International MPN News, Science & Opinion

Blood and Bone – the factory floor


The blood-bone niche and MPNs

By Zhenya Senyak

Blood cells seen through an electron scanning microscope

The identification of skeletal progenitor cells in the human bone marrow…has coincided with the recognition that the ability to transfer the hematopoietic microenvironment is an inherent property of skeletal progenitor cells…., these cells are pivotal organizers of the hematopoietic microenvironment. Bianco P, Sacchetti B, Riminucci M .Best Practice & Research Clinical Haematology, March 2011Osteoprogenitors and the hematopoietic microenvironment.” http://www.ncbi.nlm.nih.gov/pubmed/21396591

It ain’t necessarily so.

What we believe about the origin of our blood disorders, what we think about the very process of blood production itself (hematopoeisis) is undergoing rapid change.

What will they think of us down the road as genetic discoveries uncover new realities of myeloproliferative neoplasms? .

Not so long ago hematologists believed red blood cells came about from the disintegration of white cells. The process, according to one theory advanced by Georges Pouchet right before the dawning of the 20th Century was “hemoglobinic degeneration.” And Georges Hayem, a hematologist credited with the first accurate count of blood platelets, thought those same platelets were the source of RBCs. .

For all our glittering technology, we may be just as deluded. Our modern characterization of MPNs will almost certainly be revised.

Symptoms vs. causes

Although our blood cells may be proliferating in frenzied abandon, it may not be entirely accurate to say our chronic MPNs are blood diseases.. The anomalies in our blood – too many RBCs, too many platelets, etc. – are symptoms of an underlying stem cell dysfunction. It’s a bit like saying an old car that emits a trail of blue smoke from the exhaust and is burning oil has an oil problem. It would be more accurate to say the cylinders have a piston ring problem. Or even more fundamentally, the driver has a car problem.

It wouldn’t matter except placing the focus on the symptom instead of the cause can lead us up many a blind alley and leave the real problem unsolved. As it is we take our MPNs to blood doctors, hematologists, who perform blood tests to assess the state of our blood. They then prescribe drugs that to some extent or other allay the symptoms and related clinical effects. No one pretends the drugs are curative; Stop taking them and the symptoms return. To return to our old clunker, that’s like adding oil conditioner and a sealant to your crankcase along with a quart of oil once a month. It’s not elegant but it is a solution of sorts for awhile. Where do we go to solve the problem itself?

Bone: An unindicted co-conspirator

There may be a co-conspirator in our MPNs that has largely escaped identification. The origin of our MPNs is in hematopoietic stem cells that share a micro-environment with skeletal stem cells – a niche — that influences both bone and blood. They are co-dependent.

It’s a current line of research that’s gaining momentum since direct observation seems to prove it out: Dysregulation of the skeletal stem cells dysregulates blood stem cells. It’s that direct. The hematopoietic niche in this physical place within the bone medullary capsule “provides functional instruction for stem cells…” The quotes, are from Paolo Bianco’s seminal “Bone and the hematopoietic niche: a tale of two stem cells” that appeared earlier this year in Blood http://bloodjournal.hematologylibrary.org/content/117/20/5281.full

This niche of bone and blood stem cells within the marrow compartment is emerging as a possible target in developing future therapeutic approaches to MPNs.

One outcome of the current flurry of MPN clinical trials is the growing knowledge that the MPN target is elusive and it is likely some combination of drugs will be necessary to relieve symptoms. However, the drugs we are given to reduce our MPN symptoms – both the ancient standby, hydroxyurea acting on RNA transcription or the newest clinical trial stars like Ruxolitinib and Cytopia — that target the JAK-STAT pathway treat the proliferation issue, do not affect the disease state itself.

Every drug used in treating MPNs is palliative only, not curative. The only cure so far is a radical resetting of the niche by clearing out the marrow and introducing new stem cells that did not participate in the events that created the disease state. And bone stem cells are part of that niche. To actually cure MPNs, exploring the micro-environment in which it arose is the subject of extensive research

When a busy Neighborhood Goes Bad..

Consider that hematopoietic micro-environment. It’s busy, almost beyond belief. Best current estimates are 500 billion blood cells are created – and eliminated – in each of us every day. Nearly 2.5 million red blood cells are created each second. While this is taking place, bone cells are continually remodeling the niche, opening new sinuses, depositing collagen rich materials to make repairs or modify the shape of the environment.

Most of the production and destruction surrounds red blood cells, those flexible, concave and castrated cells whose nucleus has been removed to make room for its freight of oxygen-bearing heme. There is an urgency to the work since every living cell in the body needs to breathe and be nourished. Each RBC tours the body every 20 seconds and lives for nearly three months. Along the way it delivers its load of oxygen to every cell by squeezing through narrow capillaries that force discharge of its load into water based connective tissue that surrounds cells. After delivery it speeds on to the lungs to replenish its supply. We have a lot of cells; we need a lot of RBCs.

Does that immense traffic, the constant birth and destruction, cause scarring within the hematopoietic niche?

Chances are you were told fibrosis is scarring caused by overproduction of blood cells. Is that true? In the classic sense of scarring being the result of wound repair as part of the healing process, the build-up of fibrosis within the bone capsule doesn’t seem to qualify

What is fibrosis?.

While it is true that fibroblasts become active and multiply during healing — and the scar tissue they produce to replace damaged tissue is collagen — the two basic types of fibrosis within the bone marrow are different in both origin and weave from scar tissue..

Reticular fibrosis within the bone is an inevitable outcome of simply being alive. The older you are the more of these thin black fibers accumulate, an accretion of reticular fiber, broken off connective tissue, remnants of tissue washing up in the marrow capsule of bone, along with a net of parallel fibers produced by stimulation of fibroblasts by growth factor beta and platelet-derived growth factor. (Reticular fibers, made of collagen III, are net-like and appear throughout the body supporting organs and other structures.)

Another kind of fibrosis, composed of the more ominous collagen I fiber, is completely different Collagen fibrosis stuffs up the sinusoids, the blood-marrow exchange holes, crowds out productive stem cells and impairs hematopoiesis. Why does it happen? One theory implicates the increased presence of megakaryocytes with weird cargo. Another, inflammation. What we can be pretty sure of is fibroblasts can be stimulated to multiply and increase their output in response to environmental signals. And when active, fibroblasts produce and secrete gobs of collagen. Another on-going source of collagen within the marrow is the output of osteoblasts whose sole function is to deposit collagen in continual bone remodeling projects.,

. Blood stem cells don’t always remain at home in the bone marrow. And once they leave the bone-blood niche the results can be disastrous. In normal circumstances, for humans, bone marrow is the seedbed of hematopoeisis. When circumstances aren’t normal, when the marrow environment is compromised, blood stem cells move on to other locations.

Hematopoietic stem cells choose their location on a competitive basis. And when the marrow becomes inhospitable, the HSCs continue their work elsewhere, so long as there is a tunnel, a sinus – a sinusoidal connection between the breeding space and the blood system — as in bone, the spleen, liver or lymph system. .With a production quota of 500 billion a day there’s no time for a coffee break.

The… most recent mouse model of myelofibrosis has been generated by manipulating genes that control the ability of the marrow microenvironment to support hematopoiesis. In this model, development of myelofibrosis is not due to a stem cell defect but rather to a microenvironmental defect. As such, the MKs [ed. Megakaryocytes] of these mice are normal. However the microenvironment is not normal and presents many of the alterations characteristic of the microenvironment of PMF patients. This last model highlights the fact that PMF is a disease of abnormal interactions between the stem cells and their niches. [Emphasis added.]

Lilian Varricchio, A. Mancini and A.R. Migliaccio, Department of Medicine, Division of Hematology/Oncology, Mount Sinai School of Medicine, Expert Rev Hematol. 2009 June 1; 2(3): 315–334. doi: 10.1586/ehm.09.17

Bone – an active participant in hematopoiesis

Considering bone an innocent and fixed bystander in the blood production process is to miss the nature of bone itself. The obviously hard substance of bone that makes up the visible skeleton is only the outside of the bone, the extracellular matrix, essentially collagen and mineralized elements. Within it’s a plastic, evolving complex world.

Bone is basically a hollow vessel built up of concentric communicating rings.filled with supporting blood vessels, fat, marrow and a branching network of supportive thin bone collagen (trabechiae) The hematopoietic compartment exists with parts the axial skeletal

Within and around the inner surface of the bone capsule – the endosteum — are the bone stem cells, the skeletal stem cells, along with progenitor cells and their progenies, working side by side with HSCs. Literally.

The bone cells, osteocytes and osteoblasts and osteoclasts combine to shape bone: Osteocytes, the most numerous, are the end stage of an osteoblast that gets buried in its own collagenous secretions.

The population of blood stem cells are correlated with the population of the osteoblastic bone cells. While osteoblasts clearly can control the blood production niche, they are short lived and exist only on the endosteum, the thin layer of tissue that lines the surface of the hollow portion of bone, the medullary cavity. And at that, osteoblasts remain there only when the bone cavity is undergoing growth or remodeling.

Osteoclast at work

Osteocytes, although buried in bone matrix are linked through microscopic canals, lacunae, by threads of cytopolasm. They are thought to play a regulatory role in the the bone remodeling process by breaking down bone. Osteoclasts however are the heavy lifters.

The osteoclast is known to be formed by fusion of circulating mononuclear precursor cells of haematopoietic origin

Y Fukikawa, University of Oxford, United Kingdom

The clawing, chomping and bulldozing required to excavate sinusoids and clear openings for blood and marrow to exchange elements. is done by the osteoclast, a large cell with multiple nuclei found sitting by itself alongside its construction pit. And while osteocytes and osteoblasts may be indistinguishable from one another and in fact become one another at different stage of maturation, the osteoclast is a distinct population descended from the same precursor cells as macrophages, the bacteria gobbling white blood cells.

Bone and blood come together here as well since osteoclasts and macrophages are, genetically, kissing cousins.

.Osteoclasts are derived from hemopoietic stem cells and play critical roles in bone resorption and remodeling.

– Shin-Ichi Hayahi, Journal of Immunology

Rebirth

If reincarnation is a reality and you have a choice, you probably wouldn’t want to select life as an osteoblast. These little guys spend all their days in a microscopic area of bone continually generating and depositing bone forming materials, mostly collagen. And usually bury themselves in their own secretions. After a few weeks they’re given the signal to die. They turn on the appropriate genes and either commit hari kari or turn into one of two other barely preferable afterlife options. They become an osteocyte, entombed in mineralized bone until freed during an osteoclast bone remodeling project, or serve as a flattened carpet covering inert bone surface.

No, if you want to be part of the action, head for the sinusoids.

Sinusoids are a prerequisite for the establishment of hematopoeisis. Wherever blood production takes place there must be an opening, a tunnel between the niche and the circulatory system.. Bone marrow sinusoids are greater than 50 micrometers wide thin-walled blood vessels that open directly. Lined by endothelial cells and covered by skeletal stem cells over 60% of their surface, “sinusoid walls are traversed by hematopoitetic cells in both directions (blood to marrow and vice versa.”

The sinusoid is the interstate highway for hematopoiesis…and a hotbed of creation. It is a highway carved by osteoclasts and paved with skeletal stem cells. Clog the opening with collagen fibers and the rate of hematopoeisis diminishes.

Step into the sinusoid because here at last we come to the heart of the HSC niche, the holy of holies of hematopoeisis. The walls and floor and ceiling are lined with endothelial cells as are all blood vessels and within the walls are osteoprogenitors, skeletal stem cells. In the words of Bianco, “they represent the first-line extravascular environment seen by immigrant blood borne cells, with which they establish cell-cell contacts, which are integral to a niche effect.” And alongside those osteoprogenitors, the walls of the sinusoid are thickly studded with hematopoietic stem cells.

Where, exactly is this hematopoietic niche?

The hematopoietic niche is the sinusoidal wall and the endosteum – the thin layer of endothelial connective tissue lining the marrow compartment of the bone. In this niche, the meeting place of blood and bone stem cells, an on-rushing generation of blood cells is continually produced at a prodigious rate.

It is here that the blood stem cells reproduce and differentiate, here that mutations causing myeloproliferative neoplasms arise and it is here that the mutated clone is propagated.

Two final questions, one surprising answer

Two questions, at least, remain: if a specific MPN phenotype arises in the niche, why that one and not another, why PV and not ET? And, since stem cells continually renew themselves, why are MPNs generally limited to middle-aged and older populations?

Here’s a heresy. Hematopoietic stem cell are not identical. All blood stem cells are not the same. Those differences and the changing ability of the niche to support stem cells as individuals age might account for some characteristics of MPNs.

Grant Challen of the Baylor College of Medicine, isolated mouse stem cells and discovered they accepted a dye at two different rates. Sorting these cells into two classes and transplanting several hundred into mice lacking blood stem cells.his group discovered all blood stem cells were not created equal.

While each HSC was able to produce all types of blood cells they preferentially produced certain blood cells. One type of stem cell produced more RBCs and the other more immune blood cells.

Significantly, as these mice aged, the stem cells that created more RBCs began to dominate. “We believe, said Challen, ” that this phenomenon we discovered – that myeloid stem cells become increased over time – may be related to some of the myeloid proliferative diseases associated with the elderly.”

It’s possible. But deep genetic research is still in its infancy and with theories and new drugs abounding, it seems only reasonablel to question everything, take what works, and keep an eye on the research labs.

Hematopoiesis is strictly dependent on the establishment of appropriate interactions between stem cells and their niches in the marrow…. PMF is a disease in which the interactions between the stem cells and their niches in the marrow are disrupted. Therefore, the comparison of the biological properties of the stem cells and of the marrow microenvironment between PMF patients and normal donors provides the necessary tool to understand how the hematopoietic process is regulated in humans.. [Ed. Emphasis added.]

Annalisa Mancini, Anna Rita Migliaccio, Lilian Varricchio, Department of Medicine, Division of Hematology/Oncology, Mount Sinai School of Medicine, Expert Rev Hematol. 2009 June 1; 2(3): 315–334.

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© Zhenya Senyak and MPNforum.com, 2011. Unauthorized use and/or duplication of this material without express and written permission from this blog’s author and/or owner is strictly prohibited. Excerpts and links may be used, provided that full and clear credit is given to Zhenya Senyak and MPNforum.com with appropriate and specific direction to the original content.

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