Breast cancer is the most common cancer in women worldwide. Metastasis, the process by which cancer cells spread to other parts of the body, makes treatments less effective. It is important to... Show moreBreast cancer is the most common cancer in women worldwide. Metastasis, the process by which cancer cells spread to other parts of the body, makes treatments less effective. It is important to unravel the mechanism of metastasis. The epithelial-mesenchymal transition (EMT) is one such mechanism involved in cancer progression and metastasis. EMT is regulated by a network of transcription factors including Grainyhead-like 2 (GRHL2). GRHL2 is an important transcription factor for cell epithelial identity. GRHL2 has been described as capable of stimulating tumor growth but at the same time suppressing EMT and therefore could inhibit metastasis. This dissertation focuses on GRHL2 and the genes controlled by this transcription factor. I investigated the functions of GRHL2 in different subtypes of breast cancer by turning off or on the GRHL2 gene. I also analyzed the changes in EMT caused by the manipulations of GRHL2 in different subtypes of breast cancer. The results show that GRHL2 can play different, and subtype-specific roles. Although this is basic research, it provides new insights that may lead to targeted treatments. Among other things, my research shows how GRHL2 regulates the interaction of tumors with the immune system and how it affects metastasis in breast cancer. This suggests that it deserves further investigation as a potential factor in the development of therapies. Show less
Forty years after the first successful oocyte donation (OD), this technique has become a crucial part of assisted reproductive technology (ART). While OD has significantly improved pregnancy rates... Show moreForty years after the first successful oocyte donation (OD), this technique has become a crucial part of assisted reproductive technology (ART). While OD has significantly improved pregnancy rates for infertile couples, it also brings a higher risk of complications like pre-eclampsia, which occurs two to four times more often in OD pregnancies than in standard IVF pregnancies. This research investigates the unique immune challenges in OD pregnancies, where the fetus inherits genes from the donor and father, leading to greater genetic differences between the mother and fetus. By studying pathology lesions, maternal immune cell composition and their microenvironment in the placenta, along with immune cells in maternal blood, I discovered that with high fetal-maternal gene dissimilarities in healthy OD pregnancies, an immune regulatory/tolerating environment may be established locally in the placenta, together with immune alterations in maternal peripheral blood. On the other hand, in OD pregnancies without sufficient immune regulation pre-eclampsia might occur. These findings enhance our understanding of the role of maternal immune cells in OD pregnancies and offer insights into both healthy and pre-eclamptic conditions, which could potentially lead to improved clinical management strategies for these complex pregnancies in the future. Show less
Healthy aging is one of the prime goals in today's society and atherosclerosis is among the greatest causes of morbidity in elderly. Cardiovascular disease patients receiving treatment are often of... Show moreHealthy aging is one of the prime goals in today's society and atherosclerosis is among the greatest causes of morbidity in elderly. Cardiovascular disease patients receiving treatment are often of advanced aged and have an aged immune system, which limits translating experimental findings to the patient. It is therefore essential to take aging into consideration when investigating immune cells and their responses in atherosclerosis studies. This thesis describes research exploring the impact of aging on the immunological landscape in atherosclerotic cardiovascular disease using single-cell profiling. Through the use of a highly translational aging mouse model of atherosclerosis, we characterized inflammation in the plaques of young versus old mice. We discovered new cell types (T and B cells) not present in young mice with atherosclerosis. These cells secrete a variety of inflammatory factors that may contribute to the disease process and exacerbate arteriosclerosis. While the aged B cell is more prevalent in female mice, the aged T cell is more abundant in male mice. We then also found these aged cells in the blood and plaques of cardiovascular disease patients. These aged cell types could be interesting targets for future treatments against progression of atherosclerotic cardiovascular disease. Show less
This thesis contains several investigations into the contribution of complement proteins, especially C1q, in several human diseases. Additionally, human autoantibodies against C1q (anti-C1q) are... Show moreThis thesis contains several investigations into the contribution of complement proteins, especially C1q, in several human diseases. Additionally, human autoantibodies against C1q (anti-C1q) are studied, cloned and characterized in order to further the understanding of their role in autoimmune disease. Show less
Smit, V.; Mol, J. de; Bernabé Kleijn, M.N.A.B.; Depuydt, M.A.C.; Winther, M.P.J. de, Bot, I.; Kuiper, J.; Foks, A.C. 2024
Atherosclerosis, the main underlying pathology of cardiovascular disease, is a chronic inflammatory disease characterized by lipid accumulation and immune cell responses in the vascular wall,... Show moreAtherosclerosis, the main underlying pathology of cardiovascular disease, is a chronic inflammatory disease characterized by lipid accumulation and immune cell responses in the vascular wall, resulting in plaque formation. It is well-known that atherosclerosis prevalence and manifestation vary by sex. However, sexual dimorphism in the immune landscape of atherosclerotic plaques has up to date not been studied at high-resolution. In this study, we investigated sex-specific differences in atherosclerosis development and the immunological landscape of aortas at single-cell level in aged Ldlr-/- mice.We compared plaque morphology between aged male and female chow diet-fed Ldlr-/- mice (22 months old) with histological analysis. Using single-cell RNA-sequencing and flow cytometry on CD45+ immune cells from aortas of aged Ldlr-/- mice, we explored the immune landscape in the atherosclerotic environment in males and females.We show that plaque volume is comparable in aged male and female mice, and that plaques in aged female mice contain more collagen and cholesterol crystals, but less necrotic core and macrophage content compared to males. We reveal increased immune cell infiltration in female aortas and found that expression of pro-atherogenic markers and inflammatory signaling pathways was enriched in plaque immune cells of female mice. Particularly, female aortas show enhanced activation of B cells (Egr1, Cd83, Cd180), including age-associated B cells, in addition to an increased M1/M2 macrophage ratio, where Il1b+ M1-like macrophages display a more pro-inflammatory phenotype (Nlrp3, Cxcl2, Mmp9) compared to males. In contrast, increased numbers of age-associated Gzmk+CD8+ T cells, dendritic cells, and Trem2+ macrophages were observed in male aortas.Altogether, our findings highlight that sex is a variable that contributes to immunological differences in the atherosclerotic plaque environment in mice and provide valuable insights for further preclinical studies into the impact of sex on the pathophysiology of atherosclerosis. Show less
Acute cardiovascular syndromes, including myocardial infarction or stroke, are the principal cause of death in the Western society. The main underlying pathology of cardiovascular diseases is... Show moreAcute cardiovascular syndromes, including myocardial infarction or stroke, are the principal cause of death in the Western society. The main underlying pathology of cardiovascular diseases is atherosclerosis, which is caused by the accumulation of lipids and inflammatory cells in the vessel wall, in so-called atherosclerotic plaques. Current therapies mainly target the disturbed lipid homeostasis, but recent clinical trials have shown a clear benefit in treating patients with anti-inflammatory drugs. However, more specific targeting is required to avoid unwanted side effects. In this thesis, we have generated a detailed atlas of all the cells present in human atherosclerotic plaques using a novel state-of-the-art technique called single-cell RNA sequencing. This data set can be applied as a powerful tool to select potential drug targets with a functional relevance for atherosclerosis. We showed that the majority of the immune cells in the human atherosclerotic plaque consisted of T cells. Subsequently, we identified a pro-inflammatory population of T cells that likely responds to a plaque-derived antigen, suggesting that atherosclerosis has an autoimmune-like component. Finally, we have applied our single-cell atlas to define and validate targets to intervene with the recruitment and activation of mast cells and other immune cells in atherosclerosis. Show less
This work has described synthetic strategies towards well-defined structures resembling capsular polysaccharide (CPS) fragments, CPS mimics, teichoic acid (TA) fragments as well as a third... Show moreThis work has described synthetic strategies towards well-defined structures resembling capsular polysaccharide (CPS) fragments, CPS mimics, teichoic acid (TA) fragments as well as a third-generation ring-closing tandem metathesis (RCM) linker to better exploit the potential of automated synthesis. The synthesis of diheteroglycan (DHG) fragments of the Gram-positive Enterococcus faecalis have been described and preliminary biological properties shown. CPS-mimics of Neisseria meningitidis A, a Gram-negative bacterium and one of the major causes of bacterial meningitis, could provide an alternative to the inherent instability of current glycoconjugate vaccines. TA fragments presented in this thesis, typical for Gram-positive bacteria, have been equipped with labile D-alanine appendages to further investigate their biological relevance in interactions with the host immune system. A tandem-RCM linker has been developed and tested on a carbohydrate automated synthesizer, providing a proof of concept of its use. Show less
Kidney transplantation is the best treatment option for patients with kidney failure. Unfortunately many patients lose their allograft due to (chronic) rejection. Rejection is caused by the immune... Show moreKidney transplantation is the best treatment option for patients with kidney failure. Unfortunately many patients lose their allograft due to (chronic) rejection. Rejection is caused by the immune reaction of the recipient against the donor’s human leukocyte antigens (HLA). While traditional kidney allocation is based on HLA matching on the antigen level, matching on the epitope level could be more feasible. Furthermore, epitope mismatch analysis could be used for post-transplant risk stratification, enabling the personalisation of immunosuppressive treatment for kidney transplant recipients. In this thesis, the basic science and clinical application of HLA epitopes in kidney transplantation are discussed. Show less
Colorectal cancer is one of the most diagnosed cancer types worldwide and incidence remains on the rise, especially in patients under 50. The prognosis for patients with CRC differs greatly and... Show moreColorectal cancer is one of the most diagnosed cancer types worldwide and incidence remains on the rise, especially in patients under 50. The prognosis for patients with CRC differs greatly and although immunotherapy has shown promising results in a number of cancer types, not all CRC patients respond well to these treatments. This can in part be attributed to the differences in T cell infiltration between cancers but does not one on one translate to clinical response. Moreover, the activity of specific immune cells can directly influence other immune cells, both in an activating and inhibitory manner. This highlights the complexity of the tumour immune microenvironment and requires an comprehensive multiplex approach to simultaneously investigate all the players of the tumour immune microenvironment. Furthermore, the interaction between different immune cells and between those and cancer cells is essential to take into account, hence the need for an approach that combines multiplex immunophenotyping with spatial cell context. This will provide hints into the behaviour of the players of the tumour immune microenvironment and aid the understanding of CRC, but potentially of other cancer types as well. In this work we developed and applied multispectral immunophenotyping methodologies to strengthen our understanding of CRC Show less
Fosse, N.A. du; Lashley, E.E.L.O.; Anholts, J.D.H.; Beelen, E. van; Cessie, S. le; Lith, J.M.M. van; ... ; Hoorn, M.L.P. van der 2022
Background: Seminal plasma contains signaling molecules capable of modulating the maternal immune environment to support implantation and pregnancy. Prior studies indicated that seminal plasma... Show moreBackground: Seminal plasma contains signaling molecules capable of modulating the maternal immune environment to support implantation and pregnancy. Prior studies indicated that seminal plasma induces changes in gene transcription of maternal immune cells. Reduced immune suppressive capacity may lead to pregnancy loss. The aim of this study was to investigate the immunomodulating effects of seminal plasma on T cells and monocytes in the context of recurrent pregnancy loss (RPL).Methods: Female T cells and monocytes were incubated with seminal plasma of 20 males in unexplained RPL couples (RPL males) and of 11 males whose partners had ongoing pregnancies (control males). The effect of seminal plasma on messenger RNA (mRNA) expression of immune cells was measured. Levels of mRNA expression were related to key signaling molecules present in the seminal plasma. Agglomerative hierarchical cluster analysis was performed on seminal plasma expression profiles and on mRNA expression profiles. Results: Expression of CD25 and anti-inflammatory IL-10 by female T cells was significantly lower after stimulation with seminal plasma of RPL males compared to control males. Female monocytes treated with seminal plasma of RPL males showed an immune activation signature of relatively elevated HLA-DR expression. Expression of these T cell and monocyte components was particularly correlated with the amounts of TGF-beta and VEGF in the seminal plasma. Conclusion: Our findings indicate that seminal plasma has immunomodulating properties on female immune cells compatible with the induction of a more regulatory phenotype, which may be impaired in cases of unexplained RPL. Show less
Skin-penetrating parasites have something in common; they all need to evade the initial immune response in the skin in order to avoid being evicted by their hostile host and establish an infection.... Show moreSkin-penetrating parasites have something in common; they all need to evade the initial immune response in the skin in order to avoid being evicted by their hostile host and establish an infection. To do so, they are equipped with the necessary cunning stratagems. For example, they can act directly on immune cells to alter their function and they can optimize their migration patterns to their hostile environment. This thesis is aimed at unravelling those mechanisms. We study two devastating parasitic diseases: Malaria and Schistosomiasis. Both deadly and debilitating parasitic diseases, with over 200 million (malaria) and over 240 million (schistosomiasis) cases annually; the need for potent vaccines is evident. Whole weakened parasites can be used to vaccinate individuals against parasitic diseases like malaria. However, delivery of these parasites in the skin, as is commonly done in vaccinations, reduces their protectivity. We hypothesize that this reduction is caused by parasite-mediated immune-regulatory mechanisms that are initiated upon their first encounter with immune cells in the skin. We investigated whether skin penetrating parasites exploit these existing mechanisms in human skin in order to enhance their survival. Show less
This dissertation covered several relevant cycles of placebo research with the main aim to optimize placebo effects in medical contexts. Firstly, a literature review described how the immune system... Show moreThis dissertation covered several relevant cycles of placebo research with the main aim to optimize placebo effects in medical contexts. Firstly, a literature review described how the immune system can be impacted by placebo effects and their underlying learning theories. In the following chapter, these learning theories were integrated to form an optimal research design by means of pharmacological conditioning to fit a specific patient group: children with juvenile idiopathic arthritis. Secondly, this dissertation focused on developing placebo information strategies to harness placebo beliefs and educate persons about the relevancy of placebo effects in practice. These insights are valuable because treatment expectations can have a positive or negative effect on treatment outcomes. Finally, insights from placebo learning theories and placebo information strategies were combined in an integrative experimental research design. This research design employed a more ethical form of placebo use because participants were made aware of placebos, called open-label placebos. In this last study we demonstrated that open-label placebo analgesia can be induced by combining learning theories and placebo information strategies. Altogether, this dissertation provided insights in learning mechanisms, communication strategies, and research paradigms that involve the optimization of placebo effects in medical context. Show less
Agrawal, R.; Testi, I.; Lee, C.S.; Tsui, E.; Blazes, M.; Thorne, J.E.; ... ; COVID-19 Imt Study Grp 2021
Background Immunomodulatory therapy (IMT) is often considered for systemic treatment of non-infectious uveitis (NIU). During the evolving coronavirus disease-2019 (COVID-19) pandemic, given the... Show moreBackground Immunomodulatory therapy (IMT) is often considered for systemic treatment of non-infectious uveitis (NIU). During the evolving coronavirus disease-2019 (COVID-19) pandemic, given the concerns related to IMT and the increased risk of infections, an urgent need for guidance on the management of IMT in patients with uveitis has emerged. Methods A cross-sectional survey of international uveitis experts was conducted. An expert steering committee identified clinical questions on the use of IMT in patients with NIU during the COVID-19 pandemic. Using an interactive online questionnaire, guided by background experience and knowledge, 139 global uveitis experts generated consensus statements for IMT. In total, 216 statements were developed around when to initiate, continue, decrease and stop systemic and local corticosteroids, conventional immunosuppressive agents and biologics in patients with NIU. Thirty-one additional questions were added, related to general recommendations, including the use of non-steroidal anti-inflammatory drugs (NSAIDs) and hydroxychloroquine. Results Highest consensus was achieved for not initiating IMT in patients who have suspected or confirmed COVID-19, and for using local over systemic corticosteroid therapy in patients who are at high-risk and very high-risk for severe or fatal COVID-19. While there was a consensus in starting or initiating NSAIDs for the treatment of scleritis in healthy patients, there was no consensus in starting hydroxychloroquine in any risk groups. Conclusion Consensus guidelines were proposed based on global expert opinion and practical experience to bridge the gap between clinical needs and the absence of medical evidence, to guide the treatment of patients with NIU during the COVID-19 pandemic. Show less
This thesis contains a variety of information about the natural and vaccine induced immunity against the human papillomavirus. The spontaneously induced HPV-specific humoral response after... Show moreThis thesis contains a variety of information about the natural and vaccine induced immunity against the human papillomavirus. The spontaneously induced HPV-specific humoral response after infection was assessed in population-based studies. The vaccine-induced changes in HPV-seroprevalence among the HPV unvaccinated Dutch population aged 0-89 years, where we compared the HPV-seroprevalence before the introduction of the HPV vaccine with data of approximately six years post-implementation of the national HPV vaccination program. Also, the HPV immune status of the Dutch Caribbean population just after introduction of HPV vaccination was determined. Moreover, the longitudinal relation between the hr-HPV antibody levels and the prevalence of HPV infections in three-dose vaccinated girls were studied. And more insight was gained into humoral and cellular immune responses after just a one-dose of the HPV vaccine. At last, the kinetics of innate and adaptive immune responses directly after vaccination different HPV vaccines were investigated. In the coming years some important changes are expected regarding HPV screening and vaccination. The effectiveness of the one-dose schedule will become clear as clinical trials end. In the Netherlands, a sex-neutral vaccination will be implemented soon. These changes will need to be monitored to provide scientific answers about the effectiveness and immunogenicity. Show less
BackgroundInterferon-gamma release assays (IGRA) with Resuscitation promoting factor (Rpf) proteins enhanced tuberculosis (TB) screening and diagnosis in adults but have not been evaluated in... Show moreBackgroundInterferon-gamma release assays (IGRA) with Resuscitation promoting factor (Rpf) proteins enhanced tuberculosis (TB) screening and diagnosis in adults but have not been evaluated in children. Children often develop paucibacillary TB and their immune response differs from that of adults, which together affect TB disease diagnostics and immunodiagnostics. We assessed the ability of Rpf to identify infection among household TB-exposed children in The Gambia and investigated their ability to discriminate Mycobacterium tuberculosis complex (MTBC) infection from active TB disease in children.MethodsDetailed clinical investigations were done on 93 household TB-exposed Gambian children and a tuberculin skin test (TST) was administered to asymptomatic children. Venous blood was collected for overnight stimulation with ESAT-6/CFP-10-fusion protein (EC), purified protein derivative and RpfA, B, C, D and E. Interferon gamma (IFN-gamma) production was measured by ELISA in supernatants and corrected for the background level. Infection status was defined by IGRA with EC and TB disease by mycobacterial confirmation and/or clinical diagnosis. We compared IFN-gamma levels between infected and uninfected children and between infected and TB diseased children using a binomial logistic regression model while correcting for age and sex. A Receiver Operating Characteristics analysis was done to find the best cut-off for IFN-gamma level and calculate sensitivity and specificity.ResultsInterferon gamma production was significantly higher in infected (IGRA+, n=45) than in uninfected (IGRA-, n=20) children after stimulation with RpfA, B, C, and D (P=0.03; 0.007; 0.03 and 0.003, respectively). Using RpfB and D-specific IFN-gamma cut-offs (33.9pg/mL and 67.0pg/mL), infection was classified with a sensitivity-specificity combination of 73-92% and 77-72% respectively, which was similar to and better than 65-75% for TST. Moreover, IFN-gamma production was higher in infected than in TB diseased children (n=28, 5 bacteriologically confirmed, 23 clinically diagnosed), following RpfB and D stimulation (P=0.02 and 0.03, respectively).ConclusionRpfB and RpfD show promising results for childhood MTBC infection screening, and both performed similar to and better than the TST in our study population. Additionally, both antigens appear to discriminate between infection and disease in children and thus warrant further investigation as screening and diagnostic antigens for childhood TB. Show less
Pregnancy can be seen as an immunologic paradox. Even though the fetus expresses paternally inherited alloantigens it is protected from rejection by a proper regulation of the maternal immune... Show morePregnancy can be seen as an immunologic paradox. Even though the fetus expresses paternally inherited alloantigens it is protected from rejection by a proper regulation of the maternal immune system. With the studies described in this thesis, we want to get more insight in the immunologic mechanisms that play a role in pregnancy. The results of this research can help to identify underlying etiologies in patients with unexplained pregnancy complications, such as recurrent miscarriage. Identifying these causes is important for providing answers and taking away anxiety in these couples, and eventually for the development of effective therapies. Furthermore, elucidating the mechanism leading to survival or rejection of the fetal allograft is not only essential for our understanding of processes leading to normal and abnormal pregnancies, but may also result in important concepts in the field of transplantation and autoimmunity. Show less
Atherosclerosis is the most important underlying process that drives cardiovascular disease, and is characterized by an accumulation of cholesterol which triggers an inflammatory response in the... Show moreAtherosclerosis is the most important underlying process that drives cardiovascular disease, and is characterized by an accumulation of cholesterol which triggers an inflammatory response in the vessel wall. This results in the recruitment of many types of inflammatory cells towards the plaques that form in the vessel wall, among which are CD8+ T-cells. In this thesis, the role of CD8+ T-cells in the advanced stages of lesion development has been investigated, as this is the most clinically relevant stage of the disease. This thesis demonstrates that CD8+ T-cells exert a protective function. We show that the absence of CD8+ T-cells in a mouse model results in less stable atherosclerotic lesions with increased numbers of inflammatory cells. In a subsequent study, we show that CD8+ T-cells express an enzyme that inhibits the inflammatory process. We also show that injecting a specific subset of CD8+ T-cells is protective against the development of atherosclerotic lesions in mice. Importantly, we show that this data can be translated to atherosclerosis development in humans, as we demonstrate similar results using patient material obtained from endarterectomy surgery. Finally, we show that developing therapies directed towards activating CD8+ T-cells may be of value to inhibit the immune response, and thus reduce the risk of cardiovascular disease. Show less
During pregnancy a unique situation arises in which the mother's immune system accepts the fetus, which carries both maternal and paternal genes, and does not reject it as can occur in solid organ... Show moreDuring pregnancy a unique situation arises in which the mother's immune system accepts the fetus, which carries both maternal and paternal genes, and does not reject it as can occur in solid organ transplantation. The aim of this dissertation was to unravel the immunological mechanisms that ensure tolerance during a healthy pregnancy and uncover how alterations could contribute to the development of pregnancy complications, such as pre-eclampsia and preterm birth.We applied the new technique mass cytometry and the associated computational analyzes to map all immune cells of the mother during a healthy pregnancy. Furthermore, we demonstrated the presence of three types of functional regulatory CD4+ T cells, identified a phenotype of CD8+ T cells that can offer both tolerance and immunity against infections, and demonstrated potential cross-reactivity of T cells against fetal allo-antigens. The results described in this thesis have contributed to a better understanding of healthy pregnancies and form a basis on which further research can be built. Show less
