Publications by authors named "Boris Novakovic"

62 Publications

Lysine methyltransferase G9a is an important modulator of trained immunity.

Clin Transl Immunology 2021 18;10(2):e1253. Epub 2021 Feb 18.

Department of Internal Medicine Radboud Center for Infectious Diseases (RCI) Radboud University Medical Center Nijmegen The Netherlands.

Objectives: Histone methyltransferase G9a, also known as Euchromatic Histone Lysine Methyltransferase 2 (EHMT2), mediates H3K9 methylation which is associated with transcriptional repression. It possesses immunomodulatory effects and is overexpressed in multiple types of cancer. In this study, we investigated the role of G9a in the induction of trained immunity, a innate immune memory, and its effects in non-muscle-invasive bladder cancer (NMIBC) patients treated with intravesical Bacillus Calmette-Guérin (BCG).

Methods: EHMT2 expression was assessed upon induction of trained immunity by RNA sequencing and Western blotting. G9a inhibitor BIX-01294 was used to investigate the effect on trained immunity responses . Subsequent cytokine production was measured by ELISA, epigenetic modifications were measured by ChIP-qPCR, Seahorse technology was used to measure metabolic changes, and a luminescence assay was used to measure ROS release. RNA sequencing was performed on BIX-01294-treated monocytes .

Results: The expression of EHMT2 mRNA and protein decreased in monocytes during induction of trained immunity. G9a inhibition by BIX-01294 induced trained immunity and amplified trained immunity responses evoked by various microbial ligands . This was accompanied by decreased H3K9me2 at the promoters of pro-inflammatory genes. G9a inhibition was also associated with amplified trained immunity responses in circulating monocytes of NMIBC patients. Additionally, altered RNA expression of inflammatory genes in monocytes of NMIBC patients was observed upon G9a inhibition. Furthermore, intravesical BCG therapy decreased H3K9me2 at the promoter of pro-inflammatory genes.

Conclusion: Inhibition of G9a is important in the induction of trained immunity, and G9a may represent a novel therapeutic target in NMIBC patients.
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http://dx.doi.org/10.1002/cti2.1253DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7890679PMC
February 2021

Sexual Dimorphism in Innate Immunity: The Role of Sex Hormones and Epigenetics.

Front Immunol 2020 21;11:604000. Epub 2021 Jan 21.

Epigenetics Group, Infection and Immunity Theme, Murdoch Children's Research Institute, Royal Children's Hospital, Parkville, VIC, Australia.

Sexual dimorphism refers to differences between biological sexes that extend beyond sexual characteristics. In humans, sexual dimorphism in the immune response has been well demonstrated, with females exhibiting lower infection rates than males for a variety of bacterial, viral, and parasitic pathogens. There is also a substantially increased incidence of autoimmune disease in females compared to males. Together, these trends indicate that females have a heightened immune reactogenicity to both self and non-self-molecular patterns. However, the molecular mechanisms driving the sexually dimorphic immune response are not fully understood. The female sex hormones estrogen and progesterone, as well as the male androgens, such as testosterone, elicit direct effects on the function and inflammatory capacity of immune cells. Several studies have identified a sex-specific transcriptome and methylome, independent of the well-described phenomenon of X-chromosome inactivation, suggesting that sexual dimorphism also occurs at the epigenetic level. Moreover, distinct alterations to the transcriptome and epigenetic landscape occur in synchrony with periods of hormonal change, such as puberty, pregnancy, menopause, and exogenous hormone therapy. These changes are also mirrored by changes in immune cell function. This review will outline the evidence for sex hormones and pregnancy-associated hormones as drivers of epigenetic change, and how this may contribute to the sexual dimorphism. Determining the effects of sex hormones on innate immune function is important for understanding sexually dimorphic autoimmune diseases, sex-specific responses to pathogens and vaccines, and how innate immunity is altered during periods of hormonal change (endogenous or exogenous).
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http://dx.doi.org/10.3389/fimmu.2020.604000DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7873844PMC
January 2021

BCG Vaccination Induces Long-Term Functional Reprogramming of Human Neutrophils.

Cell Rep 2020 Nov;33(7):108387

Department of Internal Medicine and Radboud Center for Infectious Diseases, Radboud University Medical Center, Nijmegen, the Netherlands; Department for Genomics & Immunoregulation, Life and Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany. Electronic address:

The tuberculosis vaccine bacillus Calmette-Guérin (BCG) protects against some heterologous infections, probably via induction of non-specific innate immune memory in monocytes and natural killer (NK) cells, a process known as trained immunity. Recent studies have revealed that the induction of trained immunity is associated with a bias toward granulopoiesis in bone marrow hematopoietic progenitor cells, but it is unknown whether BCG vaccination also leads to functional reprogramming of mature neutrophils. Here, we show that BCG vaccination of healthy humans induces long-lasting changes in neutrophil phenotype, characterized by increased expression of activation markers and antimicrobial function. The enhanced function of human neutrophils persists for at least 3 months after vaccination and is associated with genome-wide epigenetic modifications in trimethylation at histone 3 lysine 4. Functional reprogramming of neutrophils by the induction of trained immunity might offer novel therapeutic strategies in clinical conditions that could benefit from modulation of neutrophil effector function.
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http://dx.doi.org/10.1016/j.celrep.2020.108387DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7672522PMC
November 2020

DNA methylation profiling identifies epigenetic differences between early versus late stages of diabetic chronic kidney disease.

Nephrol Dial Transplant 2020 Nov 4. Epub 2020 Nov 4.

Epigenetics Research, Murdoch Children's Research Institute, Melbourne, Victoria, Australia.

Background: We investigated a cross-sectional epigenome-wide association study of patients with early and late diabetes-associated chronic kidney disease (CKD) to identify possible epigenetic differences between the two groups as well as changes in methylation across all stages of diabetic CKD. We also evaluated the potential of using a panel of identified 5'-C-phosphate-G-3' (CpG) sites from this cohort to predict the progression of diabetic CKD.

Methods: This cross-sectional study recruited 119 adults. DNA was extracted from blood using the Qiagen QIAampDNA Mini Spin Kit. Genome-wide methylation analysis was performed using Illumina Infinium MethylationEPIC BeadChips (HM850K). Intensity data files were processed and analysed using the minfi and MissMethyl packages for R. We examined the degree of methylation of CpG sites in early versus late diabetic CKD patients for CpG sites with an unadjusted P-value <0.01 and an absolute change in methylation of 5% (n = 239 CpG sites).

Results: Hierarchical clustering of the 239 CpG sites largely separated the two groups. A heat map for all 239 CpG sites demonstrated distinct methylation patterns in the early versus late groups, with CpG sites showing evidence of progressive change. Based on our differentially methylated region (DMR) analysis of the 239 CpG sites, we highlighted two DMRs, namely the cysteine-rich secretory protein 2 (CRISP2) and piwi-like RNA-mediated gene silencing 1 (PIWIL1) genes. The best predictability for the two groups involved a receiver operating characteristics curve of eight CpG sites alone and achieved an area under the curve of 0.976.

Conclusions: We have identified distinct DNA methylation patterns between early and late diabetic CKD patients as well as demonstrated novel findings of potential progressive methylation changes across all stages (1-5) of diabetic CKD at specific CpG sites. We have also identified associated genes CRISP2 and PIWIL1, which may have the potential to act as stage-specific diabetes-associated CKD markers, and showed that the use of a panel of eight identified CpG sites alone helps to increase the predictability for the two groups.
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http://dx.doi.org/10.1093/ndt/gfaa226DOI Listing
November 2020

Hyper-Inflammatory Monocyte Activation Following Endotoxin Exposure in Food Allergic Infants.

Front Immunol 2020 24;11:567981. Epub 2020 Sep 24.

Murdoch Children's Research Institute, Royal Children's Hospital, Parkville, VIC, Australia.

Several recent studies have reported a key role for innate cell hyper-responsiveness in food allergy. This has predominantly been observed in early life, with evidence that innate immune function may return to baseline if food allergy resolves in later childhood. Hallmarks of hyper-responsiveness include increased circulating frequency of monocytes and altered innate cell cytokine responses to exposure with bacterial endotoxin. These features mirror the defining signatures of trained innate immunity, seen in other complex diseases. In this study, detailed immune cell and cytokine profiling was performed on peripheral blood mononuclear cells at baseline from 27 1 year old infants in the HealthNuts cohort ( = 16 egg allergic and = 11 non-allergic healthy controls) and following monocyte stimulation. We show that egg allergic infants have increased frequency of circulating monocytes, reduced numbers of regulatory CD4 T cells and increased monocyte: CD4 T cell ratios relative to healthy controls. Monocytes from both egg allergic and non-allergic infants responded to endotoxin stimulation with rapid cytokine production and downregulation of the surface receptor CD16, however monocytes from egg allergic infants were hyper-responsive, producing significantly more inflammatory cytokines (TNFα, IL-6, IL-1β, IL-8) and innate cell recruiting factors (MIP-1α) than healthy controls. This work indicates that monocytes of food allergic infants are programmed to a hyper-inflammatory phenotype and that the development of food allergy may be associated with trained immunity in early life.
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http://dx.doi.org/10.3389/fimmu.2020.567981DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7541825PMC
September 2020

Determinants of placental leptin receptor gene expression and association with measures at birth.

Placenta 2020 10 18;100:89-95. Epub 2020 Aug 18.

Murdoch Children's Research Institute, Royal Children's Hospital, Parkville, Victoria, Australia; Department of Paediatrics, University of Melbourne, Parkville, Victoria, Australia. Electronic address:

Introduction: The leptin signalling pathway is important in metabolic health during pregnancy. However, few studies have investigated the determinants and extent of leptin receptor gene (LEPR) expression in the placenta, nor the relationship with infant health in early life. Here, we investigate the genetic and maternal in utero determinants of placental LEPR expression, and whether this expression is linked to anthropometric and inflammatory measures at birth in healthy newborns in the Barwon Infant Study.

Methods: Placental LEPR expression was measured using RT-qPCR (n = 854 placentae). Associations between genetic variation in LEPR, maternal in utero factors, measures at birth and placental LEPR expression were assessed using multivariable linear regression modelling.

Results: We found that the genotype at two intronic SNPs, rs9436301 and rs9436746, was independently associated with placental LEPR expression. Maternal pre-pregnancy body mass index, gestational diabetes mellitus, weight gain and smoking in pregnancy were not associated with LEPR expression. Placental LEPR expression was negatively associated with high sensitivity C-Reactive Protein in umbilical cord blood, which persisted after adjustment for potential confounders.

Discussion: Overall, our findings suggest that genetic variation in LEPR plays a key role in regulating placental LEPR expression, which is in turn is associated with inflammatory markers in cord blood at birth. Further studies encompassing other aspects of leptin signalling are warranted to understand if these relationships are causal and have health implications.
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http://dx.doi.org/10.1016/j.placenta.2020.08.010DOI Listing
October 2020

Sex matters: XIST and DDX3Y gene expression as a tool to determine fetal sex in human first trimester placenta.

Placenta 2020 08 26;97:68-70. Epub 2020 Jun 26.

Department of Obstetrics and Gynecology, Medical University of Graz, Graz, 8036, Austria. Electronic address:

Fetal sex influences placental function as well as maternal and fetal health, being an important factor to consider in pregnancy studies. However, fetal sex determination in the first trimester of pregnancy still faces some technical limitations. Here we describe an RT-qPCR technique to determine fetal sex based on X-inactive specific transcript (XIST) and DEAD-Box helicase 3 Y-linked (DDX3Y) gene expression. This method is straightforward, reliable, fast and applicable on both, placental tissue and primary cells.
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http://dx.doi.org/10.1016/j.placenta.2020.06.016DOI Listing
August 2020

Circadian rhythm influences induction of trained immunity by BCG vaccination.

J Clin Invest 2020 10;130(10):5603-5617

Radboud Center for Infectious Diseases and Department of Internal Medicine, Radboud University Medical Center, Nijmegen, Netherlands.

BACKGROUNDThe antituberculosis vaccine bacillus Calmette-Guérin (BCG) reduces overall infant mortality. Induction of innate immune memory, also termed trained immunity, contributes toward protection against heterologous infections. Since immune cells display oscillations in numbers and function throughout the day, we investigated the effect of BCG administration time on the induction of trained immunity.METHODSEighteen volunteers were vaccinated with BCG at 6 pm and compared with 36 age- and sex-matched volunteers vaccinated between 8 am and 9 am. Peripheral blood mononuclear cells were stimulated with Staphylococcus aureus and Mycobacterium tuberculosis before, as well as 2 weeks and 3 months after, BCG vaccination. Cytokine production was measured to assess the induction of trained immunity and adaptive responses, respectively. Additionally, the influence of vaccination time on induction of trained immunity was studied in an independent cohort of 302 individuals vaccinated between 8 am and 12 pm with BCG.RESULTSCompared with evening vaccination, morning vaccination elicited both a stronger trained immunity and adaptive immune phenotype. In a large cohort of 302 volunteers, early morning vaccination resulted in a superior cytokine production capacity compared with later morning. A cellular, rather than soluble, substrate of the circadian effect of BCG vaccination was demonstrated by the enhanced capacity to induce trained immunity in vitro in morning- compared with evening-isolated monocytes.CONCLUSIONSBCG vaccination in the morning induces stronger trained immunity and adaptive responses compared with evening vaccination. Future studies should take vaccine administration time into account when studying specific and nonspecific effects of vaccines; early morning should be the preferred moment of BCG administration.FUNDINGThe Netherlands Organization for Scientific Research, the European Research Council, and the Danish National Research Foundation.
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http://dx.doi.org/10.1172/JCI133934DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641012PMC
October 2020

A Potential Role for Epigenetically Mediated Trained Immunity in Food Allergy.

iScience 2020 Jun 17;23(6):101171. Epub 2020 May 17.

Murdoch Children's Research Institute, and Department of Paediatrics, University of Melbourne, Royal Children's Hospital, Flemington Road, Parkville, VIC 3052, Australia. Electronic address:

The prevalence of IgE-mediated food allergy is increasing at a rapid pace in many countries. The association of high food allergy rates with Westernized lifestyles suggests the role of gene-environment interactions, potentially underpinned by epigenetic variation, in mediating this process. Recent studies have implicated innate immune system dysfunction in the development and persistence of food allergy. These responses are characterized by increased circulating frequency of innate immune cells and heightened inflammatory responses to bacterial stimulation in food allergic patients. These signatures mirror those described in trained immunity, whereby innate immune cells retain a "memory" of earlier microbial encounters, thus influencing subsequent immune responses. Here, we propose that a robust multi-omics approach that integrates immunological, transcriptomic, and epigenomic datasets, combined with well-phenotyped and longitudinal food allergy cohorts, can inform the potential role of trained immunity in food allergy.
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http://dx.doi.org/10.1016/j.isci.2020.101171DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7262566PMC
June 2020

β-Glucan Induces Protective Trained Immunity against Mycobacterium tuberculosis Infection: A Key Role for IL-1.

Cell Rep 2020 05;31(7):107634

Department of Internal Medicine and Radboud Center for Infectious Diseases, Radboud University Medical Center, Nijmegen, the Netherlands; Department for Genomics & Immunoregulation, Life and Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany. Electronic address:

β-glucan is a potent inducer of epigenetic and functional reprogramming of innate immune cells, a process called "trained immunity," resulting in an enhanced host response against secondary infections. We investigate whether β-glucan exposure confers protection against pulmonary Mycobacterium tuberculosis (Mtb) infection. β-glucan induces trained immunity via histone modifications at gene promoters in human monocytes, which is accompanied by the enhanced production of proinflammatory cytokines upon secondary Mtb challenge and inhibition of Mtb growth. Mice treated with β-glucan are significantly protected against pulmonary Mtb infection, which is associated with the expansion of hematopoietic stem and progenitor cells in the bone marrow and increased myelopoiesis. The protective signature of β-glucan is mediated via IL-1 signaling, as β-glucan shows no protection in mice lacking a functional IL-1 receptor (IL1R). The administration of β-glucan may be used as a novel strategy in the treatment of mycobacterial infections and possibly as an adjuvant to improve anti-tuberculosis vaccines.
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http://dx.doi.org/10.1016/j.celrep.2020.107634DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7242907PMC
May 2020

Rewiring of glucose metabolism defines trained immunity induced by oxidized low-density lipoprotein.

J Mol Med (Berl) 2020 06 30;98(6):819-831. Epub 2020 Apr 30.

Department of Internal Medicine (463), Radboud University Medical Center, PO Box 9101, 6500 HB, Nijmegen, the Netherlands.

Stimulation of monocytes with microbial and non-microbial products, including oxidized low-density lipoprotein (oxLDL), induces a protracted pro-inflammatory, atherogenic phenotype sustained by metabolic and epigenetic reprogramming via a process called trained immunity. We investigated the intracellular metabolic mechanisms driving oxLDL-induced trained immunity in human primary monocytes and observed concomitant upregulation of glycolytic activity and oxygen consumption. In two separate cohorts of healthy volunteers, we assessed the impact of genetic variation in glycolytic genes on the training capacity of monocytes and found that variants mapped to glycolytic enzymes PFKFB3 and PFKP influenced trained immunity by oxLDL. Subsequent functional validation with inhibitors of glycolytic metabolism revealed dose-dependent inhibition of trained immunity in vitro. Furthermore, in vivo administration of the glucose metabolism modulator metformin abrogated the ability for human monocytes to mount a trained response to oxLDL. These findings underscore the importance of cellular metabolism for oxLDL-induced trained immunity and highlight potential immunomodulatory strategies for clinical management of atherosclerosis. KEY MESSAGES: Brief stimulation of monocytes to oxLDL induces a prolonged inflammatory phenotype. This is due to upregulation of glycolytic metabolism. Genetic variation in glycolytic genes modulates oxLDL-induced trained immunity. Pharmacological inhibition of glycolysis prevents trained immunity.
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http://dx.doi.org/10.1007/s00109-020-01915-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7297856PMC
June 2020

A High-Fat Diet Increases Influenza A Virus-Associated Cardiovascular Damage.

J Infect Dis 2020 08;222(5):820-831

School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia.

Background: Influenza A virus (IAV) causes a wide range of extrarespiratory complications. However, the role of host factors in these complications of influenza virus infection remains to be defined.

Methods: Here, we sought to use transcriptional profiling, virology, histology, and echocardiograms to investigate the role of a high-fat diet in IAV-associated cardiac damage.

Results: Transcriptional profiling showed that, compared to their low-fat counterparts (LF mice), mice fed a high-fat diet (HF mice) had impairments in inflammatory signaling in the lung and heart after IAV infection. This was associated with increased viral titers in the heart, increased left ventricular mass, and thickening of the left ventricular wall in IAV-infected HF mice compared to both IAV-infected LF mice and uninfected HF mice. Retrospective analysis of clinical data revealed that cardiac complications were more common in patients with excess weight, an association which was significant in 2 out of 4 studies.

Conclusions: Together, these data provide the first evidence that a high-fat diet may be a risk factor for the development of IAV-associated cardiovascular damage and emphasizes the need for further clinical research in this area.
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http://dx.doi.org/10.1093/infdis/jiaa159DOI Listing
August 2020

Maternal Obesity Alters Placental Cell Cycle Regulators in the First Trimester of Human Pregnancy: New Insights for BRCA1.

Int J Mol Sci 2020 Jan 11;21(2). Epub 2020 Jan 11.

Department of Obstetrics and Gynecology, Medical University of Graz, 8036 Graz, Austria.

In the first trimester of pregnancy, placental development involves a wide range of cellular processes. These include trophoblast proliferation, fusion, and differentiation, which are dependent on tight cell cycle control. The intrauterine environment affects placental development, which also includes the trophoblast cell cycle. In this work, we focus on maternal obesity to assess whether an altered intrauterine milieu modulates expression and protein levels of placental cell cycle regulators in early human pregnancy. For this purpose, we use first trimester placental tissue from lean and obese women (gestational week 5-11, = 58). Using a PCR panel, a cell cycle protein array, and STRING database analysis, we identify a network of cell cycle regulators increased by maternal obesity in which breast cancer 1 (BRCA1) is a central player. Immunostaining localizes BRCA1 predominantly to the villous and the extravillous cytotrophoblast. Obesity-driven BRCA1 upregulation is not able to be explained by DNA methylation (EPIC array) or by short-term treatment of chorionic villous explants at 2.5% oxygen with tumor necrosis factor α (TNF-α) (50 mg/mL), leptin (100 mg/mL), interleukin 6 (IL-6) (100 mg/mL), or high glucose (25 nM). Oxygen tension rises during the first trimester, but this change in vitro has no effect on BRCA1 (2.5% and 6.5% O). We conclude that maternal obesity affects placental cell cycle regulation and speculate this may alter placental development.
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http://dx.doi.org/10.3390/ijms21020468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7014057PMC
January 2020

The emerging role of epigenetics in the immune response to vaccination and infection: a systematic review.

Epigenetics 2020 Jun - Jul;15(6-7):555-593. Epub 2020 Mar 17.

Department of Paediatrics, The University of Melbourne , Parkville, Australia.

Extensive research has highlighted the role of infection-induced epigenetic events in the development of cancer. More recently, attention has focused on the ability of non-carcinogenic infections, as well as vaccines, to modify the human epigenome and modulate the immune response. This review explores this rapidly evolving area of investigation and outlines the many and varied ways in which vaccination and natural infection can influence the human epigenome from modulation of the innate and adaptive immune response, to biological ageing and modification of disease risk. The implications of these epigenetic changes on immune regulation and their potential application to the diagnosis and treatment of chronic infection and vaccine development are also discussed.
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http://dx.doi.org/10.1080/15592294.2020.1712814DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7574386PMC
March 2020

Inhibition of Histone Demethylases LSD1 and UTX Regulates ERα Signaling in Breast Cancer.

Cancers (Basel) 2019 Dec 16;11(12). Epub 2019 Dec 16.

Department of Precision Medicine, University of Campania "Luigi Vanvitelli", 80138 Naples, Italy.

In breast cancer, Lysine-specific demethylase-1 (LSD1) and other lysine demethylases (KDMs), such as Lysine-specific demethylase 6A also known as Ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX), are co-expressed and co-localize with estrogen receptors (ERs), suggesting the potential use of hybrid (epi)molecules to target histone methylation and therefore regulate/redirect hormone receptor signaling. Here, we report on the biological activity of a dual-KDM inhibitor (MC3324), obtained by coupling the chemical properties of tranylcypromine, a known LSD1 inhibitor, with the 2OG competitive moiety developed for JmjC inhibition. MC3324 displays unique features not exhibited by the single moieties and well-characterized mono-pharmacological inhibitors. Inhibiting LSD1 and UTX, MC3324 induces significant growth arrest and apoptosis in hormone-responsive breast cancer model accompanied by a robust increase in H3K4me2 and H3K27me3. MC3324 down-regulates ERα in breast cancer at both transcriptional and non-transcriptional levels, mimicking the action of a selective endocrine receptor disruptor. MC3324 alters the histone methylation of ERα-regulated promoters, thereby affecting the transcription of genes involved in cell surveillance, hormone response, and death. MC3324 reduces cell proliferation in ex vivo breast cancers, as well as in breast models with acquired resistance to endocrine therapies. Similarly, MC3324 displays tumor-selective potential in vivo, in both xenograft mice and chicken embryo models, with no toxicity and good oral efficacy. This epigenetic multi-target approach is effective and may overcome potential mechanism(s) of resistance in breast cancer.
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http://dx.doi.org/10.3390/cancers11122027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6966629PMC
December 2019

The role of Toll-like receptor 10 in modulation of trained immunity.

Immunology 2020 03 26;159(3):289-297. Epub 2019 Nov 26.

Department of Internal Medicine, Radboud Center for Infectious Diseases (RCI), Radboud Institute of Molecular Life Sciences (RIMLS), Radboud University Medical Center, Nijmegen, The Netherlands.

Toll-like receptor 10 (TLR10) is the only member of the human Toll-like receptor family with an inhibitory function on the induction of innate immune responses and inflammation. However, its role in the modulation of trained immunity (innate immune memory) is unknown. In the present study, we assessed whether TLR10 modulates the induction of trained immunity induced by β-glucan or bacillus Calmette-Guérin (BCG). Interleukin 10 receptor antagonist production was increased upon activation of TLR10 ex vivo after BCG vaccination, and TLR10 protein expression on monocytes was increased after BCG vaccination, whereas anti-TLR10 antibodies did not significantly modulate β-glucan or BCG-induced trained immunity in vitro. A known immunomodulatory TLR10 missense single-nucleotide polymorphism (rs11096957) influenced trained immunity responses by β-glucan or BCG in vitro. However, the in vivo induction of trained immunity by BCG vaccination was not influenced by TLR10 polymorphisms. In conclusion, TLR10 has a limited, non-essential impact on the induction of trained immunity in humans.
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http://dx.doi.org/10.1111/imm.13145DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7011636PMC
March 2020

Assisted reproductive technologies are associated with limited epigenetic variation at birth that largely resolves by adulthood.

Nat Commun 2019 09 2;10(1):3922. Epub 2019 Sep 2.

Murdoch Children's Research Institute, Parkville, VIC, 3010, Australia.

More than 7 million individuals have been conceived by Assisted Reproductive Technologies (ART) and there is clear evidence that ART is associated with a range of adverse early life outcomes, including rare imprinting disorders. The periconception period and early embryogenesis are associated with widespread epigenetic remodeling, which can be influenced by ART, with effects on the developmental trajectory in utero, and potentially on health throughout life. Here we profile genome-wide DNA methylation in blood collected in the newborn period and in adulthood (age 22-35 years) from a unique longitudinal cohort of ART-conceived individuals, previously shown to have no differences in health outcomes in early adulthood compared with non-ART-conceived individuals. We show evidence for specific ART-associated variation in methylation around birth, most of which occurred independently of embryo culturing. Importantly, ART-associated epigenetic variation at birth largely resolves by adulthood with no direct evidence that it impacts on development and health.
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http://dx.doi.org/10.1038/s41467-019-11929-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6718382PMC
September 2019

Intrauterine programming of obesity and type 2 diabetes.

Diabetologia 2019 10 27;62(10):1789-1801. Epub 2019 Aug 27.

Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road, Parkville, VIC, 3052, Australia.

The type 2 diabetes epidemic and one of its predisposing factors, obesity, are major influences on global health and economic burden. It is accepted that genetics and the current environment contribute to this epidemic; however, in the last two decades, both human and animal studies have consolidated considerable evidence supporting the 'developmental programming' of these conditions, specifically by the intrauterine environment. Here, we review the various in utero exposures that are linked to offspring obesity and diabetes in later life, including epidemiological insights gained from natural historical events, such as the Dutch Hunger Winter, the Chinese famine and the more recent Quebec Ice Storm. We also describe the effects of gestational exposure to endocrine disruptors, maternal infection and smoking to the fetus in relation to metabolic programming. Causal evidence from animal studies, motivated by human observations, is also discussed, as well as some of the proposed underlying molecular mechanisms for developmental programming of obesity and type 2 diabetes, including epigenetics (e.g. DNA methylation and histone modifications) and microRNA interactions. Finally, we examine the effects of non-pharmacological interventions, such as improving maternal dietary habits and/or increasing physical activity, on the offspring epigenome and metabolic outcomes.
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http://dx.doi.org/10.1007/s00125-019-4951-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6731191PMC
October 2019

Human placental methylome in the interplay of adverse placental health, environmental exposure, and pregnancy outcome.

PLoS Genet 2019 08 1;15(8):e1008236. Epub 2019 Aug 1.

Epigenetics Group, Murdoch Children's Research Institute, Royal Children's Hospital, Parkville, Victoria, Australia.

The placenta is the interface between maternal and fetal circulations, integrating maternal and fetal signals to selectively regulate nutrient, gas, and waste exchange, as well as secrete hormones. In turn, the placenta helps create the in utero environment and control fetal growth and development. The unique epigenetic profile of the human placenta likely reflects its early developmental separation from the fetus proper and its role in mediating maternal-fetal exchange that leaves it open to a range of exogenous exposures in the maternal circulation. In this review, we cover recent advances in DNA methylation in the context of placental function and development, as well as the interaction between the pregnancy and the environment.
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http://dx.doi.org/10.1371/journal.pgen.1008236DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6675049PMC
August 2019

Long-Lasting Transcriptional Changes in Circulating Monocytes of Acute Q Fever Patients.

Open Forum Infect Dis 2019 Jul;6(7)

Australian Rickettsial Reference Laboratory, University Hospital Geelong.

Objective: Although most patients recover from acute Q fever, around 20% develop Q fever fatigue syndrome (QFS), a debilitating fatigue syndrome that lasts at least 6 months. This study investigated transcriptional profiles of circulating monocytes and circulating cytokines as a subsequent mirror of myeloid cell function, 1 and 6 months after an acute Q fever infection.

Methods: Total RNA of circulating monocytes was collected from 11 acute Q fever patients and 15 healthy controls, matched for age (±5 years) and sex. Samples were collected at a median of 27 days (baseline, interquartile range, 15-35 days) after the infection and again 6 months thereafter. Transcriptome analysis was performed using RNA sequencing. Additionally, concentrations of circulating interleukin (IL)-10, IL-1β, IL-1Ra, and IL-6 were measured in serum.

Results: At baseline, acute Q fever patients clearly show a differential transcriptional program compared with healthy controls. This is still the case at follow-up, albeit to a lesser extent. At baseline, a significant difference in levels of circulating IL-10 (P = .0019), IL-1β (P = .0067), IL-1Ra (P = .0008), and IL-6 (P = .0003) was seen. At follow-up, this difference had decreased for IL-10 (P = .0136) and IL-1Ra (P = .0017) and had become nonsignificant for IL-1β (P = .1139) and IL-6 (P = .2792).

Conclusions: We show that an acute Q fever infection has a long-term effect on the transcriptional program of circulating monocytes and, therefore, likely their myeloid progenitor cells, as well as concentrations of circulating IL-10, IL-1β, IL-1Ra, and IL-6.
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http://dx.doi.org/10.1093/ofid/ofz296DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6667718PMC
July 2019

Early-life determinants of hypoxia-inducible factor 3A gene (HIF3A) methylation: a birth cohort study.

Clin Epigenetics 2019 07 1;11(1):96. Epub 2019 Jul 1.

Murdoch Children's Research Institute, Parkville, Australia.

Background: Methylation of the hypoxia-inducible factor 3α gene (HIF3A) has been linked to pregnancy exposures, infant adiposity and later BMI. Genetic variation influences HIF3A methylation levels and may modify these relationships. However, data in very early life are limited, particularly in association with adverse pregnancy outcomes. We investigated the relationship between maternal and gestational factors, infant anthropometry, genetic variation and HIF3A DNA methylation in the Barwon Infant Study, a population-based birth cohort. Methylation of two previously studied regions of HIF3A were tested in the cord blood mononuclear cells of 938 infants.

Results: No compelling evidence was found of an association between birth weight, adiposity or maternal gestational diabetes with methylation at the most widely studied HIF3A region. Male sex (- 4.3%, p < 0.001) and pre-eclampsia (- 5.4%, p = 0.02) negatively associated with methylation at a second region of HIF3A; while positive associations were identified for gestational diabetes (4.8%, p = 0.01) and gestational age (1.2% increase per week, p < 0.001). HIF3A genetic variation also associated strongly with methylation at this region (p < 0.001).

Conclusions: Pre- and perinatal factors impact HIF3A methylation, including pre-eclampsia. This provides evidence that specific pregnancy complications, previously linked to adverse outcomes for both mother and child, impact the infant epigenome in a molecular pathway critical to several vascular and metabolic conditions. Further work is required to understand the mechanisms and clinical relevance, particularly the differing effects of in utero exposure to gestational diabetes or pre-eclampsia.
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http://dx.doi.org/10.1186/s13148-019-0687-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6604333PMC
July 2019

Treatment with Statins Does Not Revert Trained Immunity in Patients with Familial Hypercholesterolemia.

Cell Metab 2019 07 13;30(1):1-2. Epub 2019 Jun 13.

Department of Internal Medicine, and Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Center, Nijmegen 6525GA, the Netherlands. Electronic address:

Individuals with elevated LDL-cholesterol levels have an increased risk for cardiovascular disease. Despite lipid lowering strategies, however, a significant cardiovascular risk remains. Bekkering et al. show that monocytes from patients with familial hypercholesterolemia have a trained immunity phenotype and that lipid lowering with statins does not revert this pro-inflammatory phenotype.
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http://dx.doi.org/10.1016/j.cmet.2019.05.014DOI Listing
July 2019

Diabetes in pregnancy and epigenetic mechanisms-how the first 9 months from conception might affect the child's epigenome and later risk of disease.

Lancet Diabetes Endocrinol 2019 10 22;7(10):796-806. Epub 2019 May 22.

Cancer and Disease Epigenetics, Murdoch Children's Research Institute, Melbourne, VIC, Australia; Department of Pediatrics, Melbourne University, Melbourne, VIC, Australia.

Diabetes in pregnancy is not only associated with increased risk of pregnancy complications and subsequent maternal metabolic disease, but also increases the risk of long-term metabolic disease in the offspring. At the interface between genetic and environmental factors, epigenetic variation established in utero represents a plausible link between the in utero environment and later disease susceptibility. The identification of an epigenetic fingerprint of diabetes in pregnancy linked to the metabolic health of the offspring might provide novel biomarkers for the identification of offspring most at risk, before the onset of metabolic dysfunction, for targeted monitoring and intervention. In this Personal View, we (1) highlight the scale of the problem of diabetes in pregnancy, (2) summarise evidence for the variation in offspring epigenetic profiles following exposure to diabetes in utero, and (3) outline potential future approaches to further understand the mechanisms by which exposure to maternal metabolic dysfunction in pregnancy is transmitted through generations.
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http://dx.doi.org/10.1016/S2213-8587(19)30078-6DOI Listing
October 2019

A possible role for mitochondrial-derived peptides humanin and MOTS-c in patients with Q fever fatigue syndrome and chronic fatigue syndrome.

J Transl Med 2019 05 14;17(1):157. Epub 2019 May 14.

Radboud Expertise Center for Q Fever, Department of Internal Medicine, Division of Infectious Diseases 463, Radboud University Medical Center, P.O. Box 9101, 6500 HB, Nijmegen, The Netherlands.

Background: Q fever fatigue syndrome (QFS) is a well-documented state of prolonged fatigue following around 20% of acute Q fever infections. It has been hypothesized that low grade inflammation plays a role in its aetiology. In this study, we aimed to identify transcriptome profiles that could aid to better understand the pathophysiology of QFS.

Methods: RNA of monocytes was collected from QFS patients (n = 10), chronic fatigue syndrome patients (CFS, n = 10), Q fever seropositive controls (n = 10), and healthy controls (n = 10) who were age- (± 5 years) and sex-matched. Transcriptome analysis was performed using RNA sequencing.

Results: Mitochondrial-derived peptide (MDP)-coding genes MT-RNR2 (humanin) and MT-RNR1 (MOTS-c) were differentially expressed when comparing QFS (- 4.8 log2-fold-change P = 2.19 × 10 and - 4.9 log2-fold-change P = 4.69 × 10), CFS (- 5.2 log2-fold-change, P = 3.49 × 10 - 4.4 log2-fold-change, P = 2.71 × 10), and Q fever seropositive control (- 3.7 log2-fold-change P = 1.78 × 10 and - 3.2 log2-fold-change P = 1.12 × 10) groups with healthy controls, resulting in a decreased median production of humanin in QFS patients (371 pg/mL; Interquartile range, IQR, 325-384), CFS patients (364 pg/mL; IQR 316-387), and asymptomatic Q fever seropositive controls (354 pg/mL; 292-393).

Conclusions: Expression of MDP-coding genes MT-RNR1 (MOTS-c) and MT-RNR2 (humanin) is decreased in CFS, QFS, and, to a lesser extent, in Q fever seropositive controls, resulting in a decreased production of humanin. These novel peptides might indeed be important in the pathophysiology of both QFS and CFS.
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http://dx.doi.org/10.1186/s12967-019-1906-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6518812PMC
May 2019

Alkaline soda Lake Velika Rusanda (Serbia): the first insight into diatom diversity of this extreme saline lake.

Extremophiles 2019 May 19;23(3):347-357. Epub 2019 Mar 19.

University of Belgrade, Institute of Chemistry, Technology and Metallurgy, Njegoševa 12, Belgrade, 11000, Serbia.

Alkaline soda lakes are unique habitats found in specific geographic regions, usually with dry climate. The Carpathian Basin is one of those regions very important for habitat and biodiversity conservation in Europe, with natural soda lakes found in Austria, Hungary and Serbia. In comparison to other two countries from Central Europe, algal biodiversity studies of saline soda lakes in Serbia are scarce. Lake Velika Rusanda has the highest measured salinity of all saline lakes in the Carpathian Basin and there were no reports of its diatom species richness and diversity till now. We conducted 2-year investigation programme to study biodiversity and seasonal dynamics of diatoms in this lake. A total of 27 diatom taxa were found, almost all of them attached to reed and much less in benthos and plankton. Five new diatom species for Serbia were recorded, Craticula halopannonica, Navicymbula pusilla, Hantzschia weyprechtii, Nitzschia thermaloides and Navicula staffordiae. The last mentioned is new for Europe as well. Lake Velika Rusanda is inhabited mostly by alkaliphilous and halophilic diatoms. Since diatoms are used as bioindicators in soda lakes, our results will improve their further application in ecological status assessment of these fragile habitats in the Carpathian Basin.
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http://dx.doi.org/10.1007/s00792-019-01088-6DOI Listing
May 2019

Extensive epigenomic integration of the glucocorticoid response in primary human monocytes and in vitro derived macrophages.

Sci Rep 2019 02 26;9(1):2772. Epub 2019 Feb 26.

Department of Molecular Biology, Radboud Institute for Molecular Life Sciences, Faculty of Science Radboud University, PO box 9101, 6500 HG, Nijmegen, The Netherlands.

Glucocorticoid receptor is a transcription factor that is ubiquitously expressed. Glucocorticoids are circadian steroids that regulate a wide range of bodily functions, including immunity. Here we report that synthetic glucocorticoids affect 1035 mRNAs in isolated healthy human blood monocytes but only 165 in the respective six day-old monocyte-derived macrophages. The majority of the glucocorticoid response in monocytes concerns genes that are dynamic upon monocyte to macrophage differentiation, whereby macrophage-like mRNA levels are often reached in monocytes within four hours of treatment. Concomitantly, over 5000 chromosomal H3K27ac regions undergo remodelling, of which 60% involve increased H3K27ac signal. We find that chromosomal glucocorticoid receptor binding sites correlate with positive but not with negative local epigenomic effects. To investigate further we assigned our data to topologically associating domains (TADs). This shows that about 10% of macrophage TADs harbour at least one GR binding site and that half of all the glucocorticoid-induced H3K27ac regions are confined to these TADs. Our analyses are therefore consistent with the notion that TADs naturally accommodate information from sets of distal glucocorticoid response elements.
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http://dx.doi.org/10.1038/s41598-019-39395-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391480PMC
February 2019

Micromanaging human placental function: differential microRNA expression in feto-placental endothelial cells of gestational diabetes pregnancies.

Clin Sci (Lond) 2019 01 25;133(2):315-319. Epub 2019 Jan 25.

Epigenetics Research, Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road, Parkville, VIC, 3052, Australia and Department of Paediatrics, University of Melbourne, Parkville, VIC, 3052, Australia

Healthy development in utero relies on the appropriate exchange of nutrients and other signalling between the maternal and fetal circulations. Disruption to this fine balance is associated with several pregnancy and adverse birth outcomes, including gestational diabetes mellitus (GDM). This is a complex condition influenced by genetic, environment and potentially epigenetic factors in association with a range of altered developmental outcomes. A recent study, published in Clinical Science, explores miRNAs as a molecular mechanism underpinning the altered function of placental endothelial cells in GDM pregnancies.
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http://dx.doi.org/10.1042/CS20180901DOI Listing
January 2019

The Itaconate Pathway Is a Central Regulatory Node Linking Innate Immune Tolerance and Trained Immunity.

Cell Metab 2019 01 4;29(1):211-220.e5. Epub 2018 Oct 4.

Department of Internal Medicine (463) and Radboud Center for Infectious Diseases (RCI), Radboud University Nijmegen Medical Centre, Geert Grooteplein 8, Nijmegen 6500 HB, the Netherlands; Department for Genomics & Immunoregulation, Life and Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany; Human Genomics Laboratory, Craiova University of Medicine and Pharmacy, Craiova, Romania.

Sepsis involves simultaneous hyperactivation of the immune system and immune paralysis, leading to both organ dysfunction and increased susceptibility to secondary infections. Acute activation of myeloid cells induced itaconate synthesis, which subsequently mediated innate immune tolerance in human monocytes. In contrast, induction of trained immunity by β-glucan counteracted tolerance induced in a model of human endotoxemia by inhibiting the expression of immune-responsive gene 1 (IRG1), the enzyme that controls itaconate synthesis. β-Glucan also increased the expression of succinate dehydrogenase (SDH), contributing to the integrity of the TCA cycle and leading to an enhanced innate immune response after secondary stimulation. The role of itaconate was further validated by IRG1 and SDH polymorphisms that modulate induction of tolerance and trained immunity in human monocytes. These data demonstrate the importance of the IRG1-itaconate-SDH axis in the development of immune tolerance and training and highlight the potential of β-glucan-induced trained immunity to revert immunoparalysis.
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http://dx.doi.org/10.1016/j.cmet.2018.09.003DOI Listing
January 2019