While again this was written for a celiac aware audience, it does get into the various proteins in the grains that celiacs find toxic. Fits in with the recent discussion of grains versus the "gluten" grains. Don. "Genetics and What's New in Research" ------------------------------------- a talk by Martin F. Kagnoff, MD summarized by Jim Lyles Dr. Martin F. Kagnoff is the director of the Laboratory of Mucosal Immunology at the University of California at San Diego, and well-known for his research in celiac disease, particularly with respect to genetic factors. He gave a talk at the 1995 CSA/USA conference on October 5, 1995, in San Francisco, California. What follows are some highlights of Dr. Kagnoff's talk. Celiac Disease (CD) is associated with small bowel damage, which occurs when a celiac eats gluten containing foods. "Gluten" is something of a misnomer. Gluten really refers to the disease- activating proteins in wheat, and we know there are similar, related, but somewhat different proteins in other grains that activate CD as well. However, gluten-free (GF) has come to mean free of the grains which are toxic to celiacs. Nutrients are not absorbed properly in an untreated celiac, due to the damage in the small bowel. This results in a wide spectrum of different symptoms. When a celiac consumes gluten, the damage to the small intestine may be slight or it may be extensive. It depends on how sensitive that individual is to gluten, and on how much gluten is consumed. However, it is not just the small intestine that is sensitive to gluten; the entire digestive tract including the large bowel is sensitive. In a study done several years ago, wheat was placed at the very bottom of the small intestine, where celiac damage does not usually occur. This caused the type of damage that is characteristic of CD. More recently, Mike Marsh has inserted gluten into the rectum to help in diagnosing CD. He has found that inflammation and changes in the rectal lining occur in celiacs exposed to gluten in this fashion. Dr. Kagnoff showed slides with the two extremes: normal, healthy villi with small crypts and completely flattened villi with elongated crypts. He said that there are many in-between situations, where the villi are only partly gone or partly damaged. A healthy small intestine has many folds, with villi on the folds, and microvilli on the villi. Altogether, this provides a surface area equivalent to two regulation-size tennis courts; this is about 600 times as large as the surface area inside piece of tubing the same size and length as the small intestine. As the villi are damaged and the microvilli disappear in active CD, the absorptive surface is greatly reduced. The severity of the symptoms varies depending on the amount of absorptive surface that has been lost. Another point to consider is: What do you absorb in your small intestine, and where? One of the common symptoms of untreated CD is iron deficiency. Some celiacs had iron deficiency for years, but were not diagnosed until other symptoms began to show up as well. Dr. Kagnoff has a set of twins as patients that were diagnosed with iron deficiency at age seven, are short in stature, and wasn't until they were over 40 that they developed symptoms such as bloating and diarrhea which finally led to a diagnosis of CD. The reason this can happen is that iron is absorbed in the very top portion of the small bowel. If the villi damage is not severe and limited to that area, then other nutrients are absorbed further down in the intestine and only iron is malabsorbed. Another common problem with untreated celiacs is iron malabsorption coupled with calcium malabsorption. After malabsorbing calcium for years these people are susceptible to bone fractures, but really don't have that full-blown "picture" of CD which is the underlying cause of these fractures. As the small bowel lining becomes more damaged and abnormal, one starts to malabsorb fat-soluble vitamins such as D, E, A, and K. As the disease progresses, water soluble vitamins start malabsorbing as well. However, if the damage is limited to the first foot or so of the small intestine, full-blown malabsorption may never occur as the remaining 20 feet or so if small intestine will continue to absorb nutrients normally. However, in these cases iron absorption will be a continual problem as iron is only absorbed in the earliest portion of the small intestine. In CD genes, environmental factors, and the immune system all play a role. Dr. Kagnoff touched on all three of these factors. Environmental Factors --------------------- We all know that ingesting certain grains activates CD: wheat, rye, barley, and if taken in large enough quantities, oats. There is some debate in some groups as to whether or not oats are toxic to celiacs. We know that rice, corn, and sorghum are fine, as long as they are not contaminated by one of the toxic grains. When you look at the plant ancestry, you find that wheat, rye, and barley all come from a common ancestor. If you go up one more level, you find a point where oats also shares a common ancestor. These grains all have a high content of some alcohol-soluble proteins that are called prolamins. (The content is somewhat lower in oats.) These prolamins have a very high content of glutamine and proline, which are two amino acids. When we talk about a gluten-free (GF) diet, what we are really talking about are these alcohol-soluble proteins, which are named as follows: Grain Proteins ----- -------- wheat gliadins barley hordeins rye secalins oats avenins Note that we are not talking about a single protein. A variety of wheat may have 40 different gliadins encoded on multiple genes within the wheat. Some of the chromosome specialists years ago tried to engineer wheat that would lack the gliadins that active CD; but there were so many different genes encoding gliadins, on different chromosomes, that they soon realized it was an almost impossible feat. Within the wheat gliadins, there have been studies to determine which part(s) of the gliadin activate CD. Gliadins (and the corresponding proteins of the other three toxic grains) are proteins made up of many different building blocks that are called amino acids. We've found that only a small part of these proteins is needed to activate CD, about 12-15 of these amino acids; these are called gliadin peptides. Studies are being conducted all around the world, and some sort of consensus is being reached as to which peptide sequence activates a celiac response. One particular wheat gliadin (alpha-gliadin) has been studied in more detail. Don Kasarda, of the USDA research facility, and his group purified this protein, and others at the facility isolated the nucleotide and peptide sequences for alpha-gliadin. Other groups around the world have also worked with this single protein. What they've found is this protein has 266 amino acids, with over 60 glutamines and over 30 prolines. This is very unusual; most proteins have a fairly random scattering of amino acids. Tests with celiacs using alpha-gliadin have been conducted to isolate the actual amino acid sequence that triggers a celiac-type response. Groups in Norway and England have isolated the same sequence, which MAY be the part of alpha-gliadin that activates the disease. Genetic Factors --------------- Is CD a genetic disease? The answer is probably "yes"; susceptibility to CD is certainly genetic. Certain genetic factors are required; without them you don't get CD. However, even with them you might not get the disease, so CD is not entirely genetic. Within families of celiacs, depending on which studies you read, the incidence among other family members is between 2% and 15%. This is a relatively small number, but still far greater than the incidence in the population at large. What really points to a very strong genetic association is the incidence in monozygotic (identical) twins: Where one twin has CD, at least 70% of the time the other also has CD. The fact that this incidence is not 100% is another indication that there are other factors besides genetics involved in CD. Susceptibility to CD is associated with HLA genes encoded on the sixth chromosome. The HLA genes are among the most diverse set of genes encoded in humans or other mammals. They determine and govern why we are different from one another in terms of our immune system and how it reacts and responds. Celiacs nearly always have one of two HLA genes: DQ2 or DQ8. These genes are relatively common among Caucasians of European descent, occurring in about 25% of the population. These genes are not found in Japan or Africa among the blacks; consequently CD is virtually unheard of in these areas of the world. Most people with these HLA genes don't get CD, so there is still more to this puzzle than we currently know. About 95% of the celiac population carry the DQ2 gene, another 5% carry the DQ8 gene, and far less than 1% would carry anything else. If a celiac and a sibling share the same HLA genes, there is a 20-40% chance that the sibling will develop CD also. There are some studies going on which are trying to induce something similar to CD in animals. Dr. Kagnoff's group has just submitted one for publication in which they cloned DQ2 and other HLA genes and put them into mice. They are beginning to look at how the mice respond to gliadin. The Immune System ----------------- In CD, two types of T-cells come into play. The first type is intraepithelial T-lymphocytes, which exist between the epithelial cells. In active CD, one of the striking features is an increase in the number and density of these intraepithelial T-lymphocytes. These T-cells were thought to be responsible for the villi damage, but recent studies suggest that these cells actually help in the growth and development of epithelial cells. The increase in these T-cells during active CD may be an attempt to maintain the normalcy of the epithelial lining. In fact, when the gene responsible for these cells is deleted from mice, the epithelial lining is totally abnormal. The T-cells in the lamina propria are the ones responsible for the villi damage, by reacting to the presence of gliadin. During this reaction the T-cells release cytokines, and it is the cytokines which appear to cause the damage to the villi. Diagnostic Tests ---------------- CD can present with a broad array of symptoms. Often the symptoms are very subtle and appear to be far removed from the small intestine. This makes diagnosing CD difficult in many cases. The blood tests can be useful for screening. Three of the four antibody tests can be highly sensitive, but only when there is fairly marked damage to the villi. These tests generally do not come back positive when the damage is mild. Also, we have to take into account how often these tests give a false positive. For a while the endomysial antibody was felt to be nearly 100% specific, i.e., no false positives. In recent years that number has fallen off some, as there have been some false positives detected. The bottom line is: These tests are good as a screening device and to monitor compliance with the GF diet, but they cannot be used in place of the small bowel biopsy for diagnosing CD. Dr. Kagnoff does not believe these tests would be useful in screening the population at large; they are most useful in screening those in whom there is some suspicion of CD due to the symptoms. Next we need to look at HLA Class II DQ gene typing tests. These can be useful in eliminating the possibility of CD, as CD is virtually unheard of unless you have one of the two DQ markers we discussed earlier. Of course, the converse is not true: The vast majority of those which DO have one of these markers also don't have CD, so all you can say in that case is that CD remains a possibility. The HLA typing tests are also useful in determining for the siblings of a celiac if CD can be eliminated as a potential future concern. At this point Dr. Kagnoff answered some questions from the floor: Q: What is the risk of small bowel lymphoma? A: There is good (but not definitive) evidence that the increase in lymphoma is related to not being on a strict GF diet. For celiacs on a strict GF diet, the risk of small bowel lymphoma is not significantly greater than it is for non-celiacs. The question is, how much gluten is okay? Dr. Kagnoff has detected inflammation in biopsies of individuals who only eat a small amount of gluten. We know from other malignancies that ongoing inflammation is associated with increased risk. On the other hand, the overall risk is still very small, even in studies where people with active CD have eaten gluten for 30, 40, or 50 years. The risk is real, but the risk of an earthquake in California is probably higher. However, I still recommend that celiacs maintain as GF a diet as they can. Q: What is the risk of colon cancer in celiacs? A: There is no evidence that there is an increase risk of colon cancer in celiacs. The increase in risk, which is small, refers to cancers in the mouth, esophagus, oral pharynx, lymphoma of the small bowel, and cancer of the small intestine (which is very rare, even in celiacs). There is no increased risk elsewhere. Q: What is the possibility of developing a GF grain of wheat? A: Very unlikely. By the time you eliminated all of the gliadin-related genes from wheat, I'm not sure there would be much left. Q: Are you born with CD, or can it "develop" at any age? A: First it must be triggered by some event in the environment, such as a certain kind of flu, stress, etc. Then, once it is triggered, you continue to have the disease for the rest of your life.