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1.
The bivariate NRIP1/ZEB2 RNA marker permits non-invasive presymptomatic screening of pre-eclampsia.
Manders, V, Visser, A, Keijser, R, Min, N, Poutsma, A, Mulders, J, van den Berkmortel, T, Hortensius, M, Jongejan, A, Pajkrt, E, et al
Scientific reports. 2020;(1):21857
Abstract
Using genome-wide transcriptome analysis by RNA sequencing of first trimester plasma RNA, we tested whether the identification of pregnancies at risk of developing pre-eclampsia with or without preterm birth or growth restriction is possible between weeks 9-14, prior to the appearance of clinical symptoms. We implemented a metaheuristic approach in the self-learning SVM algorithm for differential gene expression analysis of normal pregnancies (n = 108), affected pregnancies (n = 34) and non-pregnant controls (n = 19). Presymptomatic candidate markers for affected pregnancies were validated by RT-qPCR in first trimester samples (n = 34) from an independent cohort. PRKG1 was significantly downregulated in a subset of pregnancies with birth weights below the 10thpercentile as shared symptom. The NRIP1/ZEB2 ratio was found to be upregulated in pregnancies with pre-eclampsia or trisomy 21. Complementary quantitative analysis of both the linear and circular forms of NRIP1 permitted discrimination between pre-eclampsia and trisomy 21. Pre-eclamptic pregnancies showed an increase in linear NRIP1 compared to circular NRIP1, while trisomy 21 pregnancies did not.
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Clinical relevance of circulating MACC1 and S100A4 transcripts for ovarian cancer.
Link, T, Kuhlmann, JD, Kobelt, D, Herrmann, P, Vassileva, YD, Kramer, M, Frank, K, Göckenjan, M, Wimberger, P, Stein, U
Molecular oncology. 2019;(5):1268-1279
Abstract
Metastasis-associated in colon cancer 1 (MACC1) and S100 calcium-binding protein A4 (S100A4) are prominent inducers of tumor progression and metastasis. For the first time, we systematically tracked circulating serum levels of MACC1 and S100A4 transcripts in the course of surgery and chemotherapy and analyzed their clinical relevance for ovarian cancer. MACC1 and S100A4 transcripts were quantified in a total of 318 serum samples from 79 ovarian cancer patients by RT-qPCR and digital droplet PCR, respectively. MACC1 and S100A4 transcripts were significantly elevated in serum of ovarian cancer patients, compared to healthy controls (P = 0.024; P < 0.001). At primary diagnosis, high levels of MACC1 or S100A4 correlated with advanced FIGO stage (P = 0.042; P = 0.008), predicted suboptimal debulking surgery and indicated shorter progression-free survival (PFS; P = 0.003; P = 0.001) and overall survival (OS; P = 0.001; P = 0.002). This is the first study in ovarian cancer to propose circulating MACC1 and S100A4 transcripts as potential liquid biopsy markers.
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Soluble Leptin Receptor Predicts Insulin Sensitivity and Correlates With Upregulation of Metabolic Pathways in Men.
Sommer, C, Lee, S, Gulseth, HL, Jensen, J, Drevon, CA, Birkeland, KI
The Journal of clinical endocrinology and metabolism. 2018;(3):1024-1032
Abstract
CONTEXT Plasma soluble leptin receptor (sOb-R) seems protective of gestational and type 2 diabetes in observational studies, but the mechanisms are unknown. sOb-R is formed by ectodomain shedding of membrane-bound leptin receptors (Ob-Rs), but its associations with messenger RNA (mRNA) expression are scarcely explored. OBJECTIVE To explore associations between plasma levels of sOb-R and (1) insulin sensitivity, (2) mRNA pathways in adipose tissue and skeletal muscle, and (3) mRNA of candidate genes for sOb-R generation in adipose tissue and skeletal muscle. DESIGN AND PARTICIPANTS The MyoGlu study included 26 sedentary, middle-aged men who underwent a 12-week intensive exercise intervention. We measured plasma sOb-R with enzyme-linked immunosorbent assay, insulin sensitivity with a hyperinsulinemic euglycemic clamp, and mRNA in skeletal muscle and adipose tissue with high-throughput sequencing. RESULTS Baseline plasma sOb-R was strongly associated with baseline glucose infusion rate (GIR) [β (95% confidence interval), 1.19 (0.57 to 1.82) mg/kg/min, P = 0.0006] and GIR improvement after the exercise intervention [0.58 (0.03 to 1.12) mg/kg/min, P = 0.039], also independently of covariates, including plasma leptin. In pathway analyses, high plasma sOb-R correlated with upregulation of metabolic pathways and downregulation of inflammatory pathways in both adipose tissue and skeletal muscle. In skeletal muscle, mRNA of LEPROT and LEPROTL1 (involved in Ob-R cell surface expression) and ADAM10 and ADAM17 (involved sOb-R-shedding) increased after the exercise intervention. CONCLUSIONS Higher plasma sOb-R was associated with improved GIR, upregulation of metabolic pathways, and downregulation of inflammatory pathways, which may be possible mechanisms for the seemingly protective effect of plasma sOb-R on subsequent risk of gestational and type 2 diabetes found in observational studies.
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Mitochondrial ferritin expression in erythroid cells from patients with alpha-thalassaemia.
Putburee, R, Jetsrisuparb, A, Fucharoen, S, Tripatara, A
Hematology (Amsterdam, Netherlands). 2018;(10):844-848
Abstract
BACKGROUND Patients with thalassaemia who received regular transfusions had increased iron accumulation, leading to iron overload, which was associated with oxidative stress. Mitochondrial ferritin, encoded by the FTMT gene is an iron-storage protein in the mitochondria. The aim of this work was to investigate the expression levels of FTMT in the reticulocytes of patients with alpha-thalassaemia who were regularly transfused and rarely transfused compared with healthy controls and to evaluate the relationships of the levels of FTMT mRNA with malondialdehyde (MDA) and ferritin in these patients. METHODS The levels of FTMT mRNA in the reticulocytes of patients (30 regularly transfused and 30 rarely transfused) and 30 healthy individuals were assessed by quantitative reverse transcription-polymerase chain reaction. The levels of ferritin and MDA were analysed by ELISA and by a thiobarbituric acid reactive substance assay, respectively. RESULTS The levels of FTMT mRNA, ferritin and MDA in both groups of patients were significantly increased compared with those in the healthy controls. In addition, the levels of FTMT mRNA, ferritin and MDA in the regularly transfused patients were significantly higher than those in the rarely transfused patients. Furthermore, the relative expression levels of FTMT in patients correlated with those of MDA and ferritin. CONCLUSION These results suggest that the elevation of expression levels of FTMT in the reticulocytes of patients with alpha-thalassaemia may be associated with iron loading and oxidative stress.
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Determinants of Visfatin/NAMPT Serum Concentration and its Leukocyte Expression in Hyperthyroidism.
Sawicka-Gutaj, N, Zybek-Kocik, A, Kloska, M, Czarnywojtek, A, Sowiński, J, Budny, B, Woliński, K, Ziemnicka, K, Mańkowska-Wierzbicka, D, Ruchała, M
Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et metabolisme. 2018;(9):653-660
Abstract
We aimed to analyze the potential influence of thyroid autoimmunity on visfatin/NAMPT serum concentration and its leukocyte expression in hyperthyroid patients. This is a single-center, cross-sectional study with consecutive enrollment. All patients with newly diagnosed overt hyperthyroidism in a course of Graves' disease or toxic nodular goiter were included in the study. They underwent physical examination, laboratory investigation, body composition analysis, and thyroid ultrasound. NAMPT mRNA leukocyte expressions were measured using RT-qPCR. Of the 173 patients, 95 were enrolled in further analysis [67 patients with Graves' disease (GD) and 28 with toxic nodular goiter (TNG)]. Control group consisted of 43 healthy volunteers adjusted for age, sex, and BMI. Higher NAMPT/visfatin serum concentration was found in patients with GD comparing with patients with TNG (p=0.03855). We found significant NAMPT leukocyte overexpression in GD patients (n=32) as compared to TNG patients (n=18) and euthyroid controls (n=24) (p=0.005965). Simple linear regression analysis revealed that NAMPT/visfatin serum concentration was significantly associated with NAMPT leukocyte expression, thyroid autoimmunity, age, HOMA-IR, and fat mass percentage (FM%). NAMPT leukocyte expression was related to thyroid autoimmunity, age, and TRAb levels. The stepwise multiple regression analysis revealed FM% and HOMA-IR as independent predictors of visfatin/NAMPT serum levels. In a separate stepwise multiple regression analysis, we confirmed the association between NAMPT leukocyte expression and TRAb levels. We found that fat mass percentage together with HOMA-IR are the most significant predictors of visfatin/NAMPT serum elevation in hyperthyroid patients.
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Effect of energy restriction and physical exercise intervention on phenotypic flexibility as examined by transcriptomics analyses of mRNA from adipose tissue and whole body magnetic resonance imaging.
Lee, S, Norheim, F, Langleite, TM, Noreng, HJ, Storås, TH, Afman, LA, Frost, G, Bell, JD, Thomas, EL, Kolnes, KJ, et al
Physiological reports. 2016;(21)
Abstract
Overweight and obesity lead to changes in adipose tissue such as inflammation and reduced insulin sensitivity. The aim of this study was to assess how altered energy balance by reduced food intake or enhanced physical activity affect these processes. We studied sedentary subjects with overweight/obesity in two intervention studies, each lasting 12 weeks affecting energy balance either by energy restriction (~20% reduced intake of energy from food) in one group, or by enhanced energy expenditure due to physical exercise (combined endurance- and strength-training) in the other group. We monitored mRNA expression by microarray and mRNA sequencing from adipose tissue biopsies. We also measured several plasma parameters as well as fat distribution with magnetic resonance imaging and spectroscopy. Comparison of microarray and mRNA sequencing showed strong correlations, which were also confirmed using RT-PCR In the energy restricted subjects (body weight reduced by 5% during a 12 weeks intervention), there were clear signs of enhanced lipolysis as monitored by mRNA in adipose tissue as well as plasma concentration of free-fatty acids. This increase was strongly related to increased expression of markers for M1-like macrophages in adipose tissue. In the exercising subjects (glucose infusion rate increased by 29% during a 12-week intervention), there was a marked reduction in the expression of markers of M2-like macrophages and T cells, suggesting that physical exercise was especially important for reducing inflammation in adipose tissue with insignificant reduction in total body weight. Our data indicate that energy restriction and physical exercise affect energy-related pathways as well as inflammatory processes in different ways, probably related to macrophages in adipose tissue.
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Accuracy of digital mRNA profiling of oesophageal biopsies as a novel diagnostic approach to eosinophilic oesophagitis.
Lexmond, WS, Hu, L, Pardo, M, Heinz, N, Rooney, K, LaRosa, J, Dehlink, E, Fiebiger, E, Nurko, S
Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology. 2015;(8):1317-1327
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Abstract
BACKGROUND Quantification of tissue eosinophils remains the golden standard in diagnosing eosinophilic oesophagitis (EoE), but this approach suffers from poor specificity. It has been recognized that histopathological changes that occur in patients with EoE are associated with a disease-specific tissue transcriptome. OBJECTIVE We hypothesized that digital mRNA profiling targeted at a set of EoE-specific and Th2 inflammatory genes in oesophageal biopsies could help differentiate patients with EoE from those with reflux oesophagitis (RE) or normal tissue histology (NH). METHODS The mRNA expression levels of 79 target genes were defined in both proximal and distal biopsies of 196 patients with nCounter® (Nanostring) technology. According to clinicopathological diagnosis, these patients were grouped in a training set (35 EoE, 30 RE, 30 NH) for building of a three-class prediction model using the random forest method, and a blinded predictive set (n = 47) for model validation. RESULTS A diagnostic model built on ten differentially expressed genes was able to differentiate with 100% sensitivity and specificity between conditions in the training set. In a blinded predictive set, this model was able to correctly predict EoE in 14 of 18 patients in distal (sensitivity 78%, 95% CI 52-93%) and 16 of 18 patients in proximal biopsies (sensitivity 89%, 95% CI 64-98%), without false-positive diagnosis of EoE in RE or NH patients (specificity 100%, 95% CI 85-100%). Sensitivity was increased to 94% (95% CI 71-100%) when either the best predictive distal or proximal biopsy was used. CONCLUSION AND CLINICAL RELEVANCE We conclude that mRNA profiling of oesophageal tissue is an accurate diagnostic strategy in detecting EoE.
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Fasting and exercise differentially regulate BDNF mRNA expression in human skeletal muscle.
Walsh, JJ, Edgett, BA, Tschakovsky, ME, Gurd, BJ
Applied physiology, nutrition, and metabolism = Physiologie appliquee, nutrition et metabolisme. 2015;(1):96-8
Abstract
Brain-derived neurotrophic factor (BDNF) gene expression was measured in human skeletal muscle following 3 intensities of exercise and a 48-h fast. No change in BDNF mRNA was observed following exercise, while fasting upregulated BDNF by ∼ 3.5-fold. These changes were dissociated from changes in peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) following exercise (+2- to 15-fold) and fasting (∼-25%). These results challenge our understanding of the response of BDNF to energetic stress and highlight the importance of future work in this area.
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Serum indoleamine 2,3-dioxygenase activity is associated with reduced immunogenicity following vaccination with MVA85A.
Tanner, R, Kakalacheva, K, Miller, E, Pathan, AA, Chalk, R, Sander, CR, Scriba, T, Tameris, M, Hawkridge, T, Mahomed, H, et al
BMC infectious diseases. 2014;:660
Abstract
BACKGROUND There is an urgent need for improved vaccines to protect against tuberculosis. The currently available vaccine Bacille Calmette-Guerin (BCG) has varying immunogenicity and efficacy across different populations for reasons not clearly understood. MVA85A is a modified vaccinia virus expressing antigen 85A from Mycobacterium tuberculosis which has been in clinical development since 2002 as a candidate vaccine to boost BCG-induced protection. A recent efficacy trial in South African infants failed to demonstrate enhancement of protection over BCG alone. The immunogenicity was lower than that seen in UK trials. The enzyme Indoleamine 2,3-dioxygenase (IDO) catalyses the first and rate-limiting step in the breakdown of the essential amino acid tryptophan. T cells are dependent on tryptophan and IDO activity suppresses T-cell proliferation and function. METHODS Using samples collected during phase I trials with MVA85A across the UK and South Africa we have investigated the relationship between vaccine immunogenicity and IDO using IFN-γ ELISPOT, qPCR and liquid chromatography mass spectrometry. RESULTS We demonstrate an IFN-γ dependent increase in IDO mRNA expression in peripheral blood mononuclear cells (PBMC) following MVA85A vaccination in UK subjects. IDO mRNA correlates positively with the IFN-γ ELISPOT response indicating that vaccine specific induction of IDO in PBMC is unlikely to limit the development of vaccine specific immunity. IDO activity in the serum of volunteers from the UK and South Africa was also assessed. There was no change in serum IDO activity following MVA85A vaccination. However, we observed higher baseline IDO activity in South African volunteers when compared to UK volunteers. In both UK and South African serum samples, baseline IDO activity negatively correlated with vaccine-specific IFN-γ responses, suggesting that IDO activity may impair the generation of a CD4+ T cell memory response. CONCLUSIONS Baseline IDO activity was higher in South African volunteers when compared to UK volunteers, which may represent a potential mechanism for the observed variation in vaccine immunogenicity in South African and UK populations and may have important implications for future vaccination strategies. TRIAL REGISTRATION Trials are registered at ClinicalTrials.gov; UK cohort NCT00427830, UK LTBI cohort NCT00456183, South African cohort NCT00460590, South African LTBI cohort NCT00480558.
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Quantitative analysis of VEGF-C mRNA of extrahepatic cholangiocarcinoma with real-time PCR using samples obtained during endoscopic retrograde cholangiopancreatography.
Dobashi, A, Imazu, H, Tatsumi, N, Okabe, M, Ang, TL, Tajiri, H
Scandinavian journal of gastroenterology. 2013;(7):848-55
Abstract
OBJECTIVE Vascular endothelial growth factor (VEGF)-C overexpression in extrahepatic cholangiocarcinoma (ECC) has been shown to be correlated with lymph node metastasis. The intensity of immunohistochemical staining of VEGF-C protein in surgical samples has been used as index of VEGF-C overexpression in previous studies. The aim of the study was to examine if VEGF-C overexpression in ECC could be preoperatively detected by using samples obtained during ERCP. METHODS Consecutive patients who underwent endoscopic retrograde cholangiopancreatography (ERCP) for biliary stricture during the study period were prospectively analyzed. VEGF-C mRNA was quantified by real-time PCR methods using endoscopic samples obtained during ERCP. The high intensity of immunohistochemical staining of VEGF-C protein in surgical samples was used for the reference standard of VEGF-C overexpression. The level of S100P mRNA which was a novel diagnostic marker of ECC was also quantified to evaluate whether the endoscopic samples contained ECC cells. RESULTS Twenty-five patients were enrolled in this study. Eighteen patients were diagnosed as ECC and seven patients were diagnosed as benign biliary structure. Nine of eighteen patients with ECC, who showed positive S100P mRNA in endoscopic samples and received surgical resection, were finally analyzed. Receiver operating characteristics analysis yielded VEGF-C mRNA cut-off value of 3.85 for detection of VEGF-C overexpression, and the diagnostic performance of VEGF-C mRNA measurement in the endoscopic sample for VEGF-C overexpression reached sensitivity of 75.0%, specificity of 100%, and accuracy of 88.9%. CONCLUSION The quantification of VEGF-C mRNA of ECC with real-time PCR using endoscopic samples was useful for preoperative detection of VEGF-C overexpression.