The Nrf2 - NF-kappaB interest thread

Started by Guest_Shinigami_* , Dec 25 2008 04:31 AM

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#61 Guest_Shinigami_*

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Posted 27 December 2008 - 05:41 PM

J Immunol. 2008 Nov 15;181(10):7090-9. <script language="JavaScript1.2">Links
Respiratory epithelial cells convert inactive vitamin D to its active form: potential effects on host defense.

Hansdottir S, Monick MM, Hinde SL, Lovan N, Look DC, Hunninghake GW. Department of Medicine, University of Iowa Carver College of Medicine, and Veterans Administration Medical Center, Iowa City, IA 52242, USA. sif-hansdottir@uiowa.edu

The role of vitamin D in innate immunity is increasingly recognized. Recent work has identified a number of tissues that express the enzyme 1alpha-hydroxylase and are able to activate vitamin D. This locally produced vitamin D is believed to have important immunomodulatory effects. In this paper, we show that primary lung epithelial cells express high baseline levels of activating 1alpha-hydroxylase and low levels of inactivating 24-hydroxylase. The result of this enzyme expression is that airway epithelial cells constitutively convert inactive 25-dihydroxyvitamin D(3) to the active 1,25-dihydroxyvitamin D(3). Active vitamin D that is generated by lung epithelium leads to increased expression of vitamin D-regulated genes with important innate immune functions. These include the cathelicidin antimicrobial peptide gene and the TLR coreceptor CD14. dsRNA increases the expression of 1alpha-hydroxylase, augments the production of active vitamin D, and synergizes with vitamin D to increase expression of cathelicidin. In contrast to induction of the antimicrobial peptide, vitamin D attenuates dsRNA-induced expression of the NF-kappaB-driven gene IL-8. We conclude that primary epithelial cells generate active vitamin D, which then influences the expression of vitamin D-driven genes that play a major role in host defense. Furthermore, the presence of vitamin D alters induction of antimicrobial peptides and inflammatory cytokines in response to viruses. These observations suggest a novel mechanism by which local conversion of inactive to active vitamin D alters immune function in the lung.

Note the paradox: the increased expression of TLR coreceptor CD14, should make it easier to detect infections and therefore activate NF-kappaB. However, the expression of an NF-kappaB driven gene, IL-8, was attenuated. Does this mean that the increased expression of TLR coreceptor CD14 first activates NF-kappaB thereby killing off some bacteria, such that the source of the problem gets eliminated and the inflammation (and activation of NF-kappaB) is subsequently attenuated? I guess I will need to read the full text on this one (I have access, and will do so later.)

Since this article appeared I started taking the Vitamin D3 supplements I got in store for a while but was hesitant to take because I knew too little about it. Taking Vitamin D3 almost immediately improves symptoms.

Taken together with the relation of NF-kappaB to bacterial infections (more on this relation coming up), this may suggest I have a bacterial infection.

The paper was accompanied by a news item on Eurekalert:

Public release date: 4-Nov-2008
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Contact: Becky Soglin
becky-soglin@uiowa.edu
319-335-6660
University of Iowa
<h1 class="title">Lung airway cells activate vitamin D and increase immune response</h1> Vitamin D is essential to good health but needs to be activated to function properly in the human body. Until recently, this activation was thought to happen primarily in the kidneys, but a new University of Iowa study finds that the activation step can also occur in lung airway cells.

The study also links the vitamin D locally produced in the lung airway cells to activation of two genes that help fight infection. The study results appear in the Nov. 15 issue of the Journal of Immunology, now online.

In addition to contributing to calcium absorption and bone health, vitamin D is increasingly recognized for its beneficial effects on the immune system. Vitamin D deficiency has been recently linked to increased risk of some infections, autoimmune diseases such as multiple sclerosis and type 1 diabetes, and some cancers.

"The more scientists have been studying vitamin D, the more we learn about new roles it plays in the human body," said the study's lead author Sif Hansdottir, M.D., fellow in internal medicine in the University of Iowa Carver College of Medicine. "The active form of vitamin D is known to affect the expression of more than 200 genes, so we were interested both in the possible lung-specific production of active vitamin D and in vitamin D-dependent production of proteins that fight infections."

The first step in vitamin D activation takes place in the liver, where an enzyme called 25-hydroxylase converts vitamin D into a "storage" form. The next step takes place typically in the kidneys, but in recent years, tissue and organs such as skin, intestines, breast and prostate have been found also to express the enzyme that completes vitamin D conversion.

The University of Iowa team, based in the laboratory of Gary Hunninghake, M.D., professor of internal medicine and the study's senior author, used cells from deceased human donors to demonstrate that the presence of the enzyme 1 alpha-hydroxylase in the airway cells helps convert the storage form of vitamin D into its active form.

"When we put the storage form of vitamin D on the lung airway cells, we saw them convert it to the active form," Hansdottir said. "The next step was to investigate whether this active form could affect the expression of genes."

The team then showed that vitamin D activated by airway cells affects two genes involved in immune defense. One gene expresses a protein called cathelicidin that can kill bacteria. The second gene, called CD14, produces a protein that helps cells recognize different kinds of pathogens that could be a threat.

"Vitamin D converted by the kidneys circulates in the bloodstream, but vitamin D converted by other organs appears to stay within those organs and protect them from infection," Hansdottir said. "We were able to see this happen in cells lining the trachea and main bronchi."

The team also found that when lung airway cells are infected by a virus, they express more of the enzyme that activates vitamin D. Hansdottir said the team is very interested in pursuing studies on the role of viral infections in vitamin D production and subsequent effects on lung infections.

"Vitamin D not only increases proteins involved in bacterial killing but also can dampen inflammation," Hansdottir said. "Controlling inflammation through vitamin D is good because too much inflammation can cause problems such as sepsis and seems to contribute to autoimmune disease."

Hansdottir noted that vitamin D insufficiencies and deficiencies (which are more severe) are fairly common, particularly for people living in northern latitudes. While vitamin D can be generated through sun exposure, such exposure is generally not recommended as a remedy because of skin cancer risks. Instead, supplements can be used.

The American Academy of Pediatrics recently recommended that the vitamin D dosage for children be increased to 400 IU (international units) per day. Optimal daily intake for adults is still being studied but may be as high as 800 to 1,000 IU.

How much is a Vitamin D3 IU in µgs?

How much time does it take after ingesting Vitamin D3 (cholicalciferol) for it to be processed by the liver into storage form which can then be converted to the active form in the lungs? Could the lungs process cholicalciferol directly into storage and subsequently active form?

#62 Guest_Shinigami_*

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Posted 27 December 2008 - 06:11 PM

How much is a Vitamin D3 IU in µgs?

one IU equals 25 ng

So,

1 IU Vitamin D3 = 25E-9 g Vitamin D3
1 g Vitamin D3 = 40E+6 IU Vitamin D3

2000 IU Vitamin D3 = 50E-6 g Vitamin D3 = 50 µg Vitamin D3

#63 Guest_Shinigami_*

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Posted 27 December 2008 - 06:50 PM

Does this mean that the increased expression of TLR coreceptor CD14 first activates NF-kappaB thereby killing off some bacteria, such that the source of the problem gets eliminated and the inflammation (and activation of NF-kappaB) is subsequently attenuated?

Actually, it obviously isn't necessary for NF-kB to be upregulated first: if the vitamin D3 induces cathelicidin thereby killing off the bacteria, the source of the problem is eliminated directly without intervention of NF-kB and NF-kB can subsequently be downregulated.

#64 Brainbox

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Posted 27 December 2008 - 07:00 PM

This is very interesting. Thanks. Needs some effort to read. And then, maybe reply if there's any need for that left. :-D

#65 Guest_Shinigami_*

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Posted 27 December 2008 - 09:54 PM

Infect Immun. 2008 Oct;76(10):4455-62. Epub 2008 Jul 21. <script language="JavaScript1.2">Links
Mycoplasma pneumoniae infection and environmental tobacco smoke inhibit lung glutathione adaptive responses and increase oxidative stress.
Kariya C, Chu HW, Huang J, Leitner H, Martin RJ, Day BJ. University of Colorado Health Sciences Center, Denver, Colorado 80206, USA.. Chronic cigarette smoking evokes a lung glutathione (GSH) adaptive response that results in elevated GSH levels in the lung epithelial lining fluid (ELF). Currently, little is known about how the lung regulates or maintains steady-state levels of ELF GSH. Pathogens such as Mycoplasma pneumoniae can exacerbate airway inflammation and oxidative stress. The present study examined whether M. pneumoniae infections synergize with environmental tobacco smoke (ETS) to disrupt lung GSH adaptive responses. Mice were exposed separately and in combination to ETS and M. pneumoniae for 16 weeks. ETS exposure resulted in a doubling of ELF GSH levels, which was blocked in the M. pneumoniae-exposed mice. In addition, the ETS-plus-M. pneumoniae-exposed mice had elevated levels of oxidized glutathione (GSSG), resulting in a dramatic change in the ELF redox state that corresponded with an increase in lung tissue DNA oxidation. Similar findings were observed in human lung epithelial cells in vitro. Cells exposed separately or in combination to cigarette smoke extract and M. pneumoniae for 48 h had elevated apical levels of GSH compared to control cells, and these increases were blocked by M. pneumoniae and were also associated with increased cellular DNA oxidation. Further studies showed that M. pneumoniae exposure blocked ETS-induced increases in GSH reductase, an enzyme that recycles GSSG back to GSH, both in vitro and in vivo. These studies suggest that M. pneumoniae infection synergizes with ETS and suppresses the lung's ability to respond appropriately to environmental challenges leading to enhanced oxidative stress.

This article also got a news item on Eurekalert:

Public release date: 19-Aug-2008
[ <script language="javascript" type="text/javascript"> Print Article | E-mail Article <script language="javascript" type="text/javascript"> | Close Window ]

Contact: William Allstetter
allstetterw@njc.org
303-398-1002
National Jewish Medical and Research Center
<h1 class="title">Infection blocks lung's protective response against tobacco smoke</h1> An infection that often goes undetected can block the lung's natural protective response against tobacco smoke, according to researchers at National Jewish Health. The findings, recently published online and scheduled to appear in the October issue of Infection and Immunity, suggest one mechanism that may cause smokers to develop chronic obstructive pulmonary disease.

"Although smoking is the overwhelming cause of chronic obstructive pulmonary disease (COPD), only 20 percent of smokers develop the disease," said Brian Day, senior author on the study and Professor of Medicine at National Jewish Health. "Our findings suggest that Mycoplasma pneumoniae (Mp) infection may be one of the co-factors that lead to COPD and other diseases among smokers."

Tobacco smoke contains more than 4,700 chemicals, which generate approximately 100 trillion reactive molecules per puff. Those molecules, known as reactive species, can damage lung tissue by chemically reacting with DNA, cell membranes and other molecules in the lung.

It has long been known that the lungs mount a strong protective response against tobacco smoke, which the National Jewish researchers confirmed in their studies in mice and cell cultures. They found that mice exposed to tobacco smoke for 16 weeks doubled the amount of the antioxidant glutathione in the fluid bathing the airways. The antioxidant reacts with the reactive species in tobacco smoke, thus preventing damaging reactions with lung tissue.

"This natural protective response actually allows people to smoke," said Day. "Without it, all smokers would suffer significantly more lung damage."

Previous work in Dr. Day's lab had suggested that lung infections might affect the lung's protective response. And work in Dr. Richard Martin's lab at National Jewish has implicated the organism Mycoplasma pneumoniae (Mp) in worsening asthma. Mp is a common lung pathogen and the most common cause of pneumonia, but can be difficult to detect because it is challenging to grow in culture. Recent tests to detect Mp DNA in the lungs have indicated that it may be more prevalent than generally recognized and can exist as a low-level chronic infection.

When Dr. Day and his colleagues infected mice with Mp it had a mild effect, slightly lowering glutathione levels in the lungs of mice breathing fresh air. When mice were exposed to tobacco smoke then infected with Mp, glutathione levels dropped even lower.

"The Mycoplasma infection completely blocked the protective response mice normally mount against tobacco smoke, reducing antioxidant levels well below even those of mice breathing fresh air," said Dr. Day.

After glutathione reacts and neutralizes reactive species in the lungs, it becomes oxidized. Under normal conditions mice and humans produce an enzyme, called glutathione reductase (GR), which recycles the oxidized glutathione into its protective, reduced state.

The researchers found that mice exposed to tobacco smoke and Mp had much higher levels of oxidized glutathione along with the low levels of reduced glutathione. The researchers also found that the Mp infection significantly lowered levels of GR in mice lungs.

"The Mycoplasma infection blocked the lungs' protective response to tobacco smoke by lowering levels of the enzyme that normally recycles oxidized glutathione back into its protective, reduced form," said Dr. Day. "This resulted in severe oxidative stress and increased tissue damage as measured by oxidized DNA.

"These higher levels of oxidative stress and damage are likely to predispose smokers with Mycoplasma infections to lung disease, such as COPD or cancer."

From an abstract I quoted previously:

We report that Nrf2 modulates the GSH redox state by regulating glutathione reductase (GSR).

I clearly thought this one had me covered. However, a blood-test for mycoplasma was negative. I'm now wondering whether the blood-test was sufficiently accurate. But how could I test this? The effect of Vitamin D certainly suggests this is a possiblity.

Edited by Shinigami, 27 December 2008 - 10:09 PM.

#66 100YearsToGo

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Posted 28 December 2008 - 01:05 AM

Hi Shinigami,

Thanks for the invitation to your thread. Yes cigarette smoke is a B*tch.

I have some questions. Do you have a strong reaction only to cigarette smoke? Or are you alergic to other stuff as well? Pollen etc...?
why are you going the Nrf2 NF-kappaB route? Can you rule out the following scenario?

As you know cigarette smoke contains a number of toxic chemicals and irritants. Lymphocyte (Mast cells) are responsable for the immune system response to alergens. A disfunctional (exaggerated) Mast cell reaction to a substance like smoke is to flood your system with histamine that on its turn causes inflamation of the mucosa. In the case of the the brionchial mucosa MAST cell function is also dependent on T-Cell activity. It is therefore not surprising that Vitamin D3 would have a positive effect on you. Vitamin D3 is a known immune system regulator. I suggest you take something between 5000iu and 10000iu daily. I'm sure you know that the DRI/RDA of 400iu is BS. How much are you taking now? Additionally I would add Quercetin ( 500mg twice daily) to your regimen. Quercetin is known to normalize MAST Cell function if used consistantly for a long period. (2 to 6 months). I'm sure you can test the veracity of above statements yourself. The recommended dosages are also a matter of public record.

Let me know if you have allready ruled this out, so we can continue on your quest.

Edited by 100YearsToGo, 28 December 2008 - 01:06 AM.

#67 Guest_Shinigami_*

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Posted 28 December 2008 - 02:10 PM

Do you have a strong reaction only to cigarette smoke? Or are you alergic to other stuff as well? Pollen etc...?

My reaction to cigarette smoke is beyond extreme (and it is thus easy to notice that the reaction is caused by cigarette smoke). It also doesn't feel too good to inhale car exhaust gasses, but the effect is not comparable. I hold my breath when crossing streets in order not to inhale car exhaust gasses, but the main car exhaust gas I am worried about is the second hand smoke produced by its occupants (the effect correlates with an open window (especially in colder weather) and the (slight) smell of cigarette smoke).

I also used to be highly allergic to pollen, and had to take anti-histamines (claritine/loratadine) to make it bearable. Interestingly, pollen no longer have any effect (or at least it is almost unnoticable) on me since the time I try to completely avoid inhaling cigarette smoke (and got good at it). I still have some claritine lying around and have tried to take it to see whether it has any effect on my reaction to cigarette smoke (inspired by the relation of cannabinoids and NF-kappaB with the immune system), but I'm not sure whether that had a significant effect (maybe I should try it again?). I also don't like the idea of taking anti-histamines to fight the problem too much, since it seems this way I'm mainly masking symptoms rather than attacking the cause of the problem, which may not be the ideal strategy.

I have visited a lung specialist which had me tested for allergies to other stuff, and it seems I'm also allergic to dust, cats and dogs (when injected subdermally), although I don't notice this allergy in practice.

why are you going the Nrf2 NF-kappaB route?

I found out about NF-kappaB when trying to find out why cannabinoids attenuated the inflammation to such a large extent. There is more relation between NF-kappaB and cannabinoids than the abstracts I posted thusfar suggest (for example also through IKK and the RelA component of NF-kappaB). I think the adenosine signalling effect of cannabinoids on NF-kappaB may be the main route of action in my case, since drinking a moderate/large amount of coffee (known to increase adenosine signalling if I remember correctly -- will try to post studies later) when under moderate cigarette smoke-induced inflammatory attack worsens symptoms dramatically (coffee doesn't have much effect when I feel fine, other than feeling caffeinated of course

).

I also read some studies which discuss the timescale of NF-kappaB activation (will try to post them later), and this seems to fit well with the timescale of cigarette smoke-induced inflammation I am experiencing.

Additionaly, chopped garlic (known to activate Nrf2) seems to have a slight protective effect (by improving basal levels of GSH?) (in contrast with cannabinoids which mainly seem to attenuate inflammation), which fits very nicely with the theory discussed in the mycoplasma article above. Vitamin C also seems to give a noticable improvement to symptoms (through improvement of GSH? I don't really remember how that was supposed to work...).

Can you rule out the following scenario?

As you know cigarette smoke contains a number of toxic chemicals and irritants. Lymphocyte (Mast cells) are responsable for the immune system response to alergens. A disfunctional (exaggerated) Mast cell reaction to a substance like smoke is to flood your system with histamine that on its turn causes inflamation of the mucosa. In the case of the the brionchial mucosa MAST cell function is also dependent on T-Cell activity. It is therefore not surprising that Vitamin D3 would have a positive effect on you. Vitamin D3 is a known immune system regulator. I suggest you take something between 5000iu and 10000iu daily. I'm sure you know that the DRI/RDA of 400iu is BS. How much are you taking now? Additionally I would add Quercetin ( 500mg twice daily) to your regimen. Quercetin is known to normalize MAST Cell function if used consistantly for a long period. (2 to 6 months). I'm sure you can test the veracity of above statements yourself. The recommended dosages are also a matter of public record.

Let me know if you have allready ruled this out, so we can continue on your quest.

That is an interesting idea. Mast cells also don't entirely seem unrelated to NF-kB though, but the relation is very unclear to me now. I will research this a little more and give you a more elaborate response to this later.

#68 Guest_Shinigami_*

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Posted 28 December 2008 - 03:52 PM

Curr Opin Pulm Med. 2006 Jan;12(1):54-9. Links
Adenosine signaling in asthma and chronic obstructive pulmonary disease.
Mohsenin A, Blackburn MR. Department of Biochemistry and Molecular Biology, University of Texas - Houston Medical School, Houston, Texas 77030, USA. PURPOSE OF REVIEW: The chronic lung diseases, asthma and chronic obstructive pulmonary disease, are pulmonary disorders in which persistent inflammation and alterations in lung structure contribute to a progressive loss of lung function. Although the exact type of inflammation and damage in each disease is distinct, they share the common feature that they are chronic in nature. Despite efforts, little is known about the cellular and molecular mechanisms that drive the chronicity of these two diseases. This review will summarize important findings regarding the role of adenosine, a signaling nucleoside implicated in the pathogenesis of these two disorders. RECENT FINDINGS: Aerosolized adenosine induces bronchoconstriction in patients with asthma and chronic obstructive pulmonary disease primarily through the release of mast cell mediators. In this setting it can not only be used to aid in diagnosis but also to monitor patient responses to steroid therapy. Adenosine levels are elevated in the lungs of asthma patients, indicating greater flux through adenosine receptor signaling pathways. In-vitro studies have shown adenosine to access pathways leading to the genesis of chronic inflammation via the release of proinflammatory cytokines and chemokines. Animal studies demonstrate that merely elevating adenosine levels in the mouse is sufficient to induce a pulmonary phenotype with features of asthma and chronic obstructive pulmonary disease. SUMMARY: Identifying mediators regulating the chronic nature of asthma and chronic obstructive pulmonary disease is critical towards advancements in treatment options. Adenosine has been implicated in promoting the inflammation and airway remodeling seen in chronic lung disease and thus makes an attractive therapeutic target.

#69 Guest_Shinigami_*

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Posted 28 December 2008 - 04:00 PM

Biochem Pharmacol. 2008 Oct 1;76(7):912-21. Epub 2008 Jul 19. <script language="JavaScript1.2">Links
A new chemical tool (C0036E08) supports the role of adenosine A(2B) receptors in mediating human mast cell activation.
Buceta M, Domínguez E, Castro M, Brea J, Alvarez D, Barcala J, Valdés L, Alvarez-Calderón P, Domínguez F, Vidal B, Díaz JL, Miralpeix M, Beleta J, Cadavid MI, Loza MI. BioFarma Research Group, Department of Pharmacology, School of Pharmacy, Instituto de Farmacia Industrial, University of Santiago de Compostela, Santiago de Compostela, Spain. Asthma is a chronic inflammatory disease of the airways that involves many cell types, amongst which mast cells are known to be important. Adenosine, a potent bronchoconstricting agent, exerts its ability to modulate adenosine receptors of mast cells thereby potentiating derived mediator release, histamine being one of the first mediators to be released. The heterogeneity of sources of mast cells and the lack of highly potent ligands selective for the different adenosine receptor subtypes have been important hurdles in this area of research. In the present study we describe compound C0036E08, a novel ligand that has high affinity (pK(i) 8.46) for adenosine A(2B) receptors, being 9 times, 1412 times and 3090 times more selective for A(2B) receptors than for A(1), A(2A) and A(3) receptors, respectively. Compound C0036E08 showed antagonist activity at recombinant and native adenosine receptors, and it was able to fully block NECA-induced histamine release in freshly isolated mast cells from human bronchoalveolar fluid. C0036E08 has been shown to be a valuable tool for the identification of adenosine A(2B) receptors as the adenosine receptors responsible for the NECA-induced response in human mast cells. Considering the increasing interest of A(2B) receptors as a therapeutic target in asthma, this chemical tool might provide a base for the development of new anti-asthmatic drugs.

The cyberbrain exists already

-- bless you, 100YearsToGo.

#70 Guest_Shinigami_*

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Posted 28 December 2008 - 04:22 PM

Vitamin D3 is a known immune system regulator. I suggest you take something between 5000iu and 10000iu daily. I'm sure you know that the DRI/RDA of 400iu is BS. How much are you taking now?

I started taking a daily dose (as indicated on the bottle), 3µg, when I found the above article. Noticing it improved symptoms, and knowing the adviced daily dose was likely much lower than it should be (3µg was indicated as 60% of adviced daily dose), I upped the dose to about 6µg/day. However, the supplements I am taking also contain calcium, 750 mg (90% adviced daily dose) for every 3µg of Vitamin D3. I guess I should find me some supplements that do not contain calcium before I increase it any further.

Additionally I would add Quercetin ( 500mg twice daily) to your regimen. Quercetin is known to normalize MAST Cell function if used consistantly for a long period. (2 to 6 months).

I'm getting quite a bit of quercetin from my diet already, and used to be a dedicated apple-eater (until they went out of season). Now they are back in season, I'll get back to eating them, and maybe find me some supplements or other food that contains it. Thanks for the suggestion.

Let me know if you have allready ruled this out, so we can continue on your quest.

Although there is clearly a link with mast cells, I still wonder what causes my mast cells to overreact and why other people do not. I will have to dig somewhat deeper into this.

Edited by Shinigami, 28 December 2008 - 05:16 PM.

#71 Guest_Shinigami_*

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Posted 28 December 2008 - 04:35 PM

Arch Pharm Res. 2008 Oct;31(10):1303-11. Epub 2008 Oct 29. Links
Flavonoids inhibit histamine release and expression of proinflammatory cytokines in mast cells.
Park HH, Lee S, Son HY, Park SB, Kim MS, Choi EJ, Singh TS, Ha JH, Lee MG, Kim JE, Hyun MC, Kwon TK, Kim YH, Kim SH. CMRI, Department of Pharmacology, Kyungpook National University, Daegu 700-422, Korea. Mast cells participate in allergy and inflammation by secreting inflammatory mediators such as histamine and proinflammatory cytokines. Flavonoids are naturally occurring molecules with antioxidant, cytoprotective, and antiinflammatory actions. However, effect of flavonoids on the release of histamine and proinflammatory mediator, and their comparative mechanism of action in mast cells were not well defined. Here, we compared the effect of six flavonoids (astragalin, fisetin, kaempferol, myricetin, quercetin, and rutin) on the mast cell-mediated allergic inflammation. Fisetin, kaempferol, myricetin, quercetin, and rutin inhibited IgE or phorbol-12-myristate 13-acetate and calcium ionophore A23187 (PMACI)-mediated histamine release in RBL-2H3 cells. These five flavonoids also inhibited elevation of intracellular calcium. Gene expressions and secretion of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-6, and IL-8 were assessed in PMACI-stimulated human mast cells (HMC-1). Fisetin, quercetin, and rutin decreased gene expression and production of all the proinflammatory cytokines after PMACI stimulation. Myricetin attenuated TNF-alpha and IL-6 but not IL-1beta and IL-8. Fisetin, myricetin, and rutin suppressed activation of NF-kappaB indicated by inhibition of nuclear translocation of NF-kappaB, NF-kappaB/DNA binding, and NF-kappaB-dependent gene reporter assay. The pharmacological actions of these flavonoids suggest their potential activity for treatment of allergic inflammatory diseases through the down-regulation of mast cell activation.

#72 Guest_Shinigami_*

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Posted 28 December 2008 - 04:38 PM

Inflamm Res. 2007 May;56(5):210-5. Links
Quercetin inhibits expression of inflammatory cytokines through attenuation of NF-kappaB and p38 MAPK in HMC-1 human mast cell line.
Min YD, Choi CH, Bark H, Son HY, Park HH, Lee S, Park JW, Park EK, Shin HI, Kim SH. Department of Surgery, Chosun University Medical School, Gwangju, 501-759, Republic of Korea. OBJECTIVE AND DESIGN: Mast cell-mediated allergic inflammation is involved in many diseases such as asthma, sinusitis, and rheumatoid arthritis. Mast cells induce production of pro-inflammatory cytokines with immune regulatory properties. We investigated the effect of quercetin on the expression of pro-inflammatory cytokines in human mast cell line, HMC-1. METHODS: HMC-1 cells were stimulated with phorbol 12-myristate 13-acetate (PMA) and calcium ionophore A23187 (PMACI). RESULTS: Quercetin decreased the gene expression and production of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6, and IL-8 in PMACI-stimulated HMC-1 cells. Quercetin attenuated PMACI-induced activation of NF-kappaB and p38 mitogen-activated protein kinase. CONCLUSION: Our study provides evidence that quercetin may suitable for the treatment of mast cell-derived allergic inflammatory diseases.

#73 Guest_Shinigami_*

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Posted 28 December 2008 - 06:35 PM

I think the adenosine signalling effect of cannabinoids on NF-kappaB may be the main route of action in my case, since drinking a moderate/large amount of coffee (known to increase adenosine signalling if I remember correctly -- will try to post studies later)

Actually, the effect of coffee on adenosine signalling stated here may not be correct. Maybe I incorrectly associated a single event of worsening inflammation with coffee consumption? I obviously tried to avoid doing that again to verify the connection...

The effect of coffee on adenosine signalling remains unclear to me for now.

Int Immunopharmacol. 2004 Oct;4(10-11):1409-17. <script language="JavaScript1.2">Links
Caffeine suppresses TNF-alpha production via activation of the cyclic AMP/protein kinase A pathway.
Horrigan LA, Kelly JP, Connor TJ. Department of Pharmacology, National University of Ireland, Galway, University Road, Galway, Ireland. louise.horrigan@nuigalway.ie This study investigated the effect of in vitro exposure to caffeine, and its major metabolite paraxanthine, at concentrations relevant to typical caffeine consumption in humans, on lipopolysaccharide (LPS)-stimulated cytokine production in human whole blood. In addition, a role for the cyclic AMP/protein kinase A (PKA) pathway in the immunomodulatory effect of caffeine was investigated. Diluted whole blood (taken following >/=15 h abstinence from caffeine-containing food and beverages) was preincubated with caffeine or paraxanthine (10-100 microM) and stimulated with LPS (1 proportional, variant g/ml) for 24 h. The proinflammatory cytokines tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-12, and the antiinflammatory cytokine IL-10 were measured in cell-free supernatants. Whilst caffeine and paraxanthine had little or no effect on IL-10, IL-1beta, or IL-12 production, TNF-alpha production was suppressed in all individuals studied. The effect was statistically significant at 100 microM and consistent across seven experiments performed. Although not statistically significant, a similar effect was observed with paraxanthine. Caffeine (100 microM) also increased intracellular cyclic AMP concentrations in LPS-stimulated monocytes isolated from whole blood. Moreover, the effect of caffeine on TNF-alpha production was abolished by pretreatment with the protein kinase A inhibitor Rp-8-Br-cAMPS (10(-4) and 10(-5)M). To conclude, this study demonstrates that concentrations of caffeine that are relevant to human consumption consistently suppress production of the proinflammatory cytokine TNF-alpha in human blood and that this effect is mediated by the cyclic AMP/protein kinase A pathway.

Edited by Shinigami, 28 December 2008 - 06:35 PM.

#74 Guest_Shinigami_*

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Posted 28 December 2008 - 06:48 PM

I think the adenosine signalling effect of cannabinoids on NF-kappaB may be the main route of action in my case, since drinking a moderate/large amount of coffee (known to increase adenosine signalling if I remember correctly -- will try to post studies later)
Actually, the effect of coffee on adenosine signalling stated here may not be correct. Maybe I incorrectly associated a single event of worsening inflammation with coffee consumption? I obviously tried to avoid doing that again to verify the connection...

Although it may give an interesting clue if the effect of coffee on my inflammation is correct, but the effect on adenosine signalling is slightly different.

Pediatr Res. 2008 Sep 17. [Epub ahead of print] Links
TITLE: Caffeine modulates TNF-alpha production by cord blood monocytes: the role of adenosine receptors.
Chavez-Valdez R, Wills-Karp M, Ahlawat R, Cristofalo EA, Nathan A, Gauda EB. Department of Pediatrics [R.C.-V., R.A., E.A.C., E.B.G.], Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; Division of Immunology [M.W.-K., A.N.]. University of Cincinnati College of Medicine, Cincinnati, OH 45229. Caffeine, a non-specific adenosine receptor (AR) antagonist is widely used to treat apnea of prematurity. Because adenosine modulates multiple biological processes including inflammation, we hypothesized that AR blockade by caffeine would increase cytokine release from neonatal monocytes. Using cord blood monocytes (CBM), we investigated 1) the changes in AR mRNA profile (qRT-PCR) and protein expression (western blot) following in-vitro culture, caffeine or lipopolysaccharide (LPS) exposure, and 2) the modulation of cytokine release and cAMP production (ELISA) induced by caffeine and specific AR antagonists: DPCPX(A1R), ZM241385(A2aR), MRS1754(A2bR), and MRS1220(A3R). Following 48h in-culture, A2aR and A2bR gene expression increased 1.9 (p=0.04) and 2.5-fold (p=0.003), respectively. A1R protein expression directly correlated with increasing LPS concentrations (p=0.01), with minimal expression pre-exposure. Only caffeine (50uM) and DPCPX (10nM) decreased TNF-alpha release from LPS activated-CBM by 20 and 25% (p=0.01) and TNF-alpha gene expression by 30 and 50%, respectively, in conjunction with a >/=2-fold increase in cAMP (p<0.05). AR blockade did not modulate other measured cytokines. The induction of A1R following LPS exposure suggests an important role of this receptor in the control of inflammation in neonates. Our findings also suggest that caffeine, via A1R blockade, increases cAMP production and inhibits pre-transcriptional TNF-alpha production by CBM.

#75 100YearsToGo

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Posted 28 December 2008 - 08:06 PM

Vitamin D3 is a known immune system regulator. I suggest you take something between 5000iu and 10000iu daily. I'm sure you know that the DRI/RDA of 400iu is BS. How much are you taking now?
I started taking a daily dose (as indicated on the bottle), 3µg, when I found the above article. Noticing it improved symptoms, and knowing the adviced daily dose was likely much lower than it should be (3µg was indicated as 60% of adviced daily dose), I upped the dose to about 6µg/day. However, the supplements I am taking also contain calcium, 750 mg (90% adviced daily dose) for every 3µg of Vitamin D3. I guess I should find me some supplements that do not contain calcium before I increase it any further.

Additionally I would add Quercetin ( 500mg twice daily) to your regimen. Quercetin is known to normalize MAST Cell function if used consistantly for a long period. (2 to 6 months).
I'm getting quite a bit of quercetin from my diet already, and used to be a dedicated apple-eater (until they went out of season). Now they are back in season, I'll get back to eating them, and maybe find me some supplements or other food that contains it. Thanks for the suggestion.

Let me know if you have allready ruled this out, so we can continue on your quest.
Although there is clearly a link with mast cells, I still wonder what causes my mast cells to overreact and why other people do not. I will have to dig somewhat deeper into this.

It will benefit you if you supplement quercetin.

An apple has about 66mg quercetin. The body can only absorb 54% of this. Giving you 35.64 mg quercetin.

Now Immune system modulation is achieved only at blood levels of 0.5 to 50 uM.

http://www.ncbi.nlm....Pubmed_RVDocSum

The following study shows that 1.5 uM is achieved by getting 1 gram quercetin a day. See the tables in this study:

http://jn.nutrition..../full/128/3/593

You would need 1000/35.64 = 28 apples to get into that range!!!

#76 Guest_Shinigami_*

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Posted 28 December 2008 - 08:10 PM

OK, so I got a little confused on adenosine signalling. Now it makes some sense again; taking this abstract (quoted previously):

Invest Ophthalmol Vis Sci. 2008 Dec;49(12):5526-31. Epub 2008 Jul 18. Links
Mediation of cannabidiol anti-inflammation in the retina by equilibrative nucleoside transporter and A2A adenosine receptor.
Liou GI, Auchampach JA, Hillard CJ, Zhu G, Yousufzai B, Mian S, Khan S, Khalifa Y. Department of Ophthalmology, Medical College of Georgia, Augusta, Georgia 30912, USA. giliou@mcg.edu

PURPOSE: Cannabidiol (CBD), a nonpsychotropic, nontoxic compound has been shown to block diabetes- and endotoxin-induced retinal damage. However, the protective mechanism of this anti-inflammatory cannabinoid is not completely understood. The goal of this study is to determine the role of adenosine signaling in retinal inflammation and its potential modulation by CBD. METHODS: The adenosine receptor (AR) subtypes expressed in rat retinal microglial cells were assessed by quantitative real-time RT-PCR. AR function was determined via in vitro and in vivo inflammatory models. Microglial cells or rats were treated with or without lipopolysaccharide (LPS) in the presence or absence of adenosine, adenosine receptor agonists/antagonists, or CBD. Adenosine uptake and tumor necrosis factor (TNF)-alpha release in cells or in retinas were determined. RESULTS: The results showed that A(2A)ARs are abundantly expressed in rat retinal microglial cells. When the cells or rats were treated with LPS, activation of the A(2A)AR was the most efficient in mediating AR agonist- or CBD-induced TNF-alpha inhibition. CBD inhibited adenosine uptake via equilibrative nucleoside transporter 1 and synergistically enhanced adenosine's TNF-alpha suppression after treatment with LPS. CONCLUSIONS: These results suggest that the activated A(2A)AR in the retinal microglial cells plays a major anti-inflammatory role in the retina and that CBD's anti-inflammatory effects are linked to the inhibition of adenosine uptake.

which suggests that A(2A)AR is indirectly activated by CBD since CBD interferes with the ENT, thereby allowing more adenosine to reach the A(2A)AR, together with the statement that caffeine is a non-specific adenosine receptor antagonist (abstract above), clarifies the opposite effect I am feeling from caffeine versus cannabinoids. (although it doesn't clarify that both have similar effects on TNF-alpha)

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Posted 28 December 2008 - 08:13 PM

You would need 1000/35.64 = 28 apples to get into that range!!!

Yeah, I am not that dedicated. I will look into it.

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Posted 29 December 2008 - 08:24 AM

Am J Respir Cell Mol Biol. 2007 Jul;37(1):3-8. Epub 2007 Apr 5. Links
Deficiency in Nrf2-GSH signaling impairs type II cell growth and enhances sensitivity to oxidants.
Reddy NM, Kleeberger SR, Cho HY, Yamamoto M, Kensler TW, Biswal S, Reddy SP. The Johns Hopkins Bloomberg School of Public Health, Department of Environmental Health Sciences, 615 North Wolfe Street, Baltimore, MD 21205, USA. sreddy@jhsph.edu Redox imbalance has been implicated in the pathogenesis of many acute and chronic lung diseases. The b-Zip transcription factor Nrf2 acts via an antioxidant/electrophilic response element to regulate antioxidants and maintain cellular redox homeostasis. Our previous studies have shown that Nrf2-deficient mice (Nrf2(-/-)) show reduced pulmonary expression of several antioxidant enzymes, which renders them highly susceptible to hyperoxia-induced lung injury. To better understand the physiologic significance of Nrf2-induced redox signaling, we have used primary cells isolated from the lungs of Nrf2(+/+) and Nrf2(-/-) mice. Our studies were focused on type II cells because these cells are constantly exposed to the oxidant environment and play key roles in host defense, injury, and repair processes. Using this system, we now report that an Nrf2 deficiency leads to defects in type II cell proliferation and greatly enhances the cells' sensitivity to oxidant-induced cell death. These defects were closely associated with high levels of reactive oxygen species (ROS) and redox imbalance in Nrf2(-/-) cells. Glutathione (GSH) supplementation rescued these phenotypic defects associated with the Nrf2 deficiency. Intriguingly, although the antioxidant N-acetyl-cysteine drastically squelched ROS levels, it was unable to counteract growth arrest in Nrf2(-/-) cells. Moreover, despite their elevated levels of ROS, Nrf2(-/-) type II cells were viable and, like their wild-type counterparts, exhibited normal differentiation characteristics. Our data suggest that dysfunctional Nrf2-regulated GSH-induced signaling is associated with deregulation of type II cell proliferation, which contributes to abnormal injury and repair and leads to respiratory impairment.

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Posted 29 December 2008 - 08:32 AM

Oncogene. 2008 Oct 2;27(44):5821-32. Epub 2008 Jun 9. Links
Genetic disruption of the Nrf2 compromises cell-cycle progression by impairing GSH-induced redox signaling.
Reddy NM, Kleeberger SR, Bream JH, Fallon PG, Kensler TW, Yamamoto M, Reddy SP. Department of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. Genetic disruption of Nrf2 greatly enhances susceptibility to prooxidant- and carcinogen-induced experimental models of various human disorders; but the mechanisms by which this transcription factor confers protection are unclear. Using Nrf2-proficient (Nrf2(+/+)) and Nrf2-deficient (Nrf2(-/-)) primary epithelial cultures as a model, we now show that Nrf2 deficiency leads to oxidative stress and DNA lesions, accompanied by impairment of cell-cycle progression, mainly G(2)/M-phase arrest. Both N-acetylcysteine and glutathione (GSH) supplementation ablated the DNA lesions and DNA damage-response pathways in Nrf2(-/-) cells; however only GSH could rescue the impaired colocalization of mitosis-promoting factors and the growth arrest. Akt activation was deregulated in Nrf2(-/-) cells, but GSH supplementation restored it. Inhibition of Akt signaling greatly diminished the GSH-induced Nrf2(-/-) cell proliferation and wild-type cell proliferation. GSH depletion impaired Akt signaling and mitosis-promoting factor colocalization in Nrf2(+/+) cells. Collectively, our findings uncover novel functions for Nrf2 in regulating oxidative stress-induced cell-cycle arrest, especially G(2)/M-checkpoint arrest, and proliferation, and GSH-regulated redox signaling and Akt are required for this process.

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Posted 29 December 2008 - 08:55 AM

Int Immunopharmacol. 2008 Dec 8. [Epub ahead of print] Links
Inhibitory effects of flavonoids extracted from licorice on lipopolysaccharide-induced acute pulmonary inflammation in mice.
Xie Y, Dong X, Wu X, Yan X, Xie Q. Zhejiang Respiratory Foods and Drugs Research Laboratory of State Drugs Administration of China, Zhejiang University, Hangzhou 310058, China. Airway inflammation plays important roles in the pathogenesis of acute respiratory distress syndrome (ARDS), asthma and chronic obstructive pulmonary disease (COPD), and anti-inflammatory treatment effectively improves the symptoms of these diseases. To develop the potentially therapeutic compounds for the treatment of pulmonary inflammation, we investigated the effects of licorice flavonoids (LF) extracted from the roots of Glycyrrhiza uralensis (licorice) on lipopolysaccharide (LPS)-induced acute pulmonary inflammation in mice. Acute pulmonary inflammation was induced by intracheal instillation with LPS, treatment with LF at dosages of 3, 10 and 30 mg/kg significantly reduced the LPS-induced inflammatory cells, including neutrophils, macrophages and lymphocytes accumulation in bronchoalveolar lavage fluids (BALF), among these inflammatory cells, LF predominately inhibited neutrophil infiltration, and the maximal effect (30 mg/kg) was as comparable as dexamethasone treatment at 1 mg/kg. Consistent with its effects on neutrophil infiltration, LF treatment significantly increased LPS-induced BALF superoxide dismutase activity, and significantly decreased lung myeloperoxidase activity as well. Furthermore, treatment with LF at 30 mg/kg significantly reduced LPS-induced lung TNFalpha and IL-1beta mRNA expression at 6 h and 24 h after LPS instillation, respectively. Finally, LF at different dosages not only significantly decreased the elevation of lung water content, but also markedly attenuated LPS-induced histological alteration. Therefore, we suggest that LF effectively attenuates LPS-induced pulmonary inflammation through inhibition of inflammatory cells infiltration and inflammatory mediator release which subsequently reduces neutrophil recruitment into lung and neutrophil-mediated oxidative injury, and this study provides with the potential rationale for development of anti-inflammatory compounds from flavonoid extracts of licorice.

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Posted 29 December 2008 - 11:39 AM

Food Chem Toxicol. 2008 Apr;46(4):1257-70. Epub 2007 Sep 15. Links
Anticarcinogenesis by dietary phytochemicals: cytoprotection by Nrf2 in normal cells and cytotoxicity by modulation of transcription factors NF-kappa B and AP-1 in abnormal cancer cells.
Gopalakrishnan A, Tony Kong AN. Graduate Program in Pharmaceutical Sciences, Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA. Cancer statistics from the American Cancer Society and other sources are a stark reminder of our failure to combat this deadly disease. Chemoprevention entails the use of specific naturally occurring dietary or synthetic agents to thwart cancer development and progression. Some of these agents are believed to do so by protecting the cells or tissues from the malicious attack of exogenous carcinogens and/or endogenous reactive oxygen/nitrogen species (RONS) by inducing several detoxifying/antioxidant enzymes that appear to form stable conjugates such as glutathione, glucuronides or sulfates thus rendering the carcinogenic species harmless. This process of inducing the cellular defense enzymes is believed to be mediated by the antioxidant response elements (ARE) within the promoter regions of these genes. Nrf2, a redox sensitive transcription factor has been documented to play a central role in ARE-driven gene expression. Nrf2, under normal unstimulated conditions, remains sequestered in the cytosol by Keap1. The putative chemopreventive agents disrupt the Nrf2-Keap1 association, thereby releasing Nrf2 which then translocates to the nucleus and drives the gene expression of detoxifying enzymes. The role of other transcription factors such as NF-kappaB and AP-1 in carcinogenesis is well established. By modulating the activity of these transcription factors and their upstream signaling molecules, naturally occurring dietary phytochemicals appear to cause apoptosis in abnormal cells that over-express these factors, thereby inhibiting the promotion and progression. This review discusses the most current and up to date understanding of the possible signaling mechanisms by which these naturally dietary phytochemicals can differentially modulate signal transduction cascades such that they can bring about apoptosis/cell death in abnormal cancer cells but at the same time induce defensive enzymes to protect against carcinogenesis in normal cells.

Edited by Shinigami, 29 December 2008 - 11:44 AM.

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Posted 29 December 2008 - 11:54 AM

Food Chem Toxicol. 2008 Apr;46(4):1257-70. Epub 2007 Sep 15. Links
Anticarcinogenesis by dietary phytochemicals: cytoprotection by Nrf2 in normal cells and cytotoxicity by modulation of transcription factors NF-kappa B and AP-1 in abnormal cancer cells.
Gopalakrishnan A, Tony Kong AN. Graduate Program in Pharmaceutical Sciences, Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA. Cancer statistics from the American Cancer Society and other sources are a stark reminder of our failure to combat this deadly disease. Chemoprevention entails the use of specific naturally occurring dietary or synthetic agents to thwart cancer development and progression. Some of these agents are believed to do so by protecting the cells or tissues from the malicious attack of exogenous carcinogens and/or endogenous reactive oxygen/nitrogen species (RONS) by inducing several detoxifying/antioxidant enzymes that appear to form stable conjugates such as glutathione, glucuronides or sulfates thus rendering the carcinogenic species harmless. This process of inducing the cellular defense enzymes is believed to be mediated by the antioxidant response elements (ARE) within the promoter regions of these genes. Nrf2, a redox sensitive transcription factor has been documented to play a central role in ARE-driven gene expression. Nrf2, under normal unstimulated conditions, remains sequestered in the cytosol by Keap1. The putative chemopreventive agents disrupt the Nrf2-Keap1 association, thereby releasing Nrf2 which then translocates to the nucleus and drives the gene expression of detoxifying enzymes. The role of other transcription factors such as NF-kappaB and AP-1 in carcinogenesis is well established. By modulating the activity of these transcription factors and their upstream signaling molecules, naturally occurring dietary phytochemicals appear to cause apoptosis in abnormal cells that over-express these factors, thereby inhibiting the promotion and progression. This review discusses the most current and up to date understanding of the possible signaling mechanisms by which these naturally dietary phytochemicals can differentially modulate signal transduction cascades such that they can bring about apoptosis/cell death in abnormal cancer cells but at the same time induce defensive enzymes to protect against carcinogenesis in normal cells.

This article looks like a must-read to anyone interested in these topics. I have access to many articles, but too little time to read them.

Edited by Shinigami, 29 December 2008 - 12:30 PM.

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Posted 30 December 2008 - 06:05 PM

Actually, adenosine signalling is still very fuzzy to me. The above information still doesn't entirely make sense.

Edited by Shinigami, 30 December 2008 - 06:37 PM.

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Posted 30 December 2008 - 06:18 PM

This is an interesting article from eurekalert on nrf2 and chemoresistance and how cancer "hijacks" this oxidative stress protection mechanism to protect itself.

Public release date: 10-Oct-2006
[ <script language="javascript" type="text/javascript"> Print Article | E-mail Article <script language="javascript" type="text/javascript"> | Close Window ]

Contact: Vanessa Wasta
wastava@jhmi.edu
410-955-1287
Johns Hopkins Medical Institutions
<h1 class="title">Key to lung cancer chemo resistance revealed</h1> Scientists at Johns Hopkins have discovered how taking the brakes off a "detox" gene causes chemotherapy resistance in a common form of lung cancer.

Products made by a gene called NRF2 normally protect cells from environmental pollutants like cigarette smoke and diesel exhaust by absorbing the materials and pumping them out of the cell. Another gene called KEAP1 encodes products that stop this cleansing process. But lung cancer cells sabotage the expression of these same genes to block assault from chemotherapy drugs.

"What we're seeing is that lung cancer cells recruit and distort NRF2 and KEAP1 expression to help tumor cells evade the toxic effects of chemotherapy," says Shyam Biswal, Ph.D., associate professor at the Johns Hopkins Bloomberg School of Public Health and Kimmel Cancer Center, who published results of cell culture studies in the October 3, 2006 issue of PLoS Medicine.

Past studies have shown that NRF2 detoxifies cells by directing proteins to absorb and pump out pollutants and chemicals. The NRF2 gene makes a "trigger" protein which starts the production of other proteins and enzymes that sweep the cell clear of toxins. To halt the detox process, proteins manufactured by KEAP1 bind to the NRF2 triggers tagging them for destruction. In cancer cells, NRF2 activity runs amok, sweeping away all cellular toxins, including chemotherapy agents.

Biswal says that blocking NRF2 activity could improve the effectiveness of standard chemotherapy drugs, particularly platinum-based compounds widely used for lung cancer.

In Biswal's study, half of 12 lung cancer cell lines and 10 of 54 tissue samples from non-small cell lung cancer patients had mutations in the KEAP1 gene rendering it inactive and unable to keep NRF2 activity in check. In addition, half of the tissue samples were missing one copy of the KEAP1 gene - cells usually have two copies of each gene. No missing genes or mutations were observed in normal lung tissues from the same patients.

NRF2 activity along with its cleansing proteins and enzymes were higher in tumor samples than normal cells, according to the researchers. Their cell culture tests also show that cancer cells with KEAP1 mutations are more resistant to chemotherapy drugs than normal lung cells.

Tumor samples with normal KEAP1 genes also show increased levels of NRF2 and its enzymes, suggesting other ways of dismantling KEAP1, such as splicing the gene to make a shortened, ineffective protein, he said.

The researchers plan to confirm their findings with a larger set of samples and then to screen for appropriate drugs. Funding for the study was provided by the National Cancer Institute Lung SPORE (Specialized Program of Research Excellence), National Heart Lung and Blood Institute, National Institute of Environmental Health Sciences Center, National Institute of Health, and the Flight Attendant Medical Research Institution.

PLoS Med. 2006 Oct;3(10):e420. <script language="JavaScript1.2">Links

Dysfunctional KEAP1-NRF2 interaction in non-small-cell lung cancer.
Singh A, Misra V, Thimmulappa RK, Lee H, Ames S, Hoque MO, Herman JG, Baylin SB, Sidransky D, Gabrielson E, Brock MV, Biswal S. Department of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland, United States of America.

BACKGROUND: Nuclear factor erythroid-2 related factor 2 (NRF2) is a redox-sensitive transcription factor that positively regulates the expression of genes encoding antioxidants, xenobiotic detoxification enzymes, and drug efflux pumps, and confers cytoprotection against oxidative stress and xenobiotics in normal cells. Kelch-like ECH-associated protein 1 (KEAP1) negatively regulates NRF2 activity by targeting it to proteasomal degradation. Increased expression of cellular antioxidants and xenobiotic detoxification enzymes has been implicated in resistance of tumor cells against chemotherapeutic drugs. METHODS AND FINDINGS: Here we report a systematic analysis of the KEAP1 genomic locus in lung cancer patients and cell lines that revealed deletion, insertion, and missense mutations in functionally important domains of KEAP1 and a very high percentage of loss of heterozygosity at 19p13.2, suggesting that biallelic inactivation of KEAP1 in lung cancer is a common event. Sequencing of KEAP1 in 12 cell lines and 54 non-small-cell lung cancer (NSCLC) samples revealed somatic mutations in KEAP1 in a total of six cell lines and ten tumors at a frequency of 50% and 19%, respectively. All the mutations were within highly conserved amino acid residues located in the Kelch or intervening region domain of the KEAP1 protein, suggesting that these mutations would likely abolish KEAP1 repressor activity. Evaluation of loss of heterozygosity at 19p13.2 revealed allelic losses in 61% of the NSCLC cell lines and 41% of the tumor samples. Decreased KEAP1 activity in cancer cells induced greater nuclear accumulation of NRF2, causing enhanced transcriptional induction of antioxidants, xenobiotic metabolism enzymes, and drug efflux pumps. CONCLUSIONS: This is the first study to our knowledge to demonstrate that biallelic inactivation of KEAP1 is a frequent genetic alteration in NSCLC. Loss of KEAP1 function leading to constitutive activation of NRF2-mediated gene expression in cancer suggests that tumor cells manipulate the NRF2 pathway for their survival against chemotherapeutic agents.

A more recent pubmed abstract on the same topic:

Cancer Res. 2008 Oct 1;68(19):7975-84. <script language="JavaScript1.2">Links

RNAi-mediated silencing of nuclear factor erythroid-2-related factor 2 gene expression in non-small cell lung cancer inhibits tumor growth and increases efficacy of chemotherapy.
Singh A, Boldin-Adamsky S, Thimmulappa RK, Rath SK, Ashush H, Coulter J, Blackford A, Goodman SN, Bunz F, Watson WH, Gabrielson E, Feinstein E, Biswal S. Department of Environmental Health Sciences, Division of Toxicology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.

Nuclear factor erythroid-2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that regulates the expression of electrophile and xenobiotic detoxification enzymes and efflux proteins, which confer cytoprotection against oxidative stress and apoptosis in normal cells. Loss of function mutations in the Nrf2 inhibitor, Kelch-like ECH-associated protein (Keap1), results in constitutive activation of Nrf2 function in non-small cell lung cancer. In this study, we show that constitutive activation of Nrf2 in lung cancer cells promotes tumorigenicity and contributes to chemoresistance by up-regulation of glutathione, thioredoxin, and the drug efflux pathways involved in detoxification of electrophiles and broad spectrum of drugs. RNAi-mediated reduction of Nrf2 expression in lung cancer cells induces generation of reactive oxygen species, suppresses tumor growth, and results in increased sensitivity to chemotherapeutic drug-induced cell death in vitro and in vivo. Inhibiting Nrf2 expression using naked siRNA duplexes in combination with carboplatin significantly inhibits tumor growth in a subcutaneous model of lung cancer. Thus, targeting Nrf2 activity in lung cancers, particularly those with Keap1 mutations, could be a promising strategy to inhibit tumor growth and circumvent chemoresistance.

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Posted 31 December 2008 - 10:31 PM

Oncogene. 2005 Oct 20;24(46):6957-69. Links
Zerumbone abolishes NF-kappaB and IkappaBalpha kinase activation leading to suppression of antiapoptotic and metastatic gene expression, upregulation of apoptosis, and downregulation of invasion.
Takada Y, Murakami A, Aggarwal BB. Cytokine Research Laboratory, Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Box 143, Houston, TX 77030, USA. Zerumbone found in subtropical ginger Zingiber zerumbet Smith exhibits antiproliferative and antiinflammatory activities but underlying molecular mechanisms are poorly understood. As several genes that regulate proliferation and apoptosis are regulated by nuclear factor (NF)-kappaB, we hypothesized that zerumbone mediates its activity through the modulation of NF-kappaB activation. We found that zerumbone suppressed NF-kappaB activation induced by tumor necrosis factor (TNF), okadaic acid, cigarette smoke condensate, phorbol myristate acetate, and H2O2 and that the suppression was not cell type specific. Interestingly, alpha-humulene, a structural analogue of zerumbone lacking the carbonyl group, was completely inactive. Besides being inducible, constitutively active NF-kappaB was also inhibited. NF-kappaB inhibition by zerumbone correlated with sequential suppression of the IkappaBalpha kinase activity, IkappaBalpha phosphorylation, IkappaBalpha degradation, p65 phosphorylation, p65 nuclear translocation, and p65 acylation. Zerumbone also inhibited the NF-kappaB-dependent reporter gene expression activated by TNF, TNFR1, TRADD, TRAF2, NIK, and IKK but not that activated by the p65 subunit of NF-kappaB. NF-kappaB-regulated gene products, such as cyclin D1, COX-2, MMP-9, ICAM-1, c-Myc, survivin, IAP1, IAP2, XIAP, Bcl-2, Bcl-xL, Bfl-1/A1, TRAF1 and FLIP, were all downregulated by zerumbone. This downregulation led to the potentiation of apoptosis induced by cytokines and chemotherapeutic agents. Zerumbone's inhibition of expression of these NF-kappaB-regulated genes also correlated with the suppression of TNF-induced invasion activity. Overall, our results indicated that zerumbone inhibits the activation of NF-kappaB and NF-kappaB-regulated gene expression induced by carcinogens and that this inhibition may provide a molecular basis for the prevention and treatment of cancer by zerumbone.

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Posted 31 December 2008 - 10:45 PM

Carcinogenesis. 2008 Apr;29(4):779-89. Epub 2007 Nov 13. Links
Curcumin downregulates the inflammatory cytokines CXCL1 and -2 in breast cancer cells via NFkappaB.
Bachmeier BE, Mohrenz IV, Mirisola V, Schleicher E, Romeo F, Höhneke C, Jochum M, Nerlich AG, Pfeffer U. Department of Clinical Chemistry and Clinical Biochemistry, Ludwig-Maximilians-University Munich, D-80336 München, Germany. bachmeier@med.uni-muenchen.de The dietary antioxidant Curcumin has been proposed for cancer chemoprevention since it induces apoptosis and inhibits the formation of breast cancer metastases. Curcumin acts through the inhibition of phosphorylation of the inhibitor of kappa B (IkappaB), which in turn reduces the nuclear translocation of nuclear factor kappa B (NFkappaB), an inflammation- and cell survival-related transcription factor. However, it is not clear whether the strong antimetastatic effect can exclusively be explained by inhibition of NFkappaB. Here, we addressed the effects of Curcumin (IC(50) = 17 muM) in MDA-MB-231 breast cancer cells using microarray gene expression analyses. Among the 62 genes whose expression was significantly altered, we found the two inflammatory cytokines CXCL1 and -2 (Groalpha and -beta) that were downregulated. Further validation of the microarray results by quantitative real-time reverse transcription-polymerase chain reaction, western blots and enzyme-linked immunosorbent assay revealed that Curcumin impairs transcription of CXCL1 and -2 >24 h and reduces the corresponding proteins. Using small interfering RNA techniques, we elucidated the underlying molecular mechanism revealing that reduction of CXCL1 and -2 messenger RNA levels is NFkappaB dependent and requires intact IkappaBalpha expression. Moreover, CXCL1 and -2 silencing leads to downregulation of several metastasis-promoting genes among which we found the cytokine receptor CXCR4. We therefore suggest that the decrease of CXCL1 and -2 mediated by Curcumin is involved in the inhibition of metastasis.

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Posted 01 January 2009 - 01:45 AM

Doesn't mention NF-kappaB explicitly, but considering previous posts on NF-kappaB, I guess this is sufficiently related:

Public release date: 31-Dec-2008
[ | E-mail Article ]

Contact: Debra Kain
ddkain@ucsd.edu
619-543-6163
University of California - San Diego
<h1 class="title">Lung cancer cells activate inflammation to induce metastasis</h1> A research team from the University of California, San Diego School of Medicine has identified a protein produced by cancerous lung epithelial cells that enhances metastasis by stimulating the activity of inflammatory cells. Their findings, to be published in the January 1 issue of the journal Nature, explain how advanced cancer cells usurp components of the host innate immune system to generate an inflammatory microenvironment hospitable for the metastatic spread of lung cancer. The discovery could lead to a therapy to limit metastasis of this most common lethal form of cancer.

The scientists – headed by Michael Karin, Ph.D., UC San Diego Distinguished Professor of Pharmacology and Pathology, who has been investigating the effects of inflammation on cancer development and progression – used a straightforward biochemical approach to identify proteins produced by metastatic cancer cells that are responsible for generation of an inflammatory microenvironment that supports the growth of metastases. Focusing on macrophages, white blood cells that are key players in the immune response to foreign invaders as well as in cancer growth and progression, they screened for factors produced by metastatic cancer cells in mice that could stimulate the activity of this inflammatory cell type.

Among the mouse cell lines screened, a highly metastatic cell line called Lewis lung carcinoma (LLC) showed particularly potent activation of macrophages. Furthermore, macrophage activation was mediated by a secreted protein. Biochemical purification of proteins secreted by LLC cells resulted in identification of an extracellular matrix protein called versican as the major macrophage activator and metastasis enhancing factor. Versican is also found in very low amounts in normal human lung epithelial cells, but is upregulated in human lung cancer, where a very large amount of this protein is found, especially in aggressive tumors.

The scientists found that versican strongly enhances LLC metastatic growth by activating receptors that lead to production of cytokines – signaling proteins that regulate the immune system. One of these receptors, TLR2, and a cytokine, TNFα, were found to be required for LLC metastasis. However, the normal function of TLR2 and TNF is in host defense-innate immunity to microbial infections. According to Karin, these findings are relevant, not just to the mouse model, but also to human lung cancer – the most common cause of cancer-related deaths worldwide. The major cause of lung cancer is tobacco smoking.

"By usurping these elements of the host immune system, versican helps generate an inflammatory environment that spurs the growth and spread of metastatic cancer," said Karin. "If we can find a way to block the production of versican or its binding to TLR2, therapeutic intervention could be used to limit metastasis of lung cancer."

Edited by Shinigami, 01 January 2009 - 01:57 AM.

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Posted 01 January 2009 - 01:50 AM

Mol Pharmacol. 2006 Jan;69(1):195-206. Epub 2005 Oct 11. <script language="JavaScript1.2">Links
Curcumin (diferuloylmethane) down-regulates expression of cell proliferation and antiapoptotic and metastatic gene products through suppression of IkappaBalpha kinase and Akt activation.
Aggarwal S, Ichikawa H, Takada Y, Sandur SK, Shishodia S, Aggarwal BB. Cytokine Research Laboratory, Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030-4009, USA. Curcumin (diferuloylmethane), an anti-inflammatory agent used in traditional medicine, has been shown to suppress cellular transformation, proliferation, invasion, angiogenesis, and metastasis through a mechanism not fully understood. Because several genes that mediate these processes are regulated by nuclear factor-kappaB (NF-kappaB), we have postulated that curcumin mediates its activity by modulating NF-kappaB activation. Indeed, our laboratory has shown previously that curcumin can suppress NF-kappaB activation induced by a variety of agents (J Biol Chem 270:24995-50000, 1995). In the present study, we investigated the mechanism by which curcumin manifests its effect on NF-kappaB and NF-kappaB-regulated gene expression. Screening of 20 different analogs of curcumin showed that curcumin was the most potent analog in suppressing the tumor necrosis factor (TNF)-induced NF-kappaB activation. Curcumin inhibited TNF-induced NF-kappaB-dependent reporter gene expression in a dose-dependent manner. Curcumin also suppressed NF-kappaB reporter activity induced by tumor necrosis factor receptor (TNFR)1, TNFR2, NF-kappaB-inducing kinase, IkappaB kinase complex (IKK), and the p65 subunit of NF-kappaB. Such TNF-induced NF-kappaB-regulated gene products involved in cellular proliferation [cyclooxygenase-2 (COX-2), cyclin D1, and c-myc], antiapoptosis [inhibitor of apoptosis protein (IAP)1, IAP2, X-chromosome-linked IAP, Bcl-2, Bcl-x(L), Bfl-1/A1, TNF receptor-associated factor 1, and cellular Fas-associated death domain protein-like interleukin-1beta-converting enzyme inhibitory protein-like inhibitory protein], and metastasis (vascular endothelial growth factor, matrix metalloproteinase-9, and intercellular adhesion molecule-1) were also down-regulated by curcumin. COX-2 promoter activity induced by TNF was abrogated by curcumin. We found that curcumin suppressed TNF-induced nuclear translocation of p65, which corresponded with the sequential suppression of IkappaBalpha kinase activity, IkappaBalpha phosphorylation, IkappaBalpha degradation, p65 phosphorylation, p65 nuclear translocation, and p65 acetylation. Curcumin also inhibited TNF-induced Akt activation and its association with IKK. Glutathione and dithiothreitol reversed the effect of curcumin on TNF-induced NF-kappaB activation. Overall, our results indicated that curcumin inhibits NF-kappaB activation and NF-kappaB-regulated gene expression through inhibition of IKK and Akt activation.

"Glutathione and dithiothreitol reversed the effect of curcumin on TNF-induced NF-kappaB activation." --> Indicating the importance of redox status on this process?

#89 davidd

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328 posts
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Location:Minnesota

Posted 14 January 2009 - 06:12 PM

Hello,

All my thunder has already been stolen in the final posts, but I'll add my 2 cents anyway.

I am glad Vitamin D has been investigated. Good stuff.

You mentioned bacterial infection part way through and you mentioned zinc supplementation helping you in another thread. This could be related, as zinc is antibacterial/antiviral. Or it could just be zinc's immune system boosting ability.

It would appear zinc has a NF-kappaB relationship as well: http://cat.inist.fr/...cpsidt=15017923

...
Zinc-induced NF-κB inhibition can be modulated by changes in the intracellular metallothionein level

Metallothionein (MT), a small metal-binding protein, is involved in the regulation of cellular metal homeostasis. Sequestration and the release of metals to and from MT plays an important role in the attenuation or amplification of signal transduction. Zinc has been suggested to be an important regulator of nuclear factor KB (NF-κB). In this study, the effect of MT expression on the zinc-induced inhibition of NF-κB activity was examined. In HeLa cells, pyrrolidine dithiocarbamate (PDTC), a zinc ionophore, and zinc itself inhibited NF-κB activity. When the cells were pretreated with MT-inducers, cadmium, or dexamethasone. PDTC did not inhibit NF-KB activity. We transfected HeLa cells with a DNA construct in which expression of MT-IIA is controlled by tet operator protein. Treatment of HeLa cells with doxycycline, a tetracycline analogue, induced the expression of MT-IIA, which attenuated the effect of PDTC on NF-κB activity. These results implicate MT in the zinc regulation of NF-KB and identify MT as one of the potential intracellular modulators of NF-KB activation.
...

That brings Metallothionein into the mix as well. I'm not sure if you touched on that, since I was mostly scanning for quercetin/D3/zinc in your posts.

Lastly, I'll second what 100YearsToGo about quercetin. It is antiflammatory. In addition, it doesn't just block histamine, like many of the drugs do, it reduces the creation of it in the first place (if my memory is correct). Lastly, it is tied to NF-kappaB as well (which you pointed out in one study). Here is another: http://www.ncbi.nlm....pubmed/16177186

...
Quercetin decreases oxidative stress, NF-kappaB activation, and iNOS overexpression in liver of streptozotocin-induced diabetic rats.

Increasing evidence in both experimental and clinical studies suggests that oxidative stress is involved in the pathogenesis and progression of diabetic tissue damage. This study investigated the protective effects of quercetin treatment on oxidative stress, nuclear factor (NF)-kappaB activation and expression of inducible nitric oxide synthase (iNOS) in streptozotocin-induced diabetic rats. Male Wistar rats were divided into 4 groups: control rats, control rats treated daily with quercetin (150 micromol/kg, i.p.), untreated diabetic rats, and diabetic rats treated with quercetin. Diabetes was induced by a single i.p. injection of streptozotocin (70 mg/kg). Eight weeks later we measured TBARS and hydroperoxide-initiated chemiluminescence (QL) in liver as markers of oxidative stress, and activities of the antioxidant enzymes catalase, superoxide dismutase (SOD), and glutathione peroxidase, NF-kappaB activation by an electrophoretic mobility shift assay and expression of IkappaB kinases (IKKalpha and IKKbeta), the inhibitor IkappaB (IkappaBalpha and IkappaBbeta), and iNOS by Western blot. The plasma glucose concentration was significantly increased in diabetic rats and was not changed by quercetin. Streptozotocin administration induced significant increases in hepatic TBARS concentration, QL, and SOD and catalase activities that were prevented by quercetin. Activation of NF-kappaB, induction of IKKalpha and iNOS protein levels, and increased degradation of IkappaBalpha were also observed in streptozotocin-treated rats. All of those effects were abolished by quercetin. These findings suggest that quercetin treatment, by abolishing the IKK/NF-kappaB signal transduction pathway, may block the production of noxious mediators involved in the development of early diabetes tissue injury and in the evolution of late complications.
...

And I'll also second that eating apples isn't going to give you the same amount of quercetin as supplements.

I took 1500 mg/day (750 morning and 750 night) for almost 3 months this past summer. That also included 750 mg of bromelain/day (approx. 17 mg/kg and 9 mg/kg respectively). I give between 7 to 13mg/kg of quercetin (3-6 mg/kg bromelain) to my daughter for alergies, as do many parents. I do give her a break from it for several weeks periodically though.

Lastly, quercetin does induce at least one phase two detoxifying enzyme, UGT1A1. One of your posts mentioned phase 2 enzymes.

David