Neutrophil function in whole blood and after purification: Changes in receptor expression, oxidase activity and responsiveness to cytokines

1992 ◽  
Vol 12 (2) ◽  
pp. 123-133 ◽  
Author(s):  
Fiona Watson ◽  
John J. Robinson ◽  
Steven W. Edwards
Keyword(s):  
Whole Blood ◽  
Neutrophil Function ◽  
Receptor Expression ◽  
Complement Receptors ◽  
Step Procedure ◽  
Plasma Membrane Receptor ◽  
One Step ◽  
Reactive Oxidants ◽  
In Cells

Neutrophil function and plasma membrane receptor expression was measured in cell suspensions isolated by two separate procedures and in unfractionated whole blood. When cells were prepared by a combined dextran/ficoll procedure, their ability to generate reactive oxidants in response to fMet-Leu-Phe was greater than in corresponding cells isolated by a one-step procedure on Mono-Poly Resolving Medium (M-PRM). Cells prepared by both methods could be primed in vitro by rGM-CSF, but the priming ratio was greater in cells prepared by the latter method. The ability of neutrophils in whole blood to generate reactive oxidants in response to fMet-Leu-Phe was extremely low, but this was increased by more than 10 fold if the blood was pre-incubated with rGM-CSF. Similarly, expression of CD 11b and CD 16 was very low (or undetectable) in neutrophils in whole blood, but this was rapidly increased upon priming. Activation by PMA resulted in a down regulation of CD 16 expression as the receptor was shed from the cell surface. Neutrophils isolated by either the dextran/ficoll or the M-PRM method showed increased expression of receptors compared with those in whole blood, although this expression was lower in cells isolated by the latter method. These data indicate that the isolation procedures used to obtain purified neutrophils prime both receptor expression and oxidase function, although these effects are minimalised in isolation procedures using M-PRM. Furthermore, as CD 16 expression on neutrophils in whole blood is rapidly up-regulated during priming, it seems likely that, as for complement receptors, rapidly-mobilisable intracellular stores of this receptor exist.

Implantable insulin pumps: diagnosis and management of decreased flow rates using radionuclide investigation

1996 ◽  
Vol 19 (12) ◽  
pp. 723-729
Author(s):  
H. Boulahdour ◽  
A. Behar ◽  
M.-J. Haardt ◽  
J-L. Selam
Keyword(s):  
Insulin Pumps ◽  
Distal Catheter ◽  
Non Invasive ◽  
Step Procedure ◽  
Catheter Obstruction ◽  
Catheter Malfunction ◽  
Invasive Method ◽  
One Step ◽  
Isotonic Sodium Chloride

The aim of this study was to develop a diagnostic procedure for pumping unit malfunction by radionuclide imaging (RI) and to validate the method by comparing the results with those obtained using more conventional methods. Fifteen radionuclide investigations were performed in 11 patients with intraperitoneal implantable insulin pumps. One mCi of 99 mTc in 1 ml isotonic sodium chloride was injected into the reservoir. The results based on catheter visualization and peritoneal accumulation were compared blindly to the efficacy of alkaline rinses and laparoscopic findings. In all RI stoppage cases except one alkaline rinses failed to restore flow. Where laparoscopy was performed, comparisons were concordant i.e. no outflow from the tip of the catheter. The RI images obtained were reproduced in vitro using a pump under normal flow conditions and complete proximal and distal catheter obstruction. RI is a safe, quick non invasive method which allows the location of the site of pump/catheter malfunction within a one step procedure and the prediction of the efficacy of sodium hydroxide rinses.

scholarly journals Correlation of α-Fetoprotein Expression in Normal Hepatocytes during Development with Tyrosine Phosphorylation and Insulin Receptor Expression

1998 ◽  
Vol 9 (5) ◽  
pp. 1093-1105 ◽  
Author(s):  
Leila Khamzina ◽  
Pierre Borgeat
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Receptor Expression ◽  
Regulatory Subunit ◽  
Β Subunit ◽  
Phosphatidylinositol 3 Kinase ◽  
Fetal Hepatocytes ◽  
In Cells ◽  
Phosphatidylinositol 3

The molecular mechanism of hepatic cell growth and differentiation is ill defined. In the present study, we examined the putative role of tyrosine phosphorylation in normal rat liver development and in an in vitro model, the α-fetoprotein-producing (AFP+) and AFP-nonproducing (AFP−) clones of the McA-RH 7777 rat hepatoma. We demonstrated in vivo and in vitro that the AFP+ phenotype is clearly associated with enhanced tyrosine phosphorylation, as assessed by immunoblotting and flow cytometry. Moreover, immunoprecipitation of proteins with anti-phosphotyrosine antibody showed that normal fetal hepatocytes expressed the same phosphorylation pattern as stable AFP+clones and likewise for adult hepatocytes and AFP− clones. The tyrosine phosphorylation of several proteins, including the β-subunit of the insulin receptor, insulin receptor substrate-1, p85 regulatory subunit of phosphatidylinositol-3-kinase, andras-guanosine triphosphatase-activating protein, was observed in AFP+ clones, whereas the same proteins were not phosphorylated in AFP− clones. We also observed that fetal hepatocytes and the AFP+ clones express 4 times more of the insulin receptor β-subunit compared with adult hepatocytes and AFP− clones and, accordingly, that these AFP+clones were more responsive to exogenous insulin in terms of protein tyrosine phosphorylation. Finally, growth rate in cells of AFP+ clones was higher than that measured in cells of AFP− clones, and inhibition of phosphatidylinositol-3-kinase by LY294002 and Wortmannin blocked insulin- and serum-stimulated DNA synthesis only in cells of AFP+ clones. These studies provide evidences in support of the hypothesis that signaling via insulin prevents hepatocyte differentiation by promoting fetal hepatocyte growth.

Embryo Rescue Culture Medium for Improved Synthesis of 4x Wheat × Rye Hybrids

1994 ◽  
Vol 42 (4) ◽  
pp. 493
Author(s):  
CH Balatero ◽  
NL Darvey
Keyword(s):  
Embryo Rescue ◽  
Favourable Effect ◽  
Embryo Formation ◽  
Step Procedure ◽  
Hybrid Embryo ◽  
One Step ◽  
Well Differentiated ◽  
Semi Solid ◽  
Optimum Recovery

The cross-incompatibility barrier between 4x wheat and rye has limited the genetic base for triticale breeding. Experiments designed to improve the synthesis of wheat-rye amphihaploids were conducted. The effects of 2,4-D on crossability and 3x hybrid embryo differentiation, and the influence of one-step and two-step media on the culture of immature 3x embryos in vitro, were investigated. Application of 10 mg L-1 2,4-D slightly improved seed set but significantly reduced the frequency of normal embryos. In contrast to the reported favourable effect of 2,4-D on haploid embryo formation in wheat × maize crosses, the application of 2,4-D in the present study offers no real advantage on amphihaploid embryo formation from 4x wheat × rye crosses. For small and immature wheat-rye hybrid (3x) embryos, optimum recovery in vitro was obtained via a two-step procedure consisting of a semi-solid MN medium followed by MS medium supplemented with IAA (1 mg L-1) and BAP (1 mg L-1). For bigger and well-differentiated embryos, the use of a one-step Gamborg's B5 medium was sufficient.

scholarly journals Immunomodulatory effects of heat-killed Mycobacterium obuense on human blood dendritic cells

2017 ◽  
Vol 23 (7) ◽  
pp. 592-605 ◽  
Author(s):  
Samer Bazzi ◽  
Helmout Modjtahedi ◽  
Satvinder Mudan ◽  
Marcel Achkar ◽  
Charles Akle ◽  
...  
Keyword(s):  
Mhc Class Ii ◽  
Human Blood ◽  
Whole Blood ◽  
Class Ii ◽  
Receptor Expression ◽  
Immunomodulatory Effects ◽  
Total Blood ◽  
Surface Receptors ◽  
Surface Receptor

Heat-killed (HK) Mycobacterium obuense is a novel immunomodulator, currently undergoing clinical evaluation as an immunotherapeutic agent in the treatment of cancer. Here, we examined the effect of in vitro exposure to HK M. obuense on the expression of different categories of surface receptors on human blood myeloid (m) and plasmacytoid (p) DCs. Moreover, we have characterized the cytokine and chemokine secretion patterns of purified total blood DCs stimulated with HK M. obuense. HK M. obuense significantly up-regulated the expression of CD11c, CD80, CD83, CD86, CD274 and MHC class II in whole-blood mDCs and CD80, CD123 and MHC class II in whole-blood pDCs. Down-regulation of CD195 expression in both DC subpopulations was also noted. Further analysis showed that HK M. obuense up-regulated the expression of CD80, CD83 and MHC class II on purified blood DC subpopulations. TLR2 and TLR1 were also identified to be engaged in mediating the HK M. obuense-induced up-regulation of surface receptor expression on whole blood mDCs. In addition, our data demonstrated that HK M. obuense augmented the secretion of CCL4, CCL5, CCL22, CXCL8, IL-6, IL-12p40 and TNF-α by purified total blood DCs. Taken together, our data suggest that HK M. obuense exerts potent differential immunomodulatory effects on human DC subpopulations.

Ultrastructure of bovine in vitro-produced blastocysts cryopreserved by vitrification

Zygote ◽  
1998 ◽  
Vol 6 (1) ◽  
pp. 17-26 ◽  
Author(s):  
Shigeki Ohboshi ◽  
Noboru Fujihara ◽  
Tatsuyuki Yoshida ◽  
Hiroshi Tomagane
Keyword(s):  
Ethylene Glycol ◽  
Cellular Damage ◽  
Bovine Embryos ◽  
Step Procedure ◽  
Direct Exposure ◽  
Subsequent Exposure ◽  
One Step ◽  
The One ◽  
Serious Injuries

SummaryThe objective of this study was to examine ultrastructural aspects of bovine in vitro-produced blastocysts associated with cryopreservation by vitrification. Morphologically good embryos were used and treated with ethylene-glycol-based vitrification solution (VS). The untreated embryos had conventional fine structure. The post-warming embryos treated with direct exposure to VS (one-step procedure) showed cellular damage structurally by cryopreservation, which included loss of microvilli, disruption of the plasma membrane, mitochondrial changes and swelling of the endoplasmic reticulum. However, nuclei and junctional regions seemed to be resistant to cryoinjury. In contrast, the post-warming embryos pre-equilibra ted with 10% ethylene glycol for 5 min and subsequent exposure to VS (two-step procedure) showed less damage than those treated by the one-step procedure. Post-warming embryos treated by the two-step procedure were cultured in vitro for 18 h. Some embryos survived and their structures re-formed to the former state, while other embryos showed serious injuries and could not reconstitute the blastocoele. Three post-warming embryos treated by the two-step procedure that survived after in vitro culture were transferred to three recipients and one of these resulted in pregnancy. These results indicate that cryopreservation by vitrification can damage membranous structures of the cells of bovine embryos, the extent and nature of this damage being dependent on the vitrification procedure.

Carbon dots obtained using hydrothermal treatment of formaldehyde. Cell imaging in vitro

Nanoscale ◽  
2014 ◽  
Vol 6 (15) ◽  
pp. 9071-9077 ◽  
Author(s):  
M. Algarra ◽  
M. Pérez-Martín ◽  
M. Cifuentes-Rueda ◽  
J. Jiménez-Jiménez ◽  
J. C. G. Esteves da Silva ◽  
...  
Keyword(s):  
Hydrothermal Treatment ◽  
Carbon Dots ◽  
Cell Imaging ◽  
Step Procedure ◽  
One Step

Highly photoluminescent carbon dots have been prepared in a one step procedure by hydrothermal treatment of formaldehyde at 180 °C.

Human monocytes express functional receptors for granulocyte colony– stimulating factor that mediate suppression of monokines and interferon-γ

Blood ◽  
2000 ◽  
Vol 95 (1) ◽  
pp. 270-276 ◽  
Author(s):  
Eva-Maria Boneberg ◽  
Lars Hareng ◽  
Florian Gantner ◽  
Albrecht Wendel ◽  
Thomas Hartung
Keyword(s):  
Flow Cytometry ◽  
Whole Blood ◽  
Receptor Expression ◽  
Human Neutrophils ◽  
Enzyme Linked Immunosorbent Assay ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Ifn Γ ◽  
Stimulating Factor

In a double-blind, placebo-controlled, randomized study, 10 healthy men received either a single dose of 480 μg granulocyte colony-stimulating factor (G-CSF) or saline. Blood taken from the volunteers was stimulated with 10 μg/mL endotoxin and released cytokines were measured by enzyme-linked immunosorbent assay. Expression of G-CSF receptors on leukocytes was examined by flow cytometry and reverse transcriptase-polymerase chain reaction. Functional activity of these receptors was tested by challenging isolated leukocyte populations to release cytokines with endotoxin in the presence of G-CSF. The G-CSF treatment attenuated the release of the proinflammatory cytokines tumor necrosis factor (TNF)-, interleukin (IL)-12, IL-1β, and interferon (IFN)-γ in ex vivo lipopolysaccharide (LPS)-stimulated whole blood. In blood from untreated volunteers the presence of G-CSF in vitro also attenuated the LPS-stimulated release of these cytokines. G-CSF in vitro also attenuated TNF- release from elutriation-purified monocytes. In the presence of 10 ng/mL recombinant TNF-, the attenuation of LPS-inducible IFN-γ release by G-CSF was blunted in whole blood. However, G-CSF had no such effect on IFN-γ release from isolated lymphocytes stimulated with anti-CD3 or a combination of TNF- and IL-12. G-CSF receptor expression was detected in human neutrophils and monocytes but not in lymphocytes by means of RT-PCR as well as flow cytometry. These results indicate that G-CSF receptors expressed on monocytes are functional in modulating monokine release. We conclude that the attenuation of IFN-γ release from lymphocytes is not a direct effect of G-CSF on these cells but is rather due to the inhibition of monocytic IL-12 and TNF- release by G-CSF. (Blood. 2000;95:270-276)

scholarly journals Expansion of Submucosal Bladder Wall TissueIn VitroandIn Vivo

2016 ◽  
Vol 2016 ◽  
pp. 1-9
Author(s):  
Gisela Reinfeldt Engberg ◽  
Clara Ibel Chamorro ◽  
Agneta Nordenskjöld ◽  
Magdalena Fossum
Keyword(s):  
Smooth Muscle ◽  
Bladder Wall ◽  
Subcutaneous Fat ◽  
Culture Conditions ◽  
Detrusor Muscle ◽  
Step Procedure ◽  
Standard Culture ◽  
One Step

In order to develop autologous tissue engineering of the whole wall in the urinary excretory system, we studied the regenerative capacity of the muscular bladder wall. Smooth muscle cell expansion on minced detrusor musclein vitroandin vivowith or without urothelial tissue was studied. Porcine minced detrusor muscle and urothelium were culturedin vitrounder standard culture conditions for evaluation of the explant technique and in collagen for tissue sectioning and histology. Autografts of minced detrusor muscle with or without minced urothelium were expanded on 3D cylinder moulds by grafting into the subcutaneous fat of the pig abdominal wall. Moulds without autografts were used as controls. Tissue harvesting, mincing, and transplantation were performed as a one-step procedure. Cells from minced detrusor muscle specimens migrated and expandedin vitroon culture plastic and in collagen.In vivostudies with minced detrusor autografts demonstrated expansion and regeneration in all specimens. Minced urothelium autografts showed multilayered transitional urothelium when transplanted alone but not in cotransplantation with detrusor muscle; thus, minced bladder mucosa was not favored by cografting with minced detrusor. No regeneration of smooth muscle or epithelium was seen in controls.

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