One of the perennial topics of this blog is the synergy between vitamins A and D. A new Japanese study published last July in the journal Immunology Letters (1) provides further evidence of this synergy, this time suggesting the dynamic duo can courageously combat the most flagellant of our inner impulses, keeping our wayward neutrophils in check and barring them from wandering too far down the winding road that leads to autoimmunity.
These investigators faced a conundrum. A number of studies suggest that the activated hormone form of vitamin D, calcitriol, has great promise for preventing and treating autoimmune diseases, but its usefulness in the clinical setting is currently limited because it promotes excessive accumulation of calcium in the blood and soft tissues. One hardly wants to cure, say, psoriasis if it means having to pass kidney stones.
The investigators thus sought to see if the activated hormone form of vitamin A, retinoic acid, might also prove useful in battling autoimmune disorders. If so, using the activated forms of both vitamins together might allow clinicians to treat these disorders using doses low enough to avoid the nasty side effects.To address this issue, the investigators examined the effects of these vitamins on the development of Th17 cells and in a mouse model of contact hypersensitivity.
Vitamins A and D Synergistically Suppress the Development of Th17 Cells
Th17 cells are helper T cells that produce a number of inflammatory chemicals, including interleukin-17. These bad boys protect against infection, but they also appear to play a role in multiple sclerosis, rheumatoid arthritis, psoriasis, and inflammatory bowel disease. The investigators took “naive” T cells from mice that had been genetically engineered to react to egg white protein, and then incubated the cells with that very protein and with different doses of the activated vitamins.
We call these cells “naive” not to mock their credulity and lack of sophistication, but simply to indicate that because of their inexperience they have remained undecided and noncommittal about the courses their lives will ultimately take. Having yet to fall into that fateful encounter with the protein of their destiny, they have yet to even decide whether that protein is friend or foe. It is the molecular environment that will bode tidings of peace or of war. Depending on these tidings the naive T cell may on that fateful day become a brother to the protein, sheltering it from the reckless and wayward assaults of the more ignorant brethren, or may instead decide to sound the battle horn and launch an all-out crusade against the protein, ready to crush even its family and friends under the banner of victory. When the time comes, the decision proves all-engrossing: it is no less than the very decision to commit to immunological tolerance or to immunologicalintolerance.
The naive T cell puts its finger to the wind, so to speak, to distill the spirit of the times, and to discern whether an alliance with the protein would prove fruitful or folly. If fruitful, the T cell becomes aregulator or suppressor T cell, and arranges a treaty of peace. If folly, the T cell becomes a helper T cell, ready to route out the enemy. But the decision does not end there; war is much more complicated than friendship. The helper T must be assigned to a unit. Whether it becomes a Th1 cell, a Th2 cell, or a Th17 cell (and perhaps we will discover others) will determine just in what ways it will assist in the war effort, and just what type of collateral damage — allergies, autoimmunity — might ensue.
Even though these mice were genetically engineered to produce a great abundance of T cells that target egg white protein, there is nothing within their genes that makes them react with tolerance orintolerance, because this is not a matter of genetics. The investigators, then, had to incubate the naive cells not only with egg white protein and different amounts and combinations of activated vitamins, but also with a biochemical cocktail to mimic the environment that would occur within the body during a time of distress and that would convince these cells to commit to enlisting in the Th17 brigade.
Lo and behold, the activated forms of both vitamins suppressed the development of Th17 cells. The more of the activated vitamin there was, the fewer Th17 cells there were. We can see this below by noting that as we move rightward along the graph, the dose of the vitamin increases, and as we move downwards, the proportion of naive cells that joined the Th17 brigade falls.
The investigators then combined the two vitamins to see whether they would prove antagonistic, additive, or synergistic. To determine the type of interaction, they created the graph below, called an “isobologram.”
Each dot represents the dose required to cut the number of Th17 cells in half. The dose required of each vitamin alone is given a value of 1.0 to make the math simpler. These doses are represented by the dots in the upper left and bottom right corners. The dots that appear inside the square represent some combination of the two vitamins. If they were to fall along the diagonal line, this would mean that the two vitamins are additive. It would mean, for example, that you could replace half of one vitamin with the other and get the exact same effect. If they were to fall somewhere in the upper right half of the graph, this would mean that the two vitamins antagonize each other and that combining them increases the dose you would need rather than decreasing it. But as we see, the dots all clearly fall in the lower left half of the graph. This means that the two vitamins are synergistic. It means that if we combine the two, we can use much lower doses of both of them to get the same effect.The investigators performed a statistical calculation from this data and concluded that the effect represents “strong synergy.”
They performed similar though less detailed experiments with human cells and obtained similar results.
Vitamins A and D Protect Against Contact Hypersensitivity in Mice
The next question was whether the activated vitamins would protect against an autoimmune condition in live mice. The investigators addressed this question with a model of contact hypersensitivity. Contact hypersensitivity is similar to a skin allergy, except that T cells do the damage rather than antibody-producing cells. The method is rather simple: rub a nasty chemical known to produce the reaction onto the ears of the mice, and see how much swelling ensues. As we see below, activated vitamins A and D cooperated together to reduce ear swelling in these mice.
The first bar represents the amount of ear swelling that occurred in the absence of either vitamin. The second bar represents the effect of activated vitamin D, the third the effect of activated vitamin A, and the last the effect of the two vitamins together. It’s not obvious that this effect is synergistic rather than additive, but it’s clear that the best effect is seen with both vitamins together.Consistent with previous research showing that Th17 cells participate in this disorder, the protection afforded by vitamins A and D was associated with a reduced number of Th17 cells in the lymph nodes draining from the ears of the mice.
Putting the Synergy in Context
Although this paper is an important step forward in the exploration of the interactions between the fat-soluble vitamins, the authors seem unaware of the small but substantial body of literature that has already begun documenting these interactions. This leads them to reject the possibility of preventing or treating autoimmune diseases with nutrition.
The authors seem unaware, for example, of the numerous studies showing that the fat-soluble vitamins each protect against the toxicity of the other. They argue that retinoic acid (activated vitamin A) could make calcitriol (activated vitamin D) safe byreducing the dose necessary to achieve the desired effect. This is, indeed, an entirely logical argument for which they provided convincing preliminary evidence. But retinoic acid protects against the soft tissue calcification induced by calcitriol even without reducing the dose used, and it does this at least in part by normalizing the production of vitamin K-dependent proteins, which is thrown awry when calcitriol is used alone (2). I hypothesized that this would be true at the end of 2006 (3). Two years later, Tufts University showed it to be true in mice (2).
The authors concluded their paper by stating that fat-soluble vitamins are toxic. They suggested that using the activated hormones as drugs would provide a safer alternative to consuming the vitamins themselves:
Vitamin A and Vitamin D are fat-soluble vitamins, which have the potential of becoming toxic if they are chronically consumed in very high doses. ATRA [activated vitamin A] has been used for the treatment of acute promyelocytic leukemia, and retinoid is used for the treatment of skin cancers and psoriasis, but a high intake of vitamin A over a long period leads to chronic vitamin A toxicity including osteoporosis [63]. The combination therapy of 1,25D3 [activated vitamin D] with ATRA would reduce the risk for these side effects and obtain a favorable clinical response with a lower dose administration.
This paragraph is difficult to interpret because of its numerous apples-to-oranges comparisons, but it seems these authors are acutely aware that they need to use the hormone forms of the vitamins together to render them safe, but wholly unaware that it may be possible to do the same with a nutritional approach. Yet the same principle demonstrated in this paper with the hormone forms is true of the vitamins themselves: when consumed together, they make each other safe and effective. I have documented these interactions in a number of articles published over the last seven years:
- Does Vitamin A Cause Osteoporosis? (Winter 2005/Spring 2006)
- From Seafood to Sunshine: A New Understanding of Vitamin D Safety (Fall 2006)
- Vitamin D Toxicity Redefined: Vitamin K and the Molecular Mechanism (December, 2006)
- Vitamin D in the Infant: Requirements for Safety (Winter, 2006)
- On the Trail of the Elusive X-Factor: A 62-Year Mystery Finally Solved, Vitamin K2 Revealed (Spring, 2007)
- The Cod Liver Oil Debate (Spring, 2009)
- Eight related blog posts
The publication of papers in the pharmacological literature like the one reviewed herein from Immunology Letters represents an opportunity of enormous magnitude: we currently have molecular biologists working out the details of how the nuclear receptors for vitamins A and D interact, pharmacologists working out the details of how their hormone forms interact, and nutrition scientists beginning to study their dietary interactions. These fields are largely fragmented at the moment. With an interdisciplinary approach, the communication barrier between these fields can be broken down and we can arrive at a fundamentally new understanding of nutritional requirements and the potential for nutrition to prevent and treat disease.
The Raw Material of Communication
In this paper, we see that vitamins A and D cooperate to tame an aberrant response of the immune system. Th17 cells don’t just participate in autoimmune diseases, however; they also, as far as we currently know, protect against infectious diseases. As I wrote about in “The Cod Liver Oil Debate,” vitamins A and D protect against infectious disease. If it turns out in time that vitamins A and D protect against autoimmunity when given in their vitamin forms and not only in their hormone forms, it may begin to appear that they help direct the immune response toward appropriate targets, suppressing autoimmunity but boosting immunity against infections.
This may seem paradoxical, but should it? Vitamins A and D themselves are not hormones. They are the raw materials from which our cells make the signals they need to communicate. If this raw material is present in insufficient amounts, we could expect communication to go awry: diplomacy in its most miserable failings, our defenses spread thin, waging half-hearted wars against everything in sight, wholly inadequate to defeat our enemies and altogether too promiscuous to maintain any lasting friendships.
When the raw material necessary for communication is present in sufficient quantity, our immune systems mount strategic responses, defending the sacred soil when necessary, but minimizing collateral damage and opting for lasting peace whenever it is in reach.
This, at least, must be part of the puzzle. As our understanding of immunity grows in the new century, it must retain a prominent position for the fat-soluble vitamins.
References
1. Ikeda U, Wakita D, Ohkuri T, Chamoto K, Kitamura H, Iwakura Y, Nishimura T. 1alpha,25-Dihydroxyvitamin D3 and all-trans retinoic acid synergistically inhibit the differentiation and expansion of Th17 cells. Immunol Lett. 2010;134(1):7-16.
2. Fu X, Wang XD, Mernitz H, Wallin R, Shea MK, Booth SL. 9-cis retinoic acid reduces 1alpha,25-dihydroxycholecalciferol-induced renal calcification by altering vitamin K-dependent gamma-carboxylation of matrix gamma-carboxyglutamic acid protein in A/J male mice. J Nutr. 2008;138(12):2337-41.
3. Masterjohn C. Vitamin D toxicity redefined: vitamin K and the molecular mechanism. Med Hypotheses. 2007;68(5):1026-34. Epub 2006 Dec 4.
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