A visit to Edmonton for islet researchers is like a visit to Mecca for Moslems or Jerusalem for Christians. Last summer Drs. Lakey and Shapiro decided that our bioartificial pancreas looked promising enough that it was worth trying in their best animal models at the laboratories of the University of Alberta. After months of preparation and preliminary studies with express mailed sheets, on March 18 we found ourselves at last flying to Vancouver then east to Edmonton. (Air Canada flies direct between Edmonton and Los Angeles but not San Francisco.)

The home of the University of Alberta and the Provincial Capital has about three-quarters of a million inhabitants. It lies on the northern reaches of the great plains, four hours east of the Rocky Mountains. The land beneath Edmonton is flat as far as one can see, relieved east-west through the middle by the meandering North Saskatchewan River. Flowing east northeast the river drops 1.5 kilometers and empties into Hudson Bay. Even in the late winter the river had an impressive flow; sometime it floods in the spring. North of the winding river is the Provincial Capitol building, a drab traditional dome, and the business district; south of the river is the University. The whole scene is reminiscent of Minneapolis-St. Paul. South Edmonton near the University was even once its own city, "Historic Strathconia." To the right is the view from our hotel, The Fairmont MacDonald (a 1900's effort at a 15th century French Chateau), across the river valley toward the University.

Three days of research were planned. We started Monday March 19th working with Christie Howie, a scientist who works for Prof. Shapiro in her newly equipped laboratory at the SMRI (Surgical Medical Research Institute). Christie had been making rat islets and shipping them to us in San Francisco. We made rat Islet Sheets for her, and she implanted them into rats in various locations. One activity on the 19th was examination of the rats. We were pleased to find that the rat Islet Sheets looked clean (free of fibrosis) after a week on the rat liver.

We also looked at electron micrographs of a rat Islet Sheet made by Ming Chen, a very skilled microscopist. He has produced some of the most striking images of artificial organs. The sheets we gave him had a rather low density of islets and the photos were not very impressive. So we told him if we got a lot of dog islets the next day we would make him a good sheet.

Another activity was preparation for making dog Islet Sheets the next day. We knew the next day would be long and complicated, so we were determined to get as prepared as we could be.

We store our highly purified alginates as dry powders. It takes a while to dissolve them in water. So as Rick and Randy set about doing that, our biggest problem appeared. Zut alors! (We were in Canada.) The alginate was pale yellow, showing it was impure! Randy and Rick went off in search of equipment we would need to do a last minute purification of the alginate. I got on the phone trying to reach the only person with a key to the Islet Sheet Medical laboratory who could help us, our friend Bill Usinger. Bill returned my call almost immediately, and borrowed a car to drive to the laboratory. He found the reserve alginate and dropped it at FedEx for next morning delivery. We crossed our fingers.

Late in the day Jon Lakey returned from a meeting at Lake Louise, and we met with him and Dr. Shapiro to plan the next day's pre clinical animal experiments. We reviewed all the latest data and decided to do one allograft using the islets from two donor dogs. Everyone was alerted to what would happen the next day and who was responsible.

At 7:30 the next morning the Edmonton team began the laborious process of making an isolate of islets of Langerhans from a dog pancreas. The pancreas is removed carefully so as not to damage the ducts and islets, then the pancreas is carefully digested with a special enzyme (that Jon Lakey helped develop). The picture shows three of the team members during a break in the action.

Christie Howie, Debra McGhee-Wilson, Jon Lakey, Scott King (masked)

Meanwhile I alerted Mary Swedberg, Jon Lakey's assistant, that our purified alginate was on the way. The FedEx web site told us that the package was in customs in Calgary. They expected to have it in our hands by mid-afternoon, Mary found out. She persuaded them to bring our package first: "'We need it for a transplant' works wonders!" she told me.

The alginate arrived at noon and the islets were ready at 1. Sheet fabrication began.

Meanwhile James Shapiro (l) and Jon Lakey (r) were preparing the recipient dog for the implant. First the pancreas was very carefully removed to minimize damage to the adjacent tissues. It is important that every fragment be removed because even a small number of host islets could muddy our results. Then, when the pancreas was out, the Edmonton metabolic team did an IVGTT: intravenous glucose tolerance test. Glucose, then L-arginine were infused and blood samples taken to look for any insulin. If there were any islets left, we would find them.

Jon's team had made a lot of tissue. In fact, we had 1,200 microliters (1.2 ml) of tissue to incorporate into sheets. This was much more than we had ever had before. Sheet fabrication went well. The picture below shows the result.


The dog Islet Sheet shown above is one of six that went into the pancreatectomized dog. The Sheets are about the size of a business card. You can see two rectangular zones (diagram at right). The islets are all within the lighter rectangle in the center. The fabric we use to strengthen sheets and prevent tearing goes all the way to the edge of the sheet. The "surgical cuff" is the zone that the surgeon sutures through to prevent the sheet from sliding off the liver. There are no islets in the cuff because we don't want to expose a single cell to the immune system when the suture needle punches a hole in the sheet.

All the sheets were sutured on to the dog's liver. The picture at left shows a dog Islet Sheet just after positioning on the liver and before suturing.

Meanwhile, back in the laboratory, Rick and Randy collected all of the islets left in the mixing flasks and crannies of our Islet Sheet making machine and made a final, small dog Islet Sheet for analysis. Ming Chen had gone home by then so we set it aside to give to him in the morning for electron microscopy.

By now it was 7 in the evening. We were happy. There were no disasters. All the problems that had come up were handled quickly. Most of all we were impressed with the professionalism of the Edmonton team. Everyone knew what they were doing and worked hard.

A final meeting with James and Jon, a review of the experiment, then off to dinner at Edmonton's fine "Characters" restaurant.

Next morning we got up and returned to the laboratory. We took our "left over" Sheet and went down to the islet laboratory to use the inverted 'scope. We were amazed at what we could see. The picture shows a high density dog Islet Sheet. The fibers are the mesh we use to impart strength and tear resistance to our sheets. The white spherical globs (except the orange one at top) are islets of Langerhans! (The yellow glob is probably pancreatic acinar tissue.) This is the highest density Islet Sheet we have ever made. To our eyes the islets look happy and healthy.

We cut the sheet in two, half for Jon and half for Ming. Jon did a static incubation assay to determine if the islets were responsive to glucose. In other words, did the entire process of making the sheet from alginate and islets damage the islets? (The islet function was found to be undiminished.)

The other half we dropped off at Ming Chen's SEM laboratory. He looked at the Sheet in his light microscope and his eyes lit up when he saw the islet density. He said maybe he could push this project to the front of the line and started fixing the sample. (Electron microscope samples are studied in vacuum, so the sample must be treated with chemicals and covered with a gold mask to permit viewing.)

We then paid a visit to the dog. Although it had been through hours of abdominal surgery the previous day, he was awake and stood up when we came in. He wagged his tail while Rick stroked his chin. Rick has an affinity for furry animals.

The remainder of the day was spent on further rat experiments, a planning meeting, and cleaning up the laboratory. And we were invited to a surprise party for a team member who had turned forty. Toward the end of the day Dr. Chen invited us to see what he had found with his electron microscope.

We all were very pleased. All of the components of the Islet Sheets were clearly visible: the islets, the core alginate, the coating alginate, and the mesh. In the micrograph below one can see two large islets and several small ones. The mesh is colored blue and is just under the alginate outer layer on the bottom of the Sheet. This photo and others show that the outer layers are perfect: not one islet or cell was exposed. (During processing for SEM the layers separate.)


On Thursday we returned to San Francisco, tired and very happy. No matter how many such experiments would be needed to demonstrate that the Islet Sheet was safe and effective, we had pulled off the first in Edmonton without mishap. Now comes the waiting to see how the dog does and what we learn.

Scott King
4 April 2001