Could some Kleefstra changes be reversible? What a mouse model study showed

Can some of the changes caused by the deficiency of EHMT1 to the brain be corrected even after birth?;

This was the main question of an important experimental study published in iScience and used a genetically modified mouse model with a deficiency of Ehmt1, the gene associated with Kleefstra Syndrome.

The work titled Derepression of inflammation-related genes link to microglia activation and neural maturation defect in a mouse model of Kleefstra syndrome reveal a potential link between EHMT1 deficiency, epigenetic regulation, inflammatory gene activation, microglia, and neuronal maturation.

But the most interesting part was a second finding. The researchers showed that some of these biological abnormalities could be reversed when they experimentally restored EHMT1/GLP expression after birth. When the restoration was done more broadly and from a relatively young age, even certain abnormal behaviors of the mice improved.

This does not mean that a cure for Kleefstra has been found. The study was conducted on mice, and the «restoration» of EHMT1 was achieved through a special genetic laboratory system.

However, it is significant evidence that certain consequences of EHMT1 deficiency are not necessarily irreversible after birth.

What does EHMT1 do

The EHMT1 encodes a protein called GLP, which cooperates with G9a and is involved in the epigenetic regulation of gene expression. One of its key mechanisms is the generation of a modification on histones called H3K9me2. Histones are proteins around which DNA is organized. Their chemical modifications can affect how easily specific genes are activated or repressed.

Therefore, EHMT1 does not only control a single gene. It is involved in the regulation of many genes simultaneously, which helps to explain why the loss of one functional copy of it can affect many different neurodevelopmental processes. In mice with deficiency Ehmt1, researchers confirmed that the amount of GLP was reduced and that H3K9me2 levels in the brain were approximately at 70% of normal levels.

A mouse model for Kleefstra Syndrome

Researchers used mice Ehmt1Δ/+, which have only one functional copy of Ehmt1.

These animals exhibit certain characteristics that are used as laboratory counterparts of aspects of human Kleefstra, such as reduced exploratory activity and increased behaviors interpreted as anxiety-related. Naturally, a mouse cannot reproduce the full human Kleefstra syndrome. However, the model allows researchers to conduct an experiment that could not be performed in humans. To reactivate the Ehmt1 at a specific time and in specific cell types and to examine which characteristics can be corrected and which cannot.

They restored the EHMT1«

It is important to clarify what exactly the researchers did.

They did not administer an EHMT1 drug to the mice, nor did they use a treatment currently available for humans. They created a special genetically modified system in which they could experimentally activate the production of GLP, the protein encoded by the Ehmt1. In this way, they could examine whether the restoration of adequate GLP at different stages of development could correct some of the consequences of its deficiency. Ehmt1.

Rehabilitation from a young age improved certain behaviors

In one of the most important experiments, the researchers broadly activated GLP in the organism after birth, when the mice were approximately 3 to 4 weeks old.

This restoration restored H3K9me2 levels and led to an improvement in some of the abnormal behaviors exhibited by the deficient mice. Ehmt1. The result is particularly interesting because it shows that, at least in the specific animal model, the possibility of intervention is not exclusively limited to embryonic development. Some brain functions could still be affected when the Ehmt1 it was restored at a postnatal stage.

However, the story was more complex when the researchers restored GLP only in the neurons.

Restoring only the neurons was not enough for the behavior

The researchers conducted a second experiment in which they activated GLP production especially in the forebrain neurons, and again after the birth.

This intervention had clear biological effects. The reduced H3K9me2 levels in neurons returned to almost normal levels. The reduced number of dendritic spines was also restored, and the abnormal activation of microglia was reduced.

However, abnormal animal behaviors They didn't improve.

Even when the researchers waited longer after activating the Ehmt1, decreased activity and increased anxiety-like behavior remained. This is one of the most important conclusions of the work. Restoring EHMT1 in only a specific population of neurons can correct certain molecular and cellular features, but it is not necessarily enough to restore a complex function such as behavior. More cell types, more brain regions, and different developmental periods are likely involved.

An unexpected finding, microglia

One of the most interesting results of the study concerned the microglia.

Microglia essentially constitute the main immune system of the brain. Their cells are not only activated in infections or injuries. They also participate in normal brain development and the remodeling of connections between neurons. In mice with deficiency Ehmt1, the researchers observed increase of activated microglia. At the same time, gene expression analysis showed increased activation of genes related to the immune and inflammatory response.

The interesting thing is that the problem did not seem to necessarily originate from the microglia themselves.

Neurons with deficiency Ehmt1 they showed increased expression of inflammatory genes, which likely subsequently affected the behavior of microglial cells.

The role of IL-1β

The researchers focused particularly on an inflammatory cytokine called IL-1β, which is encoded by the gene IL-1b.

This pathway appeared to be upregulated in the Kleefstra model. To examine whether IL-1β was actually involved in the mechanism and was not just an incidental finding, the researchers performed an additional genetic experiment. When they knocked down the IL-1b in deficient mice Ehmt1, the increase in microglia and the change in the number of dendritic spines were corrected.

This result indicates that the IL-1β inflammatory pathway may be involved in some of the neuronal changes caused by the deficiency of Ehmt1 in this specific model.

What are dendritic spines

The dendritic spines they are microscopic protrusions on the dendrites of neurons.

In many of them, synapses are formed, meaning the points through which neurons communicate with each other. Their number, shape, and maturation change during brain development and are linked to the creation and reorganization of neural networks. In mice with a deficiency Ehmt1, the researchers found reduced number and maturation defects of dendritic spines.

Postnatal GLP restoration only in neurons was able to restore the total spine number, but it did not fully restore their morphological maturation. And this shows once again that the «reversal» was not complete. Some processes remained plastic after birth, while others appear to depend more on when the intervention takes place.

From EHMT1 to inflammatory gene expression

The study proposes a possible mechanism linking the different findings.

When there is only one functional copy of Ehmt1, the activity of the G9a/GLP complex decreases and with it the epigenetic modification decreases H3K9me2. Certain regions of the genome that are normally suppressed may thus become more active. Among the genes found to be increased were genes related to inflammatory and immune pathways, like Casp1 and the IL-1b. This could lead to increased inflammatory signaling, microglial activation, and subsequently to changes in the modulation of neuronal connections. This is a potential mechanism derived from the animal model and not a proven chain of events in the human brain.

Does this mean that Kleefstra is an «inflammatory disease»?;

No.

This specific study does not prove that individuals with Kleefstra syndrome have generalized inflammation, nor that treating inflammation is a treatment for the syndrome.

The authors explicitly acknowledge as a limitation that We do not know whether the same inflammatory genes are elevated in the neurons of individuals with Kleefstra.. Therefore, the results do not constitute a reason for using anti-inflammatory drugs, supplements, or other interventions in individuals with Kleefstra. The finding is currently mainly mechanistic and investigative significance.

It opens a new avenue to examine whether the interaction between EHMT1, neurons, microglia, and inflammatory pathways is indeed involved in the biology of the syndrome.

The most important finding, the brain maintained a degree of reversibility

Perhaps the most important result of the work is not microglia or IL-1β in isolation.

Is the fact that certain molecular and cellular abnormalities that had already been established could be corrected when GLP was restored after birth.

H3K9me2 could be restored even in mature, post-mitotic neurons.

The number of dendritic spines could also be restored.

The increased presence and activation of microglia could be reversed.

And when GLP restoration occurred more broadly in the organism from a young age, certain behavioral abnormalities could also be improved.

This is an important indication biological plasticity.

But the timing seems to matter

At the same time, the study shows that not everything is equally easily reversible.

Postnatal restoration of GLP exclusively in neurons corrected many molecular and cellular markers, but was not sufficient to improve complex behaviors in mice.

The authors conclude that certain characteristics are reversible, while the possibility of improving other components depends on when the restoration is done and how extensive it is.

This is of particular importance for future therapeutic research.

If technologies are ever developed that can increase the function of EHMT1 or correct the consequences of its deficiency, it should be investigated not only if work, but also when they should be applied and to which cell types.