Microglia Stem Cells

Transplanted microglia - where did they go?

Wed, 10 May 2017 15:08:34 +0000

Hi,
yes it has been a while since I wrote something here.

Here are two papers pusblished including the work funded by you:

Leovsky C, Fabian C, Naaldijk Y, Jäger C, Jang HJ, Böhme J, Rudolph L, Stolzing A. Biodistribution of in vitro-derived microglia applied intranasally and intravenously to mice: effects of aging. Cytotherapy. 2015 Nov;17(11):1617-26
Danielyan L, Beer-Hammer S, Stolzing A, Schäfer R, Siegel G, Fabian C, Kahle P, Biedermann T, Lourhmati A, Buadze M, Novakovic A, Proksch B, Gleiter CH, Frey WH, Schwab M. Intranasal delivery of bone marrow-derived mesenchymal stem cells, macrophages, and microglia to the brain in mouse models of Alzheimer's and Parkinson's disease. Cell Transplant. 2014;23 Suppl 1:S123-39.

There will be more comming on the therapeutic effect of the cells.

So if you want to speed this rpocess up we would be happy for people to help us. You just need a computer and some freely availabe software.

Or if you have a lab, especially with a good imaging software good at counting we would be interested in a collaboration.

Also we have protein extractions and would be interested in people doing proteomics. Just send me an email and we can discuss details.

Dey

Amyloid load

Mon, 08 Oct 2012 22:14:00 +0000

As for the effect the transplanted microglia have on the pathology: I do not see a big difference in the amount of amyloid in the Alzheimer mouse brain without cells (A) and with microglia transplanted (B). So we have to wait until we get all brains analysed by sterology for the exact amyloid quantification.

Greetings,
Dey

Some transplantations done - more to come

Mon, 08 Oct 2012 22:02:00 +0000

Hi again,
so we have started with our transplantations and some of our animals have been analysed. You see a picture showing our transplanted microglia in green and the amyloid in red. You can notice that the microglia are around and in the vessels. We also know from our PCR data that we get quite some microglia into the brain. This data was gathered 4 weeks after systemic transplantation of our microglia.

Brain histology - microglia

Sun, 20 May 2012 22:37:00 +0000

To visualize microglia and amyloid plaques in vivo, we established different staining protocols (including histology and immunohistology) to later evaluate microglia number after transplantation and also amyloid load. Top left is showing an immuno staining for amyloid and top right is an overview for microglia in the hippocampus.
The bottom pictures shows an example of a co-localization of activated microglia and amyloid (red = activated microglia; green: amyloid plaque; blue= tau). You can see that many activated microglia surround the amyloid plaques, however not amyloid is co-localised with microglia, so most likely the microglia surrounding the amyloid plaques did not phagocytose any of the amyloid.
Most of our measurements will take please in the hippocampus, one of the brain regions were many of the degenerative changes happen in Alzheimer. This picture shows a good overview of a hippocampus of an Alzheimer mice stained for amyloid.
In summary we have established all necessary methods for brain staining, tested the sterology method using non-transplanted mice and are now ready to transplant. We finally got the approval from our animal guys after waiting for 10 month (there are a bit over-correct here in Germany).

Charaterisation of microglia

Sat, 19 May 2012 12:02:00 +0000

We used several markers to prove that what looks like microglia really are microglia. We used so called surface markers (CDs) including CD11b and CD45. These markers are also used to distinguish between what people all macrophages and what people think microglia are (there are a lot of discussions about the identity of microglia). The cells in the red circle are considered microglia.

Selfmade microglia

Sat, 19 May 2012 11:42:00 +0000

To start with the microglia replacement therapy we need lots of cells and they must resemble the original microglia. In the pictures you see microglia isolated from mice (left) or self-made microglia derived from bone marrow (in vitro differentiated).
In both cases they have basically the same morphology. In case of the self-made microglia we only have more in the dish.

You see that they look mostly ramified (spread out), however not as much as seen in brain tissue (in vivo). You can read all the details in our publications (Hinze and Stolzing, 2011; Hinze and Stolzing, 2012).

Background

Fri, 18 May 2012 20:25:00 +0000

As a reminder WHY we are doing this:

What are microglia:

Microglia comprise about 10% of the cell population of the brain and form the main first immune defense of the CNS. They are phagocytic, cytotoxic, present antigens and they can promote repair after injury (Simard 2007).
Microglia in the brain display normally a quiescent state in which phagocytosis, immune response and migration are downregulated and the microglia show a ramified form with long processes (Davalos 2005; Nimmerjahn 2005).
Microglia react to lesions by proliferation. They migrate actively towards amyloid-β plaques in 1-2 days (Meyer-Luehmann 2008) and take on an amoeboid morphology (Kreutzberg 1996; Stence 2001)
Microglia are thought to originate from a special progenitor cell during embryogenesis and that they are replenished during life by myeloid progenitors migrating from the bone marrow which invade the brain and mature to microglia (Lawson 1992; Rezaie 2002; Chan 2007;
Simard 2007). In fact it has been shown that bone marrow cells can be differentiated to microglia (Davoust 2006).

What role do microglia play in age:

The incidence of Alzheimer and autoimmune diseases in age has been hypothesized to result from non-removed debris (Streit 2006). The common hypothesis points to a disregulation of old microglia and holds that overreactive and constantly inflamed microglia might damage neuronal tissue (McGeer 1995). Newer findings suggest that microglia associated with tau pathologies are senescent instead of reactive and unable to perform function (Streit 2009). Microglia loose their ability to recognize and digest debris during age (Stolzing 2006). However studies involving complete microglia ablation for short periods did not result in increased plaque formation or neuritic dystrophy (Grathwohl 2009).

Replenishment of microglia during lifetime has been suggested to occur locally in the brain and by invasion of bone marrow progenitor cells (Streit 2006). Bone marrow transplantation has routinely resulted in bone marrow derived microglia in the brain (Rodriguez 2007) and
intravenously injected hematopoietic stem cells have migrated to the brain, differentiated into microglia and reduced infarct. Newer findings however show the necessity of irradiation to obtain sufficient numbers of invading cells (Ajami 2007; Mildner 2007). This was suspected to be due to irradiation either raising the permeability of the blood brain barrier or to provide signals strongly enhancing proliferation. This complements the view that a possible replenishment throughout age both by local proliferation and by invasion of progenitors do not suffice to prevent the slow deterioration of microglia cell population and function in age (Stolzing 2006; Streit 2006; Flanary 2007; Sierra 2007; Sawada 2008).

Prospects and pitfalls of a microglia therapy:

Both the hypothesis of disregulated, constantly inflamed microglia and the hypothesis of microglia senescence and non sufficient replenishment suggest the introduction of functional microglia as one possible solution.
The hypothesis of bystander damage by microglia (McGeer 1995) shows that the formation of massive plaques might only be a much later appearing symptom of the functional loss of the microglia population. In this view the actual Alzheimer disease starts already with the
accumulation of relatively small amounts that act chronically inflammatory and is at the time of massive plaque formation already long at work. While dysfunctional microglia can not easily be removed from the brain the introduction of functional microglia might remove the
causes for inflammation.
The hypothesis of microglia senescence and the association of dysfunctional, senescent microglia with plaques call even more readily for a cell therapy of microglia in age. In view of both hypotheses, a treatment of the symptoms, for example through down regulation of the inflammatory response, does not remove the underlying causes of aging. Even a degradation of plaques if not properly digested by dysfunctional microglia will not enhance Alzheimer condition. Therefore function of microglia has to be restored. Experiments on the influence of irradiation in bone marrow transplantation have shown that invasion of cells in absence of irradiation is low (Ajami 2007; Mildner 2007). Studies of non ablated, syngeneic conditions have however shown that massive amounts of cells are necessary to archive comparable levels of chimerism in bone marrow (Ramshaw 1995). This points to a competition for stem cells niches which is alleviated by irradiation (Colvin 2004). In age, on the other hand, the permeability of the blood brain barrier increases which has also been linked to Alzheimer (Farrall 2009; Valle 2009).
We have worked extensively on the aging and functional loss of the microglia cell population (Stolzing 2006). I optimized differentiation of microglia from bone marrow and showed - to my knowledge for the first time - the actual functional capacity of bone marrow derived microglia
(In progress of being published). Furthermore I could show that my bone marrow derived microglia invade brain slices in vitro.

Based on the hypothesis that the functional loss of microglia in age is responsible for Alzheimer we propose to use our microglia derived from bone marrow stem cells as a therapy for Alzheimer's and other age-related neurodegeneration.

We propose to introduce our differentiated microglia intravenously. Migration to the brain is tracked by sex based RTPCR. These functional microglia might then remove and digest the inflammatory debris in the brain,
therefore remove the root cause of inflammation of aged microglia. Over a longer follow up period a down regulation of inflammation and onset of neurogenesis might take place. To assess such effects numbers, density and activation states of microglia, numbers and densities of neuronal progenitor cells and number, densities and size of plaques are measured using stereology.

Microglia Stem Cells: Community update

Fri, 18 May 2012 20:04:00 +0000

In 2011, the Longecity community raised money to support a research project on microglia stem cell therapy.

Over the next few days, we will be reporting interim results in this blog.