Effects of Plasmin on Rabbit Platelets

1985 ◽  
Vol 53 (01) ◽  
pp. 008-014 ◽  
Author(s):  
M A Guccione ◽  
R L Kinlough-Rathbone ◽  
M A Packham ◽  
E J Harfenist ◽  
M L Rand ◽  
...  
Keyword(s):  
Prostaglandin E1 ◽  
Periodic Acid ◽  
Buoyant Density ◽  
Creatine Phosphate ◽  
Membrane Glycoproteins ◽  
High Concentration ◽  
Periodic Acid Schiff ◽  
Amine Storage ◽  
Rabbit Platelets

SummaryThe effects of plasmin have been examined because platelets may be exposed to plasmin in vivo and treatment of platelets with plasmin shortens platelet survival. Rabbit plasmin was prepared by urokinase activation of plasminogen immobilized on lysine- Sepharose. Plasmin caused rabbit platelets to aggregate and release the contents of their amine storage granules, but aggregation was slower than in response to ADP or thrombin. EDTA, prostaglandin E1, or creatine phosphate/creatine phosphokinase were inhibitory, but indomethacin was not. Deaggregation did not occur when platelets had been aggregated by a concentration of plasmin that caused extensive release of granule contents. EDTA or prostaglandin E1 caused deaggregation. Low concentrations of ADP and plasmin acted synergistically in causing platelet aggregation. Plasmin decreased the amounts of platelet membrane glycoproteins that stained with periodic acid-Schiff reagent; glycoprotein I was more susceptible than glycoproteins II and III. Concentrations of plasmin that induced the release of amine storage granule contents also released PAS- staining granule gylcoproteins.Platelets incubated with plasmin, washed and resuspended, were not aggregated by ADP, but were aggregated strongly by the combination of fibrinogen and ADP, and bound 125I-fibrinogen to a greater extent than untreated platelets. Platelets preincubated with a high concentration of plasmin were unresponsive to thrombin, but were sometimes aggregated by fibrinogen.Plasmin decreased the buoyant density and increased the median size of platelets. Thus plasmin, as well as ADP and thrombin, may contribute to the density shift observed in platelets from rabbits in which thrombosis and continuous vessel injury have been induced.

scholarly journals Evidence for processing of compact insoluble thyroglobulin globules in relation with follicular cell functional activity in the human and the mouse thyroid

2004 ◽  
pp. 73-80 ◽  
Author(s):  
AC Gerard ◽  
JF Denef ◽  
IM Colin ◽  
MF van den Hove
Keyword(s):  
Lectin Binding ◽  
Periodic Acid ◽  
Soluble Form ◽  
Graves Disease ◽  
High Concentration ◽  
Periodic Acid Schiff ◽  
Thyroid Cells ◽  
Hormone Synthesis ◽  
Normal Human

OBJECTIVE: Thyroglobulin (Tg) is stored within the follicular lumen mainly in a soluble form, but globules made of insoluble multimers are also present and considered to be a mechanism to store prohormone at high concentration. We investigated the immunohistochemical properties of these intrafollicular globules and their possible processing by thyroid cells upon stimulation in the human and in the mouse. DESIGN: Human thyroids (normal, Graves' disease and hot adenomas) and thyroids from old ICR mice without or with goitrogenic treatment were processed for light microscopy. METHODS: Immunohistochemistry for Tg with a polyclonal antibody and two monoclonal antibodies, one specific for thyroxine-rich-iodinated Tg and the other recognizing Tg independently of its iodine level, staining with periodic-acid-schiff, and binding of lectins specific for mannose and sialic acid were performed on all tIssue sections. Intrafollicular globules were quantified, with distinction between 'active' or 'hot' and 'hypofunctioning' or 'cold' follicles. RESULTS: In normal human and old mouse thyroids, the intrafollicular globules were strongly stained with PAS, but negative for the three anti-Tg antibodies and the two lectin-binding assays, while the surrounding soluble Tg was positive. In normal human tIssue, globules were more frequent in 'hypofunctioning' than in 'active' follicles. They were exceptional in Graves' disease and hot adenomas. In old mice, Tg globules were more frequent in 'cold' than in 'hot' follicles. Along with the goitrogen treatment, they became fewer, fragmented and more often present in follicles with a 'hot' aspect. CONCLUSIONS: Upon TSH stimulation, thyrocytes become able to process colloid globules suggesting that this stock of Tg can be used in vivo for thyroid hormone synthesis.

scholarly journals Ectopic atrial arrhythmias arising from canine thoracic veins during in vivo stellate ganglia stimulation

2008 ◽  
Vol 295 (2) ◽  
pp. H691-H698 ◽  
Author(s):  
Alex Y. Tan ◽  
Shengmei Zhou ◽  
Byung Chun Jung ◽  
Masahiro Ogawa ◽  
Lan S. Chen ◽  
...  
Keyword(s):  
Pulmonary Vein ◽  
Sinus Node ◽  
Periodic Acid ◽  
Sympathetic Innervation ◽  
Sympathetic Nerves ◽  
Atrial Arrhythmias ◽  
Periodic Acid Schiff ◽  
Stellate Ganglia ◽  
Ectopic Beats

The purpose of the present study was to determine whether thoracic veins may act as ectopic pacemakers and whether nodelike cells and rich sympathetic innervation are present at the ectopic sites. We used a 1,792-electrode mapping system with 1-mm resolution to map ectopic atrial arrhythmias in eight normal dogs during in vivo right and left stellate ganglia (SG) stimulation before and after sinus node crushing. SG stimulation triggered significant elevations of transcardiac norepinephrine levels, sinus tachycardia in all dogs, and atrial tachycardia in two of eight dogs. Sinus node crushing resulted in a slow junctional rhythm (51 ± 6 beats/min). Subsequent SG stimulation induced 20 episodes of ectopic beats in seven dogs and seven episodes of pulmonary vein tachycardia in three dogs (cycle length 273 ± 35 ms, duration 16 ± 4 s). The ectopic beats arose from the pulmonary vein ( n = 11), right atrium ( n = 5), left atrium ( n = 2), and the vein of Marshall ( n = 2). There was no difference in arrhythmogenic effects of left vs. right SG stimulation (13/29 vs. 16/29 episodes, P = nonsignificant). There was a greater density of periodic acid Schiff-positive cells ( P < 0.05) and sympathetic nerves ( P < 0.05) at the ectopic sites compared with other nonectopic atrial sites. We conclude that, in the absence of a sinus node, thoracic veins may function as subsidiary pacemakers under heightened sympathetic tone, becoming the dominant sites of initiation of focal atrial arrhythmias that arise from sites with abundant sympathetic nerves and periodic acid Schiff-positive cells.

scholarly journals The potential renal toxicity of silver nanoparticles after repeated oral exposure and its underlying mechanisms

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Hamed Nosrati ◽  
Manijeh Hamzepoor ◽  
Maryam Sohrabi ◽  
Massoud Saidijam ◽  
Mohammad Javad Assari ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Molecular Mechanisms ◽  
Renal Toxicity ◽  
Periodic Acid ◽  
Critical Role ◽  
Oral Exposure ◽  
Control Group ◽  
Rt Pcr ◽  
Periodic Acid Schiff

Abstract Background Silver nanoparticles (AgNPs) can accumulate in various organs after oral exposure. The main objective of the current study is to evaluate the renal toxicity induced by AgNPs after repeated oral exposure and to determine the relevant molecular mechanisms. Methods In this study, 40 male Wistar rats were treated with solutions containing 30, 125, 300, and 700 mg/kg of AgNPs. After 28 days of exposure, histopathological changes were assessed using hematoxylin-eosin (H&E), Masson’s trichrome, and periodic acid-Schiff (PAS) staining. Apoptosis was quantified by TUNEL and immunohistochemistry of caspase-3, and the level of expression of the mRNAs of growth factors was determined using RT-PCR. Results Histopathologic examination revealed degenerative changes in the glomeruli, loss of tubular architecture, loss of brush border, and interrupted tubular basal laminae. These changes were more noticeable in groups treated with 30 and 125 mg/kg. The collagen intensity increased in the group treated with 30 mg/kg in both the cortex and the medulla. Apoptosis was much more evident in middle-dose groups (i.e., 125 and 300 mg/kg). The results of RT-PCR indicated that Bcl-2 and Bax mRNAs upregulated in the treated groups (p < 0.05). Moreover, the data related to EGF, TNF-α, and TGF-β1 revealed that AgNPs induced significant changes in gene expression in the groups treated with 30 and 700 mg/kg compared to the control group. Conclusion Our observations showed that AgNPs played a critical role in in vivo renal toxicity.

THE EFFECT OF ENZYMATIC TREATMENT OF RABBIT PLATELETS ON THE BINDING OF IMMUNOGLOBULIN G TO THE SURFACE

1987 ◽  
Author(s):  
R M Evans ◽  
M A Packham
Keyword(s):  
Immunoglobulin G ◽  
Rabbit Serum ◽  
Membrane Glycoproteins ◽  
Plasmin Activity ◽  
Platelet Suspension ◽  
Platelet Survival ◽  
Igg Binding ◽  
Rabbit Igg ◽  
Rabbit Platelets

As shown previously, cleavage of surface glycoproteins on the platelet membrane shortens platelet survival. Since immunoglobulins have been implicated in the condition of idiopathic thrombocytopenic purpura in which patients have an elevated level of platelet-associated immunoglobulin G (I g G) and shortened platelet survival, we measured the binding of IgG to rabbit platelets that had been treated with neuraminidase or plasmin, to determine whether increased IgG binding to these platelets might be the signal that leads to their accelerated clearance. Rabbit platelets were washed and resuspended in calcium-free Tyrode-albumin solution (1.0 x 106/ μL), pH 6.5, and then incubated with either neuraminidase (0.1 U/mL at 22°C) or plasmin (activity: 1.6 ymol BAEE/min/mL platelet suspension at 37¼C) for 30 min. The platelets were then centrifuged and resuspended in heat-inactivated rabbit serum for 30 min to allow IgG to bind to the surface. After additional washing steps to remove non-specifically bound IgG, the amount Qt IgG that associated with the platelets was measured using an 125I-labeled goat anti-rabbit IgG binding assay. Binding data were analyzed by plotting the calculated fg of IgG bound/platelet versus the amount of 125-1 goat anti-rabbit IgG added. The binding curves for both neuraminidase and plasmin indicated significantly increased amounts of specifically bound IgG compared to control platelets. At a concentration of 30 μg/mL of labeled goat antirabbit IgG added, the amount of label associated with untreated rabbit platelets had plateaued at 0.36± 0.11 fg/platelet whereas the amount associated with neuraminidase-treated platelets was 5 times greater and with pi asmin-treated platelets, 1.5 times greater, and neither had plateaued. The results obtained with plasmin are of particular interest since this proteolytic enzyme shortens platelet survival and is present in vivo during thromboembolic events. These results indicate that IgG binds specifically to the surface of rabbit platelets following enzymatic alterations of the membrane glycoproteins and therefore may represent at least one mechanism that determines which platelets are cleared by the reticuloendothelial system.

scholarly journals Necroptosis Contributes to Urban Particulate Matter-Induced Airway Epithelial Injury

2018 ◽  
Vol 46 (2) ◽  
pp. 699-712 ◽  
Author(s):  
Feng Xu ◽  
Man Luo ◽  
Lulu He ◽  
Yuan Cao ◽  
Wen Li ◽  
...  
Keyword(s):  
Particulate Matter ◽  
Pulmonary Inflammation ◽  
Periodic Acid ◽  
Enzyme Linked Immunosorbent Assay ◽  
Periodic Acid Schiff ◽  
Mucus Production ◽  
Gm Csf ◽  
Mucin Expression

Background/Aims: Necroptosis, a form of programmed necrosis, is involved in the pathologic process of several kinds of pulmonary diseases. However, the role of necroptosis in particulate matter (PM)–induced pulmonary injury remains unclear. The objective of this study is to investigate the involvement of necroptosis in the pathogenesis of PM-induced toxic effects in pulmonary inflammation and mucus hyperproduction, both in vitro and in vivo. Methods: PM was administered into human bronchial epithelial (HBE) cells or mouse airways, and the inflammatory response and mucus production were assessed. The mRNA expressions of IL6, IL8 and MUC5AC in HBE cells and Cxcl1, Cxcl2, and Gm-csf in the lung tissues were detected by quantitative real-time RT-PCR. The secreted protein levels of IL6 and IL8 in culture supernatants and Cxcl1, Cxcl2, and Gm-csf in bronchoalveolar lavage fluid (BALF) were detected by enzyme-linked immunosorbent assay (ELISA). We used Western blot to measure the protein expressions of necroptosis-related proteins (RIPK1, RIPK3, and Phospho-MLKL), NF-κB (P65 and PP65), AP-1 (P-c-Jun and P-c-Fos) and MUC5AC. Cell necrosis and mitochondrial ROS were detected using flow cytometry. In addition, pathological changes and scoring of lung tissue samples were monitored using hemoxylin and eosin (H&E), periodic acid-schiff (PAS) and immunohistochemistry staining. Results: Our study showed that PM exposure induced RIP and MLKL-dependent necroptosis in HBE cells and in mouse lungs. Managing the necroptosis inhibitor Necrostatin-1 (Nec-1) and GSK’872, specific molecule inhibitors of necroptosis, markedly reduced PM-induced inflammatory cytokines, e.g., IL6 and IL8, and MUC5AC in HBE cells. Similarly, administering Nec-1 significantly reduced airway inflammation and mucus hyperproduction in PM-exposed mice. Mechanistically, we found PM–induced necroptosis was mediated by mitochondrial reactive oxygen species-dependent early growth response gene 1, which ultimately promoted inflammation and mucin expression through nuclear factor κB and activator protein-1 pathways, respectively. Conclusions: Our results demonstrate that necroptosis is involved in the pathogenesis of PM–induced pulmonary inflammation and mucus hyperproduction, and suggests that it may be a novel target for treatment of airway disorders or disease exacerbations with airborne particulate pollution.

Membrane Glycoproteins of Human and Rabbit Platelets: Studies In Vitro and after Circulation in Vivo

1975 ◽  
Author(s):  
J. N. George ◽  
P. C. Lewis ◽  
D. A. Sears
Keyword(s):  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Age Distribution ◽  
Human Platelets ◽  
Membrane Glycoproteins ◽  
Rabbit Platelets ◽  
Trypsin Hydrolysis ◽  
Initial Recovery

The initial events of hemostasis and thrombosis involve platelet contact interactions and may be mediated by surface glycoproteins. Human and rabbit platelets were labeled with 125I-diazotized diiodosulfanilic acid (I), which reacts covalently with proteins, and proteins were separated by SDS-polyacrylamide gel electrophoresis. Only exposed membrane proteins were labeled because: 1) protein specific activity of membranes was 4-7 times that of whole platelets, 2) different proteins were labeled when I was reacted with isolated membranes, and 3) trypsin-hydrolysis of labeled intact platelets altered the radioactive peaks. Like Phillips (Biochem. 11, 4582, 72) and Nachman et al. (JBC 248, 2928, 73) we found that lactoperoxidase iodinated the 93,000 dalton glycoprotein (GP) of human platelets. In contrast, I labeled both the 93,000 and 118,000 dalton membrane GP of human platelets, and all 3 membrane GP of rabbit platelets.Rabbit platelets labeled simultaneously with I and 51Cr had identical density and therefore age distribution of the 2 labels. After infusion into rabbits, initial recovery of I was 23% of the Cr recovery. After 3 hrs, I disappearance was exponential and more rapid (T/2 = 17 hrs) than the linear Cr disappearance (T/2 = 30 hrs, p < .01). This was due to in vivo removal of I from circulating platelets since 1 did not elute more rapidly from platelets harvested after 3 hrs circulation and incubated in plasma at 37° (T/2 of I elution = 43 hrs, Cr = 33 hrs). Platelets harvested after 14-20 hrs circulation had the same distribution of I on the membrane GP as before circulation. We postulate that this symmetrical label loss indicates uniform loss of membrane GP, suggesting that platelets lose pieces of their plasma membrane during circulation. This could occur during contact interaction in the process of hemostasis.

scholarly journals Mitochondria-Targeted Antioxidant Mito-Tempo Protects Against Aldosterone-Induced Renal Injury In Vivo

2017 ◽  
Vol 44 (2) ◽  
pp. 741-750 ◽  
Author(s):  
Wei Ding ◽  
Tingyan Liu ◽  
Xiao Bi ◽  
Zhiling Zhang
Keyword(s):  
Electron Microscopy ◽  
Renal Function ◽  
Nlrp3 Inflammasome ◽  
Renal Injury ◽  
Periodic Acid ◽  
Inflammasome Activation ◽  
Periodic Acid Schiff ◽  
Nlrp3 Inflammasome Activation

Background/Aims: Growing evidence suggests mitochondrial dysfunction (MtD) and the Nlrp3 inflammasome play critical roles in chronic kidney disease (CKD) progression. We previously reported that Aldosterone (Aldo)-induced renal injury in vitro is directly caused by mitochondrial reactive oxygen species (mtROS)-mediated activation of the Nlrp3 inflammasome. Here we aimed to determine whether a mitochondria-targeted antioxidant (Mito-Tempo) could prevent Aldo-induced kidney damage in vivo. Methods: C57BL/6J mice were treated with Aldo and/or Mito-Tempo (or ethanol as a control) for 4 weeks. Renal injury was evaluated by Periodic Acid-Schiff reagent or Masson’s trichrome staining and electron microscopy. ROS were measured by DCFDA fluorescence and ELISA. MtD was determined by real-time PCR and electron microscopy. Activation of the Nlrp3 inflammasome and endoplasmic reticulum stress (ERS) was detected via western blot. Results: Compared with control mice, Aldo-infused mice showed impaired renal function, increased mtROS production and MtD, Nlrp3 inflammasome activation, and elevated ERS. We showed administration of Mito-Tempo significantly improved renal function and MtD, and reduced Nlrp3 inflammasome activation and ERS in vivo. Conclusion: Mitochondria-targeted antioxidants may attenuate Aldo-infused renal injury by inhibiting MtD, the Nlrp3 inflammasome, and ERS in vivo. Therefore, targeting mtROS might be an effective strategy for preventing CKD.

Changes in surface morphology of pigmented retinal cells during differentiation in clonal culture

1981 ◽  
Vol 59 (1) ◽  
pp. 61-69 ◽  
Author(s):  
B. J. Crawford ◽  
Alex Yan ◽  
M. MacDonald
Keyword(s):  
Electron Microscope ◽  
Surface Morphology ◽  
Periodic Acid ◽  
Outer Edge ◽  
Gradual Change ◽  
Apical Surface ◽  
Periodic Acid Schiff ◽  
Clonal Culture ◽  
Cellular Attachment

The changes in surface morphology during the reexpression of differentiation of chick retinal pigmented epithelial cells (RPE) in clonal culture have been studied using the scanning electron microscope (SEM) and transmission electron microscope (TEM) and compared with those described in vivo. Three-week-old colonies demonstrated a gradual change in apical surface morphology along any colony radius. At the outer edge, the cell surfaces were either smooth with a few small filamentous protrusions or showed a varying number of large blebs. Toward the centre of the colony the surfaces demonstrated a gradual increase in filamentous protrusions. The apical surfaces of the most densely pigmented cells at the centre of the colony consisted mainly of small rounded protrusions. The changes in surface morphology of cells in the centre of younger colonies during redifferentiation were similar to those found along the radius of a 3-week-old colony. The results show that older colonies have all of the morphological stages of the redifferentiation process (and possibly the biochemical ones as well) arranged along any radius.The basal surfaces of all the colonies were covered by a thin acellular membrane that stained positively with periodic acid–Schiff (PAS) and which may contain fibronectins and appears to be involved in cellular attachment.

scholarly journals Gastrin Attenuates Renal Ischemia/Reperfusion Injury by a PI3K/Akt/Bad-Mediated Anti-apoptosis Signaling

2020 ◽  
Vol 11 ◽  
Author(s):  
Chao Liu ◽  
Ken Chen ◽  
Huaixiang Wang ◽  
Ye Zhang ◽  
Xudong Duan ◽  
...  
Keyword(s):  
Protective Effect ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Periodic Acid ◽  
Drug Candidate ◽  
Gastrointestinal Hormone ◽  
Akt Inhibitor ◽  
Periodic Acid Schiff

Ischemic/reperfusion (I/R) injury is the primary cause of acute kidney injury (AKI). Gastrin, a gastrointestinal hormone, is involved in the regulation of kidney function of sodium excretion. However, whether gastrin has an effect on kidney I/R injury is unknown. Here we show that cholecystokinin B receptor (CCKBR), the gastrin receptor, was significantly up-regulated in I/R-injured mouse kidneys. While pre-administration of gastrin ameliorated I/R-induced renal pathological damage, as reflected by the levels of serum creatinine and blood urea nitrogen, hematoxylin and eosin staining and periodic acid-Schiff staining. The protective effect could be ascribed to the reduced apoptosis for gastrin reduced tubular cell apoptosis both in vivo and in vitro. In vitro studies also showed gastrin preserved the viability of hypoxia/reoxygenation (H/R)-treated human kidney 2 (HK-2) cells and reduced the lactate dehydrogenase release, which were blocked by CI-988, a specific CCKBR antagonist. Mechanistically, the PI3K/Akt/Bad pathway participates in the pathological process, because gastrin treatment increased phosphorylation of PI3K, Akt and Bad. While in the presence of wortmannin (1 μM), a PI3K inhibitor, the gastrin-induced phosphorylation of Akt after H/R treatment was blocked. Additionally, wortmannin and Akt inhibitor VIII blocked the protective effect of gastrin on viability of HK-2 cells subjected to H/R treatment. These studies reveals that gastrin attenuates kidney I/R injury via a PI3K/Akt/Bad-mediated anti-apoptosis signaling. Thus, gastrin can be considered as a promising drug candidate to prevent AKI.

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