artificial and bioartificial replacement of the endocrine pancreas

Artifit id Organ.\ 16(1):61-70. Blackwell Scientific Publications. Inc., Boston 0 1992 International Society for Artificial Organs

Artificial and Bioartificial Replacement of the Endocrine Pancreas

Gkrard Reach

Biomedical Engineering and Diabetes Mellitus Research Unit, Diabetes Department, Hbtel-Dieu Hospital, Paris, France

If somebody says that some task is mechanical, it does not mean that people are incapable of doing the task, it implies that only a machine could do it over and over without ever complaining or feeling bored.

Douglas Hofstadter

DIABETES MELLITUS: THE NEED FOR ARTIFICIAL ORGANS

Type 1 , insulin-dependent, diabetes mellitus is due to the auto-immune destruction of the islets of Langerhans, which, in the pancreas, secrete insulin. lnrulin secretion by the healthy pancreas can be considered as a unique system able to permit the assimilation of nutriments during meals and, as a consequence, to achieve the homeostatic control of blood glucose concentration. Indeed, each meal triggers a peak in insulin secretion. By turning off the post-hepatic production of glucose and by increasing peripheral glucose utilization, this insulin burst prevents the occurrence of hyperglycemia following carbohydrate intake. Blood glucose concentration will increase only slightly, and then, because of the hypoglycemic effect of insulin, return to its basal level. This will turn off the stimulation of insulin secretion, avoiding thereby the occurrence of an overshoot hypoglycemia. This regulation of insulin secretion during meals is due to a direct regulatory effect of glucose on insulin secretion, which can be demonstrated in vitro on isolated islets of Langerhans, and to the fine tuning of this regulation by other nutriments (such as amino acids and free fatty acids) as well as by endocrine and neuronal afferences. It is impressive to note that this homeo-

Received September 1991. Address correspondence and reprint requests to Dr. G . Reach,

INSERM U341, Diabetes Department, Hbtel-Dieu Hospital, 1, Place du Parvis Notre Dame, 75004 Paris, France.

static system is so powerful that in normal subjects, plasma glucose concentration is kept within a very narrow range, i.e., rarely lower than 70 mgidl or higher than 120 mgidl(3.9-6.7 mmoliL), despite the discontinuous intake of carbohydrates.

In patients presenting with diabetes, the pancreatic insulin secreting beta cells have been destroyed. In the absence of insulin replacement, diabetes mellitus is a rapidly fatal disease, as the assimilation of nutriments is no longer possible. Since the discovery of insulin and its first use as a treatment of diabetes in 1922, long-term survival of diabetic patients has become possible. Thus, unlike other major organ failures (such as end-stage car- diac or liver failure), diabetes is not an immediately life-threatening disease since it can be treated with exogenous insulin. Certainly, this point must be taken into account when considering novel approaches to the cure of the disease.

However, diabetes mellitus must be considered as a chronic disease, leading potentially to severe, sometimes fatal, complications. The late complications of diabetes (blindness, end-stage renal failure, lower limb amputations, etc.) bear three characteristics: late onset, uncertainty, and a point of no return. If one considers, for instance, end-stage renal failure, which is a common complication of diabetes mellitus, the proportion of new patients presenting with diabetic nephropathy accepted for renal replacement therapy in Europe between 1976 and 1985 was found to be 10% (1). However, it was found that only 30% of diabetic patients would ever develop this complication. As there is currently no reliable marker to predict which patient will develop such a complication, this suggests that the primary prevention of these complications should be implemented very early in the course of the disease and in all diabetic patients. It is now generally

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accepted that a tight control of glycemia is the prin- cipal mode of intervention by which diabetic complications can be prevented (2). Indeed, there is considerable evidence that the quality of diabetes control is inversely correlated to the occurrence of microangiopathy and neuropathy . Unfortunately, the discontinuous administration of insulin is to this day unable to reproduce the infinitely subtle pattern of insulin secretion provided by the natural pancreas, and thus only crude correction of hyperglycemia can be obtained, with frequent overexcur- sions of blood glucose concentration either below the normal limits (hypoglycemia) or above (hyperglycemia). In order to improve the quality of diabetes control, patients must continuously adapt the insulin dosages according to the results of self-monitoring of blood glucose levels and to the presence or absence of glucose in urine samplings. They are taught how to determine their capillary blood glucose levels several times a day using a fingerprick system and glucose-oxidase reactive strips, to re- cord the data in a log, and to decide the next insulin dosage after a somewhat subtle debate (3). Not only is this system not completely efficient (for instance, severe hypoglycemia, leading to unconsciousness, cannot be always avoided, and the complete correction of hyperglycemia is rarely possible), but, in addition, it represents a task that must be accom- plished every day throughout life, which is dreadfully boring. It becomes obvious that “only a machine could do it over and over without ever complaining or feeling bored.”

A machine? More precisely, an artificial organ: The European Society for Biomaterials defined an artificial organ as “a medical device [isn’t that a machine?] that replaces, in part or in whole, the function of one of the organs of the body.” (4) Thus, the first task in designing an artificial organ is to analyze the function of the natural organ. More specifically, this will yield a reductionist view of this function. For instance, the function of the kidney can be assimilated with its aptitude to promote exchanges of solutes across a semipermeable, fil- trating membrane, and this is what the artificial kidney is supposed to do because it is the replacement of this main, vital, function of the kidney that makes patient survival possible in an otherwise fatal disease. Other important functions of the kidney, such as erythropoietin secretion, are either totally ig- nored, or substituted for by other means (i.e., trans- fusion, or more recently the administration of re- combinant erythropoietin).

The function in need of replacement in the case of insulin dependent diabetes is the insulin secretion

by pancreatic beta cells, which can be described by four characteristics: (a) continuous, even in postab- sorptive state, with rapid and transient peaks during meals; (b) automatic regulation by blood glucose levels; (c) delivery into the portal blood system; and (d) the endocrine pancreas is, of course, an internal organ placed within the body. It is important at this point to stress that none of these four characteristics are displayed by the current, discontinuous, administration of insulin by means of syringes, and that any device which can proclaim having one or several of these items could be considered as an artificial pancreas even if incomplete. When decid- ing which function of the pancreas is important, it is important to bear in mind the objectives of research toward an improvement of the treatment of diabetes mellitus (mainly, an improvement in the quality of diabetes control, and in the patient’s quality of life) since as mentioned previously, diabetes can already be treated with insulin. It is therefore from the diabetological point of view that options must be made, and this explains why diabetologists well aware by routine contact with patients of both the possibilities and the imperfections of diabetes treatment as it is currently available must be involved in the teams developing artificial pancreases where indeed an interdisciplinary approach is required. If engineers alone, without early guidance by physi- cians, took the problem in charge, it would not be surprising that the final result was a machine whose clinical applications appear impossible or useless, its long and expensive development ending up in a blind alley, without any chance of entering into the real world of medical technology.

One must, on the other hand, note that what a patient with a chronic disease is waiting for is not just an artificial organ. The artificial organ should be implantable as only this would permit the patient to consider himself cured. According to the same source (4), an implant is “a medical device made of one or more biomaterials that is intentionally placed inside the body, either totally or partially buried beneath an epithelial surface.” Is it possible to design a machine that would gather all these characteristics and be both functional and implantable? This will be the main issue addressed during this review, and it will serve as its guideline.

ARTIFICIAL ORGANS: FUNCTIONALITY VERSUS IMPLANTABILITY

The development of an implantable artificial organ is often based on the hope that a system initially designed for in vitro application can be used in vivo

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(artificial organ) and that a system first designed for short-term use can be implanted and function on a long-term basis (implantable artificial organ). For instance, to take again the example of the artificial kidney, dialysis is basically an in vitro concept (if a semipermeable membrane separates two media with different compositions, exchanges across the membrane will produce an equilibrium state at which the two media will have a similar composi- tion). The genius of pioneers as W. Kolff was to take this concept and make an in vivo application of it. This was the beginning of the discontinuous therapy of end-stage renal failure by an artificial kidney. The application of this method was at first restricted to acute anuria and later extended to chronic end- stage renal failure, either as a bridge to kidney transplantation or as a long-term therapy. How- ever, it must be considered that nowadays patients are treated with an artificial, disposable, kidney, and that they are connected several times a week to an external device. So far, the implantable artificial kidney remains a dream of the future. It can be observed that with the clinical possibilities offered by the aritificial kidney, end-stage renal failure still represents a distressing condition but is compatible with prolonged patient survival. In addition, it can be often considered as a bridge to kidney transplantation. This may explain in part the quasi-absence of progress toward the development of an implantable artificial kidney, which would necessitate the overcoming of such formidable obstacles as the permanent access to blood vessels and membrane long- term biocompatibility. As shown later on, it has been possible to design artificial pancreatic systems that work perfectly in vitro and even that function in vivo over a short period of time. The problem for diabetologists is that, as before mentioned, it is not sufficient to have a system that works for a short period of time; it is a long-term implantable artificial pancreas that is needed. An external system might even be considered by a number of patients as useless. The challenge is therefore much more difficult.

Another major difference must be stressed. It has been recognized by nephrologists, after 20 years of extensive use of artificial kidneys, that the biocompatibility issue must be considered in two different aspects. First, we must consider the artificial organ itself. Obviously, it is important that the function of the artificial kidney should not be impaired by biocompatibility reactions. For instance, as long as clotting of a limited number of hollow fibers does not alter the exchange properties of the device, they can be tolerated over the short-term period (a few hours) of a dialysis session, within the concept of

discontinuous dialysis using disposable artificial kidneys. This problem is presently resolved with the currently available artificial kidneys, generally with the help of heparin. Second, the biocompatibility issue must also be considered with respect to the patient himself It has been recently realized that some of the late complications of end-stage kidney failure replacement therapy are most likely due to the repeated contact of blood with artificial sur- faces, which elicit activation of adhering mono- cytes, producing monokines such as interleukin- 1, which might be responsible for the occurrence of amyloidosis in these patients. It is also possible that monocyte activation is triggered by bacterial lipopo- lysaccharides contained in the dialysate, crossing the membrane by back-filtration. For nephrologists, therefore, the biocompatibility issue is essentially a long-term problem threatening the health of the patient more than the function of the artificial organ itself. These two aspects of biocompatibility can therefore be considered separately because the artificial organ is not supposed to function for more than approximately 4 h. By contrast, as will be demonstrated in this review, an interesting feature of the development of an artificial pancreas is that the functionality and biocompatibility issues cannot be treated independently. As will be discussed in detail in the next section, biocompatibility reactions can rapidly alter the function of the system, and in turn, the design of the system will be determined at a great extent by considerations on its expected lifespan.

DIFFERENT APPROACHES TOWARD THE ARTIFICIAL REPLACEMENT OF THE

ENDOCRINE PANCREAS

There is but one way to do nothing and diverse ways to do something.

Ambrose Bierce

The artificial approach

Wearable and implantable pumps We have previously indicated that the first fea-

ture of insulin secretion is that unlike insulin administration by means of syringes, it is continuous with peaks occurring during meals. To mimic this property, external pumps have been designed in the late 1970s and early 1980s. They are battery driven and externally wearable; usually, insulin is delivered into the subcutaneous tissue by means of a needle, which has to be replaced every 2 days. The pump can provide the patient with a continuous basal in-

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sulin delivery rate that can be increased transiently immediately before a meal.

Interestingly, after a period of over-enthusiasm, it was recognized that only a small minority of patients (less than l%) accepted treatment on a long- term basis with such a system. A major (but not complete) explanation for this relative failure is that the system is not implantable. The patient will tell us: “When I have my syringes, I think of my disease three times a day. With the pump, I think of diabetes the whole day long, and even during the night!” This points out the fact that the fourth char- acteristic of pancreatic cells, as delineated previously (the fact that they represent an internal organ), cannot be overlooked and explains the major effort towards the development of implantable insulin delivery pumps, which started at the end of the 1970s and which has recently reached the stage of large scale clinical investigation (5) .

Most of the implantable pumps currently function along the same principle: A chamber is filled with a gas that expands at body temperature and pushes a membrane separating the gas chamber from an insulin-containing reservoir. Insulin is thus pushed through a small orifice connected to a catheter. A valve system, electronically controlled, makes it possible to control the flow rate of the liquid. This flow rate can be increased contemporaneously to meals by a remote telemetrical controller. Thus, this system displays all the features of the external pumps, and, in addition, bares the important attrib- ute of becoming an internal artificial organ. The patient no longer has to inject himself several times a day with insulin since the insulin reservoir of the pump only needs refilling once a month. In addition, an interesting feature of implantable pumps is that it becomes easier to deliver insulin either directly into a central vein (which mimicks the blood delivery of insulin by the natural pancreas), or more often, in recent years, into the peritoneal cavity. It has been convincingly demonstrated that part of the delivered insulin reaches the bloodstream through the portal vein (6), which was the third feature of insulin secretion delineated previously. This can be an advantage since it theoretically establishes a porto- sushepatic gradient in insulin concentration, which might avoid peripheral hyperinsulinemia. Hyperin- sulinemia is a subject of concern for diabetologists since it is believed to be involved in the development of diabetic macroangiopathy (7).

So far, implantable pumps have been shown to function satisfactorily for prolonged periods of time with a half-survival rate of 36 months (this means that 36 months following implantation, half of the

pumps have been explanted) (8). This is somewhat shorter than what should be expected from the battery lifespan, which is usually 5 years. The major causes for explantation were incidents located on the catheter, obstructed either by a blood clot, in the case of venous delivery, or by a fibrotic foreign body reaction, in the case of peritoneal implantation. Here, we have the first example of an interac- tion between a biocompatibility-related event and the system’s function since, obviously, if the catheter is obstructed, the whole system cannot work anymore (i.e., deliver insulin) although the function of the pump itself is still intact. Other examples follow *

Automatic rcJgulation of insulin delivery with a glucose sensor

Actually, the most important feature of insulin secretion is its automatic regulation by the blood glucose level. A true artificial beta cell should therefore incorporate a feedback system that would mon- itor blood glucose levels continuously and would control, on the basis of this monitoring, the rate of insulin delivery (9). Chemical systems are being developed. For instance, glycosylated insulin can bind to osidic receptors of a lectin, concanavalin A , and this binding can be displaced by glucose, thus re- leasing insulin according to the ambient glucose concentration (10). These chemical forms of an artificial beta cell are currently investigated at the bench level.

The electromechanical approach consists in the design of a glucose electrode, namely a system able to generate an electric current proportionally to the environing glucose concentration. Such a current would be processed by a computer that controls the flow rate of the pump delivering insulin. It is important to stress that the successful development of a glucose sensor can be used not only as a part of such a closed-loop insulin delivery system, but also, maybe before the achievement of this dream, as a part of a system for continuous glucose monitoring (reproducing, by the way, another feature of the pancreatic beta cell!) and allowing permanent access by the diabetic patient to his blood glucose concentration, or even as a part of an alarm for detecting hypoglycemia, the latter system being highly valuable for a number of diabetic patients.

A biosensor consists of two parts, a specific recognition element, which can be an enzyme, an anti- body, or a receptor, and a transduction system, which transforms the energy produced during the binding between the analyte and the recognizing element into a signal that can be processed by elec-

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tronic means, i.e., an electric current. The trans- ducer can use different principles of physics (electrochemical, enthalpic, optic). Most of the glucose electrodes are based on the electrochemical, amper- ometric, detection of the reaction between glucose and glucose-oxidase, and we will focus on this type of biosensor. The first successful glucose sensor was based on the oxidation of glucose by glucose- oxidase ( I 1). This consumes oxygen, which can be monitored by an oxygen electrode. On the other hand, the reaction generates gluconic acid and hydrogen peroxide. The latter, in the presence of a 650 mV potential, is in turn destroyed, producing elec- trons; in this case, the biosensor is a hydrogen peroxide electrode. It is possible to build a system that produces such an electric current proportionally to the glucose concentration over a range between 0 and 360 mg/dl(20 mmol/L), i.e., the range of interest for diabetologists.

This system was first used under in vitro conditions for glucose determinations. It was then ap- plied in vivo when this type of glucose sensor was incorporated inside a complex system known as an artificial pancreas, the first to be released being the Biostator in the middle 1970s. The glucose sensor consisted of a flow-through chamber in which blood, continuously drawn from the patient, was allowed to circulate in contact with a membrane. On the other side, glucose-oxidase was present in an immobilized form, separated from a platinum/ silver-silver chloride electrode by another membrane, which prevented access to the electrode of electroactive substances such as ascorbate. The current formed during the oxidation of glucose was analyzed by a computer, which controlled the flow rate of an insulin-delivery pump.

With such a system, it was possible to control perfectly the blood glucose level of diabetic patients, but only for short periods of time since the patient needed to be connected to the device, which was the size of a television set, which required an access to a venous blood vessel, and which pumped approximately 50 ml of blood per day. No wonder, therefore, that such a system could not be used for the daily treatment of diabetes and has been used mainly as a superb tool for clinical investigation. We could consider that these systems are at the same stage of development toward an artificial internal organ as are the currently available artificial kidneys. The difference is that the treatment of end- stage renal failure is compatible with discontinuous dialysis sessions, 4 h 3 times a week whereas diabetes treatment requires the continuous control of blood glucose, 7 days a week, 24 h a day.

Another difference is that even within a few hours of use, the membrane separating blood from glucose-oxidase becomes rapidly fouled with pro- tein and blood cells, thus altering the exchange properties of this membrane. Frequent recalibration of the sensor is required. We have seen that clotting of a few fibers of an artificial kidney does not hinder the function of the whole device simply because the surface of the membrane is so great. In the case of the glucose sensor’s membrane (a few square centi- meters), the fouling of a minute membrane area can be detrimental. We find here a second example in which the biocompatibility of one of the compo- nents of the system is the major determinant of the lifespan of the whole system.

These obstacles have prompted some investiga- tors, such as M. Shichiri, to propose another approach, in which a needle-type glucose sensor would be implanted in the subcutaneous tissue. Glu- cose concentrations would therefore be sensed in the interstitial fluid, and no longer in the blood. The sensor consisted of a platinum wire (the anode of the eletrode) placed inside a steel needle (the cathode). Glucose-oxidase was layered on the tip of the platinum wire, melted to form a sphere, and the electrode was covered with polyurethane to make it biocompatible (12). Sensors, measuring glucose in vitro with a short response time (shorter than 5 min), on a wide linearity range (up to 20 mmol/L), with an acceptable signalhoise ratio, can be readily prepared; the sensor works in vitro. Is the application to in vivo use possible?

First, it was possible to demonstrate that the glucose concentration in subcutaneous fluid is really identical to that in blood. For instance, U. Fischer implanted in the subcutaneous tissue of dogs a wick made of cotton threads. One hour later, the wick was removed, centrifuged, and the glucose concentration was determined in the drop of fluid collected from it. It was found to be exactly that of plasma (13). P.A. Jansson obtained similar results in man by measuring glucose in the subcutaneous interstitial fluid, collected by a microdialysis technique (14). We implanted in the subcutaneous tissue of rats a needle-type glucose sensor similar to that described by Shichiri. By using a two-point, in vivo, calibration method, we have been able to demonstrate that glucose concentrations in subcutaneous tissue follow those in blood with a time lag shorter than 5 min (15). Finally K . Rebrin, in the group of U. Fischer, demonstrated that a closed-loop insulin delivery system, monitoring glucose concentrations in the subcutaneous tissue, was able to achieve perfect regulation of blood glucose concentration dur-

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ing an oral glucose load in diabetic dogs, the results being similar to those obtained when glucose was sensed with the same machine directly in blood (16). Taken together, these results validate the choice of the subcutaneous tissue as a site for glucose monitoring. Several groups are now working on the development of such a subcutaneous needle- type glucose sensor because it is generally believed that it should be easier to avoid an adverse biocompatibility reaction with a subcutaneous implant than with a system placed in contact with blood.

However, the second question to consider again is, obviously, how long does the system work. M. Shichiri first implanted the sensor in the subcutaneous tissue of human beings and was able to demonstrate that it was operating for 3 to 4 days, but no more (12). Indeed, there was a progressive decline in the sensor’s sensitivity, which was at first im- puted to a fouling of the sensor’s membrane by pro- teins and/or cells present in the subcutaneous fluid. This hypothesis was challenged by the group of U. Fischer, in experiments performed in dogs with a similar sensor and showing the same loss of sensitivity after 2 to 3 days. The authors demonstrated that after explantation, the sensor’s characteristics determined in vitro were very similar to those found before in vivo residence. They made the important observation, by using wicks implanted over this period of time, that the glucose concentration determined in the drop of liquid collected from long-term implanted wicks was no more that of plasma, but much lower (25%). They concluded that the inflammatory reaction around the wick consumed glucose, which is obviously a disaster since the analyte has no more access to the electrode (17). Here again, we observe a direct effect of the biocompatibility issue on the function of the system. It is especially harmful that these detrimental events seem to occur within a very short period of time, too short to contemplate the glucose sensor even as a disposable device. Even if the needle-type glucose sensor is supposed to be changed every 2 days, as is the needle of an insulin wearable pump, it is obvious that a 3-day lifespan is insufficiently long enough to be considered as safe. At the end of this lecture, we will come back to the biocompatibility of glucose sensors and propose a possible solution. But before- hand, we will consider another approach toward the development of an artificial pancreas.

The Bioartificial approach

Although human ingenuity has provided the basis for a multitude of discoveries, it will never invent anything

more beautiful, more simple, or more direct than that which have been created by nature, in which nothing is missing, and nothing is superfluous.

Leonard0 da Vvnci

If insulin-dependent diabetes mellitus is due to the destruction of the cells that secrete insulin, the best solution should be the graft of pancreatic tissue. Indeed, these cells have the property of re- sponding rapidly to a glucose challenge, proportionally to the glucose concentration of the stimulating medium. There is a glucose sensor in the beta cells, and the machinery responsible for insulin exocyto- sis from the cell has a very fast response time (within I min). Moreover, the cells have sensorb for other nutriments that normally stimulate insulin secretion (amino acids, free fatty acids, hormones). In addition, unlike implantable pumps, the pancreatic beta cell has the potential of synthesizing insulin to restore the reserves of the hormone. Finally, the islet volume responsible for the secretion of the daily insulin requirement (50 IU) is extremely small: One million islets of Langerhans represent a volume smaller than 1 ml. They could therefore be readily implantable.

So far, approximately 3,000 insulin-dependent diabetic patients have received a pancreatic, vascularized graft with a rate of success that is now around 70% and a patient survival rate of 90% at 4 years (18). Only a few patients (approximately 10) were transplanted with isolated islets with success (19). These numbers are insignificant when com- pared with those of kidney transplantation, insulin- dependent diabetes being paradoxically still 10 times more frequent than end-stage renal failiire. There are two explanations to the limit on the use of pancreas or islet transplantation. The first is shared by other organ transplantations: the poor availability of transplantable tissue, especially for islet transplantation since with the available techniques, it is difficult to obtain enough islets from a single donor. The second is more particular: as previously mentioned, diabetes is not an immediately life- threatening disease, and unlike end-stage renal, car- diac, or liver failure, does not justify the lifelong use of immunosuppressive drugs. This explains why most of the grafts performed so far were carried out in patients with diabetes-induced end-stage renal failure, justifying a combined kidney and pancreas transplantation. This strategy is somewhat paradox- ical as only patients who already have diabetic complications could be cured, thus limiting the interest of pancreas transplantation for the primary prevention of diabetic complications, which is still its main rationale.

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The beauty of the bioartificial pancreas concept (20) is that it should overcome both these obstacles. Indeed, let us suppose that we can place the islets inside a bubble, made of an artificial membrane, permeable to glucose and insulin, but not to antibodies and lymphocytes. This would allow the use of xenogeneic islets (for instance porcine islets, whose availability is unlimited) in the absence of any immunosuppressive therapy.

This concept of a bioartificial organ is entering the field of artificial organs as a very promising one. It is not restricted to the replacement of the endocrine pancreatic function, but to that of any cells that do not require direct contact with the host tis- sues, namely that have an endocrine or detoxifica- tion function. Bioartificial parathyroid and liver (21) are two other applications that are currently under investigation. Bioartificial (or hybrid) organs are now present as major topics in meetings devoted to artificial organs. Is it not true that the first engineer, the great Daedalos (whose name means “worked with art”--art, as in artijci-al organs!), imprisoned with his son in the labyrinth, constructed, to get out, wings attached with wax? A bioartificial wing? Scholars say that the Daedalian work has a unique feature: duality of the materials that are used, conjunction between the living and the artificial. . . .

Coming back to the bioartificial pancreas, several approaches are possible. Vascular devices, in which blood of the host circulates in contact of a membrane (for instance, through a semipermeable hollow fiber), the islets being placed on its other side in a closed compartment, would be very similar to a hemodialyser, the “dialysate” compartment containing the islets being closed. Simpler systems, referred to as extravascular systems, including microencapsulated islets or islets placed inside a hollow fiber, may be implanted inside the peritoneal cavity. Whatever the geometry used, here it is again necessary to verify that the system performs as a closed-loop insulin delivery system, i.e., respond- ing to a glucose load by an increase in its insulin production and able to correct diabetes and that the membrane provides efficient immunoprotection for the cells. These requirements refer to the function of the system. Next, it is necessary that the system function in vivo over a long period of time. The system must also be implantable. This raises the issues of its lifespan. Again, we want to point out that, so far, most of the studies have focused on the first question, dealing with the function of the systems, and not with those related to its implantability. We will give a few examples of bioartificial pan-

creases that function satisfactorily in vitro, or even in vivo for a short period of time, but whose long- term function remains questionable.

Let us consider first the vascular systems described above. It is possible to design systems that perform as potential closed-loop insulin delivery systems, namely, that increase their insulin release in response to glucose. However, the design of the system must be carefully considered. The kinetics of the response might be inappropriately slow if the volume of the islet compartment is too large (22). We demonstrated that due to the drop in hydrostatic pressure along the fiber, at the inlet of the fiber, the hydrostatic pressure is greater than the pressure present in the islet compartment. There- fore, an ultrafiltration flux from blood to this compartment should be present. Similarly, in the second half of the fiber, the hydrostatic pressure has become lower than that in the islet compartment, and a back-filtration flux should occur from this compartment toward the bloodstream (23). Obvi- ously, if one wants to use this flux to accelerate glucose and insulin transfer, a U-shaped configura- tion should be optimal since in this case, the ultrafiltration-back-filtration flux would cross the islet compartment as a short circuit. A device constructed with this design has been able in vitro to produce insulin in response to glucose with perfect kinetics (24). We also verified, by connecting the device to normal rats (25) and to dogs over a few hours (26), that when perfused with blood the system responded to a glucose challenge by increasing its insulin production, despite the presence of a semipermeable membrane between the islets and the millieu. In addition, the short-term correction of diabetes in rats was demonstrated. Since such a membrane is impermeable to cells and antibodies, it should provide cell immunoprotection. These results indicate that this system works in vitro and in vivo for short periods of time.

A second question is how long do these systems work? The long-term survival (at least 3 months) of islets placed inside a vascular bioartificial pancreas perfused with a culture medium has been estab- lished (27). But these were in vitro studies. In vivo studies with these devices were curtailed by the occurrence of clotting. Very recently, the group of W. Chick published evidence that a vascular bioartifi- cia1 pancreas, made of a large hollow fiber, re- mained patent for several months when implanted on a vascular shunt of dogs, in the absence of hepa- rinization of the animal (28). However, the rapidity of the insulin ripost to a glucose load was far from being optimal; here, the implantability was the first

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concern, yielding a device whose functionality remains to be improved.

Microencapsulated islets of Langerhans have been proposed by the group of A. M. Sun in To- ronto as a bioartificial pancreas (29). This system does not require vascular access since the microencapsulated islets can be implanted into the peritoneal cavity, and one would thence use the exchange properties of the peritoneal membrane. Concerning the first of the questions delineated previously, we demonstrated that the size of the capsules is criti- cal. Insulin release in response to glucose by microencapsulated islets was observed under in vitro, incubation conditions only when the size of the capsules was within the 400 pm range, and not when it was 800 pm (30). We also demonstrated that the membrane of alginate-polylysine microcapsules protects the cells against the cytotoxic effect of anti-islet antibodies present in the sera of insulin- dependent diabetic patients (3 1).

Microencapsulated islets were implanted in vivo in the peritoneal cavity of rodents, dogs, and monkeys and were found to be able to correct diabetes for several months (32-34). However, it was recognized that an inflammatory reaction around the membrane occurred both with hollow fibers containing the islets (a third type of bioartificial pancreas), and with microcapsules. This reaction was prevented by treatment of the animals with dexa- methasone (35) or with superoxide scavengers (36). Is it possible, for instance, to recall the monokine hypothesis and to speculate that the outer membrane of the system activates the alternate pathway of complement, which activates macrophages to release interleukin- 1, which might promote fibrosis‘? This fibrotic reaction might limit the function of the system, not only by altering glucose and insulin transfer through the membrane (it is even possible that the inflammatory reaction may hinder the access of glucose from the milieu interieur to the bioartificial pancreas as suggested by the wick experiments described above), but also maybe via the toxic effect of some diffusible substances produced by activated macrophages, such as interleukin-1 or free radicals, which could cross the membrane and kill the islets (37). The toxic effect of interleukin-1 on islets of Langerhans has been demonstrated even when they are microencapsulated (38). Here again, either in the form of blood coagulation, in the case of a vascular system, or in the form of a foreign body reaction, in the case of an extravascular device, the biocompatibility-related phenomena would directly, and rapidly, jeopardize the function of the system. It must be understood that overcom-

ing this obstacle represents a formidable task because blood clotting and foreign body reaction are the two powerful primitive mechanisms invented by nature to protect us against the danger of wounds; by trying to resolve this problem, we are just fight- ing against evolution. The importance of this problem for all artificial organs was recognized during a workshop (Biocompatibility in Artificial Organs) or- ganized by the European Society for Artificial Or- gans, held in Erice, March 29-April 1 , 1989.

Is it an impossible challenge? It is impressive to note that in the past year, several teams, both in the academic and in the industrial field, have tackled the development of bioartificial organs, generating impressive data (most of them are not yet published) on the long-term survival of vascular and extravascular (microcapsules and hollow fibers) systems (39).

CONCLUSIONS

Artificial organs: biocompatibility or biostability?

To dissolve a conflict, the conditions that produce i t are changed so that it disappears. This can be done by changing either the environment or the opponents.

Russel L. Ackoff

Recently, in collaboration with G . S . Wilson at Kansas University and D.R. Thevenot at the IJni- versity Paris Val de Marne, we have developed a glucose sensor that was really miniaturized (40). i t consisted of a platinum wire anode, coated with teflon, except near its extremity where glucose-oxidase was layered, reticulated with glutaraldehyde, and covered with a layer of polyurethane. A coil of silver-silver chloride surrounded the platinum wire and was used as the cathode. The whole diameter of the anode was 0.25 mm. This sensor was implanted in the subcutaneous tissue of rats, and unexpect- edly was found to work for at least 10 days. Histo- logical examination of the tissue surrounding the sensor showed the presence of neocapillaries very close to the sensing area of the glucose electrode. It is possible that the small size of this sensor was responsible for this novel kind of reaction to an implant, which was different from the foreign body reaction previously observed.

We propose that in this case, the reaction to the implant, by changing the environment of the sensor, was actually beneficial to its function. Indeed, the definition of biocompatibility, given by the Euro- pean Society for Biomaterials (4) is “the ability of a material to perform with an appropriate host response in a specific application.” We have seen that

ArlifOrgclns. Vol. 16, No. 1 , 1992


the host response can be inappropriate (for instance, an inflammatory reaction that consumes the analyte, which cannot reach the biosensor, or that produces interleukin- 1 , which crosses the membrane of a bioartificial organ and kills the encapsulated cells). But it can also be appropriate (without being inert) and lead for instance to the formation of neocapillaries which improve the access of the analyte to the sensor, or the transfer of signal and nutri- ents to the encapsulated islets of Langerhans and the return of insulin. Thus, it can be expected that ways of improving angiogenesis around implants could represent a promising direction for research. Thus, the goal to reach would be the formation, around the implant, of a biostable, functional (i.e., vascularized), environment.

In conclusion, this review delineated some of the problems encountered during the design and the development of an implantable artificial organ. We hope that a major step toward the solution of this difficult challenge is the recognition of the importance of the biocompatibility issue, in addition to the functional aspect, which must itself be soundly analyzed and then never forgotten. And that, according to a connoisseur, “There always comes a time when creations of science surpass those of imagination” (Jules Verne).

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artificial and bioartificial replacement of the endocrine pancreas

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