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Intracellular Flow Cytometry

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https://www.readbyqxmd.com/read/27911410/experimental-infection-with-listeria-monocytogenes-as-a-model-for-studying-host-interferon-%C3%AE-responses
#1
Jeeyoon Jennifer Ahn, Thirumahal Selvanantham, Monan Angela Zhang, Thierry Mallevaey, Shannon E Dunn
L. monocytogenes is a gram-positive bacterium that is a cause of food borne disease in humans. Experimental infection of mice with this pathogen has been highly informative on the role of innate and adaptive immune cells and specific cytokines in host immunity against intracellular pathogens. Production of IFN-γ by innate cells during sublethal infection with L. monocytogenes is important for activating macrophages and early control of the pathogen(1-3). In addition, IFN-γ production by adaptive memory lymphocytes is important for priming the activation of innate cells upon reinfection(4)...
November 16, 2016: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/27910869/multiparameter-mechanical-and-morphometric-screening-of-cells
#2
Mahdokht Masaeli, Dewal Gupta, Sean O'Byrne, Henry T K Tse, Daniel R Gossett, Peter Tseng, Andrew S Utada, Hea-Jin Jung, Stephen Young, Amander T Clark, Dino Di Carlo
We introduce a label-free method to rapidly phenotype and classify cells purely based on physical properties. We extract 15 biophysical parameters from cells as they deform in a microfluidic stretching flow field via high-speed microscopy and apply machine-learning approaches to discriminate different cell types and states. When employing the full 15 dimensional dataset, the technique robustly classifies individual cells based on their pluripotency, with accuracy above 95%. Rheological and morphological properties of cells while deforming were critical for this classification...
December 2, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27906831/bos-is-associated-with-increased-cytotoxic-pro-inflammatory-cd8-t-nkt-like-and-nk-cells-in-the-small-airways
#3
Greg Hodge, Sandra Hodge, Aeneas Yeo, Phan Nguyen, Emily Hopkins, Holmes-Liew Cl, Paul N Reynolds, Mark Holmes
BACKGROUND: Immunosuppression therapy following lung transplantation fails to prevent bronchiolitis obliterans syndrome (BOS) in many patients, primarily a disease of the small airways. We have reported that BOS is associated with a lack of suppression of cytotoxic mediators, and pro-inflammatory cytokines, in peripheral blood T, NKT-like (particularly CD8+) and NK cells. We also showed a loss of glucocorticoid receptor (GCR) in pro-inflammatory lymphocytes following transplant. It is unknown whether these pro-inflammatory lymphocytes target the small and/or large airways in BOS...
December 1, 2016: Transplantation
https://www.readbyqxmd.com/read/27904959/differential-pulmonic-nk-and-nkt-cell-responses-in-schistosoma-japonicum-infected-mice
#4
Hefei Cha, Wenjuan Qin, Quan Yang, Hongyan Xie, Jiale Qu, Mei Wang, Daixiong Chen, Fang Wang, Nuo Dong, Longhua Chen, Jun Huang
Natural killer cells (NK cells) and natural killer T cells (NKT cells) play a role in anti-infection, anti-tumor, transplantation immunity, and autoimmune regulation. However, the role of NK and NKT cells during Schistosoma japonicum (S. japonicum) infection has not been widely reported, especially regarding lung infections. The aim of this study was to research the NK and NKT cell response to S. japonicum infection in the lungs of mice. Using immunofluorescent histological analysis, NK and NKT cells were found near pulmonary granulomas...
November 30, 2016: Parasitology Research
https://www.readbyqxmd.com/read/27904667/icaritin-induces-apoptotic-and-autophagic-cell-death-in-human-glioblastoma-cells
#5
Zhaopei Li, Xiangwen Meng, Lin Jin
BACKGROUND: GBM represents the most aggressive type of glioma which is featured by extremely aggressive invasion and destructive malignancy with a high proliferation rate. The aim of this study was to investigate the in vitro anti-tumor effect of icaritin in human GBM cell line U87. METHODS: The effect of icaritin on In vitro cell viability was determined by MTT assay and colony formation assay. The inducing effect of icaritin on cell cycle arrest, mitochondrial membrane potential loss, apoptosis, autophagy and intracellular ROS generation was assessed by flow cytometry...
2016: American Journal of Translational Research
https://www.readbyqxmd.com/read/27899395/functional-definition-of-progenitors-versus-mature-endothelial-cells-reveals-key-soxf-dependent-differentiation-process
#6
Jatin Patel, Elke J Seppanen, Mathieu P Rodero, Ho Yi Wong, Prudence Donovan, Zoltan Neufeld, Nicholas M Fisk, Mathias François, Kiarash Khosrotehrani
BACKGROUND: -During adult life, blood vessel formation is thought to occur via angiogenic processes involving branching from existing vessels. An alternate proposal suggests that neo-vessels form from endothelial progenitors able to assemble the intimal layers. We here aimed to define vessel-resident endothelial progenitors in vivo in a variety of tissues in physiological and pathological situations such as normal aorta, lungs, as well as wound healing, tumors and placenta. METHODS: -Based on protein expression levels of common endothelial markers using flow cytometry, three sub-populations of endothelial cells could be identified among VE-Cadherin+ and CD45- cells...
November 29, 2016: Circulation
https://www.readbyqxmd.com/read/27889841/the-function-of-activatable-cell-penetrating-peptides-in-human-intrahepatic-bile-duct-epithelial-cells
#7
Kui Tu, Lijin Zhao, Jin Gu, Pengzhan Yan, Fang Wang, Yu Cao
This study aimed to investigate the function of Activatable Cell-Penetrating Peptides (ACPP) in detecting the changes of human intrahepatic bile duct epithelial cell(hIBDEC). ACPP, which target matrix metalloproteinases, were constructed. All were labeled with FITC and Gd-DTPA at the N-terminal. Fluorescence microscopy was used to observe the fluorescence intensity inside hIBDEC after stimulating with different concentrations of LPS and incubating with different concentrations of ACPP to determine the optimal concentration range for LPS stimulation and the optimal concentration for FITC-ACPP effect...
November 26, 2016: Journal of Bioenergetics and Biomembranes
https://www.readbyqxmd.com/read/27884753/cell-free-dna-induced-apoptosis-of-granulosa-cells-by-oxidative-stress
#8
Yichun Guan, Wenjuan Zhang, Xingling Wang, Pengfei Cai, Qi Jia, Wenjie Zhao
BACKGROUND: Cell-free DNA is a DNA fragment that is produced by cell apoptosis which can affect the micro-environment of cell apoptosis. The levels of Cell-free DNA have been associated with successful rate of in vitro fertilization-embryo transfer (IVF-ET) and embryonic development. Our aim is to determine the relationship between cell-free DNA and embryo quality. The mechanisms of cell-free DNA in granulose and the apoptosis will be determined also. METHODS: The study enrolled patients who were undergone IVF for the first time and grouped the patients as pregnant (n=130) and non-pregnant (n=59)...
November 21, 2016: Clinica Chimica Acta; International Journal of Clinical Chemistry
https://www.readbyqxmd.com/read/27884734/nitric-oxide-coupled-signaling-in-odor-elicited-molecular-events-in-the-olfactory-center-of-the-terrestrial-snail-helix-pomatia
#9
Zoltán Serfőző, Kálmán Nacsa, Zoltán Veréb, Izabella Battonyai, Csaba Hegedűs, Csilla Balogh, Károly Elekes
Olfaction, a chemosensory modality, plays a pivotal role in the orientation and behavior of invertebrates. The central olfactory processing unit in terrestrial stylomatophoran snails is the procerebrum, which contains NO synthesizing interneurons, whose oscillatory currents are believed to be the base of odor evoked memory formation. Nevertheless, in this model the up- and downstream events of molecular cascades that trigger and follow NO release, respectively, have not been studied. Immunocytochemistry and flow cytometry studies performed on procerebral neural perikarya isolated from the snail Helix pomatia revealed cell populations with discrete DAF-2 fluorescence, indicating the release of different amounts of NO...
November 21, 2016: Cellular Signalling
https://www.readbyqxmd.com/read/27881156/klotho-sensitive-regulation-of-dendritic-cell-functions-by-vitamin-e
#10
Nguyen Thi Xuan, Phi Thi Thu Trang, Nguyen Van Phong, Nguyen Linh Toan, Do Minh Trung, Nguyen Duy Bac, Viet Linh Nguyen, Nguyen Huy Hoang, Nong Van Hai
BACKGROUND: Dendritic cells (DCs) are the most potent professional antigen-presenting cells for naive T cells to link innate and acquired immunity. Klotho, an anti-aging protein, participates in the regulation of Ca(2+) dependent migration in DCs. Vitamin E (VitE) is an essential antioxidant to protect cells from damage and elicits its inhibitory effects on NF-κB-mediated inflammatory response. However, the roles of VitE on mouse DC functions and the contribution of klotho to those effects both are unknown...
November 24, 2016: Biological Research
https://www.readbyqxmd.com/read/27879023/potential-effect-of-ezetimibe-against-mycobacterium-tuberculosis-infection-in-type-ii-diabetes
#11
I-Fang Tsai, Chiu-Ping Kuo, Andrew B Lin, Ming-Nan Chien, Hsin-Tsung Ho, Tsai-Yin Wei, Chien-Liang Wu, Yen-Ta Lu
BACKGROUND AND OBJECTIVE: Tuberculosis (TB) risk might be increased in patients with diabetes by factors other than hyperglycaemia, such as dyslipidaemia. Host lipids are essential energy sources used by mycobacteria to persist in a latent TB state. A potential therapy targeting cholesterol catabolism of mycobacteria has been proposed, but the potential of cholesterol-lowering drugs as anti-TB therapy is unclear. The purpose of this study was to determine the effects of ezetimibe, a 2-azetidinone cholesterol absorption inhibitor, on intracellular mycobacteria survival and dormancy...
November 23, 2016: Respirology: Official Journal of the Asian Pacific Society of Respirology
https://www.readbyqxmd.com/read/27878564/the-role-of-b7-family-costimulatory-molecules-and-indoleamine-2-3-dioxygenase-in-primary-sj%C3%A3-gren-s-syndrome-and-systemic-sclerosis
#12
Nóra Legány, László Berta, László Kovács, Attila Balog, Gergely Toldi
B7 costimulatory molecules are present on antigen-presenting cells (APCs) and influence intracellular expression of indoleamine 2,3-dioxygenase (IDO), a molecule with important immunoregulatory functions. We determined the frequency of activated (CD11b+) monocytes expressing B7-1, B7-2, B7-H1, and B7-H2 molecules, and that of CD3+ and CD4+ T cells expressing the corresponding CD28, CTLA-4, PD-1, and ICOS receptors in peripheral blood samples of 20 healthy adults and 9 SSc and 15 pSS patients using flow cytometry...
November 23, 2016: Immunologic Research
https://www.readbyqxmd.com/read/27878306/effects-of-interleukin-1-receptor-associated-kinase%C3%A2-1-rna-interference-in-dendritic-cells-on-inflammatory-cytokine-release-and-t-cell-proliferation
#13
Yan Gao, Xuxu Xu, Juan Feng, Ying Ma, Dongming Zheng, Yiming Meng, Fengping Shan
The present study aimed to reduce the expression of interleukin-1 receptor-associated kinase 1 (IRAK-1) in dendritic cells (DCs) by RNA interference (RNAi). Subsequently, its effects on the expression of costimulatory surface molecules, the release of inflammatory cytokines, and the proliferation of T cells during the activation process of DCs, were determined. RNAi was used to silence IRAK‑1 gene expression in DCs, followed by lipopolysaccharide stimulation. Flow cytometry was then used to detect the expression levels of DC surface molecules, including cluster of differentiation (CD)86, major histocompatibility complex class II and CD40...
November 15, 2016: Molecular Medicine Reports
https://www.readbyqxmd.com/read/27878296/pseudolaric-acid-b-inhibits-the-secretion-of-hepatitis-b-virus
#14
Jinghua Yu, Zengyan Wang, Peiyou Ren, Ting Zhong, Yue Wang, Fengmei Song, Jingwei Hou, Xiaoyan Yu, Shucheng Hua
High hepatitis B virus (HBV) load and chronic hepatitis B infection increase the risk of developing hepatocellular carcinoma (HCC), and is also associated with recurrence of HBV-related HCC. The aim of the present study was to investigate whether pseudolaric acid B (PAB), a diterpene acid isolated from the root and trunk bark of Pseudolarix kaempferi Gordon (Pinaceae), has an inhibitory role on the HBV secretion in HBV-related HCC. By detecting HBV surface antigen (HBsAg) by ELISA it was found that PAB inhibited HBV secretion in HepG2215 compared to control group, but did not decrease the intracellular HBV level, and the results were repeated in HepG2 cell transfect with HBV gene...
November 16, 2016: Oncology Reports
https://www.readbyqxmd.com/read/27877159/efficacy-of-antibacterial-peptides-against-peptide-resistant-mrsa-is-restored-by-permeabilization-of-bacteria-membranes
#15
Joshua Ravensdale, Zachary Wong, Frances O'Brien, Keith Gregg
Clinical application of antimicrobial peptides (AMPs), as with conventional antibiotics, may be compromised by the development of bacterial resistance. This study investigated AMP resistance in methicillin resistant Staphylococcus aureus, including aspects related to the resilience of the resistant bacteria toward the peptides, the stability of resistance when selection pressures are removed, and whether resistance can be overcome by using the peptides with other membrane-permeabilising agents. Genotypically variant strains of S...
2016: Frontiers in Microbiology
https://www.readbyqxmd.com/read/27873494/identificaiton-of-novel-immunogenic-human-papillomavirus-type-16-e7-specific-epitopes-restricted-to-hla-a-33-03-for-cervical-cancer-immunotherapy
#16
Sunghoon Kim, Hye Won Chung, Hoon Young Kong, Jong Baeck Lim
PURPOSE: To identify new immunogenic HLA-A*33;03-restricted epitopes from the human papillomavirus (HPV) 16 E7 protein for immunotherapy against cervical cancer. MATERIALS AND METHODS: We synthesized fourteen overlapping 15-amino acid peptides and measured intracellular interferon-γ (IFN-γ) production in PBMC and CD8+ cytotoxic T lymphocytes (CTLs) after sensitization with these peptides using flow cytometry and ELISpot assay. The immunogenicity of epitopes was verified using a ⁵¹Cr release assay with SNU1299 cells...
January 2017: Yonsei Medical Journal
https://www.readbyqxmd.com/read/27871834/ph-sensitive-pegylated-liposomes-for-delivery-of-an-acidic-dinitrobenzamide-mustard-prodrug-pathways-of-internalization-cellular-trafficking-and-cytotoxicity-to-cancer-cells
#17
Mimi M Yang, William R Wilson, Zimei Wu
This paper aims to develop and evaluate a pH-sensitive PEGylated liposomal (pPSL) system for tumor-targeted intracellular delivery of SN25860, a weakly acidic, poorly water-soluble dinitrobenzamide mustard prodrug which is activated by the E. coli nitroreductase nfB. pPSL and non pH-sensitive liposomes (nPSL), as reference, were formulated by thin-film hydration; an active drug loading method was developed with the aid of solubilizers. Cytotoxicity was evaluated in an nfsB-transfected EMT6 mouse mammary carcinoma cell line...
November 18, 2016: International Journal of Pharmaceutics
https://www.readbyqxmd.com/read/27869744/protective-effect-of-tempol-against-cisplatin-induced-ototoxicity
#18
Cha Kyung Youn, Jun Kim, Eu-Ri Jo, Jeonghyun Oh, Nam Yong Do, Sung Il Cho
One of the major adverse effects of cisplatin chemotherapy is hearing loss. Cisplatin-induced ototoxicity hampers treatment because it often necessitates dose reduction, which decreases cisplatin efficacy. This study was performed to investigate the effect of Tempol on cisplatin-induced ototoxicity in an auditory cell line, House Ear Institute-Organ of Corti 1 (HEI-OC1). Cultured HEI-OC1 cells were exposed to 30 μM cisplatin for 24 h with or without a 2 h pre-treatment with Tempol. Cell viability was determined using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and apoptotic cells were identified using terminal deoxynucleotidyl transferase dUTP nick end labeling of nuclei (TUNEL) assay and flow cytometry...
November 18, 2016: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/27869735/low-expression-of-capon-in-glioma-contributes-to-cell-proliferation-via-the-akt-signaling-pathway
#19
Shangfeng Gao, Jie Wang, Tong Zhang, Guangping Liu, Lei Jin, Daofei Ji, Peng Wang, Qingming Meng, Yufu Zhu, Rutong Yu
CAPON is an adapter protein for nitric oxide synthase 1 (NOS1). CAPON has two isoforms in the human brain: CAPON-L (long form of CAPON) and CAPON-S (short form of CAPON). Recent studies have indicated the involvement of CAPON in tumorigenesis beyond its classical role in NOS1 activity regulation. In this study, we found that the protein levels of CAPON-S, but not than CAPON-L, were significantly decreased in glioma tissues. Therefore, we established lentivirus-mediated stable cell lines with CAPON-S overexpression or down-regulation, and investigated the role of CAPON-S in the proliferation of glioma cells by using CCK8, EdU, and flow cytometry assays...
November 18, 2016: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/27866241/elevated-regulatory-t-cells-surface-and-intracellular-ctla-4-expression-and-interleukin-17-in-the-lung-cancer-microenvironment-in-humans
#20
Iwona Kwiecien, Anna Stelmaszczyk-Emmel, Malgorzata Polubiec-Kownacka, Dariusz Dziedzic, Joanna Domagala-Kulawik
Regulatory T cells (Tregs) play an important role in the suppression of the immune response in lung cancer. Cytotoxic T-lymphocyte antigen 4 (CTLA-4) expressed on T lymphocytes is capable of downregulating cytotoxic T cells and is constitutively expressed on Tregs. Little is known about the population of Tregs with two forms of CTLA-4: surface (s) and intracellular (in) in the lung cancer environment. Th17 cells defined by production of IL-17 have pleiotropic functions in anticancer immune response. Our aim was to detect the elements of immune response regulation in lung cancer in three compartments: by analysis of bronchoalveolar lavage fluid (BALF) from the lung affected by cancer (clBALF), healthy symmetrical lung (hlBALF) and peripheral blood (PB) from the same patient...
November 19, 2016: Cancer Immunology, Immunotherapy: CII
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