Microporous flow surface variation and short term thrombogenicity in dogs

Microporousflowsurfacevariationand short term thrombogenicityin dogs RodneyA White,Edwin Shops,RodrigoM. Miranda,,Stanley R Klein, I&e Goldberg,PhilipBosco,and RonaldJ. Nelson Department of Surgery, Los Angeles County Harbor/UCLA UCLA School of Medicine, Los Angeles, California, USA (Received 28 October 1981; revised 25 February 1982)

Medical

Center,

Torrance,

California

90509

and the

This study was designedto evaluatethe effect of three luminal surfacecoatingson short-termthrombogenicityin 4 mm internal diameter vascular prostheses. Microporous replamineform grafts (20-30 pm pore size) composed of silicone rubber were coated with medical grade biomaterials: Biolite @, TEDMACheparin, and [email protected] grafts were compared to each other and to control grafts of silicone rubber and Biomer that did not have coatings. Follovving three hours of implantation in the canine femoral artery, the prostheses were removed, opened longitudinally and evaluated for quantity of thrombus, % thrombus free surface, and type of thrombus. Silicone rubber grafts coated with Biolite carbon had the least thrombogenic Bow surface followed by the control Biomer grafts, Biomercoated silicone rubber grafts, TEDMAC-heparin coated silicone rubber grafts, and the control silicone rubber grafts. Due to the small number of samples, no statistical analysis was performed. Hence, the conclusions drawn are tentative.

Keywords:

Blood,

thrombogenicity,

Biolite@

carbon,

Biomer@, s/l/cone rubber,

Vascular surgeons rely on autogenous vein for repair of small internal diameter arterial defects because to date no prosthetic material has been developed which reliably retains long term patency. Many biomaterials scientists believe that for a small internal diameter vascular prostheses to function successfully it must have the non-thrombogenic characteristics of endothelial cells. Some of the variables implicated in the passivity of endothelial cells to the blood clotting mechanisms are surface charge’surface energy*, and degree of surface roughness3. Sawyer has asserted that the ideal material should have a double negative electrical layer similar to native vessels4. Baier and Abbott have postulated that a critical surface tension between 20-30 dynes/cm reduces thrombogenicity2. The porosity of the prosthesis5 and compliance6 of the graft segment have also been postulated to have effects on short and long term thrombogenicity but the exact role of these factors has not been determined. Sauvage has described a method for evaluating the thrombogenic potential of vascular prostheses by examining the blood-surface interaction following short term implantation7. In this study we used a modification of the Sauvage technique to evaluate, in viva, the effect of independent luminal surface variations on the short-term thrombogenicity of small internal diameter vascular prostheses having the same microporous wall configuration. Presented at the Thirteenth International Biomaterials Symposium, Rensselaer Polytechnic Institute, Troy, New York, May 28-31,198l.

0 1982

Butterworth

& Co (Publishers)

Ltd. 0142~9612/82/030145-05

MATERIALS

TEDMACheparin.

AND METHODS

Using the replamineform* technique, vascular prostheses were fabricated with a uniform interconnected 20-30pm porous wall*. Biomert and silicone rubbert replamineform vascular prostheses were implanted as controls and Figure 7 is a scanning electron photomicrograph of their luminal surface. Three surface coatings were applied to the luminal surface of silicone rubber grafts and evaluated for thrombogenicity. The coatings were: 1) ultralow temperature isotropic (ULTI) carbon (Biolite§),2) polyetherpolyurethane copolymer (Biomer), and 3) TEDMACheparinq. The ULTI carbon was applied by everting the compliant silicone rubber prostheses, depositing the Biolite coating, and then re-everting the prosthesis to place the coating on the luminal surface. The Biomer coating was applied by injecting a 15% solution of Biomer in dimethylacetamide through the silicone rubber lattice and then drying at 100°C for 18 h. The TEDMAC-heparin coating was applied by immersing the silicone rubber grafts in 2% TEDMAC-heparin solution and air drying. The TEDMACheparin solution caused significant swelling of the silicone *Replam Corporation, Torrance, California. f’Ethicon Corporation, Somerville, New Jersey. SSilastic, MDX4-4210, Dow Corning Corporation, Michigan. BCarbomedics, Inc., Austin, Texas. llPolysciences, Inc., Warrington, Pennsylvania.

Midland,

$03.00 Biomaterials

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Microporous

thrombogenicity:

R.A. White et al.

Figure 2 Scanning electron photomicrographs of the luminal surface of a replamineform silicone rubber prosthesis coated with TEDMACheparin (top) and a control silicone rubber prosthesis (bottom).

with thrombus deposition, and 3) type of thrombus (platelet, fibrin, or red blood cells). The quantity of thrombus was determined by inspection of the explanted grafts using a dissecting microscope and scanning electron microscope analysis. The percent of surface with thrombus deposition was determined using grid overlay.

RESULTS Figure 1 Scanning electron of the control replamineform (top), Biomer (bottom)

photomicrographs of the luminal surface vascular prostheses, Silicone rubber

rubber matrix which returned to its original configuration after the toluene evaporated. Figure 2 compares the control silicone rubber surface with the TEDMAC-heparin coated surface. All prostheses fabricated from silicone rubber were sterilized using a steam autoclave. Control Biomer prostheses were steam sterilized while immersed in saline as this delays degradation of the polyurethane matrix, although degradation would not be expected to be an important consideration in this short term implant studyg. After sterilization, the prostheses were pre-wetted by immersion in sterile saline under vacuum and agitation with ultrasound. Pre-wetting of ,;he prostheses removes entrapped air from the flow surface and improves thromboresistancelO. Fifty prostheses (4 mm internal diameter, 5-7 cm length) with 10 from each of the five groups of graft preparations were implanted bilaterally in the femoral arteries of mongrel dogs; two grafts with the same surface properties were implanted in each animal. Experimental animals were premeditated with xylazine hydrochloride and anaesthetized with chloralose. Following heparinization (1 mg/kg), the grafts were implanted as end-to-end arterial interpositions using interrupted 6-O polypropylene suture, The implants were removed after three hours of implantation and opened longitudinally to characterize the degree of thrombus deposition. Observation and description of the thrombotic response was performed by an independent observer at the completion of each experiment. Three parameters for each flow surface were determined: 1) quantity of thrombus (1 .O - no thrombus, 2.0 - limited thrombus, 3.0 - excessive thrombus, 4.0 - occluded), 2) % of surface

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The results of the study are displayed in Tab/e 7. The control silicone rubber surface had dispersed platelet and red cell aggregates and almost no thrombus free surface (Figure 3). In contrast, the silicone rubber grafts coated with Biolite had the least thrombogenic response of all the preparations with minimal thrombotic deposits and dispersed areas of thrombus free surface (Figure 4). Control Biomer prostheses also demonstrated a lowering in early thrombotic response as compared to the control silicone

Table 1.

Thrombogenicity Quantity of thrombust

of porous prosthetic % surface with thrombus

surfaces in dogs

l

Type of thrombus

deposition Platelet and fibrin aggregates: occasional red cell occlusion

Silicone rubber control

2.8

96

Silicone rubber with TEDMAC

2.7

100

Silicone rubber with Biomer

2.5

74

Red thrombus propegating from distal anastomosis; minimal platelets most of graft surface

Biomercontrol

2.1

80

Fibrin and platelet aggregates and thin red thrombus in areas

Silicone rubber with Biolite

1.5

75

Minimal, dispersed platelet aggregates

Platelet and fibrin aggregates with red cell thrombus

*Due to the small number of samples, no statistical analysis was performed. Hence, the conclusions drawn are tentative. ‘kalues are averages of number assigned to individual samples. (n = 10) Key: 1 .O - no thrombus, 2.0 - limited thrombus, 3.0 excessive thrombus, 4.0 -occluded.

Microporous

thrombogenicity:

In this study we have attempted independently

while

carbon,

temperature

TEDMAC-heparin,

copolymer

demonstrated

isotropic

Each of these materials

as a blood compatible

material

carbon-coated

silicone grafts had the least thrombogenic the graft preparations. have an acceptable

response of all

Low temperature

clinical

history

isotropic

In addition,

to experimental The Biolite

Dacron

coating

very flexible

luminal

rubber

surface coated with

response than the control was not significant. significant

effect

oared to control

prostheses which

Biomer

silicone

topography

had less thrombotic rubber

on the thrombotic

treatment

reaction

had no

of implanted

the effect

microporous

its resources to

the efficacy

prevention

of thrombus

process, we are able to study independently Studies

of the membrane

swelling of the silicone

selection

polymer

can be controlled

these two variables”.

by the appropriate

Similar

vascular grafts, however, than subcutaneous

to small diameter

The biological is considerably

implants.

Matching

graft patency 13, although ingrown

the compliance

of the internal

significant period.

of

more complicated graft wall compliance

Undoubtedly

are important

factors,

combut the

surface is, almost certainly,

paramount.

of fibrin

propagating

explanation

Biomer

grafts with a

in the

was primarily

and compliance

a

to the fact that was not identical, were essentially

5, the Biomer-coated

and had fewer surface pores. Results using a thin, solid silicone surface of grafts have shown

surfaces have less thrombus implant

Biomer

from the distal anastomosis.

coating on the luminal

in short term

Biomer

of the preparations

experiments

that smoother

rubber

and platelets

response may be attributed

surface was smoother

studies*‘.

for the disparate

There

findings

accumulation

is another between

the two

Biomer

surfaces: electron

spectroscopy

(ESCA)

studies of Biomer

surfaces revealed a greater concen-

tration

of soft segments (polyether)

than on the substrate Figure 4 Scanning electron micrograph of a representative area of the Now surface of silicone rubber prostheses coated with Biolite with minimal thrombus accumulation

;n the control

As shown in Figure

of preliminary

silicone

to the control

whereas the thrombus

the wall porosity

equivalent.

although

there were qualitative

rubber grafts coated with

although

during the implantation

comparable

composed

the surface topography

rubber

coated grafts had

of Biomer-coated

thin red cell component red cell thrombus

of

microscope

was not decreased.

The thrombus

was predominantly

when

there was no evidence

evidence that the TEDMAC

grafts, however,

This differential

transient

as a sustained drug release vehicle,

The performance

silicone

(two-fold)

hemorrhaging

grafts was quantitatively differences.

swelling

2). The TEDMAC

accumulation

replamineform

but the

Although

by scanning electron

This is circumstantial

thrombus

to the blood path of

rubber vascular grafts occurred

transmural

was functioning

in porous grafts that are decreases with time with

of the tissue response.

pliance and wall construction composition

function

(less than

in

is limited.

caused destructive

was applied,

disruption

analysis (Figure

responses have been

to be applicable

6 mm) vascular graftsI*.

of

on medical devices is as a surface coating

for local heparinization,

structure*O.

implants

Evidence

coating for

its application

of toluene

the TEDMAC-heparin

the maturation

formation

has been coupled

subcutaneous

improves

of TEDMAC-heparin

oxygenators

high concentration

Using the

on function.

TEDMAC-heparin

did not affect the

silicone surface.

It is used clinically

in dogs show that the type of tissue ingrowing

eventually

treatment

of the control

to support

membrane

to the biological

materials.

of pore size and biomaterial

demonstrated

of the prostheses were not affected

The TEDMAC-heparin thrombogenicity

has devoted

that contribute

coating as determined

analysis. Consequently

coating.

the Gott shunt but otherwise

the factors

replamineform

microscope

and is

change in the

The graft surface

by the Biolite

and compliance

not persuasive.

In recent years our laboratory function

characteristicslg.

is unaltered

by the Biolite

when com-

rubber surface.

DISCUSSION elucidating

porosity

vascular grafts with good results18.

is less than two microns thick

by scanning electron

but the difference

The TEDMAC-heparin

silicone

had the

vascular access

coatings have been applied

so that there is no significant

grafts’ mechanical rubber surface. Silicone

ULTI

coatings

in vascular surgery as a

coating for heart valves and percutaneous

Figure 3 Scanning electron micrograph of a representative area of flow surface of control silicone rubber prostheses with dispersed thrombotic deposits and almost no thrombus free surface.

has

suitable

devices14,15,16.

It is not surprising that the ULTI

devices14,17.

rubber

(ULTI)

and a polyether-poly-

(Biomer).

utility

for cardiovascular

The

we chose for coating the silicone

porous grafts were ultralow Biolite

(3 h) of three surface

keeping other variables constant.

surface preparations

urethane

to evaluate

the short term effects

preparations

R.A. White et al.

prepared

on the air-facing

analysis surface

surface22. When vascular grafts are

using the replamineform

process the blood inter-

facing surface is against the substrate, prepared

for chemical

using this technique

probably

Biomaterials

therefore,

prostheses

have a higher con-

1982,

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Julv

147

Microporous

thrombogenicity:

R.A. White et al.

ACKNOWLEDGEMENTS The authors would like to thank Bogaleth Gebre and Kenneth Johnson for their technical assistance. Replamineform vascular prostheses used in this study were prepared by Replam Corporation and were donated to the project. This work was supported in part by the Los Angeles County Affiliate of the American Heart Association Grant #641 G2-2 and the NHLBI grant #I5 ROl HL24618-02.

REFERENCES Figure 5 Scanning electron photomicrograph of the luminal surface of a silicone rubber prosthesis coated with Biomer following implantation demonstrating the smoother, yet porous surface

centration of hard segment (polyurethane) on the surface than the Biomer coated graft. The ultimate design of a small internal diameter vascular prosthesis will probably be defined by the site of implantation and its flow dynamics. The control replamineform silicone rubber grafts used in this study are too thrombogenic to attain high patency rates in dog femoral artery implants, yet they have proven to be adequate for sites with higher blood flows (i.e., aortic implants and arteriovenous fistulas). Longer term implant studies using the control replamineform Biomervascular prostheses have demonstrated improved thromboresistance and patency compared to the control silicone rubber prostheses although long term function is compromised by a slow, reactive degeneration of the implant matrix13. The silicone rubber prostheses coated with Biolite have also demonstrated a higher patency rate at two months in the canine femoral artery. This improved surface or others currently under investigation may provide the minimal thrombotic response which is required for function in small internal diameter, low flow applications. In this experiment, the main objective was to determine if there is a gross effect of surface treatment on thrombogenicity; many of the key parameters used to describe vascular surfaces were not determinedz3. Currently we are investigating the effect of surface charge, critical surface tension, and surface chemistry on the long term performance of small diameter vascular prostheses. We are also studying how these variables are affected by the physical properties of the prosthetic surface such as degree of roughness, pore size, and implant material. One limitation inherent in studies using one type of graft per animal is the variability in thrombogenic potential between dogs. In the majority of cases, patterns of thrombotic response were similar in bilateral grafts, although variability between dogs was frequently striking. Many in vivo and in vitro techniques for screening the thrombotic potential of graft surfaces have been developed, but often the correlation between these models and clinical results are poor. The marked variability in response between experimental animals, particularly dogs, accentuates this problem. An experimental design which uses vascular grafts with two surfaces in sequence has recently been described and it appears to alleviate the confusion created by the variability of responses among experimental animals*‘.

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Sharp, W.V.andTeague, PC., Pyrolytic carbon coated grafts, in Grafr Materials in Vascular Surgery: (Ed. Dardik), Chicago, Year Book Medical Publishers, 1978, pp.203-212 Haubold, A., Personal communication, Carbomedics, Inc., Austin, Texas. Flea, W.J., Whitley, D., Eberle, J.W.. Long-term membrane oxygenation without systemic heparinization, Trans. Amer. Sot. Artif. intern. Organs 1972, 18. 316-320 Goldberg, L., Bosco, P., Shors, E., Klein,S., Nelson, R.and

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