Mesenchymal Stem Cell Therapy for Cardiac Inflammation: Immunomodulatory Properties and the Influence of Toll-Like Receptors Abstract:

Mesenchymal Stem Cell Therapy for Cardiac Inflammation Background.

After myocardial infarction (MI), the inflammatory response is indispensable for initiating reparatory processes.
However, the intensity and duration of the inflammation cause additional damage to the already injured myocardium. Treatment
with mesenchymal stem cells (MSC) upon MI positively affects cardiac function. This happens likely via a paracrine mechanism.
As MSC are potent modulators of the immune system, this could influence this postinfarct immune response. Since MSC express
toll-like receptors (TLR), danger signal (DAMP) produced after MI could influence their immunomodulatory properties. Scope
of Review. Not much is known about the direct immunomodulatory efficiency of MSC when injected in a strong inflammatory
environment. This review focuses first on the interactions between MSC and the immune system. Subsequently, an overview is
provided of the effects of DAMP-associated TLR activation on MSC and their immunomodulative properties after myocardial
infarction. Major Conclusions. MSC can strongly influence most cell types of the immune system. TLR signaling can increase and
decrease this immunomodulatory potential, depending on the available ligands. Although reports are inconsistent, TLR3 activation
may boost immunomodulation by MSC, while TLR4 activation suppresses it. General Significance. Elucidating the effects of TLR
activation on MSC could identify new preconditioning strategies which might improve their immunomodulative properties.

Mesenchymal Stem Cell Therapy for Cardiac Inflammation Conclusions:

The inflammatory response after MI is essential to initiate reparative pathways and clear debris, yet these activated
immune cells cause a lot of short and long term damage to the myocardium. Broad immunosuppressive drugs were
only detrimental by reducing both the damaging and healing pathways. Stem cell therapy after MI could improve cardiac
function, most likely by the production of paracrine factors. One of the systems influenced by these paracrine factors is the
immune system. Basically every immune cell was reported to be affected by MSC in different degrees and subtypes
are induced which in turn can influence other immune cell functioning. The mechanisms by which MSC achieve these
effects remain unclear, with many groups supporting various effector molecules and pathways. In all studies, however, MSC
can influence the immune system via different pathways, thereby having a range of possible effects on their target cells.
One of the obvious reasons why different outcomes are still observed is due to the heterogeneity of the MSC and the
differences in donor, origin, isolation, culture, and coculture conditions with immune cells. A broad definition for MSC
has been defined, but this does not mean all these cells are identical. MSC from different origins can have different
capacities and can react differently to similar stimuli [131]. Even when cells are isolated from identical origins according
to a strict protocol, strong variations still exist between donors [109, unpublished own observations]. Likewise the
timing, concentration, and duration of the stimulation with TLR-ligands can influence observed effects, and the outcome
after one hour of stimulation might be entirely different from results after a day of stimulation [124].
Additionally, immunosuppression assays show a lot of variation. Some groups worked with peripheral blood mononuclear cells (PBMC), while others worked with isolated fractions of CD3+ or CD4+ T-cells. It is difficult to compare these results directly with each other, for in a PBMC mixture many
other immune cells are present which influence their environment, as shown in Figure 1. Add to this that the immunesuppressing effects on PBMC or T-cell proliferation in the untreated groups varied strongly between groups, it becomes clear that universal protocols are needed to perform this type
of assays. In addition, the experiments need to be performed with various different MSC and immune cell donors to make
the outcomes more robust. Although a great effort has been undertaken to identify the effects of TLR activation on MSC, many inconsistencies
still remain. Despite the many contradicting reports, some similarities can be found and some clues provided insights
into possible mechanisms. Many groups have established the expression of TLR by MSC, although at protein level
they are sometimes hard to detect. The effect of TLR activation on proliferation is probably minimal, while differentiation can be interfered with. Although few studies have looked at migration, improved migration might help honing in immunomodulative stem cell therapy and should be investigated further. Initial reports indicate an increase in migration, at least in the acute phase [124]. Regarding the immunomodulatory capacities of MSC, much ambiguity remains. In vitro8 Mediators of Inflammation
and in vivo work seem to indicate that TLR3 activation with poly(I:C) can boost the immunosuppressive potential of
MSC, while TLR4 activation with LPS could reduce it. Experimental studies showed TLR2 and TLR4 become activated
after MI and are correlated to ischemia-reperfusion injury and LV dysfunction [135–137]. The TLR4 activation can create an unfavorable environment for MSC, reducing their effectiveness as immunomodulatory therapeutics after MI. This in turn would make preconditioning of MSC by using
TLR3 ligands to boost immunomodulation an interesting target. These divergent effects of TLR3 and TLR4 signaling
have prompted Waterman et al. to suggest MSC can be polarized into inflammatory and anti-inflammatory subtypes by
differential TLR activation [124]. However, due to the many contradictory findings, more research will be necessary to
validate this hypothesis. The vast majority of the studies discussed in this review did not focus on cardiac inflammation, but on auto-immune
diseases or organ transplantation. This justifies the work with PAMPs, as the main concern will be an infectious threat to
a patient with a suppressed immune system. In the setting of inflammation after myocardial infarction, the inflammatory
signals consist of DAMPs. It is unlikely that DAMPs and PAMPs activate the same receptors in exactly the same way.
There are likely more receptors (PRRs) on the MSC that can recognize these ligands and it could very well be a combined
activation of receptors that leads to the activation of a specific pathway in the cell, which could differ between PAMPs and
DAMPs. To study the effectiveness of MSC therapy for postMI inflammation, it would be advisable to investigate the effect of TLR activation on MSC using DAMPs that are
released after MI. Only by investigating it this way can the role of TLR activation on MSC in the cardiac setting be truly elucidated.

Mesenchymal Stem Cell Therapy for Cardiac Inflammation – Full Study