| 1. |
- Cedergren, Jan, et al.
(author)
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Inducible nitric oxide synthase is expressed in synovial fluid granulocytes
- 2002
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In: Clinical Experimental Immunology. - : Oxford University Press (OUP). - 0009-9104 .- 1365-2249. ; 130, s. 150-155
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Journal article (peer-reviewed)abstract
- The objective of the study was to evaluate the NO-producing potential of synovial fluid (SF) cells. SF from 15 patients with arthritis was compared with blood from the same individuals and with blood from 10 healthy controls. Cellular expression of inducible nitric oxide synthase (iNOS) was analysed by flow cytometry. High-performance liquid chromatography was used to measure L-arginine and L-citrulline. Nitrite and nitrate were measured colourimetrically utilizing the Griess' reaction. Compared to whole blood granulocytes in patients with chronic arthritis, a prominent iNOS expression was observed in SF granulocytes (P < 0.001). A slight, but statistically significant, increase in iNOS expression was also recorded in lymphocytes and monocytes from SF. L-arginine was elevated in SF compared to serum (257 ± 78 versus 176 ± 65 µmol/l, P = 0.008), whereas a slight increase in L-citrulline (33 ± 11 versus 26 ± 9 µmol/l), did not reach statistical significance. Great variations but no significant differences were observed comparing serum and SF levels of nitrite and nitrate, respectively, although the sum of nitrite and nitrate tended to be elevated in SF (19.2 ± 20.7 versus 8,6 ± 6.5 µmol/l,P = 0.054). Synovial fluid leucocytes, in particular granulocytes, express iNOS and may thus contribute to intra-articular NO production in arthritis.
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| 2. |
- Cedergren, Jan, et al.
(author)
-
Inducible nitric oxide synthase (NOS II) is constitutive in human neutrophils
- 2003
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In: APMIS. - : Wiley. - 0903-4641 .- 1600-0463. ; 111:10, s. 963-968
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Journal article (peer-reviewed)abstract
- The objective was to study the expression of inducible nitric oxide synthase (NOS II) in and NO production by human blood neutrophils and in in vivo exudated neutrophils. Cellular expression of NOS II was evaluated by flow cytometry in whole blood, in isolated blood neutrophils, and in neutrophils obtained by exudation in vivo into skin chambers. Neutrophil NOS II was also demonstrated by Western blotting. Uptake of 3H-labelled L-arginine was studied in vitro and NOS activity measured in a whole cell assay by the conversion of 3H-arginine to 3H-citrulline. In contrast to unseparated blood cells, NOS II was demonstrable both in isolated blood neutrophils and exudated cells. The failure to detect NOS II by flow cytometry in whole blood cells thus proved to be due to the quenching effect of hemoglobin. Western blotting revealed a 130 kD band corresponding to NOS II in isolated blood neutrophils, but detection was dependent on diisopropylfluorophosphate for proteinase inhibition. L-arginine was taken up by neutrophils, but enzymatic activity could not be demonstrated. We conclude that human neutrophils constitutively express NOS II, but that its demonstration by FITC-labelling is inhibited by hemoglobin-mediated quenching in whole blood samples.
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| 3. |
- Forslund, Tony, 1956-, et al.
(author)
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Nitric Oxide Regulates the Aggregation of Stimulated Human Neutrophils
- 2000
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In: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 274:2, s. 482-487
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Journal article (peer-reviewed)abstract
- Neutrophil aggregation is mediated by both CD18 integrin and L-selectin. Nitric oxide attenuates the integrin-mediated adhesion of neutrophils to collagen and to endothelium and may therefore affect aggregation as well. FMLP-stimulated neutrophils exposed to -arginine showed increased and prolonged aggregation, whereas cells pretreated with L-NAME did not differ from FMLP-stimulated controls. Nitric oxide is known to induce ADP ribosylation of G-actin, which inhibits polymerization. We detected equivalent levels of total F-actin in cells pretreated with -arginine or L-NAME and non-pretreated controls. However, neutrophils pretreated with -arginine and stimulated by CD18 integrin cross-linking exhibited a more limited increase in total F-actin, compared to control and L-NAME-pretreated cells. Thus at least two signaling pathways may be involved FMLP-stimulated aggregation, mediated by CD18 integrins. More specifically, it is plausible that FMLP-receptor signaling upregulates CD18 integrins and endogenous NO subsequently modulates CD18-mediated signaling to prolong aggregation, possibly through ADP-ribosylation of actin.
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| 4. |
- Loitto, Vesa-Matti, et al.
(author)
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Neutrophil leukocyte motility requires directed water influx
- 2002
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In: Journal of Leukocyte Biology. - 0741-5400 .- 1938-3673. ; 71:2, s. 212-222
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Journal article (peer-reviewed)abstract
- The ability of neutrophils to sense and move to sites of infection is essential for our defense against pathogens. For motility, lamellipodium extension and stabilization are prerequisites, but how cells form such membrane protrusions is still obscure. Using contrast-enhanced video microscopy and Transwell® assays, we show that water-selective aquaporin channels regulate lamellipodium formation and neutrophil motility. Addition of anti-aquaporin-9 antibodies, HgCl2, or tetraethyl ammonium inhibited the function(s) of the channels and blocked motility-related shape changes. On human neutrophils, aquaporin-9 preferentially localized to the cell edges, where N-formyl peptide receptors also accumulated, as assessed with fluorescence microscopy. To directly visualize water fluxes at cell edges, cells were loaded with high dilution-sensitive, self-quenching concentrations of fluorophore. In these cells, motile regions always displayed increased fluorescence compared with perinuclear regions. Our observations provide the first experimental support for motility models where water fluxes play a pivotal role in cell-volume increases accompanying membrane extensions.
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