Reductions in Serum Phosphate Induced by Supplementation With Free-Form Amino Acids: Interactions of Phosphate Homeostasis with Renal Ammoniagenesis

These articles [Heidland et al., 1978: (http://www.ajcn.org/cgi/reprint/31/10/1784.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/707333); Lamiell et al., 1990: (http://www.ncbi.nlm.nih.gov/pubmed/2108005)] show that supplementation with free-form amino acids, particularly in conjunction with a low- or "no-phosphate" diet (Lamiell et al., 1990), can fairly drastically reduce serum phosphate and also increase serum calcium. I've discussed some of these articles in past postings (http://hardcorephysiologyfun.blogspot.com/2009/02/potential-for-hypophosphatemia-or.html). Heidland et al. (1978) found that the serum calcium levels varied inversely with the serum phosphorus levels, in the people who had been given supplemental essential amino acids, and the authors suggested that that inverse relationship might have resulted from the increases in serum 1alpha,25-dihydroxyvitamin D3 (calcitriol, hormonal vitamin D, HVD) and the resorptive effect of an increase in serum HVD (?). These are really good articles, and I've never seen anyone mention that increase in "calcium mobilization from bone" (Heidland et al., 1978, p. 1791) in response to an increase in serum HVD. It's not widely-recognized, but there is a lot of research showing that increases in serum HVD can produce effects that are basically opposite to those of increases in autocrine or paracrine HVD (HVD that is formed in response to increases in extracellular 25-hydroxyvitamin D and that acts on nearby cells or in the same cell in which it is formed), and those paradoxical effects seem to show up more in relation to the calcemic or calcium-transport-modifying effects of HVD. For example, increases in HVD are known to be somewhat permissive with respect to soft-tissue calcification in animals, but increases in 25-hydroxyvitamin D, in the absence of concomitant increases in serum calcium (effects that are potentially more likely to occur in response to UV-induced increases in vitamin D than in response to oral vitamin D, in my opinion, given that oral vitamin D is likely to be more calcemic, etc.), have generally not increased soft tissue calcification and may have the potential to decrease soft-tissue calcification by, in theory, helping to prevent the osteoblastic differentiation of smooth muscle cells, etc. But combining high-dose vitamin D3 with high-dose calcium supplements has the potential to cause problems, and Heidland et al. (1978) suggested that the association of the amino acid supplementation with hypercalcemia, in some people, had been a result of the extra calcium supplementation (coupled with the relative absence of dietary phosphate).

A lot of these articles on these strategies for managing hyperparathyroidism and hyperphosphatemia in people who have kidney failure only look at serum phosphate or serum parathyroid hormone (PTH) levels and don't consider the levels of intracellular phosphate or the context in which the increases in PTH levels are occurring, and these are problematic aspects of a lot of these articles (I'm not talking about the ones I cited above). Lamiell et al. (1990) found, for example, that, after the second administration of the zero-phosphate parenteral nutrition formula was given to a person (they gave it and discontinued it a few times before they determined that hypophosphatemia or intracellular phosphate depletion, in addition to hyperammonemia, was causing the encephalopathy, and this is understandable), the encephalopathy occurred when the serum phosphate was normal. Lamiell et al. (1990) attributed that encephalopathic episode to hyperammonemia, and that's plausible, but it's important to remember that the intracellular inorganic and organic (i.e. ATP and ADP, phosphocreatine, etc.) phosphate levels can be significantly depleted in a person whose steady-state serum inorganic phosphate levels are normal [for example, Ambuhl et al., 1999: (http://www.ncbi.nlm.nih.gov/pubmed/10561144)]. Many most articles whose authors have measured the intracellular 2,3-diphosphoglycerate (2,3-DPG) concentrations in red blood cells have shown that have shown that those concentrations don't correlate at all with serum phosphate levels. I also think there's a danger in thinking that these derangements in calcium and phosphate homeostasis will only occur in people who have renal failure, but supplemental phosphate decreased urinary calcium excretion in normal people who had evidently not been exhibiting hypercalciuria [Heaney and Recker, 1987: (http://www.ncbi.nlm.nih.gov/pubmed/2851341)]. So some of these unspoken assumptions, such as the assumption that massive doses of calcium are going to not cause hypercalcemia or pathological effects in people who do not have kidney disease, in short-term trials with calcium supplements have the potential to be invalid.

The mechanisms by which free-form amino acids can decrease serum phosphate are not well-understood, but Heidland et al. (1978) noted that the persistence of the decreases in serum phosphate over the long term, in some people who are given supplemental amino acids, argues against the idea that the hypophosphatemic effect is a result of the "refeeding syndrome" or even something akin to it. I think it's caused by changes in the interactions of renal ammoniagenesis with the mechanisms governing phosphate reabsorption in the proximal tubules, and those interactions basically boil down to changes in acid-base homeostasis. For example, the acute, supplemental-glutamine-induced increases in serum bicarbonate that can occur in humans could be expected to favor an increase in phosphate transport into cells. There probably is an increase in phosphate uptake into cells, in response to some amino acids, but it's noteworthy that excessive or high doses of some mixtures of free-form amino acids seem to be more likely to produce hyperammonemia than protein does. Lamiell et al. (1990) attributed that to the absence of arginine in some mixtures, and that's conceivable. But it might be a result of the kinetics of the absorption of free-form amino acids. Their absorption is going to be much more rapid than the absorption of protein-derived amino acids, and that could overwhelm the liver's capacity for ureagenesis. It's also possible that increases in phosphate utilization (or loss of phosphate in the urine, induced by the amino acids) decreases the availability of intracellular inorganic and organic phosphate in a way that impairs the activities of hepatic urea cycle enzymes. But one possibility that seems plausible to me is that amino acids increase urinary phosphate loss and that the loss of phosphate increases renal ammoniagenesis to a degree that is significant enough to disturb the systemic acid-base homeostatic mechanisms and actually contribute to hyperammonemia. Under some circumstances, the overall urea cycle activity is thought to be an important factor in influencing systemic acid-base homeostasis, and Haussinger et al. (1990) [cited and discussed here: (http://hardcorephysiologyfun.blogspot.com/2009/02/urea-cycle-renal-glutaminase-activity.html); Haussinger et al., 1990: ([Haussinger et al., 1990: (http://www.springerlink.com/content/l2vx314521367706/)] basically found that even mild liver dysfunction was associated with a failure of renal ammoniagenesis to downregulate in response to systemic metabolic alkalosis. So, in a normal person, glutamine supplementation might acutely increase serum bicarbonate or only increase it on some days, such as on a day of intense exercise (in response to acidosis), but massive doses of glutamine or other essential amino acids that increase renal glutamine availability (glutaminase and the glutamine cycle are important in the regulation of renal ammoniagenesis, in response to metabolic acidosis) could contribute to the persistently-alkalotic state that can occur even in compensated liver disease, as discussed by Haussinger et al. (1990).

It's worth noting that alkalosis doesn't just increase phosphate uptake into cells (it tends to decrease intracellular phosphate availability in the long-term or even short term, because the uptake into cells provides phosphate to some cells at the expense of others, given the usual, alkalosis-induced reduction in serum phosphate) but impairs the unloading of oxygen from hemoglobin in a "2,3-DPG-depletion-independent" manner. That's one reason why bicarbonate administration can be so disastrous in a person who is hypophosphatemic, as discussed in past postings. One would think that the alkalosis-induced uptake (it's thought to partially be a result of the alkalosis-induced activation of glycolytic enzymes) of phosphate would increase 2,3-DPG levels, but it tends to not be the case in hypophosphatemia. It might be that more 2,3-DPG is driven into skeletal muscle myocytes or other cells that are not red blood cells, or it might be that the 2,3-DPG-independent "impairment" in the unloading of oxygen from hemoglobin tends to offset any potential for an increase in 2,3-DPG formation to occur in response to the alkalosis-induced increase in phosphate uptake by red blood cells. In any case, an inappropriate and persistent increase in renal ammoniagenesis could cause an ammonia-mediated impairment of TCA cycle activity in proximal tubule cells, given that ammonia excesses are known to inhibit TCA cycle enzymes and other mitochondrial enzymes and to interfere with energy metabolism by all sorts of mechanisms. That metabolic toxicity could reduce phosphate reabsorption by proximal tubule cells and exacerbate phosphaturia. Some of these articles by Ambuhl and colleagues (http://scholar.google.com/scholar?hl=en&q=ambuhl+phosphate) look promising as sources of information on those types of mechanisms.

Royal Rambler on Vitamin D, Hypercalciuria, Phosphate, and Fairly Boring "Minerals"

The authors of this article [Hathcock et al., 2007: (http://www.ajcn.org/cgi/content/full/85/1/6)(http://www.ncbi.nlm.nih.gov/pubmed/17209171?dopt=Abstract)] reviewed a number of clinical trials in which researchers had used dosages of vitamin D ranging from 2,000-100,000 IU/day of vitamin D3, and the research generally shows that hypercalciuria hardly ever occurs at dosages of vitamin D3 of 2,000-4,000 IU/day. Hypercalcemia hardly occurred at any of the reasonable dosages and didn't even occur at 100,000 IU/day. There probably isn't a dose of vitamin D3 that has zero potential to elevate urinary calcium, and the slight elevations in serum calcium that oral vitamin D3, in particular, tends to cause (in my opinion) have the potential to exacerbate thrombogenic conditions, as discussed in past postings (http://hardcorephysiologyfun.blogspot.com/2009/01/calcium-magnesium-serum-calcium-vitamin.html).

Sayer et al. (2004) [Sayer, 2004: (http://cs.portlandpress.com/cs/106/0549/1060549.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/15027893)] analyzed a lot of research on the mechanisms underlying nephrocalcinosis, which technically refers to an increase in the calcium contents of the kidneys but is generally taken to mean calcification, meaning the deposition of calcium phosphate (usually in the form of apatite), in parts of the kidneys, and Sayer et al. (2004) basically argued that hypercalciuria is the factor that seems to be associated with nephrocalcinosis. Even relatively small increases in the intake of phosphate have generally decreased the rate of urinary calcium excretion, and phosphate (from sodium or potassium phosphate) at something like 1,000 mg/day has been used to prevent kidney stones in recent decades. Then I guess everyone decided that phosphate is a "vice" or something and had to be removed from the diets of everyone. The research that I've discussed in recent postings indicates that taking very high dosages of phosphate (generally supplemental phosphate at greater than 70-100 mg/kg/day), along with hormonal vitamin D at dosages that cause intermittent hypercalcemia and hypercalciuria, creates the conditions that can cause nephrocalcinosis. But vitamin D itself generally does not cause either hypercalciuria or hypercalcemia, and there's some moderate dosage (i.e. 2,000-4,000 IU/day, as a conservative dosage) that could help keep the parathyroid hormone levels low but could do so without elevating urinary calcium unnecessarily. There's likely to be some middle ground, in my opinion.

One approach would be to use adequate dosages of magnesium (the research is really disorganized in magnesium, and that's the reason I keep avoiding citing all the articles I have on it), given that magnesium has the potential to help prevent nephrocalcinosis (http://scholar.google.com/scholar?hl=en&q=magnesium+nephrocalcinosis), in my opinion, and avoid calcium supplements and get calcium from milk or something like that. (There's some research claiming to show that magnesium decreases apatite formation but increases amorphous calcium phosphate formation, which is not really dangerous, but the research in actual animals generally shows that urinary magnesium wasting, from any number of causes, and decreases in magnesium status/intakes can increase the risk of calcification, in the kidneys and other sites.) I don't take supplemental calcium anymore and get enough calcium from milk that I use to "eat cereal." But anyway, the dosage range of magnesium is large, and I don't really feel like getting into a discussion of it and then have to "wade" through the research. The research on magnesium is not reader-friendly, really, and is surprisingly complex. Magnesium oxide (MgO) is absorbed relatively slowly, throughout much of the small intestine, and chelated magnesium aspartate (MgAsp) or MgAsp HCl (they're not the same) are absorbed more rapidly and are probably absorbed through either the amino acid or dipeptide transporters or by passive diffusion. Magnesium from magnesium oxide is likely to be absorbed by divalent metal transporters or by passive diffusion and the solvent drag mechanisms that enhance the absorption of some cations by passive diffusion. Some people report in the literature that the bioavailability of chelated MgAsp is basically too high for some people to be able to tolerate, and there might be something to that. If it enters the brain too rapidly, it can cause sort of drowsiness or mental sluggishness. The last time I tried chelated MgAsp, I experienced that sort of problem with it and went back to MgO. Some people can't tolerate MgO, however, at higher dosages, given the nausea and GI effects that it can cause. In that case, MgAsp might be preferable. Magnesium orotate would potentially be problematic, in my opinion, because of the orotate, as I discussed, indirectly, in one of my old papers (http://hardcorephysiologyfun.blogspot.com/2009/08/some-more-old-papers-of-mine.html).

I've never seen a report of true toxicity from magnesium at anything resembling a reasonable dosage (even 1500-2000 mg/day have been used in some research), and the occasional reports of hypermagnesemia have generally occurred in people taking 2500-3000 mg/day or more. The intravenous magnesium dosages used in various therapeutic contexts produce really high serum magnesium levels, but any excess is likely to mainly end up being excreted in the urine almost immediately. There could be some problems with fasting blood glucose levels decreasing in diabetics, in response to Mg supplementation, and it can transiently lower blood pressure and elevate serum potassium (and decrease urinary sodium reabsorption). The changes in sodium and potassium excretion might produce the acute decreases in blood pressure, but, supposedly, the low-level calcium channel blocking effect can reduce blood pressure a little bit. But the blood pressure changes seem to not really persist to as great a degree as one might think. In people who are hypertensive, there might be more of a lasting effect, but I don't know that I'd expect much in that area. The research is sort of mixed on that, but there can be a transient decrease in blood pressure that does not last longer than a day or two, after any increase in the dosage. Obviously, one would want to talk about these things with one's doctor. Magnesium, like many things, could cause side effects (such as electrolyte abnormalities, etc.) that could be detrimental, in conjunction with the effects of medications, for example, but that would not really constitute toxicity, in my opinion, at reasonable dosages. Even in the cases of supposed death by magnesium dosages of 15,000-30,000 mg per day, the person had been drinking the whole bottle of the OTC magnesium supplement for a long time, in one case. And it wasn't clear to me that the existing kidney failure, not the bizarrely high dosages of magnesium, hadn't actually been the cause of death. Magnesium could conceivably increase phosphate turnover or the dietary phosphate requirement, either by binding to phosphate in the GI tract, thereby potentially increasing the requirement, or by increasing the activities of glycolytic enzymes. Reporting on the research on magnesium is like pulling teeth, though, because of the variable qualities of the bioavailabilities and even percent absorption of different magnesium salts or chelated forms. The only other thing I can think of to say is that magnesium aspartate may not mean chelated magnesium aspartate, depending on the manufacturer's labeling practices. Chelated MgAsp is not the same thing as MgAsp as a salt. In MgAsp, the Asp(2-) and Mg2+ are ionized. In chelated MgAsp, the Mg is bound to the aspartate by coordinate covalent bonds. A chelate tends to be absorbed (a significant percentage of a dosage) intact and transported by dipeptide or amino acid transporters or by passive diffusion, but a salt (an ionic pair in crystalline form that dissociates upon entry into solution) dissociates in water.

I should mention that I don't take a multivitamin, because of all of the potentially problematic "ingredients" in it. The only "minerals" or metals I take in supplemental form are selenium, from sodium selenite, at about 150 ug/day, 150 ug of iodine (from potassium iodide), magnesium, and a low dose of molybdenum. Iodine can basically shut down thyroid function at dosages slightly higher than the RDA of 150 ug/day, especially some people, and selenium can also decrease thyroid hormone levels (and cause obesity or something and decrease serum IGF-1 levels, etc.) at dosages of even 200-300 ug/day, according to some research [(http://scholar.google.com/scholar?q=selenium+thyroid+adverse&hl=en); Hawkes et al., 2008: (http://www.ncbi.nlm.nih.gov/pubmed/18565425)]. I can't get the articles showing IGF-1 decreases to show up, but one has to ask oneself what the mechanism would be for that. It doesn't sound like a good effect to me and doesn't sound like a great way of supposedly preventing cancer, but the decreases in IGF-1 may (or may not) only occur at the higher dosage range. But overt selenium deficiency (it's probably uncommon) can also decrease thyroid hormone formation. The function of selenium, in my opinion, is to serve as a cofactor of thioredoxin reductase enzyme complexes, glutaredoxin reductase enzyme(s), glutathione peroxidase enzymes, etc. (and ribonucleotide reductase). It seems like it can have relatively significant effects on antioxidant enzymes, but I'd suggest watching out for those adverse effects. The dosage that causes those effects in one person might be lower than the dosage that causes them in another person. One approach would be to stick to the low end of the range or evaluate how much selenium one gets from his or her diet Manganese is abundant in foods and has been shown to cause psychiatric and neurological derangements, when given in excess, in supplemental form. So I definitely don't supplement with that. Copper and zinc are very problematic in supplemental form, in my view, and are abundant in foods, and I don't take those. And then there's all the potentially neurotoxic or "choroid-plexus-epithelial-cell-toxic" vitamin A and beta-carotene in some multivitamins. I take a small amount of iron protein succinylate, because I work out like a @#$%&%$ madman. That's an exaggeration, but it's a bare-bones approach. In any case, I can't advise anyone on these more boring topics, even, or on any other topics. One would want to talk with one's doctor about these things.

Oculocereborenal Syndrome of Lowe (OCRL): Potential Involvement of Disturbances in Megalin Fnxn's & Relevance to the Effects of Phosphate Depletion

(I meant to type oculocerebrorenal in the title, but my mind isn't working especially well this week, due to the intranasal flu vaccine.) This article [Ramanathan et al., 2009: (http://www.sajaa.co.za/index.php/sajaa/article/viewPDFInterstitial/387/428)] is maybe a poorly-chosen example of an article about Lowe syndrome [the "oculocerebrorenal syndrome of Lowe" (OCRL)], but there are plenty of articles on it (http://scholar.google.com/scholar?q=oculocerebrorenal+Lowe&hl=en). On the surface, the manifestations of OCRL look very similar to Fanconi's syndrome and to the manifestations of intracellular phosphate depletion, but, apparently, many people who have OCRL, caused by hypofunctionality of the OCRL protein(s), or who have a subtype of Dent's disease that is caused by mutations in the OCRL protein (the OCRL protein is an inositol polyphosphate 5-phosphatase that hydrolyzes mainly phosphatidylinositol 4,5-bisphosphate, or PtdIns(4,5)P2, into phosphatidylinositol 4-phosphate but also hydrolyzes other phosphatidylinositols, including phosphatidylinositol 3,4,5-triphosphate, or PtdIns(3,4,5)P3, into phosphatidylinositol 4,5-bisphosphate, etc.) do not display the phosphaturia or hypophosphatemia and rickets that characterize X-linked hypophosphatemic rickets and that occur or can occur in Fanconi's syndrome [Kleta, 2008: (http://www.ncbi.nlm.nih.gov/pubmed/18667737)]. But there's still a lot of overlap with the manifestations of intracellular phosphate depletion and with Fancon's syndrome and other causes of hypophosphatemia. There's hypercalciuria, proteinuria, hypercalciuria, etc., and the proximal tubules are relatively selectively affected, in terms of the effects of the disorder on the kidneys. OCRL also causes vacuolar changes in myelin that fall short of overt demyelination, and OCRL causes cataracts and glaucoma, etc. The encephalopathy and mental retardation in OCRL is seemingly more severe than the effects of hypophosphatemia, but I've cited articles in past postings showing that rather devastating central nervous system damage can result from hypophosphatemia. It's conceivable that intracellular phosphate depletion in neurons and astrocytes, in parts of the brain, is more common than is recognized and that the depletion of phosphate does, in fact, commonly cause vacuolar myelopathy (which, incidentally, is not a very specific neuropathological change and occurs in a variety of contexts, including vitamin B12 depletion, etc.). It's not all *that* generic, as a neuropathological manifestation, however, and OCRL is still similar to Fanconi's syndrome, in my opinion and the opinion of others [Erdmann et al., 2008: (http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=2025683)(http://www.ncbi.nlm.nih.gov/pubmed/17765681)]. It's not really useful to say that it's a totally different condition, when there's so much overlap of the manifestations.

My first thought was that the mutations in OCRL were causing intracellular phosphate depletion with something like intermittent hypophosphatemia, but the most direct manifestation of OCRL (the syndrome) is the aberrant accumulation of PtdIns(4,5)P2, primarily. One possibility is that, in humans who display significant intracellular phosphate depletion and develop proximal tubular acidosis and other changes that overlap with those found in OCRL or Dent's disease caused by mutations in the OCRL protein (Dent's disease can also result from mutations in chloride transporters), there's a generalized depletion of different phosphatidylinositols that, by some mechanism, leads to a similar skewing of the abundances of different phosphatidylinositols in favor of PtdIns(4,5)P2, as in OCRL. Or it's possible that, in OCRL, the impairment in the hydrolysis of PtdIns(4,5)P2 leads to a reduction in the availability of inorganic phosphate for use in ATP formation, much as the sequestration of phosphate in fructose 2,6-bisphosphate can cause ATP depletion following a fructose load. Or, maybe the formation or turnover of mutiple PtdIns's are upregulated in a way that sequesters inorganic phosphate, in multiple pools of PtdIns's, in a way that's maladaptive and detrimental to energy metabolism. Or, it may have nothing to do with energy metabolism.

But Erdmann et al. (2008) found, basically, that the accumulation of PtdIns(4,5)P2 in endocytic vesicles apparently deranged the trafficking of megalin to the apical membranes of proximal tubule epithelial cells. In any case, there can be abnormalities in the uptake of calcium and other constituents of tubular fluid and in receptor-mediated endocytosis by megalin in response to the conditions that occur in people who have OCRL mutations (and the OCRL syndrome) (Erdmann et al., 2008). Erdmann et al. (2008) mention that deranged megalin signalling could account for the CNS abnormalities and that patients with Dent's disease (even the forms due to mutations in chloride transporters) exhibit abnormalities in the functioning of megalin. Erdmann et al. (2008) didn't mention it, but megalin transports vitamin B12 bound to transcobalamin (http://scholar.google.com/scholar?hl=en&q=transcobalamin+megalin) and also transports vitamin D bound to vitamin D binding protein (http://scholar.google.com/scholar?hl=en&q=%22vitamin+D+binding+protein%22+megalin). Megalin serves a transport function across the blood-brain and blood-CSF barriers (http://scholar.google.com/scholar?hl=en&q=megalin+%22blood-brain%22+OR+%22blood-CSF%22), and the "spongy" changes in myelin or "pallor" of myelin (http://scholar.google.com/scholar?hl=en&q=myelin+OCRL+pallor+OR+spongy) seen in people who have OCRL hypofunctionality are reminiscent, in my mind, of the vacuolar myelopathy seen in subacute combined degeneration, due to vitamin B12 depletion (http://scholar.google.com/scholar?hl=en&q=vacuolar+myelopathy+B12), or in humans who have methionine adenosyltransferase deficiency, etc. [(http://scholar.google.com/scholar?hl=en&q=vacuolar+myelopathy+methionine+OR+%22S-adenosylmethionine%22+OR+%22S-adenosyl-L-methionine%22); (http://scholar.google.com/scholar?hl=en&q=myelin+deficiency+%22methionine+adenosyltransferase%22)].

It's interesting that cycloleucine, an inhibitor of methionine adenosyltransferase (MAT), the enzyme that synthesizes S-adenosylmethionine (SAM-e), causes "vacuolation" of myelin (http://scholar.google.com/scholar?hl=en&q=myelin+cycloleucine+vacuolation+OR+vacuolar). It's likely that "vacuolar myelopathy," which can sometimes be characterized by pathological changes in the myelin and also in oligodendrocytes or other cells [such as inclusion bodies in the nuclei of different cell types (http://scholar.google.com/scholar?hl=en&q=%22vacuolar+myelopathy%22+inclusion+body+vacuolation+OR+vacuolar)], is heterogeneous, but one interpretation would be to say that phosphate depletion can reduce SAM-e levels by reducing ATP and adenosine nucleotide pools in oligodendrocytes and other cell types. ATP depletion is known to be capable of causing SAM-e depletion [see either Morrison et al., 1997, or Eto et al., 2002, both of whom showed that SAM-e levels were decreased in the brains of people who had had Alzheimer's disease (the authors in at least one group were saying, correctly, in my view, that the SAM-e depletion was really likely to have been caused by ATP depletion): (http://scholar.google.com/scholar?hl=en&q=ATP+%22severely+decreased%22+Alzheimer%27s+%22S-adenosylmethionine%22)], and that, together with derangements in the abundance of PtdIns(4,5)P2 and other phosphatidylinositols (causing reduced vitamin B12 transport into the brain by reducing the megalin-mediated uptake of B12, etc.), could account for the web of associations I've discussed in this article. It's interesting that Reed et al. (2007) [Reed et al., 2007: (http://www.ncbi.nlm.nih.gov/pubmed/17392004)] found that cats that displayed low serum vitamin B12 and low serum folate levels also tended to display low serum phosphate levels (Reed et al., 2007). One could attribute that to any number of changes and say that the cats had some kind of Fanconi's syndrome that impaired reabsorption of folate binding protein, transcobalamin, and also inorganic phosphate from the tubular fluid. It's known that megalin knockout mice display low-molecular weight proteinuria (http://scholar.google.com/scholar?hl=en&q=megalin+proteinuria), as discussed by Erdmann et al. (2008), and lose different vitamins and other proteins in their urine, and megalin also transports folate binding protein (http://scholar.google.com/scholar?hl=en&q=megalin+folate+binding+protein). But phosphate depletion per se can cause metabolic acidosis or ATP depletion without acidosis in the proximal tubules and could, in my opinion, be a cause and consequence of proximal tubule pathologies. Here's another article that describes an association of B12 depletion with phosphate depletion and that could be explained by the fact that malabsorption, as in liver disease, can cause hypophosphatemia and cobalamin deficiency and also folate depletion [Wojtyczka, 1998: (http://cs.portlandpress.com/cs/095/0735/cs0950735.htm)(http://www.ncbi.nlm.nih.gov/pubmed/9831699)]. Those types of effects, such as loss of vitamin B12 and reduced folates and phosphate in the urine, could explain some of the post-infectious mono issues that people have [(http://scholar.google.com/scholar?hl=en&q=infectious+mono+nephritis+OR+tubular+OR+tubulointerstitial+OR+%22proximal+tubule%22); (http://scholar.google.com/scholar?hl=en&q=infectious+mono+adverse+OR+complication)]. It could be similar to the research showing that cerebral folate deficiency can result from expansion of the pools of antibodies that bind to the reduced folate carrier and other folate transporters at the blood-CSF barrier, given that the immune infiltration of the EBV-infected proximal tubule epithelial cells (http://scholar.google.com/scholar?hl=en&q=infectious+mono+EBV+%22proximal+tubule%22) could create a mess of immune-mediated impairments in proximal tubule functioning (such as by cytokine-mediated disturbances in energy metabolism, etc.). Supposedly, EBV doesn't infect choroid plexus epithelial cells, but I wouldn't be surprised if it did (http://scholar.google.com/scholar?hl=en&q=EBV+%22choroid+plexus%22). There are some significant problems with the notions that a lot of people have about the cell types that EBV supposedly can or can't infect. Here are some more searches [(http://scholar.google.com/scholar?hl=en&q=%22choroid+plexus%22+CD21+OR+C3d+OR+C3R); (http://scholar.google.com/scholar?hl=en&q=EBV+C3R+OR+C3d+OR+CD21)]. Everyone assumes that CD21 isn't likely to be expressed by cells in the CNS and that EBV must infect cells by binding to CD21, but what if it isn't true. A lot of viruses can infect cells using multiple transport mechanisms, some of which have only recently been discovered for influenza, for example. Also, there are significant problems with detecting EBV proteins during autopsies, and many articles look for EBV DNA or viremia (there's not going to be a bunch of viral DNA floating around, all over the place, in a cell latently-infected with EBV). EBV infects epithelial cells in basically every other organ, and it probably infects astrocytes and microglia (http://scholar.google.com/scholar?hl=en&q=EBV+infection+astrocyte+OR+%22human+monocytes%22) and pericytes (http://scholar.google.com/scholar?hl=en&q=resident+macrophage+pericyte+brain) and fibroblasts (i.e. meningeal fibroblasts, probably) [see Koide et al., 1997: (http://scholar.google.com/scholar?hl=en&q=EBV+fibroblasts)]. That type of effect on the proximal tubules could reduce phosphate and vitamin D and reduced folate and vitamin B12 reabsorption by the kidneys and could produce similar impairments at the blood-CSF barrier, etc. (http://scholar.google.com/scholar?hl=en&q=infectious+mono+brain+complication+OR+adverse).

I don't claim to be able to explain all of the different manifestations of these conditions, but the overlap of the effects of OCRL mutations with the effects of idiopathic Fanconi's syndrome and also the effects of intracellular phosphate depletion are fairly difficult to ignore completely. There must be some explanation, but it's interesting, in any case.

Hypercalciuria and Nephrocalcinosis as Presenting Signs/Symptoms in HHRH: Potential Relevance to Phosphate Homeostasis in Normal People

In this article [Bergwitz et al., 2006: (http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=1380228)(http://www.ncbi.nlm.nih.gov/pubmed/16358214)], Bergwitz et al. (2006) discussed the fact that nephrocalcinosis is often but not always a presenting symptom in hereditary hypophosphatemic rickets with hypercalciuria (HHRH). This is the third form of hypophosphatemia that I've read about and that results from mutations in nuclear DNA. In HHRH, there's hypofunctionality of one or another of the phosphate transporters that reabsorb filtered phosphate from the tubular fluid. The fact that nephrocalcinosis occurs before the people are ever treated with exogenous phosphate and occurs as a result of the phosphate depletion-induced elevations in calcitriol (1alpha,25-dihydroxyvitamin D, or 1,25-VD) provides some more indirect evidence that hypercalciuria is likely to contribute prominently to the nephrocalcinosis that can occur in people taking high doses of phosphate and hormonal vitamin D (or high doses of vitamin D) in combination [i.e. in X-linked hypophosphatemic rickets (XLHR) and autosomal dominant hypophosphatemic rickets]. Given that the physiological (endocrinological, etc.) conditions in HHRH are likely to, arguably, more closely mimic the conditions that would be present in a condition such as Fanconi syndrome (or even dietary phosphate depletion, to the extent that phosphate depletion could produce acidosis and mitochondrial damage in the proximal tubules), the research on HHRH suggests that the hypercalciuria that reliably accompanies significant dietary phosphate depletion (see past postings) (and the intracellular phosphate depletion that ultimately can result from it) could itself cause nephrocalcinosis.

In XLHR, part of the rationale for using calcitriol is to suppress the parathyroid hormone (PTH) release that can occur in response to the phosphate-induced decreases in serum calcium. In HHRH, there's no need to use calcitriol, because the serum calcium levels tend to be normal or elevated. And people who have HHRH display elevations in serum calcitriol levels, unless they're vitamin D deficient (severe but not mild vitamin D depletion decreases serum calcitriol, and vitamin D supplementation only increases serum calcitriol in people who are severely vitamin D deficient). The fact that normal vitamin D doesn't increase serum calcitriol past a certain point in normal people and can, at high dosages, actually decrease serum calcitriol (perhaps partly by increasing phosphate retention) is relevant to phosphate homeostasis in normal people. In my opinion, vitamin D (cholecalciferol, not calcitriol) at reasonable dosages (i.e. 2000-4000 IU/day or dosages that aren't high enough to potentially increase serum calcium and urinary calcium by much, if at all) could help to maintain low PTH levels in the context of reasonable increases in the dietary phosphate intake, in relation to the calcium intake. One reason it's desirable to maintain low serum PTH levels is that high PTH levels can increase urinary phosphate excretion and even, under extreme conditions, decrease the steady-state serum phosphate levels to a range that is below the baseline phosphate level (i.e. before the phosphate supplementation started). But that sort of derangement is more likely to occur, in my opinion, if one neglects magnesium homeostasis and doesn't take some reasonable but not massive amount of dietary calcium. I'm not much up for citing a bunch of articles today, but the point is that vitamin D can suppress PTH levels to a reasonable extent, much as calcitriol can [Barger-Lux et al., 1998: (http://scholar.google.com/scholar?hl=en&q=Barger-Lux+graded)], but can do so without the hypercalcemia and hypercalciuria that accompany calcitriol administration. Anyway, it's important to remember the potential that exists, in my view, for thrombogenic effects to result from even slight increases in serum calcium (such as in response to vitamin D, particularly orally-administered vitamin D) in people who have thrombogenic disorders or who are susceptible to that type of thing. Obviously, one would want to discuss this with one's doctor.

Phosphate Depletion Associated With Hypoxia, Autonomic Neuropathy, Hypoventilation, or Paralysis: Potential Relevance to Sudden Infant Death Syndrome

These [Siddiqui and Bertorini, 1998: (http://www.ncbi.nlm.nih.gov/pubmed/9572247); Gravelyn et al., 1988: (http://deepblue.lib.umich.edu/bitstream/2027.42/27325/1/0000348.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/3364446); Steckman et al., 2006: (http://www.ncbi.nlm.nih.gov/pubmed/16642427); Heames and Cope, 2006: (http://www.ncbi.nlm.nih.gov/pubmed/17090245)] are some interesting articles that show some of the variegated manifestations of hypophosphatemia. A crucial fact that I've taken from the research on phosphate homeostasis (it's arguably the most crucial point) is that neither the steady-state nor the between-dosage (in the context of phosphate infusions in animals or phosphate supplementation in humans) intracellular phosphate levels, in either muscle cells or red blood cells, correlates with the serum phosphate levels. For example, Chobanian et al. (1995) [Chobanian et al., 1995: (http://www.ncbi.nlm.nih.gov/pubmed/7900836)] found that the intracellular ATP concentrations in cells in the proximal tubules correlated positively with the intracellular inorganic phosphate (Pi) concentrations, and artificially-induced changes in extracellular Pi concentrations produced changes in the intracellular Pi concentrations. But in human studies, the intracellular Pi values generally do not correlate with serum Pi values, and the intracellular Pi concentrations can be significantly depleted in a person who has a normal serum Pi level.

That usual absence of a correlation between intracellular and serum Pi concentrations means, in my opinion, that intracellular phosphate depletion, in "normophosphatemic" people, should be considered as a possible factor contributing to some of these conditions that have been associated with hypophosphatemia. Siddiqui and Bertorini (1998) cited research showing that phosphate depletion can produce neuropathy that mimics Guillain-Barre syndrome, and the authors described the symptoms of a patient who developed neurological symptoms after she had been given parenteral nutrition without phosphate. The manifestations of neuropathy were suggestive of demyelinating polyneuropathy but were rapidly reversed by phosphate supplementation, meaning that there wasn't demyelination. The authors also discussed the fact that an increase in hexokinase activity, in response to insulin that has been released after the intake of carbohydrates, is thought to be an important factor that mediates the carbohydrate-induced increase in the transport of phosphate into cells and the decrease in serum phosphate that can result from that transport (Siddiqui and Bertorini, 1998). The authors also cited research showing cognitive dysfunction and encephalopathy in hypophosphatemic or (merely) intracellular-phosphate-depleted people (Siddiqui and Bertorini, 1998). One interpretation of the article by Steckman et al. (2006), in which gallstone-induced pancreatitis occurred in conjunction with hypophosphatemia and improved in response to phosphate administration, is that the phosphate depletion was causing neuropathy and interfering with gallbladder contractions. Neuropathy is known to be associated with gallbladder disease, and the normal functioning and contraction of the gallbladder is regulated by its autonomic (and sensory) innervation [(http://scholar.google.com/scholar?q=neuropathy+gallbladder+gallstone&hl=en);
the visceral sensory innervation can influence mast cell degranulation in the gallbladder, via the efferent-action-potential-mediated release of neuropeptides, and changes in mast cell degranulation and neuropeptide release can influence the autonomic regulation of gallbladder functioning, etc.: (http://scholar.google.com/scholar?hl=en&q=%22mast+cell%22+gallbladder+CGRP+OR+%22substance+P%22+OR+%22vasoactive+intestinal+peptide%22)]. Another interpretation would be to say that the phosphate depletion caused ATP depletion in the liver and led to cholestasis, etc. Similarly, the respiratory muscle weakness found in association with hypophosphatemia or low serum phosphate levels (Gravelyn et al., 1988, cited above) could be a result of autonomic dysfunction, particularly given that hypophosphatemia can cause reversible quadriparesis (paralysis, meaning the people are transiently quadripalegics) (http://scholar.google.com/scholar?hl=en&q=quadriparesis+hypophosphatemia). The hypoventilation that can accompany hypophosphatemia could also be due to autonomic neuropathy and ATP depletion in parts of the brain (http://scholar.google.com/scholar?hl=en&q=hypoventilation+hypophosphatemia). Hypophosphatemia has also shown up in association with extrapontine myelinolysis (which is central "pontine" myelinolysis that doesn't occur in the pons, essentially), one of the forms of osmotic demyelination that can result from the excessively-rapid correction of hyponatremia with intravenous, hypertonic saline [Qadir et al., 2005: (http://www.jpma.org.pk//PdfDownload/759.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/16045098)]. The authors suggested that ATP depletion in glial cells in parts of the brain might have contributed to the case, but I'm not sure that the authors actually said that the phosphate might have contributed to or caused the ATP depletion. The intracellular phosphate may well have been depleted in parts of the brain, and that depletion may have impaired volume regulation and predisposed to the osmotic demyelination.

In any case, I found this article showing "sinusoidal" seasonal changes in the incidence of sudden infant death syndrome (SIDS) (the seasonal change in the incidence shows up in the Southern and Northern hemispheres, and SIDS was found to peak in the winter in both hemispheres) [Douglas et al., 1996: (http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2351134)(http://www.ncbi.nlm.nih.gov/pubmed/8646093)], and there's old research suggesting an association of SIDS with vitamin D depletion or differences in vitamin D metabolism or rickets, etc. [Schluter, 1996: (http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=2352183&blobtype=pdf)(http://www.ncbi.nlm.nih.gov/pubmed/8842097); (http://scholar.google.com/scholar?hl=en&q=%22sudden+infant+death%22+%22vitamin+D%22)] (or with other light-associated changes, such as involving changes in melatonin levels induced by sleeping on the back as opposed to the side, etc.) (Douglas et al., 1996). There's also research showing that infants who were experiencing apnea were more likely to be hypercalcemic than infants not experiencing apnea [Kooh and Binet, 1990: (http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1452283&blobtype=pdf)(http://www.ncbi.nlm.nih.gov/pubmed/2207905)]. I couldn't get results to show up on a quick search, but hypercalcemia has been found to occur in hypophosphatemic people. Although Kooh and Binet (1990) didn't find that serum phosphate levels were associated with apnea in any way, the serum Pi levels wouldn't have to. Given that intracellular Pi levels do not reliably correlate with serum Pi levels and that phosphate depletion is known to be capable of causing respiratory paralysis/hypoventilation and hypoxia and neuropathy [see this and many others, some of which I discussed above: Weber et al., 2000: (http://www.ncbi.nlm.nih.gov/pubmed/10663486)] [and given that vitamin D depletion is known to be a cause of phosphate depletion (and that vitamin D supplementation, even in the absence of any genetic defect specifically involving vitamin D receptor signalling)], one possibility is that intracellular phosphate depletion in parts of the brain (and in the red blood cells, causing low-level hypoxia that might gradually have more severe consequences) could contribute to some cases of SIDS. Although there was one small study showing no apparent depletion of 25-hydroxyvitamin D levels in the context of SIDS, there could very easily be different degrees of intracellular phosphate depletion among infants with the same 25(OH)D levels. And looking at the serum phosphate levels wouldn't necessarily show anything, given the lack of correlation of intracellular and serum Pi levels. Someone would have to use MRS scans or look at the intracellular 2,3-DPG or Pi levels in red blood cells in infants, instead of just looking at the serum Pi. It's interesting that Heames and Cope (2006) (cited above) found that they could reduce the rate of infusion of noradrenaline in a manner that was proportional to the increase in serum phosphate, in a person who had developed transient heart failure from postsurgical phosphate depletion. The phosphate depletion basically caused hypotension, and the interactions with noradrenaline are really interesting (the usual thing people discuss is the fact that adrenergic drugs decrease serum phosphate by promoting phosphate uptake into cells). Given the changes in the autonomic regulation of blood pressure that occur in response to changes in the orientation of the body, such as in a baby sleeping prone vs. supine (http://scholar.google.com/scholar?q=autonomic+orthostatic+prone+supine&hl=en), it's possible that there's a kind of feed-forward depletion of intracellular phosphate in parts of the brain that can lead to apnea and then increased ventilation to compensate (and then phosphate depletion because of that and because of the noradrenaline released in response to that, as in the stress response to hypoxia, and to the potential vitamin D-depletion-induced renal phosphate wasting, etc.).

Arguably, the most well-established cause of hypophosphatemia is alkalosis induced by hyperventilation (http://scholar.google.com/scholar?q=hyperventilation+alkalosis+hypophosphatemia&hl=en), and apnea commonly occurs in response to post-hyperventilation alkalosis (http://scholar.google.com/scholar?q=hyperventilation+apnea&hl=en). So the alkalosis, in response to hyperventilation (as in response to autonomic dysfunction during sleep, resulting from changes in the sleep position and from phosphate depletion in neurons or smooth muscle cells or muscle cells in the diaphragm), could drive phosphate into cells outside the brain, thereby reducing phosphate availability to the brain, and then that could gradually set the stage for more severe episodes of hypoxia, more autonomic dysfunction due to the phosphate depletion in the brain, etc. There's evidence of repeated episodes of hypoxia in some research on SIDS [see Takashima et al. (1978) and Rognum et al. (1991): (http://scholar.google.com/scholar?hl=en&q=%22sudden+infant+death%22+hypoxia)]. A decrease in the responsiveness of smooth muscle cells (or other cell types, as in neurons in the brainstem, in the context of phosphate depletion) to noradrenaline occurs in people who have orthostatic hypotension and other derangements of baroreceptor functioning, and L-threo-3,4-dihydroxyphenylserine (DOPS) has been researched as a treatment for orthostatic hypotension and orthostatic tachycardia (DOPS is a precursor of noradrenaline) (http://scholar.google.com/scholar?hl=en&q=orthostatic+DOPS). Hypophosphatemia has been associated with instability in blood pressure, in association with postural hypotension and other problems with the sensitivity and functioning of the baroreceptor reflexes (http://scholar.google.com/scholar?hl=en&q=orthostatic+hypophosphatemia). Anyway, I just put those types of crude thoughts up on this blog.

Supplemental Phosphate (and Calcitriol) in Hereditary Forms of Hypophosphatemia: Potential Relevance to Phosphate Dosages & Responses in Normal Humans

This article [Reusz et al., 1990: (http://fetalneonatal.com/cgi/reprint/65/10/1125.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/2248503)], along with other articles that describe the adverse effects or absence of adverse effects of different dosages of supplemental phosphate in people who have X-linked hypophosphatemic rickets (XLHR) (or autosomal dominant hypophosphatemic rickets), are likely to be relevant to an understanding of the risks (or lack thereof) of phosphate supplementation in humans who don't have genetic disorders. Sitara et al. (2004) [Sitara et al., 2004: (http://www.geocities.com/razzaquems/MatrixBiology.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/15579309)] noted that the mechanisms underlying the hypophosphatemia in those two sets of genetic disorders are not perfectly understood, but the actions or serum levels of FGF-23 (fibroblast growth factor-23) are augmented in both sets of genetic disorders. The PHEX gene product is an endopeptidase, a protease enzyme, and mutations in that gene evidently are the root cause of XLHR and, among other phenotypic changes, serve to augment the actions of FGF-23 (http://scholar.google.com/scholar?q=phex+hypophosphatemia&hl=en). One could make the argument that the hyperphosphaturia in people who have those genetic disorders would cause those people to be at a lesser risk of developing ectopic calcification, as in response to any given dosage of supplemental phosphate. But I don't think that's true. Researchers have reported many cases of nephrocalcinosis, which is calcification of parts of the kidneys and would be the main risk of (particularly excessive) phosphate supplementation (in my opinion), in people with XLHR who have taken the combination of phosphate and hormonal vitamin D (HVD), which is calcitriol (1alpha,25-dihydroxyvitamin D3), that has been the standard therapeutic approach to treating the hypophosphatemia in those disorders. FGF-23, a protein that is "hyperfunctional" in these genetic forms of hypophosphatemia, decreases renal HVD formation and decreases phosphate reabsorption by proximal tubule epithelial cells. With regard to HVD formation, one could make the argument that the decreases in serum HVD, in many people who have these genetic disorders, would make the supplemental HVD less toxic than it would be in normal people, thereby confounding an attempt to sort through the risks of HVD vs. supplemental phosphate and to get a sense of the risks of different dosages of phosphate in normal people. But I don't think that's likely to be a valid reason for ignoring the data in some of these articles, either, because HVD seems to have been causing the same hypercalciuria and hypercalcemia in people with genetic hypophosphatemia as it tends to in normal humans.

The dosages of phosphate that have been associated with nephrocalcinosis in humans, as described by Reusz et al. (1990), are really high (a mean of 136.4 mg/kg bw/day, or 9548 mg/day, for a 70-kg human), and the "lower" range of dosages of phosphate (50-100 mg/kg bw/day, which is about 3500-7000 mg/day, or a mean of 69.9 mg/kg bw/day, which is 4893 mg/day) were not associated with nephrocalcinosis but were still quite high. Those dosages (more than 4000-5000 mg of phosphate/day, from any supplemental phosphate and food-derived phosphate, combined) are similar to the dosages that, for example, Heaney (2004) [Heaney, 2004: (http://www.mayoclinicproceedings.com/content/79/1/91.full.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/14708952)] was saying would potentially cause problems in humans. But almost no one ingests anywhere near those amounts of phosphate (which were, as discussed, not associated with nephrocalcinosis) per day, and a maximum of only 2300 mg/day of supplemental phosphate was required to treat people (who did not have genetic disorders) who displayed idiopathic (cause-unknown) phosphate depletion ("phosphate diabetes"). My point is that it's not a choice between the use of massive amounts of phosphate and the appalling consequences of the phosphate depletion that could occur, in the 21st century, here, in people who ingest only sources of "phytates," in whole grains and other vegetable- and plant-derived foods, that may provide little utilizable phosphate. There's a middle ground between the use of high doses of phosphate (and the state of blind terror, at the prospect of phosphate-induced nephrocalcinosis, that could go along with that) and the sense of "comfort in the majority viewpoint" that seems to potentially go along with phosphate deprivation and with the development of hypoxic brain injuries and osteomalacia and arthropathy (potentially, neuropathic, degenerative arthropathy/osteopathy) (http://scholar.google.com/scholar?hl=en&q=hypophosphatemia+osteopathy+OR+arthropathy) (it seems to me that the lower back pain and lumbar vertebral collapse/degeneration that characterize phosphate depletion are somewhat reminiscent of the neuropathic arthropathy seen in Charcot foot disease, for example, meaning that the symptoms and manifestations could be partially neuropathic in origin) that can result from intracellular phosphate depletion.

Also, Goodyer et al. (1987) [Goodyer et al., 1987: (http://www.ncbi.nlm.nih.gov/pubmed/2822887)] discussed the dosage range of HVD (40 ng/kg/day, or 2800 ng/day, for a 70-kg human) that had been associated with the development of nephrocalcinosis in people with XLHR or autosomal dominant hypophosphatemic rickets (ADHR), and researchers have generally used very high dosages of either vitamin D2, vitamin D3, and/or HVD in people who have had those disorders. Goodyer et al. (1987) supposedly found adverse effects associated with an intake of 4000 IU/day of vitamin D2 in people with XLHR or ADHR, but one wonders if, given all of the problems, reported in old articles, with vitamin D supplements containing ten times the labeled content of vitamin D, the dosage was actually higher. That dosage range (4000 IU/day) of vitamin D has not been reported to cause hypercalcemia in studies in normal humans. Another possibility is that the people were taking vitamin D and HVD and that the hypercalcemia was attributed to the vitamin D (as opposed to the HVD, which is the more likely culprit, in my opinion). I say that because I've never seen any case report in which a person with XLHR or ADHR was given, as a standalone treatment, only a low dosage of 4000 IU/day of vitamin D3 or vitamin D2. In most cases, the dosages have been massive, and hypercalciuria seems more likely to be a cause of the nephrocalcinosis than phosphate supplementation per se, in most of these people. Gross et al. (1998) [Gross et al., 1998: (http://www.ncbi.nlm.nih.gov/pubmed/9598513)] found that 2.5 ug HVD/day (2500 ng/day), in normal humans who had prostate cancer, caused hypercalciuria in everyone, at dosages ranging from 1500-2500 ng/day. Reisz et al. (1990) argued, despite the past research that had associated hypercalciuria with nephrocalcinosis and that they cited, that hypercalciuria had been associated more with the development of kidney stones than with the development of nephrocalcinosis, but, in most trials in people who have not had XLHR or ADHR, the participants have not taken both HVD and phosphate supplements, in massive dosages. The dosages of vitamin D (198-1370 IU/kg/day, or 13860-95900 IU/day) and HVD (5-35 ng/kg/day , or 350-2450 ng/day) are large and, perhaps not surprisingly, the people who displayed nephrocalcinosis had been the ones who had experienced multiple episodes of hypercalciuria or hypercalcemia. Nephrocalcinosis requires pathologically-increased concentrations of both calcium and phosphate, usually, to occur. Additionally, Seikaly et al. (1996) [Seikaly et al., 1996: (http://www.ncbi.nlm.nih.gov/pubmed/8545232)] found that nephrocalcinosis was more common in people who were taking HVD and phosphate and who had renal tubular acidosis. Metabolic acidosis, in the proximal tubule epithelial cells that reabsorb most of the phosphate from the tubular fluid, can cause urinary phosphate loss, but intracellular phosphate depletion can also be an important cause of metabolic acidosis. Thus, metabolic acidosis can be both a cause and a consequence of intracellular phosphate depletion, and it's important to remember these types of complexities. The insulin resistance and mitochondrial toxicity that can result from chronic phosphate depletion have the potential to actually increase the risk of calcification, because inorganic phosphate is constantly going to be "dumped" from its "storage" in intracellular phosphocreatine and adenosine nucleotide pools, etc. That's been suggested to be one mechanism for tissue-specific calcification in any number of disorders, in people who do not have XLHR. When there are these constant metabolic crises, such as can occur in response to intracellular phosphate depletion, there will tend to be these frequent, intermittent "episodes" in which phosphate is lost from cells or from its intracellular binding to organic compounds (such as creatine) and elevated, pathologically, in the extracellular or intracellular (cytosolic or mitochondrial) fluid. More specifically, for example, rhabdomyolysis is a fairly common result of intracellular phosphate depletion, and rhabdomyolysis can cause both wild elevations in serum phosphate (and, hence, phosphate concentrations in the kidneys, potentially contributing to nephrocalcinosis) and elevations in myoglobin and other proteins, released from necrotic muscle cells. Chronic rhabdomyolysis, such as in response to exercise in a person who is phosphate-depleted, has the potential to cause more kidney damage than phosphate supplementation ever would, and there are reports of people dying from rhabdomyolysis that was associated with (and likely to have been a consequence of, in my opinion) intracellular phosphate depletion in muscle cells and other cells (http://scholar.google.com/scholar?hl=en&q=hypophosphatemia+rhabdomyolysis+renal+failure).

Furthermore, "high" intakes of phosphate are well known to actually decrease urinary calcium excretion [Hegsted et al., 1981: (http://jn.nutrition.org/cgi/reprint/111/3/553.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/7205408); LaFlamme and Jowsey, 1972: (http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=292432&blobtype=pdf)(http://www.ncbi.nlm.nih.gov/pubmed/5080411)], and the high urinary calcium excretion that accompanies phosphate depletion is thought to be due, in large part, to the ongoing breakdown of hydroxyapatite in the bone tissue [Laroche et al., 1993: (http://www.ncbi.nlm.nih.gov/pubmed/8358977)]. Incidentally, I scaled those dosages of phosphate that were given to dogs and that were associated with calcification, in the study by LaFlamme and Jowsey (1972), and the equivalent human dosages would be massive (I did the calculations awhile ago, and it works out to 8000-some mg/day or more of phosphate). I honestly don't understand what the problem is with those types of dosage considerations in research in animals. The disregard for the physiological norms, in the context of dosages of so many nutrients or compounds that are given to animals, is significant, in my opinion, and is an ongoing issue in animal research.

Incidentally, Laroche et al. (1993) discussed the neuropsychiatric and pain-related manifestations of intracellular phosphate depletion and also found that people who had phosphate diabetes (intracellular phosphate depletion) displayed symptoms consistent with reflex sympathetic dystrophy. That's a condition that's mysterious and that causes bizarre, extreme pain and other symptoms. I don't have time to go into all of that, but it's basically more evidence that neuropathy and neurological damage can sometimes be one manifestation of phosphate depletion, in my opinion (and is evidence that the "back pain" or "bone pain" of phosphate depletion may be neuropathic in origin and may not have to do with bone problems per se, independent of the central nervous system).

It's also important to remember that increases in the phosphate intake could bind magnesium in the GI tract and produce adverse effects by that mechanism. Supplemental magnesium could conceivably reduce some of the supposed risk of increases in the dietary phosphate/(Ca+Mg) intake ratio, in my opinion, although I can't make any guarantees, at all, about safety in individuals or even in general. All I can offer is my sense of things. Even though supplemental magnesium increases phosphate reabsorption in animals and can decrease parathyroid hormone release (magnesium only increases PTH levels, up to a point, when a person has been grossly deficient in magnesium) [Thumfart et al., 2008: (http://www.ncbi.nlm.nih.gov/pubmed/18701629)], magnesium has been shown to reduce the incidence or extent of calcification in animals taking massive amounts of phosphate. I'll collect some of those articles, but the point is that the use of supplemental magnesium is worthwhile, in my opinion, and is likely to be especially worthwhile in the context of an increase in the phosphate intake, from food or another source, in relation to the intakes of magnesium and calcium, etc. That said, one would want to monitor one's electrolytes and discuss these issues with one's doctor. Magnesium can elevate serum potassium and cause natriuresis (an increase in urinary sodium excretion) at high dosages, even though a high-magnesium diet (Thumfart et al., 2008) decreased urinary sodium loss in animals in that article.

Vitamin D and Phosphate: Relevance to Muscle Weakness in Vitamin D Depletion & Significance of Lipoprotein Binding of Orally-Administered Vitamin D

Chudley et al. (1981) [Chudley et al., 1981: (http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1862641&blobtype=pdf)(http://www.ncbi.nlm.nih.gov/pubmed/6793223)] noted that muscle weakness and low-level indications of peripheral neuropathy (and overt neuropathy, but the authors don't really discuss that) can occur in people who are hypophosphatemic, and it's interesting that researchers have commonly found muscle weakness and balance problems (predisposing to falls in elderly people) and muscle pain to be associated with vitamin D deficiency (given that vitamin D supplementation is used to treat hypophosphatemia due to a number of different causes). There's actually evidence that the phosphate depletion that results (in part) from the secondary hyperparathyroidism that, in turn, results from vitamin D deficiency does contribute to muscle weakness and the associated problems (muscle pain, balance problems) (http://scholar.google.com/scholar?q=phosphate+%22vitamin+D%22+muscle+weakness&hl=en). It actually seems likely that the muscle weakness could partly be due to neuropathy induced, in part, to the phosphate depletion that accompanies vitamin D deficiency. I should mention that I don't think vitamin D (or rather the elevations in 25-hydroxyvitamin D) produced in response to UVB is as calcemic as oral vitamin D is, and I base that statement on various lines of evidence. I think it might be that oral vitamin D is converted into 25-hydroxyvitamin D and then into 1alpha,25-dihydroxyvitamin D locally, in enterocytes in the GI tract (the cytochrome P450 enzyme(s) that display(s) vitamin D 25-hydroxylase activity is/are expressed in a wide variety of tissues), or it may be a result of the fact that the oral vitamin D becomes more highly bound to lipoproteins [Haddad et al., 1993: (http://www.pubmedcentral.nih.gov/picrender.fcgi?pmid=8390483&blobtype=pdf)(http://www.ncbi.nlm.nih.gov/pubmed/8390483)]. That could cause its transport into cells to be regulated in different ways, etc. That article by Haddad et al. (1993) is great, and they say that vitamin D3 produced in the skin (and not just 25-hydroxyvitamin D produced in the liver) is primarily bound to vitamin D binding protein (Gc globulin), but orally-administered vitamin D becomes bound to chylomicrons and lipoproteins and enters the liver much more rapidly than skin-derived vitamin D. The issue wouldn't just be the rate of transport into the liver, though, because the lipoprotein-bound vitamin D and 25-hydroxyvitamin D would enter cells in ways that would not be subject to the same endosomal regulatory mechanisms, etc., as the transport of vitamin-D-binding-protein-bound vitamin D or 25-hydroxyvitamin D would be subject to.

Regardless of the mechanisms, there does seems to be some difference that, in my opinion, makes oral vitamin D more calcemic, and that could be important in the context of some of these issues related to the effects of vitamin D repletion on phosphate homeostasis. I remember reading an old article in which someone suggested that some of the neuroprotective effects of vitamin D, in the context of in vitro or animal experiments, might be mediated by its effects on phosphate homeostasis (on the preservation of ATP, as a result of its phosphate-"sparing" effects and effects on phosphate transport, etc.), but I forget what the article was specifically testing (and I can't find it right now). There would be a limit to the supposed beneficial effects of vitamin D repletion on phosphate homeostasis, and the concomitant elevations in serum calcium could, to some extent, negate the benefits associated with the vitamin D-mediated reductions in urinary phosphate loss (an effect that is, in part, secondary to the localized conversion of 25-hydroxyvitamin D into hormonal vitamin D, in the parathyroid tissue, and autocrine/paracrine suppression of parathyroid hormone release, etc.).

A Glimmer of Clarity and Understanding About the Significance of Serum Alkaline Phosphatase, in Relation to Vitamin B6 and Vitamin D

This is a complex and difficult-to-understand area of research, for various reasons, but some of these articles have helped me to sort of understand the alkaline phosphatase issue. The authors of this article [Lomashvili et al., 2004: (http://jasn.asnjournals.org/cgi/content/full/15/6/1392)(http://www.ncbi.nlm.nih.gov/pubmed/15153550?dopt=Abstract)] discuss evidence that elevated alkaline phosphatase activity, either on the plasma membranes of smooth muscle cells or in soluble form, apparently, in the plasma, hydrolyzes pyrophosphate (P03-O-PO3)(4-) (PPi) and thereby prevents the inhibition of vascular calcification that PPi is thought to confer, even in the presence of elevated levels of free calcium Ca2+ and inorganic PO4(3-) (PO4). Vitamin B6 repletion/supplementation and vitamin D3 repletion generally decrease serum alkaline phosphatase (ALP) and may reduce calcification, in part, by those mechanisms (to the extent that reductions in free ALP activity reflect some changes at the sites of calcification). Matias et al. (2009) [Matias et al., 2009: (http://www.ncbi.nlm.nih.gov/pubmed/18775809?dopt=Abstract)], for example, found that 25-hydroxyvitamin D concentrations correlated inversely with the extents of vascular calcification among patients with renal failure. The PPI is thought to bind to sites of existing calcification and block PO4 from binding and forming more hydroxyapatite crystals. I cited some of the articles showing the vitamin B6 associations and mechanisms, in relation to ALP, in some past postings [(http://hardcorephysiologyfun.blogspot.com/2009/01/another-article-mentioning-plp-in.html); (http://hardcorephysiologyfun.blogspot.com/2009/01/pyridoxine-calcium-channels-and.html)].

This is a really confusing area of research, and it's still not clear to me what the origin of serum ALP is. Supposedly serum ALP decreases as bone turnover decreases, and Regidor et al. (2008) [Regidor et al., 2008: (http://www.asn-online.org/press/pdf/2008-Media/Kalantar-Zadeh-Bone%20Disease%20Study.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/18667733)] discussed the fact that extreme and pathological decreases in bone turnover (such as might result from some of the osteoclast-and-monocyte-macrophage-lineage-cell-cytotoxic approaches to treating bone demineralization) can decrease serum ALP to pathologically low levels. I don't think those kinds of decreases would occur in response to reasonable dosages of supplemental vitamin D, but the increases in serum calcium that tend to result from either excessive vitamin D or calcium supplementation could, in my opinion, promote thrombogenic effects (hypercoagulability, etc.) (http://hardcorephysiologyfun.blogspot.com/2009/01/calcium-magnesium-serum-calcium-vitamin.html). The article by Seelig (1990) is especially good, and here are two articles that discuss those issues and that are cited in that old posting [Ruttmann et al., 2007:(http://www.anesthesiaanalgesia.org/cgi/content/full/104/6/1475) (http://www.ncbi.nlm.nih.gov/pubmed/17513645); Seelig, 1990: (http://www.ncbi.nlm.nih.gov/pubmed/2132751)]. Supposedly the actual serum ALP activity is partially a result of ALP, which acts extracellularly, on the plasma membranes of neutrophils, etc. I just read through an editorial and a couple of articles on ALP and calcification, and the researchers talk about the way no one really understands research on ALP. They don't say that, but they discuss the fact that it's not entirely clear if it's active on neutrophils or other cell types and then cleaved into a soluble form or what is even going on. It sounds like serum ALP is sort of like serum soluble transferrin receptor (sTfR), in the sense that serum ALP is normally produced, released upon cleavage of the membrane-anchored form during apoptosis (?), at some rate that correlates with the rate of turnover of osteoclasts and osteoblasts. But it can be elevated in cholestatic liver disease also (see 2nd old posting on vitamin B6 and ALP), etc. I guess this one article that I don't have time to cite right now says that serum ALP is enzymatically active but doesn't contribute to PPi cleavage. The authors also say that serum ALP don't correlate with serum PPi and that it's mainly the ALP expressed by smooth muscle cells that hydrolyzes PPi locally and is thought to thereby contribute to calcification (by forming PO4 locally, from PPi). The authors say no one knows why vascular calcification nonetheless seems to correlate with changes in bone turnover. I'm not sure what the correlation is that they're referring to, but presumably they're saying that increases in calcification accompany extremely low levels of bone turnover (presumably as a result of localized decreases in extracellular PPi at sites of vascular calcification, if one accepts the validity of these associations and mechanisms). They're saying they don't know how the expression and activity of smooth-muscle-cell ALP could be changed in association with changes in osteoblast apoptosis (and with the associated changes in serum ALP that do not contribute significantly to the localized cleavage of PPi, on the plasma membranes of the smooth muscle cells).

The research on vitamin B6 and ALP is just really confusing to everyone who reads it, seemingly, and to me. I can understand the basics--that plasma-membrane ALP cleaves albumin-bound pyridoxal-5'-phosphate (PLP) into pyridoxal, which then enters cells and is rephosphorylated, to PLP, by pyridoxal kinase. Humans and animals that have genetic mutations that decrease the activities of one or more of their alkaline phosphatase isoforms apparently have elevated serum PLP levels but have functional B6 deficiency, because ALP is required for the uptake of pyridoxal into cells (as discussed above). But it's not at all clear to me what the mechanism is by which an increase in B6 intake would decrease serum ALP. It's not an especially enjoyable topic to read about. I also think that excesses of B6 could produce neuropathy, in part, by decreasing ALP excessively, but that's just my opinion. And excesses of vitamin D intake could promote vascular calcification by elevating serum calcium, given that the traditional focus on the [Ca] x [PO4] product, as a factor whose elevation is associated with an increase in the extent calcification, shouldn't just be completely discounted or ignored. Everyone still seems to think taking a lot of calcium supplementation, beyond the RDA or whatever, is a good idea. I don't think it is, and I think people don't realize the manifold factors that can prevent the absorption or retention of dietary magnesium. I've discussed some of them in past postings and have discussed some of the dosage ranges that have been applied to extreme states, such as liver disease, in which magnesium absorption can become compromised. The dosage range for magnesium is fairly large, and I don't know what the right dosage is. That's the type of thing one would want to discuss with one's doctor.

Neuroprotective and Supposed Antidepressant-Like Effects of Sodium Butyrate: Relevance to HMB Research and Energy Metabolism

A lot of these articles showing that butyrate (usually administered or used in vitro as sodium butyrate, or SB), a short-chain fatty acid similar in structure to HMB (3-hydroxy-3-methylbutyrate or 3-hydroxyisovalerate, discussed in the two previous postings), reduces the degradation of numerous proteins by proteasomes are relevant to research on HMB. There are many similarities among the effects of butyrate and HMB. HMB is thought to exert its anticatabolic effects by inhibiting proteasomal activity (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=proteasome+methylbutyrate+OR+%223-hydroxyisovalerate%22) and also by acting as a precursor of HMG-CoA and of cholesterol. The extent to which an HMB-induced increase in cholesterol formation contributes to the HMB-induced inhibition of proteasomal activity is unknown. SB is a nonselective inhibitor of histone deacetylase enzymes in vitro, and its histone deacetylase inhibitory effect, at least in vitro, is thought to contribute to its inhibition of TNF-alpha-induced NFkappaB (NFkB) transcription factor [a.k.a. the "Rel" family of subunits that form the dimers that comprise NFkB transcription factors: (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=RelA+RelB)] activation in the cytosol (Yin et al., 2001: (http://www.jbc.org/cgi/reprint/276/48/44641)(http://www.ncbi.nlm.nih.gov/pubmed/11572859?dopt=Abstract); (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=%22sodium+butyrate%22+proteasome)]. Butyrate doesn't prevent the ubiquitination of IkB proteins (inhibitors of NFkB activation) but causes them to acccumulate as ubiquitin-conjugated proteins, without being degraded in proteasomes, evidently (Yin et al., 2001). HMB is also thought to exert anti-inflammatory effects by suppressing NFkB activation, as a result of the HMB-induced suppression of "proteasomal activity" [Baxter et al., 2005: (http://www.ncbi.nlm.nih.gov/pubmed/16006030)].

I don't doubt that some of those mechanisms are important, and a decrease in the activation of NFkB transcription factors can be antiproliferative and can downregulate the expression of numerous pro-inflammatory cytokines (cytokines that suppress mitochondrial functioning), etc., but SB is produced by microorganisms in the GI tract and is known to be the major energy substrate for colonocytes in the submucosal layers (I forget the terminology) of the colon (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=%22sodium+butyrate%22+energy). The in vitro research probably uses bizarre conditions and shows that SB can induce apoptosis of colon cancer cells. It looks like SB is pro-apoptotic at high but not low concentrations (0.5 mM to 2 mM) [Singh et al., 1997: (http://carcin.oxfordjournals.org/cgi/reprint/18/6/1265.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/9214612)]. But SB supposedly doesn't produce very strong histone deacetylase inhibition in the brain in vivo in animals, but it does produce neuroprotective effects in all sorts of different models (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=%22sodium+butyrate%22+neurodegenerative+OR+neurological+OR+neuroprotective+OR+Parkinson%27s+OR+Huntington%27s+OR+ischemia+OR+ischaemia+OR+hypoxia+OR+anoxia). Sodium butyrate has also produced some "antidepressant-like" effects in animal models of depression (http://scholar.google.com/scholar?q=%22sodium+butyrate%22+antidepressant&hl=en&lr=). Sodium butyrate is also sold as a supplement (http://www.google.com/products?q=sodium+butyrate&hl=en&aq=f).

Anyway, I just put this information up here, but I have no idea what the dosage range would be. One would obviously want to discuss this type of thing with one's doctor, and the most obvious, potential problem would be the disturbances in phosphate or calcium homeostasis in response to something like this. The infusion of 3-hydroxybutyrate, a "ketone" that doesn't have a carbonyl group but is defined as being a ketone, and acetate, for example, can increase plasma bicarbonate, and this effect appears to be the result of the metabolism of the organic acids/fatty acids and not from effects on phosphate homeostasis, in some articles. But these organic anions can just have strange effects, and it's something to be aware. Many medications can affect acid-base homeostasis and could interact with sodium butyrate or HMB. Some anticonvulsants act as carbonic anhydrase inhibitors, for example, and could interact with these types of short-chain fatty acids (such as sodium butyrate) or branched-chain organic acids/fatty acids (such as HMB).

In my opinion, sodium butyrate probably acts mostly as an energy substrate, but that doesn't exclude other mechanisms. I also think the research on sodium butyrate is likely to be relevant to future research on the mechanisms of action of HMB. Both compounds inhibit proteasomal activity and may have overlapping or similar effects, but I don't think it's going to be as simple as testing HMB as a "histone deacetylase inhibitor." Histone acetylation is extraordinarily complex and dynamic, and to think that one can treat a multitude of conditions with histone deacetylase inhibitors is not realistic, in my opinion. Vitamin D receptor activation can increase histone acetylation (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=%22vitamin+D%22+histone+acetyltransferase+OR+acetylation), much as sodium butyrate supposedly does (histone deacetylase inhibition leads to increases in the acetylation of histone proteins). But it's clear, in my opinion, that a lot of the effects of sodium butyrate cannot be explained in terms of histone acetylation.

Epstein-Barr Virus Infection of Astrocytes and Monocytes: Potential Relevance to Research on Multiple Sclerosis and Astrocyte Cell Cycle Re-Entry

These articles [Chaudhuri, 2005: (http://www.ncbi.nlm.nih.gov/pubmed/15617877); Behan et al., 2002: (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=Behan+Chaudhuri+Roep+%22THE+PATHOGENESIS+OF+MULTIPLE+SCLEROSIS+REVISITED%22); VanAmerongen et al., 2004: (http://www.direct-ms.org/pdf/VitDMS/VanAmerongenVitDMSreview.pdf)(http://www.ncbi.nlm.nih.gov/pubmed/15054436); Cepok et al., 2005: (http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1077174)(http://www.ncbi.nlm.nih.gov/pubmed/15841210); Diesel et al., 2005: (http://clincancerres.aacrjournals.org/cgi/content/full/11/15/5370)(http://www.ncbi.nlm.nih.gov/pubmed/16061850); Sanders et al., 1996: (http://www.ncbi.nlm.nih.gov/pubmed/8799216); Prokova et al., 2002: (http://www.jbc.org/cgi/content/full/277/11/9342)(http://www.ncbi.nlm.nih.gov/pubmed/11781310?dopt=Abstract); Koch et al., 2006: (http://www.ncbi.nlm.nih.gov/pubmed/17050217)] are really good, and Chaudhuri (2005) suggests that vitamin D repletion during brain development may protect against abnormal astrocyte apoptosis later in life and thereby confer protection against multiple sclerosis. This is interesting and is similar to the vitamin D hypothesis of schizophrenia [McGrath and colleagues: (http://scholar.google.com/scholar?q=%22vitamin+D%22+schizophrenia&hl=en&lr=)], in the sense that there's this concept of vitamin D deficiency, during development, creating abnormalities in brain development that do not manifest themselves until relatively later in life than one might expect them to. For example, vitamin D depletion during brain development drastically decreases the expression and protein content of the low-affinity neurotrophin receptor (p75NTR), which binds all of the neurotrophins and plays crucial roles in the regulation of not only apoptosis or protection against apoptosis, by NGF and other neurotrophins (NT-3, NT-4, BDNF, etc.), but in the trophic effects of NGF in the adult brain.

Holmoy (2008) suggested that vitamin D repletion could protect against brain damage due to late Epstein-Barr Virus (EBV) infection (i.e. after early childhood, when infection is often asymptomatic or less destructive to the brain), which tends to produce an expansion of autoreactive T-cell populations [Holmoy, 2008: (http://www.ncbi.nlm.nih.gov/pubmed/17574770)]. There's actually research showing that EBV can infect astrocytes [Menet et al., 1999: (http://www.ncbi.nlm.nih.gov/pubmed/10438862)] and monocytes and other cells of the monocyte-macrophage lineage [Savard et al., 2000: (http://www.ncbi.nlm.nih.gov/pubmed/10684275); (http://scholar.google.com/scholar?num=100&hl=en&lr=&cites=16392233215499070431)], which means that EBV may very well infect microglia and perivascular macrophages, etc. There still seems to be a popular sentiment that EBV only infects B-cells and epithelial cells, but there is overwhelming evidence that this is not the case and that EBV infects cells in the brain en masse during infectious mononucleosis (the term mononucleosis refers to the characteristic finding of monouclear phagocyte, or monocyte, infiltration of tissues infected by EBV; most cases of infectious mono are the result of primary EBV infection, although some can be from primary CMV infection or EBV infection that causes polyclonal, EBV-infected B-cells to start producing anti-CMV IgM and make it look like a person who had previously been infected with CMV has a primary CMV infection). I don't feel like going through papers and discussing them, but here are some hastily-done searches showing vast numbers of articles on the subject (http://scholar.google.com/scholar?num=100&hl=en&lr=&q=mononucleosis+brain+OR+encephalitis+OR+encephalopathy+OR+meningitis+OR+meningeoencephalitis); (http://scholar.google.com/scholar?q=mononucleosis++brain+OR+encephalitis+OR+encephalopathy+OR+meningitis+OR+meningeoencephalitis&num=100&hl=en&lr=&scoring=r&as_ylo=2004)]. The pro-inflammatory response during infectious mono is massive, and the notion that the blood-brain barrier would be impermeable to infiltration by EBV-infected, polyclonal B-cells is not reasonable. Also, an important distinguishing feature of infectious mono is enlargement or lymphadenopathy in the posterior cervical lymph nodes that provide lymphatic drainage to the brain, producing a stiff neck, etc. The oligoclonal IgG antibodies in the CSF of people with multiple sclerosis have repeatedly been shown to bind EBV proteins (Cepok et al., 2005), and Cepok et al. (2005) go into all the research showing that type of thing. It's possible that the immune response is being directed against other latently-infected B-cells, etc., but the evidence is pretty substatial that late EBV infection plays some role in the etiology of multiple sclerosis, in my opinion. To think that astrocytes and probably microglia and other cell types in the central nervous system would be spared infection makes no sense to me. So it probably occurs in many or most people who are infected with EBV (90-95 percent of the US population, by age 26-27), and one might look for some differences in the degree of ongoing damage or in the pattern of gene expression by EBV (i.e. the latency pattern) in astrocytes or microglia, etc. (discussed below) of people who go on to develop multiple sclerosis, in comparison to controls.

Behan et al. (2002) discuss a lot of evidence that inappropriate astrocytic cell-cycle re-entry plays a prominent role in the etiology of multiple sclerosis, and the authors, one of whom is Chaudhuri (see Chaudhuri, 2005), also discuss the association of multiple sclerosis with glioblastoma multiforme and with rare, diffuse forms of gliomas, etc. That article is superb and is really brilliant, and yet it's not even indexed in Medline. The fact that vitamin D analogs have been used to treat glioblastoma multiforme is interesting, and the effects of vitamin D receptor (VDR) ligands, including calcitriol itself, on the astrocytic cell cycle could suggest that they could protect against astrocytic cell cycle re-entry and apoptosis in people with multiple sclerosis. I tend to think they wouldn't be all that effective in that regard and that the focus of Chaudhuri (2005) on the developing brain makes more sense. But the focus on astrocytes (Chaudhuri, 2005; Behan et al., 2005) is really intriguing, and it suggests to me that other measures might protect against abnormal astrocyte proliferation and apoptosis (i.e. guanosine and other intravenously-administered purine nucleotides or those in combination with energy substrates, etc.). That's just my opinion. It's interesting that VDR activation leads to very complex interactions with the transforming growth factor-beta signalling cascade, such as by forming heterodimers with Smad3 and potentiating many Smad3-induced transcriptional changes (VanAmerongen et al., 2004), and that the EBV latent membrane protein-1 suppresses Smad3-dependent transcriptional changes (Prokova et al., 2002). Smad3 is phosphorylated by type I TGFbeta receptors and is thereby activated as a transcription factor. Smad3 interacts with many proteins, but the suppression by LMP1 of the TGFbeta-induced and Smad3-mediated increase in p21WAF1/Cip1 expression (Prokova et al., 2002) is a relatively specific intersection with the transcriptional program that tends to be induced by VDR activation. The p21WAF1/Cip1 gene is a major cell-cycle-regulatory gene whose expression is responsive to and increased by VDR activation. The gene product allows for enhanced DNA repair before cell division, etc., and contributes to the antiproliferative and differentiating effects of VDR activation. That's just one example, but it lends credence to the hypothesis of Holmoy (2008) and suggests that the interactions of VDR-ligand-induced transcriptional changes with EBV-induced transcriptional changes may be relatively direct and may go beyond the realm of VDR-ligand-induced increases in interleukin-10 output from monocytes, etc. It might be possible to look for the effects of vitamin D or its analogs on EBV-infected, cultured monocytes or astrocytes or to look for associations of 25-hydroxyvitamin D levels with the incidences of glioblastoma among patients with multiple sclerosis? That sounds pretty difficult. There are some recent articles discussing all the problems with detecting herpesviruses in the brain during autopsies. Serafini et al. (2007) [Serafini et al., 2007: (http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2118531)(http://www.ncbi.nlm.nih.gov/pubmed/17984305)] found that cells in perivascular regions of the brains of people with multiple sclerosis were immunoreactive for LMP1 and other latency-associated EBV proteins, and it doesn't sound like that can be casually attributed to infiltrating, EBV-infected B-cells, etc. Someone could look for an association between 25-hydroxyvitamin D levels at death and the latency pattern of EBV infection in the brains of people with MS (or just look for different latency patterns in people with MS). I'm just thinking out loud with this.