PATHOGENESIS OF INFLAMMATION

Material obtained from the in vitro incubation of granulocytes from saline-induced peritoneal exudates of rabbits has been shown to produce inflammation and fever in rabbits. The supernatant material from cells incubated in saline has been termed granulocytic substance (GS) and is heat-labile. Its production is temperature dependent, occurring at 37°C but not at 4°C, requires viable cells, and is inhibited by potassium ions. A similar material is liberated when cells are incubated in a more physiologic medium. Freezing and thawing of granulocytes does not release GS and the active principle cannot be obtained from the incubation of lymphocytes. GS produces a delayed inflammatory response as measured by leucocyte sticking and emigration in the rabbit ear chamber and the leakage of protein-conjugated dye at the site of intradermal injection. The former response can be accurately quantitated by calculation of the inflammatory index from reactions observed in the ear chamber. The inflammatory reaction and the properties of GS distinguish it from a variety of previously described mediators of inflammation, but GS appears to be identical with leucocytic pyrogen. The possible role of GS in delayed and protracted inflammation and its relationship to the pathogenesis of fever are discussed.

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Mutations affecting the tRNA-splicing endonuclease activity of Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/118.4.609 ◽

1988 ◽

Vol 118 (4) ◽

pp. 609-617

Author(s):

M Winey ◽

M R Culbertson

Keyword(s):

Growth Defect ◽

Temperature Sensitive ◽

Gene Products ◽

Endonuclease Activity ◽

Temperature Dependent ◽

Direct Role ◽

Trna Splicing

Abstract Two unlinked mutations that alter the enzyme activity of tRNA-splicing endonuclease have been identified in yeast. The sen1-1 mutation, which maps on chromosome 12, causes temperature-sensitive growth, reduced in vitro endonuclease activity, and in vivo accumulation of unspliced pre-tRNAs. The sen2-1 mutation does not confer a detectable growth defect, but causes a temperature-dependent reduction of in vitro endonuclease activity. Pre-tRNAs do not accumulate in sen2-1 strains. The in vitro enzyme activities of sen1-1 and sen2-1 complement in extracts from a heterozygous diploid, but fail to complement in mixed extracts from separate sen1-1 and sen2-1 haploid strains. These results suggest a direct role for SEN gene products in the enzymatic removal of introns from tRNA that is distinct from the role of other products known to affect tRNA splicing.

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Contributions of arachidonic acid derivatives and substance P to the sensitization of cutaneous nociceptors

Journal of Neurophysiology ◽

10.1152/jn.1990.64.2.457 ◽

1990 ◽

Vol 64 (2) ◽

pp. 457-464 ◽

Cited By ~ 61

Author(s):

R. H. Cohen ◽

E. R. Perl

Keyword(s):

Arachidonic Acid ◽

Substance P ◽

Receptive Fields ◽

Lipoxygenase Inhibitor ◽

Arterial Perfusion ◽

Mediators Of Inflammation ◽

Rabbit Ear ◽

C Fiber

1. The role of presumed chemical mediators of inflammation in the heat-induced sensitization of cutaneous C-polymodal nociceptors (CPNs) was examined in a rabbit ear preparation maintained in vitro by intra-arterial perfusion with a solution free of protein and cellular elements. 2. In this preparation, CPNs consistently showed enhanced responsiveness after repeated exposure of their receptive fields to noxious levels of heat. The average magnitude of sensitization was quantitatively similar to that observed in vivo, suggesting that blood-born factors are not essential for development of sensitization. 3. Sensitization in one-half of randomly selected CPNs was blocked or reduced when the perfusate contained a cyclooxygenase inhibitor, indomethacin or dipyrone, or the dual cyclooxygenase/lipoxygenase inhibitor, BW755C, even though initial responsiveness to heat and pressure was unaltered. These observations suggest that arachidonic acid breakdown products, possibly prostaglandins, are intermediaries in the sensitization of some, but not all, C-fiber nociceptors of the skin. In addition, heat-induced sensitization for some C-fiber cutaneous nociceptors is the result of processes that are at least partially independent of those involved in excitation. 4. Substance P (SP) or the putative SP antagonists, [D-Pro2, D-Trp7.9]-SP or [D-Pro2, D-Phe7, D-Trip9]-SP, produced no significant effect on heat-responsiveness or sensitization, although ongoing activity may have marginally increased over control levels after repeated heat stimulations. We conclude that SP in an in vitro preparation is not involved in the enhancement of cutaneous C-fiber nociceptor responsiveness after repeated thermal insults.

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THE ACUTE INFLAMMATORY REACTION IN THE RABBIT EAR CHAMBER WITH PARTICULAR REFERENCE TO THE PHENOMENON OF LEUKOCYTIC MIGRATION

Journal of Experimental Medicine ◽

10.1084/jem.124.4.543 ◽

1966 ◽

Vol 124 (4) ◽

pp. 543-556 ◽

Cited By ~ 20

Author(s):

W. J. Cliff

Keyword(s):

Inflammatory Responses ◽

Specific Activity ◽

Time Lapse ◽

Amoeboid Movement ◽

Inflammatory Reactions ◽

Rabbit Ear ◽

Rabbit Ear Chamber ◽

Venular Endothelium

Responses to injections of various materials into rabbit ear chambers were studied by in vivo microscopy. The acute inflammatory responses provoked by injections of antibody-antigen complexes were both quantitatively and qualitatively different from the responses obtained after injections of either homologous sera or the antigens alone. The sticking of leukocytes to endothelium during these responses occurred only in the venules draining the injection sites and was frequently present only on the sides of the venules towards the injection sites. An explanation of this finding was proposed in terms of absorption by the minute vessels related to the injection sites of postulated mediator(s) with specific activity on venular endothelium. Analysis of the rates and direction of movement of leukocytes during the reactions produced by the antibody-antigen complexes was performed with the aid of time-lapse cinemicroscopy. The leukocytes that were sticking to the venular endothelium frequently exhibited amoeboid locomotion within the vessels. Twice as many of these cells moved against the direction of blood flow as with it. This finding was discussed and an explanation proposed. A method for detecting a drift in the overall population of emigrated leukocytes within the inflamed tissue was described and revealed that four times as many amoeboid cells moved away from the injection sites as towards them. This result was discussed in the light of the in vitro chemotactic properties of antibody-antigen complexes demonstrated for rabbit leukocytes. An alternative explanation was proposed in terms of variation in the population density of these cells and their random movements and collisions. The rates of amoeboid movement of leukocytes during the acute inflammatory reactions produced by antibody-antigen complexes were similar to the rates found during turpentine inflammation and were compared to other published values.

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Fluorescence analysis of the interaction of isometamidium with Trypanosoma congolense

Biochemical Journal ◽

10.1042/bj2920031 ◽

1993 ◽

Vol 292 (1) ◽

pp. 31-35 ◽

Cited By ~ 8

Author(s):

D Zilberstein ◽

J Wilkes ◽

H Hirumi ◽

A S Peregrine

Keyword(s):

Time Course ◽

Fluorescence Analysis ◽

Trypanosoma Congolense ◽

Sub Saharan Africa ◽

Temperature Dependent ◽

Intact Cells ◽

Uptake Process ◽

Sub Saharan

Isometamidium chloride (Samorin) is the only compound recommended for prophylaxis against bovine trypanosomiasis in sub-Saharan Africa. The fluorescence property of this compound was used to investigate the interaction of the molecule with in vitro-derived bloodstream forms of Trypanosoma congolense IL 1180. Incubation of isometamidium with trypanosomes at 37 degrees C for 180 min resulted in a gradual alteration of the lambda max. with time (from 600 to 584 nm) and an increase in the intensity of trypanosome-associated fluorescence of approx. 2-fold. The alteration in fluorescence was temperature-dependent and inhibited by the addition of N-ethylmaleimide. In contrast, with intact cells addition of digitonin caused a rapid increase in fluorescence intensity to approximately four times that observed with intact cells. Uptake of isometamidium was also determined using radiolabelled drug; the results indicated that the time course of the uptake process resembled the fluorescence profile and was temperature-dependent. The results therefore indicate that the alteration of fluorescence is due to interaction of isometamidium with an intracellular component(s) and that isometamidium is transported across the plasma membrane via a protein carrier. The data also indicate that the described fluorescence technique can be used to investigate the role of membrane transport in resistance to isometamidium.

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Signaling through FcγRIIA and the C5a-C5aR pathway mediates platelet hyperactivation in COVID-19

10.1101/2021.05.01.442279 ◽

2021 ◽

Author(s):

Sokratis A. Apostolidis ◽

Amrita Sarkar ◽

Heather M. Giannini ◽

Rishi R. Goel ◽

Divij Mathew ◽

...

Keyword(s):

Cardiovascular Disease ◽

Platelet Activation ◽

Clinical Manifestations ◽

Vascular Complications ◽

Surface Expression ◽

Microfluidic Chamber ◽

Mediators Of Inflammation ◽

Syk Inhibitor

Patients with COVID-19 present with a wide variety of clinical manifestations. Thromboembolic events constitute a significant cause of morbidity and mortality in patients infected with SARS-CoV-2. Severe COVID-19 has been associated with hyperinflammation and pre-existing cardiovascular disease. Platelets are important mediators and sensors of inflammation and are directly affected by cardiovascular stressors. In this report, we found that platelets from severely ill, hospitalized COVID-19 patients exhibit higher basal levels of activation measured by P-selectin surface expression, and have a poor functional reserve upon in vitro stimulation. Correlating clinical features to the ability of plasma from COVID-19 patients to stimulate control platelets identified ferritin as a pivotal clinical marker associated with platelet hyperactivation. The COVID-19 plasma-mediated effect on control platelets was highest for patients that subsequently developed inpatient thrombotic events. Proteomic analysis of plasma from COVID-19 patients identified key mediators of inflammation and cardiovascular disease that positively correlated with in vitro platelet activation. Mechanistically, blocking the signaling of the FcγRIIa-Syk and C5a-C5aR pathways on platelets, using antibody-mediated neutralization, IgG depletion or the Syk inhibitor fostamatinib, reversed this hyperactivity driven by COVID-19 plasma and prevented platelet aggregation in endothelial microfluidic chamber conditions, thus identifying these potentially actionable pathways as central for platelet activation and/or vascular complications in COVID-19 patients. In conclusion, we reveal a key role of platelet-mediated immunothrombosis in COVID-19 and identify distinct, clinically relevant, targetable signaling pathways that mediate this effect. These studies have implications for the role of platelet hyperactivation in complications associated with SARS-CoV-2 infection.

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The Effect of Serotonin Inhibition on Initial Microvessel Hemostasis and Platelet Aggregation In Vivo

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657355 ◽

1983 ◽

Vol 49 (03) ◽

pp. 173-175 ◽

Cited By ~ 6

Author(s):

D Bergqvist ◽

S Arvidsson ◽

C O Esquivel ◽

B Lindblad ◽

U Haglund

Keyword(s):

Adrenergic Receptor ◽

Formation Time ◽

Platelet Activity ◽

Receptor Affinity ◽

Rabbit Ear ◽

Rabbit Ear Chamber ◽

Plug Formation ◽

Mesenteric Microcirculation

SummaryThe role of serotonin (5-HT) in initial microvascular hemostasis is not fully understood. This study was made to evaluate the effect on hemostatic plug formation and laser-induced arteriolar microembolism of different substances which counteract the effect of 5-HT. Hemostatic plug formation time and stability was measured in the rabbit mesenteric microcirculation and laserinduced embolism in the rabbit ear chamber. Ketanserine, a selective 5-HT2-receptor blocker shortened arteriolar hemostatic plug formation time. Dihydroergotamine, an unselective blocker (with 5-HT- and α-adrenergic receptor affinity) increased venular hemostatic plug formation time and also decreased the hemostatic plug stability. Laser-induced platelet embolism was unaltered after both ketanserine and dihydroergotamine administration. The magnitude of these changes seems to exclude an important effect of 5-HT in initial microvessel hemostasis or on platelet activity.

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Human Platelets as a Model for the Binding and Degradation of Thrombopoietin

Blood ◽

10.1182/blood.v89.8.2782 ◽

1997 ◽

Vol 89 (8) ◽

pp. 2782-2788 ◽

Cited By ~ 107

Author(s):

Paul J. Fielder ◽

Philip Hass ◽

Mark Nagel ◽

Eric Stefanich ◽

Ramon Widmer ◽

...

Keyword(s):

Specific Binding ◽

Platelet Rich Plasma ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Human Platelets ◽

Scatchard Analysis ◽

Temperature Dependent ◽

Degradation Pattern

Abstract Recent studies have shown that plasma thrombopoietin (TPO) levels appear to be directly regulated by platelet mass and that removal of plasma TPO by platelets via binding to the c-Mpl receptor is involved in the clearance of TPO in rodents. To help elucidate the role of platelets in the clearance of TPO in humans, we studied the in vitro specific binding of recombinant human TPO (rhTPO) to human platelet-rich plasma (PRP), washed platelets (WP), and cloned c-Mpl. Using a four-parameter fit and/or Scatchard analysis, the approximate affinity of rhTPO for its receptor, which was calculated from multiple experiments using different PRP preparations, was between 128 and 846 pmol/L, with ∼25 to 224 receptors per platelet. WP preparations gave an affinity of 260 to 540 pmol/L, with ∼25 to 35 receptors per platelet, and erythropoietin failed to compete with 125I-rhTPO for binding to WP. Binding and dissociation studies conducted with a BiaCore apparatus yielded an affinity of 350 pmol/L for rhTPO binding to cloned c-Mpl receptors. The ability of PRP to bind and degrade 125I-rhTPO was both time- and temperature-dependent and was blocked by the addition of excess cold rhTPO. Analysis of platelet pellets by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that 125I-rhTPO was degraded into a major fragment of ∼45 to 50 kD. When 125I-rhTPO was incubated with a platelet homogenate at pH = 7.4, a degradation pattern similar to intact platelets was observed. Together, these data show that human platelets specifically bind rhTPO with high affinity, internalize, and then degrade the rhTPO.

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Scopoletin Augments DJ1/Nrf2 Signalling and Prevents Protein Aggregation in Parkinson’s disease

10.1101/260521 ◽

2018 ◽

Author(s):

S Narasimhan Kishore Kumar ◽

Jayakumar Deepthy ◽

Velusamy Prema ◽

Srinivasan Ashokkumar ◽

Mohan Thangarajeswari ◽

...

Keyword(s):

Oxidative Stress ◽

Protein Aggregation ◽

Nuclear Translocation ◽

Morinda Citrifolia ◽

Active Principle ◽

Antioxidant Genes ◽

Induced Apoptosis ◽

Target Side

AbstractGiven the role of oxidative stress in PD pathogenesis and off-target side effects of currently available drugs, several natural phytochemicals seems to be promising in the management of PD. Here, we tested the hypothesis that scopoletin, an active principle obtained from Morinda citrifolia (MC), efficiently quenches oxidative stress through DJ-1/Nrf2 signalling and ameliorates rotenone-induced PD. Despite reducing the oxidative stress, administration of MC extract (MCE) has lessened the protein aggregation as evident from decreased levels of nitrotyrosine and α-Synuclein. In vitro studies revealed that scopoletin lessened rotenone-induced apoptosis in SH-SY5Y cells through preventing oxidative injury. Particularly, scopoletin markedly up-regulated DJ-1, which then promoted the nuclear translocation of Nrf2 and transactivation of antioxidant genes. Furthermore, we found that scopoletin prevent the nuclear exportation of Nrf2 by reducing the levels of Keap1 and thereby enhancing the neuronal defence system. Overall, our findings suggest that scopoletin acts through DJ-1 mediated Nrf2 signalling to protect the brain from rotenone-induced oxidative stress and PD. Thus, we postulate that scopoletin could be a potential drug to treat PD.

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Morinda citrifolia and Its Active Principle Scopoletin Mitigate Protein Aggregation and Neuronal Apoptosis through Augmenting the DJ-1/Nrf2/ARE Signaling Pathway

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/2761041 ◽

2019 ◽

Vol 2019 ◽

pp. 1-13 ◽

Cited By ~ 8

Author(s):

Kishore Kumar S. Narasimhan ◽

Deepthy Jayakumar ◽

Prema Velusamy ◽

Ashokkumar Srinivasan ◽

Thangarajeswari Mohan ◽

...

Keyword(s):

Oxidative Stress ◽

Protein Aggregation ◽

Nuclear Translocation ◽

Morinda Citrifolia ◽

Active Principle ◽

Antioxidant Genes ◽

Induced Apoptosis ◽

Target Side

Given the role of oxidative stress in PD pathogenesis and off-target side effects of currently available drugs, several natural phytochemicals seem to be promising in the management of PD. Here, we tested the hypothesis that scopoletin, an active principle obtained from Morinda citrifolia (MC), efficiently quenches oxidative stress through DJ-1/Nrf2 signaling and ameliorates rotenone-induced PD. Despite reducing oxidative stress, the administration of MC extract (MCE) has lessened protein aggregation as evident from decreased levels of nitrotyrosine and α-synuclein. In vitro studies revealed that scopoletin lessened rotenone-induced apoptosis in SH-SY5Y cells through preventing oxidative injury. Particularly, scopoletin markedly upregulated DJ-1, which then promoted the nuclear translocation of Nrf2 and transactivation of antioxidant genes. Furthermore, we found that scopoletin prevents the nuclear exportation of Nrf2 by reducing the levels of Keap1 and thereby enhancing the neuronal defense system. Overall, our findings suggest that scopoletin acts through DJ-1-mediated Nrf2 signaling to protect the brain from rotenone-induced oxidative stress and PD. Thus, we postulate that scopoletin could be a potential drug to treat PD.

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Effects of Bilirubin on Cerebral Arterial Tone in vitro

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.1989.94 ◽

1989 ◽

Vol 9 (5) ◽

pp. 666-674 ◽

Cited By ~ 29

Author(s):

Frederick Jia-Pei Miao ◽

Tony Jer-Fu Lee

Keyword(s):

Cerebral Vasospasm ◽

Cerebral Arteries ◽

Active Principle ◽

Cerebral Vessel ◽

Vasoactive Substances ◽

High Concentrations ◽

Arterial Tone

Hemoglobin and its metabolite, bilirubin, have been shown to be present in high concentrations in CSF following subarachnoid hemorrhage (SAH). Several reports have indicated that hemoglobin is a potent cerebral vasoconstrictor and therefore is considered to be an active principle in the genesis of cerebral vasospasm. The possible role of bilirubin on the genesis of cerebral vasospasm, however, has not been clarified. The effect of bilirubin on cerebral vessel tone was therefore examined using in vitro tissue bath techniques. Bilirubin (10−5-3 × 10−5 M) induced strong constriction of cerebral arteries from the cat, dog, and pig. The vasoconstriction returned to baseline after bilirubin was washed by prewarmed Krebs solution. Vasomotor responses to various vasoactive substances were then examined after bilirubin was washed away (the bilirubin postwash effect). Norepinephrine (NE)–induced but not serotonin- or acetylcholine (ACh)–induced constrictions were significantly potentiated by bilirubin postwash effect. The potentiated NE-induced constriction was attenuated by yohimbine but not by prazosin. This enhanced vasoconstriction was mimicked by clonidine but not by phenylephrine, suggesting that the potentiation of NE-induced constriction by the bilirubin postwash effect was mediated by the α2-adrenoceptor subtype. The bilirubin postwash effect also resulted in blockade of endothelium-dependent vasodilations induced by A-23147, ACh, and ATP without affecting relaxations induced by direct muscle relaxants such as β-adrenoceptors, papaverine, and sodium nitroprusside. These results indicate that bilirubin induces direct vasoconstriction, potentiates α2-adrenoceptormediated vasoconstriction, and inhibits endothelium-dependent vasodilation. These actions of bilirubin may promote an enhanced overall vasoconstriction in vivo. Bilirubin, therefore, may be involved in the genesis of cerebral vasospasm following SAH.

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