Helper Function Research Articles

Brainchop: Providing an Edge Ecosystem for Deployment of Neuroimaging Artificial Intelligence Models.

Deep learning has proven highly effective in various medical imaging scenarios, yet the lack of an efficient distribution platform hinders developers from sharing models with end-users. Here, we describe brainchop, a fully functional web application that allows users to apply deep learning models developed with Python to local neuroimaging data from within their browser. While training artificial intelligence models is computationally expensive, applying existing models to neuroimaging data can be very fast; brainchop harnesses the end user's graphics card such that brain extraction, tissue segmentation, and regional parcellation require only seconds and avoids privacy issues that impact cloud-based solutions. The integrated visualization allows users to validate the inferences, and includes tools to annotate and edit the resulting segmentations. Our pure JavaScript implementation includes optimized helper functions for conforming volumes and filtering connected components with minimal dependencies. Brainchop provides a simple mechanism for distributing models for additional image processing tasks, including registration and identification of abnormal tissue, including tumors, lesions and hyperintensities. We discuss considerations for other AI model developers to leverage this open-source resource.

SSN2: The next generation of spatial stream network modeling in R.

The SSN2 R package provides tools for spatial statistical modeling, parameter estimation, and prediction on stream (river) networks. SSN2 is the successor to the SSN R package (Ver Hoef, Peterson, Clifford, & Shah, 2014), which was archived alongside broader changes in the R-spatial ecosystem (Nowosad, 2023) that included 1) the retirement of rgdal (Bivand, Keitt, & Rowlingson, 2021), rgeos (Bivand & Rundel, 2020), and maptools (Bivand & Lewin-Koh, 2021) and 2) the lack of active development of sp (Bivand, Pebesma, & Gómez-Rubio, 2013). SSN2 maintains compatibility with the input data file structures used by the SSN R package but leverages modern R-spatial tools like sf (Pebesma, 2018). SSN2 also provides many useful features that were not available in the SSN R package, including new modeling and helper functions, enhanced fitting algorithms, and simplified syntax consistent with other R generic functions.

RACK1 enhances STAT3 stability and promotes T follicular helper cell development and function during blood-stage Plasmodium infection in mice.

CD4+ T cells are central mediators of protective immunity to blood-stage malaria, particularly for their capacity in orchestrating germinal center reaction and generating parasite-specific high-affinity antibodies. T follicular helper (Tfh) cells are predominant CD4+ effector T cell subset implicated in these processes, yet the factors and detailed mechanisms that assist Tfh cell development and function during Plasmodium infection are largely undefined. Here we provide evidence that receptor for activated C kinase 1 (RACK1), an adaptor protein of various intracellular signals, is not only important for CD4+ T cell expansion as previously implied but also plays a prominent role in Tfh cell differentiation and function during blood-stage Plasmodium yoelii 17XNL infection. Consequently, RACK1 in CD4+ T cells contributes significantly to germinal center formation, parasite-specific IgG production, and host resistance to the infection. Mechanistic exploration detects specific interaction of RACK1 with STAT3 in P. yoelii 17XNL-responsive CD4+ T cells, ablation of RACK1 leads to defective STAT3 phosphorylation, accompanied by substantially lower amount of STAT3 protein in CD4+ T cells, whereas retroviral overexpression of RACK1 or STAT3 in RACK1-deficient CD4+ T cells greatly restores STAT3 activity and Bcl-6 expression under the Tfh polarization condition. Further analyses suggest RACK1 positively regulates STAT3 stability by inhibiting the ubiquitin-proteasomal degradation process, thus promoting optimal STAT3 activity and Bcl-6 induction during Tfh cell differentiation. These findings uncover a novel mechanism by which RACK1 participates in posttranslational regulation of STAT3, Tfh cell differentiation, and subsequent development of anti-Plasmodium humoral immunity.

Mitoclone2: an R package for elucidating clonal structure in single-cell RNA-sequencing data using mitochondrial variants.

Clonal cell population dynamics play a critical role in both disease and development. Due to high mitochondrial mutation rates under both healthy and diseased conditions, mitochondrial genomic variability is a particularly useful resource in facilitating the identification of clonal population structure. Here we present mitoClone2, an all-inclusive R package allowing for the identification of clonal populations through integration of mitochondrial heteroplasmic variants discovered from single-cell sequencing experiments. Our package streamlines the investigation of this phenomenon by providing: built-in compatibility with commonly used tools for the delineation of clonal structure, the ability to directly use multiplexed BAM files as input, annotations for both human and mouse mitochondrial genomes, and helper functions for calling, filtering, clustering, and visualizing variants.

Molecular design of controllable recombinant adeno-associated virus (AAV) expression systems for enhanced vector production.

Recombinant adeno-associated virus (rAAV) is the leading vector for the delivery of gene therapies. However, low viral genome (VG) titers are common and the proportion of "full" capsids containing the therapeutic gene payload can be highly variable. The coordinated molecular design of plasmids encoding viral components and Helper functions remains a major challenge for rAAV manufacturing. Here we present the design of improved Rep/Cap and Helper plasmids for rAAV2/8 production, (i) a Rep/Cap expression vector harboring independently controllable rep and cap genes and (ii) an improved Helper plasmid harboring E4 gene deletion variants. First, an optimized Rep/Cap vector utilized a truncated p5 promoter, a p5 cis-regulatory element at the 3' end in combination with a heterologous promoter to drive Cap expression and an additional copy of the rep52/40 gene to overexpress short Rep proteins. We demonstrate that Rep78 is essential for efficient rAAV2/8 production in HEK293 cells, and a higher ratio of short Rep to long Rep proteins enhances genome packaging. Second, we identified regulators and open reading frames within the Helper plasmid that contribute to increased rAAV2/8 production. While L4-33k/22k is integral to optimal production, the use of E4orf6-6/7 subset significantly enhanced VG titer. Together, an optimal combination of engineered Rep/Cap and Helper plasmid variants increased VG titer by 3.1-fold. This study demonstrates that configuring and controlling the expression of the different AAV genetic elements contributes toward high rAAV production and product quality (full/empty capsid ratio).

Cytotoxic Tph subset with low B-cell helper functions and its involvement in systemic lupus erythematosus

T peripheral helper (Tph) cells are thought to contribute to extra-follicular B cell activation and play a pathogenic role in autoimmune diseases. However, the role of Tph subsets is not fully elucidated. Here, we investigate the immunological functions of Tph subsets and their involvement in systemic lupus erythematosus (SLE). We have defined four Tph subsets (Tph1: CXCR3+CCR6−, Tph2: CXCR3−CCR6−, Tph17: CXCR3−CCR6+, and Tph1-17: CXCR3+CCR6+) and performed RNA sequencing after cell sorting. Tph1 and Tph17 subsets express substantial levels of IL21, indicating B cell helper functions. However, Tph2 and Tph1-17 subsets express low IL21. Interestingly, we have found Tph2 subset express high levels of CX3CR1, GZMB, PRF1, GLNY, S1PR5, TBX21, EOMES, ZNF863, and RUNX3, indicating a feature of CD4+ cytotoxic T lymphocytes. In SLE patients, the frequency of Tph1 and Tph2 subsets are significantly increased and positively correlated with SLE disease activity indexes. Tph1 cells expansion has been observed in patients with cutaneous and musculoskeletal manifestations. On the other hand, Tph2 cell expansion has been found in patients with lupus nephritis in addition to the above manifestations. Our findings imply that Tph1 and Tph2 subsets exert distinct immunological functions and are contributed to the complexity of clinical manifestations in SLE.

Effects of autoimmune abnormalities on fertility and placental morphology in mice

Autoimmune diseases (AIDs) alter the placental immune environment leading to fetal loss. This study investigated the effects of AIDs on pregnancy and the placenta in AID-prone MRL/MpJ-Faslpr/lpr mice and wild-type MRL/MpJ, which were mated with male MRL/MpJ and MRL/MpJ-Faslpr/lpr at five months and defined as moLpr and moMpJ, respectively. AID indices (spleen weight and serum autoantibody levels) and fertility status (number and size of fetuses, morphology, and comprehensive gene expression of placentas) were evaluated on gestational day 15.5. Both strains showed equivalent fertility, but moLpr showed lighter placentas and fetuses than moMpJ, and decreased fertility with AID severity. moLpr placentas had a higher number of T cells, higher expression of genes associated with T helper 2 and T follicular helper functions, and altered expression of genes (Krt15, Slc7a3, Sprr2a3) that significantly regulate pregnancy or immunity. The gene expression of T cell migration-associated chemokines (Ccl5, Cxcl9) was significantly increased in moLpr placentas, and CCL5 and CXCL9 were detected in moLpr placentas, particularly in T cells and placenta-component cells, respectively. Thus, AID altered placental morphofunction and fertility in mice; however, fertility was maintained at the examined time points. This study enhances our understanding of placental alterations and gestational risk due to AIDs.

Code and Data Repository for Quadratic Optimization Models for Balancing Preferential Access and Fairness: Formulations and Optimality Conditions

This directory provides code used for the computations associated with the quadratic optimization models in the context of undesirable facility location problems for the abovementioned article. src contains scripts for the optimization of the MIQP model and visualization of the results: model.py contains functions for building and optimizing the MIQP model, while greedy_heuristic.py applies a greedy heuristic to achieve a feasible solution. plotting.py and results.py contain functions tasked with visualizing results through various different plots as well as excel tables. They also contain superordinate functions that create the corresponding results first by running functions from model.py and/or greedy_heuristic.py before creating the corresponding visualization. These functions are called by functions in tables_and_figures.py to create the exact same figures and tables that are included in the paper from scratch. Lastly, utils.py contains helper and utility functions. catchment_population presents an efficient algorithm to compute the "catchment population" of each recycling center that was used to estimate the capacities. An implementation of this algorithm is contained in catchment_population.py. Further, this directory contains two corresponding input data files in csv format: bavaria_grid_population.csv is a file containing the latitude and longitude of the centroid of each 100m x 100m grid in Bavaria as well as the residing population. rc_locations.csv contains the latitude and longitude of each recycling center in Bavaria. data contains the two input data files that are used in the MIQP model: users_and_facilities.xlsx contains all ZIP codes and recycling centers related data like the population, centroid and regional spatial type (rural/urban) of each ZIP code as well as the capacity, centroid and regional spatial type of each recycling center. travel_dict.json.pbz2 is a compressed json file that contains the travel probabilities from each ZIP code to each recycling center. results contains the excel tables and figures visualizing the results that are included in the paper.

Age-associated CD4+ T cells with B cell-promoting functions are regulated by ZEB2 in autoimmunity.

Aging is a significant risk factor for autoimmunity, and many autoimmune diseases tend to onset during adulthood. We conducted an extensive analysis of CD4+ T cell subsets from 354 patients with autoimmune disease and healthy controls via flow cytometry and bulk RNA sequencing. As a result, we identified a distinct CXCR3midCD4+ effector memory T cell subset that expands with age, which we designated "age-associated T helper (THA) cells." THA cells exhibited both a cytotoxic phenotype and B cell helper functions, and these features were regulated by the transcription factor ZEB2. Consistent with the highly skewed T cell receptor usage of THA cells, gene expression in THA cells from patients with systemic lupus erythematosus reflected disease activity and was affected by treatment with a calcineurin inhibitor. Moreover, analysis of single-cell RNA sequencing data revealed that THA cells infiltrate damaged organs in patients with autoimmune diseases. Together, our characterization of THA cells may facilitate improved understanding of the relationship between aging and autoimmune diseases.

Human Bocavirus 1 NP1 acts as an ssDNA-binding protein to help AAV2 DNA replication and cooperates with RPA to regulate AAV2 capsid expression.

Adeno-associated virus (AAV) requires co-infection with helper virus for efficient replication. We previously reported that Human Bocavirus 1 (HBoV1) genes, including NP1, NS2, and BocaSR, were critical for AAV2 replication. Here, we first demonstrate the essential roles of the NP1 protein in AAV2 DNA replication and protein expression. We show that NP1 binds to single-strand DNA (ssDNA) at least 30 nucleotides (nt) in length in a sequence-independent manner. Furthermore, NP1 colocalized with the BrdU-labeled AAV2 DNA replication center, and the loss of the ssDNA-binding ability of NP1 by site-directed mutation completely abolished AAV2 DNA replication. We used affinity-tagged NP1 protein to identify host cellular proteins associated with NP1 in cells cotransfected with the HBoV1 helper genes and AAV2 duplex genome. Of the identified proteins, we demonstrate that NP1 directly binds to the DBD-F domain of the RPA70 subunit with a high affinity through the residues 101-121. By reconstituting the heterotrimer protein RPA in vitro using gel filtration, we demonstrate that NP1 physically associates with RPA to form a heterologous complex characterized by typical fast-on/fast-off kinetics. Following a dominant-negative strategy, we found that NP1-RPA complex mainly plays a role in expressing AAV2 capsid protein by enhancing the transcriptional activity of the p40 promoter. Our study revealed a novel mechanism by which HBoV1 NP1 protein supports AAV2 DNA replication and capsid protein expression through its ssDNA-binding ability and direct interaction with RPA, respectively.IMPORTANCERecombinant adeno-associated virus (rAAV) vectors have been extensively used in clinical gene therapy strategies. However, a limitation of these gene therapy strategies is the efficient production of the required vectors, as AAV alone is replication-deficient in the host cells. HBoV1 provides the simplest AAV2 helper genes consisting of NP1, NS2, and BocaSR. An important question regarding the helper function of HBoV1 is whether it provides any direct function that supports AAV2 DNA replication and protein expression. Also of interest is how HBoV1 interplays with potential host factors to constitute a permissive environment for AAV2 replication. Our studies revealed that the multifunctional protein NP1 plays important roles in AAV2 DNA replication via its sequence-independent ssDNA-binding ability and in regulating AAV2 capsid protein expression by physically interacting with host protein RPA. Our findings present theoretical guidance for the future application of the HBoV1 helper genes in the rAAV vector production.

DendroX: multi-level multi-cluster selection in dendrograms

BackgroundCluster heatmaps are widely used in biology and other fields to uncover clustering patterns in data matrices. Most cluster heatmap packages provide utility functions to divide the dendrograms at a certain level to obtain clusters, but it is often difficult to locate the appropriate cut in the dendrogram to obtain the clusters seen in the heatmap or computed by a statistical method. Multiple cuts are required if the clusters locate at different levels in the dendrogram.ResultsWe developed DendroX, a web app that provides interactive visualization of a dendrogram where users can divide the dendrogram at any level and in any number of clusters and pass the labels of the identified clusters for functional analysis. Helper functions are provided to extract linkage matrices from cluster heatmap objects in R or Python to serve as input to the app. A graphic user interface was also developed to help prepare input files for DendroX from data matrices stored in delimited text files. The app is scalable and has been tested on dendrograms with tens of thousands of leaf nodes. As a case study, we clustered the gene expression signatures of 297 bioactive chemical compounds in the LINCS L1000 dataset and visualized them in DendroX. Seventeen biologically meaningful clusters were identified based on the structure of the dendrogram and the expression patterns in the heatmap. We found that one of the clusters consisting of mostly naturally occurring compounds is not previously reported and has its members sharing broad anticancer, anti-inflammatory and antioxidant activities.ConclusionsDendroX solves the problem of matching visually and computationally determined clusters in a cluster heatmap and helps users navigate among different parts of a dendrogram. The identification of a cluster of naturally occurring compounds with shared bioactivities implicates a convergence of biological effects through divergent mechanisms.

Multi-omics kinetic analysis of recombinant adeno-associated virus production by plasmid transfection of HEK293 cells.

Recombinant adeno-associated virus (rAAV) is among the most commonly used vectors for gene therapy. It is commonly produced by transfection of HEK293 cells with three plasmids each containing the vector genome including gene of interest (GOI), helper functions, and rep and cap genes for genome replication and capsid formation. To meet the potential clinical needs, the productivity of the production system needs to be enhanced. A better process characterization of the production system will further advance our insights into ways to enhance productivity. Here, we employed transcriptomic analysis to quantify the dynamics of different isoforms of viral transcripts and to assess the shift of cellular physiology, and deployed targeted proteomic analysis for absolute quantification of viral proteins and tandem mass tags (TMTs) for assessing cellular responses at the protein level. Functional analysis at transcriptome and proteome levels identified defense and immune response, unfolded protein response, p53 signaling as enriched. The small molecule additive intervention study based on functional analysis showed the potential of such omics-guided productivity enhancement. Together, multi-omics analysis advanced understanding of rAAV production and provided insight into enhancing rAAV production by plasmid transfection.

The Role of Sales Accounting Information System in PT. Petro Gasindo Intiniaga

The role of sales accounting information systems in a company is very important Importantly, having a sales accounting information system can maximize profits obtained, achieve sales volume, and support sales growth. So that If the company achieves maximum sales levels, the company needs to pay attention sales system in the company. This research focuses on (1) How the sales accounting information system is implemented at PT. Petro Gasindo Intiniaga, (2) What is the internal control system at PT. Petro Gasindo Intiniaga (3) What is the role of the sales information system at PT. Petro Gasindo Intiniaga to improve the internal control system. This research uses a qualitative approach with a descriptive research type. The data sources used are Primary Data and Secondary Data. Research procedure The data used is field search (a) interviews (b) documentation (c) Observation. To check the validity of the data findings, three triangulations were carried out (1) source triangulation (2) technique triangulation (3) time triangulation. The result of this research is an internal control system in Information Systems Sales Accounting at PT. Petro Gasindo Intiniaga is not doing well. In the The company still has several weaknesses, including; (1) There still exists a dual function in the sales system, between the warehouse helper function and admin functions which are only carried out by the warehouse department. (2) The Authorization System for Approval for Delivery of goods is still authorized by the production and warehouse departments. (3) Deep sales system in which the cash received is not fully deposited into the bank on the date the transaction takes place.

CD39 expression defines exhausted CD4+ Tcells associatedwith poor survival and immune evasion in human gastric cancer.

CD4+ Tcell helper and regulatory function in human cancers has been well characterised. However, the definition of tumor-infiltrating CD4+ Tcell exhaustion and how it contributes to the immune response and disease progression in human gastriccancer (GC) remain largely unknown. A total of 128 GC patients were enrolled in the study. The expression of CD39 and PD-1 on CD4+ Tcells in the different samples was analysed by flow cytometry. GC-infiltrating CD4+ Tcell subpopulations based on CD39 expression were phenotypically and functionally assessed. The role of CD39 in the immune response of GC-infiltrating Tcells was investigated by inhibiting CD39 enzymatic activity. In comparison with CD4+ Tcells from the non-tumor tissues, significantly more GC-infiltrating CD4+ Tcells expressed CD39. Most GC-infiltrating CD39+CD4+ Tcells exhibited CD45RA-CCR7- effector-memory phenotype expressing more exhaustion-associated inhibitory molecules and transcription factors and produced less TNF-α, IFN-γ and cytolytic molecules than their CD39-CD4+ counterparts. Moreover, ex vivo inhibition of CD39 enzymatic activity enhanced their functional potential reflected by TNF-α and IFN-γ production. Finally, increased percentages of GC-infiltrating CD39+CD4+ Tcells were positively associated with disease progression and patients' poorer overall survival. Our study demonstrates that CD39 expression defines GC-infiltrating CD4+ Tcell exhaustion and their immunosuppressive function. Targeting CD39 may be a promising therapeutic strategy for treating GC patients.

Heterologous booster vaccination enhances antibody responses to SARS-CoV-2 by improving Tfh function and increasing B-cell clonotype SHM frequency.

Heterologous prime-boost has broken the protective immune response bottleneck of the COVID-19 vaccines. however, the underlying mechanisms have not been fully elucidated. Here, we investigated antibody responses and explored the response of germinal center (GC) to priming with inactivated vaccines and boosting with heterologous adenoviral-vectored vaccines or homologous inactivated vaccines in mice. Antibody responses were dramatically enhanced by both boosting regimens. Heterologous immunization induced more robust GC activation, characterized by increased Tfh cell populations and enhanced helper function. Additionally, increased B-cell activation and antibody production were observed in a heterologous regimen. Libra-seq was used to compare the differences of S1-, S2- and NTD-specific B cells between homologous and heterologous vaccination, respectively. S2-specific CD19+ B cells presented increased somatic hypermutations (SHMs), which were mainly enriched in plasma cells. Moreover, a heterologous booster dose promoted the clonal expansion of B cells specific to S2 and NTD regions. In conclusion, the functional role of Tfh and B cells following SARS-CoV-2 heterologous vaccination may be important for modulating antibody responses. These findings provide new insights for the development of SARS-CoV-2 vaccines that induce more robust antibody response.

AAV production in stable packaging cells requires expression of adenovirus 22/33K protein to allow episomal amplification of integrated rep/cap genes

Efficient manufacture of recombinant adeno-associated virus (rAAV) vectors for gene therapy remains challenging. Packaging cell lines containing stable integration of the AAV rep/cap genes have been explored, however rAAV production needs to be induced using wild-type adenoviruses to promote episomal amplification of the integrated rep/cap genes by mobilizing a cis-acting replication element (CARE). The adenovirus proteins responsible are not fully defined, and using adenovirus during rAAV manufacture leads to contamination of the rAAV preparation. ‘TESSA’ is a helper adenovirus with a self-repressing Major Late Promoter (MLP). Its helper functions enable efficient rAAV manufacture when the rep and cap genes are provided in trans but is unable to support rAAV production from stable packaging cells. Using rAAV-packaging cell line HeLaRC32, we show that expression of the adenovirus L4 22/33K unit is essential for rep/cap amplification but the proteins are titrated away by binding to replicating adenovirus genomes. siRNA-knockdown of the adenovirus DNA polymerase or the use of a thermosensitive TESSA mutant decreased adenovirus genome replication whilst maintaining MLP repression, thereby recovering rep/cap amplification and efficient rAAV manufacture. Our findings have direct implications for engineering more efficient adenovirus helpers and superior rAAV packaging/producer cells.

Abstract B003: Communication is key: Understanding the modulating effect of immune cell crosstalk on T cell therapy function

Abstract T cell therapies have shown great promise, especially for the treatment of hematological malignancies, but this success has not been translatable for most solid tumors. Therefore, efforts have been put towards improving their efficacy, including optimization of the subset composition of the T cell product and development of strategies to overcome immunosuppression by the tumor microenvironment. Here, we combined advanced immune-organoid co-cultures and live 3D imaging to understand how different T cell subsets and immune cells in the tumor microenvironment can affect T cell therapy efficacy. We found that CD4+ and CD8+ subsets of therapeutic T cells cooperate to boost anti-tumor function, while tumor-associated macrophages (TAMs) have an opposing effect and reduce tumor-targeting by engineered T cells. Through our dynamic imaging-readout (Dekkers, Alieva Nature Biotech. 2022), we observed that a high level of interactions are involved in these modulating events, indicating direct crosstalk between the different cell types. To further investigate the impact of these interactions, we developed a 3D interactome mapping tool which enables us to profile all occurring cell-cell interactions in our 3D imaging datasets and determine their functional outcome. With this tool we identified a specific interaction between CD4+ and CD8+ T cells, where a subset of CD4+ T cells acquired a potent helper function over time by first engaging with tumor organoids and subsequently inducing CD8+ T cell target localization via direct contact. Multi-omics integration with single cell transcriptomic data allowed us to study the underlying molecular pathways involved in this interaction, leading to the identification of a crucial binding partner involved in this cooperative tumor-targeting behavior. Using the same strategy, we are currently investigating how interactions between TAMs and therapeutic T cells modulate their tumor-targeting activity. Together, these data highlight how cellular interactions can play an important role in engineered T cell function and we are currently exploring strategies to target these axes of immune cell communication to maximize therapy efficacy. Citation Format: Amber K.L. Wezenaar, Celina Honhoff, Helena van Eck, Elena Jiménez Curiel, Annelisa M Cornel, Femke C.A. Ringnalda, Tineke Aarts-Riemens, Jürgen Kuball, Zsolt Sebestyen, Hans Clevers, Stefan Nierkens, Johanna F. Dekkers, Ellen J. Wehrens, Maria Alieva, Anne C. Rios. Communication is key: Understanding the modulating effect of immune cell crosstalk on T cell therapy function [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr B003.

A forskolin-mediated increase in cAMP promotes T helper cell differentiation into the Th1 and Th2 subsets rather than into the Th17 subset

The adenylyl cyclase (AC) signaling pathway is suggested to be a key regulator of immune system functions. However, specific effects of cyclic adenosine monophosphate (cAMP) on T helper (Th) cell differentiation and functions are unclear. The involvement of cAMP in the Th cell differentiation program, in particular the development of Th1, Th2, and Th17 subsets, was evaluated employing forskolin (FSK), a labdane diterpene well known as an AC activator. FSK mediated an elevation in Th1-specific markers reinforcing the Th1 cell phenotype. The Th2 differentiation was supported by FSK, though cell metabolism was negatively affected. In contrast, the Th17 immunophenotype was severely suppressed leading to the highly specific upregulation of CXCL13. The causality between FSK-elicited cAMP production and the observed reinforcement of Th2 differentiation was established by using AC inhibitor 2′,5′-dideoxyadenosine, which reverted the FSK effects. Overall, an FSK-mediated cAMP increase affects Th1, Th2 and Th17 differentiation and can contribute to the identification of novel therapeutic targets for the treatment of Th cell-related pathological processes.

Application of fast marching method and quality map to well trajectory optimization with a novel well parametrization

This paper proposes a novel parametrization technique for deviated and horizontal wells and investigates the potential of combining the Fast Marching Method (FMM) with Particle Swarm Optimization (PSO) to facilitate optimization of the trajectory of vertical, deviated, and horizontal wells. The parametrization technique is based on practical drilling parameters, which can be controlled easily. Results showed that the PSO-FMM could find similar solutions to the direct optimization (PSO-DIRECT) on simple, synthetic cases with few number wells, but not on complex cases with a large number of wells. The FMM was hybridized with a reservoir simulator (PSO-HYBRID) that resolved the shortcomings of the PSO-FMM by filtering the solutions of the PSO-FMM based on the net present value (NPV). Also, initializing the optimizer by a quality map (QM) and the PSO-FMM method significantly improved the solutions. Initializing the PSO-DIRECT method by the PSO-FMM increased the NPV by 12% compared to the PSO-FMM method, and an average growth of 8.4% compared to the PSO-HYBRID. Also, the PSO-DIRECT + QM could find better results than the direct optimization, with an average increase of 1.4% in the NPV; however, its computational cost was two times the computational cost of the PSO-DIRECT + FMM and even higher than the computational cost of the direct optimization. The novelty of this paper is proposing a new well trajectory parametrization technique and comprehensively investigating the application of the FMM in well location and trajectory optimization on synthetic and real-field cases. It was shown that it is better to use the FMM as a helper function for the reservoir simulator rather than using it to completely substitute the reservoir simulator.

Mitochondrial ROS-dependent CD4+PD-1+T cells are pathological expansion in patients with primary immune thrombocytopenia.

Aberrant-activated T cells, especially CD4+T cells, play a crucial part in the pathogenetic progress of immune thrombocytopenia (ITP). PD-1-mediated signals play a negative part in the activation of CD4+T cells. However, knowledge is limited on the pathogenic characteristics and function of CD4+PD-1+T cells in ITP. The frequency and phenotype including cell activation, apoptosis, and cytokine production of CD4+PD-1+T cells were evaluated by flow cytometry. PD-1 Ligation Assay was performed to assess the function of PD-1 pathway in CD4+T cells. Mitochondrial reactive oxygen species (mtROS) were detected by MitoSOX Red probe. Compared with healthy controls (HC), the frequencies of CD4+PD-1+T cells were significantly increased in ITP patients. However, these cells are not exhausted despite PD-1 expression. Besides retaining cytokine-producing potential, these CD4+PD-1+T cells also had a possible B-cell helper function including expressing ICOS, CD84, and CD40L. Moreover, the CD4+PD-1+T cell subset contained higher levels of mitochondrial ROS than CD4+PD-1-T cell subset in patients with ITP. And mtROS inhibition could reduce the secretion of the inflammatory cytokines and regulate the function of CD4+PD-1+T cells. Upon in-vitro T cell receptor (TCR) stimulation of CD4+T cells in the presence of plate-bound PD-L1 fusion protein (PD-L1-Ig), CD4+T cells from ITP patients appeared resistant to such PD-1-mediated inhibition of interferon (IFN)-γ secretion. The CD4+PD-1+T cells were more abundant in patients with ITP. Additionally, this CD4+PD-1+T cell subset may be a potential etiology of ITP and a potential immune therapeutic target for ITP patients in the future.