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Ruth Goold
Member in Phase 3
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Posted: Wed Jan 9th, 2008 04:18 |
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The role of the VDR and other nuclear hormone receptors in inflammation and the production of antimicrobials is becoming better known. Unfortunately, the promotion of dietary vitamin D supplementation is based on the belief that higher levels of 25-D will enable the VDR to function better. One key point that needs to be demonstrated is that 25-D is a VDR antagonist, and therefore high levels are immunosuppressive (shutting down the VDR and its transcriptional activities instead of enhancing them).
This free access paper http://www.pnas.org/cgi/content/short/105/1/64
describes a neat protocol for accurately and precisely monitoring the activation of any given steroid hormone receptor by an agonist and its lack of activation by an antagonist. A simple explanation is that in a (human) cell line one can introduce the hormone receptor of interest (e.g. the VDR) which is joined, across a cleavage site, to a transcriptional activator (TA). A modified ligand (agonist) for that receptor is made (1, 25-D in this case) which is attached to an enzyme that can recognize the cleavage site between the VDR and its attached TA.
When the modified agonist (1, 25-D plus enzyme) binds to the modified receptor (VDR plus TA), the agonist changes the conformation of the VDR homo- or hetero-dimer and the enzyme cleaves the TA from the VDR. The released TA travels to the nucleus and activates a stable reporter gene (that can be measured). If a modified antagonist (25-D plus enzyme) is made and binds to the modified VDR, the conformation of the VDR does not change and cleavage does not take place. Thus, no TA is released and the reporter gene is not activated.
A very neat grad student project – anyone with access to a molecular lab and a student or two out there? This method could also be used to show that 25-D is an antagonist for other nuclear hormone receptors, as it appears to be.
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03/02/07 Ph 1 MP; 2001: Pulmonary sarc; 01/04/07: 125 D=110pmol/L(45.8 pg/ml)| 25D=20.8 ng/ml: 04/07 19.2: 07/07 11?: 09/07 16.5: 11/07 <10.0: 01/08 <10.0: 05/08 10 ng/ml. Ca. Elocom (ears). diphenhydramine 25 mg. Adidas EE glasses outside. NoIRs
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Dr Trevor Marshall
Research Team
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Posted: Wed Jan 9th, 2008 04:36 |
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Ruth,
I wish it was as simple as that, but there are known to be (at least) 2 TAs for the VDR, binding at different sites. Secondly, there is a liklihood of functional duplication between (at least) the PXR and VDR. We also know that viruses, at least the HIV virus, directly co-opt the VDR, apparently unactivated by its ligand. Finally, there are the issues of stability of the ligand-receptor-cofactor complex, which I have been examining with my molecular dynamics, and observing in-vivo by noticing the very short time that Olmesartan continues to do its job effectively. Certainly there have to be reasons for this disassociation, and I suspect they are associated with "bumping" into particular proteins and enzymes which have the energy to break apart the complexes.
I may be wrong - but I would opine that this paper is neglecting so much of the knowledge we already have about the way the VDR works. If I had studied the other nuclear receptors as much as the VDR, I am sure that I would be able to pinpoint errors in their thesis with respect to those as well.
One of the things that Molecular Biology needs to understand is that many problems are imponderable, they are too complex too be resolved to definitive answers at this point in time. It seems that today's graduates do not have the ability to stand back and see the forest, but keep butting their heads up against the nearest tree. I prefer to skip around those tree-trunks, and look backwards from the changed perspective  Many have told me that I am "not thorough enough." I would say that I just know how to recognize the magnitude of a problem without first needing to apply for a 5 year study grant
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Dr Trevor Marshall
Research Team
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Posted: Wed Jan 9th, 2008 04:46 |
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Ruth,
There are several studies which I cite in my upcoming paper that conclude the Vitamin D metabolites are, on the whole, immunosuppressive (although I argue that they activate a receptor transcribing over a thousand genes, and to expect an immunosuppressive/non-immunosupressive mode of operation is over-simplification).
You might also like to review this paper:
"Additive immunosuppressive effects of 1,25-dihydroxyvitamin D3 and corticosteroids on TH1, but not TH2, responses"
http://tinyurl.com/3dfenv
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Ruth Goold
Member in Phase 3
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Posted: Wed Jan 9th, 2008 16:48 |
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Dr Trevor Marshall wrote: Ruth,
I wish it was as simple as that, but there are known to be (at least) 2 TAs for the VDR, binding at different sites. Secondly, there is a liklihood of functional duplication between (at least) the PXR and VDR.
Whoa tiger – this technique provides a simple quantitative way to monitor the activation (or lack thereof) of any chosen NHR with candidate agonist (and antagonist) ligands. I wasn’t suggesting that it answered all the mysteries of immuno suppression/activation.
Everyone agrees that proper functioning of the VDR (and other NHRs) is essential for good health. It doesn’t matter whether, in the end, one concludes that on balance, proper functioning of the VDR is described as immuno-suppressive or immuno-acitvating (clearly, at different stages in inflammation and cancer it serves both functions).
Unfortunately, many people are assuming that high levels of 25-D will promote proper activation of the VDR. Your molecular modelling has put you away ahead of the field in terms of understanding the conditions required for ‘proper activation of the VDR’ and these do not include high concentrations of 25-D. The technique described in this paper simply provides a simple tool to confirm on an experimental basis your deductions regarding activation/inacitvation of the VDR without following the thousands of transient transcriptional events that indicate VDR activation in real life.
It doesn’t matter how many how many TAs there are for the VDR because in this protocol you introduce the modified VDR into your cell line and also introduce the reporter gene that produces a single stable (i.e. measurable) product when the the introduced VDR (and only the introduced VDR – not any endogenous NHRs in the cell line) is activated. You now have an experimental system in which to monitor the effect of ligands on activation of the VDR. It could be used to demonstrate that 1, 25-D activates the VDR and 25-D doesn’t. Moreover, it could be used to show that even 1, 25-D doesn’t activate the VDR effectively when in the presence of high concentrations (i.e. >18 ng/ml) of 25-D. It could also be used to confirm the action of new candidate agonists/antagonists (such as capnine) on the VDR.
These demonstrations might seem trivial to you because you are so familiar with the concepts revealed by your modelling work. However, the world of experimental biology is quite far behind and primarily incapable of assessing the veracity of your molecular models (and therefore just ignores them). This protocol provides an experimental tool to verify the predictions of your modelling in terms that even the most simple-minded biologist can understand.
I know this is just a beginning but I think it is a critical step to undertake if you (we all) want to stem the tide of vitamin D promotion such as that exemplified in the NBC clip. Imagine a headline proclaiming “ Vitamin D supplements shut down the Body’s Cancer Protection System.”
Next, the world would have to be made to understand why folks with Th1 disease have high levels of 1, 25-D, low levels of 25-D and still don’t have functional VDRs. Now I see experiments to show that capnine (and perhaps cell homogenates from Th1 compromised patients containing other bacterial ligands) can also out-compete 1, 25-D for binding to the VDR but fail to activate it. (Of course, since the VDR is a self-regulating molecule the sustained high levels of 1, 25-D in Th1 people is an indication in itself that the VDR is not functioning – otherwise it would shut its own production off. But, again, you are a long way ahead of others in this understanding).
Anyway, this is but a simple tool I know, but I think you are underestimating its potential usefulness.
Ruth
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03/02/07 Ph 1 MP; 2001: Pulmonary sarc; 01/04/07: 125 D=110pmol/L(45.8 pg/ml)| 25D=20.8 ng/ml: 04/07 19.2: 07/07 11?: 09/07 16.5: 11/07 <10.0: 01/08 <10.0: 05/08 10 ng/ml. Ca. Elocom (ears). diphenhydramine 25 mg. Adidas EE glasses outside. NoIRs
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Ruth Goold
Member in Phase 3
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Posted: Wed Jan 9th, 2008 17:06 |
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Dr Trevor Marshall wrote: You might also like to review this paper:
"Additive immunosuppressive effects of 1,25-dihydroxyvitamin D3 and corticosteroids on TH1, but not TH2, responses"
http://tinyurl.com/3dfenv
Interesting article on a couple of fronts (I have only read the abstract so far) - one comment that catches my attention is:
"However, this hormone [1, 25-D] upregulates T(H)2 cytokines and ..."
So, since the most significant result of 10 months of MP for me has been a big increase in allergic responses, I suppose that I can indeed conclude that this is a sign that my VDR is becoming functional 
Thanks,
Ruth
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03/02/07 Ph 1 MP; 2001: Pulmonary sarc; 01/04/07: 125 D=110pmol/L(45.8 pg/ml)| 25D=20.8 ng/ml: 04/07 19.2: 07/07 11?: 09/07 16.5: 11/07 <10.0: 01/08 <10.0: 05/08 10 ng/ml. Ca. Elocom (ears). diphenhydramine 25 mg. Adidas EE glasses outside. NoIRs
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wrotek
Member in Phase 3
| Joined: | Fri Dec 31st, 2004 |
| Location: | Wroclaw, Poland |
| Posts: | 1121 |
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Posted: Sun Jan 13th, 2008 13:57 |
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You might also like to review this paper: "Additive immunosuppressive effects of 1,25-dihydroxyvitamin D3 and corticosteroids on TH1, but not TH2, responses" http://tinyurl.com/3dfenv I understand 1,25-D exerts their immunosupressive actions through some other receptor when it's concentration becomes too high ? Is this some kind of protection mechanism ?
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Lyme reflux chronic pain fatigue depression 125D36 Ph1Sep05 Ph2Oct06 Ph3Apr07 homebound in low lux NoIRs 25D<7 Oct06
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NickBowler
Member in Phase 3
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Posted: Mon May 12th, 2008 15:38 |
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This study apparently shows a statistical bias in death rates from cancer for people with lower levels of vitamin 25D, and also a statistical correlation between higher levels of vitamin 25D and reduced risk of death from cancer. No correlation was found with 1,25D levels and cancer death rates according to the report.
http://www.nutraingredients.com/news/ng.asp?n=85199&c=TSqlTaJaZt7tHjhP0OP9gw%3D%3D
On the face of it, this would seem to be very hard to refute evidence, and makes it difficult to explain how 25D could possibly be immunosuppressive. What would be the best articles to cite as a counter argument? I could try and get something published in CAM (UK equivalent of Townsend letter) if some guidance on this were available.
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Sarcoirodis CIDP, MP start 11/07, NoIRs, 02/08 25D-8, Ph3 since 07/08|
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Dr Trevor Marshall
Research Team
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Posted: Mon May 12th, 2008 15:53 |
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Nick, the problem is that Medicine doesn't understand Figure 1 of my recent Bioessay. Fig.1 describes how the body's production 25-D is down-regulated by the PXR receptor in Th1 disease.
So a high level of 25-D means healthy people, low levels means people with Th1 disease. You wouldn't think that would be so very complex...
What these studies are seeing is the depression of 25-D production by the disease state, and has nothing to do with 'deficiency.' The D metabolites are sterols and seco-steroids, produced by the body itself, and have nothing to do with nutrition.
Oh... I didn't say Th1 inflammation causes cancer? Well, we don't have enough evidence yet for me to say that. A total lack of metastatic cancer in our (high risk) cohort leaves me with an absence of data - something which is hard to build upon... The key here is the gene MTSS1, which is transcribed by the VDR. When the VDR is blocked by the pathogens the body cannot produce the MIM protein from this gene. That protein is called "Missing in Metastasis" (also called "Metastasis Suppressor #1")and the names are self-explanatory, I think...
Last edited on Mon May 12th, 2008 15:57 by Dr Trevor Marshall
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NickBowler
Member in Phase 3
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Posted: Mon May 12th, 2008 16:03 |
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Trevor,
Are there any studies you know of which show that people using Sartans or even ACE inhibitors long term (possibly unblocking their VDR's), have reduced incidence of metastatic cancer?
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Sarcoirodis CIDP, MP start 11/07, NoIRs, 02/08 25D-8, Ph3 since 07/08|
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Dr Trevor Marshall
Research Team
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Posted: Mon May 12th, 2008 16:27 |
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No, and I wouldn't expect any. As I wrote to the Lancet (some time ago) it is a terrible mistake to treat these drugs as "families." Each drug is different. Only Olmesartan has the ability to activate the VDR, and then only if dosed as we do in the MP. The chance that anybody would stumble across this during an Evidence-based trial is zero, and zero people have. The ONLY way to make this breakthrough is by understanding, and exploring, the molecular biology, and then applying that knowledge to the clinical setting
http://www.thelancet.com/journals/lancet/article/PIIS0140673606695093/fulltext
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NickBowler
Member in Phase 3
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Posted: Thu May 15th, 2008 09:33 |
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There is some new information on the cancer and aging protective aspects of vitamin D here:
http://www.nutraingredients.com/news/ng.asp?n=85255&c=TSqlTaJaZt7lbz2Hcz8dCg%3D%3D
At least they are referring to the right molecule this time!!
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Sarcoirodis CIDP, MP start 11/07, NoIRs, 02/08 25D-8, Ph3 since 07/08|
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Dr Trevor Marshall
Research Team
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Posted: Thu May 15th, 2008 16:32 |
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Those wet biologists are thrashing around in the dark. They need to learn about in-silico methodology...
G6PD is transcribed by the VDR (Table 1 of Wang, et al)
Of course it is stimulated by 1,25-D in healthy cells, but not in cancer (infected) cells.
..Trevor..
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