Johns Hopkins Magazine - June 1995 Issue

Pig Hearts in Human Chests?

For heart patients awaiting transplants, pigs may pose the most promising prospect - - not too many years from now. That's where Bill the monkey comes in.

By Elise Hancock

March 13

Bill is one unhappy cynomolgus monkey. Head drooping, he huddles in the back of his cage, facing the corner. He ignores Liz Davis, who is attempting to feed him a slice of black-ripe banana - - a major treat, most days.

"Come on, Bill," she coaxes, "here's some banana, just the way you like it. The others would kill for this."

The banana touches his muzzle, but Bill shakes it off, then swivels his head around and bares his teeth. He turns back into his corner, with a certain air of finality, and begins to scratch, reaching up under a vest that protects his surgical wound. But he can't qui-ite reach that itch, try though he may. It's driving him crazy. No, he does not want any banana. Nor will he take a Primatreat. Not even a pink one. And just in case anyone has missed the message, he continues to look around now and then, to bare his teeth.

Bill probably doesn't feel very well. Four days ago, he spent six hours under full anesthesia, while surgeons transplanted the heart of a week-old pig into his abdomen, seaming it into the abdominal aorta. Now the pig heart beats alongside his own, bubom, bubom, bubom.

"This room is his intensive care," says Davis, a third-year resident at Hopkins. A Cornell math major (Phi Beta Kappa), she graduated from Hopkins medical school at the top of the class in 1992, then did two years of general surgical residency. In six more years, she'll be a cardiac surgeon. At this moment, though, she is "Bill's ICU doctor, and his nurse too. And his cage cleaner." A tall, slender woman with long dark-brown hair, she is dressed for the job, wearing scrubs and a mask.

If Bill had received a true transplant, a replacement for his own heart, the surgery would have been longer and more complex - - a needless risk, for now. Running the pig heart in Bill's abdomen, pumping Bill's blood, will be good enough. It will allow a team of Hopkins researchers, headed by pathology chair Fred Sanfilippo, to assess how well an experimental protein, given with standard therapy, can ward off the so-called "complement cascade" of the immune system - - the major barrier to transplantation from one species to another.

Sanfilippo, 45, is a baseball fan (let's get first things first), a compact man with springy black hair and mustache. He came to Hopkins in 1993 from Duke, where he'd been full professor of pathology, immunology, and surgery. The three fields come together in this research, which Sanfilippo hopes will ultimately solve the growing problem of donor organ shortage.

The problem is that each year more and more people need transplant hearts, kidneys, or livers, while the number of people who donate organs remains much the same. In 1993, in the United States alone, 28,000 people died while waiting for a human organ. "The greatest risk for patients needing a transplant is death waiting for the transplant," says Sanfilippo. "And if you're the parent of a child needing a transplant, your child's greatest risk of death is lack of a donor. So, where's the solution? Well, I would maintain that a key option that ought to be explored is xenotransplant, using a food source as a donor."

Pigs, in a word.

Why pigs? Because - - unlike wild primates - - they carry no viruses that are deadly to people. Because they breed prolifically. Because most peoples of the world already eat pork, so the ethical problem is small. And to top it all off, of all the hearts in the animal kingdom, those of pigs are most like those of people. That's true even down to size, says William ("Wink") Baldwin, an MD-PhD immunopathologist with whom Sanfilippo has been collaborating for almost 10 years. "Pig hearts come from pediatric size to full-sized adults. In children, we assume the pig heart would grow to respond to the workload as the child grows."

"Yeah!" says Davis, enthusiastically. "If you need a heart, you'll just order a number 2, or whatever." There'd be no shortage of donor hearts, and no wait.

That prospect assumes, however, that transplant rejection can be prevented, which is hard enough in human-to-human transplants (or any other allograft, a transplant within a single species). That's mostly because of cellular immunity, the process by which T-cells orchestrate the banishment of tumors, fungi, viruses, and tissue inflammation - - as well as non-self tissue, the transplant organ. Unless prevented by antiimmune drugs, cellular immunity will slowly but surely destroy any allograft.

With xenografts, tissue transplanted across species lines, the problem is even worse, for the host's immune system reacts with all-out vigor, as if torching an oil field. Known as hyperacute rejection, it's the body's chief weapon against bacteria.

Evidently, to the immune system, a xenograft looks like one great big blob of infection, so within seconds, pre-formed naturally occurring antibodies (NAb) bind with antigens from the transplant. That triggers the complement cascade, a series of 20 proteins in the blood serum that clog arteries and kill any graft cells the blood can reach. It's both quick and lethal. Sanfilippo's lab has found that pig hearts run on human blood, in vitro, will live only 25 to 34 minutes before they stop dead, mottled with hemorrhage. Even at first, the hearts beat half-speed.

Enter sCR1, a recombinant (and soluble, hence the s) form of Complement Receptor 1, a natural protein that keeps your complement from eating up your organs every time it attacks bacteria. The drug was developed at Hopkins in the late 1980s by distinguished immunologist Douglas Fearon, and it is now licensed and produced by T-Cell Sciences.

Happily, Sanfilippo's group has found that Fearon's stepped-up version of the protein seems to protect xeno-grafts and to have real clinical potential. With sCR1, the NAb still bind the antigen, but there's no active complement to be triggered. The proof: Based on five years of work with rats and other experimental animals, Hopkins postdoc Scott Pruitt treated two monkey colleagues of Bill's with infusions of sCR1 after they were given pig hearts. In these animals, the grafts survived five and seven days, respectively.

Bill represents the next step. He is the first xenograft "patient" to be treated with both sCR1, to stave off the complement cascade, and a trio of standard drugs - - the "triple therapy" - - that prevent cellular rejection: cyclosporine, cyclophosphamide, and a steroid. "We don't know what immune response Bill will mount," says Baldwin. "Nobody has any idea. Will it be like an allograft, or will it be something new?"

Davis, Baldwin, and Sanfilippo are hoping that Bill will live for at least two weeks. Then they'll know they're on the right track.

Davis still wants Bill to eat. In the long run, his nutrition will be important. A green Primatreat? No. A purple one? Bill shakes it off.

"The principal investigator for the entire program is Dr. Sanfilippo," she explains, "but this is basically my experiment. I participated in the planning, and I make the decisions day to day. Doses, schedules, the IVs, figuring out the regimen of drugs, exactly how to do it, how to keep Bill warm after surgery." Davis stays with Bill around the clock. On Day 1 and Day 2 after surgery, she even slept in the same room. "I've been home only twice," she says, "just to take a shower."

Already, she has the swift, decisive speech and manner of a surgeon. (Or is it that surgeons are born, not made?) When she speaks to Bill, however, her words come more slowly, and she slides into that unmistakable, softly emphatic speech that people use with their very small children. "Just think," she tells him. "You're in the jungle, foraging for grapes.... nice grapes... Now wouldn't that be good?"

No, it would not. Every line of the little body says no, no, no.

Actually, given that he's four days out of surgery, Bill is doing well. The second heart in his abdomen beats strongly, and he's active. He comes often to the front of the cage, eager to see who's coming in the door, and he seems to enjoy socializing with people. He often "talks" back - - smacking his lips, pch pch pch, like a series of quick dry kisses to the air. His wound is healing well; there's no sign of infection, even though his immune system is profoundly depressed. In fact, explains Davis, his present foul mood is a very good sign. Bill now feels well enough to act feisty.

The question is, how to help him do even better? How to make him more comfortable? How to get him eating more? "I wonder if the Betadine [antiseptic] is making him itch," worries Davis. Or perhaps, despite all the human attention, Bill is lonely. Should she bring in a TV set? "The vets tell me that monkeys really like the talk shows."

Bill is the first xenograft "patient" to be treated with both sCR1 and a trio of standard drugs. The researchers are hoping that he will live for at least two weeks.

Day 5

Good news - - Bill is eating and has had a bowel movement. The intestinal tract often goes on vacation after surgery. Now Bill's is back at work. And he's grooming himself, absorbed in plucking invisible specks off his left arm. That, too, is a sign of well-being.

In fact veterinarian Bob Adams, doing his morning rounds, was very pleased with his patient. Betadine does itch, he told Davis. They sprayed Bill with water from a squeeze bottle, washing down his body, and he turned and pranced in the stream. From now on, Davis will gently rinse off the Betadine after she cleans his wound, as she is now about to do.

It's a cautious process. Bill may be friendly - - "he's the sweetest," interjects Davis - - but he's still a monkey. If he feels threatened, he may bite, and he may carry herpes virus B, which causes fatal encephalitis in 70 percent of humans. To handle him, Davis wears heavy bright-blue gloves, elbow-high, and she needs help from Tuan Lam, a Hopkins medical student who is spending a fellowship year in the lab. (When he is not "paying his dues" as bottom man on the totem pole, he is developing molecular means of inhibiting complement.)

Lam spends a lot of time in this room, too, monkey-sitting while Davis works in the lab, devising a way to prevent neutrophil binding (another immune mechanism) on guinea pig hearts transplanted into rats. So Bill knows Lam well, and likes him.

Everyone who comes into the room wears a mask, yet Bill can tell people apart. He comes forward. "Pch pch," he says in greeting. He accepts a grape, judiciously nibbling off the skin (which is hard to digest) before he pops in the morsel.

Then he sees that Lam and Davis are removing his hard rubber toys, and he pees and backs to the far side of the cage. "He knows the cage is going to squeeze him," says Davis.

Squeeze it does. Lam slowly pulls forward Bill's perch, two bars that run front to back, so that the back wall nudges the cringing monkey forward, forward, forward, to the waiting hypodermic. "It's a light dose of ketamine," explains Davis. "It's a dissociative anesthetic. You're awake, but you don't care what's happening."

Soon Bill is out cold, belly-up on a sterile pad on the floor. Kneeling beside him, Davis removes his protective vest, then checks his suture and treats it with Betadine. (And rinses it.) Good. No signs of infection. She checks the metal-covered lines that carry Bill's immune drugs through the top of the cage and into his bloodstream - - good. Nothing perforated or leaking. She lays a gentle hand on Bill's abdomen, feeling the donor heart while Lam watches the clock - - good, 172 beats per minute. "The strength of the beat is nice," she approves.

She gives him a shot of antibiotic, and also takes a blood sample, just as small as possible. "He lost blood in surgery, and you can't take a lot each day. Each day you've got to think, which ones do I really have to know?... If his hematocrit were low, there'd be nothing I could do about it." So, no hematocrits today. She does check cyclosporine levels every day, and platelets and kidney function every other day.

Finally, she turns the limp body onto his stomach ("I'm sorry, Bill," she whispers) and puts his vest back on. It must fit just right: loose enough not to block his catheter, tight enough that he can't scratch the incision. Still out, he's replaced in the cage, lying on his side. Back in go the toys, and Lam locks the door. Then Lam and Davis wait, watching, till Bill rouses and sits up. He is somewhat wobbly.

It will be half an hour before he's back to normal, pch-ing and eating and viewing himself in Davis's hand mirror. Or just sitting. Like anyone else, Bill needs some alone time. When he wants to be alone, he goes to the back right- hand corner of the cage and turns away. "He's picked that for his sleeping place," says Davis. "He cleaned it today."

While Bill rouses, she explains the thinking behind his drug regimen: sCR1 is extremely powerful, not something you'd want to take for years. Indeed, it's doubtful that anyone could survive long-term with complement suppressed - - you'd die of infections. "So the hope is that at some point we could stop the sCR1 and send patients home on triple therapy."

But what would happen when the complement recovers? NAb would still be binding antigen from the graft. Wouldn't that trip off the complement cascade?

Probably not, says Wink Baldwin. He and Sanfilippo think xenografts are analogous to transplants between people with incompatible blood types - - A and B, for instance. In the past, such a "discordant" transplant would activate the complement cascade and shrivel, just like a xenograft. But about six years ago, it was discovered that if the host's levels of NAb can be drastically reduced around the time of the operation, hyperacute rejection does not occur. "It's referred to as graft adaptation," says Baldwin, "which means that we don't know what's going on. But it's reliable."

In addition, Baldwin and Sanfilippo expect to combine the use of sCR1 with other approaches, such as transgenic pigs. In this approach, pioneered by David White, in Cambridge, England, you genetically engineer pigs to express certain human complement regulators, which could include CR1, in their blood vessels. The heart from such a pig, once in the patient, would presumably be able to protect itself from the patient's complement, while leaving the immune system intact to deal with infection elsewhere. While immune suppression would still be needed, that would be a big step forward, says Baldwin.

Baldwin and Sanfilippo are also enthusiastic about "mixed chimerism" ("chimer-" as in "chimera," one animal having parts from two). David Sachs at Harvard has shown the way here, working with pig hearts in cynomolgus monkeys. At the time of the transplant, says Baldwin, "you'd take bone marrow from the pig and put it in the human." The pig bone marrow will provide immune protection for the pig heart, "and eventually the pig cells will be accepted as self. The hope is that eventually one wouldn't have to immuno- suppress." So sCR1 plus chimerism may be the next step for Sanfilippo's team, once they've learned how far they can go with sCR1 and anti-immune drugs. Sanfilippo and David Sachs are talking it over.

Both Baldwin and Sanfilippo are optimistic about combining these techniques, because chimerism does occur in human-to-human transplants. Baldwin says that in most cases where a graft is entirely successful, such as a kidney that lasts for 30 years, chimerism has developed, at least to some degree. That is, blood studies show that the person's marrow produces some leukocytes and other immune cells that match those of the organ donor. The chimerism happens by accident. Apparently donor stem cells lurking in the organ's leftover blood migrated to the bone marrow and set up a colony.

Of course, researchers immediately set out to reproduce the effect, seeking to improve it. Last year surgeons from the University of Alabama found that marrow transfusion helps transplants dramatically. In 35 patients getting kidney-plus-marrow, where chimerism occurred, 91 percent had no rejection episodes; without chimerism, only 9 percent were trouble-free.

A high degree of accidental chimerism may explain why a few lucky people who stop taking their immune-suppressing drugs (they're expensive, and Medicare/Medicaid do not pay indefinitely) still don't reject the organ.

If you have had a transplant, Sanfilippo and Baldwin urge that you note those words "few" and "lucky," and do not stop your drugs on your own. "Once you've lost ground," warns Baldwin, "it's very hard to regain it."

Day 7

It is 7:45 p.m. - - close to Bill's bedtime. Lights are out, except for a glow to the left, where Liz Davis sits on her "bed" - - a foamblock chair that folds out. Light from a small lamp reflects cozily off her charts and papers, and in the warm gray dimness, Bill drowses in his sleeping corner, head between his knees. Goodnight, chair. Goodnight, room.

Shock! Seeing the visitor, Davis flicks on the overheads. Bill sits up, shaking his head.

Night before last, she reports, she'd had a glitch. Davis checks Bill's lines at regular intervals, to make sure his IV and drugs are flowing at the right rates. (For sCR1, "right" means exactly .04 cc. per hour.) For the middle-of-the-night check, she sets an alarm. And that night, in the middle of the night, she found Bill's sCR1 line kinked off. He'd probably missed the drug for two to three hours. Quickly, she sedated him and unkinked everything.

She says it was "no big deal, probably, because the half-life of the drug is eight hours." He'd never been even close to without it. She hopes. And yesterday was a great day - - a 27-grape day. Bill ate well; he was highly alert and active, climbing his cage; and his platelet count came back to normal, 250,000. Since the body uses platelets (crudely speaking) to plug up injuries and stop any loss of blood, the normal count means he's healing well. It also means there's no cardiac thrombosis - - a sign of rejection, and another condition that uses up platelets and would drop the count.

Still, Davis is cautious. "I like him too much and there's too much [money and effort] invested in him. My theory is, anything that could go wrong will go wrong. So last night I was overly anal and I woke up every two hours and checked his lines. Today both he and I are exhausted." Tonight, she'll check every three hours.

She went to the Superfresh yesterday and bought Bill $7 worth of goodies. One of his drugs causes food aversion, so he's finicky. Variety matters. Today he seemed bored with grapes, and she no longer bothers with bananas. The new hit is raspberries. As she talks, Davis is scratching Bill's back, reaching into the cage with a long, long set of tongs.

Drowsy though he is, Bill looks positively rakish. Before his surgery, the veterinary anesthesiologist, Deborah VanKempen, had shaved around his neck and ears with maybe excessive zeal, and Bill looks as though he'd had a Mohawk. On top of his head, spiky dark- and light-brown hair sticks straight up. His pink ears are mottled with irregular dark blotches. One looks exactly like Australia.

When he tires of having his back scratched, Bill turns and lowers his head to invite the tongs elsewhere. Davis knows just where he wants it: that ridge along the shoulder where muscles get tight.

"I love animals, I really, really do," she says. "I certainly have a hard time hurting animals, putting them in positions they don't enjoy, like with the vest." Even with rats, she finds animal research difficult.

Bill reaches around and taps his back, and the obedient tongs move again, now stroking up and down either side of the spinal cord. The animal leans into the stroke, while Davis's voice murmurs on, repeating thoughts and doubts worn smooth with repetition. "I get a little bit of comfort, though not complete, if I make sure they don't experience any pain or discomfort.... I don't like that he's in a confined place. But if we set him free, he wouldn't survive. He wouldn't know how."

Click, lights out. It is 8:30, Bill's bedtime. Liz Davis's, too. She hauls her chair-bed into the next door cubicle and spreads it out with the hint of a slap.

Day 8 - - Good news! Bill is doing well. He's gone longer than both the sCR1-only monkeys. He is still infection free, and the pig heart is beating away strongly.

In the 1960s and '70s, transplant researchers had assumed that organs from monkeys, chimpanzees, and other non-human primates would be the best non-human transplants for people, since all primates are genetically quite similar. Between chimpanzees and people, just over 1 percent of the genome differs. Primate transplants were even tried a few times, with resounding failure. (Do you remember the short-lived "Baby Fae," to whom surgeons at Loma Linda gave the heart of a baboon?)

By the 1980s, however, the community of transplant researchers had grown uneasy about this approach. Wild primates often carry viruses that can be lethal to people, they breed slowly, and many species are endangered. But the major issue was ethical: "You can't work with these animals and not feel it," says Sanfilippo. "They're so much like us. You've only to look at their hands." So in 1989 the researchers held one of those quiet little seminars - - no press, only world leaders invited - - where research directions get changed.

At one such conference, in 1974, the world's DNA researchers called a halt to their own research for more than a year, until they could ensure that mutant DNA wouldn't get away and run amok. At this conference, the subject was xenotransplantation, including several sessions on the use of primates. "We invited Jane Goodall," says Sanfilippo, "and after we talked over what she had to say, well..."

The famous primatologist's presentation had centered around some of the chimps she's known in her 29 years observing them at Gombe National Park, Tanzania. (For more, see her book, The Chimpanzees of Gombe, Harvard University Press 1986.) She told stories of Flo, that "tolerant, affectionate, protective and playful mother." Of neurotic Flint, a spoiled brat who insisted even at the age of 6 that his mother carry him, and who died of grief when she died. Of 12-year-old Spindle, who adopted the orphaned Mel, sharing food and nest. Of Mike, who became alpha male despite his small size, by means of his brain: He taught himself to menace the other males by rushing at them kicking three empty kerosene cans all at once, which made such a gaudy racket that he never had to fight. Goodall closed with the tale of Old Man, a chimpanzee in Florida who saved his keeper's life, protecting the man from other chimpanzees. We humans might care to reciprocate, she suggested.

"They're like us," repeats Sanfilippo. "They think, they have emotions, they have families, they have societies, they can be deceptive, they can kill each other, they can be altruistic." His voice trails off.

In their discussion (quite heated, says Sanfilippo), most conferees came to agree that the other primates' less-than-human intelligence does not justify their use for transplants: "Healthy chimpanzees have higher IQs than many retarded humans," as Goodall pointed out - - yet no one would advocate the use of retarded humans as organ donors or experimental models. Question closed.

Since that conference, research has largely ceased on ways to use primates as organ donors. Pigs are where the action is.

Primates remain essential to transplant (and other) research, however, because they are the closest possible stand-ins for humans. To abandon their use entirely would be to halt the work; one can only go so far with rats et al. But researchers now use primates as sparingly as possible. Sanfilippo believes that most accept the general criterion that sacrificing a single primate life is justified only when there is potential to benefit large numbers of human patients. By law, all research protocols in the United States are reviewed by an ethics committee at the researcher's institution, and the performance of the committees is itself reviewed by government agencies.

Day 8

Wow! Bill is doing well. He's gone longer than both the sCR1-only monkeys. He is still infection-free, and the pig heart is beating strongly.

Monkeys do not smile, so it's hard to be sure, but Bill certainly appears to enjoy teasing Davis. She is offering a glob of indeterminate brown goo, holding the spoon in a long clamp. (It's Nutracal, a high-calorie nutritional supplement.) Bill pushes the spoon away, several times, but finally comes to the front and moves his face slowly forward, forward, toward the spoon. He backs away and shows his teeth, then relents and nibbles a little. Hmm. Not bad. He takes more. But it's sticky. By now blobs of brown goo ring his mouth, quivering. One drops on his hand; he looks at it with distaste, then smears it on the cage.

Now Davis is chasing his face around the cage with a syringe full of orange juice. "This morning you liked it, Bill - - it will wash down this stuff." He finally takes it from his water bottle, sucking, and Davis makes notes. So far this afternoon he's eaten four and a half grapes, 10 cc of orange juice, and two smearsful of Nutracal.

She dangles a bunch of grapes before his mouth. "Feed me grapes, my sweet." He takes one more grape, then cleans his hand and face, a matter of meticulous licking, and deigns to accept a purple Primatreat, tucking it into his right cheek pouch. Every once in a while he palms the cheek to pop out the purple wafer, inspect it, smell it, and chew off another piece - - snap! it's hard, like a big Necco wafer.

He refuses a raisin, refuses apple sauce, and goes to the back of the cage, where he eyes Davis with what appears to be a twinkle. She laughs. "Bill must think I'm like an Italian grandmother. `Eat! Eat! ...(reproachful) It's good! Don't you like my cooking?...Eat!'"

It's Friday. Will Davis please keep the writer posted over the weekend? Sure. She takes the phone number and tapes it to the wall with others: Wink Baldwin. Fred Sanfilippo. Tuan Lam. Anesthesiologist VanKempen (a daily visitor). The vet. And the veterinary pathologist. "F.O.B.," says Davis. "Friends of Bill."

Tuan Lam arrives. "Who's that?" she says. "Who's that? Is that Tuan-y? Is that Tuan-y?" Bill comes to the front of the cage. "Pch," he remarks.

Lam will monkey-sit while Davis goes for a shower - - and to drop in on a friend, too, just for a minute. It's been spring out all week, people tell her, while she's been shut up in cinderblock, never seeing daylight. She'll be gone about two hours, she tells Lam. She shows him an air bubble in the syringe of sCR1 - - "Just make sure it doesn't get down there, okay?" - - and strides off, brisk as ever.

Day 9

Davis calls. Bill's feeling fine, though his platelets are down to 50,000. (That may be a sign that he's getting too little cyclophosphamide - - something they can correct. Or it may signal rejection taking place.) He's eating well. Today he reached right out of the cage to take three spoonfuls of trail mix. Most of the food from the Superfresh was going bad, so she took it over to the other monkeys. Boy, they loved it. Mostly, they get monkey biscuits, she says. "Bill eats at the Prime Rib."

Bill is alert, and kidney function is good. He shows no signs of toxicity or infection.

Day 11

The voice on the phone is flat. "Bill is no longer with us." When Davis sedated Bill this morning for his normal checkup, "the pig heart was not beating. It was stone-cold." He had rejected overnight.

"As soon as I saw that," says Davis, "my heart sank. Some because he had in fact rejected - - we only got to 11 days. More because I knew what that meant. We were going to have to sacrifice him." (Why not just remove the pig heart? Because the team needs to check for impact on Bill's major organs, says Sanfilippo. In addition, Bill would face a protracted post-surgery period of feeling miserable, only to spend the remainder of his long, dull life in a cage.)

Bill was still groggy from his sedative, but Davis and Tuan Lam tried to feed him - - "his last meal, I guess." They also took some pictures, to remember him by, and Davis scheduled O.R. time immediately. When Debbie VanKempen came down to do the anesthesia, she saw Davis's teary eyes and hugged her. "Oh Liz," she said, "don't start, you're going to make me cry. Think clinical, Liz, think clinical."

"So we pushed ahead," says Davis. With help from Lam and fellow postdoc Zhiping Qian, she performed the autopsy herself, tears running down her face. Bill died on the table in his sleep; he was infused with euthanasia solution, a barbiturate.

Inside, Bill looked good: no infections, no signs of obstruction. His liver, lungs, kidney, spleen, and his own heart looked entirely normal. "That's really nice," explains Davis. "If the animal were miserable and had infections everywhere - - that's not a viable therapeutic option."

The pig heart, she says, showed "what Dr. Sanfilippo calls an accelerated acute rejection - - an intense amount of hemorrhage, edema, dead cells and neutrophil infiltration." Though the small and medium-sized blood vessels in the pig heart were thrombosed shut (hence Bill's dropping platelet counts), the large ones still admitted blood flow.

For the following week, each morning Liz Davis woke up crying. "My God," she'd think, "I've killed my friend."

Why does she do research that involves animals, then? "I ask that question to myself," she answers. "I guess…Well, it's for an important reason." She is already excited about the next experiment, especially some things she's found about cyclophosphamide.

To an outside observer, this experience looks like one more rite of passage for Davis, not unlike the rocky time young interns go through the first time one of their patients dies.

T-Cell Sciences analyzed Bill's blood levels of sCR1, while Sanfilippo, Baldwin, and Davis analyzed blood and tissues, looking at immune response and its effect on both the graft and organs. After nearly five weeks of work, their broad findings were:

As in the earlier monkeys, sCR1 works to prevent hyperacute rejection. Complement was successfully blocked.

The triple therapy also worked. It not only prevented acute (cellular) rejection - - the pig heart showed few infiltrating T-cells - - but also delayed the acute accelerated rejection.

Yet Bill's immune system rejected the heart, generating substantial amounts of antibody, both IgM and IgG. "We've learned a lot from this experiment," says Sanfilippo: Obviously the team must shut down yet another immune mechanism, one triggered by evoked antibody response.

Sanfilippo has concluded that Bill's rejection was caused by a new surge of NAb, as well as by antibodies to the pig's Major Histocompatibility Complex (MHC). "It's like an immunization," explains Davis. "You get immunized, then antibody titers go up against whatever it is. We think Bill directed antibodies against the MHC and the natural antigens of the graft."That would explain why rejection did not begin right away; it takes time to build antibodies.

Well, Sanfilippo and Baldwin know ways to deal with evoked antibodies. In rats, anyway. Hmm. As a first step, in the next experiment, they will fine-tune the timing of cyclosporine and cyclo- phosphamide. Later, they may apply Davis's basic research on preventing neutrophil binding, which also pertains to accelerated acute rejection.

Sanfilippo is optimistic, Baldwin more guarded. "If we knew all the answers," he says, "it wouldn't be science."

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