bluuebunny/crossref_metadata_2025 huggingface dataset:
Plasma Concentration of Eltrombopag Correlated with the Efficacy in the Treatment of Refractory Aplastic Anemia: A Single-Center Study in China
Authors: Zuo Wei , Zhang Bo , Ruan Jing , Chen Miao , Li Jian , Han Bing
Published: February 2021

Background and purpose: Eltrombopag (ELT) was effective in the treatment of relapse/refractory aplastic anemia (AA) patients. Response and adverse drug reaction (ADR) differed greatly among individuals even at the same dosage of ELT.

Methods: Patients diagnosed with non-severe aplastic anemia (NSAA) between January 2018 to January 2019 in Peking Union Medical Colleague Hospital who were refractory to immunosuppressive therapy were treated with ELT and followed up for at least 6 months. Plasma concentrations of ELT were detected by the high-performance liquid chromatography-mass spectrometry after at least two months of ELT treatment and staying for that dosage for at least two weeks. Dose-concentration, concentration-response and concentration- ADR relation were evaluated.

Results: Among the 72 patients treated with ELT during that period, 44 patients with complete data were enrolled. 6 (13.6%) were males and 38 were females (86.4%) with a median age of 54 years (IQR: 38.5-63). By the time of plasma concentration detected, the median dosage of ELT was 75(IQR 50-100) mg/d and median time for total ELT exposure was 3 (IQR 2.0-6.0) months, and 37 (70.5%) patients responded to ELT. The median concentration of ELT was 10.4μg/ml (IQR 3.7-24.4μg/ml). The concentration of ELT was positively correlated with the daily dose of ELT (r = 0.68, p < 0.001). Multivariate logistic regression analysis showed that the risk of inefficacy of ELT with a concentration between 11.2-15.2 μg/ml was 0.028-fold (95% CI: 0.001-0.864; P=0.041) to those with a concentration between 3.2-7.2μg/ml. The cutoff value of concentration for ELT efficacy was 12.50μg/ml by receiver operation characteristic curve. A higher risk of ADR was related to a longer total exposure to ELT (P=0.012). Although not significant, odd ratio (OR) increased with ELT concentration, suggesting a possible elevated risk of ADR correlated with blood concentration.

Conclusions: ELT is effective for the treatment of NSAA with accepTable side effects. Plasma concentration of ELT correlated with the dose and the effects of ELT.

Disclosures

No relevant conflicts of interest to declare.

OffLabel Disclosure:

Eltrombopag (ELT) has been proven to induce trilinear hematopoietic responses in relapse/refractory AA patients and has been approval as a monotherapy in relapsed/refractory SAA in the USA and Europe. However, the use of ELT is only approved for idiopathic thrombocytopenic purpura (ITP) in China by the National Medical Products Administration (NMPA), for AA it is still "off-label".
Incidence and Outcomes of Acute Transfusion Reactions in Hospitalized Patients in the United States
Authors: Kumar Rohit , Malapati Sindhu , Singh Sunny R K , Mamedov Bokhodir , Shah Myra R , Bhandari Shruti
Published: February 2021

Introduction

Acute transfusion reactions (ATRs) have a broad spectrum of presentations ranging from benign to life-threatening. Due to the rarity of these reactions, there is a paucity of data regarding their incidence and clinical outcomes. The objectives of this study were to determine the incidence of ATRs, its risk factors, and associated mortality.

Methods:

We reviewed the National Inpatient Sample (NIS) database 2014 for admissions where the patient (>=18 years old) was transfused blood products. The NIS is a large publicly available all-payer inpatient healthcare database designed to produce U.S. regional and national estimates of inpatient utilization, access, charges, quality, and outcomes. ATRs were identified using ICD-9 CM codes for transfusion-associated circulatory overload (TACO), transfusion-related acute lung injury (TRALI), febrile non-hemolytic transfusion reactions (FNHTR), acute infections, anaphylaxis, and acute hemolytic reaction. Pearson's chi-square and student's t-test were used to compare categorical and continuous variables between hospitalizations with versus without ATRs, respectively. Multivariate logistic regression analysis was done to determine the risk factors for common ATRs (TACO, TRALI, and FNHTR). A multivariate cox proportional model was built to compare the mortality of two study groups. A 2-sided p-value ≤ 0.05 was considered significant.

Results:

A total of 2,134,691 hospitalizations were associated with the transfusion of blood products. ATRs were documented in 0.2% of the hospitalizations (TACO 0.08%, TRALI 0.06%, FNHTR 0.09%, others 0.003%). The group that had ATRs was slightly younger (median age 67 vs 68 years, p=0.002), had the same proportion of females (58.3% vs 55.3%, p=0.055), less comorbidity score (28.7% vs 31.7% had Charlson Comorbidity Index >3, p=0.042) and more critically ill (17.8% vs 10.5% on mechanical ventilation, p<0.001) compared to group without ATRs. Hospitalizations with ATRs had longer median length of stay (7 vs 6 days, p<0.001) and higher median hospital cost ($64,399 vs $53,912, p<0.001) compared to without ATRs. The risk factors for common ATRs (odds ratio, OR) are mentioned in the table. ATRs were not associated with increased risk of mortality (combined HR 0.89 95%CI 0.71-1.12, p=0.321).

Conclusions:

Nationally, the incidence of ATRs is low in hospitalized patients and it is not associated with increased mortality. This large database analysis gives insight into the risk factors associated with different ATRs.

Disclosures

No relevant conflicts of interest to declare.
Endothelial Microparticles Delivering Lncrna NEAT1 Are Involved in the Pathophysiology of aGVHD Following Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Zhang Ran , Cao Weijie , Li Li , Wan Dingming , Jiang Zhongxing , Sun Kai , Xia Linghui
Published: February 2021

Endothelial microparticles (EMPs) upregulation has been observed in the pathological process of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). The biological role and therapeutic potential of long non-coding RNAs (lncRNAs) carried by EMPs in aGVHD after allo-HSCT are still open questions. Herein, we investigated the functional significance of the lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) delivered by EMPs in aGVHD. To address this question, in the present study, we performed a comprehensive analysis of the lncRNA NEAT1 expression and EMPs in aGVHD by a series of in vivo and in vitro experiments. We found that expressions of lncRNA NEAT1 in MPs were remarkably elevated in aGVHD mice than in non-aGVHD mice. Furthermore, the lncRNA NEAT1 levels in the MPs were remarkably higher than those in the plasma of the aGVHD mice. We then used TNF-α (100 ng/ml) to stimulate primary mouse aortic endothelial cells (MAECs) to shed EMPs, and the expression of lncRNA NEAT1 in the EMPs and their maternal cells was compared. The lncRNA NEAT1 levels in the EMPs produced by TNF-α-stimulated MAECs were significantly higher than that in both their maternal cells and non-stimulated EMPs. Our study demonstrates that EMPs could assemble and concentrate lncRNA NEAT1. Taken together, NEAT1 delivered by EMPs could be used as a diagnostic and prognostic surrogate marker for aGVHD, and its targeting could therefore represent a promising strategy for novel therapeutic options in life-threatening aGVHD after allo-HSCT.

Disclosures

No relevant conflicts of interest to declare.
The Serine Protease Matriptase Acts As a Tumour Suppressor in Multiple Myeloma
Authors: Steiro Ida , Abdollahi Pegah , Børset Magne , Slørdahl Tobias S.
Published: February 2021

Both in newly diagnosed multiple myeloma (MM) and during progression of the disease, malignant plasma cells are found circulating in peripheral blood as well as in the bone marrow (BM). The disseminated nature of MM is strongly dependent on the interplay between the cancer cells and the BM microenvironment, promoting myeloma cell migration in the BM. Matriptase (ST14), a type-II transmembrane serine protease primarily found in epithelial tissues, is overexpressed in a variety of human malignancies and is sufficient to induce tumour formation in mice. Frequently, a concomitant reduction in the levels of its cognate inhibitor hepatocyte growth factor activator inhibitor (HAI)-1 (SPINT1) is observed in carcinomas, while expression and function of the related inhibitor HAI-2 (SPINT2) is yet to be clarified. Dysregulated expression causing increased matriptase proteolytic activity has been associated with cancer growth, survival and metastasis. Here, we show for the first time a role of matriptase as a possible tumour suppressor in myeloma pathogenesis. Gene expression analysis of primary cells from MM patients (n=24) and human myeloma cell lines (n=8) revealed highly variable levels of matriptase, HAI-1 and HAI-2. This observation prompted us to investigate the functional role of matriptase in vitro. We showed that stable overexpression of matriptase in INA-6, a MM cell line with no endogenous ST14 expression, reduced migration by more than 50% in response to the combination of the pro-migratory cytokines stromal cell-derived factor-1 alpha (SDF-1α) and hepatocyte growth factor (HGF, Fig. 1A). Conversely, stable knockdown of matriptase in two MM cell lines with high endogenous matriptase expression (RPMI-8226 and JJN-3) significantly enhanced migration in vitro. Mechanistically, matriptase overexpression blocked activation of Src kinase (Fig. 1B), well-known as a critical player in metastasis formation promoting cancer cell motility, invasiveness and angiogenesis. In agreement with our result, previous studies have demonstrated the activation of Src family kinases (SFK) downstream SDF-1/CXCR4-signaling. Finally, we performed survival analyses in the public available MMRF CoMMpass trial database (release version IA14). Low ST14 expression was associated with significant worse overall survival (P=0.05, Fig. 1C) and progression-free survival (P=0.02, Fig. 1D). Altogether, our data are in marked contrast to the role ascribed to matriptase in epithelial and certain non-epithelial tumours as an oncogenic protein and an unfavourable prognostic marker. In conclusion, these findings suggest a novel role of matriptase as a tumour suppressor in MM pathogenesis.

Disclosures

Slørdahl: Celgene: Consultancy; Janssen and Celgene: Honoraria.
The Neurotransmitter Receptor Gabbr1 Regulates Proliferation and Function of Hematopoietic Stem and Progenitor Cells
Authors: Elujoba-Bridenstine Adedamola , Shao Lijian , Zink Katherine , Sanchez Laura , Pajcini Kostandin V. , Tamplin Owen J.
Published: February 2021

Hematopoietic stem and progenitor cells (HSPCs) are multipotent cells which differentiate to maintain and replenish blood lineages throughout life. Due to these characteristics, HSPC transplants represent a cure for patients with a variety of hematological disorders. HSPC function and behavior is tightly regulated by various cell types and factors in the bone marrow niche. The nervous system has been shown to indirectly influence hematopoiesis by innervating the niche; however, we present a direct route of HSPC regulation via expression of neurotransmitter receptors on HSPC surface. We have identified Gamma Aminobutyric acid (GABA) receptor B subunit 1 (Gabbr1), a hitherto unknown hematopoietic player, as a regulator of HSPC function. GABBR1 is known to be expressed on human HSPCs (Steidl et al., Blood 2004), however its function in their regulation remains unknown. Based on published RNA-seq data (Nestorowa et al., Blood 2016), we discovered that Gabbr1 is expressed on a subset of HSPCs. We confirmed this expression using RT-qPCR to assay hematopoietic populations in the bone marrow (BM). Surface receptor expression analysis showed that Gabbr1 protein is expressed on a subset of BM HSPCs. To detect GABA, the ligand for Gabbr1 in the BM microenvironment, we utilized imaging mass spectrometry (IMS). We detected regionally specific GABA signal in the endosteal region of the BM. We further identified B cells as a cellular source of GABA in the BM. To understand the role of Gabbr1 in hematopoiesis, we generated CRISPR-Cas9 Gabbr1 null mutants on a C57/BL6 background suitable for hematopoietic studies and studied their hematopoietic phenotype. We discovered a decrease in the absolute number of Lin-Sca1+cKit+ (LSK) HSPCs, but the long-term hematopoietic stem cells (LT-HSCs) remain unaffected. Further analysis of peripheral blood of Gabbr1 null mutants showed decreased white blood cells due to reduced B220+ cells. This differentiation defect was confirmed in an in vitro differentiation assay where Gabbr1 null HSPCs displayed an impaired ability to produce B cells. We show that Gabbr1 null HSCs show diminished reconstitution ability when transplanted in a competitive setting. Reduced Gabbr1 null HSC reconstitution persisted in secondary transplant recipients indicating a cell autonomous role for Gabbr1 in regulating reconstitution of HSCs in transplant recipients. Our results show a crucial role for Gabbr1 in HSPC regulation and may translate to human health as a rare human SNP within the GABBR1 locus that correlates with altered leukocyte counts has been reported (Astle et al., Cell 2016). Our studies indicate an important role for Gabbr1 in HSPC reconstitution and differentiation into B cell lineages.

Disclosures

No relevant conflicts of interest to declare.
Low-Dose Triptolide Promotes MDM2 Inhibitor Nutlin3a to Induce Acute Myeloid Leukemia Cell Death Via p53-Dependent and -Independent Mechanisms
Authors: Chen Qinwei , Deng Suqi , Lin Zhijuan , Deng Manman , Ding Lihong , Zha Jie , Zhou Yong , Jiang Yuelong , Carter Bing Z , Xu Bing
Published: February 2021

The tumor suppressor p53 is central to hematopoietic stem cell function. Loss of p53 function is associated with poor prognosis of acute myeloid leukemia (AML). P53 negative regulator MDM2 is frequently overexpressed in AML, thus it becomes an attractive therapeutic target for the treatment of AML with wild-type p53 (Wt-p53). Targeting MDM2 to restore p53 activity has been assessed in AML, but clinical outcomes are less promising. Notably, p53-mutated AML still lacks effective treatment approaches. Therefore, our study strived to explore potent therapeutics to target AML with p53 wild-type and mutations.

Triptolide (TPL), an ancient herbal medicine, has been used to treat immune disorders for centuries. Accumulating evidence including ours have proven that low dose of TPL potentiates the efficacy of chemotherapies in a broad range of tumor types. In this study, we evaluated the anti-leukemic efficacy of combining low dose TPL with Nutlin3a, a MDM2 inhibitor, against AML with p53 wild-type and mutations. We found that low-dose TPL synergized with Nutlin3a to induce mitochondrial apoptosis in AML cells with Wt-p53 bothin vitroandin vivo. Underlying mechanism for the synergy showed that Nutlin3a upregulated pro-apoptosis factors (e.g. PUMA, p21), while low-dose TPL suppressed MDM2 transcription and reduced anti-apoptosis proteins (e.g. XIAP, Mcl-1) in Wt-p53 AML cells. Interestingly, we also observed that TPL but not Nutlin3a induced cell death in p53 shRNA knockdown, p53 deficiency cell lines (e.g. THP-1, HL-60) and primary samples. Thus, the p53-independent role of TPL was consequently focused. Our RNA-Seq analysis identified 621 differential expression genes (DEGs) after TPL treatment. These genes, including MYC, ATF4 and 4EBP1, were enriched in intrinsic apoptosis pathway responding to ER stress and thus we concluded that Triptolide played its p53 independent role via MYC-ATF4 axis.

Collectively, these findings suggest that coadministration of Nutlin3a with low dose TPL would benefit for AML patients.

Disclosures

Carter: AstraZeneca:Research Funding;Syndax:Research Funding;Ascentage:Research Funding;Amgen:Research Funding.

OffLabel Disclosure:

Triptolide (TPL), an ancient herbal medicine, is applying to treat immune disorders.
LYG1 Deficiency Attenuates the Severity of Acute Graft-Versus-Host Disease Via Skewing T Cells Polarization Towards Treg Cells
Authors: Liu Huihui , Yu Zhengyu , Tang Bo , Miao Shengchao , Qin Chenchen , Li Yuan , Liang Zeyin , Zhang Yang , Ren Hanyun , Dong Yujun
Published: February 2021

Acute graft-versus-host disease (aGVHD) is a lethal complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT). As a complex immunopathology, aGVHD depends on the recognition of host antigens by donor T cells and induces augmented response of alloreactive T cells. Despite considerable achievements in the treatment of aGVHD, it remains a major clinical problem for the patients undergoing allo-HSCT. Therefore, it is necessary to further illustrate new mechanisms and develop novel therapeutic strategies of aGVHD. Previously we reported LYG1 (Lysozyme G-like 1) as a novel classical secretory protein promoted antitumor function of T cell. In this study, the role of LYG1 in aGVHD was investigated. Firstly, we examined whether LYG1 affected the alloreactivity of CD4+ T cells in vitro by MLR assay and discovered that LYG1 deficiency reduced the activation of CD4+ T cells and Th1 ratio, but increased Treg ratio. Then we confirmed these observations using a major MHC mismatched aGVHD model by transferring T cells sorting from WT B6 or Lyg1-/- mice with bone marrow cells from WT B6 mice into lethally irradiated BALB/c mice. The alloreactive CD4+ T cells and the proportions of Th1 cells decreased whereas the proportions of Treg cells increased in spleens and livers in mice receiving Lyg1-/- T cells. LYG1-deficient T cells attenuated aGVHD severity, inhibited the expression of CXCL9 and CXCL10 and restrained CD4+ T cells infiltrating in livers. Furthermore, administration of recombinant LYG1 protein intraperitoneally aggravated aGVHD by promoting IFN-γ production. More importantly, LYG1 deficiency did not affect GVT (graft-versus-tumor) effects. In summary, we demonstrate LYG1 regulates aGVHD via modulating the alloreactivity of CD4+ T cells and differentiation of Th1/Treg cells. Our study indicates that LYG1 may be a novel target in aGVHD by mitigating aGVHD without impairing GVT function. The therapeutic effect of targeting LYG1 is required in future investigations.

Funding

This study was supported by grant from The National Natural Science Foundation of China (NSFC) (Grant Number 81600144) and grant from Research Foundation of Peking University First Hospital.

Disclosures

No relevant conflicts of interest to declare.
N-glycan–mediated shielding of ADAMTS13 prevents binding of pathogenic autoantibodies in immune-mediated TTP
Authors: Ercig Bogac , Graça Nuno A. G. , Kangro Kadri , Arfman Tom , Wichapong Kanin , Hrdinová Johana , Kaijen Paul , van Alphen Floris P. J. , van den Biggelaar Maartje , Vanhoorelbeke Karen , Veyradier Agnès , Coppo Paul , Reutelingsperger Chris , Nicolaes Gerry A. F. , Männik Andres , Voorberg Jan
Published: February 2021

Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is an autoimmune disorder caused by the development of autoantibodies targeting different domains of ADAMTS13. Profiling studies have shown that residues R568, F592, R660, Y661, and Y665 within exosite-3 of the spacer domain provide an immunodominant region of ADAMTS13 for pathogenic autoantibodies that develop in patients with iTTP. Modification of these 5 core residues with the goal of reducing autoantibody binding revealed a significant tradeoff between autoantibody resistance and proteolytic activity. Here, we employed structural bioinformatics to identify a larger epitope landscape on the ADAMTS13 spacer domain. Models of spacer-antibody complexes predicted that residues R568, L591, F592, K608, M609, R636, L637, R639, R660, Y661, Y665, and L668 contribute to an expanded epitope within the spacer domain. Based on bioinformatics-guided predictions, we designed a panel of N-glycan insertions in this expanded epitope to reduce the binding of spacer domain autoantibodies. One N-glycan variant (NGLY3-ADAMTS13, containing a K608N substitution) showed strongly reduced reactivity with TTP patient sera (28%) as compared with WT-ADAMTS13 (100%). Insertion of an N-glycan at amino acid position 608 did not interfere with processing of von Willebrand factor, positioning the resulting NGLY3-ADAMTS13 variant as a potential novel therapeutic option for treatment of iTTP.
BAFF promotes heightened BCR responsiveness and manifestations of chronic GVHD after allogeneic stem cell transplantation
Authors: Jia Wei , Poe Jonathan C. , Su Hsuan , Anand Sarah , Matsushima Glenn K. , Rathmell Jeffrey C. , Maillard Ivan , Radojcic Vedran , Imai Kazuhiro , Reyes Nancy J. , Cardona Diana M. , Li Zhiguo , Suthers Amy N. , Curry-Chisolm Itaevia M. , DiCioccio Rachel A. , Saban Daniel R. , Chen Benny J. , Chao Nelson J. , Sarantopoulos Stefanie
Published: February 2021

Patients with chronic graft-versus-host disease (cGVHD) have increased B cell–activating factor (BAFF) levels, but whether BAFF promotes disease after allogeneic bone marrow transplantation (allo-BMT) remains unknown. In a major histocompatibility complex–mismatched model with cGVHD-like manifestations, we first examined B-lymphopenic μMT allo-BMT recipients and found that increased BAFF levels in cGVHD mice were not merely a reflection of B-cell number. Mice that later developed cGVHD had significantly increased numbers of recipient fibroblastic reticular cells with higher BAFF transcript levels. Increased BAFF production by donor cells also likely contributed to cGVHD, because BAFF transcript in CD4+ T cells from diseased mice and patients was increased. cGVHD manifestations in mice were associated with high BAFF/B-cell ratios and persistence of B-cell receptor (BCR)–activated B cells in peripheral blood and lesional tissue. By employing BAFF transgenic (Tg) mice donor cells, we addressed whether high BAFF contributed to BCR activation in cGVHD. BAFF increased NOTCH2 expression on B cells, augmenting BCR responsiveness to surrogate antigen and NOTCH ligand. BAFF Tg B cells had significantly increased protein levels of the proximal BCR signaling molecule SYK, and high SYK protein was maintained by BAFF after in vitro BCR activation or when alloantigen was present in vivo. Using T cell–depleted (BM only) BAFF Tg donors, we found that BAFF promoted cGVHD manifestations, circulating GL7+ B cells, and alloantibody production. We demonstrate that pathologic production of BAFF promotes an altered B-cell compartment and augments BCR responsiveness. Our findings compel studies of therapeutic targeting of BAFF and BCR pathways in patients with cGVHD.
Factor VIIa induces extracellular vesicles from the endothelium: a potential mechanism for its hemostatic effect
Authors: Das Kaushik , Keshava Shiva , Ansari Shabbir A. , Kondreddy Vijay , Esmon Charles T. , Griffin John H. , Pendurthi Usha R. , Rao L. Vijaya Mohan
Published: February 2021

Recombinant factor FVIIa (rFVIIa) is used as a hemostatic agent to treat bleeding disorders in hemophilia patients with inhibitors and other groups of patients. Our recent studies showed that FVIIa binds endothelial cell protein C receptor (EPCR) and induces protease-activated receptor 1 (PAR1)-mediated biased signaling. The importance of FVIIa-EPCR-PAR1–mediated signaling in hemostasis is unknown. In the present study, we show that FVIIa induces the release of extracellular vesicles (EVs) from endothelial cells both in vitro and in vivo. Silencing of EPCR or PAR1 in endothelial cells blocked the FVIIa-induced generation of EVs. Consistent with these data, FVIIa treatment enhanced the release of EVs from murine brain endothelial cells isolated from wild-type (WT), EPCR-overexpressing, and PAR1-R46Q–mutant mice, but not EPCR-deficient or PAR1-R41Q–mutant mice. In vivo studies revealed that administration of FVIIa to WT, EPCR-overexpressing, and PAR1-R46Q–mutant mice, but not EPCR-deficient or PAR1-R41Q–mutant mice, increased the number of circulating EVs. EVs released in response to FVIIa treatment exhibit enhanced procoagulant activity. Infusion of FVIIa-generated EVs and not control EVs to platelet-depleted mice increased thrombin generation at the site of injury and reduced blood loss. Administration of FVIIa-generated EVs or generation of EVs endogenously by administering FVIIa augmented the hemostatic effect of FVIIa. Overall, our data reveal that FVIIa treatment, through FVIIa-EPCR-PAR1 signaling, releases EVs from the endothelium into the circulation, and these EVs contribute to the hemostatic effect of FVIIa.
A combination of cyclophosphamide and interleukin-2 allows CD4+ T cells converted to Tregs to control scurfy syndrome
Authors: Delville Marianne , Bellier Florence , Leon Juliette , Klifa Roman , Lizot Sabrina , Vinçon Hélène , Sobrino Steicy , Thouenon Romane , Marchal Armance , Garrigue Alexandrine , Olivré Juliette , Charbonnier Soëli , Lagresle-Peyrou Chantal , Amendola Mario , Schambach Axel , Gross David , Lamarthée Baptiste , Benoist Christophe , Zuber Julien , André Isabelle , Cavazzana Marina , Six Emmanuelle
Published: February 2021

Immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is caused by mutations in forkhead box P3 (FOXP3), which lead to the loss of function of regulatory T cells (Tregs) and the development of autoimmune manifestations early in life. The selective induction of a Treg program in autologous CD4+ T cells by FOXP3 gene transfer is a promising approach for curing IPEX. We have established a novel in vivo assay of Treg functionality, based on adoptive transfer of these cells into scurfy mice (an animal model of IPEX) and a combination of cyclophosphamide (Cy) conditioning and interleukin-2 (IL-2) treatment. This model highlighted the possibility of rescuing scurfy disease after the latter’s onset. By using this in vivo model and an optimized lentiviral vector expressing human Foxp3 and, as a reporter, a truncated form of the low-affinity nerve growth factor receptor (ΔLNGFR), we demonstrated that the adoptive transfer of FOXP3-transduced scurfy CD4+ T cells enabled the long-term rescue of scurfy autoimmune disease. The efficiency was similar to that seen with wild-type Tregs. After in vivo expansion, the converted CD4FOXP3 cells recapitulated the transcriptomic core signature for Tregs. These findings demonstrate that FOXP3 expression converts CD4+ T cells into functional Tregs capable of controlling severe autoimmune disease.
Visual4DTracker: a tool to interact with 3D + t image stacks
Authors: Cordelli Ermanno , Soda Paolo , Iannello Giulio
Published: February 2021

Background

Biological phenomena usually evolves over time and recent advances in high-throughput microscopy have made possible to collect multiple 3D images over time, generating $$3D+t$$

3
D
+
t

(or 4D) datasets. To extract useful information there is the need to extract spatial and temporal data on the particles that are in the images, but particle tracking and feature extraction need some kind of assistance.

Results

This manuscript introduces our new freely downloadable toolbox, the Visual4DTracker. It is a MATLAB package implementing several useful functionalities to navigate, analyse and proof-read the track of each particle detected in any $$3D+t$$

3
D
+
t

stack. Furthermore, it allows users to proof-read and to evaluate the traces with respect to a given gold standard. The Visual4DTracker toolbox permits the users to visualize and save all the generated results through a user-friendly graphical user interface. This tool has been successfully used in three applicative examples. The first processes synthetic data to show all the software functionalities. The second shows how to process a 4D image stack showing the time-lapse growth of Drosophila cells in an embryo. The third example presents the quantitative analysis of insulin granules in living beta-cells, showing that such particles have two main dynamics that coexist inside the cells.

Conclusions

Visual4DTracker is a software package for MATLAB to visualize, handle and manually track $$3D+t$$

3
D
+
t

stacks of microscopy images containing objects such cells, granules, etc.. With its unique set of functions, it remarkably permits the user to analyze and proof-read 4D data in a friendly 3D fashion. The tool is freely available at https://drive.google.com/drive/folders/19AEn0TqP-2B8Z10kOavEAopTUxsKUV73?usp=sharing
DRPnet: automated particle picking in cryo-electron micrographs using deep regression
Authors: Nguyen Nguyen Phuoc , Ersoy Ilker , Gotberg Jacob , Bunyak Filiz , White Tommi A.
Published: February 2021

Background

Identification and selection of protein particles in cryo-electron micrographs is an important step in single particle analysis. In this study, we developed a deep learning-based particle picking network to automatically detect particle centers from cryoEM micrographs. This is a challenging task due to the nature of cryoEM data, having low signal-to-noise ratios with variable particle sizes, shapes, distributions, grayscale variations as well as other undesirable artifacts.

Results

We propose a double convolutional neural network (CNN) cascade for automated detection of particles in cryo-electron micrographs. This approach, entitled Deep Regression Picker Network or “DRPnet”, is simple but very effective in recognizing different particle sizes, shapes, distributions and grayscale patterns corresponding to 2D views of 3D particles. Particles are detected by the first network, a fully convolutional regression network (FCRN), which maps the particle image to a continuous distance map that acts like a probability density function of particle centers. Particles identified by FCRN are further refined to reduce false particle detections by the second classification CNN. DRPnet’s first CNN pretrained with only a single cryoEM dataset can be used to detect particles from different datasets without retraining. Compared to RELION template-based autopicking, DRPnet results in better particle picking performance with drastically reduced user interactions and processing time. DRPnet also outperforms the state-of-the-art particle picking networks in terms of the supervised detection evaluation metrics recall, precision, and F-measure. To further highlight quality of the picked particle sets, we compute and present additional performance metrics assessing the resulting 3D reconstructions such as number of 2D class averages, efficiency/angular coverage, Rosenthal-Henderson plots and local/global 3D reconstruction resolution.

Conclusion

DRPnet shows greatly improved time-savings to generate an initial particle dataset compared to manual picking, followed by template-based autopicking. Compared to other networks, DRPnet has equivalent or better performance. DRPnet excels on cryoEM datasets that have low contrast or clumped particles. Evaluating other performance metrics, DRPnet is useful for higher resolution 3D reconstructions with decreased particle numbers or unknown symmetry, detecting particles with better angular orientation coverage.
Esmraldi: efficient methods for the fusion of mass spectrometry and magnetic resonance images
Authors: Grélard Florent , Legland David , Fanuel Mathieu , Arnaud Bastien , Foucat Loïc , Rogniaux Hélène
Published: February 2021

Background

Mass spectrometry imaging (MSI) is a family of acquisition techniques producing images of the distribution of molecules in a sample, without any prior tagging of the molecules. This makes it a very interesting technique for exploratory research. However, the images are difficult to analyze because the enclosed data has high dimensionality, and their content does not necessarily reflect the shape of the object of interest. Conversely, magnetic resonance imaging (MRI) scans reflect the anatomy of the tissue. MRI also provides complementary information to MSI, such as the content and distribution of water.

Results

We propose a new workflow to merge the information from 2D MALDI–MSI and MRI images. Our workflow can be applied to large MSI datasets in a limited amount of time. Moreover, the workflow is fully automated and based on deterministic methods which ensures the reproducibility of the results. Our methods were evaluated and compared with state-of-the-art methods. Results show that the images are combined precisely and in a time-efficient manner.

Conclusion

Our workflow reveals molecules which co-localize with water in biological images. It can be applied on any MSI and MRI datasets which satisfy a few conditions: same regions of the shape enclosed in the images and similar intensity distributions.
Intensive care unit discharge: mind the gap!
Authors: Deana Cristian , Sermann Giovanni , De Monte Amato
Published: February 2021

Mortality after intensive care discharge is a hot topic in critical care medicine. Many factors probably play a role: patient’s comorbidities and severity of the disease may have great impact on mortality. However it should be taken into account also the level of care that characterizes the ward in which the patient is discharged to. A soft transition from intensive care units to the other hospital wards is desirable to avoid the traumatic step that the fragile post-ICU patient has to face with.
Effects of an individualized analgesia protocol on the need for medical interventions after adenotonsillectomy in children: a randomized controlled trial
Authors: Guo Jian , Zhuang Peijun , Liu Kun , Wan Yuanyuan , Wang Xuan
Published: February 2021

Background

It has been proposed that the dose of rescue opioids should be individually titrated to the severity of obstructive sleep apnea after adenotonsillectomy. However, a sleep study is not always available before adenotonsillectomy. This randomized, controlled and blinded trial evaluated a strategy of pain control individualized to the results of a fentanyl test, rather than the results of polysomnography, in children after adenotonsillectomy.

Methods

A total of 280 children (3–10 years old) undergoing elective adenotonsillectomy were randomized into an individualized protocol (IP) group or a conservative protocol (CP) group. All patients received a fentanyl test before extubation. Pain was assessed every 10 min in the recovery room, and rescue morphine was given when the Children’s Hospital of Eastern Ontario Pain Scale (CHEOPS) score was > 6. In the IP group, the dose of rescue morphine was individualized to the results of the fentanyl test (10 μg/kg in the case of a positive result and 50 μg/kg in the case of a negative result). In the CP group, the dose was fixed (25 μg/kg). The primary outcome was the percentage of patients requiring at least one medical intervention. The secondary outcome was the median duration of CHEOPS scores > 6.

Results

Fewer patients in the IP group than in the CP group required medical interventions [11.9% (16/134) vs 22.3% (29/130),P = 0.025]. The median duration of CHEOPS scores > 6 was shorter in the IP group than in the CP group [20 (95% CI: 17 to 23) min vs 30 (95% CI: 28 to 32) min,P <  0.001].

Conclusions

Compared with a conservative dosing approach, this individualized protocol may improve analgesia without a significant increase in respiratory adverse events.

Trial registration

ClinicalTrials.govNCT02990910, registered on 13/12/2016.
Development of an eHealth programme for self-management of persistent physical symptoms: a qualitative study on user needs in general practice
Authors: Rask Mette Trøllund , Jakobsen Pernille Ravn , Clemensen Jane , Rosendal Marianne , Frostholm Lisbeth
Published: February 2021

Background

Persistent physical symptoms (PPS) are estimated to be present in 17% of patients in general practice. Hence, general practitioners (GPs) play a key role in both the diagnostic assessment and the management of PPS. However, research indicates a need to improve their strategies to support self-help in patients, and eHealth tools may serve as an opportunity. This study aimed to explore patients’ and GPs’ needs related to self-management of PPS. The study was designed to inform the future development of eHealth interventions in this field.

Methods

This qualitative study was based on 20 semi-structured interviews (6 GPs and 14 patients with PPS). Interviews were audiotaped, transcribed verbatim and analysed through a five-step thematic analysis approach. First, we conducted an inductive analysis to identify and explore emerging subthemes. Second, using a deductive mapping strategy, we categorised the derived subthemes according to the COM-B behaviour change model and its three domains: capability, opportunity and motivation.

Results

We identified eleven subthemes in the patient interviews and seven subthemes in the GP interviews. Several unmet needs emerged. First, we identified a need to consider PPS early in the illness trajectory by taking a bio-psycho-social approach. Second, both patients and GPs need better skills to manage uncertainty. Third, hope is important for the patients. Fourth, patients need guidance from their GP in how to self-manage their PPS.

Conclusions

This study provides important insight into key issues and needs related to capability, opportunity and motivation that should be addressed in the design of future eHealth self-management interventions targeting patients with PPS in general practice in order to support and improve care.
Blood angiopoietin-2 predicts liver angiogenesis and fibrosis in hepatitis C patients
Authors: Osawa Yosuke , Yoshio Sachiyo , Aoki Yoshihiko , Korenaga Masaaki , Imamura Masatoshi , Oide Takashi , Okawara Miku , Kawai Hironari , Tsutsui Yuriko , Yoshida Yuichi , Yoshikawa Shiori , Mori Taizo , Yamazoe Taiji , Kanto Tatsuya
Published: February 2021

Background

Pathological angiogenesis is involved in the development of hepatocellular carcinoma. In patients with chronic hepatitis C (CHC), the level of angiogenic factor angiopoietin (ANGP)-2 is reported to be increased in the blood, correlating with fibrosis. In this study, we aimed to clarify whether blood ANGP-2 is useful as a biomarker for liver angiogenesis and fibrosis in CHC patients and to further reveal the relationship between such pathology in a carbon tetrachloride (CCl4)-treated liver fibrosis mouse model.

Methods

Plasma levels of ANGP-2, expression of a liver sinusoidal endothelial cell (LSEC) marker (CD31), collagen deposition (Sirius Red staining) in the liver, clinical fibrosis markers (Mac-2 binding protein glycosylation isomer, virtual touch quantification, and liver stiffness measurement), and liver function (albumin bilirubin score) were examined in CHC patients. To determine the effects of an anti-angiogenic agent on liver fibrosis in vivo, sorafenib was administered to the CCl4-treated mice (BALB/c male).

Results

The plasma levels of ANGP-2 were increased in CHC patients compared to healthy volunteers and decreased by the eradication of hepatitis C with direct-acting antivirals. In addition, plasma ANGP-2 levels were correlated with CD31 expression, collagen deposition, clinical fibrosis markers, and liver function. Sorafenib inhibited liver angiogenesis and fibrosis in the CCl4-treated mice and was accompanied by decreased ANGP-2 expression in LSECs.

Conclusions

ANGP-2 may serve as a useful biomarker for liver angiogenesis and fibrosis in CHC patients. In addition, angiogenesis and fibrosis may be closely related.
Multiple antimicrobial resistance and outcomes among hospitalized patients with complicated urinary tract infections in the US, 2013–2018: a retrospective cohort study
Authors: Zilberberg Marya D. , Nathanson Brian H. , Sulham Kate , Shorr Andrew F.
Published: February 2021

Background

Complicated urinary tract infection (cUTI) is common among hospitalized patients. Though carbapenems are an effective treatment in the face of rising resistance, overuse drives carbapenem resistance (CR). We hypothesized that resistance to routinely used antimicrobials is common, and, despite frequent use of carbapenems, associated with an increased risk of inappropriate empiric treatment (IET), which in turn worsens clinical outcomes.

Methods

We conducted a retrospective cohort study of patients hospitalized with a culture-positive non-CR cUTI. Triple resistance (TR) was defined as resistance to > 3 of the following: 3rd generation cephalosporins, fluoroquinolones, trimethoprim-sulfamethoxazole, fosfomycin, and nitrofurantoin. Multivariable models quantified the impact of TR and inappropriate empiric therapy (IET) on mortality, hospital LOS, and costs.

Results

Among 23,331 patients with cUTI, 3040 (13.0%) had a TR pathogen. Compared to patients with non-TR, those with TR were more likely male (57.6% vs. 47.7%, p < 0.001), black (17.9% vs. 13.6%, p < 0.001), and in the South (46.3% vs. 41.5%, p < 0.001). Patients with TR had higher chronic (median [IQR] Charlson score 3 [2, 4] vs. 2 [1, 4], p < 0.001) and acute (mechanical ventilation 7.0% vs. 5.0%, p < 0.001; ICU admission 22.3% vs. 18.6%, p < 0.001) disease burden. Despite greater prevalence of empiric carbapenem exposure (43.3% vs. 16.2%, p < 0.001), patient with TR were also more likely to receive IET (19.6% vs. 5.4%, p < 0.001) than those with non-TR. Although mortality was similar between groups, TR added 0.38 (95% CI 0.18, 0.49) days to LOS, and $754 (95% CI $406, $1103) to hospital costs. Both TR and IET impacted the outcomes among cUTI patients whose UTI was not catheter-associated (CAUTI), but had no effect on outcomes in CAUTI.

Conclusions

TR occurs in 1 in 8 patients hospitalized with cUTI. It is associated with an increase in the risk of IET exposure, as well as a modest attributable prolongation of LOS and increase in total costs, particularly in the setting of non-CAUTI.
The monitoring of vancomycin: a systematic review and meta-analyses of area under the concentration-time curve-guided dosing and trough-guided dosing
Authors: Tsutsuura Moeko , Moriyama Hiromu , Kojima Nana , Mizukami Yuki , Tashiro Sho , Osa Sumika , Enoki Yuki , Taguchi Kazuaki , Oda Kazutaka , Fujii Satoshi , Takahashi Yoshiko , Hamada Yukihiro , Kimura Toshimi , Takesue Yoshio , Matsumoto Kazuaki
Published: February 2021

Background

This systematic review and meta-analysis explored the relationship between vancomycin (VCM) monitoring strategies and VCM effectiveness and safety.

Methods

We conducted our analysis using the MEDLINE, Web of Sciences, and Cochrane Register of Controlled Trials electronic databases searched on August 9, 2020. We calculated odds ratios (ORs) and 95% confidence intervals (CIs).

Results

Adult patients with methicillin-resistant Staphylococcus aureus (MRSA) bacteraemia with VCM trough concentrations ≥15 μg/mL had significantly lower treatment failure rates (OR 0.63, 95% CI 0.47–0.85). The incidence of acute kidney injury (AKI) increased with increased trough concentrations and was significantly higher for trough concentrations ≥20 μg/mL compared to those at 15–20 μg/mL (OR 2.39, 95% CI 1.78–3.20). Analysis of the target area under the curve/minimum inhibitory concentration ratios (AUC/MIC) showed significantly lower treatment failure rates for high AUC/MIC (cut-off 400 ± 15%) (OR 0.28, 95% CI 0.18–0.45). The safety analysis revealed that high AUC value (cut-off 600 ± 15%) significantly increased the risk of AKI (OR 2.10, 95% CI 1.13–3.89). Our meta-analysis of differences in monitoring strategies included four studies. The incidence of AKI tended to be lower in AUC-guided monitoring than in trough-guided monitoring (OR 0.54, 95% CI 0.28–1.01); however, it was not significant in the analysis of mortality.

Conclusions

We identified VCM trough concentrations and AUC values that correlated with effectiveness and safety. Furthermore, compared to trough-guided monitoring, AUC-guided monitoring showed potential for decreasing nephrotoxicity.
Validity and reliability of the Portuguese version of the modified Migraine Disability Assessment
Authors: Ferreira Pedro L. , Luzeiro Isabel , Lopes Margarida , Jorge André , Silva Bruno , Ferreira Lara
Published: February 2021

Background

Migraine Disability Assessment Scale (MIDAS) is a useful tool to measure headache-related disability. Modified MIDAS with 4-week recall period reduces recall bias and improves accuracy of the results. This study aimed at validating mMIDAS in Portuguese.

Methods

Studied population consisted of adult migraine patients attending a headache outpatient clinic. Reliability was assessed by internal consistency and reproducibility in a 3-week test-retest. Content validity was evaluated by two expert panels. Construct validity was tested by comparing mMIDAS-P index in socioeconomic and clinical patient groups and scale unidimensionality was evidenced by factor analysis. Criterion validity was tested using EQ-5D-5L and HADS.

Results

Ninety-two patients, 88% female, mean age of 44 years, participated. They had, in average, 9.7 headache days in previous month, pain averaging 7.5/10. About 69.9% were on a migraine prophylactic treatment, and 42.4% had severe disability; 29.4 and 13.0% showed, respectively, moderate/severe anxiety and depression. Content validity showed that mMIDAS-P is simple and clinically useful. It did not show to be determined by patient’s sociodemographic characteristics and it was correlated with depression scale and EQ-5D-5L. Test-retest demonstrated high reproductive reliability and good internal consistency.

Conclusion

mMIDAS-P is valid and reliable. We strongly recommend it for clinical and research use.
The diagnostic challenge of coexistent sarcoidosis and thyroid cancer – a retrospective study
Authors: Wenter Vera , Albert Nathalie L. , Ahmaddy Freba , Unterrainer Marcus , Hornung Julia , Ilhan Harun , Bartenstein Peter , Spitzweg Christine , Kneidinger Nikolaus , Todica Andrei
Published: February 2021

Background

Sarcoid lesions may mimic metastatic disease or recurrence in thyroid cancer (TC) patients as both diseases may affect the lungs and lymph nodes. We present the first study to systematically evaluate the clinical course of patients with (TC) after adjuvant radioactive iodine therapy (RIT) and concomitant sarcoidosis of the lung or the lymph nodes.

Methods

We screened 3285 patients and retrospectively identified 16 patients with TC (11 papillary thyroid cancer (PTC), 3 follicular thyroid cancer (FTC), 1 oncocytic PTC, 1 oncocytic FTC) and coexisting sarcoidosis of the lung and/or the lymph nodes treated at our institute. All patients had undergone thyroidectomy and initial adjuvant RIT. Challenges in diagnosing and the management of these patients were evaluated during long term follow-up (median 4.9 years (0.8–15.0 years)).

Results

Median age at first diagnosis of TC was 50.1 years (33.0–71.5 years) and of sarcoidosis 39.4 years (18.0–63.9 years). During follow-up, physicians were able to differentiate between SA and persistent or recurrent TC in 10 of 16 patients (63%). Diagnosis was complicated by initial negative thyroglobulin (Tg), positive Tg antibodies and non-specific imaging findings. Histopathology can reliably distinguish between SA and TC in patients with one suspicious lesion.

Conclusion

Physicians should be aware of the rare coexistence of sarcoidosis and TC. Lymphadenopathy and pulmonary lesions could be metastases, sarcoidosis or even a mix of both. Therefore, this rare patient group should receive a thorough work up including histopathological clarification and, if necessary, separately for each lesion.