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Intracellular pH flow cytometry

Dishi Wang, Yanxia Zhou, Xinru Li, Xiaoyou Qu, Yunqiang Deng, Ziqi Wang, Chuyu He, Yang Zou, Yiguang Jin, Yan Liu
pH-responsive polymeric micelles have shown promise for the targeted and intracellular delivery of antitumor agents. The present study aimed to elucidate the possible mechanisms of pH-sensitivity and cellular internalization of PEOz-b-PLA micelles in detail, further unravel the effect of hydrophilic/hydrophobic ratio of the micelles on their cellular internalization, and examine the intracellular trafficking routes and fate of PEOz-b-PLA after internalization of the micelles. The results of variations in the size and Zeta potential of PEOz-b-PLA micelles and cross-sectional area of PEOz-b-PLA molecules with pH values suggested that electrostatic repulsion between PEOz chains resulting from ionization of the tertiary amide groups along PEOz chain at lower pH than its pKa was responsible for pH-sensitivity of PEOz-b-PLA micelles...
February 10, 2017: ACS Applied Materials & Interfaces
Shoubing Zhou, Dan Wu, Xiaodong Yin, Xiaoxiao Jin, Xiu Zhang, Shiya Zheng, Cailian Wang, Yanwen Liu
BACKGROUND: One of the main problems in B cell lymphoma treatment is severe adverse effects and low therapeutic efficacy resulting from systemic chemotherapy. A pH-sensitive controlled drug release system based on mesoporous silica nanoparticles was constructed for targeted drug delivery to tumor cells to reduce systemic toxicity and improve the therapeutic efficacy. METHODS: In this study, the doxorubicin (DOX) was filled into the mesopores of the functional MSNs (DMSNs)...
February 6, 2017: Journal of Experimental & Clinical Cancer Research: CR
Sergii Kalytchuk, Kateřina Poláková, Yu Wang, Jens P Froning, Klara Cepe, Andrey L Rogach, Radek Zbořil
Nanoscale biocompatible photoluminescence (PL) thermometers that can be used to accurately and reliably monitor intracellular temperatures have many potential applications in biology and medicine. Ideally, such nanothermometers should be functional at physiological pH across a wide range of ionic strengths, probe concentrations, and local environments. Here, we show that water-soluble N,S-co-doped carbon dots (CDs) exhibit temperature-dependent photoluminescence lifetimes and can serve as highly sensitive and reliable intracellular nanothermometers...
January 31, 2017: ACS Nano
Wenyan Li, Kevin D Houston, Jessica P Houston
Phase-sensitive flow cytometry (PSFC) is a technique in which fluorescence excited state decay times are measured as fluorescently labeled cells rapidly transit a finely focused, frequency-modulated laser beam. With PSFC the fluorescence lifetime is taken as a cytometric parameter to differentiate intracellular events that are challenging to distinguish with standard flow cytometry. For example PSFC can report changes in protein conformation, expression, interactions, and movement, as well as differences in intracellular microenvironments...
January 16, 2017: Scientific Reports
Yinyang Hu, Fang Zeng
The clinical applications of tradition prodrug are restricted to a certain extent due to its some defects such as side effects, low selectivity and single function. Herein, we designed a theranostic strategy based on the fluorescence resonance energy transfer (FRET) mechanism. The theranostic prodrug was constructed with an anticancer drug camptothecin (CPT), a cleavable linker based on disulfide bonds (SS) and a fluorophore naphthalimide derivative (NAP). For this drug delivery and reporting system (NAP-SS-CPT), FRET occurs between CPT (energy donor) and NAP (energy receptor); and upon cellular uptake by GSH-overexpressing cancer cells, disulfide bonds could be successfully cleaved by the high concentration of GSH, and FRET process was interrupted to achieve dual fluorescence response and specifically release of CPT...
March 1, 2017: Materials Science & Engineering. C, Materials for Biological Applications
Mouna Dammak, Sandra Mareike Haase, Ramzi Miladi, Faten Ben Amor, Mohamed Barkallah, David Gosset, Chantal Pichon, Bernhard Huchzermeyer, Imen Fendri, Michel Denis, Slim Abdelkafi
BACKGROUND: The increasing demand for microalgae lipids as an alternative to fish has encouraged researchers to explore oleaginous microalgae for food uses. In this context, optimization of growth and lipid production by the marine oleaginous V2-strain-microalgae is of great interest as it contains large amounts of mono-unsaturated (MUFAs) and poly-unsaturated fatty acids (PUFAs). METHODS: In this study, the isolated V2 strain was identified based on 23S rRNA gene...
December 5, 2016: Lipids in Health and Disease
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...
January 10, 2017: International Journal of Pharmaceutics
Wei Chen, Yong Hou, Zhaoxu Tu, Lingyan Gao, Rainer Haag
pH-Degradable PVA nanogels, which are prepared by photo-crosslinking thermo-preinduced PVA nanoaggregates in water without any surfactants or toxic organic solvents, are used for intracellular PTX release and anticancer treatment. These nanogels fast degraded at mildly acidic conditions with a pH-triggered PTX release, and the degradation products are only native PVA and poly(hydroxyethyl acrylate) (PHEA) as well as acetaldehyde without any toxic byproducts. The nanogel sizes could be tailored by different temperatures during the crosslinking process...
October 31, 2016: Journal of Controlled Release: Official Journal of the Controlled Release Society
M Carmen Vílchez, Marina Morini, David S Peñaranda, Víctor Gallego, Juan F Asturiano, Luz Pérez
The role of potassium from the seminal plasma and/or the activation media was examined by selectively removing K(+) from this media, and by testing the use of K(+) channel inhibitors and a K-ionophore. Sperm motility was measured using a CASA system, intracellular K(+) and pH were measured by flow cytometry, and sperm head area was measured by ASMA: Automated Sperm Morphometry Analyses. Sperm motility was notably inhibited by the removal of K(+) from the seminal plasma and by treatment with the K(+) ionophore valinomycin...
January 2017: Comparative Biochemistry and Physiology. Part A, Molecular & Integrative Physiology
Natalia Rios, Lucía Piacenza, Madia Trujillo, Alejandra Martínez, Verónica Demicheli, Carolina Prolo, María Noel Álvarez, Gloria V López, Rafael Radi
The specific and sensitive detection of peroxynitrite (ONOO(-)/ONOOH) in biological systems is a great challenge due to its high reactivity towards several biomolecules. Herein, we validated the advantages of using fluorescein-boronate (Fl-B) as a highly sensitive fluorescent probe for the direct detection of peroxynitrite under biologically-relevant conditions in two different cell models. The synthesis of Fl-B was achieved by a very simply two-step conversion synthetic route with high purity (>99%) and overall yield (∼42%)...
December 2016: Free Radical Biology & Medicine
Yogesh Singh, Yuetao Zhou, Xiaolong Shi, Shaqiu Zhang, Anja T Umbach, Madhuri S Salker, Karl S Lang, Florian Lang
CD4(+) T helper 9 (Th9) cells are a newly discovered Th cell subset that produce the pleiotropic cytokine IL-9. Th9 cells can protect against tumors and provide resistance against helminth infections. Given their pivotal role in the adaptive immune system, understanding Th9 cell development and the regulation of IL-9 production could open novel immunotherapeutic opportunities. The Na(+)/H(+) exchanger 1 (NHE1; gene name Slc9α1)) is critically important for regulating intracellular pH (pHi), cell volume, migration, and cell survival...
November 4, 2016: Journal of Biological Chemistry
Xinlong Zang, Huaiwei Ding, Xiufeng Zhao, Xiaowei Li, Zhouqi Du, Haiyang Hu, Mingxi Qiao, Dawei Chen, Yuihui Deng, Xiuli Zhao
Therapeutic delivery of small interfering RNA (siRNA) is a major challenge that limits its potential clinical application. Here, a pH-sensitive cholesterol-Schiff base-polyethylene glycol (Chol-SIB-PEG)-modified cationic liposome-siRNA complex, conjugated with the recombinant humanized anti-EphA10 antibody (Eph), was developed as an efficient nonviral siRNA delivery system. Chol-SIB-PEG was successfully synthesized and confirmed with FTIR and (1)H-NMR. An Eph-PEG-SIB-Chol-modified liposome-siRNA complex (EPSLR) was prepared and characterized by size, zeta potential, gel retardation, and encapsulation efficiency...
2016: International Journal of Nanomedicine
Cora Roehlecke, Monika Valtink, Annika Frenzel, Doris Goetze, Lilla Knels, Henning Morawietz, Richard H W Funk
PURPOSE: Intracellular formation of advanced glycation end products (AGEs) is a crucial pathological process in retinal diseases such as age-related macular degeneration (AMD) or diabetic retinopathy (DR). Glyoxal is a physiological metabolite produced during formation of AGEs and has also been shown to derive from photodegraded bisretinoid fluorophores in aging retinal pigment epithelial (RPE) cells. METHODS: Flow cytometry was combined with either: 1) immunocytochemical staining to detect glyoxal induced formation of N(ε)-carboxymethyllysine (CML)-modifications of intracellular proteins (AGEs) and changes in the production of stress response proteins; or 2) vital staining to determine apoptosis rates (annexin V binding), formation of intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and changes in intracellular pH upon treatment of cells with glyoxal...
December 2016: Graefe's Archive for Clinical and Experimental Ophthalmology
Sara Basbous, Anaïs Levescot, Nathalie Piccirilli, Françoise Brizard, François Guilhot, Lydia Roy, Nicolas Bourmeyster, Jean-Marc Gombert, André Herbelin
CD1d-restricted iNKT cells are believed to play a key role in cancer immune surveillance and are functionally deficient in chronic myeloid leukaemia (CML). Herein, we have hypothesized that this defect might originate from BCR-ABL-dependent dysfunctions in myeloid dendritic cells (mDCs). Indeed, flow cytometry and confocal microscopy revealed that cell surface expression of CD1d was downregulated in CML mDCs, relative to healthy donor (HD) controls. The decreased cell surface display of CD1d could not be ascribed to defective mDC differentiation, as attested by normal expression of HLA-DR and the CD86 maturation marker...
August 11, 2016: Journal of Pathology
A Kaszuba, A Konior, T Mikolajczyk, A Prejbisz, M Kabat, I Cendrowska-Demkow, P Dobrowolski, B Pregowska-Chwala, K Michel-Rowicka, A Klisiewicz, P Michalek, S Toutounchi, Z Galazka, M Reincke, A Januszewicz, Tj Guzik
OBJECTIVE: Role of the T cells in pathogenesis of hypertension has been demonstrated in numerous mouse models, limited data is available in human hypertension. Therefore the aim of our study was to quantify blood T lymphocyte subsets, characterized by expression of CD25 markers (CD3 + , CD8 + and CD3 + CD4-CD8- cells) and expression of perforin and granzyme (in CD8 + cells) in subjects with primary hyperaldosteronism (PA), resistant hypertension (RHT) characterized by increased cardiovascular risk and excessive amounts of circulating aldosterone in comparison to controls matched for age, sex and BMI(control group, CG)...
September 2016: Journal of Hypertension
Venera Kuci Emruli, Roger Olsson, Fredrik Ek, Sara Ek
BACKGROUND: Mantle cell lymphoma (MCL) is an aggressive disease with short median survival. Molecularly, MCL is defined by the t(11;14) translocation leading to overexpression of the CCND1 gene. However, recent data show that the neural transcription factor SOX11 is a disease defining antigen and several involved signaling pathways have been pin-pointed, among others the Wnt/β-catenin pathway that is of importance for proliferation in MCL. Therefore, we evaluated a compound library focused on the Wnt pathway with the aim of identifying Wnt-related targets that regulate growth and survival in MCL, with particular focus on SOX11-dependent growth regulation...
2016: BMC Cancer
C Fourgeaud, S Derieux, S Mirshahi, S Besbes, J Chidiac, I Mahé, G Contant, M Pocard, J Soria, M Mirshahi
INTRODUCTION: Gastric signet ring cell carcinoma (GSRC) is a distinct entity among of other gastric cancer. With unknown etiopathology, their incidence is increasing and it presents a low sensitivity to chemotherapy. AIM: Here, we studied the expression of the heparanase (HPA) in cancer tissues from GSRC patients and several cancer cell lines. HPA is an endo-β-D-glucuronidase, capable of cleaving the lateral chains of heparan sulfate on cell surfaces and the extracellular matrix at acidic pH...
April 2016: Thrombosis Research
A Cisáriková, Z Barbieriková, L Janovec, J Imrich, L Hunáková, Z Bačová, H Paulíková
Acridin-3,6-dialkyldithiourea hydrochlorides (AcrDTUs) have been evaluated as a new group of photosensitizers (PSs) for photodynamic antitumor therapy (PDT). Mouse leukemia cells L1210 were used for testing of AcrDTUs as the new PSs. The irradiation (UV-A light (365 nm), 1.05 J/cm(2)) increased cytotoxicity of all derivatives against L1210 cells more than ten times. The highest photocytotoxicity was found for propyl-AcrDTU with IC50=0.48±0.03 μM after 48 h incubation. A generation of the superoxide radical anion upon UV-A irradiation of propyl-AcrDTU was confirmed by in situ photochemical EPR experiments...
May 1, 2016: Bioorganic & Medicinal Chemistry
Gunda Hansen, Claus Lindvald Johansen, Gunvor Marten, Jacqueline Wilmes, Lene Jespersen, Nils Arneborg
In this study, we investigated the influence of three extracellular pH (pHex) values (i.e., 5.5, 6.5, and 7.5) on the growth, viability, cell size, acidification activity in milk, and intracellular pH (pHi) of Lactococcus lactis subsp. lactis DGCC1212 during pH-controlled batch fermentations. A universal parameter (e.g., linked to pHi) for the description or prediction of viability, specific acidification activity, or growth behavior at a given pHex was not identified. We found viability as determined by flow cytometry to remain high during all growth phases and irrespectively of the pH set point...
July 2016: Applied Microbiology and Biotechnology
Huang Rongbin, Xiang Lei, Liu Ying, Deng Xiangping, Cao Xuan, Li Lanfang, Yu Cuiyun, Chen Yanming, Tang Guotao
OBJECTIVES: To develop an acid trigger release of antitumour drug delivery carriers, pH-sensitive amphiphilic poly (ethyleneglycol)-imine-benzoic-dipalmitate (PEG-I-dC16 ) polymers were designed and synthesized and the drug-loaded micelles were evaluated in vitro. METHODS: PEG-I-dC16 synthesized by Schiff base synthetic method and characterized by (1) H-NMR. To determine the drug-loading capacity, doxorubicin (DOX) was encapsulated in the micelles using membrane dialysis method...
June 2016: Journal of Pharmacy and Pharmacology
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