Publications by authors named "Berit Sletbakk Brusletto"

10 Publications

  • Page 1 of 1

Uptake of circulating extracellular vesicles from rectal cancer patients and differential responses by human monocyte cultures.

FEBS Open Bio 2021 Mar 8;11(3):724-740. Epub 2021 Feb 8.

Department of Oncology, Akershus University Hospital, Lørenskog, Norway.

Extracellular vesicles (EVs) released by tumor cells can directly or indirectly modulate the phenotype and function of the immune cells of the microenvironment locally or at distant sites. The uptake of circulating EVs and the responses by human monocytes in vitro may provide new insights into the underlying biology of the invasive and metastatic processes in cancer. Although a mixed population of vesicles is obtained with most isolation techniques, we predominantly isolated exosomes (small EVs) and microvesicles (medium EVs) from the SW480 colorectal cancer cell line (established from a primary adenocarcinoma of the colon) by sequential centrifugation and ultrafiltration, and plasma EVs were prepared from 22 patients with rectal adenoma polyps or invasive adenocarcinoma by size-exclusion chromatography. The EVs were thoroughly characterized. The uptake of SW480 EVs was analyzed, and small SW480 EVs were observed to be more potent than medium SW480 EVs in inducing monocyte secretion of cytokines. The plasma EVs were also internalized by monocytes; however, their cytokine-releasing potency was lower than that of the cell line-derived vesicles. The transcriptional changes in the monocytes highlighted differences between adenoma and adenocarcinoma patient EVs in their ability to regulate biological functions, whereas the most intriguing changes were found in monocytes receiving EVs from patients with metastatic compared with localized cancer.
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http://dx.doi.org/10.1002/2211-5463.13098DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7931235PMC
March 2021

Extensive Changes in Transcriptomic "Fingerprints" and Immunological Cells in the Large Organs of Patients Dying of Acute Septic Shock and Multiple Organ Failure Caused by .

Front Cell Infect Microbiol 2020 19;10:42. Epub 2020 Feb 19.

Department of Medical Biochemistry, Oslo University Hospital, Oslo, Norway.

Patients developing meningococcal septic shock reveal levels of (10-10/mL) and endotoxin (10-10 EU/mL) in the circulation and organs, leading to acute cardiovascular, pulmonary and renal failure, coagulopathy and a high case fatality rate within 24 h. To investigate transcriptional profiles in heart, lungs, kidneys, liver, and spleen and immunostain key inflammatory cells and proteins in post mortem formalin-fixed, paraffin-embedded (FFPE) tissue samples from meningococcal septic shock patients. Total RNA was isolated from FFPE and fresh frozen (FF) tissue samples from five patients and two controls (acute non-infectious death). Differential expression of genes was detected using Affymetrix microarray analysis. Lung and heart tissue samples were immunostained for T-and B cells, macrophages, neutrophils and the inflammatory markers PAI-1 and MCP-1. Inflammatory mediators were quantified in lysates from FF tissues. The transcriptional profiles showed a complex pattern of protein-coding and non-coding RNAs with significant regulation of pathways associated with organismal death, cell death and survival, leukocyte migration, cellular movement, proliferation of cells, cell-to-cell signaling, immune cell trafficking, and inflammatory responses in an organ-specific clustering manner. The canonical pathways including acute phase response-, EIF2-, TREM1-, IL-6-, HMBG1-, PPAR signaling, and LXR/RXR activation were associated with acute heart, pulmonary, and renal failure. Fewer genes were regulated in the liver and particularly in the spleen. The main upstream regulators were TNF, IL-1β, IL-6, RICTOR, miR-6739-3p, and CD3. Increased numbers of inflammatory cells (CD68+, MPO+, CD3+, and CD20+) were found in lungs and heart. PAI-1 inhibiting fibrinolysis and MCP-1 attracting leukocyte were found significantly present in the septic tissue samples compared to the controls. FFPE tissue samples can be suitable for gene expression studies as well as immunostaining of specific cells or molecules. The most pronounced gene expression patterns were found in the organs with highest levels of DNA. Thousands of protein-coding and non-coding RNA transcripts were altered in lungs, heart and kidneys. We identified specific biomarker panels both protein-coding and non-coding RNA transcripts, which differed from organ to organ. Involvement of many genes and pathways add up and the combined effect induce organ failure.
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http://dx.doi.org/10.3389/fcimb.2020.00042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7045056PMC
June 2021

Urine -2-Microglobulin, Osteopontin, and Trefoil Factor 3 May Early Predict Acute Kidney Injury and Outcome after Cardiac Arrest.

Crit Care Res Pract 2019 7;2019:4384796. Epub 2019 May 7.

Institute of Clinical Medicine, University of Oslo, P.O.Box 1072 Blindern, 0316 Oslo, Norway.

Purpose: Acute kidney injury (AKI) is a common complication after out-of-hospital cardiac arrest (OHCA), leading to increased mortality and challenging prognostication. Our aim was to examine if urine biomarkers could early predict postarrest AKI and patient outcome.

Methods: A prospective observational study of resuscitated, comatose OHCA patients admitted to Oslo University Hospital in Norway. Urine samples were collected at admission and day three postarrest and analysed for -2-microglobulin (2M), osteopontin, and trefoil factor 3 (TFF3). Outcome variables were AKI within three days according to the Kidney Disease Improving Global Outcome criteria, in addition to six-month mortality and poor neurological outcome (PNO) (cerebral performance category 3-5).

Results: Among 195 included patients (85% males, mean age 60 years), 88 (45%) developed AKI, 88 (45%) died, and 96 (49%) had PNO. In univariate analyses, increased urine 2M, osteopontin, and TFF3 levels sampled at admission and day three were independent risk factors for AKI, mortality, and PNO. Exceptions were that 2M measured at day three did not predict any of the outcomes, and TFF3 at admission did not predict AKI. In multivariate analyses, combining clinical parameters and biomarker levels, the area under the receiver operating characteristics curves (95% CI) were 0.729 (0.658-0.800), 0.797 (0.733-0.861), and 0.812 (CI 0.750-0.874) for AKI, mortality, and PNO, respectively.

Conclusions: Urine levels of 2M, osteopontin, and TFF3 at admission and day three were associated with increased risk for AKI, mortality, and PNO in comatose OHCA patients. This trail is registered with NCT01239420.
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http://dx.doi.org/10.1155/2019/4384796DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6530154PMC
May 2019

Molecular studies of meningococcal and pneumococcal meningitis patients in Ethiopia.

Innate Immun 2019 04;25(3):158-167

3 Department of Clinical Chemistry, Oslo University Hospital, Norway.

Neisseria meningitidis infections in sub-Saharan Africa usually present with distinct symptoms of meningitis but very rarely as fulminant septicemia when reaching hospitals. In Europe, development of persistent meningococcal shock and multiple organ failure occurs in up to 30% of patients and is associated with a bacterial load of >10/ml plasma or serum. We have prospectively studied 27 Ethiopian patients with meningococcal infection as diagnosed and quantified with real-time PCR in the cerebrospinal fluid (CSF) and serum. All presented with symptoms of meningitis and none with fulminant septicemia. The median N. meningitidis copy number (NmDNA) in serum was < 3.5 × 10/ml, never exceeded 1.8 × 10/ml, and was always 10-1000 times higher in CSF than in serum. The levels of LPS in CSF as determined by the limulus amebocyte lysate assay were positively correlated to NmDNA copy number ( r = 0.45, P = 0.030), levels of IL-1 receptor antagonist, ( r = 0.46, P = 0.017), and matrix metallopeptidase-9 (MMP-9; r = 0.009). We also compared the inflammatory profiles of 19 mediators in CSF of the 26 meningococcal patients (2 died and 2 had immediate severe sequelae) with 16 patients with Streptococcus pneumoniae meningitis (3 died and 3 with immediate severe sequelae). Of 19 inflammatory mediators tested, 9 were significantly higher in patients with pneumococcal meningitis and possibly linked to worse outcome.
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http://dx.doi.org/10.1177/1753425918806363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6830936PMC
April 2019

Transcriptomic data from two primary cell models stimulating human monocytes suggest inhibition of oxidative phosphorylation and mitochondrial function by N. meningitidis which is partially up-regulated by IL-10.

BMC Immunol 2017 10 27;18(1):46. Epub 2017 Oct 27.

Blood Cell Research Group, Section for Research, Department of Medical Biochemistry, Oslo University Hospital, Oslo, Norway.

Background: Biological interpretation of DNA microarray data may differ depending on underlying assumptions and statistical tests of bioinformatics tools used. We used Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA) to analyze previously generated DNA microarray data from human monocytes stimulated with N. meningitidis and IL-10 ("the model system"), and with meningococcal sepsis plasma before and after immunodepletion of IL-10 ("the patient plasma system"). The objectives were to compare if the two bioinformatics methods resulted in similar biological interpretation of the datasets, and to identify whether GSEA provided additional insight compared with IPA about the monocyte host response to meningococcal activation.

Results: In both experimental models, GSEA and IPA identified genes associated with pro-inflammatory innate immune activation, including TNF-signaling, Toll-like receptor signaling, JAK-STAT-signaling, and type I and type II interferon signaling. GSEA identified genes regulated by the presence of IL-10 with similar gene sets in both the model system and the patient plasma system. In the model system, GSEA and IPA in sum identified 170 genes associated with oxidative phosphorylation/mitochondrial function to be down-regulated in monocytes stimulated with meningococci. In the patient plasma system, GSEA and IPA in sum identified 122 genes associated with oxidative phosphorylation/mitochondrial dysfunction to be down-regulated by meningococcal sepsis plasma depleted for IL-10. Using IPA, we identified IL-10 to up-regulate 18 genes associated with oxidative phosphorylation/mitochondrial function that were down-regulated by N. meningitidis.

Conclusions: Biological processes associated with the gene expression changes in the model system of meningococcal sepsis were comparable with the results found in the patient plasma system. By combining GSEA with IPA, we discovered an inhibitory effect of N. meningitidis on genes associated with mitochondrial function and oxidative phosphorylation, and that IL-10 partially reverses this strong inhibitory effect, thereby identifying, to our knowledge, yet another group of genes where IL-10 regulates the effect of LPS. We suggest that relying on a single bioinformatics tool together with an arbitrarily chosen filtering criteria for data analysis may result in overlooking relevant biological processes and signaling pathways associated with genes differentially expressed between compared experimental conditions.
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http://dx.doi.org/10.1186/s12865-017-0229-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5659018PMC
October 2017

Traceability and distribution of DNA in archived post mortem tissue samples from patients with systemic meningococcal disease.

BMC Clin Pathol 2017 16;17:10. Epub 2017 Aug 16.

Blood Cell Research Group, Section for Research, Department of Medical Biochemistry, Oslo University Hospital HF, Ullevål Hospital, PO Box 4956 Nydalen, 0424 Oslo, Norway.

Background: The pathophysiology and outcome of meningococcal septic shock is closely associated with the plasma level of lipopolysaccharides (LPS, endotoxin) and the circulating level of meningococcal DNA. The aim of the present study was to quantify the number of in different formalin-fixed, paraffin-embedded (FFPE) tissue samples and fresh frozen (FF) tissue samples from patients with systemic meningococcal disease (SMD), to explore the distribution of in the body.

Methods: DNA in FFPE and FF tissue samples from heart, lungs, liver, kidneys, spleen and brain from patients with meningococcal shock and controls (lethal pneumococcal infection) stored at variable times, were isolated. The bacterial load of DNA was analyzed using quantitative real-time PCR (qPCR) and primers for the capsule transport A (ctrA) gene (1 copy per DNA). The human beta-hemoglobin (HBB) gene was quantified to evaluate effect of the storage times (2-28 years) and storage method in archived tissue.

Results: DNA was detected in FFPE and FF tissue samples from heart, lung, liver, kidney, and spleen in all patients with severe shock. In FFPE brain, DNA was only detected in the patient with the highest concentration of LPS in the blood at admission to hospital. The highest levels of DNA were found in heart tissue (median value 3.6 × 10 copies DNA/μg human DNA) and lung tissue (median value 3.1 × 10 copies DNA/μg human DNA) in all five patients. DNA was not detectable in any of the tissue samples from two patients with clinical meningitis and the controls (pneumococcal infection). The quantity of HBB declined over time in FFPE tissue stored at room temperature, suggesting degradation of DNA.

Conclusions: High levels of DNA were detected in the different tissue samples from meningococcal shock patients, particularly in the heart and lungs suggesting seeding and major proliferation of meningococci in these organs during the development of shock, probably contributing to the multiple organ failure. The age of archived tissue samples appear to have an impact on the amount of quantifiable DNA.
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http://dx.doi.org/10.1186/s12907-017-0049-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559868PMC
August 2017

Urine biomarkers give early prediction of acute kidney injury and outcome after out-of-hospital cardiac arrest.

Crit Care 2016 10 5;20(1):314. Epub 2016 Oct 5.

Institute of Clinical Medicine, University of Oslo, Oslo, Norway.

Background: Post-resuscitation care after out-of-hospital cardiac arrest (OHCA) is challenging due to the threat of organ failure and difficult prognostication. Our aim was to examine whether urine biomarkers could give an early prediction of acute kidney injury (AKI) and outcome.

Methods: This was a prospective observational study of comatose OHCA patients at Oslo University Hospital Ullevål, Norway. Risk factors were clinical parameters and biomarkers measured in spot urine (cystatin C, neutrophil gelatinase-associated lipocalin (NGAL) and the product of tissue inhibitor of metalloproteinase 2 (TIMP-2) and insulin-like growth factor-binding protein 7 (IGFBP7)) at admission and day 3. Outcome variables were AKI within 3 days using the Kidney Disease Improving Global Outcomes definition, 6-month mortality, and poor neurological outcome (PNO) defined as cerebral performance category 3-5.

Results: Among 195 included patients (85 % males, mean age 60 years), 88 (45 %) died, 96 (49 %) had PNO, and 88 (45 %) developed AKI. In univariate analysis, increased urine cystatin C and NGAL concentration sampled at admission and day 3 were independent risk factors for AKI, mortality and PNO. Increased urine TIMP-2 × IGFBP7 levels was associated with AKI only at admission. In multivariate analyses combining clinical parameters and biomarker concentrations, the area under the receiver operating characteristics curve (AuROC) with 95 % confidence interval (CI) were 0.774 (0.700-0.848), 0.812 (0.751-0.873), and 0.819 (0.759-0.878) for AKI, mortality and PNO, respectively.

Conclusions: In comatose OHCA patients, urine levels of cystatin C and NGAL at admission and day 3 were independent risk factors for AKI, 6-month mortality and PNO.

Trial Registration: Clinicaltrials.gov NCT01239420 . Registered 10 November 2010.
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http://dx.doi.org/10.1186/s13054-016-1503-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5052716PMC
October 2016

IL-10 immunodepletion from meningococcal sepsis plasma induces extensive changes in gene expression and cytokine release in stimulated human monocytes.

Innate Immun 2015 May 18;21(4):429-49. Epub 2014 Sep 18.

Blood Cell Research Group, Section for Research, Department of Medical Biochemistry, Oslo University Hospital, University of Oslo, Oslo, Norway.

The severity of systemic meningococcal disease (SMD) correlates to plasma concentrations of LPS and IL-10, with the highest levels detected in non-survivors. Here, plasma from patients with SMD containing high and low concentrations of LPS were incubated with human monocytes before and after immunodepletion of IL-10 to study the effect of IL-10 on gene expression and cytokine release. Patient plasma containing IL-10 induced the expression of 1657 genes in human monocytes when compared with gene expression induced by low LPS plasma. After immunodepletion of IL-10, this number increased to 2260. By directly comparing the gene expression profiles induced before and after immunodepletion of IL-10, the presence of IL-10 differentially regulated 373 genes. Functional classes associated with these genes were cellular function and maintenance, cellular development, cellular growth and proliferation, cell-cell signaling and interaction and cellular movement. Immunodepletion of IL-10 resulted in down-regulation of genes of the leukocyte immunoglobulin-like receptor family, and up-regulation of genes of type I IFN signaling, TLR signaling, the inflammasomes, coagulation and fibrinolysis. Finally, immunodepletion of IL-10 increased the protein levels of IL-1β, IL-8, TNF-α, MIP-1α and MIP-1β. Data suggest that IL-10 in meningococcal sepsis plasma regulates a variety of genes and signaling pathways, likely leading to an overall inhibitory effect on the inflammatory response induced in meningococcal sepsis.
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http://dx.doi.org/10.1177/1753425914547743DOI Listing
May 2015

Transcriptional profiling of mRNAs and microRNAs in human bone marrow precursor B cells identifies subset- and age-specific variations.

PLoS One 2013 30;8(7):e70721. Epub 2013 Jul 30.

Department of Medical Biochemistry, Oslo University Hospital, Oslo, Norway.

Background: Molecular mechanisms explaining age-related changes in the bone marrow with reduced precursor B cell output are poorly understood.

Methods: We studied the transcriptome of five precursor B cell subsets in individual bone marrow samples from 4 healthy children and 4 adults employing GeneChip® Human Exon 1.0 ST Arrays (Affymetrix®) and TaqMan® Array MicroRNA Cards (Life Technologies™).

Results: A total of 1796 mRNAs (11%) were at least once differentially expressed between the various precursor B cell subsets in either age group (FDR 0.1%, p≤1.13×10(-4)) with more marked cell stage specific differences than those related to age. In contrast, microRNA profiles of the various precursor B cell subsets showed less hierarchical clustering as compared to the corresponding mRNA profiles. However, 17 of the 667 microRNA assays (2.5%) were at least once differentially expressed between the subsets (FDR 10%, p≤0.004). From target analysis (Ingenuity® Systems), functional assignment between postulated interacting mRNAs and microRNAs showed especially association to cellular growth, proliferation and cell cycle regulation. One functional network connected up-regulation of the differentiation inhibitor ID2 mRNA to down-regulation of the hematopoiesis- or cell cycle regulating miR-125b-5p, miR-181a-5p, miR-196a-5p, miR-24-3p and miR-320d in adult PreBII large cells. Noteworthy was also the stage-dependent expression of the growth promoting miR-17-92 cluster, showing a partly inverse trend with age, reaching statistical significance at the PreBII small stage (up 3.1-12.9 fold in children, p = 0.0084-0.0270).

Conclusions: The global mRNA profile is characteristic for each precursor B cell developmental stage and largely similar in children and adults. The microRNA profile is much cell stage specific and not changing much with age. Importantly, however, specific age-dependent differences involving key networks like differentiation and cellular growth may indicate biological divergence and possibly also altered production potential with age.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0070721PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3728296PMC
April 2014

Increased ID2 levels in adult precursor B cells as compared with children is associated with impaired Ig locus contraction and decreased bone marrow output.

J Immunol 2013 Aug 3;191(3):1210-9. Epub 2013 Jul 3.

Department of Medical Biochemistry, Oslo University Hospital, 0407 Oslo, Norway.

Precursor B cell production from bone marrow in mice and humans declines with age. Because the mechanisms behind are still unknown, we studied five precursor B cell subsets (ProB, PreBI, PreBII large, PreBII small, immature B) and their differentiation-stage characteristic gene expression profiles in healthy individual toddlers and middle-aged adults. Notably, the composition of the precursor B cell compartment did not change with age. The expression levels of several transcripts encoding V(D)J recombination factors were decreased in adults as compared with children: RAG1 expression was significantly reduced in ProB cells, and DNA-PKcs, Ku80, and XRCC4 were decreased in PreBI cells. In contrast, TdT was 3-fold upregulated in immature B cells of adults. Still, N-nucleotides, P-nucleotides, and deletions were similar for IGH and IGK junctions between children and adults. PreBII large cells in adults, but not in children, showed highly upregulated expression of the differentiation inhibitor, inhibitor of DNA binding 2 (ID2), in absence of changes in expression of the ID2-binding partner E2A. Further, we identified impaired Ig locus contraction in adult precursor B cells as a likely mechanism by which ID2-mediated blocking of E2A function results in reduced bone marrow B cell output in adults. The reduced B cell production was not compensated by increased proliferation in adult immature B cells, despite increased Ki67 expression. These findings demonstrate distinct regulatory mechanisms in B cell differentiation between adults and children with a central role for transcriptional regulation of ID2.
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http://dx.doi.org/10.4049/jimmunol.1203462DOI Listing
August 2013