Evidence of the role of hemolysis in experimental cerebral vasospasm

1990 ◽  
Vol 72 (5) ◽  
pp. 775-781 ◽  
Author(s):  
John W. Peterson ◽  
Lawrence Roussos ◽  
Byung-Duk Kwun ◽  
John D. Hackett ◽  
Christopher J. Owen ◽  
...  
Keyword(s):  
Cerebral Vasospasm ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Canine Model ◽  
Initial Reaction ◽  
Basal Cistern ◽  
Content Type ◽  
Vascular Reaction ◽  
Erythrocyte Lysis

✓ The short-term (≤ 72-hour) reaction to subarachnoid injections of various blood components was determined in a canine model of cerebral vasospasm. Platelet-rich plasma (PRP) formed durable clots in the basal cistern surrounding the basilar artery and provoked no vascular reaction in 72 hours or more. Freshly isolated autologous erythrocytes resuspended in PRP likewise provoked no vasoconstriction in 72 hours, although a second injection of fresh erythrocytes in PRP induced significant reaction, as in the conventional “double subarachnoid hemorrhage (SAH)” canine model. Hemolysate of fresh erythrocytes led to a severe immediate vascular reaction after introduction into the basal cistern using PRP as the carrier/clotting medium, as did the injection of intact erythrocytes incubated ex vivo for 72 hours. Resolution of the initial reaction was rapid for hemolysate, but slow and (depending on hematocrit) incomplete for intact “aged” erythrocytes. In vitro measurements of erythrocyte lysis in these media and histological examination indicate that the production of erythrocyte lysate was responsible for the vascular reaction observed, suggesting that the rate of lysis of erythrocytes in the subarachnoid clot is a major factor in the genesis of vasospasm after SAH.

The role of inflammation in experimental cerebral vasospasm

1990 ◽  
Vol 72 (5) ◽  
pp. 767-774 ◽  
Author(s):  
John W. Peterson ◽  
Byung-Duk Kwun ◽  
John D. Hackett ◽  
Nicholas T. Zervas
Keyword(s):  
Cerebral Vasospasm ◽  
Rapid Onset ◽  
Canine Model ◽  
Basal Cistern ◽  
Autologous Plasma ◽  
Content Type ◽  
Vascular Reaction ◽  
Blood Clots ◽  
Subarachnoid Blood

✓ The short-term (≤ 72-hour) cerebral vascular reaction to subarachnoid injectates of various specific blood components was determined by angiography in a canine model of cerebral vasospasm. Cell-free subarachnoid clots of autologous plasma in the basal cistern were found to produce no significant reaction of the basilar artery, while whole-blood clots induced a small (15%) chronic constriction after 24 hours. Because the plasma clots were not well retained in the basal cistern, however, small beads (dextran or latex) were added to stabilize them. Injection of beads and plasma led to moderate-to-severe chronic vasoconstriction (35% to 40%) with rapid onset. Control experiments demonstrated that these foreign bodies (beads) alone induced this vascular reaction. Histological examination showed that severe inflammation followed the introduction of subarachnoid beads. The experiments demonstrate that inflammation alone, in the absence of other processes associated with subarachnoid hemorrhage, may induce persistent and severe cerebroarterial constriction and raises the possibility that inflammation in response to subarachnoid blood may play a role in clinical vasospasm.

Possible role of the erythrocyte in causing prolonged cerebral vasospasm

1979 ◽  
Vol 51 (6) ◽  
pp. 773-778 ◽  
Author(s):  
Nobuyuki Ozaki ◽  
Sean Mullan
Keyword(s):  
Cerebral Vasospasm ◽  
Contractile Activity ◽  
Platelet Rich Plasma ◽  
Red Cells ◽  
Significant Activity ◽  
Sephadex Column ◽  
Content Type ◽  
Canine Basilar Artery

✓ Contractile activity of the various fractions of fresh and incubated blood was studied in vitro using the isolated canine basilar artery. Of the various fractions of fresh blood, significant contraction was induced by serum, but moderate contraction was induced by platelet-rich plasma and lysed red cells, while intact red cells and platelet-poor plasma had no significant activity. The contractions induced by serum and platelet-rich plasma were blocked by phenoxybenzamine, while those induced by lysed red cells were not. Whereas serum and platelet-rich plasma lost their contractile activity after 24 hours of incubation, lysed red cells retained activity up to 7 days after incubation. Biochemical analysis of the hemolysate by means of Sephadex column chromatography revealed that the contractile substance(s) possessed a molecular weight above 5000. These results suggest the possibility that the above substance(s) may play a role in prolonged cerebral vasospasm.

Reaction of Acetaldehyde with Human Platelets

1992 ◽  
Vol 67 (01) ◽  
pp. 126-130 ◽  
Author(s):  
Olivier Spertini ◽  
Jacques Hauert ◽  
Fedor Bachmann
Keyword(s):  
Platelet Aggregation ◽  
Inhibitory Action ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Shape Change ◽  
Reaction Medium ◽  
Human Platelets ◽  
Membrane Glycoproteins ◽  
Neutral Ph

SummaryPlatelet function defects observed in chronic alcoholics are not wholly explained by the inhibitory action of ethanol on platelet aggregation; they are not completely reproduced either in vivo by short-term ethanol perfusion into volunteers or in vitro by the addition of ethanol to platelet-rich plasma. As acetaldehyde (AcH) binds to many proteins and impairs cellular activities, we investigated the effect of this early degradation product of ethanol on platelets. AcH formed adducts with human platelets at neutral pH at 37° C which were stable to extensive washing, trichloracetic acid hydrolysis and heating at 100° C, and were not reduced by sodium borohydride. The amount of platelet adducts formed was a function of the incubation time and of the concentration of AcH in the reaction medium. At low AcH concentrations (<0.2 mM), platelet bound AcH was directly proportional to the concentration of AcH in the reaction medium. At higher concentrations (≥0.2 mM), AcH uptake by platelets tended to reach a plateau. The amount of adducts was also proportional to the number of exposures of platelets to pulses of 20 pM AcH.AcH adducts formation severely impaired platelet aggregation and shape change induced by ADP, collagen and thrombin. A positive correlation was established between platelet-bound AcH and inhibition of aggregation.SDS-PAGE analysis of AcH adducts at neutral pH demonstrated the binding of [14C]acetaldehyde to many platelet proteins. AcH adduct formation with membrane glycoproteins, cytoskeleton and enzymes might interfere with several steps of platelet activation and impair platelet aggregation.This in vitro study shows that AcH has a major inhibitory action on platelet aggregation and may account for the prolonged ex vivo inhibition of aggregation observed in chronic alcoholics even in the absence of alcoholemia.

Early Platelet-Collagen Interactions in Whole Blood and Their Modifications by Aspirin and Dipyridamole Evaluated by a New Method (Basic Wave)

1985 ◽  
Vol 54 (04) ◽  
pp. 799-803 ◽  
Author(s):  
José Luís Pérez-Requejo ◽  
Justo Aznar ◽  
M Teresa Santos ◽  
Juana Vallés
Keyword(s):  
Whole Blood ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
White Blood Cells ◽  
Reversible Aggregation ◽  
Inhibitory Compounds ◽  
Rapid Generation ◽  
Stimulation Of ◽  
Collagen Stimulation

SummaryIt is shown that the supernatant of unstirred whole blood at 37° C, stimulated by 1 μg/ml of collagen for 10 sec, produces a rapid generation of pro and antiaggregatory compounds with a final proaggregatory activity which can be detected for more than 60 min on a platelet rich plasma (PRP) by turbidometric aggregometry. A reversible aggregation wave that we have called BASIC wave (for Blood Aggregation Stimulatory and Inhibitory Compounds) is recorded. The collagen stimulation of unstirred PRP produces a similar but smaller BASIC wave. BASIC’s intensity increases if erythrocytes are added to PRP but decreases if white blood cells are added instead. Aspirin abolishes “ex vivo” the ability of whole blood and PRP to generate BASIC waves and dipyridamole “in vitro” significantly reduces BASIC’s intensity in whole blood in every tested sample, but shows little effect in PRP.

Dipyridamole Inhibits Platelet Aggregation in Whole Blood

1983 ◽  
Vol 50 (04) ◽  
pp. 852-856 ◽  
Author(s):  
P Gresele ◽  
C Zoja ◽  
H Deckmyn ◽  
J Arnout ◽  
J Vermylen ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Whole Blood ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Significant Negative Correlation ◽  
Significant Inhibition ◽  
Oral Drug ◽  
Human Blood Samples ◽  
Induced Aggregation

SummaryDipyridamole possesses antithrombotic properties in the animal and in man but it does not inhibit platelet aggregation in plasma. We evaluated the effect of dipyridamole ex vivo and in vitro on platelet aggregation induced by collagen and adenosine- 5’-diphosphate (ADP) in human whole blood with an impedance aggregometer. Two hundred mg dipyridamole induced a significant inhibition of both ADP- and collagen-induced aggregation in human blood samples taken 2 hr after oral drug intake. Administration of the drug for four days, 400 mg/day, further increased the antiplatelet effect. A significant negative correlation was found between collagen-induced platelet aggregation in whole blood and dipyridamole levels in plasma (p <0.001). A statistically significant inhibition of both collagen (p <0.0025) and ADP-induced (p <0.005) platelet aggregation was also obtained by incubating whole blood in vitro for 2 min at 37° C with dipyridamole (3.9 μM). No such effects were seen in platelet-rich plasma, even after enrichment with leukocytes. Low-dose adenosine enhanced in vitro inhibition in whole blood.Our results demonstrate that dipyridamole impedes platelet aggregation in whole blood by an interaction with red blood cells, probably involving adenosine.

Anti-Inflammatory Effects of Novel Standardized Platelet Rich Plasma Releasates on Knee Osteoarthritic Chondrocytes and Cartilage in vitro

2019 ◽  
Vol 20 (11) ◽  
pp. 920-933 ◽  
Author(s):  
Lucía Gato-Calvo ◽  
Tamara Hermida-Gómez ◽  
Cristina R. Romero ◽  
Elena F. Burguera ◽  
Francisco J. Blanco
Keyword(s):  
Gene Expression ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Cartilage Explants ◽  
Ex Vivo Model ◽  
Chondrocyte Viability ◽  
Platelet Concentration ◽  
The Absolute ◽  
Anti Inflammatory

Background: Platelet Rich Plasma (PRP) has recently emerged as a potential treatment for osteoarthritis (OA), but composition heterogeneity hampers comparison among studies, with the result that definite conclusions on its efficacy have not been reached. Objective: 1) To develop a novel methodology to prepare a series of standardized PRP releasates (PRP-Rs) with known absolute platelet concentrations, and 2) To evaluate the influence of this standardization parameter on the anti-inflammatory properties of these PRP-Rs in an in vitro and an ex vivo model of OA. Methods: A series of PRPs was prepared using the absolute platelet concentration as the standardization parameter. Doses of platelets ranged from 0% (platelet poor plasma, PPP) to 1.5·105 platelets/µl. PRPs were then activated with CaCl2 to obtain releasates (PRP-R). Chondrocytes were stimulated with 10% of each PRP-R in serum-free culture medium for 72 h to assess proliferation and viability. Cells were co-stimulated with interleukin (IL)-1β (5 ng/ml) and 10% of each PRP-R for 48 h to determine the effects on gene expression, secretion and intra-cellular content of common markers associated with inflammation, catabolism and oxidative stress in OA. OA cartilage explants were co-stimulated with IL-1β (5 ng/ml) and 10% of either PRP-R with 0.75·105 platelets/µl or PRP-R with 1.5·105 platelets/µl for 21 days to assess matrix inflammatory degradation. Results: Chondrocyte viability was not affected, and proliferation was dose-dependently increased. The gene expression of all pro-inflammatory mediators was significantly and dose-independently reduced, except for that of IL-1β and IL-8. Immunoblotting corroborated this effect for inducible NO synthase (NOS2). Secreted matrix metalloproteinase-13 (MMP-13) was reduced to almost basal levels by the PRP-R from PPP. Increasing platelet dosage led to progressive loss to this anti-catabolic ability. Safranin O and toluidine blue stains supported the beneficial effect of low platelet dosage on cartilage matrix preservation. Conclusion: We have developed a methodology to prepare PRP releasates using the absolute platelet concentration as the standardization parameter. Using this approach, the composition of the resulting PRP derived product is independent of the donor initial basal platelet count, thereby allowing the evaluation of its effects objectively and reproducibly. In our OA models, PRP-Rs showed antiinflammatory, anti-oxidant and anti-catabolic properties. Platelet enrichment could favor chondrocyte proliferation but is not necessary for the above effects and could even be counter-productive.

scholarly journals Application of the 1-µsec pulsed-dye laser to the treatment of experimental cerebral vasospasm

1991 ◽  
Vol 75 (2) ◽  
pp. 271-276 ◽  
Author(s):  
Atsushi Teramura ◽  
Robert Macfarlane ◽  
Christopher J. Owen ◽  
Ralph de la Torre ◽  
Kenton W. Gregory ◽  
...  
Keyword(s):  
Cerebral Vasospasm ◽  
Basilar Artery ◽  
Laser Energy ◽  
Autologous Blood ◽  
Preliminary Investigation ◽  
Dye Laser ◽  
Pulsed Dye Laser ◽  
Content Type

✓ Laser energy of 480 nm was applied in 1-µsec pulses varying between 2.2 and 10 mJ to in vitro and in vivo models of cerebral vasospasm. First, the pulsed-dye laser was applied intravascularly via a 320-µm fiber to basilar artery segments from six dogs. The segments were mounted in a vessel-perfusion apparatus and constricted to, on average, 70% of resting diameter by superfusion with dog hemolysate. Immediate increase in basilar artery diameter occurred to a mean of 83% of control. In a second model, the basilar artery was exposed transclivally in the rabbit. In three normal animals, superfusion of the artery with rabbit hemolysate resulted in a reduction of mean vessel diameter to 81% of control. Following extravascular application of the laser, vessels returned to an average of 106% of the resting state. In six rabbits, the basilar artery was constricted by two intracisternal injections of autologous blood, 3 days apart. Two to 4 days after the second injection, the basilar artery was exposed. Extravascular laser treatment from a quartz fiber placed perpendicular to the vessel adventitia resulted in an immediate 53% average increase in caliber to an estimated 107% of control. No reconstriction was observed over a period of up to 5 hours. Morphologically, damage to the arterial wall was slight. This preliminary investigation suggests that the 1-µsec pulsed-dye laser may be of benefit in the treatment of cerebral vasospasm.

Cerebrovascular characterization of clazosentan, the first nonpeptide endothelin receptor antagonist clinically effective for the treatment of cerebral vasospasm. Part I: Inhibitory effect on endothelinA receptor—mediated contraction

2005 ◽  
Vol 102 (6) ◽  
pp. 1101-1107 ◽  
Author(s):  
Hartmut Vatter ◽  
Michael Zimmermann ◽  
Veronika Tesanovic ◽  
Andreas Raabe ◽  
Lothar Schilling ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Cerebral Vasospasm ◽  
Isometric Force ◽  
Inhibitory Effect ◽  
Affinity Constant ◽  
Dependent Manner ◽  
Content Type ◽  
The Right ◽  
Fine Print

Object. The central role of endothelin (ET)—1 in the development of cerebral vasospasm after subarachnoid hemorrhage is indicated by the successful treatment of this vasospasm in several animal models by using selective ETA receptor antagonists. Clazosentan is a selective ETA receptor antagonist that provides for the first time clinical proof that ET-1 is involved in the pathogenesis of cerebral vasospasm. The aim of the present investigation was, therefore, to define the pharmacological properties of clazosentan that affect ETA receptor—mediated contraction in the cerebrovasculature. Methods. Isometric force measurements were performed in rat basilar artery (BA) ring segments with (E+) and without (E−) endothelial function. Concentration effect curves (CECs) were constructed by cumulative application of ET-1 or big ET-1 in the absence or presence of clazosentan (10−9, 10−8, and 10−7 M). The inhibitory potency of clazosentan was determined by the value of the affinity constant (pA2). The CECs for contraction induced by ET-1 and big ET-1 were shifted to the right in the presence of clazosentan in a parallel dose-dependent manner, which indicates competitive antagonism. The pA2 values for ET-1 were 7.8 (E+) and 8.6 (E−) and the corresponding values for big ET-1 were 8.6 (E+) and 8.3 (E−). Conclusions. The present data characterize clazosentan as a potent competitive antagonist of ETA receptor—mediated constriction of the cerebrovasculature by ET-1 and its precursor big ET-1. These functional data may also be used to define an in vitro profile of an ET receptor antagonist with a high probability of clinical efficacy.

scholarly journals Ex VivoMaturation Assay for Testing Antimalarial Sensitivity of Rodent Malaria Parasites

2016 ◽  
Vol 60 (11) ◽  
pp. 6859-6866 ◽  
Author(s):  
Zi Wei Chang ◽  
Benoit Malleret ◽  
Bruce Russell ◽  
Laurent Rénia ◽  
Carla Claser
Keyword(s):  
Ex Vivo ◽  
Single Step ◽  
Parasite Development ◽  
Ring Stage ◽  
Malaria Parasites ◽  
Rodent Malaria ◽  
Content Type ◽  
Parasite Biology

ABSTRACTEx vivoassay systems provide a powerful approach to studying human malaria parasite biology and to testing antimalarials. For rodent malaria parasites, short-termin vitroculture andex vivoantimalarial susceptibility assays are relatively cumbersome, relying onin vivopassage for synchronization, since ring-stage parasites are an essential starting material. Here, we describe a new approach based on the enrichment of ring-stagePlasmodium berghei,P. yoelii, andP. vinckei vinckeiusing a single-step Percoll gradient. Importantly, we demonstrate that the enriched ring-stage parasites develop synchronously regardless of the parasite strain or species used. Using a flow cytometry assay with Hoechst and ethidium or MitoTracker dye, we show that parasite development is easily and rapidly monitored. Finally, we demonstrate that this approach can be used to screen antimalarial drugs.

scholarly journals Correlates between Models of Virulence for Mycobacterium tuberculosis among Isolates of the Central Asian Lineage: a Case for Lysozyme Resistance Testing?

2015 ◽  
Vol 83 (6) ◽  
pp. 2213-2223 ◽  
Author(s):  
Claire Pardieu ◽  
Nicola Casali ◽  
Simon O. Clark ◽  
Richard Hooper ◽  
Ann Williams ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Ex Vivo ◽  
Resistance Testing ◽  
Snp Analysis ◽  
Single Nucleotide ◽  
Content Type ◽  
Differential Outcomes ◽  
Central Asian ◽  
Strain H37rv

Virulence factors (VFs) contribute to the emergence of new humanMycobacterium tuberculosisstrains, are lineage dependent, and are relevant to the development ofM. tuberculosisdrugs/vaccines. VFs were sought withinM. tuberculosislineage 3, which has the Central Asian (CAS) spoligotype. Three isolates were selected from clusters previously identified as dominant in London, United Kingdom. Strain-associated virulence was studied in guinea pig, monocyte-derived macrophage, and lysozyme resistance assays. Whole-genome sequencing, single nucleotide polymorphism (SNP) analysis, and a literature review contributed to the identification of SNPs of interest. The animal model revealed borderline differences in strain-associated pathogenicity.Ex vivo, isolate C72 exhibited statistically significant differences in intracellular growth relative to C6 and C14. SNP candidates inducing lower fitness levels included 123 unique nonsynonymous SNPs, including three located in genes (lysX,caeA, andponA2) previously identified as VFs in the laboratory-adapted reference strain H37Rv and shown to confer lysozyme resistance. C72 growth was most affected by lysozymein vitro. A BLAST search revealed that all three SNPs of interest (C35F, P76Q, and P780R) also occurred in Tiruvallur, India, and in Uganda. Unlike C72, however, no single isolate identified through BLAST carried all three SNPs simultaneously. CAS isolates representative of three medium-sized human clusters demonstrated differential outcomes in models commonly used to estimate strain-associated virulence, supporting the idea that virulence varies within, not just across,M. tuberculosislineages. Three VF SNPs of interest were identified in two additional locations worldwide, which suggested independent selection and supported a role for these SNPs in virulence. The relevance of lysozyme resistance to strain virulence remains to be established.

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