Regulation Of Cell Activity Research Articles

Self-activated microbatteries to promote cell death through local electrical stimulation

Abstract Electrical stimulation of cells offers a viable alternative route to treat diseases like cancer that account for several deaths despite sophisticated treatments. While micro/nanoscale systems have demonstrated to regulate cell activity by inducing electrical signals, many of these rely on external excitation sources for operation. In this work, we present an array of Al-H2O based microbatteries of dimensions smaller than a single cell to induce localized electrical impulses by involving no excitation sources outside the cellular environment, i.e., self-stimulation, for a limited operation time. The fabricated structure allows microbattery activation, i.e., produces a voltage or current flow, on contact with the ionic species in biological medium. The study explores the response of human osteosarcoma cells (Saos-2) under the influence of these generated electrical impulses. The microbatteries allowed proliferation of Saos-2 cells when configured to operate in an open-circuit fashion, while they induced cell death in a short-circuited operation due to a current flow causing a drastic reduction in cell viability to 13.5%. The self-stimulated current produced a profound cytotoxic effect up to 24 h of incubation before the microbatteries became inactive. An up-scaled model and fabricated test-structure further explores the electrochemical characteristics of Al-H2O based batteries and their effect on relevant parameters like pH. The overall results suggest prospects for a non-invasive treatment in cancer therapy, neuromuscular system, neurological disorders, chronic treatments, tissue regeneration and other electrically active systems by the application of a self-stimulated electrical signal.

Abnormal promoter DNA hypermethylation of the integrin, nidogen, and dystroglycan genes in breast cancer

Cell transmembrane receptors and extracellular matrix components play a pivotal role in regulating cell activity and providing for the concerted integration of cells in the tissue structures. We have assessed DNA methylation in the promoter regions of eight integrin genes, two nidogen genes, and the dystroglycan gene in normal breast tissues and breast carcinomas (BC). The protein products of these genes interact with the basement membrane proteins LAMA1, LAMA2, and LAMB1; abnormal hypermethylation of the LAMA1, LAMA2, and LAMB1 promoters in BC has been described in our previous publications. In the present study, the frequencies of abnormal promoter hypermethylation in BC were 13% for ITGA1, 31% for ITGA4, 4% for ITGA7, 39% for ITGA9, 38% for NID1, and 41% for NID2. ITGA2, ITGA3, ITGA6, ITGB1, and DAG1 promoters were nonmethylated in normal and BC samples. ITGA4, ITGA9, and NID1 promoter hypermethylation was associated with the HER2 positive tumors, and promoter hypermethylation of ITGA1, ITGA9, NID1 and NID2 was associated with a genome-wide CpG island hypermethylated BC subtype. Given that ITGA4 is not expressed in normal breast, one might suggest that its abnormal promoter hypermethylation in cancer is non-functional and is thus merely a passenger epimutation. Yet, this assumption is not supported by our finding that it is not associated with a hypermethylated BC subtype. ITGA4 acquires expression in a subset of breast carcinomas, and methylation of its promoter may be preventive against expression in some tumors. Strong association of abnormal ITGA4 hypermethylation with the HER2 positive tumors (p = 0.0025) suggests that simultaneous presence of both HER2 and integrin α4 receptors is not beneficial for tumor cells. This may imply HER2 and integrin α4 signaling pathways interactions that are yet to be discovered.

Epsins 1 and 2 promote NEMO linear ubiquitination via LUBAC to drive breast cancer development.

Estrogen receptor-negative (ER-negative) breast cancer is thought to be more malignant and devastating than ER-positive breast cancer. ER-negative breast cancer exhibits elevated NF-κB activity, but how this abnormally high NF-κB activity is maintained is poorly understood. The importance of linear ubiquitination, which is generated by the linear ubiquitin chain assembly complex (LUBAC), is increasingly appreciated in NF-κB signaling, which regulates cell activation and death. Here, we showed that epsin proteins, a family of ubiquitin-binding endocytic adaptors, interacted with LUBAC via its ubiquitin-interacting motif and bound LUBAC's bona fide substrate NEMO via its N-terminal homolog (ENTH) domain. Furthermore, epsins promoted NF-κB essential modulator (NEMO) linear ubiquitination and served as scaffolds for recruiting other components of the IκB kinase (IKK) complex, resulting in the heightened IKK activation and sustained NF-κB signaling essential for the development of ER-negative breast cancer. Heightened epsin levels in ER-negative human breast cancer are associated with poor relapse-free survival. We showed that transgenic and pharmacological approaches eliminating epsins potently impeded breast cancer development in both spontaneous and patient-derived xenograft breast cancer mouse models. Our findings established the pivotal role epsins played in promoting breast cancer. Thus, targeting epsins may represent a strategy to restrain NF-κB signaling and provide an important perspective into ER-negative breast cancer treatment.

Circular RNA FOXP1 relieves trophoblastic cell dysfunction in recurrent pregnancy loss via the miR-143-3p/S100A11 cascade

ABSTRACT Recurrent pregnancy loss (RPL) is closely associated with insufficient functions of trophoblastic cells. Circular RNA forkhead box P1 (circFOXP1) can regulate cell activities in different types of diseases. However, its effects on trophoblastic cells and its role in RPL development remain unknown. In this study, gene expressions were detected by RT-qPCR. Protein levels were detected by Western blotting. Trophoblastic cell viability, apoptosis, invasion, and migration were respectively analyzed via CCK-8, flow cytometry, wound healing, and transwell assays. The association between miR–143–3p and circFOXP1 or S100A11 (S100 calcium binding protein A11) was explored and confirmed by bioinformatics prediction and luciferase reporter assay. Herein, miR–143–3p was upregulated in RPL. Furthermore, miR–143–3p upregulation induced apoptosis and suppressed proliferation, epithelial-to-mesenchymal transition (EMT) process, and metastatic capabilities of trophoblastic cells; whereas, miR–143–3p inhibition exert opposite effects. MiR–143–3p targeted S100A11 and was adversely regulated by circFOXP1 expression. S100A11 inhibition partially offset the effect of miR–143–3p knockdown on trophoblastic cell viability, apoptosis, EMT, invasion, and migration. In addition, circFOXP1 competitively combined with miR–143–3p, thus regulating S100A11 expression. Moreover, circFOXP1 regulated trophoblastic cell functions through the miR–143–3p/S100A11 cascade. To sum up, our study, for the first time, demonstrated that circFOXP1 could improve dysfunction of trophoblastic cells through the miR–143–3p/S100A11 axis, providing novel biomarkers and diagnostic targets for RPL.

Interacting Genetic Lesions of Melanoma in the Tumor Microenvironment: Defining a Viable Therapy.

Melanoma is the most aggressive form of skin cancer with an estimated 106,110 newly diagnosed cases in the United States of America in 2021 leading to an approximated 7180 melanoma-induced deaths. Cancer typically arises from an accumulation of somatic mutations and can be associated with mutagenic or carcinogenic exposure. A key characteristic of melanoma is the extensive somatic mutation rate of 16.8 mutations/Mb, which is largely attributed to UV exposure. Bearing the highest mutational load, many of them occur in key driver pathways, most commonly the BRAFV600E in the mitogen-activated protein kinase (MAPK) pathway. This driver mutation is targeted clinically with FDA-approved therapies using small molecule inhibitors of oncogenic BRAFV600E and MEK, which has greatly expanded therapeutic intervention following a melanoma diagnosis. Up until 2011, therapeutic options for metastatic melanoma were limited, and treatment typically fell under the spectrum of surgery, radiotherapy, and chemotherapy.Attributed to the extensive mutation rate, as well as having the highest number of neoepitopes, melanoma is deemed to be extremely immunogenic. However, despite this highly immunogenic nature, melanoma is notorious for inducing an immunosuppressive microenvironment which can be relieved by checkpoint inhibitor therapy. The two molecules currently approved clinically are ipilimumab and nivolumab, which target the molecules CTLA-4 and PD-1, respectively.A plethora of immunomodulatory molecules exist, many with redundant functions. Additionally, these molecules are expressed not only by immune cells but also by tumor cells within the tumor microenvironment. Tumor profiling of these cell surface checkpoint molecules is necessary to optimize a clinical response. The presence of immunomodulatory molecules in melanoma, using data from The Cancer Genome Atlas and validation of expression in two model systems, human melanoma tissues and patient-derived melanoma cells, revealed that the expression levels of B and T lymphocyte attenuator (BTLA), TIM1, and CD226, concurrently with the BRAFV600E mutation status, significantly dictated overall survival in melanoma patients. These molecules, along with herpesvirus entry mediator (HVEM) and CD160, two molecules that are a part of the HVEM/BTLA/CD160 axis, had a higher expression in human melanoma tissues when compared to normal skin melanocytes and have unique roles to play in T cell activation. New links are being uncovered between the expression of immunomodulatory molecules and the BRAFV600E genetic lesion in melanoma. Small molecule inhibitors of the MAPK pathway regulate the surface expression of this multifaceted molecule, making BTLA a promising target for immuno-oncology to be targeted in combination with small molecule inhibitors, potentially alleviating T regulatory cell activation and improving patient prognosis.

Microglia Control CNS T Regulatory Cell Activity During Remission From EAE Pathology

Microglia, the parenchymal brain macrophages of the central nerve system (CNS), have emerged as critical players in brain development and homeostasis. However, immune functions of these cells remain less well defined. We investigated contributions of microglia in a relapsing remitting (RR) multiple sclerosis paradigm, experimental autoimmune encephalitis (RR-EAE) in C57BL/6 / SJL F1 mice. Fate mapping-assisted translatome profiling during the RR disease course revealed the potential of microglia to interact with T cells through antigen presentation, co-stimulation and co-inhibition. Abundant microglia / T cell aggregates, as observed by histology and flow cytometry, supported the notion of functional interactions during remission, with a bias towards T regulatory cells. Finally, microglia ablation and microglia-restricted Ifnγ receptor mutagenesis, while compatible with EAE onset, significantly affected the T cell compartment of the animals and remission. Collectively, our data establish critical interactions of microglia with T cells during autoimmune neuro-degeneration.

SARS-CoV-2 sculpts the immune system to induce sustained virus-specific naïve-like and memory B-cell responses.

ObjectivesSARS‐CoV‐2 infection induces virus‐reactive memory B cells expressing unmutated antibodies, which hints at their emergence from naïve B cells. Yet, the dynamics of virus‐specific naïve B cells and their impact on immunity and immunopathology remain unclear.MethodsWe longitudinally profiled SARS‐CoV‐2‐specific B‐cell responses in 25 moderate‐to‐severe COVID‐19 patients by high‐dimensional flow cytometry and isotyping and subtyping ELISA. We also explored the relationship of B‐cell responses to SARS‐CoV‐2 with the activation of effector and regulatory cells from the innate or adaptive immune system.ResultsWe found a virus‐specific antibody response with a broad spectrum of classes and subclasses during acute infection, which evolved into an IgG1‐dominated response during convalescence. Acute infection was associated with increased mature B‐cell progenitors in the circulation and the unexpected expansion of virus‐targeting naïve‐like B cells. The latter further augmented during convalescence together with virus‐specific memory B cells. In addition to a transitory increase in tissue‐homing CXCR3+ plasmablasts and extrafollicular memory B cells, most COVID‐19 patients showed persistent activation of CD4+ and CD8+ T cells along with transient or long‐lasting changes of key innate immune cells. Remarkably, virus‐specific antibodies and the frequency of naïve B cells were among the major variables defining distinct immune signatures associated with disease severity and inflammation.ConclusionAside from providing new insights into the complexity of the immune response to SARS‐CoV‐2, our findings indicate that the de novo recruitment of mature B‐cell precursors into the periphery may be central to the induction of antiviral immunity.

Characterization of the Cell Surface Phenotype and Regulatory Activity of B-Cell IgD Low (BDL) Regulatory B Cells.

B-cell IgD Low (BDL) B cells have been shown to promote immunological tolerance by inducing proliferation of CD4+Foxp3+ T-regulatory cells (Treg) in a glucocorticoid-induced tumor necrosis factor receptor-related protein ligand (GITRL, Tnfsf18)-dependent manner. BDL cells constitute a small subset of splenic B lymphocytes that, in mice, are characterized by the B220+IgMintCD21intCD23+CD93-IgDlow/- cell surface expression profile. In this chapter, we show the flow cytometry gating strategy developed to identify and purify BDL. In addition, we describe an in vitro assay and two in vivo assays to assess BDL regulatory activity by quantitating Treg expansion/proliferation and indicate how they can be used in mouse models of disease. Collectively, these methods are useful to track and quantitate BDL and Treg numbers and assess their regulatory activity in inflammatory disease models.

Inhibition of BMP9 and BMP10 signalling by the ALK1-Fc Ligand Trap enhances systemic inflammation and increases vein graft atherosclerosis

Abstract Background Vein graft patency rapidly declines within 5 years due to extensive intimal hyperplasia (IH) and instable atherosclerosis. The transforming growth factor-β (TGF-β) superfamily is involved in vein graft failure by regulating cell activation and inflammation. Activin receptor–like kinase-1 (ALK1), a TGFβ superfamily type I receptor, induces signalling through the Smad1/5/8 pathway upon binding of BMP9/10. ALK1-Fc, a soluble chimeric protein, consisting of both the Fc-part of an antibody and the extracellular domain of ALK1, this functions as a ligand trap for BMP9/10 thereby inhibiting downstream signalling. Purpose To identify whether BMP signalling blockade enhances inflammatory responses and IH formation resulting in plaque destabilization. Methods All animal experiments were performed in compliance with Dutch government guidelines and the Directive 2010/63/EU of the European Parliament. Hypercholesterolemic male ApoE3*Leiden mice underwent an interposition of a donor caval vein in the carotid artery and were treated twice weekly with ip injections of 10 mg/kg control-Fc y or ALK1-Fc. Non-invasive ultrasound imaging was performed weekly until the day of sacrifice (d28). Flow cytometry was performed on blood on d7 and d28. Vein grafts were processed for histological analysis. Results The long-axis lumen diameter (relative to d7) in the control group decreased significantly at d21 with 5% and 11% at d28 compared to an increase of 5% (d21) and 2% (d28) in the ALK1-Fc group. D28 short-axis analyses demonstrated a significant difference in lumen area between a decrease of 8% (control-Fc) and a 30% increase (Alk1-Fc). Moreover, d28 short-axis analyses showed a significant difference in wall area between an 8% decrease in the controls and a 27% increase in the Alk1-Fc group. Histological assessment confirmed that ALK1-Fc resulted in increased outward remodelling and a 45% significant increase in lesion size. Lesion composition analyses showed a significant reduction in relative collagen, a trend in relative smooth muscle cell content reduction, and more destabilizing plaque dissections, characterized by blood filled gaps in the IH from the lumen towards the adventitia. At 7 and 28 days after surgery Ly6C monocytes in the blood were significantly increased in the ALK1-Fc group compared to control-FC. In the control-FC group the Ly6Chigh monocytes were significantly reduced at t28 in comparison to t7 whereas the Ly6Clow monocytes were significantly increased. In the ALK1-Fc treated mice, both Ly6C subpopulations stayed high at both time points. ALk1-Fc treatment thus results in a sustained pro-inflammatory response. Conclusions Ultrasound and histological assessment demonstrated an increase in outward remodelling, increased IH, a sustained inflammatory response and plaque destabilization upon ligand trapping of the ALK1-Smad1/5 signalling pathway. This identifies BMP9/10 signalling as an attractive target to treat vein graft failure. Funding Acknowledgement Type of funding source: Public hospital(s). Main funding source(s): Leiden University Medical Center

MicroRNA-936/ERBB4/Akt axis exhibits anticancer properties of gastric cancer through inhibition of cell proliferation, migration, and invasion.

Gastric cancer is one of the most common cancers globally and has a poor prognosis. MiR-936 has been reported to regulate cell activity and tumor progression in non-small cell lung cancer, glioma, and epithelial ovarian cancer. However, the specific role and mechanism of miR-936 in gastric cancer have not been explored. In present study, gastric cancer cells were transfected with miR-936 mimic, and cell proliferation, cell cycle distribution, cell apoptosis, migration and invasion were assessed via cell-counting kit-8, flow cytometry, wound healing, and transwell assay, respectively. Dual luciferase reporter assay was used to check miR-936 binding to its downstream target. It was shown that miR-936 was downregulated in gastric cancer tissues and cells. Erb-B2 Receptor Tyrosine Kinase 4 (ERBB4) was confirmed as a direct target of miR-936 and negatively regulated its expression by miR-936. Overexpression of miR-936 suppressed cell proliferation, cell cycle progression, cell migration and invasion, and enhanced cell apoptosis in gastric cancer cells, which could be reversed by further ERBB4 overexpression. Western blot results showed that miR-936/ERBB4 axis regulated Akt-related pathways to control gastric cancer cell activities. Therefore, our data suggest that miR-936 overexpression inhibits cell proliferation and invasion and promotes cell apoptosis through Akt-related pathways by targeting ERBB4, which provides novel insight to target miR-936 or miR-936/ERBB4 axis for the treatment of gastric cancer.

An update on the pathophysiology of immune thrombocytopenia.

To summarize the literature relating to the pathophysiology of ITP including the working mechanisms of therapies, T-cell and B-cell physiology, protein/RNA/DNA biochemistry, and animal models in an attempt to unify the perceived abnormal immune processes. The most recent pathophysiologic irregularities associated with ITP relate to abnormal T-cell responses, particularly, defective T regulatory cell activity and how therapeutics can restore these responses. The robust literature on T cells in ITP points to the notion that ITP is a disease initiated by faulty self-tolerance mechanisms very much like that of other organ-specific autoimmune diseases. There is also a large literature on new and existing animal models of ITP and these will be discussed. It appears that understanding how to specifically modulate T cells in patients with ITP will undoubtedly lead to effective antigen-specific therapeutics. ITP is predominately a T cell disorder which leads to a breakdown in self tolerance mechanisms and allows for the generation of anti-platelet autoantibodies and T cells. Novel therapeutics that target T cells may be the most effective way to perhaps cure this disorder.

Abstract CT238: An open-label, phase 1 study of IDO inhibitor KHK2455 in combination with avelumab in adult subjects with locally advanced or metastatic urothelial carcinoma

Abstract Background: Indoleamine 2,3-dioxygenase 1 (IDO1) degrades tryptophan to kynurenine, suppressing T cell effector activity and increasing T regulatory cell activity. IDO1 expression is induced in some cancer cells and has been associated with suppression of effector T cell responses toward tumors. In nonclinical studies, IDO1 inhibition combined with other clinically established immune modulating agents (eg, anti-CTLA-4 or anti-PD-1 antibodies) showed a significant improvement in tumor growth suppression. KHK2455 is a long acting and selective IDO1 inhibitor under development as a new approach to enhance anti-tumor immunity. KHK2455 is being combined with different anti-cancer therapies to explore its ability to enhance anti-tumor immunity and overcome tumor immune evasion. Avelumab is a PD-L1 checkpoint inhibitor approved for treatment of urothelial carcinoma. This study represents an opportunity to assess the complementary action of KHK2455 IDO1 inhibition and avelumab checkpoint inhibition in the treatment of urothelial carcinoma. Methods: Study 2455-002 is a two-part, multicenter, open-label Phase 1 study of KHK2455 in combination with avelumab in adult subjects with locally advanced or metastatic urothelial carcinoma (including bladder, urethra, ureters, and renal pelvis), who are checkpoint inhibitor naїve and previously treated with platinum-based therapy. Part 1 (dose-escalation) has a 3+3 design to evaluate safety and tolerability and identify the maximum tolerated dose (MTD). Dose de-escalation based on dose limiting toxicities is permitted. Part 2 (cohort expansion) further explores the safety, pharmacokinetics, pharmacodynamics, and anti-tumor activity for the combination regimen at the MTD or highest tested dose. All subjects will receive oral KHK2455 at their assigned dose and avelumab IV. The primary study endpoints are safety and tolerability, and the secondary endpoints are efficacy (based on RECIST v1.1 criteria), pharmacokinetics, and immunogenicity. Exploratory endpoints include examination of pre and post treatment biopsies for intratumoral suppression of IDO1 activity to correlate peripheral and intratumoral suppression, along with pre and post treatment serum markers including, but not limited to, immune mediators and related cytokines. Descriptive statistics (n, mean, standard deviation, median, minimum, and maximum) for continuous variables and frequency distributions and percentages for discrete variables will be utilized. Approximately 44 subjects are planned for enrollment (12 subjects in Part 1 and 32 subjects in Part 2). Clinical trial information: NCT03915405 (sponsor: Kyowa Kirin Pharmaceutical Development, Inc.) Citation Format: Pratibha Desai, Santosh Rao, Yuta Ikawa, Barbara Kapelan, Sergey Efuni, Robert Latek, Takashi Sato, Denis Healy, Yousef Zakharia. An open-label, phase 1 study of IDO inhibitor KHK2455 in combination with avelumab in adult subjects with locally advanced or metastatic urothelial carcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr CT238.

NF1, Neurofibromin and Gene Therapy: Prospects of Next-Generation Therapy.

Neurofibromatosis type 1 [NF1] is an autosomal dominant genetic disorder affecting multiple organs. NF1 is well known for its various clinical manifestations, including café-au-late macules, Lisch nodules, bone deformity and neurofibromas. However, there is no effective therapy for NF1. Current therapies are aimed at alleviating NF1 clinical symptoms but not curing the disease. By altering pathogenic genes, gene therapy regulates cell activities at the nucleotide level. In this review, we described the structure and functions of neurofibromin domains, including GAP-related domain [GRD], cysteine-serine rich domain [CSRD], leucine-rich domain [LRD] and C-terminal domain [CTD], which respectively alter downstream pathways. By transfecting isolated sequences of these domains, researchers can partially restore normal cell functions in neurofibroma cell lines. Furthermore, recombinant transgene sequences may be designed to encode truncated proteins, which is functional and easy to be packaged into viral vectors. In addition, the treatment effect of gene therapy is also determined by various factors such as the vectors selection, transgene packaging strategies and drug administration. We summarized multiple NF1 gene therapy strategies and discussed their feasibility from multiple angles. Different protein domains alter the function and downstream pathways of neurofibromin.

Knock-Down of Long Non-Coding RNA ANRIL Suppresses Mouse Mesangial Cell Proliferation, Fibrosis, Inflammation via Regulating Wnt/β-Catenin and MEK/ERK Pathways in Diabetic Nephropathy.

Our study aimed to investigate the role of long non-coding RNA ANRIL (lnc-ANRIL) knock-down in regulating cell activities, inflammation and downstream signaling pathways in mouse mesangial cellular diabetic nephropathy (DN) model.: METHODS: The mouse mesangial cells (SV40-MES13 cells) were treated with high-glucose (HG) to construct cellular DN model. Lnc-ANRIL knock-down plasmid and control knock-down plasmid were transfected into HG-treated SV40-MES13 cells as Sh-ANRIL group and Sh-NC group respectively. Lnc-ANRIL expression was significantly higher in HG-treated SV40-MES13 cells compared with normal glucose-treated SV40-MES13 cells and osmotic control-treated SV40-MES13 cells. Lnc-ANRIL knock-down suppressed cell proliferation and promoted cell apoptosis in HG-treated SV40-MES13 cells. As for fibrosis, lnc-ANRIL knock-down reduced fibronectin and collagen I expressions in HG-treated SV40-MES13 cells. Besides, the expressions of supernatant tumor necrosis factor-alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1), interleukin (IL)-1β, IL-6, IL-8 and IL-18 were reduced in Sh-ANRIL group compared with Sh-NC group. Furthermore, Wnt3, β-catenin, p-MEK1 and p-ERK1 expressions were suppressed in Sh-ANRIL group compared with Sh-NC group, which suggested that lnc-ANRIL knock-down inhibited Wnt/β-catenin and MEK/ERK pathways in HG-treated SV40-MES13 cells. Lnc-ANRIL knock-down suppresses mouse mesangial cell proliferation, fibrosis, inflammation, Wnt/β-catenin and MEK/ERK pathways in DN.

Chiral CuxCoyS Nanoparticles under Magnetic Field and NIR Light to Eliminate Senescent Cells

In the present study, chiral Cux Coy S nanoparticles (NPs) were developed to selectively induce apoptosis of senescent cells using both an alternating magnetic field (AMF) and near infrared (NIR) photon illumination. The chiral effects on living cells were investigated, and d-Cux Coy S NPs showed about 2.5 times higher of internalized ability than l-NPs. By modifying beta 2 macroglobulin (MG), senescent cells were effectively eliminated by d-Cux Coy S NPs without damaging the activities of normal cells under AMF and photon illumination. Compared to the individual application of NIR illumination and AMF, their synergistic effect induced the production of caspase-3 with a much shorter treatment time and higher efficiency due to the more serious photon-induced cellular redox and mechanical damage of cellular skeleton. Moreover, the developed strategy was successfully used to remove senescent cells in vivo. This study developed a controllable way of regulating cell activities using chiral NPs, which will provide a valuable way for treating diseases and promoting health.

Chiral CuxCoyS Nanoparticles under Magnetic Field and NIR Light to Eliminate Senescent Cells

AbstractIn the present study, chiral CuxCoyS nanoparticles (NPs) were developed to selectively induce apoptosis of senescent cells using both an alternating magnetic field (AMF) and near infrared (NIR) photon illumination. The chiral effects on living cells were investigated, and d‐CuxCoyS NPs showed about 2.5 times higher of internalized ability than l‐NPs. By modifying beta 2 macroglobulin (MG), senescent cells were effectively eliminated by d‐CuxCoyS NPs without damaging the activities of normal cells under AMF and photon illumination. Compared to the individual application of NIR illumination and AMF, their synergistic effect induced the production of caspase‐3 with a much shorter treatment time and higher efficiency due to the more serious photon‐induced cellular redox and mechanical damage of cellular skeleton. Moreover, the developed strategy was successfully used to remove senescent cells in vivo. This study developed a controllable way of regulating cell activities using chiral NPs, which will provide a valuable way for treating diseases and promoting health.

MicroRNA-377 Alleviates Myocardial Injury Induced by Hypoxia/Reoxygenation via Downregulating LILRB2 Expression.

Background:miR-377 is closely related to myocardial regeneration. miR-377-adjusted mesenchymal stem cells abducted ischemic cardiac angiogenesis. Nevertheless, there were rarely reports about the impact of miR-377 on myocardial ischemia injury. The purpose of this work is that whether miR-377 can protect against myocardial injury caused by hypoxia/reoxygenation (H/R).Methods:Gene expression omnibus database (http://www.ncbi.nlm.nih.gov/geo/; no. GSE53211) was utilized to study the differential expression of miR-377 in patients with an acute ST-segment elevation myocardial infarction and healthy controls. The luciferase activity was determined utilizing the dual-luciferase reporter system. Quantitative real-time polymerase chain reaction and Western blotting were used to measure the messenger RNA and protein level.Results:Low expression of miR-377 and high expression of leukocyte immunoglobulin-like receptor B2 (LILRB2) were identified in patients with myocardial infarction from analyzing the Gene Expression Omnibus data set. Besides, miR-377 expression was downregulated in cardiomyocyte exposed to H/R. Additionally, overexpression of miR-377 could visibly improve cardiomyocyte injury by regulating cell activity and apoptosis.Conclusions:In short, our findings suggested that miR-377/LILRB2 might regard as a hopeful therapeutic target for myocardial ischemic.

Analysis of genetic information from the antlers of Rangifer tarandus (reindeer) at the rapid growth stage.

Reindeer is the only deer species in which both males and females regularly grow antlers, providing an excellent model for studying the rapid growth and annual regeneration of antlers. The study of genetic information from reindeer is the basis for revealing the unique mechanism of antler growth. In the present study, we obtained 18.86 GB of clean reads, which were assembled to obtain 94,575 unigenes (average length: 704.69). Among these reads, 30,980 sequences were identified by searching a database of known proteins and then annotated with Gene Ontology (GO) terms, Clusters of Orthologous Groups (COG) classifications and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. All 7,480 simple sequence repeats (SSRs) were detected. A total of 84,435 and 82,226 high-quality single-nucleotide polymorphisms (SNPs) were identified in male and female reindeer, respectively. We identified 31 genes that were highly expressed in reindeer antlers. These genes regulate cell activities that are closely associated with the process of rapid tissue growth. Our results provide a basis for studying reindeer antlers and for further studying the molecular genetics, population genetics, and functional genomics of reindeer.

Cnp Promoter-Driven Sustained ERK1/2 Activation Increases B-Cell Activation and Suppresses Experimental Autoimmune Encephalomyelitis.

The ERK1/2 signaling pathway promotes myelin wrapping during development and remyelination, and sustained ERK1/2 activation in the oligodendrocyte (OL) lineage results in hypermyelination of the CNS. We therefore hypothesized that increased ERK1/2 signaling in the OL lineage would 1) protect against immune-mediated demyelination due to increased baseline myelin thickness and/or 2) promote enhanced remyelination and thus functional recovery after experimental autoimmune encephalomyelitis (EAE) induction. Cnp-Cre;Mek1DD-eGFP/+ mice that express a constitutively active form of MEK1 (the upstream activator of ERK1/2) in the OL lineage, exhibited a significant decrease in EAE clinical severity compared to controls. However, experiments using tamoxifen-inducible Plp-CreERT;Mek1DD-eGFP/+ or Pdgfrα-CreERT;Mek1DD-eGFP mice revealed this was not solely due to a protective or reparative effect resulting from MEK1DD expression specifically in the OL lineage. Because EAE is an immune-mediated disease, we examined Cnp-Cre;Mek1DD-eGFP/+ splenic immune cells for recombination. Surprisingly, GFP+ recombined CD19+ B-cells, CD11b+ monocytes, and CD3+ T-cells were noted when Cre expression was driven by the Cnp promoter. While ERK1/2 signaling in monocytes and T-cells is associated with proinflammatory activation, fewer studies have examined ERK1/2 signaling in B-cell populations. After in vitro stimulation, MEK1DD-expressing B-cells exhibited a 3-fold increase in CD138+ plasmablasts and a 5-fold increase in CD5+CD1dhi B-cells compared to controls. Stimulated MEK1DD-expressing B-cells also exhibited an upregulation of IL-10, known to suppress the initiation of EAE when produced by CD5+CD1dhi regulatory B-cells. Taken together, our data support the conclusion that sustained ERK1/2 activation in B-cells suppresses immune-mediated demyelination via increasing activation of regulatory B10 cells.

Abstract B060: Side-by-side comparison of small molecule IDO1 inhibitors in biochemical and cell-based assays and development of a IDO1-expressing mouse model to evaluate target modulation

Abstract Introduction: Indoleamine 2,3-dioxygenase (IDO1) is a heme-containing oxidoreductase enzyme that converts L-tryptophan (Trp) into N’-formylkynurenine (NFK). IDO1 is broadly expressed by tumor cells as well as immune cells in the tumor microenvironment of many cancers, which is often correlated with poor prognosis. Depletion of Trp and increased L-kynurenine (Kyn) levels induce immune tolerance by suppression of effector T-cell and natural killer cell functions, and activation of regulatory T-cells and myeloid-derived suppressor cells. Four small molecule inhibitors are currently investigated in phase III (linrodostat/BMS-986205), phase II (epacadostat/INCB024360) or phase I (MK7162 and LY3381916) clinical trials. Linrodostat, epacadostat and LY3381916 are reported as being selective for IDO1 over TDO, while details on MK7162 have not yet been disclosed. The compounds inhibit IDO1 with different mechanisms, with epacadostat binding to the heme-group in the catalytic center of IDO1, while linrodostat and LY3381916 bind to apo-IDO1 and compete with heme for the active site [1-3]. Experimental procedures Clinical and reference IDO1 inhibitors were characterized in biochemical assays for IDO1 and tryptophan 2,3-dioxygenase (TDO), a structurally different enzyme, which also catalyzes the conversion of Trp to NFK. Furthermore, the compounds were profiled in a panel of functional cell-based assays, including human cancer cell lines and assays based on IDO1- or TDO2-overexpressing HEK293 cells. A IDO1-expressing subline of the mouse B16F10 melanoma cell line was generated and used to develop a syngeneic mouse model at Charles River Laboratories (USA) to determine target modulation in vivo [4]. Stable gene expression was confirmed by qPCR and target modulation was examined by measurement of Trp and Kyn levels using LC-MS/MS. Results: Linrodostat has sub-nanomolar cellular potency, despite the absence of any biochemical activity on the timescale of our assays, which is consistent with its reported heme-competing mechanism of inhibition [1]. Linrodostat also inhibits different Cytochrome P450 enzymes with micromolar activity. In our biochemical assays, epacadostat was not selective for IDO1 over TDO, whereas in the IDO1- and TDO2-overexpressing HEK293 cell lines it was 2000 times selective for IDO1. High level expression of IDO1 in B16F10 cells did not result in enhanced tumor growth after grafting in syngeneic mice, which contrasts published data with a similar model [4]. Nonetheless, we observed strong modulation of Trp and Kyn levels in plasma and in tumors of the IDO1-overexpressing mouse model, compared to non-tumor bearing mice. Treatment of the B16F10-IDO1 model with epacadostat did not result in a reduction of tumor growth, though epacadostat did induce clear changes in Trp and Kyn in both plasma and tumor tissue. Conclusion: Our comparative study of the potencies and selectivities of IDO1 inhibitors, as well as our model for measuring in vivo target modulation, helps to identify strengths and weaknesses of current IDO1 inhibitors, and supports the development of new inhibitors. [1] Nelp et al. (2018) Proc. Natl. Acad. Sci. U.S.A. 115, 3249-3254; [2] Yue et al. (2017) ACS Med. Chem. Lett. 8, 486-491; [3] Dorsey et al. (2018) Proceedings: AACR Annual Meeting 2018, Abstract nr. 5245; [4] Holmgaard et al. (2015) Cell Rep. 13, 412-424. Citation Format: Yvonne Grobben, Joost C.M. Uitdehaag, Antoon M. van Doornmalen, Nicole Willemsen-Seegers, Diep Vu-Pham, Jos de Man, Rogier C. Buijsman, Guido J.R. Zaman. Side-by-side comparison of small molecule IDO1 inhibitors in biochemical and cell-based assays and development of a IDO1-expressing mouse model to evaluate target modulation [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B060. doi:10.1158/1535-7163.TARG-19-B060