Comments on: Life Expectancy of Encapsulated Islets http://www.solvingdiabetes.org/2009/08/17/life-expectancy-of-encapsulated-islets/ A site dedicated to solving diabetes. Sat, 14 Aug 2010 23:12:33 +0000 http://wordpress.org/?v=2.9.2 hourly 1 By: Scott King http://www.solvingdiabetes.org/2009/08/17/life-expectancy-of-encapsulated-islets/comment-page-1/#comment-92 Scott King Mon, 17 Aug 2009 14:25:11 +0000 http://www.solvingdiabetes.org/?p=331#comment-92 In a healthy mammal nutrients are distributed through the body by the circulatory system. Blood has high levels of sugars, amino acids, fats, iron, proteins etc. In the fine capillaries these nutrients diffuse through the capillary walls into the interstitial spaces then to the hungry cells. Generally cells are less than 150 micrometers from a capillary. It follows that many cells are not touched by a capillary but are dependent on nutrients that may diffuse past several intermediate cells. Diffusion of nutrients through the Islet Sheet to the islets inside raises two different questions. First, can the nutrients diffuse the full distance effectively? And second, given that the Sheet is designed to keep immune effectors out, are any important nutrients accidentally excluded? The first question can be answered fairly precisely. The limiting nutrient in mammals (assuming nutrition is adequate for the whole animal) is oxygen. Cells become stressed within minutes of oxygen deprivation. Something like 40% of the blood is made up of specialized bodies that transport oxygen, red blood cells. Engineers at MIT have developed computer models that model the diffusion of oxygen through the body and the rate that cells consume oxygen; the concentration of oxygen in each cell is predicted by the model. These models can be used to probe subtle features, but the rule of thumb is pretty simple: the islets should be less than 150 microns from the surface of the capsule. That is why the Islet Sheet is about 300 microns thick. (And it explains why islets in 800 micron capsules die: they are starved for oxygen.) So we can be sure from diffusion of oxygen models that islets in the sheet behave almost as well as vascularize islets. The second issue is illustrated in the figure. (See above link for figure) This feature of capsule membranes is usually called the molecular weight cutoff, an unfortunate borrowing from electronics, where there are such things as bandpass filters with a sharp cutoff. But the analogous molecular filters do not exist in biology. The alginate gels used in encapsulation are mostly water, in fact > 95% water, and the alginate gel forms a random network resulting in tortuous paths of various sizes. A really small molecule like water or glucose diffuses randomly through the water essentially unimpeded. (Diffusion of glucose through the Islet Sheet is identical to diffusion through unstirred water.) Something very big like a cell does not get through at all. But as the molecular size grows the diffusion rate naturally drops, and the number of paths begins to decline (the paths are of random widths). There is no size that is absolutely excluded; it just takes longer and longer to get through. Our goal for the Islet Sheet is shown in the red line. The higher the permeability value the faster the diffusion through the Sheet. On the left are small nutrients including glucose and ions; these are unimpeded. Insulin is quite small at 7kD, and travels easily through the Sheet. Things get more interesting in the middle. One often reads that a successful capsule must completely exclude IgG antibody to prevent immune destruction of the islets. We don’t see it that way for a couple of reasons. First, IgG cannot damage cells; it functions as a ‘tag’ to identify target cells. So IgG needs a cell or serum complement to kill. Serum complement is a suite of proteins that kill cells by punching a hole in the cell membrane, and it is triggered by binding to 2 IgG molecules in close proximity. Moreover, because antibody bound to the cell surface is dynamically consumed by the process of ligand induced endocytosis, the chance of 2 IgG molecules being sufficiently close to mutually bind C1q (even if it were present) is infinitesimal if the diffusion of IgG is substantially modulated. On the right of the diagram (molecular weight 400kD) is C1q, an essential component of the efficient classic complement cascade. Without C1q IgG is benign. IgM on the other hand does not require complement to kill cells. Fortunately IgM is even larger than C1q so if C1q is excluded so will be IgM. So all we need do is keep C1q and IgM out. This is fortunate: just to the left of IgG (167kD) is transferrin (80 kD). Now iron is an essential nutrient. But iron is toxic, so is never un-chaperoned in the body. The protein that transports iron to cells is called transferrin (ferrous is Latin for iron). Because there is no membrane with a molecular cutoff, we could not allow transferrin in but not IgG. To summarize, the Islet Sheet let enough transferrin through to provide needed iron, but excludes C1q to prevent complement cell killing. Transferrin is not consumed so a substantial retardation of diffusion is acceptable. In a healthy mammal nutrients are distributed through the body by the circulatory system. Blood has high levels of sugars, amino acids, fats, iron, proteins etc. In the fine capillaries these nutrients diffuse through the capillary walls into the interstitial spaces then to the hungry cells. Generally cells are less than 150 micrometers from a capillary. It follows that many cells are not touched by a capillary but are dependent on nutrients that may diffuse past several intermediate cells.

Diffusion of nutrients through the Islet Sheet to the islets inside raises two different questions. First, can the nutrients diffuse the full distance effectively? And second, given that the Sheet is designed to keep immune effectors out, are any important nutrients accidentally excluded?

The first question can be answered fairly precisely. The limiting nutrient in mammals (assuming nutrition is adequate for the whole animal) is oxygen. Cells become stressed within minutes of oxygen deprivation. Something like 40% of the blood is made up of specialized bodies that transport oxygen, red blood cells. Engineers at MIT have developed computer models that model the diffusion of oxygen through the body and the rate that cells consume oxygen; the concentration of oxygen in each cell is predicted by the model. These models can be used to probe subtle features, but the rule of thumb is pretty simple: the islets should be less than 150 microns from the surface of the capsule. That is why the Islet Sheet is about 300 microns thick. (And it explains why islets in 800 micron capsules die: they are starved for oxygen.) So we can be sure from diffusion of oxygen models that islets in the sheet behave almost as well as vascularize islets.

The second issue is illustrated in the figure. (See above link for figure)

This feature of capsule membranes is usually called the molecular weight cutoff, an unfortunate borrowing from electronics, where there are such things as bandpass filters with a sharp cutoff. But the analogous molecular filters do not exist in biology. The alginate gels used in encapsulation are mostly water, in fact > 95% water, and the alginate gel forms a random network resulting in tortuous paths of various sizes. A really small molecule like water or glucose diffuses randomly through the water essentially unimpeded. (Diffusion of glucose through the Islet Sheet is identical to diffusion through unstirred water.) Something very big like a cell does not get through at all. But as the molecular size grows the diffusion rate naturally drops, and the number of paths begins to decline (the paths are of random widths). There is no size that is absolutely excluded; it just takes longer and longer to get through.

Our goal for the Islet Sheet is shown in the red line. The higher the permeability value the faster the diffusion through the Sheet. On the left are small nutrients including glucose and ions; these are unimpeded. Insulin is quite small at 7kD, and travels easily through the Sheet. Things get more interesting in the middle. One often reads that a successful capsule must completely exclude IgG antibody to prevent immune destruction of the islets. We don’t see it that way for a couple of reasons. First, IgG cannot damage cells; it functions as a ‘tag’ to identify target cells. So IgG needs a cell or serum complement to kill. Serum complement is a suite of proteins that kill cells by punching a hole in the cell membrane, and it is triggered by binding to 2 IgG molecules in close proximity. Moreover, because antibody bound to the cell surface is dynamically consumed by the process of ligand induced endocytosis, the chance of 2 IgG molecules being sufficiently close to mutually bind C1q (even if it were present) is infinitesimal if the diffusion of IgG is substantially modulated. On the right of the diagram (molecular weight 400kD) is C1q, an essential component of the efficient classic complement cascade. Without C1q IgG is benign. IgM on the other hand does not require complement to kill cells. Fortunately IgM is even larger than C1q so if C1q is excluded so will be IgM. So all we need do is keep C1q and IgM out. This is fortunate: just to the left of IgG (167kD) is transferrin (80 kD). Now iron is an essential nutrient. But iron is toxic, so is never un-chaperoned in the body. The protein that transports iron to cells is called transferrin (ferrous is Latin for iron). Because there is no membrane with a molecular cutoff, we could not allow transferrin in but not IgG. To summarize, the Islet Sheet let enough transferrin through to provide needed iron, but excludes C1q to prevent complement cell killing. Transferrin is not consumed so a substantial retardation of diffusion is acceptable.

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