Örebro University
School of Medical Sciences Degree Project
15 ECTS January 2019
REGIONAL SPECIALIZATION OF THE
ADAPTIVE IMMUNE SYSTEM WITHIN THE
HUMAN GUT
Version 2
Author: Yolanda Vikberg Martínez Supervisor: Elisabeth Hultgren-Hörnquist, PhD, Professor
Abstract Background
The human intestine is the largest immune organ in the human body and is constantly exposed to a great number of antigens and bacteria. This requires a balance between tolerogenic and immunogenic responses to avoid inflammation. The small and large intestine have different anatomy, physiological functions and are affected differently by diseases. Many studies on intestinal immunity do not distinguish between the small and large intestine, and most studies are performed on mice. To fully understand human intestinal immunity, there is a need for more studies on humans that also treat the different intestinal regions as separate compartments.
Aim
To summarize possible differences in the adaptive immune system in the human intestine, with special focus on antigen presenting dendritic cells and T lymphocytes.
Methods
A systematic review was performed on previously published articles that examined human intestinal immunity with focus on adaptive immunity. The articles were obtained from the database PubMed using MeSH terms. The articles were selected based on certain inclusion and exclusion criteria and their quality was evaluated.
Results
The collected data from included articles clearly show a difference in small and large intestinal adaptive immunity regarding distribution and functions of dendritic cells and T lymphocytes.
Conclusion
More knowledge about the differences in the properties of the immune system in different regions of the intestine would probably contribute to a greater understanding of the immunological basis of diseases affecting the intestines and might be valuable for developing new treatment possibilities.
Abbreviations
APC: Antigen presenting cells DC: Dendritic cells
DN: Double negative DP: Double positive
FACS: Fluorescence-activated cell sorter.
FITC: Fluorescein isothiocyanate
FOXP3: Forkhead box protein 3
GALT: Gut-associated lymphoid tissue ILT3: Immunoglobulin-like transcript 3 IFN-g: Interferon g
IL: Interleukin
IEL: Intraepithelial lymphocytes LPL: Lamina propria lymphocytes NK: Natural killer
TH cells: T helper cells Th cells
Treg: T regulatory cells
TGF-β: Transforming growth factor β TNF-a : Tumor necrosis factor a
1 BACKGROUND ... 4
1.1 THE HUMAN INTESTINES ... 4
1.2 DENDRITIC CELLS ... 5
1.3 T LYMPHOCYTE SUBTYPES ... 6
2 AIM ... 7
3 MATERIALS AND METHODS ... 7
3.1 SEARCH APPROACH AND SELECTION CRITERIA ... 7
3.2 SCIENTIFIC ASSESSMENT ... 9 3.3 ETHICAL CONSIDERATIONS ... 9 4 RESULTS ... 9 4.1 DENDRITIC CELLS ... 9 4.2 T LYMPHOCYTES ... 11 5 DISCUSSION ... 14 6 CONCLUSION ... 16 7 SPECIAL THANKS TO ... 16 8 REFERENCES ... 17 9 APPENDIX ... 20
1 BACKGROUND
1.1 THE HUMAN INTESTINES
The intestine comprises the largest immunological tissue in the human body [1] and is always exposed to a large variety of antigens from dietary sources or the microbiota. The interest in how the intestines are affected by its contents is increasing and numerous studies are performed to map the properties of the immune system in this part of the body. There are not many studies that distinguish between the small and large intestine, even though these two regions both have their own characteristic anatomy and physiology [1]. For instance, the bacterial load is much larger in the colon versus the ileum. The ileum is constantly exposed to dietary antigens and digestive enzymes. Because of this high exposure to antigens and bacteria in the intestines, there is a great need for homeostasis between the tolerogenic and immunogenic responses to prevent inflammation to occur in the healthy intestine [2]. The two intestinal regions are also affected differently by diseases. For instance, malignant disease is much more prevalent in the large intestine, and inflammatory bowel diseases have different anatomical localizations: whereas ulcerative colitis only affects the colon, Crohn’s disease can affect the whole gastrointestinal tract. Celiac disease is a disease restricted to the small bowel and can also affect the skin in dermatitis herpetiformis [1,3]. Pathogenic infections also show anatomical restriction: for example, Salmonella infection usually occurs in the small intestine whereas Shigella infection occurs in the colon, and different strains of Escherichia coli (E. coli) have distinct anatomical sites of action [4,5]. However, it is still unclear how the different distribution of these diseases is connected to the different proportions of immune cell subtypes in the small and large intestine [1]. Most studies on intestinal immunity are performed on mice and primarily the ileum, and this might not be an accurate model for describing human immunity in neither the ileum nor the colon. In addition, laboratory animals are on a defined diet and have a less diverse microbiota in comparison to the human colon [6]. Differences in the microbiota are likely to vary due to differences in diet and hygiene routines such as coprophagy in mice [7], a habit which exposes the small intestine to microbe-associated molecular patterns in feces that normally mainly exists in the large intestine in humans. To fully understand the pathogenesis of intestinal diseases and the function, or dysfunction, of the immune system in these processes, there is a great need for examining the properties of the human intestinal immune system in different anatomical regions, both in healthy individuals and also objects affected by disease. The intestinal immune system can be divided into the gut-associated lymphoid tissue (GALT), and effector sites. The GALT is composed of Peyer’s patches, isolated lymphoid follicles, the
appendix and gut draining mesenteric lymph nodes and effector sites include the epithelium and the lamina propria [8]. The GALT is the major site of antigen exposure [9] and is important in creating the tolerogenic milieu in the intestines. Two very important players in the game of balancing tolerance and immune responses are dendritic cells (DC) and T lymphocytes [10]. These are, along with other cells of the immune system, located in the lamina propria, the epithelia and the GALT [8,11,12].
1.2 DENDRITIC CELLS
DC are antigen presenting cells (APC) with the ability to activate naïve T cells [6] but in addition to that, they are also indispensable in making sure that even though there is a high antigen exposure in the intestine, there is no inflammation occurring in the healthy intestine [2]. There are various DC populations throughout the intestinal lymphoid tissues. DC are essential in generating low-level immune responses in the steady state which creates a tolerogenic milieu towards the commensal microbiota and avoiding pathological immune responses [13]. Immature DC are crucial in processing and presenting self-antigens to T lymphocytes which leads to peripheral tolerance [14]. DC mature upon exposure to microbes or pro-inflammatory cytokines and this usually induces changes regarding their morphology, phenotype and function. After this has occurred, the DC can perform their function of priming and activating naïve T lymphocytes [15].
Maturation of DC subsets is followed by a switch in the expression of chemokine receptors (CCR). Exposure to lipopolysaccharide (LPS) or tumor necrosis factor a (TNF-a) causes a downregulation of receptors for inflammatory chemokines, such as CCR1, CCR2, CCR5 and CXCR1. These are probably important for the recruitment of DC with an immature phenotype to tissues in vivo. On the other hand, maturation causes upregulation of the homeostatic chemokine receptor CCR7. CCR7 is important for DC migration to regional lymph nodes irrespective of their maturation status. In the steady state these migrating DC that are not activated by pathogens or inflammation present self-Ag to naïve T lymphocytes. This process induces peripheral tolerance and thus inhibit autoreactive, potentially dangerous T lymphocytes [16]. One factor for inducing the maturation and survival for DC is leptin [6]. The major known role for leptin is to stimulate the feeling of satiety, but it also has immunomodulatory effects. For example, leptin initiates activation of the NFkB signaling pathway which has an antiapoptotic effect on DC, promoting their maturation and survival. Leptin can also improve the ability among DC to stimulate T cells and create a Th1 response [6].
1.3 T LYMPHOCYTE SUBTYPES
Intestinal T lymphocytes are located in three major compartments: the GALT, the epithelium and the lamina propria [8]. There are many different functional subsets of T lymphocytes. Some of these will be presented here.
The T lymphocytes of the GALT are naïve cells that have not yet been primed by an APC. In the effector sites, T lymphocytes are known as lamina propria lymphocytes (LPLs) and intraepithelial lymphocytes (IELs), respectively, and have either a memory or effector phenotype [17]. IELs seem to be involved in oral tolerance and keeping the integrity of the epithelial barrier whereas LPLs, among other things, control the local production of antibodies. Both CD8+ IELs and LPLs express perforin and Fas ligand (FasL), both being cytotoxic
markers [17]. LPLs also include the CD4+ T helper 1 cells (TH1 cells), T helper 2 cells (TH2
cells), T helper 17 cells (TH17 cells) and T regulatory cells (Tregs), all being effector cells.
TH1 cells are involved in immunological responses towards intracellular pathogens. The major
cytokine they produce is IFN-g.
TH2 cells are specialized for initiating immune responses to parasites, such as helminths, and
are also involved in allergic diseases [8]. The major cytokine associated with TH2 cells is IL-4.
TH17 cells are important for mucosal immunity [18] and are often associated with autoimmune
diseases [19,20]. They also provide protection against bacteria and fungi [8]. TH17 cells are
characterized by secretion of the proinflammatory cytokine IL-17 [18] that attracts and activates neutrophils and other immune cells, and they also secrete IL-21 and IL-22.
Regulatory T cells (Treg) are CD4+CD25+ cells, characterized by expression of the transcription
factor Forkhead box protein 3 (FOXP3) [21]. Treg are important for intestinal homeostasis and
dysregulation of this cell subset is very common in immune mediated diseases [20]. They produce the regulatory cytokines IL-10 and TGF-b [8]. Treg and TH17 cells share some features:
for example, their differentiation is induced by TGF-b and they are both present at high levels in the intestine [22]. Treg have the ability to turn into TH17 cells in the presence of APCs and
IL-2 and IL-15 [20]. This means that there is a great plasticity of Tregs, regulated by cytokines
T lymphocytes are the dominant immune cell in the intestinal epithelium [1,23]. They express either the T-cell receptor (TCR) ab or gd heterodimer [8,23]. Around 50% of the IELs are CD8ab+TCRab+ (CD8+TCRab+) and the rest are unconventional (not MHC class I or II
restricted) forms of T lymphocytes [23]. Such unconventional T lymphocytes are for example TCRgd+, either double negative (DN) for both CD4 and CD8, or CD8aa+ with any of the TCRs
mentioned [23]. TCRgd+ T lymphocytes may have a regulatory role in the intestine, a possible
function supported by the fact that they respond with secretion of TGF-b and IL-10 when stimulated with non-self-phospholipids (PLs) [23].
2 AIM
The aim of this essay was, through a systematic review of the literature, to summarize possible differences in the adaptive immune system in the human small versus the large intestine, with special focus on antigen presenting dendritic cells and T lymphocytes.
3 MATERIALS AND METHODS
3.1 SEARCH APPROACH AND SELECTION CRITERIA
A systematic review was performed on previously published articles concerning the adaptive immune system in either the small or large intestine, or articles that studied both these tissues. The articles were collected from the database PubMed. Medical subject headings (MeSH) terms were used to create four different block searches using the MeSH-terms below. To further limit the search, articles with the MeSH term “mice” were excluded using the boolean operator >NOT<.
1. Small intestine, antigen presenting cells. 2. Small intestine, T lymphocytes.
3. Large intestine, antigen presenting cells. 4. Large intestine, T lymphocytes.
No exclusions were used based on the publication date. The filter “humans” was applied. When reading the abstracts generated from this search, articles only focusing on pathology were excluded and so were also articles presenting studies performed on other species than humans. Studies covering both pathology and healthy controls were not excluded.
The search generated 448 articles, and all the abstracts were read and either selected or discarded based on inclusion and exclusion criteria. Inclusion criteria in this systematic literature study were: Studies performed on healthy patients or studies using healthy controls and studies reporting features of either of the cell populations used as MeSH-terms. Exclusion criteria were: studies performed only on other species than humans, studies focusing only on pathology, studies where the anatomical region of the intestine was not specified to the small intestine or the colon. Review articles were also discarded to avoid reporting bias [24]. Figure 1 describes the search process.
Figure 1. Flow chart of the search process.
448 abstracts
43 abstracts kept
Included fulltexts:
12 articles. fulltexts:31Excluded
Excluded because main focus on pathology or inflammation, or because did not go beyond textbook
knowledge
405 abstracts discarded
Discarded because main focus on pathology or inflammation,
or study performed in non-human species
3.2 SCIENTIFIC ASSESSMENT
A template from the Swedish agency for health technology assessment and assessment of social services (SBU) was used as inspiration for a checklist to evaluate the quality of the articles included [24]. The quality of the included articles was based on the following questions:
• Was there an explicit aim?
• Was the study population clearly defined? • Was the method clearly described?
• Was the anatomical region examined declared? • Were the results clearly described and discussed?
• Did the articles have any ethical considerations and/or patient consent?
Based on the answer to the questions above (yes, no or unclear) the articles were given a quality of low (none of the articles), moderate (8 of the articles) or high (4 of the articles). Table 1 in appendix shows a summary of included articles and their evaluated quality.
3.3 ETHICAL CONSIDERATIONS
9 of the 12 included articles had ethical approval [2,6,17,18,25–29]. 3 of the articles did not mention ethical consideration [12,23,30]. 8 articles declared consent from patients [2,6,17,18,23,25,27,29]. 4 articles did not mention consent from patients [12,26,28,30]. This was a systematic literature study based on previously published articles and further ethical considerations were not considered necessary.
4 RESULTS
4.1 DENDRITIC CELLS
CD103+CD11b+ DC are an unique subset of dendritic cells in the intestines and are the
predominant DC subset in the small intestine [10]. CD103 is also known as integrin αE, which is an important DC marker and involved in inducing Tregs and tolerance [2,31]. CD11b, also
known as Sirpa or integrin a, is a member of the integrin b2 family which mediates cell adhesion and signaling [19]. TGF-b is present at high levels in the intestines and is indispensable for imprinting of LP CD103+CD11b+ DC [10]. Characterization of human DC
showed that both ileal and colonic DC express CD103 [2]. Most of these CD103+ DC
co-expressed CD11b [2]. There was a higher proportion of CD103+CD11b+ DC in the ileum than
in the colon. The proportion of CD103+CD11b- DC was higher in the colon compared to the
The gut-homing marker integrin b7 is expressed with either integrin αE or integrin α4. There was no difference in DC expression of the gut-homing marker integrin b7 and DC from both intestinal compartments were able to generate integrin b7 expressing T lymphocytes [2]. The expression of the DC activation marker CD40 was examined by Mann et al. [2] and demonstrated equal expression in the ileum and the colon. They also found no significant differences in expression of Toll like receptors 2 and 4 (TLR2 and TLR4) when comparing ileal and colonic DC [2]. They concluded that the maturation and activation status of DC was equal in both intestinal compartments [2].
Maturation and migration properties of colonic LP DC were examined by Bell et al. [12] using multicolour flow cytometry. DCs were collected either by enzymatic digestion of colonic biopsies by collagenase or by letting cells migrate out from colonic biopsies during overnight culture. Freshly isolated DCs had a more immature phenotype compared to DCs cultured overnight, based on expression of CD25 (IL-2Ra), CD40, CD80 and CD86 that were all higher on cultured cells. The freshly isolated cells were CD40+ but had low expression of the other
markers. The maturation state of the DCs was reflected in their endocytic capability: immature DC had a higher endocytic capability compared to cultured DC, based on higher ability to take up fluorescein isothiocyanate (FITC)-dextran. Migratory behaviour was considered higher among mature DC because these were the cells that were recovered after migration out of the colonic biopsies [12].
Leptin was found by Al-Hassi et al. [6] to have a mechanistic role in LP DC migration and induction of CCR7 expression on DC in the normal human ileum but not in the colon, as seen in paired samples comparing ileal DC and colonic DC. This difference was not related to the maturation status of the DC because both intestinal compartments had immature DC with low expression of CD40, CD80 and CD86 [6]. Ileal CCR7+ DC migrated towards CCL19, a CCR7
chemoattractant [32]. Leptin signals via LepRb and the expression of this receptor mirrored the expression of CCR7, in that only CCR7+ DC expressed LepRb [6]. This shows that leptin might
have a regulatory role on DC migration from the ileum [6]. Mann et al. [2] also examined the expression of CCR7 on CD103+CD11b+ DC and found that expression of CCR7 on this subset
was higher in the colon than in the ileum. Both CD103+ and CD103- colonic DC showed an
increased endocytic capacity compared to ileal DC [2]. This study did however not relate the expression of CCR7 to leptin.
Immunoglobulin-like transcript 3 (ILT3), an inhibitory receptor expressed on myeloid APC such as DC [33], gives rise to tolerogenic APC when expressed at high levels. This receptor was expressed at a greater proportion on colonic compared to ileal DC. The major DC subset expressing ILT3 was CD103- [2].
Colonic DC also showed greater regulatory traits compared to their ileal counterparts in that they had an enhanced ability to induce IL-10 producing Tregs [2]. Both migratory DC and DC
with endocytic capability were enhanced in the colon compared to the ileum. The ileal DC had, compared to colonic DC, a higher inflammatory cytokine profile with higher production of TNF-a and IL-1b. In contrast, there were no differences in TGF-b, IL-12, IL-22 and IL-23 production from the DCs from the two different intestinal compartments [2].
Finally, expression of homing receptors differed between ileal and colonic DC [2]. DC from the ileum were CCR9+ whereas the colonic DC were CCR4+. This was reflected in their
imprinting ability on T lymphocytes: The ileal CCR9+ DC were able to generate CCR9+ T
lymphocytes whereas colonic DC had a greater ability to generate CCR4+ T lymphocytes. The
majority of T lymphocytes generated from both intestinal regions were however CCR9- [2].
4.2 T LYMPHOCYTES
Expression of CCR9 on T lymphocytes was examined by Papadakis et al. [28]. Immunofluorescence showed that both CD4+ and CD8+ ileal LPLs had higher expression of
CCR9 compared to colonic T lymphocytes. T lymphocytes in the mesenteric lymph nodes draining the ileum also expressed high levels of CCR9 [28]. Even though mRNA transcripts of CCR9 were not present in the colon, a very small fraction of colonic T lymphocytes were CCR9+, as shown by flow cytometry[28]. The authors suggest that these colonic CCR9+ T
lymphocytes only express this receptor by coincidence or through stimulation by other chemokines than those that stimulate CCR9 expression in the small intestine. The authors further suggest that induction of CCR9 is restricted to the small intestinal mucosa [28].
Colonic TH17 cells were found by Carrasco et al. [18] to be present at significantly higher
numbers compared with paired terminal ileal counterparts. In contrast, there were no differences in the numbers of TH1 cells in the two gut compartments [18]. These results were
based on examining a mixture of intraepithelial (IEL) and lamina propria lymphocytes (LPL) using IHC [18].
CD4+CD8+, double positive (DP), and CD4-CD8-, double negative (DN), T lymphocytes are
rare subsets that are more or less restricted to the intestines except for the thymus. The healthy ileal mucosa contained a higher proportion of DP T lymphocytes in comparison to the colonic mucosa. On the other hand, DN T lymphocytes were present at higher proportions in the healthy colon (not specified whether DP and DN were located in the LP or epithelium) [18]. There were also slightly higher numbers of CD4+CD25+FOXP3+ T lymphocytes (T
reg) in the right colon
compared to the left colon and the terminal ileum, based on examining a mixture of IELs and LPLs collectively [18].
In summary, TH17 effector cells, Treg and DN T cells were all increased in the colon compared
to the ileum, and thus both tolerogenic and effector mechanisms were found to be increased in the colon [18].
Cytotoxic properties of IELs and LPLs in the ileum and the colon were examined by Melgar et al. [17]. Methods used were immunomorphometry and reverse transcriptase PCR. The effector cells were not treated ex vivo and thus, the in vivo cytolytic activity was examined. In both intestinal regions, both IELs and LPLs were positive for the two cytotoxic proteins Fas ligand (FasL) and perforin. Perforin+ LPLs were as common in the ileum as in the colon, but among
IELs they were more prevalent in the ileum compared to the colon. FasL+ LPLs were more
prevalent in the colon compared to the ileum but regarding IELs they were present at the same proportions in both intestinal regions. When examining cytolytic activity among IELs and LPLs from the ileum, the results varied and only about half of the samples contained cells with significant TCR- or CD3-mediated cytotoxicity. However, in the colon, IELs and LPLs had no or low TCR- or CD3-mediated cytotoxic activity [17]. The authors hypothesize that the colonic Perforin+ cells may in fact be natural killer (NK) T lymphocytes and require stimulation via a
NK-receptor in order to acquire a cytotoxic activity [17].
In summary: both ileal and colonic IELs and LPLs express FasL and Perforin but only the ileal cells had TCR- or CD3-mediated cytotoxicity. The colonic cells might need other signals for activating their cytotoxic properties [17].
T lymphocyte mRNA expression for the cytokines IFN-g, a major Th1 cytokine, and IL-17A
showed higher values in the terminal ileum compared to the colon based on examining a mixture of IELs and LPLs collectively, but no statistical significance was obtained because of great inter-individual variability [18]. IFN-g and IL-2 secreting colonic T lymphocytes (not specified whether these were IEL or LPL) were examined by Breese et al. [26] using
plaque-forming cell assay (PFC) and Norhern blot. Both cytokines were found in only low or undetectable concentrations in most healthy controls (9/13 and 10/14 respectively). Concordantly, Northern blot showed that 0/9 controls had detectable IFN-g-mRNA and no signal was detected for IL-2 [26]. The number of IFN-g and IL-4 secreting IELs and LPLs in the duodenum and right colon in healthy biopsies were enumerated by Carol et al. [25] with enzyme-linked immunospot technique (ELISPOT) without further in vitro stimulation, and they found a higher proportion of cytokine secreting IELs and LPLs and also a higher total cytokine secretion in the duodenum compared to the colon. In both intestinal compartments IELs showed a higher cytokine secretion compared to LPLs [25].
In summary: the duodenum showed a higher T lymphocyte secretion of IFN-g and IL-4 compared to the colon, where secretion of mentioned cytokines and also IL-2 was low or absent [25,26]. IELs seem to have a higher total cytokine secretion compared to LPLs in both intestinal compartments [25].
Production of TNF-a from jejunal IELs and LPLs was studied by Ebert et al. [30] by ELISA, and they also examined the ability among IELs to respond to this cytokine. Jejunal LPLs had a greater TNF-a production compared to IELs, especially CD4+ T cells [30].
Duodenal IEL secretion of the cytokines TNF-a, IFN-g, TGF-b1, IL-4 and IL-10 in response to CD1 transfected peripheral blood DC was measured by Russano et al. [23]. The authors hypothesized that the CD1 molecule was involved in recognition of PL by T lymphocytes in the intestine. Some of the transfectants were loaded with phospholipid (PL) Ags to examine if the IELs responded differently to that. Both conventional TCRab+ and unconventional TCRgd+
andDN TCRab+ were analyzed. Most TCRab+ T lymphocytes showed IFN-g and TNF-a
secretion. TCRab+ cells responded to PL stimulation with large IL-4 and TGF-b1 secretion. A
larger proportion of TCRgd+ compared to TCRab+ T lymphocytes were responsive to PL Ag
stimulation, resulting in IL-4 secretion, dose-dependent secretion of IFN-g and a strong secretion of TGF-b [23]. In contrast, there was no significant difference in secretion of IL-10 when comparing PL stimulated and unstimulated cells [23]. Colonic LPL cytokine secretion was examined by Makita et al. [27] using intracytofluorometric analysis. When stimulated with PMA and CaI2, LP CD4+CD25- cells secreted IL-2 and IFN-g but not IL-10, whereas LP
In summary: most duodenal TCRab+ T lymphocytes secrete IFN-g and TNF-a, and upon PL
stimulation they also secrete large amounts of IL-4 and TGF-b1. TCRgd+ T lymphocytes
secreted large amounts of TGF-b1 and dose-dependent amounts of IFN-g after PL stimulation [23]. Colonic CD4+CD25-, but not CD4+CD25bright, cells LPLs secrete IL-2 and IFN-g. Neither
type of colonic LPL secreted IL-10 [27].
Ileal and colonic Vg9/Vd2 T lymphocytes (not specified whether these were IELs or LPLs) were analyzed by Tyler et al. [29] using FACS, quantitative PCR and ELISA. This cell subset is previously known to have characteristics of professional APCs. The authors examined their ability to induce IL-22 secretion by CD4+ T lymphocytes. They found that IL-15, but not IL-6,
is required for driving gd T lymphocytes to become CCR9+, or gut-homing, and also for the
induction of IL-22 secretion [29]. Vg9/Vd2 T lymphocytes were able to induce expression of IL-22 and calprotectin from the mucosa, but they did not induce expression of IL-17 [29].
5 DISCUSSION
The results described above clearly show that there are differences in the properties of the immune system when comparing different anatomical regions of the human intestine. In summary, the small intestine seems to have a more pro-inflammatory and cytotoxic profile compared to the regulatory and tolerogenic colonic milieu.
Two studies showed that DCs in the ileum and colon have an immature phenotype [2,12]. There were, however, differences regarding other properties of DCs from the two compartments, such as cytokine profile and expression of cell surface markers. ILT3 was expressed at higher levels on colonic DC compared to ileal DC which supports a more regulatory role for colonic DC [2]. What is interesting is that the major subtype of DC expressing ILT3 were also negative for CD103, a marker involved in inducing Tregs and tolerance. However, CD103+ DCs were present
in both the ileum and he colon [2]. A regulatory milieu in the colon is probably necessary for the possibility of having a diverse microbiota that does not induce inflammation and is beneficial for intestinal homeostasis. The small intestine does not have the high bacterial load that is seen in the colon and might therefore not be in need of as strong regulatory properties. Colonic DC had an increased ability to induce Treg [2] and these were found in a higher
proportion in the colon compared to the ileum, further supporting a regulatory milieu [18]. Other DC properties that were increased in the colon compared to the ileum were endocytosis and migration [2]. Migratory colonic DC showed a higher maturation status compared to
endocytic colonic DC [12]. There was also a higher proportion of TH17 effector cells and DN
T cells compared in colon compared to the ileum [18], but bias might occur due to the analysis of a mixture of IELs and LPLs.
The higher proinflammatory profile of ileal compared to colonic DCs and the higher cytotoxicity of ileal compared to T lymphocytes [2] might be a clue to why the small intestine is rarely affected by malignant disease: This could maybe contribute to a better functioning immunosurveillance and inhibit the initiation of small intestinal metaplasia.
Ileal, but not colonic, DC were able to respond to leptin stimulation, causing them to express CCR7 and become migratory [6]. IELs had a higher total cytokine secretion compared to LPLs [25], possibly because of closer proximity to luminal antigens.
Expression of homing receptors showed anatomical differences: Whereas ileal DC were CCR9+, colonic DC were CCR4+, markers that were reflected in their imprinting ability on T
lymphocytes: Ileal DC imprinted CCR9 expression whereas colonic DC imprinted CCR4 expression on T lymphocytes [2].
T lymphocyte subsets showed differences in cytokine secretion in the ileum compared to the colon. In the small intestine there was a higher secretion of IFN-g, IL-17A and IL-4 compared to the colon, were secretion of these cytokines and also IL-2 was low or absent [18,25,26]. However, one study saw that colonic CD4+ LPLs did secrete IL-2 and IFN-g when stimulated
with PMA [27]. The different results might be explained by the use of different methods and the examination of different cell subtypes such as IELs and LPLs or a mixture of these. Albeit this study shows that there are some anatomical differences in the distribution and function of DCs and T lymphocytes when comparing the small and large intestine, the results should be looked at with a critical eye. The included articles have used different methods for their analyses and they examined different cell subtypes, cytokines and surface markers which means that the results are not directly comparable. Different laboratory methods are used for different purposes and they all have their advantages and disadvantages. Another aspect that might be a limitation to this study is the fact that not all the studies were considered high quality in the scientific assessment. Also, all the articles included were searched only from PubMed which might have contributed to selection bias. Additional possible weaknesses with the present study are that all the articles were read and evaluated by one person only and that not all the included articles examined both the small and large intestine.
In the introduction the microbiota of the intestine was briefly presented as a source of antigen exposure. There are large individual variations in the microbiota, depending on diet and other environmental factors, which is highly likely to contribute to differences in the physiology of the intestines and the function of the immune system. The geographical areas of the included articles are mostly part of the Western world, except from one that was from Japan. If studies from other parts of the world would have been included in this systematic review the results might have become different because of differences in lifestyle habits resulting in other profiles of the microbiota.
6 CONCLUSION
The present study shows that there are differences in the distribution and properties and DCs and T lymphocytes in the small intestine compared to the colon, with a more pro-inflammatory and cytotoxic profile in the small intestine compared to the more regulatory and tolerogenic milieu in the colon. More studies investigating these differences could contribute to better understanding on the immunological properties that lead to disease in the human intestine. New findings might contribute to the development of better treatment possibilities of diseases affecting the intestines.
7 SPECIAL THANKS TO
I express special thanks to my supervisor Elisabeth Hultgren Hörnquist, who has helped and supported me with her time and knowledge throughout the whole project. Thank you for all the helpful encouragement. I feel fortunate to have had you has my supervisor.
8 REFERENCES
1. Mowat AM, Agace WW. Regional specialization within the intestinal immune system. Nature Reviews Immunology 2014;14:667–85.
2. Mann ER, Bernardo D, English NR, Landy J, Al-Hassi HO, Peake STC, et al. Compartment-specific immunity in the human gut: properties and functions of dendritic cells in the colon versus the ileum. Gut 2016;65:256–70.
3. Zhang Y-Z, Li Y-Y. Inflammatory bowel disease: Pathogenesis. World J Gastroenterol 2014;20:91–9.
4. Gorbach SL. Microbiology of the Gastrointestinal Tract [Internet]. In: Baron S, editor. Medical Microbiology. Galveston (TX): University of Texas Medical Branch at Galveston; 1996 [cited 2019 Jan 2]. Available from: http://www.ncbi.nlm.nih.gov/books/NBK7670/
5. Croxen MA, Law RJ, Scholz R, Keeney KM, Wlodarska M, Finlay BB. Recent
Advances in Understanding Enteric Pathogenic Escherichia coli. Clin Microbiol Rev 2013;26:822–80. 6. Al-Hassi HO, Bernardo D, Murugananthan AU, Mann ER, English NR, Jones A, et al. A mechanistic role for leptin in human dendritic cell migration: differences between ileum and colon in health and Crohn’s disease. Mucosal Immunol 2013;6:751–61.
7. Syed SA, Abrams GD, Freter R. Efficiency of Various Intestinal Bacteria in Assuming Normal Functions of Enteric Flora After Association with Germ-Free Mice. Infect Immun
1970;2:376–86.
8. van Wijk F, Cheroutre H. Intestinal T cells: Facing the mucosal immune dilemma with synergy and diversity. Semin Immunol 2009;21:130–8.
9. Weiner HL, da Cunha AP, Quintana F, Wu H. Oral tolerance. Immunol Rev 2011;241:241–59.
10. Bain CC, Montgomery J, Scott CL, Kel JM, Girard-Madoux MJH, Martens L, et al. TGFβR signalling controls CD103+CD11b+ dendritic cell development in the intestine. Nat Commun [Internet] 2017 [cited 2019 Jan 2];8. Available from:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5607002/
11. Wang C, McDonald KG, McDonough JS, Newberry RD. Murine isolated lymphoid follicles contain follicular B lymphocytes with a mucosal phenotype. Am J Physiol Gastrointest Liver Physiol 2006;291:G595–604.
12. Bell SJ, Rigby R, English N, Mann SD, Knight SC, Kamm MA, et al. Migration and maturation of human colonic dendritic cells. J. Immunol. 2001;166:4958–67.
13. Coombes JL, Powrie F. Dendritic cells in intestinal immune regulation. Nat Rev Immunol 2008;8:435–46.
14. Hopp A-K, Rupp A, Lukacs-Kornek V. Self-Antigen Presentation by Dendritic Cells in Autoimmunity. Front Immunol [Internet] 2014 [cited 2019 Jan 2];5. Available from:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3923158/
Mediators Inflamm [Internet] 2016 [cited 2018 Dec 29];2016. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4789473/
16. Scheinecker C, McHugh R, Shevach EM, Germain RN. Constitutive Presentation of a Natural Tissue Autoantigen Exclusively by Dendritic Cells in the Draining Lymph Node. J Exp Med 2002;196:1079–90.
17. Melgar S, Hammarström S, Oberg A, Danielsson A, Hammarström M-L. Cytolytic capabilities of lamina propria and intraepithelial lymphocytes in normal and chronically inflamed human intestine. Scand. J. Immunol. 2004;60:167–77.
18. Carrasco A, Fernández-Bañares F, Pedrosa E, Salas A, Loras C, Rosinach M, et al. Regional Specialisation of T Cell Subsets and Apoptosis in the Human Gut Mucosa: Differences Between Ileum and Colon in Healthy Intestine and Inflammatory Bowel Diseases. J Crohns Colitis 2016;10:1042–54.
19. Stevanin M, Busso N, Chobaz V, Pigni M, Ghassem-Zadeh S, Zhang L, et al. CD11b regulates the Treg/Th17 balance in murine arthritis via IL-6. European Journal of Immunology 2017;47:637–45.
20. Koenen HJPM, Smeets RL, Vink PM, Rijssen E van, Boots AMH, Joosten I. Human CD25highFoxp3pos regulatory T cells differentiate into IL-17–producing cells. Blood
2008;112:2340–52.
21. Weitkamp J-H, Rudzinski E, Koyama T, Correa H, Matta P, Alberty B, et al. Ontogeny of FOXP3(+) regulatory T cells in the postnatal human small intestinal and large intestinal lamina propria. Pediatr. Dev. Pathol. 2009;12:443–9.
22. Gagliani N, Vesely MCA, Iseppon A, Brockmann L, Xu H, Palm NW, et al. Th17 cells transdifferentiate into regulatory T cells during resolution of inflammation. Nature 2015;523:221–5. 23. Russano AM, Bassotti G, Agea E, Bistoni O, Mazzocchi A, Morelli A, et al. CD1-restricted recognition of exogenous and self-lipid antigens by duodenal gammadelta+ T lymphocytes. J. Immunol. 2007;178:3620–6.
24. Method [Internet]. [cited 2019 Jan 3];Available from: https://www.sbu.se/en/method/ 25. Carol M, Lambrechts A, Van Gossum A, Libin M, Goldman M, Mascart-Lemone F. Spontaneous secretion of interferon gamma and interleukin 4 by human intraepithelial and lamina propria gut lymphocytes. Gut 1998;42:643–9.
26. Breese E, Braegger CP, Corrigan CJ, Walker-Smith JA, MacDonald TT. Interleukin-2- and interferon-gamma-secreting T cells in normal and diseased human intestinal mucosa. Immunology 1993;78:127–31.
27. Makita S, Kanai T, Oshima S, Uraushihara K, Totsuka T, Sawada T, et al. CD4+CD25bright T cells in human intestinal lamina propria as regulatory cells. J. Immunol. 2004;173:3119–30.
28. Papadakis KA, Prehn J, Nelson V, Cheng L, Binder SW, Ponath PD, et al. The Role of Thymus-Expressed Chemokine and Its Receptor CCR9 on Lymphocytes in the Regional
29. Tyler CJ, McCarthy NE, Lindsay JO, Stagg AJ, Moser B, Eberl M. Antigen-Presenting Human γδ T Cells Promote Intestinal CD4+ T Cell Expression of IL-22 and Mucosal Release of Calprotectin. J. Immunol. 2017;198:3417–25.
30. Ebert EC. Tumour necrosis factor-alpha enhances intraepithelial lymphocyte proliferation and migration. Gut 1998;42:650–5.
31. Roe MM, Swain S, Sebrell TA, Sewell MA, Collins MM, Perrino BA, et al.
Differential regulation of CD103 (αE integrin) expression in human dendritic cells by retinoic acid and Toll-like receptor ligands. J Leukoc Biol 2017;101:1169–80.
32. MIP-3 beta | CCL19 | Human Macrophage Inflammatory Protein-3 beta | Cell
Applications [Internet]. [cited 2018 Dec 29];Available from: https://www.cellapplications.com/mip-3-ccl19-human-macrophage-inflammatory-protein-3-beta
33. Cella M, Döhring C, Samaridis J, Dessing M, Brockhaus M, Lanzavecchia A, et al. A Novel Inhibitory Receptor (ILT3) Expressed on Monocytes, Macrophages, and Dendritic Cells Involved in Antigen Processing. J Exp Med 1997;185:1743–51.
9 APPENDIX
Table 1: Summary of articles included.
Articles and
year of publication
Geografic area
Intestinal region Cell type(s)
examined Scientific assessment Bell, 2001 [12] United Kingdom Colon DC Moderate Mann, 2016 [2] United Kingdom Ileum Colon DC High Al-Hassi, 2013 [6] United Kingdom Ileum Colon DC High Papadakis, 2000 [28] USA Ileum Colon LPL High Carrasco, 2016 [18]
Spain Terminal ileum Colon TH17, TH1, DP, DN, Treg Moderate Carol, 1998 [25] Belgium Duodenum Colon IEL, LPL Moderate Melgar, 2004 [17] Sweden Ileum Colon IEL, LPL High Breese, 1993 [26] United Kingdom
Colon T-cells Moderate
Ebert, 1997 [30]
USA Jejunum IEL, LPL Moderate
Russano, 2007 [23]
USA Duodenum IEL Moderate
Tyler, 2017 [29] United Kingdom Terminal ileum Colon T-cells Moderate Makita, 2004 [27]
