IL-25 and B cells in atherosclerosis

Abstract

Atherosclerosis is a chronic inflammatory disease of medium and large vessels of the body which is mainly characterized by the formation of lipid rich plaques within the arterial tissue. It is the most common cause of cardiovascular disease leading to clinical manifestations of coronary artery and cerebrovascular disease. Immune responses targeting modified low density lipoprotein (LDL) play an important role in atherosclerosis. Moreover, Th1 immune responses have inextricably been associated with disease progression while the role of Th17 and Th2 related immune responses is less clear.

In the present work we report that different B cell subsets could potentially be used as biomarkers for the prediction of future stroke events. Specifically, our findings demonstrate an association between high levels of CD19+CD40+ cells and a decreased risk of stroke while high levels of CD19+CD86+ B cells were associated with an increased risk.

Another main focus was the investigation of the role of IL-25 in atherosclerosis. In order to tackle this, the cytokine was both administered and blocked in hypercholesterolaemic apoE-/- mice. Interestingly, an atheroprotective effect of the cytokine was observed. Regarding exogenous IL-25 administration, the proposed mechanism of atheroprotection implicates the increase of IgM antibodies targeting phosphorylcholine (PC), a major epitope on oxidized LDL (oxLDL) via innate lymphoid cell type 2 (ILC2)-derived IL-5. We suggest that the essential trigger for this mechanism is the expansion of ILC2s.

Additionally, blockade of endogenous IL-25 at the onset of atherosclerosis led to increased plaque formation of unstable phenotype, accompanied by a Th1/Th17 shift of the cytokine balance in the spleen of the mice. The above mentioned findings suggest that signaling through IL-25 during early plaque formation possibly regulates protective immune mechanisms and that disruption of that signaling leads to the dominance of pro-inflammatory immune responses.

Due to the atheroprotective effect of IL-25 in mice we also investigated its effect on human peripheral blood mononuclear cells (hPBMCs). IL-25 was shown to dampen Th17 and Th1-related immune responses in hPBMCs while in the presence of oxLDL it dampens Th1 and promotes Th2-related immune responses. The above mentioned findings indicate that IL-25 could potentially have a protective role in human atherosclerosis.

Abstract (Swedish)

Popular Abstract in English

Atherosclerosis is a disease of medium and large sized vessels of our bodies that is mainly characterized by the deposition of lipids in the vessel walls. Increased lipid levels in the blood of an individual, because of lifestyle habits but also because of genetics, can lead to the formation of an “atherosclerotic plaque”. Thus, although lipids are important for the normal function of cells, when in excess can lead to atherosclerosis.

As oil is insoluble in water the same thing happens with the lipids in our blood, making difficult their essential distribution to the various tissues. But our body has found a very efficient solution; the lipids “get packed” to form spherical particles, surrounded by a membrane that enables them to float freely in aqueous “solutions” such as the blood. These particles are called lipoproteins and according to their composition they are categorized into different types. Low density lipoprotein (LDL) particles have mostly been associated with atherosclerosis therefore LDL is known as the “bad cholesterol”.

An atherosclerotic plaque is created when the lipids of the blood (mostly LDL) get trapped in the vessel wall and get retained at that location. This gradually increasing, un-physiological homing of lipids and cells leads to the formation of a gruel material that grows within the vascular wall making the vessel less flexible upon physical movement. If the individual that has such a plaque will not regulate his lipid levels through medication and change of lifestyle habits, there is an increased risk that the plaque will increase and rupture. When this happens the material of the plaque gets exposed to the blood flow leading to the formation of a blood clot. This clot can then stop the flow of the blood to the rest of the body and this may lead to death.

When plaques of the heart vessels get ruptured then the individual experiences a heart attack but when the same thing happens for a plaque of the carotid arteries then a stroke event takes place. Unfortunately, nowadays atherosclerosis is the leading cause of death worldwide. Statins are drugs that were discovered in the '70s and they act by decreasing lipid levels in the blood but cannot fully protect from atherosclerosis.

The last few decades, scientists working in the field of atherosclerosis have acknowledged that in addition to the contribution of lipids, the immune system also participates to the progression as well as the resolution of the disease. The term “immune system” refers to specific cell types and proteins of our bodies which exist in order to protect us from viruses, infections and any microscopic “threat” that the body may encounter.

Some of these cells that work as “soldiers” protecting us, are called B cells, T helper cells and phagocytes. B cells can secrete substances called “antibodies” that can capture anything “unwanted” and remove it from the body. Phagocytes can “eat-up” harmful particles, bacteria and dead cells and remove them from the surroundings while T helper cells can communicate with other cells either through direct contact or by secretion of substances that neighboring cells perceive as signals, telling them to do something very specific.

When lipids increase in the blood and LDL accumulates in the arterial wall, the immune system realizes that something strange and unusual is going on. This accumulation and retention also leads to subsequent modification of LDL. Then the cells of the immune system recognize strange patterns on the modified lipoprotein particles and target them. This is a mechanism of protection but when the control is lost and the accumulation and modification of lipids is great then the promotion of “danger” signals from such cells can lead to increased inflammation and further damage of the arterial wall.

One way to possibly treat atherosclerosis is to try to direct the immune system in such a way so that the inflammation and subsequent disease-promoting immune responses in the atherosclerotic plaque are dampened. Scientists have attempted to do that many times with great success throughout the last decades through administration of specific immune cell types or substances in experimental animal models.

Interleukin-25 (IL-25) is a cytokine (protein) that normally exists in our bodies. In the present work, exogenous IL-25 was administered in mice that have increased lipid levels through genetic modification, thus being prone to atherosclerotic plaque formation. Interestingly, IL-25 administration could decrease atherosclerotic plaques in the aortas of both young and old mice, the latter ones having already established plaques (paper II). This means that IL-25 is effective in preventing to some extent the early formation of atherosclerotic plaques as well as decreasing the progression of already established plaques. We suggest that the way that IL-25 acts in order to confer this protective effect is through the induction of a cell type, the existence of which the scientific community was not aware of, until 2010. This cell type is named ILC2s (innate lymphoid cells type 2) and has the capability to secrete large amounts of another protein called IL-5. IL-5 works as a signal for B cells to get activated and secrete antibodies against unwanted particles which in our case are molecular patterns on modified LDL. By the generation of these antibodies, modified LDL particles get removed thus conferring protection from the disease.

In another set of experiments including the same mouse model we administered an antibody that blocked IL-25, leading to increased atherosclerosis (paper III). This result was observed only when we treated young mice that did not have established plaques. When older mice underwent this treatment we did not observe any difference in the formation of plaques. The results of these experiments indicate that endogenous IL-25 is important in the protection against atherosclerosis during early plaque formation since blockade of the protein led to increased atherosclerotic plaques only in young mice.

Since we established in mice that IL-25 protects against atherosclerosis via the induction of immune mechanisms, we also investigated the effect of the cytokine on human cells of the immune system (paper IV). For that reason we isolated mononuclear cells that exist in the blood of humans. When we incubated the cells in vitro (in the lab under sterile conditions) we saw that the cells secreted less pro-inflammatory cytokines. With the term pro-inflammatory we mean promoting inflammation and atherosclerosis-related immune responses. These results indicate that IL-25 can potentially have a protective role in human atherosclerosis.

In order to be able to treat atherosclerosis we also need to find ways to predict future events such as strokes and heart attacks. In paper I we have shown that the existence of specific B cell subsets can be related with increased or decreased risk of a future stroke event. We had the opportunity to characterize and quantify specific B cell subsets in the blood of 700 individuals that were randomly selected from a clinical study called “Malmö Diet and Cancer Study”. In this particular study, 28449 individuals were recruited between 1991 and 1994. Some of these individuals were also asked to participate in a sub-study focusing on cardiovascular risk (number of individuals=6103). The 700 individuals that we included in our study (paper I) were randomly obtained from the 6103 participants.

All the participants were followed from baseline examination until the first cardiovascular event such as myocardial infarction (heart attack) or stroke either of them being fatal or non-fatal. Participants were followed until 2008. At the baseline examination, blood was collected from these individuals and the isolated mononuclear cells were frozen at -140oC. In 2011 we thawed the cells that were obtained at the basal examination and analyzed them focusing on B cell subsets with a technique called “flow cytometry”. We next had the opportunity to correlate the manifestation of strokes or myocardial infarctions with the amounts of the B cell subsets that were analyzed. Interestingly, we saw that high levels of CD19+CD86+ B cells correlated with an increased risk of a future stroke which was greater for men. Additionally, high levels of another B cell subset (identified as CD19+CD40+ B cells) were associated with a decreased risk of a future stroke event. This is the very first study to report an association between B cells and cardiovascular events. We hope that in the future other scientists will have similar observations so that this finding could actually be used in the clinic. If so, this would mean that with a blood sample the physician would be able to quantify the levels of these B cell subsets and depending on the results, the patient could then be categorized as being either in a low or a high risk of experiencing a future stroke event. Patients at high risk will have to be monitored more closely and treated more aggressively in order to prevent such an event.

• author: Mantani, Polyxeni
• supervisor: Gunilla Nordin Fredrikson; Harry Björkbacka; Jan Nilsson
• opponent: Dr Miller, Ashley, University of Glasgow
• organization: EXODIAB: Excellence of Diabetes Research in Sweden; Cardiovascular Research - Immunity and Atherosclerosis (research group)
• publishing date: 2014
• type: Thesis
• publication status: published
• subject: Cardiac and Cardiovascular Systems
• keywords: IL-25, B cells, ILC2s, IL-5, anti-PC IgM, atherosclerosis, stroke, cardiovascular events
• in: Lund University Faculty of Medicine Doctoral Dissertation Series
• volume: 2014:77
• pages: 173 pages
• defense location: "Lilla aulan", entrance 59, Skåne University Hospital, Malmö
• defense date: 2014-06-12 13:00:00
• ISSN: 1652-8220
• ISBN: 978-91-7619-006-7
• language: English
• LU publication?: yes
• id: f1d3bb95-25c2-491e-9723-7b57c1b260b8 (old id 4437822)
• date added to LUP: 2016-04-01 13:49:11
• date last changed: 2019-05-22 05:15:32