November 1980 Are Hybrid Artificial Organs Necessary?

Hybrid Artificial Organs: Are They Really Necessary? Yukihiko Nosd, Paul S. Malchesky, and James W. Smith

Received: October 1980

ABSTRACT In order to reproduce completely the function of an organ, past research has considered hybrid artificial organs as the only method to achieve this goal. Various attempts have been made to use cell components, wholecells, tissue, and organ sections in conjunction with artificial organ systems. However, utilization of biological preparations or tissues is not clinically practical today. In this paper disadvantages of using natural tissue preparations are outlined. Recent progress in membrane technology such as membrane plasmapheresis, particularly on-line plasma treatment by multiple reactors, on-line plasma treatment by cryogelation, or the use of a cascade membrane scheme for the selective removal of macromolecules, will provide the tools to treat and remove macromolecules which were once considered extremely difficult to treat. The historical development of membrane plasmapheresis is described, including potential applications of this type of artificial organ for practical clinical treatment of metabolic, toxic and immunologic disease states.

Key words: hybrid artificial organs, liver slices, freeze- dried liver granules, membrane plasmapheresis, on-line plasma treatment, multiple reactors, double-membrane filters, cryogelation, cascade membrane technology

INTRODUCTION Reproduction of the complicated functions of the body is almost impossible by entirely artificial means. Simple artificial organs such as a cardiac prosthesis, which is merely a pump, or a n artificial kidney, which is merely a dialyzer adjusting electro- lytes, removing small molecules, and providing water balance, are simple mechanical substitutes for these natural organs. While simple in design, these systems are efficient as witnessed, for example, by the dialyzer’s clinical application and widespread use. It should be remembered, however, tha t the artificial organ does not reproduce all functions of the organ and therefore h a s limitations for long-term use. The reproduction of all functions of an organ system is too complex at the present to be accom- plished. This is, in part, due to the lack of complete information on how the natural organ system operates in the normal and disease states. The functions of the complex organ systems such as the

From the Department o f Artificial Organs, the Cleveland Clinic Foundation. Cleveland, Ohio 44’106, U.S .A .

liver a n d pancreas, which perform the sophisticated adjustment of various chemicals including not only small molecules but also macromolecules and protein- bound materials, are almost impossible to simulate by artificial means. I n the case of the liver the duplication of i ts function is extremely difficult since much liver function is still unknown to the most knowledgeable gastroenterologist or hepatophysiolo- gist. The concept of hybrid artificial organs, utilizing some form of biological material combined with non- biological material to make a substitution organ, is the only possible alternative to make complex replacement devices.

HYBRID HEPATIC ASSIST DEVICE: A BRIEF HISTORICAL REVIEW

I n the 1950s efforts were made to simulate the function of the liver by a hybrid artificial liver. To study the function of the liver, it is most ideal to use healthy, living subjects. Co-perfusion was a n initial attempt to achieve this goal. I n 1957 Hori and Kimoto a t the University of Tokyo utilized co-perfusion of the patient with healthy dogs for the treatment of hepatic coma.’ For their studies, four dogs were used. The blood of the healthy dogs was introduced into a four-coil apparatus of semipermeable membranes (Fig. 1). I n each cell another coil was also incorporated and into this coil the patient’s blood was introduced. Any excess metabolites in the patient’s blood would be picked up by the healthy dog’s blood and be metabolized. Any deficiency of metabolites in the patient’s blood would also be adjusted by the dogs. I n general, about two liters of metabolic fluid, containing various amino acids, vitamins, and ATP to stimulate the TCA cycle, was used. Ion-exchange resins were also incorporated into this system. The University of Tokyo group called this device a “biological artificial liver.” Limited clinical studies were performed and the results were reported.2 However, the use of living animals and their connection to the patient proved somewhat cumbersome and highly impractical.

I n 1959, Mikami, Mito a n d Nos6 introduced a device utilizing a liver tissue preparation3 (Fig. 2a). Originally liver tissue homogenate was utilized based upon the very brief report of S ~ r r e n t i n o . ~

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FIG. 1. Kimoto’s, Sugiura’s, and Hori’s biological artificial liver* (1959). Four healthy dogs were used and a pump recirculated the metabolic fluid past separate semipermeable membrane coils con- taining patient’s blood and healthy dog’s blood.

However, it was extremely difficult to achieve urea synthesis from this tissue preparation, in agreement with Krebs, who had found synthesis of urea impossible to achieve by liver tissue homogenate. Based upon these biochemical studies from many years ago, utilization of fresh liver slices incorporated into a hybrid type of device was introduced.’ Unfortunately, it was extremely difficult to produce a device incorporating a few hundred grams of liver tissue when thickness of 0.2-0.4 mm was needed for tissue viability. I t could be done, but i t was not practical.

A more feasible utilization of liver tissue is tissue culture or liver tissue preparations having longer shelf life, such a s enzyme preparations or freeze- dried liver. Because of difficulties with the use of tissue culture cells a t tha t time, the decision was made to use freeze-dried liver tissue preparations. After this form of processing, enzyme activity was reduced quite substantially. However, it was found tha t certain liver functions were maintained for a t least two hours.’ Utilizing this type of liver pre- paration, the hybrid artificial liver was used for a limited number of patient treatments (Fig. 2b).6 Overall effectiveness is still unknown, but with such experience, the hybrid artificial organ and its applications and limitations was conceived.

THE HYBRID ARTIFICIAL ORGAN: ITS LIMITATIONS

Among limitations involved in the use of hybrid artificial organs is the necessity of using tissue or cells with their attendant inherent immunological problems. In order to prevent immunological problems, permeable membranes with porosity allowing passage

FIG. 2. a) Mikami’s, Mito’s and NosB’s device’ ( ;959). Metabolic fluid was recirculated in the chamber containing gel-type cellulose membrane which was connected to liver insufficiency subject. The metabolic circuit included a bubble oxygenatcr m d a chamber containing fresh liver slices. b) Mikami’s and NoEf ’ 5 ; devicesv6 (1960). Metabolic fluid (5-10 liters) contained freeze-dii<?d canine liver granules. For experimental studies, gel-type cellulose membrane was used; however, standard cellulose membranz for hemodialysis was utilized for the clinical application.

of albumin but not globulin were needed. It was very difficult to find ideal membranes for t h k purpose, so most of the membranes used for this application had porosities in the range of less than I00 A, which allowed only a small percentage of albumin to pass. With limited sievingof albumin, any of the metabolites or toxins bound to protein will not be effectively contacted by the biologic material. For the initial studies conducted by Nos6 et al. in 1960, gel-type cellulose membrane of 40-50 A was used. This membrane was considered to be two to three times more permeable than the standard hemodialysis membrane. Efforts were made to utilize more porous cellulosic membranes, including Japanese papers,’ but the technology of 1960 did not provide the ideal membrane for this application. I t was considered a t that time that if homologous or possibly autologous tissues were used, this immunological barrier would not need to be incorporated if the treatment was limited to end-stage liver failure. However, in general, a method to establish this immunological barrier by mechanical means was and remains the

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major limiting factor for effective hybrid artificial organs.

The best method to obtain biological tissue from other living subjects and maintain its activity is by tissue culture. However, during maintenance in tissue culture, the metabolic cycle of the cell may change so t h a t the original hepatocyte functions may not be the same. The hepatoma cell grows well in tissue culture preparations, but we are not sure if this cell has completely normal metabolic capabilities. In addition, some toxic or carcinogenic factors might be released from this preparation.

If we use a natural tissue preparation such a s fresh slices, shelf life problems exist. Frozen sliced tissues will have longer shelf life and further extension is possible by freeze-drying liver slices or liver granules; however, substantial enzyme activity is lost during this process. Tissue condition is suboptimal due to processing, and abnormal metabolic cycles may be activated. In addition, widespread application of hybrid artificial organs requires readily available off-the-shelf preparations under a range of varying environmental conditions. Storage should be possible without complex preparations. Because cells and tissues are used, contamination with various toxic and pathogenic factors must be avoided in ibis type of tedious preparation, and the need for a preservation period is difficult to avoid. Complex maintenance and stringent environmental controls require extremely expensive methods of preparation and storage. Thus, even if a hybrid artificial organ is made available at a major health center, i t will be a t a substantial cost. If this is so, this type of approach would be very difficult for widespread use in the future.

Tissue culture technology h a s advanced recently and devices for tissue culture or preservation are improving, leading to a rebirth of hybrid artificial organs. At the same time we have also made considerable progress in membrane and artificial organ technology. Thus we must carefully consider the relative strengths and weaknesses of various devices to determine which will fulfill existing needs most practically. We should have a step-by-step reproduction of organ functions in a rather simple way so tha t we know what we are doing. The artificial organ may not be perfect, but a t least certain parts of the physiological functions of the various organs can be artificially reproduced.

After total hepatectomy, a n animal will go through various pathophysiological states before succumbing to hepatic coma. Initially, the animal will have liypoglycemic coma. At this time if we introduce the proper level of glucose, which can be done by mechanical means, the animal will live

longer. Then, various electrolyte imbalances will occur. If we adjust electrolytes during this period, the animal will live longer. After a certain period of time, blood coagulation abnormalities will take place. Adjustment of those abnormalities promotes longer survival. Then, various toxic factors such as ammonia increase in the blood. If these toxins areremoved, the animal lives somewhat longer still. By this step-by- step approach to replace the known function of the liver, we can keep anhepatic animals alive for one to two days instead of a few hours.

In the past, our technology was such that only small molecules could be processed. Recently, large porosity membranes have become available and the adjustment of middle-size molecules is possible utilizing recently developed hem~dia lyzers .~ However, these devices are not designed for permeability of proteins and large molecules.

MACROMOLECULAR TREATMENT BY MEMBRANE DEVICE

By 1972, development of membrane technology had been such that membranes with a porosity of 0.2 to 0.5 pm were available. Utilizing these more porous membranes and using homologous fresh liver slices, our group rejuvenated the experimental hybrid hepatic assist device concept with membrane plas- mapheresis and reported on i t initially in 19749 and

This was the first experimental attempt of on-line membrane plasmapheresis for therapeutic application (Fig. 3). These higher porosity membranes were shown to be permeable to protein-bound or macromolecular toxins as these occur in hepatic failure. While the immunological problems with this methodology may not be a severe restriction for end- stage liver insufficiency patients, various technical problems associated with handling liver tissue preparations and the need to find a practical, useful, clinical therapy caused u s to concentrate on the nonbiological reactor systems.”’ l 4 In 1974, Williams’ group also attempted centrifugal plasmapheresis and its combination with a charcoal cartridge.15 The initial plate-type device required operating conditions within a narrow range of shear rate and trans- membrane pressure, did not operate efficiently, and was difficult to fabricate.12 For this reason, with the availability of hollow-fiber devices, efforts were concentrated in this area.

Hollow-fiber devices using membranes of 0.2 to 0.5 pm pore size were developed into clinically acceptable designs. The Monsanto porous, hollow- fiber oxygenator device was used during 1972 and proved not applicable for plasmapheresis. Since 1976, the Asahi hollow-fiber devices for ascites treatment have been evaluated for their applicability

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methods described in Figures 4 through 6 or combinations of the methods described in Figures 4 and 5.

It is our opinion tha t at this time we should not overemphasize rejuvenating complex hybrid artificial organs tha t were attempted many years ago. This is not to say tha t we should not work in this area; the developments for tomorrow must be the research of today. But for practical applications at present, the non-hybrid artificial organ should be stressed. With the background in hybrid artificial organs research spanning more t h a n 20 years, the shortcomings of these devices as tools for safe, easy-to-use, and reasonably priced therapeutic methods are quite obvious. We should rather concentrate on the effort to have more practical, non-hybrid artificial organs for metabolic assist devices.

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