Heterogeneous accumulation of senescent cells expressing the senescence-associated secretory phenotype (SASP) affects tissue homeostasis which leads to diseases, such as osteoarthritis (OA). In... Show moreHeterogeneous accumulation of senescent cells expressing the senescence-associated secretory phenotype (SASP) affects tissue homeostasis which leads to diseases, such as osteoarthritis (OA). In this study, we set out to characterize heterogeneity of cellular senescence within aged articular cartilage and explored the presence of corresponding metabolic profiles in blood that could function as representative biomarkers. Hereto, we set out to perform cluster analyses, using a gene-set of 131 senescence genes (N = 57) in a previously established RNA sequencing dataset of aged articular cartilage and a generated metabolic dataset in overlapping blood samples. Using unsupervised hierarchical clustering and pathway analysis, we identified two robust cellular senescent endotypes. Endotype-1 was enriched for cell proliferating pathways, expressing forkhead box protein O4 (FOXO4), RB transcriptional corepressor like 2 (RBL2), and cyclin-dependent kinase inhibitor 1B (CDKN1B); the FOXO mediated cell cycle was identified as possible target for endotype-1 patients. Endotype-2 showed enriched inflammation-associated pathways, expressed by interleukin 6 (IL6), matrix metallopeptidase (MMP)1/3, and vascular endothelial growth factor (VEGF)C and SASP pathways were identified as possible targets for endotype-2 patients. Notably, plasma-based metabolic profiles in overlapping blood samples (N = 21) showed two corresponding metabolic clusters in blood. These non-invasive metabolic profiles could function as biomarkers for patient-tailored targeting of senescence in OA. Show less
Background: Non-coding genetic variants that influence gene transcription in pancreatic islets play a major role in the susceptibility to type 2 diabetes (T2D), and likely also contribute to type 1... Show moreBackground: Non-coding genetic variants that influence gene transcription in pancreatic islets play a major role in the susceptibility to type 2 diabetes (T2D), and likely also contribute to type 1 diabetes (T1D) risk. For many loci, however, the mechanisms through which non-coding variants influence diabetes susceptibility are unknown. Results: We examine splicing QTLs (sQTLs) in pancreatic islets from 399 human donors and observe that common genetic variation has a widespread influence on the splicing of genes with established roles in islet biology and diabetes. In parallel, we profile expression QTLs (eQTLs) and use transcriptome-wide association as well as genetic co-localization studies to assign islet sQTLs or eQTLs to T2D and T1D susceptibility signals, many of which lack candidate effector genes. This analysis reveals biologically plausible mechanisms, including the association of T2D with an sQTL that creates a nonsense isoform in ERO1B, a regulator of ER-stress and proinsulin biosynthesis. The expanded list of T2D risk effector genes reveals overrepresented pathways, including regulators of G-protein-mediated cAMP production. The analysis of sQTLs also reveals candidate effector genes for T1D susceptibility such as DCLRE1B, a senescence regulator, and lncRNA MEG3. Conclusions: These data expose widespread effects of common genetic variants on RNA splicing in pancreatic islets. The results support a role for splicing variation in diabetes susceptibility, and offer a new set of genetic targets with potential therapeutic benefit. Show less
Netrin-4, recognized in neural and vascular development, is highly expressed by mature endothelial cells. The function of this netrin-4 in vascular biology after development has remained unclear.... Show moreNetrin-4, recognized in neural and vascular development, is highly expressed by mature endothelial cells. The function of this netrin-4 in vascular biology after development has remained unclear. We found that the expression of netrin-4 is highly regulated in endothelial cells and is important for quiescent healthy endothelium. Netrin-4 expression is upregulated in endothelial cells cultured under laminar flow conditions, while endothelial cells stimulated with tumor necrosis factor alpha resulted in decreased netrin-4 expression. Targeted reduction of netrin-4 in endothelial cells resulted in increased expression of vascular cell adhesion molecule 1 and intercellular adhesion molecule 1. Besides, these endothelial cells were more prone to monocyte adhesion and showed impaired barrier function, measured with electric cell-substrate impedance sensing, as well as in an ?organ-on-achip? microfluidic system. Importantly, endothelial cells with reduced levels of netrin-4 showed increased expression of the senescence-associated markers cyclin-dependent kinase inhibitor-1 and -2A, an increased cell size and decreased ability to proliferate. Consistent with the gene expression profile, netrin-4 reduction was accompanied with more senescent associated ?-galactosidase activity, which could be rescued by adding netrin-4 protein. Finally, using human decellularized kidney extracellular matrix scaffolds, we found that pre-treatment of the scaffolds with netrin-4 increased numbers of endothelial cells adhering to the matrix, showing a pro-survival effect of netrin-4. Taken together, netrin-4 acts as an anti-senescence and anti-inflammation factor in endothelial cell function and our results provide insights as to maintain endothelial homeostasis and supporting vascular health. Show less
Mohamed, S.A.; Grewal, N.; Gittenberger-de Groot, A.C. 2018
Cell culture models play an important role in biomedical research and will continue to do so given the growing opposition to vivisection and the limited predictive value of animal models for... Show moreCell culture models play an important role in biomedical research and will continue to do so given the growing opposition to vivisection and the limited predictive value of animal models for human disease. Moreover, cell culture models can be easily established to mimic physiological or pathological processes, which is difficult to accomplish using in silico models. While non-cellular in vitro models are highly suitable for studying simple biochemical processes, cell culture models recapitulate many of the complex regulatory circuits governing protein activity in vivo and hence allow investigation of diverse physiological processes. Also, cell culture models offer the possibility to address fundamental research questions in a much more simplified, specific and controllable manner than can be achieved using in vivo models. Show less
Giacconi, R.; Costarelli, L.; Piacenza, F.; Basso, A.; Burkle, A.; Moreno-Villanueva, M.; ... ; Malavolta, M. 2018
The development of methods for the genetic modification of plants a few decades ago has provided a tremendous boost for molecular plant science. Crop plants have been generated that are... Show moreThe development of methods for the genetic modification of plants a few decades ago has provided a tremendous boost for molecular plant science. Crop plants have been generated that are resistant to insects or herbicides, or that produce useful sugars or healthy nutrients. Although the ban on growing GM crops in Europe has considerably limited the application of GM technologies, they have still contributed considerably to fundamental plant science. Especially by using the natural and very efficient mechanism of DNA transfer by the soil born bacterium Agrobacterium tumefaciens, many collections of mutant lines of model plant species such as Arabidopsis and rice have been generated, in which genes are disrupted or overexpressed by the insertion of an Agrobacterium transfer DNA (T-DNA) construct. These collections have been used in forward or reverse genetics studies to unravel the function of a gene or a family of genes in plant defense or development, and to identify the key regulators in these processes. The study described in this thesis focused on the use of one of these key regulators, the Arabidopsis AT-HOOK MOTIF NUCLEAR LOCALIZED PROTEIN 15/REJUVENATOR (AHL15/RJV), to alter developmental processes such as flowering, senescence and regeneration. Show less
In this thesis, senescence is measured in human populations according to its definition of an increase in the risks of dysfunction, disease, and death with chronological age. Part I of this thesis... Show moreIn this thesis, senescence is measured in human populations according to its definition of an increase in the risks of dysfunction, disease, and death with chronological age. Part I of this thesis investigates how a population__s senescence rate can be measured through the increase in mortality rate with age. Part II of this thesis investigates how senescence can be measured through the increase in morbidity - with a focus on cardiovascular disease - in a non-western population and thus be compared with the senescence process in western populations. Show less
The aim of this thesis was to investigate the radiological phenotype of the human brain in familial longevity with regard to brain structure. This study was performed as part of the Leiden... Show moreThe aim of this thesis was to investigate the radiological phenotype of the human brain in familial longevity with regard to brain structure. This study was performed as part of the Leiden Longevity Study __ a study population consisting of offspring of long-lived Dutch people who are genetically predisposed to become long-lived as well and their environmentally and age-matched spouses. Dedicated MR imaging techniques were applied to study the phenotype of the human brain on the macrostructural as well as microstructural level. Both features which are most likely caused by early development and a lower susceptibility to known age-related structural brain changes on the macro- as well as microstructural level are hallmarks of the brain phenotype in longevity. The contributing mechanisms have yet to be identified but may involve homeostatic control and body function. Keeping in mind that there is a gradual transition from healthy brain aging to what is generally accepted as __normal__ brain aging and pathological brain changes as disease correlates, our study results implicate that the brains of offspring of nonagenarian siblings are less susceptible to neurodegenerative brain changes such as (vascular) dementia, Alzheimer__s disease and cerebrovascular disease. Our findings should be considered as starting points for future studies on the functional implications of the presented results and studies on the underlying pathways of brain structure preservation in human longevity. Show less
The evolution of ageing is a field flush with misconceptions, misunderstandings, and hiatuses. In this thesis I address the most important misunderstanding and misconceptions, and develop new... Show moreThe evolution of ageing is a field flush with misconceptions, misunderstandings, and hiatuses. In this thesis I address the most important misunderstanding and misconceptions, and develop new theory to fill the gaps. This work directly leads to the restatement of the central question in the evolutionary theory of ageing. Rather than evaluating evolutionary forces in models that are at best weakly rooted in (patho-) physiological mechanisms, usually phrased in terms of __age-specific genes__ that are not further specified, as is the current practice, the most pressing question becomes why an organism cannot, or does not, do in itself what it is perfectly capable of doing outside itself in the form of reproduction, namely producing a perfectly healthy __young__ organism. Evolutionary forces cannot answer this question. If anything, this is a mechanistic question. I suggest investigating __the evolution of unretainability__: why and how has our form of life evolved, in which it is mechanistically impossible to bring ageing to a halt, and what are the responsible mechanistic constraints? Show less
Koopman, Jacob J.E.; Rozing, Maarten P.; Kramer, Anneke; Abad, José M.; Finne, Patrik; Heaf, James G.; ... ; Westendorp, Rudi G.J. 2015
The rate of senescence can be inferred from the acceleration by which mortality rates increase over age. Such a senescence rate is generally estimated from parameters of a mathematical model fitted... Show moreThe rate of senescence can be inferred from the acceleration by which mortality rates increase over age. Such a senescence rate is generally estimated from parameters of a mathematical model fitted to these mortality rates. However, such models have limitations and underlying assumptions. Notably, they do not fit mortality rates at young and old ages. Therefore, we developed a method to calculate senescence rates from the acceleration of mortality directly without modeling the mortality rates. We applied the different methods to age group–specific mortality data from the European Renal Association—European Dialysis and Transplant Association Registry, including patients with end-stage renal disease on dialysis, who are known to suffer from increased senescence rates (n = 302,455), and patients with a functioning kidney transplant (n = 74,490). From age 20 to 70, senescence rates were comparable when calculated with or without a model. However, when using non-modeled mortality rates, senescence rates were yielded at young and old ages that remained concealed when using modeled mortality rates. At young ages senescence rates were negative, while senescence rates declined at old ages. In conclusion, the rate of senescence can be calculated directly from non-modeled mortality rates, overcoming the disadvantages of an indirect estimation based on modeled mortality rates. Show less
Waaijer, M.E.C.; Wieser, M.; Grillari-Voglauer, R.; Heemst, D. van; Grillari, J.; Maier, A.B. 2014
With the ever-increasing life expectancies of people, the prevalence of aging-related diseases is also increasing. We studied a unique cohort of people; offspring of nonagenarian siblings with the... Show moreWith the ever-increasing life expectancies of people, the prevalence of aging-related diseases is also increasing. We studied a unique cohort of people; offspring of nonagenarian siblings with the propensity for longevity were also recruited and compared to their partners, representing the general population. The offspring (~60 year old) already show lower prevalence of cardiovascular disease and diabetes. In the thesis __Cellular stress in vitro and longevity on vivo__ it is reported how cells (dermal fibroblasts) derived from these offspring were compared with cells from their partners. It was shown that they react differenly in vitro under both non stressed and under conditions of oxidative stress. Future research is warranted to further elucidate the genes involved in these differences. Show less
Replicative ageing of fibroblasts has often been used as a model for organismal ageing. The general assumption that the ageing process is mirrored by cellular senescence in vitro is based on lower... Show moreReplicative ageing of fibroblasts has often been used as a model for organismal ageing. The general assumption that the ageing process is mirrored by cellular senescence in vitro is based on lower replicative capacity of human fibroblasts from donors of higher chronological age, but these inverse relations have not been reported unequivocally. The relation between chronological age and fibroblast growth characteristics was assessed in nonagenarian subjects of the Leiden 85+ Study. A high remaining replicative capacity impressively showed that even in the very elderly a crucial number of cells with high mitotic capacity are left to give rise to fibroblast strains with the capacity for more than 100 population doublings. During the course of fibroblast growth in vitro, beta-galactosidase activity has been shown to be a reliable biomarker for replicative senescence. In myoblast cultures the relation between mixed cultures and clonal cultures was studied, showing marked heterogeneity between clonal cultures that all had a significantly lower replicative capacity when compared to mixed cultures, indicating heterogeneity of cells within one tissue compartment in their in vivo history. In a formal review on the replicative capacity of fibroblasts from patients suffering from accelerated ageing syndromes, age related diseases and donor age it was found that except for premature ageing syndromes, the replicative capacity of fibroblasts in vitro does not mirror key characteristics of human life histories. Show less
