scholarly journals Uptake and intracellular activity of AM-1155 in phagocytic cells.

1996 ◽  
Vol 40 (12) ◽  
pp. 2756-2759 ◽  
Author(s):  
T Yamamoto ◽  
H Kusajima ◽  
M Hosaka ◽  
H Fukuda ◽  
Y Oomori ◽  
...  
Keyword(s):  
Cell Types ◽  
Intracellular Concentration ◽  
Nucleoside Transport ◽  
Metabolic Inhibitors ◽  
Phagocytic Cells ◽  
Penetration Process ◽  
Extracellular Concentration ◽  
Intracellular Activity ◽  
Human Polymorphonuclear Leukocytes

The uptake and intracellular activity of AM-1155 in murine J774.1 macrophages and human polymorphonuclear leukocytes were investigated. AM-1155 penetrated phagocytic cells rapidly and reversibly, although the penetration process was not affected by metabolic inhibitors such as sodium fluoride, cyanide m-chlorophenylhydrazone, or ouabain or by nucleoside transport system inhibitors such as adenosine. The intracellular concentration-to-extracellular concentration ratio of AM-1155 in both cell types of phagocytes ranged from 5 to 7. These ratios were almost equal to those for sparfloxacin. The intracellular activity of AM-1155 in J774.1 macrophages, examined with Staphylococcus aureus 209P as a test bacterium, was dependent on the extracellular concentration. AM-1155 at a concentration of 1 microgram/ml reduced the number of viable cells of S. aureus ingested by more than 90%. The intracellular activity of AM-1155 was more potent than those of sparfloxacin, ofloxacin, ciprofloxacin, flomoxef, and erythromycin. These results suggest that the potent intracellular activity of AM-1155 might mainly be due to the high intracellular concentration and its potent in vitro activity.

scholarly journals Uptake and intracellular activity of NM394, a new quinolone, in human polymorphonuclear leukocytes.

1996 ◽  
Vol 40 (3) ◽  
pp. 739-742 ◽  
Author(s):  
M Ozaki ◽  
K Komori ◽  
M Matsuda ◽  
R Yamaguchi ◽  
T Honmura ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Infection ◽  
Polymorphonuclear Leukocytes ◽  
Concentration Ratio ◽  
Extracellular Concentration ◽  
Intracellular Activity ◽  
Subsequent Elution ◽  
Active Transport System ◽  
Human Polymorphonuclear Leukocytes ◽  
Kinetics Of

The uptake of NM394, a new quinolone, by and its subsequent elution from human polymorphonuclear leukocytes were studied and compared with those of ofloxacin and ciprofloxacin. The kinetics of the uptake of NM394 was similar to that of ciprofloxacin. The maximum intracellular-to-extracellular concentration ratio was 12.3, compared with 8.6 for ciprofloxacin and 4.9 for ofloxacin at the extracellular concentration of 20 micrograms/ml. The elution of NM394 from human polymorphonuclear leukocytes occurs relatively slowly; 5 min after the removal of extracellular NM394, nearly 100% still remained in polymorphonuclear leukocytes, compared with ofloxacin, which was so rapidly eluted that only 12% remained. The uptake of NM394 was significantly decreased at 4 degrees C and by the presence of NaCN but was not affected by the presence of L-glycine, L-leucine, L-serine, adenosine, or NaF. NM394 showed intracellular activity at a concentration of 0.1 microgram/ml that significantly reduced the number of phagocytosed Pseudomonas aeruginosa cells with 2 h of incubation. These results suggest that uptake of NM394 by human polymorphonuclear leukocytes occurs via an active transport system differing from that of ofloxacin, whose uptake is affected by the presence of L-glycine and L-leucine, and that once accumulated, NM394 remains intracellularly active and participates in protection against bacterial infection.

scholarly journals Cellular Pharmacokinetics and Intracellular Activity of Gepotidacin againstStaphylococcus aureusIsolates with Different Resistance Phenotypes in Models of Cultured Phagocytic Cells

2018 ◽  
Vol 62 (4) ◽  
Author(s):  
Frédéric Peyrusson ◽  
Paul M. Tulkens ◽  
Françoise Van Bambeke
Keyword(s):  
Bacterial Infections ◽  
Maximum Effect ◽  
Cell Fractionation ◽  
Phagocytic Cells ◽  
Content Type ◽  
Cellular Pharmacokinetics ◽  
Extracellular Concentration ◽  
Infected Cells ◽  
Maximal Reduction

ABSTRACTGepotidacin (GSK2140944), a novel triazaacenaphthylene bacterial topoisomerase inhibitor, is currently in clinical development for the treatment of bacterial infections. This study examinedin vitroits activity against intracellularStaphylococcus aureus(involved in the persistent character of skin and skin structure infections) by use of a pharmacodynamic model and in relation to cellular pharmacokinetics in phagocytic cells. Compared to oxacillin, vancomycin, linezolid, daptomycin, azithromycin, and moxifloxacin, gepotidacin was (i) more potent intracellularly (the apparent bacteriostatic concentration [Cs] was reached at an extracellular concentration about 0.7× its MIC and was not affected by mechanisms of resistance to the comparators) and (ii) caused a maximal reduction of the intracellular burden (maximum effect) of about −1.6 log10CFU (which was better than that caused by linezolid, macrolides, and daptomycin and similar to that caused by moxifloxacin). After 24 h of incubation of infected cells with antibiotics at 100× their MIC, the intracellular persisting fraction was <0.1% with moxifloxacin, 0.5% with gepotidacin, and >1% with the other drugs. The accumulation and efflux of gepotidacin in phagocytes were very fast (kinandkout, ∼0.3 min−1; the plateau was reached within 15 min) but modest (intracellular concentration-to-extracellular concentration ratio, ∼1.6). In cell fractionation studies, about 40 to 60% of the drug was recovered in the soluble fraction and ∼40% was associated with lysosomes in uninfected cells. In infected cells, about 20% of cell-associated gepotidacin was recovered in a sedimentable fraction that also contained bacteria. This study highlights the potential for further study of gepotidacin to fight infections where intracellular niches may play a determining role in bacterial persistence and relapses.

scholarly journals ARHGAP25, a novel Rac GTPase-activating protein, regulates phagocytosis in human neutrophilic granulocytes

Blood ◽  
2012 ◽  
Vol 119 (2) ◽  
pp. 573-582 ◽  
Author(s):  
Roland Csépányi-Kömi ◽  
Gábor Sirokmány ◽  
Miklós Geiszt ◽  
Erzsébet Ligeti
Keyword(s):  
Actin Cytoskeleton ◽  
Rho Gtpases ◽  
Active Form ◽  
Cell Types ◽  
Small Gtpases ◽  
Negative Regulator ◽  
Phagocytic Cells ◽  
Rac Gtpase

Members of the Rac/Rho family of small GTPases play an essential role in phagocytic cells in organization of the actin cytoskeleton and production of toxic oxygen compounds. GTPase-activating proteins (GAPs) decrease the amount of the GTP-bound active form of small GTPases, and contribute to the control of biologic signals. The number of potential Rac/RhoGAPs largely exceeds the number of Rac/Rho GTPases and the expression profile, and their specific role in different cell types is largely unknown. In this study, we report for the first time the properties of full-length ARHGAP25 protein, and show that it is specifically expressed in hematopoietic cells, and acts as a RacGAP both in vitro and in vivo. By silencing and overexpressing the protein in neutrophil model cell lines (PLB-985 and CosPhoxFcγR, respectively) and in primary macrophages, we demonstrate that ARHGAP25 is a negative regulator of phagocytosis acting probably via modulation of the actin cytoskeleton.

scholarly journals Pseudomonas aeruginosa Cytotoxin ExoU Is Injected into Phagocytic Cells during Acute Pneumonia

2010 ◽  
Vol 78 (4) ◽  
pp. 1447-1456 ◽  
Author(s):  
Maureen H. Diaz ◽  
Alan R. Hauser
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Immune Cells ◽  
Type Iii Secretion ◽  
Cell Types ◽  
Secretion System ◽  
Host Cells ◽  
Phagocytic Cells ◽  
Acute Pneumonia

ABSTRACT ExoU, a cytotoxin translocated into host cells via the type III secretion system of Pseudomonas aeruginosa, is associated with increased mortality and disease severity. We previously showed that impairment of recruited phagocytic cells allowed survival of ExoU-secreting P. aeruginosa in the lung. Here we analyzed types of cells injected with ExoU in vivo using translational fusions of ExoU with a β-lactamase reporter (ExoU-Bla). Cells injected with ExoU-Bla were detectable in vitro but not in vivo, presumably due to the rapid cytotoxicity induced by the toxin. Therefore, we used a noncytotoxic ExoU variant, designated ExoU(S142A)-Bla, to analyze injection in vivo. We determined that phagocytic cells in the lung were frequently injected with ExoU(S142A). Early during infection, resident macrophages constituted the majority of cells into which ExoU was injected, but neutrophils and monocytes became the predominant types of cells into which ExoU was injected upon recruitment into the lung. We observed a modest preference for injection into neutrophils over injection into other cell types, but in general the repertoire of injected immune cells reflected the relative abundance of these cells in the lung. Our results indicate that phagocytic cells in the lung are injected with ExoU and support the hypothesis that ExoU-mediated impairment of phagocytes has a role in the pathogenesis of pneumonia caused by P. aeruginosa.

scholarly journals Differentiation stimuli induce receptors for plasma fibronectin on the human myelomonocytic cell line HL-60

Blood ◽  
1984 ◽  
Vol 64 (4) ◽  
pp. 858-866
Author(s):  
CG Pommier ◽  
J O'Shea ◽  
T Chused ◽  
T Takahashi ◽  
M Ochoa ◽  
...  
Keyword(s):  
Cell Line ◽  
Human Peripheral Blood ◽  
Cell Types ◽  
Plasma Fibronectin ◽  
Blood Monocytes ◽  
Before And After ◽  
Differentiating Agents ◽  
Membrane Antigens ◽  
Human Polymorphonuclear Leukocytes

Plasma fibronectin (Fn) induces phagocytosis of C3b-opsonized sheep erythrocytes (EC3b) by human peripheral blood monocytes. However, Fn does not induce erythrophagocytosis of EC3b by human polymorphonuclear leukocytes (PMN), unless the PMN have been exposed to C5a or N-formyl- methionyl-leucyl-phenylalanine. Because of this difference, it is of great interest to examine Fn binding to cells that possess the capacity to differentiate into either granulocytes or monocytes. Hence, we have examined the consequences of Fn binding to the human myelomonocytic cell line, HL-60, both before and after in vitro differentiation of the HL-60, along a monocytoid or a granulocytoid pathway. Fn receptors were not found on undifferentiated HL-60, but several differentiating agents promoted the HL-60 binding of Fn-coated microspheres (Fn-ms). The peak of Fn-ms binding occurred four to five days after the induction of differentiation with dimethylsulfoxide (DMSO), and two days after induction by PMA. In addition, cells that differentiated along either the monocytoid or the granulocytoid pathway showed a marked increase in the phagocytosis of both IgG-coated erythrocytes (EA) and EC3b when they were exposed to Fn. Comparison of the effects of anti-Fn monoclonals on the binding of Fn-ms to the monocytes, PMN, and HL-60 showed that the same monoclonals block Fn-ms-binding and Fn-induced EC3b phagocytosis by all three cell types. Two monoclonal antibodies, M1/70 and A6F10, directed against membrane antigens on PMN and monocytes, inhibited Fn-ms binding. Both also blocked Fn-induced EC3b ingestion by these cells. However, neither antibody blocked Fn-ms binding or EC3b ingestion by differentiated HL-60. We conclude that differentiated HL-60 cells express functionally active Fn receptors, similar to monocytes and activated PMN, which, nonetheless, differ from normal cells in their association with the antigens recognized by M1/70 and A6F10.

scholarly journals Intracellular Penetration and Activity of DX-619 in Human Polymorphonuclear Leukocytes

2006 ◽  
Vol 50 (9) ◽  
pp. 3173-3174 ◽  
Author(s):  
Isabel García ◽  
Sofía Ballesta ◽  
Concepción Murillo ◽  
Evelio J. Perea ◽  
Álvaro Pascual
Keyword(s):  
Staphylococcus Aureus ◽  
Polymorphonuclear Leukocytes ◽  
Environmental Temperature ◽  
Metabolic Inhibitors ◽  
Intracellular Activity ◽  
Intracellular Concentrations ◽  
Human Polymorphonuclear Leukocytes

ABSTRACT The intracellular penetration and activity of DX-619 in human polymorphonuclear leukocytes have been evaluated. DX-619 reached intracellular concentrations 10 times higher than the extracellular concentrations reached. Uptake was rapid, reversible, nonsaturable, and affected by environmental temperature, some metabolic inhibitors, and a soluble membrane activator. DX-619 showed intracellular activity against Staphylococcus aureus.

scholarly journals Differentiation stimuli induce receptors for plasma fibronectin on the human myelomonocytic cell line HL-60

Blood ◽  
1984 ◽  
Vol 64 (4) ◽  
pp. 858-866 ◽  
Author(s):  
CG Pommier ◽  
J O'Shea ◽  
T Chused ◽  
T Takahashi ◽  
M Ochoa ◽  
...  
Keyword(s):  
Cell Line ◽  
Human Peripheral Blood ◽  
Cell Types ◽  
Plasma Fibronectin ◽  
Blood Monocytes ◽  
Before And After ◽  
Differentiating Agents ◽  
Membrane Antigens ◽  
Human Polymorphonuclear Leukocytes

Abstract Plasma fibronectin (Fn) induces phagocytosis of C3b-opsonized sheep erythrocytes (EC3b) by human peripheral blood monocytes. However, Fn does not induce erythrophagocytosis of EC3b by human polymorphonuclear leukocytes (PMN), unless the PMN have been exposed to C5a or N-formyl- methionyl-leucyl-phenylalanine. Because of this difference, it is of great interest to examine Fn binding to cells that possess the capacity to differentiate into either granulocytes or monocytes. Hence, we have examined the consequences of Fn binding to the human myelomonocytic cell line, HL-60, both before and after in vitro differentiation of the HL-60, along a monocytoid or a granulocytoid pathway. Fn receptors were not found on undifferentiated HL-60, but several differentiating agents promoted the HL-60 binding of Fn-coated microspheres (Fn-ms). The peak of Fn-ms binding occurred four to five days after the induction of differentiation with dimethylsulfoxide (DMSO), and two days after induction by PMA. In addition, cells that differentiated along either the monocytoid or the granulocytoid pathway showed a marked increase in the phagocytosis of both IgG-coated erythrocytes (EA) and EC3b when they were exposed to Fn. Comparison of the effects of anti-Fn monoclonals on the binding of Fn-ms to the monocytes, PMN, and HL-60 showed that the same monoclonals block Fn-ms-binding and Fn-induced EC3b phagocytosis by all three cell types. Two monoclonal antibodies, M1/70 and A6F10, directed against membrane antigens on PMN and monocytes, inhibited Fn-ms binding. Both also blocked Fn-induced EC3b ingestion by these cells. However, neither antibody blocked Fn-ms binding or EC3b ingestion by differentiated HL-60. We conclude that differentiated HL-60 cells express functionally active Fn receptors, similar to monocytes and activated PMN, which, nonetheless, differ from normal cells in their association with the antigens recognized by M1/70 and A6F10.

scholarly journals STUDY ON GROWTH OF RICKETTSIAE

1959 ◽  
Vol 109 (3) ◽  
pp. 271-292 ◽  
Author(s):  
Zanvil A. Cohn ◽  
F. Marilyn Bozeman ◽  
Janis M. Campbell ◽  
James W. Humphries ◽  
Thomas K. Sawyer
Keyword(s):  
Tissue Culture ◽  
Divalent Cations ◽  
Biochemical Properties ◽  
Metabolic Inhibitors ◽  
Penetration Process ◽  
Factors Affecting ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Fluid Environment

A system has been described in which the penetration of Rickettsia tsutsugamushi (Karp strain) into tissue culture cells can be quantitated, and the factors affecting this process studied. The results indicated that rickettsial penetration in vitro depended largely on the viability of the organisms. Certain components of the fluid environment such as the divalent cations and protein were found to be of importance. The temperature dependence of the penetration process was found to vary with the nature of the suspending medium. A number of compounds related to L-glutamic acid enhanced penetration, whereas metabolic inhibitors depressed this process. Aureomycin at concentrations between 50 and 250 µg./ml. inhibited the penetration of rickettsiae while chloramphenicol at similar concentrations was ineffective. The results are discussed in terms of the biological and biochemical properties of this group of agents.

scholarly journals Sensitivity of Acute Leukemia Cells to Cytarabine Is a Correlate of Cellular es Nucleoside Transporter Site Content Measured by Flow Cytometry With SAENTA-Fluorescein

Blood ◽  
1997 ◽  
Vol 90 (1) ◽  
pp. 346-353 ◽  
Author(s):  
Wendy P. Gati ◽  
Alan R.P. Paterson ◽  
Loree M. Larratt ◽  
A. Robert Turner ◽  
Andrew R. Belch
Keyword(s):  
Flow Cytometry ◽  
Acute Leukemia ◽  
Cell Types ◽  
Transport Processes ◽  
Nucleoside Transport ◽  
Leukemia Cells ◽  
Nucleoside Transporter ◽  
Flow Cytometric Method ◽  
Ic50 Values

Abstract Cytarabine (araC) is converted to araC 5′-triphosphate after entering leukemia cells as a substrate for nucleoside transport processes. This study tested the relationship between araC cytotoxicity, measured in an in vitro tetrazolium dye reduction assay of cell viability, and the cellular abundance of es nucleoside transport elements, assayed by a flow cytometric method that used the es-specific stain, 5-(SAENTA-x8)-fluorescein (5-(Sx8)-F), in cultured leukemia cells and in myeloblasts and lymphoblasts (blasts) from leukemia patients. Cellular es site abundance (Bmax value for 5-(Sx8)-F binding) varied sixfold among nine leukemic myeloblast samples from patients. In cultured OCI/AML-2 myeloblasts and CCRF-CEM T-lymphoblasts, and in fresh leukemic blasts, es sites were fractionally blocked by treatment with graded concentrations of nitrobenzylthioinosine (NBMPR), an inhibitory es site ligand, to simulate the variation in es expression found in leukemic blasts from patients with acute myeloid leukemia. When the cytotoxicity of a single concentration of araC was determined in NBMPR-treated leukemia cells, cell kill correlated closely with the intensity of 5-(Sx8)-F fluorescence (r = .92 to .99), a measure of the cell surface abundance of functional es nucleoside transporter sites. Concentrations of NBMPR that achieved half-maximal reduction (4.3 to 12 nmol/L) of cellular 5-(Sx8)-F fluorescence (measured by flow cytometry) approximated IC50 values (1 to 10 nmol/L) previously found for inhibition by NBMPR of es-mediated nucleoside fluxes in several cell types, supporting the view that 5-(Sx8)-F interacted with the es transporter. The correlation of araC cytotoxicity and the Bmax for 5-(Sx8)-F binding to es sites in cultured leukemia cells and in leukemic blasts from acute leukemia patients (r = .95) suggests that the flow cytometry assay of es capacity may be useful in predicting clinical response to araC.

scholarly journals Uptake and intracellular activity of trovafloxacin in human phagocytes and tissue-cultured epithelial cells.

1997 ◽  
Vol 41 (2) ◽  
pp. 274-277 ◽  
Author(s):  
A Pascual ◽  
I García ◽  
S Ballesta ◽  
E J Perea
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Polymorphonuclear Leukocytes ◽  
Peritoneal Macrophages ◽  
Extracellular Concentration ◽  
Opsonized Zymosan ◽  
Intracellular Activity ◽  
Cultured Epithelial Cells ◽  
Energy Dependent ◽  
Human Polymorphonuclear Leukocytes

The penetration of trovafloxacin into human polymorphonuclear leukocytes (PMNs), human peritoneal macrophages, and tissue-cultured epithelial cells (McCoy cells) was evaluated. The cellular concentration to extracellular concentration (C/E) ratios of trovafloxacin were greater than 9 for extracellular concentrations ranging from 0.5 to 25 micrograms/ml. The uptake of trovafloxacin by PMNs was rapid, reversible, nonsaturable, not energy dependent, and significantly increased at 4 degrees C. Ingestion of opsonized zymosan, but not opsonized Staphylococcus aureus, significantly increased the amount of PMN-associated trovafloxacin. This agent at concentrations of 0.5 and 1 microgram/ml induced a greater reduction in the survival of intracellular S. aureus in PMNs than ciprofloxacin and ofloxacin. It was concluded that trovafloxacin reaches concentrations within phagocytic and nonphagocytic cells several times higher than the extracellular ones, while it remains active in PMNs.

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