Background: Radiostereometric analysis (RSA) is a highly accurate tool to detect implant migration and predict loosening following total knee arthroplasty (TKA). However, little is known about the... Show moreBackground: Radiostereometric analysis (RSA) is a highly accurate tool to detect implant migration and predict loosening following total knee arthroplasty (TKA). However, little is known about the predisposing risk factors for implant migration, nor which migration profile should be considered physiological (i.e., merely part of an implant-settling phase) and which should be considered pathological (i.e., having a high probability for implant loosening). By pooling individual participant data from long-term follow-up RSA studies, we aimed to identify predisposing risk factors for tibial component loosening. Methods: Individual data were collected for 630 patients from 11 RSA studies. The repeated measurements were analyzed with use of a linear mixed-effects model, determining the effect of age, sex, body mass index, diagnosis, preoperative and postoperative limb alignment, and prosthesis characteristics on tibial component migration over time, taking into account the clustering of patients within studies. Results: High initial migration was found to result in early mechanical loosening in 18 cases (2.9%) and septic loosening in 2 cases (0.3%), whereas stabilization of high initial migration occurred in 17 cases (2.7%). Late loosening occurred in 13 cases (2.1%). All other 580 cases (92.1%) showed early stabilization and remained stable over time. Mixed-effects model analyses showed that for cemented prostheses, sex, diagnosis, and posterior cruciate ligament type had an effect on migration, but these differences were nonsignificant when analyzing migration from 3 months onwards. Uncemented prostheses aligned in varus showed more migration than neutrally and valgus-aligned TKAs (p = 0.031), and this difference increased over time (p < 0.001). Significantly higher migration was observed following uncemented TKA without an osseointegration-promoting surface (p < 0.001). Conclusions: For cemented prostheses, increased migration during the first 3 postoperative months was observed for female patients, patients with rheumatoid arthritis, and patients who underwent a posterior-stabilized TKA. For uncemented prostheses, both postoperative varus alignment of the lower limb and the absence of an osseointegration-promoting surface significantly increased postoperative tibial component migration. Show less
We propose a top-down approach for pathway analysis of longitudinal metabolite data. We apply a score test based on a shared latent process mixed model which can identify pathways with... Show moreWe propose a top-down approach for pathway analysis of longitudinal metabolite data. We apply a score test based on a shared latent process mixed model which can identify pathways with differentially progressing metabolites. The strength of our approach is that it can handle unbalanced designs, deals with potential missing values in the longitudinal markers, and gives valid results even with small sample sizes. Contrary to bottom-up approaches, correlations between metabolites are explicitly modeled leveraging power gains. For large pathway sizes, a computationally efficient solution is proposed based on pseudo-likelihood methodology. We demonstrate the advantages of the proposed method in identification of differentially expressed pathways through simulation studies. Finally, longitudinal metabolite data from a mice experiment is analyzed to demonstrate our methodology. Show less
Signorelli, M.; Ebrahimpoor, M.; Veth, O.; Hettne, K.; Verwey, N.; Garcia-Rodriguez, R.; ... ; Spitali, P. 2021
DMD is a rare disorder characterized by progressive muscle degeneration and premature death. Therapy development is delayed by difficulties to monitor efficacy non-invasively in clinical trials. In... Show moreDMD is a rare disorder characterized by progressive muscle degeneration and premature death. Therapy development is delayed by difficulties to monitor efficacy non-invasively in clinical trials. In this study, we used RNA-sequencing to describe the pathophysiological changes in skeletal muscle of 3 dystrophic mouse models. We show how dystrophic changes in muscle are reflected in blood by analyzing paired muscle and blood samples. Analysis of repeated blood measurements followed the dystrophic signature at five equally spaced time points over a period of seven months. Treatment with two antisense drugs harboring different levels of dystrophin recovery identified genes associated with safety and efficacy. Evaluation of the blood gene expression in a cohort of DMD patients enabled the comparison between preclinical models and patients, and the identification of genes associated with physical performance, treatment with corticosteroids and body measures. The presented results provide evidence that blood RNA-sequencing can serve as a tool to evaluate disease progression in dystrophic mice and patients, as well as to monitor response to (dystrophin-restoring) therapies in preclinical drug development and in clinical trials. Show less
Background Blockade of cardiac sympathetic fibers by thoracic epidural anesthesia (TEA) was previously shown to reduce right and left ventricular systolic function and effective pulmonary arterial... Show moreBackground Blockade of cardiac sympathetic fibers by thoracic epidural anesthesia (TEA) was previously shown to reduce right and left ventricular systolic function and effective pulmonary arterial elastance. At conditions of constant paced heart rate, cardiac output and systemic hemodynamics were unchanged. In this study, we further investigated the effect of cardiac sympathicolysis during physical stress and increased oxygen demand.Methods In a cross-over design, 12 patients scheduled to undergo thoracic surgery performed dynamic ergometric exercise tests with and without TEA. Hemodynamics were monitored and biventricular function was measured by transthoracic two-dimensional and M-mode echocardiography, pulsed wave Doppler and tissue Doppler imaging.Results TEA attenuated systolic RV function (TV SMODIFIER LETTER PRIME: - 21%, P < 0.001) and LV function (MV SMODIFIER LETTER PRIME: - 14%, P = 0.025), but biventricular diastolic function was not affected. HR (- 11%, P < 0.001), SVI (- 15%, P = 0.006), CI (- 21%, P < 0.001) and MAP (- 12%, P < 0.001) were decreased during TEA, but SVR was not affected. Exercise resulted in significant augmentation of systolic and diastolic biventricular function. During exercise HR, SVI, CI and MAP increased (respectively, + 86%, + 19%, + 124% and + 17%, all P < 0.001), whereas SVR decreased (- 49%, P < 0.001). No significant interactions between exercise and TEA were found, except for RPP (P = 0.024) and MV E DT (P = 0.035).Conclusion Cardiac sympathetic blockade by TEA reduced LV and RV systolic function but did not significantly blunt exercise-induced increases in LV and RV function. These data indicate that additional mechanisms besides those controlled by the cardiac sympathetic nervous system are involved in the regulation of cardiac function during dynamic exercise. Show less
Purpose To describe the treatment outcomes and recurrence risk of chronic central serous chorioretinopathy (cCSC) in patients who had complete resolution of subretinal fluid (SRF) after either... Show morePurpose To describe the treatment outcomes and recurrence risk of chronic central serous chorioretinopathy (cCSC) in patients who had complete resolution of subretinal fluid (SRF) after either primary half-dose photodynamic therapy (PDT) or high-density subthreshold micropulse laser (HSML) in the PLACE trial.Methods This multicentre prospective follow-up study evaluated cCSC patients at 1 year after completion of the PLACE trial. Outcomes included: complete resolution of SRF on OCT, best-corrected visual acuity (BCVA) in Early Treatment of Diabetic Retinopathy Study (ETDRS) letters, retinal sensitivity on microperimetry and a visual function questionnaire (NEI-VFQ25).Results Twenty-nine out of 37 patients who received half-dose PDT and 15 out of 17 patients who received HSML could be evaluated at final visit. At final visit, 93% of the patients treated with half-dose PDT had complete resolution of SRF, compared with 53% of HSML-treated patients (p = 0.006). At final visit, the mean estimate increase in the PDT group compared with the HSML group was + 2.1 ETDRS letters, +0.15 dB for the retinal sensitivity and + 5.1 NEI-VFQ25 points (p = 0.103, p = 0.784 and p = 0.071, respectively). The mean estimated central retinal thickness in the half-dose PDT group was -7.0 mu m compared with the HSML group (p = 0.566). The mean estimated subfoveal choroidal thickness in the half-dose PDT group was -16.6 mu m compared with the HSML group (p = 0.359).Conclusion At 20 months after treatment, cCSC patients successfully treated with half-dose PDT are less likely to have recurrences of SRF compared with those successfully treated with HSML. However, functional outcomes did not differ. Show less
Signorelli, M.; Mason, A.G.; Mul, K.; Evangelista, T.; Mei, H.; Voermans, N.; ... ; Spitali, P. 2020
Facioscapulohumeral muscular dystrophy (FSHD) is caused by the expression of DUX4 in skeletal muscles. A number of therapeutic approaches are being developed to antagonize the events preceding and... Show moreFacioscapulohumeral muscular dystrophy (FSHD) is caused by the expression of DUX4 in skeletal muscles. A number of therapeutic approaches are being developed to antagonize the events preceding and following DUX4 expression that leads to muscular dystrophy. Currently, the possibility to evaluate treatment response in clinical trials is hampered by the lack of objective molecular biomarkers connecting the disease cause to clinical performance. In this study we employed RNA-seq to examine gene expression in PAXgene tubes obtained from two independent cohorts of FSHD patients. Analysis of gene expression profiles did not lead to the identification of genes or pathways differentially expressed in FSHD patients, or associated with disease severity. In particular, we did not find evidence that the DUX4 and PAX7 signatures were differentially expressed. On the other hand, we were able to improve patient classification by including single genes or groups of genes in classification models. The best classifier was ROPN1L, a gene known to be expressed in testis, coincidentally the typical location of DUX4 expression. These improvements in patient classification hold the potential to enrich the FSHD clinical trial toolbox. Show less
COPD risk is jointly determined by fetal lung development, lung growth rate and lung growth duration leading to the maximally attained level of lung function in early adulthood. Bronchopulmonary... Show moreCOPD risk is jointly determined by fetal lung development, lung growth rate and lung growth duration leading to the maximally attained level of lung function in early adulthood. Bronchopulmonary dysplasia (BPD) is considered a developmental arrest of alveolarisation. Long-term outcome studies of adult survivors born before the introduction of surfactant therapy ("old BPD") showed impaired lung function. We aimed to predict adult lung function and lung density in a cohort of premature infants born in the surfactant era, representing "new BPD".We studied a cohort of young adults born between 1987 and 1998, with (n=36) and without (n=28) BPD, treated in a single centre. Their perinatal characteristics and pulmonary function in infancy were studied by regression analysis for correlation with adult lung function and tissue lung density, all expressed by z-scores, at a mean age of 19.7 +/- 1.1 and 21 +/- 2.2 years, respectively.Although BPD adults had on average lower forced expiratory volume in 1 s (zFEV(1))/forced vital capacity (FVC) and zFEV(1) than those without, 55% of the BPD group had zFEV1/FVC values above the lower limit of normal (LLN). Moreover, above LLN values of diffusing capacity of the lung for carbon monoxide (zD(LCO)) was present in 89% of BPD adults and lung density in 71%. Only higher oxygen supply (F-IO2) at 36 weeks post-conception of BPD subjects had a trend with lower zFEV(1) (B=-6.4; p=0.053) and lower zD(LCO) (B=-4.1; p=0.023) at adulthood.No statistically significant predictors of new BPD were identified. Show less
We present a new modelling approach for longitudinal overdispersed counts that is motivated by the increasing availability of longitudinal RNA-sequencing experiments. The distribution of RNA-seq... Show moreWe present a new modelling approach for longitudinal overdispersed counts that is motivated by the increasing availability of longitudinal RNA-sequencing experiments. The distribution of RNA-seq counts typically exhibits overdispersion, zero-inflation and heavy tails; moreover, in longitudinal designs repeated measurements from the same subject are typically (positively) correlated. We propose a generalized linear mixed model based on the Poisson-Tweedie distribution that can flexibly handle each of the aforementioned features of longitudinal overdispersed counts. We develop a computational approach to accurately evaluate the likelihood of the proposed model and to perform maximum likelihood estimation. Our approach is implemented in theRpackageptmixed, which can be freely downloaded from CRAN. We assess the performance ofptmixedon simulated data, and we present an application to a dataset with longitudinal RNA-sequencing measurements from healthy and dystrophic mice. The applicability of the Poisson-Tweedie mixed-effects model is not restricted to longitudinal RNA-seq data, but it extends to any scenario where non-independent measurements of a discrete overdispersed response variable are available. Show less
Imbalanced transforming growth factor beta (TGF beta) and bone morphogenetic protein (BMP) signaling are postulated to favor a pathological pulmonary endothelial cell (EC) phenotype in pulmonary... Show moreImbalanced transforming growth factor beta (TGF beta) and bone morphogenetic protein (BMP) signaling are postulated to favor a pathological pulmonary endothelial cell (EC) phenotype in pulmonary arterial hypertension (PAH). BMP9 is shown to reinstate BMP receptor type-II (BMPR2) levels and thereby mitigate hemodynamic and vascular abnormalities in several animal models of pulmonary hypertension (PH). Yet, responses of the pulmonary endothelium of PAH patients to BMP9 are unknown. Therefore, we treated primary PAH patient-derived and healthy pulmonary ECs with BMP9 and observed that stimulation induces transient transcriptional signaling associated with the process of endothelial-to-mesenchymal transition (EndMT). However, solely PAH pulmonary ECs showed signs of a mesenchymal trans-differentiation characterized by a loss of VE-cadherin, induction of transgelin (SM22 alpha), and reorganization of the cytoskeleton. In the PAH cells, a prolonged EndMT signaling was found accompanied by sustained elevation of pro-inflammatory, pro-hypoxic, and pro-apoptotic signaling. Herein we identified interleukin-6 (IL6)-dependent signaling to be the central mediator required for the BMP9-induced phenotypic change in PAH pulmonary ECs. Furthermore, we were able to target the BMP9-induced EndMT process by an IL6 capturing antibody that normalized autocrine IL6 levels, prevented mesenchymal transformation, and maintained a functional EC phenotype in PAH pulmonary ECs. In conclusion, our results show that the BMP9-induced aberrant EndMT in PAH pulmonary ECs is dependent on exacerbated pro-inflammatory signaling mediated through IL6. Show less
PURPOSE: To assess whether chronic central serous chorioretinopathy (cCSC) patients without a complete resolution of subretinal fluid (SRF) after either half-dose photodynamic therapy (PDT) or high... Show morePURPOSE: To assess whether chronic central serous chorioretinopathy (cCSC) patients without a complete resolution of subretinal fluid (SRF) after either half-dose photodynamic therapy (PDT) or high-density subthreshold micropulse laser (HSML) treatment may benefit from crossover treatment.DESIGN: Multicenter prospective interventional case series.METHODS: cCSC patients with persistent SRF at the final visit of the PLACE trial were included. Patients received crossover treatment with either half-dose PDT or HSML.RESULTS: Thirty-two patients received PDT and 10 patients received HSML. At the first evaluation visit (6-8 weeks after treatment), 81% of patients in the PDT group had complete resolution of SRF, while none of the HSML-treated patients had complete resolution of SRF. At final visit (1 year after baseline), 78% (P = .030) and 67% (P = .109) of the patients, respectively, had a complete resolution of SRF. The mean retinal sensitivity in the PDT group increased from 21.7 dB (standard error [SE]: 0.9) to 23.4 dB (SE: 0.8) at evaluation visit 1 (P = .003), to 24.7dB (SE: 0.8) at final visit (P < .001), while there were no significant changes in the HSML group (23.7 dB [SE: 1.6] at baseline, 23.8 dB [SE: 1.4] at evaluation 1, and 23.3 dB [SE: 1.4] at final visit). The mean visual acuity and mean visual quality-of-life questionnaire score did not change significantly in both groups.CONCLUSIONS: Crossover to half-dose PDT after previous unsuccessful HSML treatment for cCSC may lead to improved anatomic and functional endpoints, while crossover to HSML after half-dose PDT does not seem to significantly affect these endpoints. (C) 2020 The Authors. Published by Elsevier Inc. Show less
Ebrahimpoor, M.; Spitali, P.; Hettne, K.; Tsonaka, R.; Goeman, J. 2020
Studying sets of genomic features is increasingly popular in genomics, proteomics and metabolomics since analyzing at set level not only creates a natural connection to biological knowledge but... Show moreStudying sets of genomic features is increasingly popular in genomics, proteomics and metabolomics since analyzing at set level not only creates a natural connection to biological knowledge but also offers more statistical power. Currently, there are two gene-set testing approaches, self-contained and competitive, both of which have their advantages and disadvantages, but neither offers the final solution. We introduce simultaneous enrichment analysis (SEA), a new approach for analysis of feature sets in genomics and other omics based on a new unified null hypothesis, which includes the self-contained and competitive null hypotheses as special cases. We employ closed testing using Simes tests to test this new hypothesis. For every feature set, the proportion of active features is estimated, and a confidence bound is provided. Also, for every unified null hypotheses, a P-value is calculated, which is adjusted for family-wise error rate. SEA does not need to assume that the features are independent. Moreover, users are allowed to choose the feature set(s) of interest after observing the data. We develop a novel pipeline and apply it on RNA-seq data of dystrophin-deficient mdx mice, showcasing the flexibility of the method. Finally, the power properties of the method are evaluated through simulation studies. Show less
Tsonaka, R.; Signorelli, M.; Sabir, E.; Seyer, A.; Hettne, K.; Aartsma-Rus, A.; Spitali, P. 2020
Duchenne muscular dystrophy is a severe pediatric neuromuscular disorder caused by the lack of dystrophin. Identification of biomarkers is needed to support and accelerate drug development.... Show moreDuchenne muscular dystrophy is a severe pediatric neuromuscular disorder caused by the lack of dystrophin. Identification of biomarkers is needed to support and accelerate drug development. Alterations of metabolites levels in muscle and plasma have been reported in pre-clinical and clinical cross-sectional comparisons. We present here a 7-month longitudinal study comparing plasma metabolomic data in wild-type and mdx mice. A mass spectrometry approach was used to study metabolites in up to five time points per mouse at 6, 12, 18, 24 and 30 weeks of age, providing an unprecedented in depth view of disease trajectories. A total of 106 metabolites were studied. We report a signature of 31 metabolites able to discriminate between healthy and disease at various stages of the disease, covering the acute phase of muscle degeneration and regeneration up to the deteriorating phase. We show how metabolites related to energy production and chachexia (e.g. glutamine) are affected in mdx mice plasma over time. We further show how the signature is connected to molecular targets of nutraceuticals and pharmaceutical compounds currently in development as well as to the nitric oxide synthase pathway (e.g. arginine and citrulline). Finally, we evaluate the signature in a second longitudinal study in three independent mouse models carrying 0, 1 or 2 functional copies of the dystrophin paralog utrophin. In conclusion, we report an in-depth metabolomic signature covering previously identified associations and new associations, which enables drug developers to peripherally assess the effect of drugs on the metabolic status of dystrophic mice. Show less
Schwach, V.; Fernandes, M.G.; Maas, S.; Gerhardt, S.; Tsonaka, R.; Weerd, L. van der; ... ; Salvatori, D.C.F. 2020
Aims Cardiovascular diseases caused by loss of functional cardiomyocytes (CMs) are a major cause of mortality and morbidity worldwide due in part to the low regenerative capacity of the adult human... Show moreAims Cardiovascular diseases caused by loss of functional cardiomyocytes (CMs) are a major cause of mortality and morbidity worldwide due in part to the low regenerative capacity of the adult human heart. Human pluripotent stem cell (hPSC)-derived cardiovascular progenitor cells (CPCs) are a potential cell source for cardiac repair. The aim of this study was to examine the impact of extensive remuscularization and coincident revascularization on cardiac remodelling and function in a mouse model of myocardial infarction (MI) by transplanting doxycycline (DOX)-inducible (Tet-On-MYC) hPSC-derived CPCs in vivo and inducing proliferation and cardiovascular differentiation in a drug-regulated manner.Methods and results CPCs were injected firstly at a non-cardiac site in Matrigel suspension under the skin of immunocompromised mice to assess their commitment to the cardiovascular lineage and ability to self-renew or differentiate in vivo when instructed by systemically delivered factors including DOX and basic fibroblast growth factor (bFGF). CPCs in Matrigel were then injected intra-myocardially in mice subjected to MI to assess whether expandable CPCs could mediate cardiac repair. Transplanted CPCs expanded robustly both subcutis and in the myocardium using the same DOX/growth factor inducing regime. Upon withdrawal of these cell-renewal factors, CPCs differentiated with high efficiency at both sites into the major cardiac lineages including CMs, endothelial cells, and smooth muscle cells. After MI, engraftment of CPCs in the heart significantly reduced fibrosis in the infarcted area and prevented left ventricular remodelling, although cardiac function determined by magnetic resonance imaging was unaltered.Conclusion Replacement of large areas of muscle may be required to regenerate the heart of patients following MI. Our human/mouse model demonstrated that proliferating hPSC-CPCs could reduce infarct size and fibrosis resulting in formation of large grafts. Importantly, the results suggested that expanding transplanted cells in situ at the progenitor stage maybe be an effective alternative causing less tissue damage than injection of very large numbers of CMs. Show less
Hoekstra, P.T.; Casacuberta Partal, M.; Lieshout, L. van; Corstjens, P.L.A.M.; Tsonaka, R.; Assare, R.K.; ... ; Dam, G.J. van 2020
BackgroundPreventive chemotherapy with praziquantel (PZQ) is the cornerstone of schistosomiasis control. However, a single dose of PZQ (40 mg/kg) does not cure all infections. Repeated doses of PZQ... Show moreBackgroundPreventive chemotherapy with praziquantel (PZQ) is the cornerstone of schistosomiasis control. However, a single dose of PZQ (40 mg/kg) does not cure all infections. Repeated doses of PZQ at short intervals might increase efficacy in terms of cure rate (CR) and intensity reduction rate (IRR). Here, we determined the efficacy of a single versus four repeated treatments with PZQ on Schistosoma mansoni infection in school-aged children from Cote d'Ivoire, using two different diagnostic tests.MethodsAn open-label, randomized controlled trial was conducted from October 2018 to January 2019. School-aged children with a confirmed S. mansoni infection based on Kato-Katz (KK) and point-of-care circulating cathodic antigen (POC-CCA) urine cassette test were randomly assigned to receive either a single or four repeated doses of PZQ, administered at two-week intervals. The primary outcome was the difference in CR between the two treatment arms, measured by triplicate KK thick smears 10 weeks after the first treatment. Secondary outcomes included CR estimated by POC-CCA, IRR by KK and POC-CCA, and safety of repeated PZQ administration.Principal findingsDuring baseline screening, 1,022 children were assessed for eligibility of whom 153 (15%) had a detectable S. mansoni infection, and hence, were randomized to the standard treatment group (N = 70) and the intense treatment group (N = 83). Based on KK, the CR was 42% (95% confidence interval (CI) 31-52%) in the standard treatment group and 86% (95% CI 75-92%) in the intense treatment group. Observed IRR was 72% (95% CI 55-83%) in the standard treatment group and 95% (95% CI 85-98%) in the intense treatment group. The CR estimated by POC-CCA was 18% (95% CI 11-27%) and 36% (95% CI 26-46%) in the standard and intense treatment group, respectively. Repeated PZQ treatment did not result in a higher number of adverse events.Author summaryThe previously established efficacy of the widely used drug praziquantel (PZQ) against schistosomiasis might have been overestimated due to the use of inaccurate diagnostic methods. Repeated PZQ treatment at short intervals in areas with ongoing transmission could more effectively target non-susceptible schistosomula as they will have matured into drug susceptible worms within a few weeks. In the current study, we aimed to determine the cure rate (CR) of repeated PZQ, measured by the Kato-Katz (KK) technique and the point-of-care circulating cathodic antigen (POC-CCA) test, respectively. An open-label, randomized controlled trial was conducted assigning 153 school-aged children with a confirmed Schistosoma mansoni infection to two groups, one receiving a single PZQ treatment, while the second group received four repeated PZQ treatments, given at two-week intervals. Based on the KK test, the CR was significantly higher after four repeated treatments compared to a single treatment. When using POC-CCA, a diagnostic method that has not been utilized before in studies assessing the efficacy of four repeated PZQ treatments, the CR was much lower, even after four repeated PZQ treatments. Our results indicate that worms are still present after multiple PZQ treatments and that PZQ might be less efficacious than previously published. Show less
Signorelli, M.; Ayoglu, B.; Johansson, C.; Lochmuller, H.; Straub, V.; Muntoni, F.; ... ; Spitali, P. 2019
