[ACS Symposium Series] Polymeric Materials and Artificial Organs Volume 256 || Artificial Cells

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  • 11 Artificial Cells

    THOMAS MING SWI CHANG Artificial Cells and Organs Research Centre, McGill University, Faculty of Medicine, 3655 Drummond St., Montreal, PQ, Canada H3G 1Y6

    Artificial ce l l s were first prepared in 1957. Since then, there has been increasing basic, applied and clinical research i n this area. At present, artificial ce l l s are being investigated clinically as blood substitutes; artificial kidney; detoxifier; artificial l i v e r ; drug carriers; immunosorbent; hemoperfusion and other areas. They are also being investigated for their appl i -cations in biotechnology in the areas of immobilized enzymes, biologicals , and cells.

    Since artificial ce l l s were first prepared by the author i n 1957 [1,2] an increasing number of approaches to their use are now available. Thus, artificial c e l l membranes can now be formed using a variety of synthetic or biological materials, resulting i n variations of permeability, surface properties and blood compati-bility. Almost any material can be included within artificial cells. These include enzyme systems, c e l l extracts, b iological cells, magnetic material, isotopes, antigens, antibodies, vaccines, hormones, adsorbents and others. A number of potential appl i -cations, suggested ear l ie r , have now reached the clinical trial or clinical application stage. Detailed reviews are available [4-7].

    RED BLOOD CELL SUBSTITUTE

    Since 1956 we have investigated the f eas ib i l i t y of using a r t i f i c i a l red blood ce l l s for use i n blood transfusion [1-3]. I n i t i a l l y , we prepared a r t i f i c i a l ce l l s i n the form of microencap-

    0097-6156/84/0256-0171S06.00/0 1984 American Chemical Society

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    In Polymeric Materials and Artificial Organs; Gebelein, C.; ACS Symposium Series; American Chemical Society: Washington, DC, 1984.

  • 172 POLYMERIC MATERIALS AND ARTIFICIAL ORGANS

    sulated hemoglobin. I n - v i t r o , these a r t i f i c i a l c e l l s d i d not i n t e r a c t with blood group antibodies, but they could transport oxygen and carbon dioxide. However, a f t e r i n f u s i o n , they were

    removed r a p i d l y from the c i r c u l a t i o n .

    We also prepared a r t i f i c i a l red blood c e l l s with an organic m a t e r i a l , s i l i c o n e rubber, as the major component [8,9]. S i l i c o n e rubber microspheres can transport oxygen. However, they were removed r a p i d l y from the c i r c u l a t i o n . Other groups were t e s t i n g a s i m i l a r material i n the form of s i l i c o n e o i l and fluorocarbon f l u i d fo r transporting of oxygen. Later, they prepared a f i n e emulsion of fluorocarbon o i l as a red blood c e l l s u b stitute [10]. By coating the surface with phospholipids to form an a r t i f i c i a l c e l l membrane, the fluorocarbon emulsion could become s t a b i i l i z e d and blood compatible. C l i n i c a l t r i a l s have been i n i t i a t e d i n Japan and U.S.A. However the long-term in-vivo e f f e c t s of fluorocarbon are not known.

    We e a r l i e r demonstrated that hemoglobin could be cross-linked by using a b i f u n c t i o n a l agent, forming a large polyhemoglobin. This permitted the a r t i f i c i a l c e l l s to be made much smaller than microencapsulated hemoglobin [2-4,9]. This has now been developed further so that hemoglobin can be cross-linked i n t o an even smaller polyhemoglobin which remains i n s o l u t i o n . Studies, being c a r r i e d out at t h i s Research Centre, have shown that the small polyhemoglobin survived much longer i n the c i r c u l a t i o n , as compared to stroma-free hemoglobin [11].

    THE ROLE OF ARTIFICIAL CELLS IN ARTIFICIAL ORGANS

    The rate of e q u i l i b r a t i o n i n 10 ml of 20 diameter a r t i f i c i a l c e l l s i s 400 times higher than f o r a standard hemodialysis machine [3,5,8]. Furthermore, membrane properties of a r t i f i c i a l c e l l s can be varied over a wide range to allow for changes i n permeability c h a r a c t e r i s t i c s and surface properties. The very small volume of a r t i f i c i a l c e l l s required, and the v a r i a t i o n s possible, r e s u l t i n d i f f e r e n t types of miniaturized a r t i f i c i a l organs. Both t h e o r e t i c a l analyses and animal studies have demonstrated the f e a s i b i l i t y of using the p r i n c i p l e of a r t i f i c i a l c e l l s to form extremely compact, e f f i c i e n t and simple a r t i f i c i a l organs. By varying the contents of the a r t i f i c i a l c e l l s (adsorbents, detoxicants, enzymes, c e l l e x t r a c t s , c e l l s and other b i o l o g i c a l l y a c t i v e m a t e r i a l s ) , the a r t i f i c i a l organs can be adjusted to carry out d i f f e r e n t biochemical or d e t o x i f i c a t i o n functions. The p r i n c i p l e of a r t i f i c i a l c e l l s has already been used to form blood compatible charcoal and r e s i n hemoperfusion systems for use i n p a t i e n t s , as a blood d e t o x i f i e r (poisoning), a r t i f i c i a l kidney, a r t i f i c i a l l i v e r , or immunosorbent. These w i l l be described below.

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    In Polymeric Materials and Artificial Organs; Gebelein, C.; ACS Symposium Series; American Chemical Society: Washington, DC, 1984.

  • 11. CHANG Artificial Cells 173

    ARTIFICIAL CELLS FOR ARTIFICIAL KIDNEY

    The large surface to volume r e l a t i o n s h i p and the u l t r a t h i n membrane allows rapid e q u i l i b r a t i o n of metabolites i n t o the a r t i f i c i a l c e l l s . By placing enzymes, ion exchange r e s i n and activated charcoal i n s i d e a r t i f i c i a l c e l l s , i t was demonstrated that the a r t i f i c i a l c e l l s could be used for hemoperfusion, a new form of a r t i f i c i a l kidney [8]. The p r i n c i p l e of a r t i f i c i a l c e l l s , con-t a i n i n g activated charcoal for hemoperfusion, was developed further to a stage for c l i n i c a l a p p l i c a t i o n [12,13]. In t r e a t i n g uremic p a t i e n t s , we have reported improvements i n nausea, vomiting, p r u r i t i s , f e e l i n g of well-being, and i n peripheral neuropathy [l4,15]. This approach i s much more e f f i c i e n t i n removing organic uremic waste metabolites than standard hemodialysis. However, i t does not remove water, e l e c t r o l y t e s or urea. As a r e s u l t , hemo-perfusion has been combined i n se r i e s with hemodialysis. This approach solves the problems of e l e c t r o l y t e , urea and f l u i d removal [16]. We i n i t i a l l y demonstrated that a 2-hour ACAC hemoperfusion-hemodialysis can replace 6 hours of standard hemodialysis. Furthermore, t h i s approach resulted i n improvements i n nerve conduction v e l o c i t y . This has stimulated i n t e r e s t i n t h i s combined approach and, i n the most recent large-scale studies c a r r i e d out i n I t a l y , the effectiveness of t h i s approach has been c l e a r l y demon-strated [17]. Another approach i s the combined use of hemoperfusion and a small (0.2 m2) u l t r a f i l t r a t o r , the l a t t e r f o r removal of water and sodium chloride [16]. In t h i s approach, no hemodialysis equipment i s required and the u l t r a f i l t r a t o r i s used only to remove and discard a l l the f i l t e r e d f l u i d (up to 2.7 l i t e r s ) , based e n t i r e l y on the hydrostatic pressure c o n t r o l l e d by the blood pump. A preliminary long-term c l i n i c a l evaluation has demonstrated the safety and effectiveness of t h i s approach [18]. The completion of t h i s system w i l l have to wait for the development of a urea removal system. Recently, a composite a r t i f i c i a l kidney has been formed, combining a r t i f i c i a l c e l l s and d i a l y s i s or a r t i f i c i a l c e l l s and an u l t r a f i l t r a t o r [19].

    ARTIFICIAL CELLS IN POISONING

    Hemoperfusion, using a r t i f i c i a l c e l l s containing activated charcoal, i s e f f e c t i v e i n t r e a t i n g severe acute drug poisoning f or those drugs which can be adsorbed, and which have a small volume d i s t r i b u t i o n [20]. High drug clearances have been obtained: glutethimide 230 ml/min, phnobarbital 228 ml/min, methyprylon 230 ml/min, methaqualone 230 ml/min, and secobarbital 200 ml/min. The effectiveness of hemoperfusion f o r those drugs with a high volume d i s t r i b u t i o n , e.g., t r i c y c l i c antidepressants and digoxin, i s s t i l l c o n t r o v e r s i a l .

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  • 174 POLYMERIC MATERIALS AND ARTIFICIAL ORGANS

    This approach has also been used i n the treatment of acute i n t o x i c a t i o n i n p e d i a t r i c patients [21]. The amberlite r e s i n hemoperfusion system has a high clearance for many drugs [22]. However, the problem of p l a t e l e t depletion was noted with t h i s system. More recently, our approach of using an albumin-coating to make the p a r t i c l e surface more blood compatible for hemoperfusion [12] has been applied to the coating of res i n s . As a r e s u l t , hemoperfusion with albumin-coated amberlite does not deplete p l a t e l e t s .

    ARTIFICIAL CELLS AS ARTIFICIAL LIVER

    Our i n i t i a l observation that hemoperfusion can r e s u l t i n the temporary recovery of consciousness i n grade IV hepatic coma patients [23] i s now conclusively supported by other centers [5-7]. However, i t s e f f e c t on long-term s u r v i v a l has not yet been estab-l i s h e d . A galactosamine-induced fulminant hepatic coma rat model has been used to study i n d e t a i l the e f f e c t s of charcoal hemo-perfusion on the s u r v i v a l rates of the animals [24-28]. ACAC hemoperfusion has been conclusively demonstrated to increase both s u r v i v a l time and s u r v i v a l rate of rats i n the earl y stages of hepatic coma. ACAC hemoperfusions c a r r i e d out i n the l a t e r stage of hepatic coma increased the s u r v i v a l time but not the s u r v i v a l rate. The same r e s u l t s were obtained through homologous l i v e r perfusion which increased the s u r v i v a l rate of rats i n the early stages of hepatic coma, but not those i n the l a t e r stage of coma. The present r e s u l t s would indi c a t e that hemoperfusion plays an important r o l e for the support of early stages of hepatic coma. In a l a t e r stage of coma, perhaps i r r e v e r s i b l e changes have already taken place. The r e s u l t s of experimental studies i n animals from t h i s laboratory [24-28] demonstrating the need for e a r l i e r treatment have now been corroborated by an i n i t i a l c l i n i c a l t r i a l [29]. Thus, larger scale c l i n i c a l t r i a l i s now ready to investigate t h i s a r t i f i c i a l l i v e r concept further, based on a r t i f i c i a l c e l l s , f o r the treatment of patients i n the e a r l i e r stages of fulminant hepatic f a i l u r e [30].

    We have also investigated the use of a r t i f i c i a l c e l l s con-t a i n i n g tyrosinase to carry out some metabolic functions of the l i v e r [31]. Results i n animals show that tyrosinase a r t i f i c i a l c e l l s , retained i n extracorporeal shunts perfused by blood, can e f f e c t i v e l y lower the systemic blood tyrosine l e v e l s of l i v e r f a i l u r e r a t s . A r t i f i c i a l c e l l s , to carry out other metabolic functions of the l i v e r , are also being studied. For instance, using a r t i f i c i a l c e l l s containing multienzyme systems with cofactor r e c y c l i n g [32], we have studied the i n - v i t r o conversion of ammonia sequentially i n t o d i f f e r e n t types of amino acids. This way, ammonia

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  • 11. CHANG Artificial Cells 175

    has been converted into glutamate and then sequentially i n t o alanine or other amino acids.

    ARTIFICIAL CELLS AND IMMUNOSORBENT

    The ACAC hemoperfusion system developed by the author consists of albumin-collodion coated charcoal [12]. Albumin makes the surface blood compatible and also takes part i n i n t e r a c t i n g with material i n the c i r c u l a t i n g blood, including the f a c i l i t a t e d transport of loosely protein-bound substances [5]. Terman reported the i n t e r e s t i n g f i n d i n g that these ACAC microcapsules can also be used to remove antibodies to albumin from dog plasma [33]. He proceeded further to incorporate other types of antigens or antibodies onto the surface of t h i s ACAC a r t i f i c i a l c e l l system, r e s u l t i n g i n immunosorbents fo r d i f f e r e n t types of a p p l i c a t i o n s . In very preliminary studies he found that, by replacing the albumin of the ACAC system with protein A for use i n plasma perfusion, he was able to s i g n i f i c a n t l y reduce the s i z e of breast carcinomas i n patients.

    Another area involves the use of synthetic immunosorbents. In order to prevent the problem of p a r t i c u l a t e release and blood i n c o m p a t i b i l i t y , we coated synthetic immunosorbents for blood group A & with albumin and c o l l o d i o n . I t was demonstrated that the synthetic immunosorbent could s t i l l remove the anti-A and anti-B blood group but did not a f f e c t p l a t e l e t s or release p a r t i c u l a t e s [33]. An albumin-coated system has since been used c l i n i c a l l y by another centre to remove anti-A and anti-B from plasma of patients p r i o r to bone marrow transplantation.

    ARTIFICIAL CELLS CONTAINING ENZYME SYSTEM

    The i n j e c t i o n of free enzyme of heterogenous o r i g i n may r e s u l t i n h y p e r s e n s i t i v i t y reactions, production of antibodies, and rapid removal and i n a c t i v a t i o n . Furthermore, free enzymes cannot be kept at the desired s i t e s of action and are l e s s stable at a body temperature of 37C. The use of a r t i f i c i a l c e l l s containing enzymes and proteins has been investigated [3,4]. Some examples of research being c a r r i e d out w i l l be b r i e f l y mentioned.

    The f i r s t demonstration of the use of a r t i f i c i a l c e l l...

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