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Intracellular Flow Cytometry

Joseph B Margolick, Jay H Bream, Tricia L Nilles, Huifen Li, Susan J Langan, Shane Deng, Ruibin Wang, Nikolas Wada, Sean X Leng
Background: Both aging and treated human immunodeficiency virus (HIV)-infected populations exhibit low-level chronic immune activation of unknown etiology, which correlates with morbidity and mortality. Cytomegalovirus (CMV) infection is common in both populations, but its relation to immune activation is unknown. Methods: T cells from men who have sex with men (22 virologically suppressed HIV+, 20 HIV-) were stimulated with peptides spanning 19 CMV open reading frames, and intracellular cytokine responses were assessed...
February 26, 2018: Journal of Infectious Diseases
Jiye Sun, Xuemei Chen, Ting Liu, Xushun Jiang, Yue Wu, Shan Yang, Wei Hua, Zhengdong Li, Huizhe Huang, Xiongzhong Ruan, Xiaogang Du
BACKGROUND Increased lipid accumulation in renal tubular epithelial cells (TECs) contributes to their injury and dysfunction and progression of tubulointerstitial fibrosis. Berberine (BBR), a natural plant alkaloid isolated from traditional medicine herbs, is effective in lowing serum lipid, and has a protective effect on chronic kidney disease (CKD) with dyslipidemia, including diabetic nephropathy. The aim of this study was to investigate the effect of BBR on palmitate (PA)-induced lipid accumulation and apoptosis in TECs...
March 12, 2018: Medical Science Monitor: International Medical Journal of Experimental and Clinical Research
Xiaojie Zang, Guangji Wang, Qinngyun Cai, Xiao Zheng, Jingwei Zhang, Qianying Chen, Baojin Wu, Xiong Zhu, Haiping Hao, Fang Zhou
Multi-drug resistance (MDR) is a common limitation for the clinical use of microtubule-targeting chemotherapeutic agents and it is the main factor for poor prognoses in cancer therapy. Here, we report on deoxypodophyllotoxin (DPT), a promising microtubule inhibitor in phase I, as a promising candidate to circumvent the obstacle. DPT remarkably suppressed the tumor growth in xenograft mice bearing either Paclitaxel (PTX)-sensitive MCF-7/S or acquired resistance MCF-7/Adr (MCF-7/A) cells. Also, DPT exhibited a similar accumulation in both tumors, while PTX displayed much a lower accumulation in the resistant tumors...
March 9, 2018: Drug Metabolism and Disposition: the Biological Fate of Chemicals
Vida Sheikh, Pinar Kasapoglu, Alireza Zamani, Zahra Basiri, Ahmad Tahamoli-Roudsari, Mahdi Alahgholi-Hajibehzad
1 α, 25-dihydroxyvitamin D3 (VitD3) has been suggested to have strong modulatory properties in the immune system. Researchers in the present study primarily aimed to understand the effect of VitD3 on human CD4+ T cell proliferation in antigen presenting cells (APCs) free condition in vitro. The effect of VitD3 on intracellular cytokine responses trend to Th1, Th2, Th17 and Th22 was evaluated using the flow cytometry. Moreover the effect of VitD3 on the expression of inhibitory markers such as PD1, PD-L1, and CTLA4 which are induced upon polyclonal T cell receptor (TCR) activation on CD4+ T cells, was assessed...
March 6, 2018: Human Immunology
Ri-Zhen Huang, Xiao-Chao Huang, Bin Zhang, Hai-Yang Jia, Zhi-Xin Liao, Heng-Shan Wang
BACKGROUND: Hepatocellular carcinoma (HCC) is the leading cause of cancer death, and novel chemotherapeutic drugs for treating HCC are urgently needed. 16-O-caffeoyl-16-hydroxylhexadecanoic acid (CHHA) is a new phenylpropanoid isolated by our group from Euphorbia nematocypha which is commonly used to treat solid tumors. However, the underlying mechanisms responsible for the CHHA-induced apoptosis in cancer cells, particularly in HCC, remain unknown. PURPOSE: In the present work, we evaluated the growth inhibitory effect of CHHA on HCC cells and explored the underlying molecular mechanisms...
March 1, 2018: Phytomedicine: International Journal of Phytotherapy and Phytopharmacology
Lihua Zhu, Jia Li, Wu Dayang, Li Bing
PURPOSE: To investigate the protective effect of beta-casomorphin-7 (β-CM-7) in oxidative stressed human lens epithelial cells (HLECs) and to explore the possible mechanism for oxidative stress in human lens epithelial cells induced by high glucose. METHODS: We used HLECs to determine the effect of different concentrations of β-CM-7 on cell viability by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolimol/L bromide (MTT) assay. We used flow cytometry to determine the content of reactive oxygen species (ROS) induced by oxidative stress and a bioassay kit to determine the oxidant malondialdehyde (MDA) and antioxidant enzyme superoxide dismutase (SOD) levels...
March 8, 2018: Cutaneous and Ocular Toxicology
Mustafa Aziz Hatiboglu, Abdurrahim Kocyigit, Eray Metin Guler, Kerime Akdur, Arife Nalli, Ersin Karatas, Saffet Tuzgen
BACKGROUND: Prognosis of patients with melanoma brain metastasis is poor despite various chemotherapeutic agents. Researchers focus on finding effective treatment with low risk of toxicity. Thymoquinone (TQ) has been found to be effective on different types of cancer. However, no data exists on the effect of TQ in intracerebral melanoma. The purpose of this study was to assess the effect of TQ in B16-F10 melanoma cell in vitro and intracerebral melanoma in vivo. METHODS: The mechanisms of efficacy were investigated by ATP assay for cytotoxicity, flow cytometry and Acridine-orange staining for apoptosis, Comet assay for genotoxicity, CM-H2 DCF-DA (2,7-dichlorodihydrofluorescein) for intracellular reactive oxygen species (ROS) generation and ELISA methods for inflammatory cytokines...
March 3, 2018: World Neurosurgery
Kevin Marron, John F Kennedy, Conor Harrity
Background: Raised intracellular cytokine ratios (CKR) are proposed as a significant risk factor for adverse reproductive outcome. An elevated cytokine ratio, such as between TNFa and/or IFNg to IL-10 is associated with recurrent miscarriage (RM). The use of pharmacological immunomodulators such as TNFα inhibitors in these patients is controversial and not generally recommended due to a lack of conclusive data supporting their use. We evaluated whether the use of anti-oxidants/dietary supplements as an alternative could positively influence CKR's in ART patients...
2018: Fertility Research and Practice
Yun-Xin Lu, Qi-Nian Wu, Dong-Liang Chen, Le-Zong Chen, Zi-Xian Wang, Chao Ren, Hai-Yu Mo, Ya Chen, Hui Sheng, Ying-Nan Wang, Yun Wang, Jia-Huan Lu, De-Shen Wang, Zhao-Lei Zeng, Feng Wang, Feng-Hua Wang, Yu-Hong Li, Huai-Qiang Ju, Rui-Hua Xu
Rationale : The antitumor activity of high-dose ascorbate has been re-evaluated recently, but the mechanism underlying cell-specific sensitivity to ascorbate has not yet been clarified. Methods: The effects of high-dose ascorbate on gastric cancer were assessed using cancer cell lines with high and low expression of GLUT1 via flow cytometry and colony formation assays in vitro and patient-derived xenografts in vivo . Results : In this study, we demonstrated that gastric cancer cells with high GLUT1 expression were more sensitive to ascorbate treatment than cells with low GLUT1 expression...
2018: Theranostics
E N Lyukmanova, M L Bychkov, G V Sharonov, A V Efremenko, M A Shulepko, D S Kulbatskii, Z O Shenkarev, A V Feofanov, D A Dolgikh, M P Kirpichnikov
BACKGROUND AND PURPOSE: Nicotinic acetylcholine receptors (nAChRs) are a promising target for development of new anticancer therapies. Our purpose is to study effects of human proteins SLURP-1 and SLURP-2, antagonists of nAChRs, on human epithelial cancer cells. EXPERIMENTAL APPROACH: Growth of epithelial cancer cells (A431, SKBR3, MCF-7, A549, HT-29) exposed to SLURP-1, SLURP-2, mecamylamine, atropine, timolol and gefitinib was investigated by WST-1 test. Expression levels of SLURP-1, α7-nAChR and EGF receptors and their distribution in cancer cells were studied by confocal microscopy and flow cytometry...
March 5, 2018: British Journal of Pharmacology
Eanna P Connaughton, Serika Naicker, Shirley A Hanley, Stephanie M Slevin, John K Eykelenboom, Noel F Lowndes, Timothy O'Brien, Rhodri Ceredig, Matthew D Griffin, Michael C Dennedy
Human blood monocytes are sub-classified as classical, intermediate and non-classical. In this study, it was shown that conventionally-defined human intermediate monocytes (IM) can be divided into two distinct subpopulations with mid- and high-level surface expression of HLA-DR (referred to as DRmid and DRhi IM). These IM subpopulations were phenotypically and functionally characterized in healthy adult blood by flow cytometry, migration assays and lipoprotein uptake assays. Their absolute numbers and proportions were then compared in blood samples from obese and non-obese adults...
March 5, 2018: Immunology and Cell Biology
Preeti Viswanathan, Yogeshwar Sharma, Priya Gupta, Sanjeev Gupta
OBJECTIVES: Acetaminophen hepatotoxicity is a leading cause of hepatic failure with impairments in liver regeneration producing significant mortality. Multiple intracellular events, including oxidative stress, mitochondrial damage, inflammation, etc., signify acetaminophen toxicity, although how these may alter cell cycle controls has been unknown and was studied for its significance in liver regeneration. MATERIALS AND METHODS: Assays were performed in HuH-7 human hepatocellular carcinoma cells, primary human hepatocytes and tissue samples from people with acetaminophen-induced acute liver failure...
March 5, 2018: Cell Proliferation
Shixiang Wang, Nan Tan, Cuicui Ma, Jie Wang, Pu Jia, Jianghong Liu, Yang Yang, Zhanxiong Xie, Kun Zhao, Xiaohui Zheng
AIMS: In clinical practice, herbal medicines have played an important role in the modulation of drug transporters through the combination of conventional prescription drugs, which necessitates the elucidation of herb-drug interactions. The present study was designed to investigate the inhibitory effects and mechanisms of benzaldehyde, vanillin, muscone, and borneol on P-glycoprotein (P-gp). METHODS: The effects of the 4 compounds on the intracellular accumulation of rhodamine-123 (Rho-123) in vinblastine-treated Caco-2 (VB-Caco-2) cells were studied by monitoring fluorescence intensity through a flow cytometry assay, and the effects of these compounds on Rho-123 transport through VB-Caco-2 monolayers and Rho-123 intestinal absorption in the rat everted gut sac were investigated by high-performance liquid chromatography...
March 2, 2018: Pharmacology
Qiu-Li Zhang, Wei Wang, Yan Jiang, A-Tuya, Dongmei, Lan-Lan Li, Zhan-Jun Lu, He Chang, Tian-Zi Zhang
OBJECTIVE: To determine the effectiveness of GRGM-13 on oxidative stress induced apoptosis of retinal ganglion cells (RGCs) and revealed its possible mechanism. MATERIALS AND METHODS: Caspase-3 activity, MDA level, and glutathione peroxidase level were detected by Caspase-3 assay kit, Lipid Peroxidation MDA Assay Kit, and Total Glutathione Peroxidase Assay Kit, respectively. Protein levels of Bax, Bcl-2, p-p38 and p38 were observed by Western Blot. Reactive oxygen species assay kit was used to determine intracellular ROS level...
March 1, 2018: Biomedicine & Pharmacotherapy, Biomédecine & Pharmacothérapie
Xinya Xu, Yue Zheng, Yunfen Huang, Jian Chen, Zijian Gong, Yuying Li, Chun Lu, Wei Lai, Qingfang Xu
BACKGROUND: The deposition of advanced glycation end products (AGEs) is accelerated in photoaged skin, but the underlying mechanisms remain elusive. Intracellular degradation has been recently considered to play an important role in AGEs removal. Although lysosomal cathepsin D (CatD), B (CatB), L(CatL) and proteasomes are found to degrade internalized AGEs, it remains unknown which protease degrades internalized AGEs in human dermal fibroblasts (HDFs), and whether a decrease in intracellular degradation contributes to enhanced AGEs deposition in photoaged skin...
February 19, 2018: Journal of Dermatological Science
John Lucas, Mary Goetsch, Matt Fischer, Steven Forst
Xenorhabdus nematophila are Gram-negative bacteria that engage in mutualistic associations with entomopathogenic nematodes. To reproduce, the nematodes invade insects and release X. nematophila into the haemolymph where it functions as an insect pathogen. In complex medium, X. nematophila cells produce two distinct types of intracellular crystalline inclusions, one composed of the methionine-rich PixA protein and the other composed of the PixB protein. Here we show that PixB crystalline inclusions were neither apparent in X...
March 2, 2018: Microbiology
Abdul Hakeem, Fouzia Zahid, Guiting Zhan, Ping Yi, Hai Yang, Lu Gan, Xiangliang Yang
Background: Nanotechnology-based drug delivery systems exhibit promising therapeutic efficacy in cancer chemotherapy. However, ideal nano drug carriers are supposed to be sufficiently internalized into cancer cells and then release therapeutic cargoes in response to certain intracellular stimuli, which has never been an easy task to achieve. Objective: This study is to design mesoporous silica nanoparticles (MSNs)-based pH-responsive nano drug delivery system that is effectively internalized into cancer cells and then release drug in response to lysosomal/endosomal acidified environment...
2018: International Journal of Nanomedicine
Ajda Coker Gurkan, Elif Damla Arisan, Pinar Obakan Yerlikaya, Halime Ilhan, Narcin Palavan Unsal
PURPOSE: One of the recently developed polyamine (PA) analogues, N1 ,N11 -diethylnorspermine (DENSpm), has been found to act as an apoptotic inducer in melanoma, breast, prostate and colon cancer cells. Also, its potential to induce autophagy has been established. Unfolded protein responses and starvation of amino acids are known to trigger autophagy. As yet, however, the molecular mechanism underlying PA deficiency-induced autophagy is not fully clarified. Here, we aimed to determine the apoptotic effect of DENSpm after autophagy inhibition by 3-methyladenine (3-MA) or siRNA-mediated Beclin-1 silencing in colon cancer cells...
February 28, 2018: Cellular Oncology (Dordrecht)
Vibe Ballegaard, Peter Brændstrup, Karin Kaereby Pedersen, Nikolai Kirkby, Anette Stryhn, Lars P Ryder, Jan Gerstoft, Susanne Dam Nielsen
In people living with HIV (PLWHIV), coinfection with cytomegalovirus (CMV) has been associated with inflammation, immunological ageing, and increased risk of severe non-AIDS related comorbidity. The effect of CMV-specific immune responses on systemic inflammation, immune activation and T-cell senescence was evaluated in 53 PLWHIV treated with combination antiretroviral therapy (cART). Activated-, terminally differentiated-, naïve-, and senescent T-cells were assessed by flow cytometry, and plasma levels of CMV IgG, interleukin-6, tumor necrosis factor-α, high-sensitivity C-reactive protein and soluble-CD14 were measured...
February 28, 2018: Scientific Reports
Xiaoyan Zhou, Jie Weng, Jing Xu, Qiulin Xu, Weiju Wang, Weijin Zhang, Qiaobing Huang, Xiaohua Guo
BACKGROUND/AIMS: Disruption of endothelial barrier integrity in response to advanced glycation end products (AEGs) stimulation contributes to vasculopathy associated with diabetes mellitus. Mammalian diaphanous-related formin (mDia1) has been reported to bind to the cytoplasmic domain of the receptor for advanced glycation end products (RAGE), which induces a series of cellular processes. This study directly evaluated the participation of mDia1 in AGE-induced hyperpermeability and revealed the precise intracellular signal transductions of this pathological process...
February 23, 2018: Cellular Physiology and Biochemistry
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