently available data are discussed.Availability: The data sets used, which are the most important part, are available at the UCI Machine Learning Repository. The tools used are all standard and easily available. © 2014 The Author.
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| 28. |
- Venge, Per, et al.
(författare)
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HNL (Human Neutrophil Lipocalin) and a multimarker approach to the distinction between bacterial and viral infections
- 2019
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Ingår i: JIM - Journal of Immunological Methods. - : Elsevier. - 0022-1759 .- 1872-7905. ; 474
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVES: The distinction between bacterial and viral causes of acute infections is a major clinical challenge. In this report we investigate the diagnostic performance in this regard of nine candidate biomarkers together with HNL (Human Neutrophil Lipocalin).METHODS: Blood was obtained from patients with symptoms of infectious (n = 581). HNL was measured in whole blood (B-HNL) after pre-activation with the neutrophil activator fMLP or in plasma (P-HNL). Azurocidin also known as heparin-binding protein (HBP), Calprotectin, PMN-CD64, CRP (C-reactive protein), IP-10 (Interferon γ-induced Protein 10 kDa), PCT (Procalcitonin), TK1 (Thymidine kinase 1), TRAIL (TNF-related apoptosis-inducing ligand) were measured in plasma/serum. Area under the ROC (receiver operating characteristics) curve (AuROC) was used for the evaluation of the clinical performance of the biomarkers.RESULTS: Side-by-side comparisons of the ten biomarkers showed large difference in the AuROC with B-HNL being the superior biomarker (0.91, 95% CI 0.86-0.95) and with the other nine biomarkers varying from AuROC of 0.63-0.79. The combination of B-HNL with IP-10 and/or TRAIL increased the diagnostic performance further to AuROCs of 0.94-0.97. The AuROCs of the combination of CRP with IP-10 and/or TRAIL were significantly lower than combinations with B-HNL 0.87 (95% CI 0.83-0.91).CONCLUSION: The diagnostic performance of whole blood activated HNL was superior in the distinction between bacterial or viral infections. The addition of IP-10 and/or TRAIL to the diagnostic algorithm increased the performance of B-HNL further. The rapid analysis of HNL, reflecting bacterial infections, together with biomarkers reflecting viral infections may be the ideal combination of diagnostic biomarkers of acute infections.
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| 29. |
- Venge, Per, et al.
(författare)
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Human Neutrophil Lipocalin as a Superior Diagnostic Means To Distinguish between Acute Bacterial and Viral Infections
- 2015
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Ingår i: Clinical and Vaccine Immunology. - 1556-6811 .- 1556-679X. ; 22:9, s. 1025-1032
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Tidskriftsartikel (refereegranskat)abstract
- The distinction between causes of acute infections is a major clinical challenge. Current biomarkers, however, are not sufficiently accurate. Human neutrophil lipocalin (HNL) concentrations in serum or whole blood activated by formyl-methionine-leucine-phenylalanine (fMLP) were shown to distinguish acute infections of bacterial or viral cause with high accuracy. The aim was therefore to compare the clinical performance of HNL with currently used biomarkers. Seven hundred twenty-five subjects (144 healthy controls and 581 patients with signs and symptoms of acute infections) were included in the study. C-reactive protein (CRP), the expression of CD64 on neutrophils, procalcitonin (PCT), and blood neutrophil counts were measured by established techniques, and HNL concentrations were measured in whole-blood samples after activation with fMLP. All tested biomarkers were elevated in bacterial as opposed to viral infections (P<0.001). CRP, PCT, and CD64 expression in neutrophils was elevated in viral infections compared to healthy controls (P<0.001). In the distinction between healthy controls and patients with bacterial infections, the areas under the receiver operating characteristic (ROC) curves were >0.85 for all biomarkers, whereas for the distinction between bacterial and viral infections, only HNL concentration in fMLP-activated whole blood showed an area under the ROC curve (AUROC) of >0.90 and superior clinical performance. The clinical performance of HNL in fMLP-activated whole blood was superior to current biomarkers and similar to previous results of HNL in serum. The procedure can be adopted for point-of-care testing with response times of <15 min.
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| 30. |
- Venge, Per, et al.
(författare)
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Human neutrophil lipocalin in fMLP-activated whole blood as a diagnostic means to distinguish between acute bacterial and viral infections
- 2015
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Ingår i: JIM - Journal of Immunological Methods. - : Elsevier BV. - 0022-1759 .- 1872-7905. ; 424, s. 85-90
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Tidskriftsartikel (refereegranskat)abstract
- The distinction between causes of acute infections is a major clinical challenge. Current biomarkers, however, are not sufficiently accurate. Human neutrophil lipocalin (HNL) in serum distinguishes acute infections with high accuracy, but in the emergency setting the assay time should be <15-20 min, which excludes the use of serum samples. The aim was therefore to develop a novel rapid assay principle and test its clinical performance. Methods: Serum and neutrophils obtained from 84 infected and 20 healthy subjects were used in the experimental study. 725 subjects (144 healthy controls and 581 patients with signs and symptoms of acute infections) were included in the clinical study. HNL was measured in EDTA-plasma by ELISA or in heparinized whole blood after fMLP activation by a prototype point-of-care assay. Results: Increased release of HNL from neutrophils after activation with fMLP was seen already after 5 min incubation. The release of HNL from purified neutrophils after 15 min incubation with fMLP was significantly correlated to the HNL concentrations in serum obtained from the same patient (r = 0.74, p < 0.001). In the distinction between healthy controls and patients with bacterial infections, the areas under the ROC-curves were 0.95 (95% CI 0.91-0.97) and 0.88 (95% CI 0.84-0.91) for HNL in fMLP-activated whole blood and EDTA-plasma, respectively, (p <0.001) and in the distinction between bacterial and viral infections 0.91 (95% CI 0.86-0.95) and 0.76 (95% CI 0.70-0.81), respectively (p <0.001). Conclusion: The clinical performance of HNL in fMLP-activated whole blood was superior to HNL in EDTA-plasma and similar to HNL in serum. The procedure can be adopted for point-of-care testing with response times of <15 min.
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| 31. |
- You, Liwen, et al.
(författare)
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Comprehensive bioinformatic analysis of the specificity of human immunodeficiency virus type 1 protease.
- 2005
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Ingår i: J Virol. - Washington, DC : The American Society for Microbiology. - 0022-538X .- 1098-5514. ; 79:19, s. 12477-86
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Tidskriftsartikel (refereegranskat)abstract
- Rapidly developing viral resistance to licensed human immunodeficiency virus type 1 (HIV-1) protease inhibitors is an increasing problem in the treatment of HIV-infected individuals and AIDS patients. A rational design of more effective protease inhibitors and discovery of potential biological substrates for the HIV-1 protease require accurate models for protease cleavage specificity. In this study, several popular bioinformatic machine learning methods, including support vector machines and artificial neural networks, were used to analyze the specificity of the HIV-1 protease. A new, extensive data set (746 peptides that have been experimentally tested for cleavage by the HIV-1 protease) was compiled, and the data were used to construct different classifiers that predicted whether the protease would cleave a given peptide substrate or not. The best predictor was a nonlinear predictor using two physicochemical parameters (hydrophobicity, or alternatively polarity, and size) for the amino acids, indicating that these properties are the key features recognized by the HIV-1 protease. The present in silico study provides new and important insights into the workings of the HIV-1 protease at the molecular level, supporting the recent hypothesis that the protease primarily recognizes a conformation rather than a specific amino acid sequence. Furthermore, we demonstrate that the presence of 1 to 2 lysine residues near the cleavage site of octameric peptide substrates seems to prevent cleavage efficiently, suggesting that this positively charged amino acid plays an important role in hindering the activity of the HIV-1 protease.
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