Platelet Factor 4 (PF4) And β-Thromboglobulin (BTG) In Atherosclerosis And Diabetes Mellitus

Distinguishing between in vitro and in vivo release of platelet specific proteins has led to problems in interpretation of clinical studies with PF4, and with BTG.As part of a prospective study of peripheral occlusive arterial disease in diabetes mellitus PF4 and BTG were measured together by specific radioimmunoassay in four groups of subjects age 50-70 years. Groups included (1) 106 normal subjects (NC); (2) 126 with clinical and objective evidence (transcutaneous doppler ultrasound and treadmill exercise) of peripheral arterial occlusive disease (ASO); (3) 244 with diabetes mellitus without ASO and (4) 101 with both diabetes mellitus and ASO.PF4 did not distinguish among the four groups: NC vs ASO p=. 18, NC vs DM p=.52, NC vs DM+ASO p=.73, NC vs all diabetics p=.61, and NC vs all ASO p=.46. However, BTG did distinguish the groups with vascular disease from NC: NC vs ASO p=.02, NC vs DM+ASO p<.001, NC vs DM p=.32, DM vs DM+ASO p<.01, all ASO vs all nonASO p<.001.Normal range for PF4 is 0-13 ng/ml with a median of 3.0 ng/ml, a mean of 3.0 ng/ml, and SD 1.97. Normal range for BTG is 14-46 ng/ml with a median of 26 ng/ml, a mean of 27.2 ng/ml, and SD 10.3. When PF4 and BTG are correlated (all groups included) r=.41 p<.001.PF4 and BTG levels should be performed together. When done together low PF4 levels support the absence of in vitro platelet release due to sample collection and handling. BTG levels are significantly increased in subjects with peripheral arterial disease, supporting the active role of platelets in thrombotic occlusive arterial disease.

Serial Platelet Survival Half-Life (PS) In Peripheral Vascular Disease And Diabetes Mellitus

10.1055/s-0038-1652857 ◽

1981 ◽

Cited By ~ 1

Author(s):

F J Kazmier ◽

V Fuster ◽

J H Chesebro ◽

J H O’Fallon ◽

P J Palumbo

Keyword(s):

Diabetes Mellitus ◽

Half Life ◽

Normal Subjects ◽

Arterial Disease ◽

Least Square ◽

Computer Assisted ◽

Occlusive Arterial Disease ◽

Platelet Survival ◽

Objective Evidence ◽

Mayo Clinic, Mayo Foundation, Rochester, MinnesotaPlatelets are believed to play an important role in the pathogenesis of atherosclerosis and thrombotic occlusive arterial disease. As part of a prospective study of peripheral occlusive arterial disease in diabetes mellitus, platelet survival half-life (PS) was determined in years 1 and 2 from the disappearance pattern of 51cr-labeled platelets obtained by measuring radioactivity in serial samples over 8 days in four groups of patients age 50-70. Using computer assisted least-square analysis, a single exponent is fitted to the data and half-life determined.Groups included (1) normal subjects (NC); (2) subjects with clinical and objective evidence (transcutaneous doppler ultrasound and treadmill exercise) of peripheral arterial occlusive disease (AS0); (3) subjects with diabetes mellitus without AS0 (DM); (A) subjects with both diabetes mellitus and AS0 (DM+AS0). 112 subjects were tested in year 1 and 75 of these were available for retesting in year 2.In year 1, there is a difference in the distribution of platelet survival (PS) for the four groups (p=.002). PS is significantly short in groups AS0 and DM+AS0 compared with controls (NC), Year 2 values for PS (75 subjects) were compared to year 1. Using the t test on mean differences PS for each group was not significantly different year 2 vs year 1; mean percent differences year 2 vs year 1 were 1.2% (NC), 0.2% (ASO), 3.2% (DM), and -9.3% (9-3% lower than year 1 for DM+ ASO).Platelet survival half-life (PS) does distinguish popula tion groups and is reproducible when repeated.

Abstract 5281: Bioluminescence Imaging of X-Ray Visible Microencapsulated Mesenchymal Stem Cells for Cell Based Therapy of Peripheral Arterial Disease

Circulation ◽

10.1161/circ.118.suppl_18.s_519-a ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Dorota A Kedziorek ◽

Piotr Walczak ◽

Yingli Fu ◽

Nicole Azene ◽

Aravind Arepally ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Arterial Disease ◽

X Ray ◽

Encapsulated Cells ◽

X Ray Imaging ◽

Introduction: Therapeutic angiogenesis in Peripheral Arterial Disease (PAD) using stem cell therapy is potentially complicated by immunorejection. To overcome this problem, microen-capsulation using the alginate-poly-L-lysine (PLL)-alginate (APA) method was developed to provide a protective porous bubble to block antibodies but allow exchange of small molecules. Recently, we have developed a method to enable X-ray detection of these capsules. However, cell survival within the capsules could not be determined. Plus PLL can be mildly cytotoxic. In the present study, we combined reporter gene methods to verify cell survival with X-ray detection of the microcapsules in a rabbit PAD model and studied the PLL impact on cell viability. Methods: Rabbit mesenchymal stem cells (MSCs) were transfected with triple fusion (TF) reporter gene for bioluminescence (firefly luciferase), fluorescence (red fluorescent protein) and PET (truncated thymidine kinase). TF-MSCs were encapsulated in the perfluorooctyl bromide (PFOB) capsules to enable computed tomographic detection. Capsule crosslinking was performed with three PLL concentrations, i.e., 0.005%, 0.025% and 0.05%. Bioluminescent imaging (BLI) was used to monitor cells survival for one week in vitro and after intramuscular injection in vivo . Results: Serial in vitro BLI enabled the detection of viable encapsulated MSCs without detrimental signal degradation (~13% decrease of BLI signal intensity after PFOB encapsulation comparing to equal number of naked MSCs). PLL did not result in cell death; higher PLL concentrations were correlated with stronger BLI signal. BLI signal production was only slightly reduced by second layer of alginate (~80% for 0.05% PLL). In vivo BLI demonstrated the detection of naked, APA, and PFOB-encapsulated TF-MSCs. X-ray imaging enabled PFOB microcapsules detection relative to vasculature. Conclusion: BLI allows monitoring of encapsulated cells survival. PLL concentrations ≤ 0.05% appear safe for encapsulated cells with higher concentration being associated with enhanced crosslinking and capsule stability. MSCs expressing TF reporter in PFOB microcapsules enables dual monitoring of cell delivery/capsule tracking by X-ray imaging and cell viability with BLI.

Platelet Activation Markers in Patients with Peripheral Arterial Disease

Thrombosis and Haemostasis ◽

10.1055/s-0038-1665429 ◽

1997 ◽

Vol 78 (06) ◽

pp. 1434-1437 ◽

Cited By ~ 33

Author(s):

Paolo Gresele ◽

Mariella Catalano ◽

Carlo Giammarresi ◽

Raul Volpato ◽

Rosanna Termini ◽

...

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Platelet Function ◽

Arterial Disease ◽

Platelet Function Test ◽

Activation Markers ◽

Significant Difference ◽

SummaryPeripheral vascular disease (PVD) is an indicator of diffuse atherosclerosis and is associated with a greatly increased incidence of coronary heart and cerebrovascular disease. Although several studies have assessed whether in vivo platelet activation takes place in patients with PVD, no data are available comparing different platelet function tests in this patient population.We have compared prospectively four tests for the measurement of in vivo platelet activation (plasma βTG, plasma PF4, intraplatelet (βTG and urinary excretion of 11-dehydro-TXB2) and one in vitro platelet function test (ADP-induced platelet aggregation) in 63 well-characterized patients with intermittent claudication and in 18 age- and sex- matched healthy volunteers.No statistically significant difference was found between patients and controls for plasma βTG (20.0 ± 11.8 vs. 18.8 ± 9.0 ng/ml, respectively), plasma PF4 (5.2 ± 2.9 vs. 6.3 ± 3.5 ng/ml), βTG/PF4 ratio (4.0 ± 2.9 vs. 3.6 ± 1.8), intraplatelet pTG (4503 ± 1482 vs. 4059 ± 1065 ng/ml), and threshold aggregatory concentration of ADP (1.7 ± 0.72 vs. 1.45 ± 0.56 μM).Urinary 11-dehydro-TXB2 was instead significantly higher in the PVD group (55.4 ± 27.5 vs. 26.7 ± 7.0 ng/h, p <0.001).Our study shows that urinary 11-dehydro-TXB2 is a more sensitive index of in vivo platelet activation than the measurement of either platelet specific proteins or of in vitro platelet aggregation in patients with PVD.

In vitro evaluation of nanoparticle drug-coated balloons: a pectin-RGDS-OC8H17-paclitaxel solution

Applied Nanoscience ◽

10.1007/s13204-021-01736-4 ◽

2021 ◽

Author(s):

Peng Wang ◽

Lin Gui ◽

Yuji Wang ◽

Sheng Wang

Keyword(s):

Clinical Importance ◽

Arterial Disease ◽

Transluminal Angioplasty ◽

Control Group ◽

Effective Therapeutic Strategy ◽

Artery Disease ◽

Peripheral Arterial ◽

Novel Drug

AbstractDrug-coated balloons have proved to be an effective technology in percutaneous transluminal angioplasty in treating peripheral artery disease. Paclitaxel-based coating is mainly used. Solutions to such problems as drug loss and inefficient drug release during operations, however, have not been found yet. This study aims to explore the activity of a newly designed paclitaxel-coated balloon in vitro using pectin as the excipient (pectin-paclitaxel) compared with the commercially available shellac excipient balloon, and to characterize the novel nanoparticle paclitaxel-coated balloon with peptide (Arg-Gly-Asp-Ser, RGDS) derivative RGDS-OC8H17 (pectin-RGDS-OC8H17-paclitaxel). Two coating solutions, pectin-paclitaxel and pectin-RGDS-OC8H17-paclitaxel, were successively designed and prepared. The morphology of both coating solutions was first characterized compared with the control group, the commercially available paclitaxel-coated balloon. Then the in vitro experiments were conducted to determine the drug-releasing profiles of both pectin-paclitaxel and pectin-RGDS-OC8H17-paclitaxel coatings. The pectin-RGDS-OC8H17-paclitaxel-coated balloon was smoother and more homogeneous compared with the commercially available paclitaxel-coated balloon and the pectin-paclitaxel-coated balloon. This difference was more obvious when paclitaxel was at low concentration. During the in vitro trial, the drug-releasing curve of the pectin-RGDS-OC8H17-paclitaxel model showed an adjustable paclitaxel-releasing: more than 90% of the paclitaxel released in 2 h at 300 rpm and more than 99% released in 10 min at 1200 rpm. Compared to the performance of the current commercially available shellac excipient products and the pectin-paclitaxel coating, pectin-RGDS-OC8H17-paclitaxel coating provided higher drug-releasing speed. However, the clinical outcomes of this finding need to be further demonstrated. Paclitaxel-coated balloons as an effective therapeutic strategy currently in treating peripheral arterial disease need to be further improved in terms of its efficiency in anti-proliferative drug delivery and release. The pectin-RGDS-OC8H17-paclitaxel coating solution developed in this study exhibited excellent drug-releasing properties. Further experiments are still needed to demonstrate the performance of this novel drug-coated balloon in vivo and its clinical importance.

m7G Methyltransferase METTL1 Promotes Post-ischemic Angiogenesis via Promoting VEGFA mRNA Translation

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.642080 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yongchao Zhao ◽

Lingqiu Kong ◽

Zhiqiang Pei ◽

Fuhai Li ◽

Chaofu Li ◽

...

Keyword(s):

Mrna Translation ◽

Arterial Disease ◽

Therapeutic Angiogenesis ◽

Tube Formation ◽

Dependent Manner ◽

Global Increase ◽

Plasmid Transfection ◽

Post-transcriptional modifications play pivotal roles in various pathological processes and ischemic disorders. However, the role of N7-methylguanosine (m7G), particularly m7G in mRNA, on post-ischemic angiogenesis remains largely unknown. Here, we identified that methyltransferase like 1 (METTL1) was a critical candidate responsible for a global decrease of m7G within mRNA from the ischemic tissues. The in vivo gene transfer of METTL1 improved blood flow recovery and increased angiogenesis with enhanced mRNA m7G upon post-ischemic injury. Increased METTL1 expression using plasmid transfection in vitro promoted HUVECs proliferation, migration, and tube formation with a global increase of m7G in mRNA. Mechanistically, METTL1 promoted VEGFA mRNA translation in an m7G methylation-dependent manner. Our findings emphasize a critical link between mRNA m7G and ischemia and provide a novel insight of targeting METTL1 in the therapeutic angiogenesis for ischemic disorders, including peripheral arterial disease.

ADAM12: a genetic modifier of preclinical peripheral arterial disease

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00803.2014 ◽

2015 ◽

Vol 309 (5) ◽

pp. H790-H803 ◽

Cited By ~ 16

Author(s):

Ayotunde O. Dokun ◽

Lingdan Chen ◽

Mitsuharu Okutsu ◽

Charles R. Farber ◽

Surovi Hazarika ◽

...

Keyword(s):

Inbred Mouse ◽

Genetic Modifier ◽

Arterial Disease ◽

Endothelial Cell Proliferation ◽

Mouse Strains ◽

Trait Locus ◽

In prior studies from multiple groups, outcomes following experimental peripheral arterial disease (PAD) differed considerably across inbred mouse strains. Similarly, in humans with PAD, disease outcomes differ, even when there are similarities in risk factors, disease anatomy, arteriosclerotic burden, and hemodynamic measures. Previously, we identified a locus on mouse chromosome 7, limb salvage-associated quantitative trait locus 1 (LSq-1), which was sufficient to modify outcomes following experimental PAD. We compared expression of genes within LSq-1 in Balb/c mice, which normally show poor outcomes following experimental PAD, with that in C57Bl/6 mice, which normally show favorable outcomes, and found that a disintegrin and metalloproteinase gene 12 ( ADAM12) had the most differential expression. Augmentation of ADAM12 expression in vivo improved outcomes following experimental PAD in Balb/c mice, whereas knockdown of ADAM12 made outcomes worse in C57Bl/6 mice. In vitro, ADAM12 expression modulates endothelial cell proliferation, survival, and angiogenesis in ischemia, and this appeared to be dependent on tyrosine kinase with Ig-like and EGF-like domain 2 (Tie2) activation. ADAM12 is sufficient to modify PAD severity in mice, and this likely occurs through regulation of Tie2.

Delivery of hepatocyte growth factor mRNA from nanofibrillar scaffolds in a pig model of peripheral arterial disease

Regenerative Medicine ◽

10.2217/rme-2020-0023 ◽

2020 ◽

Vol 15 (6) ◽

pp. 1761-1773

Author(s):

Tatiana S Zaitseva ◽

Guang Yang ◽

Dimitris Dionyssiou ◽

Maedeh Zamani ◽

Steve Sawamura ◽

...

Keyword(s):

Therapeutic Potential ◽

Arterial Disease ◽

Translational Efficiency ◽

Collagen Scaffolds ◽

Vascular Regeneration ◽

Peripheral Arterial ◽

Hepatocyte Growth Factor Mrna

Background: Chemical modification of mRNA (mmRNA) substantially improves their stability and translational efficiency within cells. Nanofibrillar collagen scaffolds were previously shown to enable the spatially localized delivery and temporally controlled release of mmRNA encoding HGF both in vitro and in vivo. Materials & methods: Herein we developed an improved slow-releasing HGF mmRNA scaffold and tested its therapeutic efficacy in a porcine model of peripheral arterial disease. Results & conclusion: The HGF mmRNA was released from scaffolds in a temporally controlled fashion in vitro with preserved transfection activity. The mmRNA scaffolds improved vascular regeneration when sutured to the ligated porcine femoral artery. These studies validate the therapeutic potential of HGF mmRNA delivery from nanofibrillar scaffolds for treatment of peripheral arterial disease.

Change on Blood Coagulation, Platelet Function and Fibrinolysis in Diabetes Mellitus - Preliminary Report

10.1055/s-0039-1682387 ◽

1977 ◽

Author(s):

R. Giustolisi ◽

R. Musso ◽

T. Lombardo ◽

M. Russoand ◽

E. Cacciola

Keyword(s):

Diabetes Mellitus ◽

Degradation Products ◽

Vascular Wall ◽

Diabetic Patients ◽

Platelet Adhesiveness ◽

Platelet Factor ◽

Protamine Sulphate ◽

At Iii

Some coagulation aspects are studied in diabetes mellitus because this dismetabo-lic disease represents a “high risk factor” of predisposition leading to classical lesions of the vascular wall and thrombosis. Were studied 24 diabetic patients between 16 and 68 years old and 14 healthy subjects. Tests performed are followed: partial thromboplastyn time(PTT), plasma coagulation time RVV(RVV-T), antithrombin III(At-III), alpha2macroglobulin(a2M), fibrin/fibrinogen degradation products(FDP), ethanol gelation(EG) and protamine sulphate(PS), euglobulin lysis time(ELT), platelet adhesiveness to glass(PAG), platelet adhesiveness in vivo (PAV), platelet factor-3 availability(PF-3), platelet aggregation by ristocetin 1, 2-1, 5-1, 8 mg/ml(RIPA),Diabetics showed a fall in At-III, increase a2M, a significant decrease ELT and increase FDP with often positivity EG. We also noted a shortening of PTT, PF-3 rate and RVV-T. In vitro platelets adhesiveness rises more than it does in vivo. Besides the PPP from diabetics increased the control subjects PAG. The RIPA is increased. Our findings showed, therefore, in diabetic patients a thrombophilic pattern by blood hypercoagulability and fibrinolytic activity decreased.

Thrombotect: An Optimized Sample Collection System For The Inhibition Of In Vitro Platelet Release

10.1055/s-0038-1652356 ◽

1981 ◽

Author(s):

E F Workman ◽

D M Clark

Keyword(s):

In Vitro Release ◽

Blood Collection ◽

Sample Collection ◽

Collection System ◽

Platelet Factor ◽

Thrombotic Disease ◽

Disease States ◽

Platelet Release

Platelet activation occurs in a variety of thrombotic disease states and may be monitored via measurement of Platelet Factor 4 (PF4) levels in plasma. In order for PF4 levels to reflect in vivo activation, care must be taken to eliminate in vitro α -granule release during the collection and preparation of blood specimens. Thrombotect is an evacuated blood collection tube containing an anticoagulant (EDTA) and two platelet inhibitors (2-chloroadenosine and procaine-HCl). This system is effective in preventing in vitro release under a variety of centrifugation conditions. Subsequent to a 30 minute icing step, whole blood samples may be centrifuged at room temperature and one of three centrifugation speeds - 2500 × g for 20 minutes,1500 × g for 30 minutes, and 1000 × g for 40 minutes. Plasma specimens from normal individuals were assayed after being prepared under the above conditions. PF4 levels were as follows: 2500 × g, *2.79 ± 1.00 (73); 1500 × g, 3.84 ±1.78 (20); 1000 × g, 3.35 ± 1.65 (20). Variations in the temperature of centrifugation had little effect on the ability of Thrombotect to inhibit PF4 release, whereas other commonly used blood collection cocktails became progressively less effective with increasing temperature.*(Mean PF4 in ng/ml ± 1 S.D., “n” values in parenthesis). Results similar to those obtained for EDTA/Theophylline were obtained when EDTA was used in conjunction with adenosine and/or aspirin. The Thrombotect system has been employed in an examination of PF4 levels in a variety of thrombotic disease states. It also provides an excellent collection system for use with other “platelet-release” assays.

Platelet Activation During Treadmill Exercise In Patients With Chronic Peripheral Arterial Disease

10.1055/s-0038-1652386 ◽

1981 ◽

Cited By ~ 1

Author(s):

G Baele ◽

H Bogaerts ◽

D L Clement ◽

R Pannier ◽

F Barbier

Keyword(s):

Platelet Activation ◽

Plasma Levels ◽

Treadmill Exercise ◽

Arterial Disease ◽

Mean Value ◽

Platelet Factor ◽

Normal Individuals ◽

β-Thromboglobulin (β-TG) and platelet factor 4 (PF-4) are specific platelet proteins released during in vivo platelet activation. An increase in PF-4 after exercise- induced myocardial ischemia was reported by Green, L.H. et al.. This observation prompted us to measure β-TG and PF-4 in patients with chronic occlusive arterial disease of the lower limbs and to look for increments during treadmill exercise. β-TG was measured using the Amersham test kit and PF-4 using the radioimmunoassay kit of Abbott Diagnostics Division. Plasma levels in 28 normal individuals ranged for β-TG from 7 to 39 ng/ml with a mean value of 21.0 ng/ml, for PF-4 from 1 to 19 ng/ml with a mean value of 6.0 ng/ml. β-TG and PF-4 were measured in 59 patients with chronic peripheral arterial disease before and 5 min. after treadmill exercising till occurrence of claudication. Plasma levels of β-TG before treadmill exercising ranged from 24 to 260 ng/ml with a mean of 77.9 ng/ml, PF-4 levels ranged from 2 to 240 ng/ml with a mean of 30.4 ng/ml. These levels were significantly higher than those measured in normal individuals.After treadmill exercise β-TG levels showed a statistically significant increase to a mean value of 87.3 ng/ml but PF-4 did not rise significantly (mean value : 32.4 ng/ml). The supplementary increase of already elevated β-TG levels may be explained by enhanced in vivo platelet activation during treadmill exercising till occurrence of claudication. As the clearance of PF-4 from human plasma has been shown to be much faster than the clearance of β-TG increases in PF-4 levels may be more difficult to detect during dynamic explorations of the vascular system.