• The Korean Journal of Physiology and Pharmacology
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• v.8 no.1
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• pp.43-50
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• 2004

Ischemia followed by reperfusion in the presence of polymorphonuclear leukocytes (PMNs) results in a marked cardiac contractile dysfunction. Amrinone, a specific inhibitor of phosphodiesterase 3, has an antioxidant activity against PMNs. Therefore, we hypothesized that amrinone could attenuate PMNs-Induced cardiac dysfunction by suppression of reactive oxygen species (ROS) produced fby PMNs. In the present study, we examined the effects of amrinone on isolated ischemic (20 min) and reperfused (45 min) rat hearts perfused with PMNs. Amrinone at $25\;{\mu}M$, given to hearts during the first 5 min of reperfusion, significantly improved coronary flow, left ventricular developed pressure (P<0.001), and the maximal rate of development of left ventricular developed pressure (P<0.001), compared with ischemic/reperfused hearts perfused with PMNs in the absence of amrinone. In addition, amrinone significantly reduced myeloperoxidase activity by 50.8%, indicating decreased PMNs infiltration (p< 0.001). Superoxide radical and hydrogen peroxide production were also significantly reduced in fMLP- and PMA-stimulated PMNs pretreated with amrinone. Hydroxyl radical was scavenged by amrinone. fMLP-induced elevation of $[Ca^{2+}]_i$ was also inhibited by amrinone. These results provide evidence that amrinone can significantly attenuate PMN-induced cardiac contractile dysfunction in the ischemic/reperfused rat heart via attenuation of PMNs infiltration into the myocardium and suppression of ROS release by PMNs.

• Journal of Veterinary Clinics
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• v.34 no.2
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• pp.70-75
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• 2017

Fucoidan increases the chemotactic activity of peripheral blood polymorphonuclear cells (PMNs) through interleukin (IL)-8 produced by peripheral blood mononuclear cells (PBMCs). It has been demonstrated that fucoidan can regulate the chemotaxis of PMNs by activating F-actin polymerization. The objectives of this study are to investigate the direct effect of fucoidan on the chemotaxis of porcine PMNs and to examine whether this effect is associated with changes in phosphoinositide 3-kinase (PI3K) activity. The chemotactic activity of porcine PMNs was evaluated by modified Boyden chamber assay. Akt phosphorylation activity, a main downstream of PI3K, was measured by Western blotting assay. Fucoidan itself has no chemoattractant effect for PMNs. However, direct treatment of PMNs with fucoidan showed higher chemotactic activity to porcine recombinant (pr) IL-8 than that of PMNs without fucoidan. The increased chemotactic activity of fucoidan-treated PMNs to pr IL-8 was suppressed by treatment of wortmannin, an inhibitor of PI3K. Treatment of PMNs with fucoidan also increased Akt phosphorylation level. This increase was also suppressed by wortmannin. These results suggested that fucoidan can upregulate chemotactic activity of porcine PMNs to IL-8, which is associated with PI3K activation.

• Journal of Veterinary Clinics
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• v.39 no.2
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• pp.51-58
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• 2022

Quercetin, a flavonoid found in fruits and vegetables, exhibits a strong anti-inflammatory activity. The objective of this study was to examine the effect of quercetin on chemotactic activity of peripheral blood polymorphonuclear cells (PMNs) to culture supernatant from peripheral blood mononuclear cells (PBMCs) stimulated with lipopolysaccharide (LPS). In addition, we determined whether this effect is related to interleukin (IL)-8 and changes in cytoskeleton. The chemotactic activity of PMNs was evaluated by a modified Boyden chamber assay. Total cellular filamentous (F)-actin levels were measured by method of fluorescence microscopy. The levels of IL-8 mRNA and protein were measured by real time polymerase reaction method and enzyme-linked immunosorbent assay, respectively. Quercetin (0-50 µM) itself has no chemoattractant effect for PMNs. The culture supernatant from PBMCs (2 × 10⁶ cells/mL) treated with LPS (1 ㎍/mL) showed remarkable increase in chemotaxis of PMNs. However, this effect was reduced dose-dependently by treatment with quercetin. In addition, PBMCs treated with LPS revealed enhanced levels in IL-8 protein and mRNA. Co-treatment of LPS with quercetin (50 µM) in PBMCs decreased IL-8 production and expression. Treatment of quercetin (0-50 µM) on PMNs to rpIL-8 (10 nM) decreased dose-dependently the chemotactic activity of PMNs. Treatment of quercetin on PMNs to IL-8 also reduced their total cellular F-actin level. These results suggested that quercetin attenuates chemotactic activity of PMNs, which is mediated by down-regulation of IL-8 production from LPS-stimulated PBMCs and inhibition of F-actin polymerization in PMNs.

• Biomolecules & Therapeutics
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• v.1 no.2
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• pp.131-136
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• 1993

In the present study, the effect of $HgCl_2$on the function of human peripheral polymorphonuclear leukocytes(PMNs) was examined. PMNs were isolated from human peripheral blood with density centrifugation in Ficoll-Paque. The cells were then incubated with $0.5{\sim}5{\mu}M\;HgCl_2$and glass adherence, chemotactic activity and erythrocyte-antibody rosette forming activity were measured. $HgCl_2$ decreased the function of PMNs in all three aspects tested. $HgCl_2$significantly diminished glass adherence(40.5 {\mu}M: 92{\pm}12%$ (percentage of control, $mean{\pm}$ S.D.); 41 {\mu}M: 46{\pm}11%,$ P<0.01; $3{\mu}M: 35{\pm}7%,$P<0.01;$5{\mu}M:49{\pm}10%,$ P<0.01). Similarly, significant differences were observed in chemotactic activity after $HgCl_2$treatment compared with control (control: $0.95{\pm}0.14mm; 0.5 {\mu}M: 0.91{\pm}0.11 mm; 1 {\mu}M: 0.77{\pm}0.16mm, P<0.05; 3{\mu}M: 0.61{\pm} 0.06mm, P<0.01; 5{\mu}M: 0.15{\pm}0.03 mm, P<0.01).$ Also, 4HgCl_2$decreased the percentage of rosette-forming PMNs, indicating diminished phagocytic activity of PMNs upon $HgCl_2$ exposure compared with control (control: $58{\pm}4%; 1{\mu}M: 53{\pm}4%, p<0.05; 3{\mu}M: 49{\pm}3%, P<0.01; 5{\mu}M: 46{\pm}3%, P<0.01).$ Cell viability was not antered after $HgCl_2$treatment (483{\pm}5%$ viability in control PMNs versus $81{\pm}8%$ viability in $5{\mu}M$ Hg-treated PMNs), suggesting that the impaired PMN function after $HgCl_2$treatment was not due to nonspecific cytotoxicity induced by $HgCl_2$. $HgCl_2$-induced decrease in the function of PMNs may have some implications in depressed host susceptibilityupon bacterial challenge after mercury exposure.

• The Korean Journal of Physiology and Pharmacology
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• v.8 no.6
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• pp.319-327
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• 2004

This study was aimed at evaluating the effect of defibrotide on the development of the surgically induced reflux esophagitis, on gastric secretion, lipid peroxidation, polymorphonuclear leukocytes (PMNs) accumulation, polymorphonuclear leukocytes adherence, superoxide anion and hydrogen peroxide production in PMNs, scavenge of hydroxyl radical and hydrogen peroxide, cytokine (interleukin-1 ${\beta}$, tumor necrosis $factor-{\alpha}$) production in blood, and intracelluar calcium mobilization in PMNs. Defibrotide did not inhibit the gastric secretion and not change the gastric pH. Treatment of esophagitis rats with defibrotide inhibited lipid peroxidation, and myeloperoxidase (MPO) in the esophagus in comparison with untreated rats. Defibrotide significantly decreased the PMN adherence to superior mesenteric artery endothelium in a dose-dependent manner, Superoxide anion and hydrogen peroxide production in $1{\mu}M$ formylmethionylleucylphenylalanine (fMLP)- or $0.1{\mu}g/ml$ N-phorbol 12-myristate 13-acetate (PMA)-activated PMNs was inhibited by defibrotide in a dose-dependent fashion. Defibrotide effectively scavenged the hydrogen peroxide but did not scavenge the hydroxyl radical. Treatment of esophagitis rats with defibrotide inhibited interleukin-1 ${\beta}$ production in the blood in comparison with untreated rats, but tumor necrosis $factor-{\alpha}$ production was not affected by defibrotide. The fMLP-induced elevation of intracellular calcium in PMNs was inhibited by defibrotide. The results of this study suggest that defibrotide may have partly beneficial protective effects against reflux esophagitis by the inhibition lipid peroxidation, PMNs accumulation, PMNs adherence to endothelium, reactive oxygen species production in PMNs, inflammatory cytokine production(i.e. interleukin-1 ${\beta}$), and intracellular calcium mobilization in PMNs in rats.

• Journal of Veterinary Clinics
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• v.37 no.2
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• pp.61-66
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• 2020

Nickel is a nutritionally essential trace element that plays an important role in the immune system of several animal species. The aim of this study was to examine the effect of nickel chloride on chemotactic activity of peripheral blood polymorphonuclear cells (PMNs) and whether this effect is associated with interleukin (IL)-8 and a nuclear factor-kappa B (NF-κB)-dependent pathway. Peripheral blood mononuclear cells (PBMCs) and PMNs were isolated by Percoll solution (Specific gravity; 1.080) and 1.5% dextran treatment, respectively. A modified Boyden chamber assay was used to measure the chemotactic activity of PMNs. The level of IL-8 in culture supernatant from PBMCs was measured by enzyme-linked immunosorbent assay (ELISA). Both of PBMCs and PMNs exhibited a low viability when cultured with concentration of greater than 1,000 μM of nickel chloride for 24 h. Thus, nickel chloride was used at concentration of 500 μM, which preserved cell viability. Treatment with nickel did not directly affect the chemotactic activity of PMNs. However, the chemotactic activity of PMNs was remarkably increased by culture supernatant from PBMCs treated with nickel chloride (500 μM) for 24 h. Recombinant porcine IL-8 polyclonal antibody (pAb) neutralized the enhancing effect on the chemotactic activity of PMNs by culture supernatant from PBMCs treated with nickel and this culture supernatant had higher IL-8 levels than the culture supernatant from untreated PBMCs. In addition, n-tosyll-phenylalanine chloromethyl ketone (TPCK), a NF-κB inhibitor, antagonized the enhancing effect on the chemotactic activity of PMNs by the culture supernatant from PBMCs treated with nickel. These results suggested that nickel stimulates porcine PBMCs to produce IL-8, which increases the chemotaxis of PMNs via NF-κB-dependent pathway.

• Journal of Veterinary Clinics
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• v.35 no.5
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• pp.195-199
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• 2018

Zinc is necessary for normal functions in the immune system. The objective of the study is to examine the effect of zinc on the chemotactic activity of porcine peripheral blood polymorphonuclear cells (PMNs). A modified Boyden chamber was used to determine the directional migration distance of PMNs. Various concentrations of zinc showed no chemotactic activity to PMNs. However, culture supernatant from peripheral blood mononuclear cells (PBMCs) treated with zinc remarkably increased the chemotactic activity of PMNs when compared with culture supernatant from PBMCs treated without zinc. Culture supernatant from PBMCs treated without zinc also increased the migration distance of PMNs relative to vehicle control (medium alone). Increasing effect in chemotactic activity of PMNs by culture supernatant from PBMCs treated with zinc was inhibited by treatment of porcine anti-interleukin (IL)-8 polyclonal antibody (pAb). This effect was not affected by heat treatment ($4-85^{\circ}C$). This corresponded with heat stable physical characteristics of IL-8. These results suggest that zinc can upregulate the chemotaxis of PMNs, which is primary mediated by IL-8 chemotactic factor released from PBMCs treated with zinc.

• Journal of Veterinary Clinics
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• v.37 no.5
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• pp.249-254
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• 2020

Neutrophil extracellular trap (NET) formation is an immune response for the invasion of microbes. The purpose of this study is to examine the effect of zinc on NET formation of porcine peripheral blood polymorphonuclear cells (PMNs). The NET formation of PMNs was measured by fluorescence microplate reader. The production of tumor necrosis factor (TNF)-α in the culture supernatants from zinc-treated peripheral blood mononuclear cells (PBMCs) was measured by enzyme-linked immunosorbent assay (ELISA). Zinc itself did not have no effect on NET formation. However, the NET formation of PMNs was increased by culture supernatants from PBMCs treated with zinc. Also, the NET formation of PMNs was increased by recombinant porcine (rp) TNF-α. The production of TNF-α in PBMCs culture supernatants was shown to increase upon zinc treatments. These NET formations of PMNs increased by either culture supernatant from PBMCs treated with zinc or rpTNF-α were inhibited by treatment of anti-rpTNF-α polyclonal antibody (pAb). These results suggested that zinc has an immunostimulating effect on the NET formation of PMNs, which is mediated by TNF-α released from zinc-treated PBMCs. Therefore, zinc may play an important role for NET formation in the defense of porcine inflammatory diseases.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.6
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• pp.515-523
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• 2000

Polymorphonuclear leukocytes (PMNs) play an important role in myocardial ischemia/reperfusion (MI/R) injury. Moreover, platelets are also important blood cells that can aggravate myocardial ischemic injury. This study was designed to test the effects of PMNs and platelets separately and together in provoking cardiac dysfunction in isolated perfused rat hearts following ischemia and reperfusion. Additional control rat hearts were perfused with $75{\times}10^6$ PMNs, with $75{\times}10^6$ platelets, or with $75{\times}10^6\;PMNs+75{\times}10^6$ platelets over a five minute perfusion followed by a 75 min observation period. No significant reduction in coronary flow (CF), left ventricular developed pressure (LVDP), or the first derivative of LVDP (dP/dt max) was observed at the end of the observation period in any non-ischemic group. Similarly, global ischemia (I) for 20 min followed by 45 minutes of reperfusion (R) produced no sustained effects on the final recovery of any of these parameters in any group of hearts perfused in the absence of blood cells. However, I/R hearts perfused with either PMNs or platelets alone exhibited decreases in these variables of $5{\sim}10%$ (p＜0.05 from control). Furthermore, I/R hearts perfused with both PMNs and platelets exhibited decreases of 50 to 60% in all measurements of cardiac function (p＜0.01). These dual cell perfused I/R hearts also exhibited marked increases in cardiac myeloperoxidase (MPO) activity indicating a significant PMN infiltration, and enhanced P-selectin expression on the coronary microvascular endothelium. All cardiaodynamic effects as well as PMN accumulation and P-selectin expression were markedly attenuated by a recombinant soluble PSGL-1 which inhibits selectin mediated cell adhesion. These results provide evidence that platelets and PMNs act synergistically in provoking post-reperfusion cardiac dysfunction, and that this may be largely due to cell to cell interactions mediated by P-selectin. These results also demonstrate that a recombinant soluble PSGL-1 reduces myocardial reperfusion injury by platelet and PMNs interaction.

• Journal of Veterinary Clinics
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• v.28 no.2
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• pp.183-189
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• 2011

The objective of this study was to examine the effect of fucoidan on the phagocytic capapcity and oxidative burst activity (OBA) of canine peripheral blood polymorphonuclear cells (PMNs). The phagocytic capacity and OBA of PMNs were evaluated simultaneously by using a flow cytometer. Fucoidan itself did not cause any direct effect on the phagocytic capacity and OBA of PMNs. However, the phagocytic capacity and OBA of PMNs were enhanced by the culture supernatant from PBMCs treated with fucoidan. The phagocytic capacity and OBA of PMNs were also increased by treatment with recombinant canine (rc) tumor necrosis factor (TNF)-${\alpha}$. The ability of the culture supernatant from fucoidan-treated PBMCs to stimulate the phagocytic capacity and OBA of PMNs was inhibited by addition of anti-rc TNF-${\alpha}$ polyclonal antibody (PAb) prior to the culture. The amount of TNF-${\alpha}$ in the culture supematant from PBMCs was shown to increase upon treatment of fucoidan as compared with that of vehicle-treated PBMCs culture supematant. The level of TNF-${\alpha}$ mRNA expression in PBMCs was also up-regulated by the fucoidan treatment. These results suggest that fucoidan has an immunoenhancing effect on the phagocytic capacity and OBA of canine PMNs, which is mainly mediated by TNF-${\alpha}$ released from fucoidan-stimulated PBMCs.