Platelet retention

Another study [114] on the role of protonated amine of HA copolymers in platelet retention showed that minimum retention was again observed on the surface of the copolymer containing N+ in 0.1 0.3 wt%. In other words, cell attachment scarcely takes place on the surface of HA2, in contrast to poly-HEMA, on the surface of which more than 90 percent cell attachment was observed. 

Schwartz BS, Leis LA, Johnson GJ. In vivo platelet retention in hianan bleeding-time wounds II. Effect of aspirin ingestion. J Lab Clin Med 1979 94 574-84... 

Bailly. A.L, Laurent A, Lu. H, Elalami. I, Jacob. P, Mundler. O, MerlancL J.J, Lautier. A, Soria J and Soria C, Fibrinogen binding and platelet retention relationdiip with die thrombogenicaty of catheters, J. Biomed. Mater. Res., 30,101-108 (1996). 

Figure 9 Degree of platelet retention in unheparinized rabbits undergoing simulated extracorporeal membrane oxygenation (ECMO) whose extracorporeal blood conduits are lined with MAHMA/NO-containing PVC to inhibit platelet adhesion ( ) as compared with controls whose tubing is made of undiazeniumdiolated PVC ( ). [Adapted from Reference 13 with permission.]...

A rapid, minimally invasive in vivo animal model was developed to evaluate continuously the process of thrombosis on biomaterials. Goats subjected acutely to bilateral polyethylene catheters are monitored for net retention of luln-oxine platelets on the separate catheters. Comparisons between continuous scintigraphic monitoring of platelet retention and thrombus recovered at catheter retrieval show that platelet retention and thrombus growth are dynamic processes for at least the first few hours of implantation, and that platelet retention is predictive for thrombus size. Mean recovered thrombus weight is 23 mg cm, which corresponds to a 44% increase in effective radius of the catheter-thrombus combination. 

Members of the biomaterials community continue to search for experimental animal models that assess the thromboresistance of polymers. We developed a new animal model involving rapid, simple retrograde cannu-lation of the goat s carotid arteries. The method promises to assess potential biomaterials, evaluate drugs that may decrease thrombus growth, and measure real-time thrombus growth and dissolution. A critical aspect of this new experimental model is that the continuously monitored net platelet retention data can be modeled mathematically. 

Circulating radiolabeled platelets adhered and aggregated on two polyethylene catheters inserted retrograde into the carotid arteries. Increasing platelet retention is shown in the sequence A through EThe beginning of thrombus dissolution is seen in F . The small platelet collection is best shown in the precatheter sample (A). Blood was drawn... 

Figure 3. Net platelet retention behavior on polyethylene catheters in four goats. Thrombus platelet concentration catheter length was used to calibrate externally detected platelet retention curves. The repeatability of the initial slope ana the variable times to peak are shown. Key I, Goat 32 2, Goat 36 ...

A model that predicts accurately thromboembolic risk associated with the use of biomaterials is still needed. Utilizing minimal surgical techniques, catheters were placed in goat carotid arteries, and subsequent analysis in real time was performed noninvasively of net platelet retention of autologously labeled nlIn platelets. We concentrated on acute animal studies using polyethylene catheters. Price et al. used a very similar model in dogs, obtaining similar results (9). 

The reversal in net platelet retention is very rapid, is never complete, and is exponential in quality. 

Despite the fact that we are measuring net platelet retention, we can probably predict thrombus weight from platelets retained (r = 0.9). A similar correlation (r = 0.88) exists for dogs, comparing clot weight to platelet activity (%/centimeter) (9). 

Several polymers were evaluated in the form of a surface coating on glass beads packed in columns to determine their ability to retain platelets when whole human blood passes over the surface. This ability was measured as the platelet retention index p, the fraction of platelets retained on the column. Lowest values of p were found for poly(ethylene oxide), polypropylene oxide), poly(tetramethylene oxide) (in the form of polyurethanes), and polydimethylsiloxane. Highest values (around 0.8) were found for cross-linked poly(vinyl alcohol) and the copolymers of ethylenediamine with diisocyanates. Intermediate values were found for polystyrene and its copolymers with methyl acrylate, for polyacrylate, and for poly(methyl methacrylate). The results are interpreted in terms of possible hydrophobic and hydrogen bonding interactions with plasma proteins. 

This chapter deals with a specific test of blood-surface interaction in vitro platelet retention in a column of beads (due to platelet adhesion and aggregation). Protein adsorption precedes platelet adsorption, and thus the in vitro platelet retention test involves competitive and sequential adsorption of proteins, the outcome of which produces surfaces having widely varying degrees of platelet retention. Except in the case of thrombin (3), plasma protein absorption on these surfaces has not been studied. 

Figure 1. In vitro test for platelet retention, using plastic columns, packed with fine glass beads whose surfaces are thereafter coated with test polymer (ambient temperature = 37°C).

Ten polymers were studied in respect to platelet retention, as summarized in Figure 2. 

Figure 2. Composite display of average platelet retention index p for selected polymer types. The acrylates, but not polystyrenes, are significantly passivated by pretreatment with plasma (n = 6). Key , lowest value observed in each case and PPP, preincubated in platelet-poor plasma.

We tested the hypothesis that the platelet retention index should increase as polymer composition varied from 100 mol % methyl acrylate to 100 mol % styrene, the respective platelet retention indices p being about 0.25 and 0.55 for the homopolymers. The results are shown in Figure 3, wherein p increases to a maximum near 40% styrene. When the surfaces were incubated in platelet-poor plasma before contact with whole blood, the values of p were much reduced (from 0.25 to 0.05 for methyl acrylate) for copolymers containing up to 60 mol % styrene. Copolymers of higher styrene content were not rendered significantly less retentive by plasma pretreatment. 

Thus, time of water exposure prior to challenging platelets is probably important, but the matter needs further study. Our tests suggest that increasing the hydrophobic side group length increases platelet retention in these experiments surfaces were exposed to isotonic saline solution for less... 

Figure 3. Average platelet retention index p as a function of mole fraction of styrene in a series of copolymers of methyl acrylate and styrene. Key —, control and —, pretreated in platelet-poor plasma.

Preincubation of surfaces with platelet-poor plasma substantially reduced the platelet retention index of polyacrylates and polymethacrylates, but hardly altered the retention index of polystyrene or copolymers with methyl acrylate that are rich in styrene. Even without plasma pretreatment, the surface, when exposed to whole blood, was probably first contacted by molecular elements (including the proteins) before the cellular elements arrived. What then is the mode of action of plasma pretreatment Several hypotheses, none conclusive, can be advanced, such as ... 

The evidence summarized in Figure 2 shows lowest platelet retention indices for polyethylene oxide) (PEO), polypropylene oxide) (PPO), and PDMS. This result implies low degrees of adsorption of specific critical plasma proteins and/or adsorption without conformation alteration of the protein. The molecular attributes these three polymers have in common appear to be ... 

These observations indicate a need to refine the meaning of hydro-phobicity in connection with protein interaction and platelet retention. 

Before long-term hemocompatibility can be expected for any material, the nature of polymer surface-protein interaction must be established in more detail the way in which the polymer surface alters itself (rotation of segments, side groups, chain refolding, etc.) in response to the protein species the way the protein is altered in conformation (if at all) upon adsorption by the surface and how this conformational change provokes platelet retention. Of course, longer-term ex vivo or in vivo studies also will be necessary. 

Molecular models (2) of the hard-segment phase of segmented polyether polyurethanes tested in the in vitro bead column (4) show a high platelet retention index (p = 0.8) regardless of molecular composition. The variable p is defined as the fraction of platelets in whole citrated human blood entering a test column that are retained on the bead surfaces, averaged for... 

Upon contact with blood, polyethers absorb varying amounts of water and undergo partial dissolution while remaining partially crystalline. They show platelet retention index values around 0.2-0.3. 

Parallel tests show that thrombin adsorption is minimal on well-prepared polyurethanes containing amorphous PEO, greater on PTMO, and very high on analogues of the hard segment (diisocyanate-diamine copolymers), thus paralleling the trends in platelet retention index p. This result is consistent with the postulate that protein adsorption must precede platelet adsorption. 

These polyurethanes (and their precursors or analogues) were cast from an appropriate solvent, usually DMF, onto the glass bead surfaces used in the in vitro test for platelet retention (4), or for the thrombin absorption test used previously (2). Crystals of KBr for Fourier transform infrared (FTIR) spectroscopy and glass microscopic slides for examination by XPS (ESCA) served as supports for polymers cast from the same solvents. Concentration of polymer (5 wt %), temperature of casting... 

C), and subsequent slow evaporation conditions under argon gas (40°C, 21 days) were maintained as closely identical as possible in preparing surfaces for each test (FTIR, ESC A, and platelet retention). The air side of the film was the side exposed to platelets and examined by XPS. The other side (against the glass substrate) was not exposed. 

Figure 3. Average platelet retention index vs. , where = fraction of C Is...

Second, the highly crystalline and hydrogen-bonding alternating copolymer of diisocyanate and diamine, an analog of the hard segment of polyurethanes, shows high platelet retention (p 0.8). Therefore, this molecular species will cause platelet retention, to the extent that it appears at the surface of polyurethanes. 

Thus, ESCA C Is spectra present a remarkable correlation of platelet retention indices, provided that crystalline materials are excluded from the analysis. However, at any value of <(> in Figure 3, a range of p exists. 

By XPS, much less nitrogen was found in their surfaces than in most of the polymers reported in Table III, and < >, the fraction of ethereal carbon, ranged upward from 0.8. When PEO 3500 was used as the diol, the XPS scans of the segmented polyurethane were identical with the PEO calibration standard i.e., no nitrogen signal could be detected at any level of amplification, and the expanded C Is scan corresponded to only carbon bonded to ether i.e., cf> = 1. Platelet retention indices were 0.05 or less. 

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