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Stem Cells


Proton Head

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Instead of wasting time on arguing about the use of stem cells, why don't people try and find methods to reverse the specialization of differentiated cells.

That would give us access to stem cells without the need for cloning embryos.

 

What's the big deal. All you have to do is determine the chromosome structure, protein - content and localization of a stem cell.

 

Then you just induce some changes on whatever cells you have at hand and there you go.

 

Its so easy you know.

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I don't think it would be as easy as you are making it out to be. It's much easier to take stem cells and turn them into specialized cells than it is to take specialized cells and turn them into stem cells. The cells are already preprogrammed to become specialized when exposed to certain hormones and there are built-in mechanisms to carry out the specialization. There are no mechanisms for turning the specialized cells back into stem cells and this kind of procedure is way beyond current knowledge or technology. (disregarding any unusual cases where specialized cells naturally turn back into stem cells if there is such a thing.)

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Instead of wasting time on arguing about the use of stem cells' date=' why don't people try and find methods to reverse the specialization of differentiated cells.

That would give us access to stem cells without the need for cloning embryos.

 

What's the big deal. All you have to do is determine the chromosome structure, protein - content and localization of a stem cell.

 

Then you just induce some changes on whatever cells you have at hand and there you go.

 

Its so easy you know.[/quote']

 

Thx to our buddy introns and extrons, gene expression makes it very difficult to reverse the process. Imagine taking a 3,200,000 word essay, randomly cutting out words so only 320,000 are left, and then try to piece it back together. (It would not be fun)

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I agree its not going to be easy, but a time will come when we need to develop such methods, so instead of just complaining why don't give it a try.

 

I mean basically if we disregard the different organelle and protein content of specialized cells compared to stem cells (something which would probably change over time if we could make specialized cells express their genome the way stem cells do), the only difference between stem and specialized cells is the difference in their chromosome arrangement.

 

This difference in chromosome structure is largely determined by the nucleosome structures associated with DNA. Nucleosomes compose of histidine protein subunits, and the chemical modification of their N -terminal tails sticking out of the nucleosome core is a factor which largely regulates the nucleosome structure. Also associated with chromosome structure are different regulatory elements which block or assist nucleosome formation and a group of structure modifying proteins called non-histidine proteins.

 

Analyzing the stem cell nucleus protein content could give us hints as to what proteins function to organize the nucleosomes. Also analysis of the mRNA content of the stem cell could give us usefull hints of what parts of the genome are under transcription. This in turns gives light on how the nucleosome units are modified at certain areas.

 

Through trial and error we could find ways to return some specialized cells into a state (not necessarily stem cell) where they could under the information content of other tissues turn into cells of that tissue.

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  • 1 month later...

I think that as cancer research goes on, we may find out more and more about how our cells work. For instance, with diabetes I've always wondered why they can't get the islet cells in the pancreas to re-grow. I wonder why they can't investigate the cells of pancreatic cancer to see if any of them are islet cells. Then they could figure out a way to 'turn on' those cells again so that they can re-grow, then turn them off so they aren't a 'cancer'.

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  • 4 weeks later...

 

I mean basically if we disregard the different organelle and protein content of specialized cells compared to stem cells (something which would probably change over time if we could make specialized cells express their genome the way stem cells do)' date=' the only difference between stem and specialized cells is the difference in their chromosome arrangement.[/quote']

 

I think by the phrase "chromosome arrangement" you mean DNA modification. Differentiated cells are thought not to irreversibly modify their DNA. So we can assume the sequence is the same in all cells. However, rapid modifications of genes and nucleotides occurs on a second-to-second basis in cells, according to regulatory proteins in the nucleus. This allows easier or more difficult access of transcription factors to their binding sites on DNA, which has everything to do with the fate of the cells. For example, it has been shown that treatment of bone-marrow derived stem cells with a chemical that demethylates DNA (opens up sequenes for binding) stimulates cardiogenic differentiation sporadically. So the "chromosome arrangement" is actually the same, but the "organelle and protein content" of the cell is in constant flux with DNA modification.

 

It might be possible to dedifferentiate specialized cells, but it would require a disassembly of all the structural and biochemical features that cell had acquired. If a "natural" pathway does not exist for the disassembly of some of these, it would probably be difficult if not impossible to reverse the free energy cascade of the enzymatic pathways. Kind of equivalent to comparing knocking over a glass of water (loss of free energy) versus reassembling all the droplets and glass shards into the glass of water again (gain of free energy). You see the difficulty.

 

Keep coming up with ideas! We need em :)

 

Dave

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  • 3 months later...

I don't get the problem...Take egg samples from 12 women, put it on a colorful spinner. Take sperm samples from 12 guys, put it on a colorful spinner. Spin both spinners and pair them off. Make embryos. Get stem cells. Done. Where's the controversy?

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  • 2 months later...
I don't get the problem...Take egg samples from 12 women, put it on a colorful spinner. Take sperm samples from 12 guys, put it on a colorful spinner. Spin both spinners and pair them off. Make embryos. Get stem cells. Done. Where's the controversy?

It's not that easy. You still have the controvery of taking a life, which is a big no-no according to religion and the Universal Declaration of Human Rights [article 3]. I believe it's said that once fertilization occurs [involving an egg & sperm], that is when life begins.

 

Hence why researchers are using other methods such as growing stem cells from human skin or from 2 eggs. They're even exploring potential stem cells from adults [although embryos are the most versatile type].

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  • 2 years later...
I think that as cancer research goes on, we may find out more and more about how our cells work. For instance, with diabetes I've always wondered why they can't get the islet cells in the pancreas to re-grow. I wonder why they can't investigate the cells of pancreatic cancer to see if any of them are islet cells. Then they could figure out a way to 'turn on' those cells again so that they can re-grow, then turn them off so they aren't a 'cancer'.

 

The company I work for is currently preparing an animal study for the treatment of type II diabetes. The developed technology can dedifferentiate PBMCs to insulin secreting islet cells. In the pipeline is to take the dedifferentiate PBMCs to heart muscle. On step closer...

 

You do touch on a point I think is often over looked in the stem cell debate. Can they truely be controlled? Do people realize the the incredible potential of stem cells carries an equally incredible risk?

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Instead of wasting time on arguing about the use of stem cells, why don't people try and find methods to reverse the specialization of differentiated cells.

That would give us access to stem cells without the need for cloning embryos.

 

What's the big deal. All you have to do is determine the chromosome structure, protein - content and localization of a stem cell.

 

Then you just induce some changes on whatever cells you have at hand and there you go.

 

Its so easy you know.

 

I don't know how easy that is to do. Take some paint, and you can draw some pretty pictures, but you can't put the paint back in the jar. Stem cells have mechanisms to specialize into other cell types, but it might not work the other way. It might be easier to find a method to keep them from specializing than to find a way to turn them back. Only type that I know can turn back into stem cells are gametes, but that is where the controversy starts. Perhaps we could trick their precursors before they turn into sperm and eggs?

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Instead of wasting time on arguing about the use of stem cells, why don't people try and find methods to reverse the specialization of differentiated cells.

That would give us access to stem cells without the need for cloning embryos.

 

What's the big deal. All you have to do is determine the chromosome structure, protein - content and localization of a stem cell.

 

Then you just induce some changes on whatever cells you have at hand and there you go.

 

Its so easy you know.

 

Now you are into my area of expertise: stem cells.

 

1. It's not "easy" to try to de-differentiate cells. In fact, it seems that differentiation happens only one way: from undifferentiated to differentiated. For instance, skeletal muscle cells are multinucleated and there is no mechanism to get them to break apart into the mononucleated muscle stem cells. Chondrocytes are differentiated and embedded in their matrix. There is no way to get them to de-differentiate, remove the matrix, and start proliferating. It appears that in many cases the DNA is methylated in the process of differentiation, permanently shutting down those genes by that covalent bond. There isn't a way to remove those bonds.

 

2. There are stem cells in the adult. When you hear the words "stem cells" the first question you have to ask is WHICH stem cell? You are obviously talking about embryonic stem cells or ESCs for short.

 

3. The people working with ESCs have made the claim that ESCs are the ONLY stem cells capable of differentiating into all the cells in the body. They claim that adult stem cells have limited differentiation capability. There is a building body of work that there are adult stem cells that have the same differentiation capability of ESCs. Verfaillie, Prockop, Miller, and Lucas all have papers out documenting the ability of adult stem cells to differentiate into phenotypes of all 3 dermal lineages.

 

the only difference between stem and specialized cells is the difference in their chromosome arrangement.

 

Where did you get this idea? Proton Head, when you have a question regarding medicine, you need to go to PubMed and do a search. In this case, do the search on "stem, cell, epigenetic, differentiation" Below are just 2 papers in the literature:

 

1: J Cell Mol Med. 2007 Jul-Aug;11(4):602-20.

 

Programming the genome in embryonic and somatic stem cells.

 

Collas P, Noer A, Timoskainen S.

 

Institute of Basic Medical Sciences, Department of Biochemistry, Faculty of

Medicine, University of Oslo, 0317 Oslo, Norway. philippe.collas@medisin.uio.no

 

In opposition to terminally differentiated cells, stem cells can self-renew and

give rise to multiple cell types. Embryonic stem cells retain the ability of the

inner cell mass of blastocysts to differentiate into all cell types of the body

and have acquired in culture unlimited self-renewal capacity. Somatic stem cells are found in many adult tissues, have an extensive but finite lifespan and can differentiate into a more restricted array of cell types. A growing body of evidence indicates that multi-lineage differentiation ability of stem cells can be defined by the potential for expression of lineage-specification genes. Gene expression, or as emphasized here, potential for gene expression, is largely controlled by epigenetic modifications of DNA and chromatin on genomic regulatory and coding regions. These modifications modulate chromatin organization not only on specific genes but also at the level of the whole nucleus; they can also affect timing of DNA replication. This review highlights how mechanisms by which genes are poised for transcription in undifferentiated stem cells are being uncovered through primarily the mapping of DNA methylation, histone modifications and transcription factor binding throughout the genome. The combinatorial

association of epigenetic marks on developmentally regulated and

lineage-specifying genes in undifferentiated cells seems to define a pluripotent state.

 

1: Stem Cells. 2007 Jan;25(1):2-9. Epub 2006 Oct 5.

 

Concise review: epigenetic mechanisms contribute to pluripotency and cell lineage

determination of embryonic stem cells.

 

Gan Q, Yoshida T, McDonald OG, Owens GK.

 

Department of Molecular Physiology and Biological Physics, University of

Virginia, Charlottesville, Virginia 22908, USA.

 

Epigenetic mechanisms, such as histone modifications and DNA methylation, have been shown to play a key role in the regulation of gene transcription. Results of recent studies indicate that a novel "bivalent" chromatin structure marks key developmental genes in embryonic stem cells (ESCs), wherein a number of untranscribed lineage-control genes, such as Sox1, Nkx2-2, Msx1, Irx3, and Pax3, are epigenetically modified with a unique combination of activating and repressive histone modifications that prime them for potential activation (or repression) upon cell lineage induction and differentiation. However, results of these studies also showed that a subset of lineage-control genes, such as Myf5 and Mash1, were not marked by these histone modifications, suggesting that distinct epigenetic mechanisms might exist for lineage-control genes in ESCs. In this review article, we summarize evidence regarding possible mechanisms that control these unique histone modifications at lineage-control gene loci in ESCs and consider their possible contribution to ESC pluripotency. In addition, we propose a novel "histone modification pulsing" model wherein individual pluripotent stem cells within the inner cell mass of blastocysts undergo transient asynchronous histone modifications at these developmental gene loci,thereby conferring differential responsiveness to environmental cues and morphogenic gradients important for cell lineage determination. Finally, we consider how these rapid histone modification exchanges become progressively more stable as ESCs undergo differentiation and maturation into specialized cell lineages.

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ARGH! I hate reading over my posts and seeing how poorly I stated thoughts.

 

Let's try again:

The company I work for is currently preparing an animal study for the treatment of type II diabetes. The developed technology can de-differentiate PBMCs and direct them into insulin secreting islet cells. In the pipeline is to take PBMCs to heart muscle.

 

Better...but not much.

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ARGH! I hate reading over my posts and seeing how poorly I stated thoughts.

 

Let's try again:

The company I work for is currently preparing an animal study for the treatment of type II diabetes. The developed technology can de-differentiate PBMCs and direct them into insulin secreting islet cells. In the pipeline is to take PBMCs to heart muscle.

 

Better...but not much.

 

I would submit that the PBMCs are circulating undifferentiated stem cells. It is known that hematopoietic stem cells circulate. I can't find any reference to "PBMC, de-differentiate" in a PubMed search. Do you have the peer-reviewed article describing this "de-differentiation"?

 

In terms of PBMCs and islet cells, I see several articles indicating the PBMCs inhibit differentiation of islet cells, but none where PBMCs can be induced to differentiate to islet cells. Again, any peer-reviewed publications?

 

My experience is that there is a LOT of hype that comes out of companies but we don't see the solid science. Which is why so much of research in companies fails and the companies themselves go under.

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Very true!

In fact the technology was bought without solid review of the data and independant testing. So when they handed us the "Procedure" and we followed it exactly we went from Monocytes to multi-nucleated osteocytes... Not good! But research was done and corrections were made.

 

Here is one publication I had on hand:

 

Zhao Y, Glesne D, Huberman E. A human peripheral blood monocyte-derived subset acts as pluripotent stem cells. Proc Natl Acad Sci U S A. 2003; 100: 2426-2431

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Here is one publication I had on hand:

 

Zhao Y, Glesne D, Huberman E. A human peripheral blood monocyte-derived subset acts as pluripotent stem cells. Proc Natl Acad Sci U S A. 2003; 100: 2426-2431

 

Mirabelle, if we look just at the Abstract of the publication we get:

"We have identified, cultured, characterized, and propagated adult pluripotent stem cells (PSC) from a subset of human peripheral blood monocytes. These cells, which in appearance resemble fibroblasts, expand in the presence of macrophage colony-stimulating factor and display monocytic and hematopoietic stem cell markers including CD14, CD34, and CD45. We have induced these cells to differentiate into mature macrophages by lipopolysaccharide, T lymphocytes by IL-2, epithelial cells by epidermal growth factor, endothelial cells by vascular endothelial cell growth factor, neuronal cells by nerve growth factor, and liver cells by hepatocyte growth factor. The pluripotent nature of individual PSC was further confirmed by a clonal analysis. The ability to store, expand, and differentiate these PSC from autologous peripheral blood should make them valuable candidates for transplantation therapy."

 

There is nothing here about de-differentiation. Instead, it is as I said: "I would submit that the PBMCs are circulating undifferentiated stem cells." That's what the abstract of the PNAS paper says they are.

 

They are isolated using the same procedure to isolate mononuclear cells from blood: a Ficoll-Hypaque gradient. If you read the paper the authors got 2 populations -- monocytes that were about 90-95% of the population and another smaller population they called f-monocytes. It is the f-monocytes that proliferated, took over the culture, and eventually responded in differentiation assays to make new phenotypes.

 

I can understand your problem: if you didn't either 1) treat with CSF and LIF (particularly the LIF) and 2) allow the f-monocytes to overgrow the culture, then yes, you would get osteoclasts! After all, monocytes are the precursor cells to oosteoclasts.

 

As for "solid evidence", I don't see that Huberman has any other publications on these cells! He did it once and quit. If you want to see "solid review of the data" and "independent testing", look at Darwin Prockop or PA Lucas. Both labs have had many publications and collaborations with other labs that grew the same cells.

 

By the procedure in this paper, Huberman could have isolated a number of adult stem cells: Prockop's mesenchymal stem cells, Osiris' (Caplan's) mesenchymal stem cells, Goodell's "side population", Verfaillie's multipotent adult progenitor cells, or Lucas' multipotent adult stem cells. All have CD34 and Huberman did not do a more extensive CD profile of the cells (MASCs, for instance, are CD14 positive), all adhere to plastic, and all have roughly the morphology in culture of f-monocytes. The use of M-CSF and uptake of particles confuses the issue, since none of the other researchers used this growth factor or performed this assay. Therefore there is no way to tell whether the f-monocytes (or PSCs) are identical or different from the other adult stem cells.

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As I read this topic I remembered the issue that bothered me not long ago, maybe somebody here has an answer for me, I will apriciate it very much.

 

what is the difference between primery macrophages extracted from mice peritoneum, and monocytes extracted from BM of the same mice and stimulated for one week with M-CSF?!

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I would like to help you ,Tearfeather but the nomenclature is a foreign language to me. I can visualize chemicals structures much easier.

 

The best approach for making transitional cells, from stable cells, is to think in terms of hydrogen bonding. Any DNA differentiation, in terms of its active packed and unpacked shape and content, implies a different hydrogen potential configuration. The protein grid creates a H-potential capacitance, which parallels this, which causes the DNA configuration to maintain a steady state default, with some tweak, flexibility. That is how daughter cells reset the DNA. The protein grid makes the chromosomes default toward the equilibrium shape.

 

One simple technique for making transition cells, is to add the nucleus of one cell differentiation, into the protein grid of another cell differentiation. The result will be a configurational H-potential between the nucleus and the odd protein grid, with the system trying to reach a steady state. The trick is to use the two extreme H-potential states (differentiations). This gives us the widest bandwidth and the most time for manipulation.

 

If you look at stems cells advancing toward differentiated cells, the stem cells use the same principle of the nucleus and the protein grid being out of synch. The protein grid is seeing the input affect of neighboring cells, and is always one step ahead of the DNA. Eventually steady state forms, with both the DNA and the protein grid finally on the same page. At that point, we need to change partners to reset the nonequilibrium.

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I would like to help you ,Tearfeather but the nomenclature is a foreign language to me. I can visualize chemicals structures much easier.

 

The best approach for making transitional cells, from stable cells, is to think in terms of hydrogen bonding. Any DNA differentiation, in terms of its active packed and unpacked shape and content, implies a different hydrogen potential configuration. The protein grid creates a H-potential capacitance, which parallels this, which causes the DNA configuration to maintain a steady state default, with some tweak, flexibility. That is how daughter cells reset the DNA. The protein grid makes the chromosomes default toward the equilibrium shape.

 

One simple technique for making transition cells, is to add the nucleus of one cell differentiation, into the protein grid of another cell differentiation. The result will be a configurational H-potential between the nucleus and the odd protein grid, with the system trying to reach a steady state. The trick is to use the two extreme H-potential states (differentiations). This gives us the widest bandwidth and the most time for manipulation.

 

If you look at stems cells advancing toward differentiated cells, the stem cells use the same principle of the nucleus and the protein grid being out of synch. The protein grid is seeing the input affect of neighboring cells, and is always one step ahead of the DNA. Eventually steady state forms, with both the DNA and the protein grid finally on the same page. At that point, we need to change partners to reset the nonequilibrium.

 

Thank you very much for your answer :),

I think I didn't described my problem correct.

I want to know the difference between this two types of the macrophages having the same source.

Peritoneal macrophages went throw the similar maturation pathway like Bone Marrow (BM) derived ones, the difference is that BM derived cells were

maturated in vitro and Peritoneal cells were extracted from mice,

so, BM macrophages are like ''virgin'' cells that never met pathogen before, and the peritoneal cells already may have some ''life'' experience.

My question refered to people who had some experience with this two kinds of macrophages and may tell me if there is some difference in biochemical level between this two types of cells.

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I want to know the difference between this two types of the macrophages having the same source.Peritoneal macrophages went throw the similar maturation pathway like Bone Marrow (BM) derived ones, the difference is that BM derived cells were maturated in vitro and Peritoneal cells were extracted from mice, so, BM macrophages are like ''virgin'' cells that never met pathogen before, and the peritoneal cells already may have some ''life'' experience. My question refered to people who had some experience with this two kinds of macrophages and may tell me if there is some difference in biochemical level between this two types of cells.

 

I don't have the answer you want, but some insight that might help. The analogy of virgin and worldly macrophages brought an analogy to mind that seems very appropriate to the question you asked.

 

The virgin macrophages are sort of like a virgin bride, in the sense, she is a clean slate that can adapt to almost any type of husband she loves. She doesn't go into the marriage with baggage or even fixed expectations.

 

The worldly is similar to a bride who has already been married before. She has already developed a high level of adaptation. If she marries a similar type of guy, she would be up to steam, almost immediately. But if she married a guy who was different in many ways, her previous training could cause a problem adapting, since she will carry certain biases and expectations, that may not be the most appropriate for the new guy.

 

The virgin needs more time to adapt, but is capable any type of adaptation. The worldly is can adapt quickly to similar situations, but when something totally new appears, she has to both deprogram and readapt, with the result the adaptation can take longer than the virgin.

 

The body produces a constant supply of virgin macrophages, so it is ready to adapt to any new thing that comes down the pike. Although these are virgin, they need to travel within the body or blood, where they gather some indirect experience due to the immune factors that already there.

 

This is sort of like the sisters, mother and aunts of the bride, giving the bride advice and stories of what to expect, before she becomes married. But it is not first hand or worldly experience, but nevertheless can teach her many things. She is still in a fantasy world of expectations.

 

In the case of HIV, all this good advice is not appropriate. The analogy is her support staff are all married to businessmen, but her husband to be, is an artist. So she goes into the marriage, a virgin, but with too much well intentioned advice/expectation that has little to do with her husband. Under these circumstance, she may adapt easier with less advice.

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As I read this topic I remembered the issue that bothered me not long ago, maybe somebody here has an answer for me, I will apriciate it very much.

 

what is the difference between primery macrophages extracted from mice peritoneum, and monocytes extracted from BM of the same mice and stimulated for one week with M-CSF?!

 

Macrophages are fused monocytes. Macrophages are multinuclear cells and arise from the precursor monocytes. M-CSF is a mitogen for monocytes. That is, it causes them to divide (proliferate) and increases their number.

 

I understand your view of Huberman's work but since it was done at Argonne National Laboratory I'll just smile and wave at the government.

 

I worked at Argonne for 6 months as an undergraduate. The "government" has nothing to do with it! There has been some fantastic work come out of Argonne (one paper with my name on it :) ) We are dealing with this specific paper, not the institution where it was produced.

 

I would note that a lot worse work comes out of small companies.

 

One simple technique for making transition cells, is to add the nucleus of one cell differentiation, into the protein grid of another cell differentiation. The result will be a configurational H-potential between the nucleus and the odd protein grid, with the system trying to reach a steady state. The trick is to use the two extreme H-potential states (differentiations). This gives us the widest bandwidth and the most time for manipulation.

 

Pioneer, will you please stop trying to pass off your "theory" as fact? It's not. We've shown the fallacies of this idea several times.

 

BTW, what are "transition cells"?

 

If you look at stems cells advancing toward differentiated cells, the stem cells use the same principle of the nucleus and the protein grid being out of synch.

 

Oh good grief. Please stop trying to make up your own nonsensical terms. Or is your aim to confuse people about what the science really is?

 

The protein grid is seeing the input affect of neighboring cells, and is always one step ahead of the DNA. Eventually steady state forms, with both the DNA and the protein grid finally on the same page. At that point, we need to change partners to reset the nonequilibrium.

 

The correct way to say this is that stem cells respond to inductive factors that bind to specific receptors on the cell membrane. Those receptors, in turn, activate specific signalling pathways that translate the signal to the nulceus. The end result is to specifically turn some genes on and other genes off by means of transcription factors. Once high level control genes are turned on (such as Sox-9 or sonic hedgehog), the proteins made from those genes then bind to other control sites on the DNA to cause the expression of still other genes. For instance, Sox-9 controls the expression of type II collagen and proteoglycan core protein in differentiating chondrocytes. Synthesis of type II collagen and PG core protein is a major component of making a chondrocyte be a chondrocyte.

 

They should get anyomous donors of sperm and eggs and then combine them in a test tube. Out of the blue I know, but that way they can make the stem cell without them calling it human. I'm prolife by the way.

 

Uh, they already do this in making embryonic stem cells! :doh: BTW, the prolife people still insist that the resulting blastocyst is "human". You need to research what the position of "prolife" is before you accept the label.

 

However, we are talking about ADULT stem cells here. That's apples and oranges. The f-monocytes of Haberman come from the blood of adults. IOW, they can be found in YOUR blood.

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