567 research outputs found
Morpholino Analogues of Fingolimod as Novel and Selective S1P1 Ligands with In Vivo Efficacy in a Mouse Model of Experimental Antigen-Induced Encephalomyelitis.
No full textMultiple sclerosis (MS) is a chronic, inflammatory, autoimmune disease of the central nervous system (CNS) which is associated with lower life expectancy and disability. The experimental antigen-induced encephalomyelitis (EAE) in mice is a useful animal model of MS, which allows exploring the etiopathogenetic mechanisms and testing novel potential therapeutic drugs. A new therapeutic paradigm for the treatment of MS was introduced in 2010 through the sphingosine 1-phosphate (S1P) analogue fingolimod (FTY720, Gilenya®), which acts as a functional S1P1 antagonist on T lymphocytes to deplete these cells from the blood. In this study, we synthesized two novel structures, ST-1893 and ST-1894, which are derived from fingolimod and chemically feature a morpholine ring in the polar head group. These compounds showed a selective S1P1 activation profile and a sustained S1P1 internalization in cultures of S1P1-overexpressing Chinese hamster ovary (CHO)-K1 cells, consistent with a functional antagonism. In vivo, both compounds induced a profound lymphopenia in mice. Finally, these substances showed efficacy in the EAE model, where they reduced clinical symptoms of the disease, and, on the molecular level, they reduced the T-cell infiltration and several inflammatory mediators in the brain and spinal cord. In summary, these data suggest that S1P1-selective compounds may have an advantage over fingolimod and siponimod, not only in MS but also in other autoimmune diseases
Novel compounds with dual S1P receptor agonist and histamine H3 receptor antagonist activities act protective in a mouse model of multiple sclerosis.
No full textThe sphingosine 1-phosphate (S1P) receptor 1 (S1P1) has emerged as a therapeutic target for the treatment of multiple sclerosis (MS). Fingolimod (FTY720) is the first functional antagonist of S1P1 that has been approved for oral treatment of MS. Previously, we have developed novel butterfly derivatives of FTY720 that acted similar to FTY720 in reducing disease symptoms in a mouse model of experimental autoimmune encephalomyelitis (EAE). In this study, we have synthesized a piperidine derivative of the oxazolo-oxazole compounds, denoted ST-1505, and its ring-opened analogue ST-1478, and characterised their in-vitro and in-vivo functions. Notably, the 3-piperidinopropyloxy moiety resembles a structural motif of pitolisant, a drug with histamine H3R antagonistic/inverse agonist activity approved for the treatment of narcolepsy. Both novel compounds exerted H3R affinities, and in addition, ST-1505 was characterised as a dual S1P1+3 agonist, whereas ST-1478 was a dual S1P1+5 agonist. Both multitargeting compounds were also active in mice and reduced the lymphocyte numbers as well as diminished disease symptoms in the mouse model of MS. The effect of ST-1478 was dependent on SK-2 activity suggesting that it is a prodrug like FTY720, but with a more selective S1P receptor activation profile, whereas ST-1505 is a fully active drug even in the absence of SK-2. In summary, these data suggest that the well soluble piperidine derivatives ST-1505 and ST-1478 hold promise as novel drugs for the treatment of MS and other autoimmune or inflammatory diseases, and by their H3R antagonist potency, they might additionally improve cognitive impairment during disease
Sphingosine-1-phosphate and macrophage biology—how the sphinx tames the big eater
No full textThe sphingolipid sphingosine-1-phosphate (S1P) is produced by sphingosine kinases to either signal through intracellular targets or to activate a family of specific G-protein-coupled receptors (S1PR). S1P levels are usually low in peripheral tissues compared to the vasculature, forming a gradient that mediates lymphocyte trafficking. However, S1P levels rise during inflammation in peripheral tissues, thereby affecting resident or recruited immune cells, including macrophages. As macrophages orchestrate initiation and resolution of inflammation, the sphingosine kinase/S1P/S1P-receptor axis emerges as an important determinant of macrophage function in the pathogenesis of inflammatory diseases such as cancer, atherosclerosis, and infection. In this review, we therefore summarize the current knowledge how S1P affects macrophage biology
Effective inhibition of acid and neutral ceramidases by novel B-13 and LCL-464 analogues
No full textInduction of apoptosis mediated by the inhibition of ceramidases has been shown to enhance the efficacy of conventional chemotherapy in several cancer models. Among the inhibitors of ceramidases reported in the literature, B-13 is considered as a lead compound having good in vitro potency towards acid ceramidase. Furthermore, owing to the poor activity of B-13 on lysosoamal acid ceramidase in living cells, LCL-464 a modified derivative of B-13 containing a basic omega-amino group at the fatty acid was reported to have higher potency towards lysosomal acid ceramidase in living cells. In a search for more potent inhibitors of ceramidases, we have designed a series of compounds with structural modifications of B-13 and LCL-464. In this study, we show that the efficacy of B-13 in vitro as well as in intact cells can be enhanced by suitable modification of functional groups. Furthermore, a detailed SAR investigation on LCL-464 analogues revealed novel promising inhibitors of aCDase and nCDase. In cell culture studies using the breast cancer cell line MDA-MB-231, some of the newly developed compounds elevated endogenous ceramide levels and in parallel, also induced apoptotic cell death. In summary, this study shows that structural modification of the known ceramidase inhibitors B-13 and LCL-464 generates more potent ceramidase inhibitors that are active in intact cells and not only elevates the cellular ceramide levels, but also enhances cell death
The WNK1-SPAK/OSR1 signaling pathway in pneumonia-induced lung barrier failure
No full textObjective Despite the availability of effective antibiotics and pneumococcal vaccines, lung
barrier failure remains a major complication of pneumonia, with high mortality and morbidity
rates. Loss of function of the endothelial cystic fibrosis transmembrane conductance regulator
(CFTR) has been associated with lung barrier failure in pneumonia. Additionally,
pharmacological restoration of CFTR function with ivacaftor has shown barrier-protective
properties. The regulatory cascade of CFTR includes lysine-deficient protein kinase 1 (WNK1),
which has been suggested as a possible mediator of barrier breakdown. Yet, the mechanisms
by which CFTR contributes to lung barrier integrity and its functional interaction with WNK1
remain incompletely understood. To address this, this study has three main objectives: (i) to
test whether in vivo WNK1 activation could mimic the effect of ivacaftor-mediated CFTR
potentiation; (ii) to investigate the role of WNK1 and its downstream target kinases,
SPS/Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress-responsive kinase
1 (OSR1), in pneumonia-induced lung barrier disruption, and (iii) to determine whether the
regulation of WNK1-SPAK/OSR1 is cell-type specific and differs between airway epithelial and
endothelial cells.
Methods A murine model (C57BL/6J wild-type) of severe pneumococcal pneumonia was used
to study WNK1 activation and its effects on vascular leakage and inflammatory response.
Furthermore, a heterozygous knockout of WNK1 (Wnk1+/-) and a homozygous knock-in of
SPAK (SpakL502A/L502A) were used to investigate the functional role of the kinases in the murine
infection model. To further characterize cell-type specific regulation of WNK1-SPAK/OSR1
signaling, human primary pulmonary microvascular endothelial cells (HPMEC) and primary
human pulmonary alveolar epithelial cells (HPAEpiC) were used.
Transepithelial/transendothelial electrical resistance (TEER) was used to measure the cellular
integrity after pharmacological inhibition of WNK1 or SPAK. In addition, the effect of WNK1
inhibition (WNK-In-11) and nonspecific WNK1 activation (temozolomide) in uninfected or S.
pneumoniae-infected conditions and their consequence on cell-specific regulation were
investigated by proteomic and phosphoproteomic profiling.
Results Pharmacological WNK1 activation prior to S. pneumoniae infection partly prevented
severe barrier disruption, improved body temperature, and reduced bacterial burden. Genetic
deficiency of WNK1 and SPAK in mice, however, did not affect lung permeability distinctly.
Whereas, under in vitro conditions, inhibition of WNK1 decreased cellular resistance HPMEC but not in HPAEpiC. Notably, SPAK inhibition decreased resistance in both cell types. Although
no obvious changes in the global proteome were observed, the phosphoproteomic profiling
revealed that inhibitory WNK1 phosphorylation is increased in HPMEC but not in HPAEpiC
after targeted WNK1 inhibition and S. pneumoniae infection.
Conclusion The results suggest an integral role for WNK1-SPAK/OSR1 signaling in lung
barrier function. Activation of WNK1 in vivo was shown to have beneficial effects on barrier
stability. Furthermore, the signaling was shown to have distinct and cell-type-specific
characteristics. In addition to WNK1 and SPAK/OSR1, the regulation of pulmonary
permeability is likely to depend on other kinases, including other members of the WNK family
Author Co-Citation Analysis (ACA): a powerful tool for representing implicit knowledge of scholar knowledge workers
No full textIn the last decade, knowledge has emerged as one of the most important and valuable organizational assets. Gradually this importance caused to emergence of new discipline entitled ―knowledge management‖. However one of the major challenges of knowledge management is conversion implicit or tacit knowledge to explicit knowledge. Thus Making knowledge visible so that it can be better accessed, discussed, valued or generally managed is a long-standing objective in knowledge management. Accordingly in this paper author co- citation analysis (ACA) will be proposed as an efficient technique of knowledge visualization in academia (Scholar knowledge workers)
Label-free, quantitative detection of drugs and nanocarriers in cells and skin
No full textIn dieser Arbeit wurden Wirkstoffe mithilfe von labelfreien Methoden ortsaufgelöst in biologischen Proben detektiert. Die Detektion erfolgte durch eine Simulation der gemessenen Spektren hyperspektraler Messungen im spektralen Bereich an der O 1s Kante (520-565 eV) und im mittleren Infrarotbereich (1190-1320 cm^-1 und 1600-1850 cm^-1) von behandelten Proben mithilfe einer numerischen Singulärwertzerlegung auf Basis von einzeln gemessenen Reinsubstanzspektren und Spektren der unbehandelten biologischen Spezies.
Der erste Teil der Arbeit beinhaltet die labelfreie Detektion des Wirkstoffs Rapamycin und der redoxsensitiven Nanocarrier ex vivo topisch behandelter Humanhaut. Hierbei wurden in Epoxidharz eingebettete Proben mithilfe von Röntgenmikroskopie an der O 1s-Kante untersucht. Weitere Proben wurden mithilfe von photothermischer Expansionsspektroskopie (AFM-IR) untersucht. Es kann gezeigt werden, dass eine Vorbehandlung mit der Serinprotease Trypsin die Membranen der Corneocyten öffnet und der Wirkstoff intrazellulär in das Stratum Corneum transportiert werden kann. Der Wirkstoffnachweis erfolgte für diese Proben quantitativ. Es lässt sich auch die transportverstärkende Wirkung der Vaseline mit hoher Ortsauflösung beobachten. Beide genutzten Nanocarrier zeigen einen Transport des Wirkstoffs Rapamycin in die Haut. Langzeitbehandlungen zeigen eine Anreicherung der lipophilen Nanocarrier in den Lipidmembranen der Zellkerne der viablen Epidermis. Die Messung einer Kryoprobe mittels AFM-IR zeigt vergleichbare Ergebnisse zu den fixierten und eingebetteten Proben. Dies zeigt, dass die umfangreiche Probenvorbereitung fixierter Haut für die Röntgenmikroskopie keinen Einfluss auf die lokale Wirkstoffverteilung hat.
Im zweiten Teil der Arbeit werden die Wirkstoffe Cetuximab und Docetaxel in SCC-25-Kollagen-Zellkulturen auf Basis von hyperspektralen AFM-IR-Messungen ortsaufgelöst und labelfrei nachgewiesen. Die Auswertung erfolgt ebenfalls auf Basis einer numerischen Singulärwertzerlegung mit den jeweiligen Spektren der Reinsubstanzen im mittleren Infrarotbereich zur Bestimmung der lokalen Wirkstoffverteilung. Die Gabe des Wirkstoffs Cetuximab erfolgt bei einer Zellkultur topisch auf den Tumorzellen und bei der anderen über das Nährmedium an der Seite der Kollagene, um eine systemische Behandlung zu simulieren. Es soll untersucht werden, ob eine topische Behandlung als Ergänzung oder Alternative zu einer systemischen dienen kann. Der Wirkstoff Cetuximab kann bei beiden Autragungsmethoden detekiert werden. Messungen mit hoher räumlicher Auflösung deuten darauf hin, dass eine zelluläre Aufnahme des Wirkstoffs stattfindet. Docetaxel kann in zwei Zellkulturen, welche mit unterschiedlichen Konzentrationen topisch behandelt wurden, labelfrei nachgewiesen werden.In this thesis, drugs were detected spatially resolved in biological samples using label-free methods. The detection was performed by simulating the experimental spectra taken in hyperspectral measurements in the spectral in the regime of O 1s edge (520-565 eV) and in the mid-infrared regime (1190-1320 cm^-1 and 1600-1850 cm^-1) using a numerical singular value decomposition based on individually measured reference spectra and spectra of the untreated biological species.
The first part of this work involves label-free detection of the drug rapamycin and the redoxsensitive nanocarriers in topically treated human skin ex vivo. Here, samples embedded in epoxy resin were examined using scanning X-ray microscopy at the O 1s edge. Additionally the samples were examined using photothermal expansion spectroscopy (AFM-IR). It is shown that pretreatment with the serine protease trypsin opens the membranes of the corneocytes and the drug can be transported intracellularly into the stratum corneum. Quantitative drug detection was accomplished for these samples. The transport enhancing effect of petrolatum was also investigated with high spatial resolution. Redoxsensitive core-multishell nanocarriers show enhanced rapamycin penetration into the top skin layers. Long-term treatments show an accumulation of the lipophilic nanocarriers in the lipid membranes of the cell nuclei in the viable epidermis. Studies on a cryosample by AFM-IR shows comparable results to fixed and embedded skin samples. This supports the validity of extensive sample preparation protocols used along with scanning X-ray microscopy indicating that there is no influence on the local drug distribution.
In the second part of this work, the drugs cetuximab and docetaxel are spatially resolved and label-free detected in SCC-25 collagen cell cultures based on hyperspectral AFM-IR measurements. Singular value decomposition is also used for determining the local drug distribution using reference spectra of the pure substances in the mid-infrared regime. The drug cetuximab is topically applied on a cellular tumor model to prove the efficacy of this approach. Alternatively, the drug is administered via the culture medium on the side of the collagen base of the model in the other to simulate a systemic treatment. The aim of this approach wasto investigate whether topical treatment can serve as an adjunct or alternative to the established systemic treatment. The results indicate that drug cetuximab can be detected by both systemic and topical treatment. High spatialresolution measurements indicate that cellular uptake of the drug takes place. Label-free detection of docetaxel was also accomplished in cell cultures that were topically treated with different drug concentrations
Mucin-assoziierte regulatorische Faktoren in C. jejuni Infektionen und Fortschritte in der Sphäroid-Modellierung
No full textCampylobacter jejuni is a prevalent zoonotic pathogen and the most common source of bacterial intestinal infections worldwide. However, the pathogenesis of the infection is not fully understood and detailed research on the underlying molecular mechanisms of many host-pathogen factors involved in colonization and infection is needed. C. jejuni has adapted several methods to penetrate the double mucus layer in the human colon, infect the epithelium and cause acute inflammation processes. Main component of this protecting barrier is MUC2, which is highly glycosylated by mucin-type O-glycosylations through specific glycosyltransferases. The expression of several glycosyltransferases involved in mucin-type O-glycosylation is dysregulated in C. jejuni infections, which may affect the mucosal defense mechanisms and influence the severity of the infection.
miRNAs play an important role in controlling mammalian host cell responses to bacterial infections. It was shown that the mucin-associated miRNAs miR-125a-5p and miR-615-3p are dysregulated in C. jejuni infections. Moreover, a potential regulation of miR-125a-5p on the expression of the sialyltransferase St3gal1 and the glycosyltransferase B4galt1 was demonstrated by combining in silico, in vitro and in vivo approaches. St3gal1 and B4galt1 are both involved in mucin-type O-glycosylations and significantly upregulated in an infection-dependent manner in the colon of C. jejuni infected mice compared to naïve control animals. Hence, miR-125a-5p possibly participates in regulating the glycosylation pattern of important host cell response proteins like MUC2 upon infection.
For further investigations, human structured multicellular intestinal spheroids (SMIS) were generated. This advanced 3D cell culture model comprises of four relevant cell lines, with a fibroblast core that is surrounded by a single layer of enterocytes, goblet cells and activated monocytes. SMIS thereby replicate the microarchitecture of the luminal surface of the human intestinal mucosa. Further, the spheroids show differentiated morphological characteristic like microvilli after only two days in culture. SMIS were specifically designed to study molecular mechanisms following the infection by intestinal pathogens. After C. jejuni infections, they exhibit regulatory patterns of immunological markers that surpass 2D monolayers and closely align with in vivo responses to the infection. The human SMIS protocol is adaptable to the species mouse and pig, demonstrating its versatility.Campylobacter jejuni ist ein weit verbreitetes zoonotisches Pathogen und die häufigste Ursache bakterieller intestinaler Infektionen weltweit. Die Pathogenese der Infektion ist jedoch unzureichend erforscht und bedarf weiterführender Untersuchungen hinsichtlich der zugrundeliegenden molekularen Mechanismen von Wirt und Pathogen, die an Kolonisation und Infektion beteiligt sind. C. jejuni hat verschiedene Methoden entwickelt, um die schützende doppelte Mucus-Schicht im menschlichen Colon zu durchdringen, um dann das Epithelium zu infizieren und akute Inflammationsprozesse auszulösen. Hauptbestandteil dieser schützenden Barriere ist MUC2, das durch muzinartige O-Glykosylierungen, die von spezifischen Glykosyltransferasen katalysiert werden, stark glykosyliert ist. Die Expressionen mehrerer Glykosyltransferasen, die an muzinartigen O-Glykosylierungen beteiligt sind, sind bei C. jejuni Infektionen dysreguliert. Diese Veränderung kann die mukosalen Abwehrmechanismen beeinträchtigen und die Schwere der Infektion beeinflussen.
miRNAs spielen in Säugetieren bei der Kontrolle der Wirtszellantwort auf bakterielle Infektionen eine wichtige Rolle. Es konnte gezeigt werden, dass die muzin-assoziierten miRNAs miR-125a-5p and miR-615-3p in C. jejuni Infektionen dysreguliert sind. Darüber hinaus konnte eine mögliche Regulation von miR-125a-5p auf die Expression von der Sialyltransferase St3gal1 und die Glykosyltransferase B4galt1 durch die Kombination von in silico, in vitro und in vivo Methoden nachgewiesen werden. St3gal1 und B4galt1 sind beide an muzinartigen O-Glykosylierungen beteiligt und bei einer Infektion im Colon von C. jejuni infizierten Mäusen verglichen mit nicht-infizierten Kontrolltieren signifikant hochreguliert. Daraus ist zu schließen, dass miR-125a-5p Teil der Regulationsprozesse von Glykosylierungsmustern wichtiger Wirtszellenproteine als Antwort auf Infektionen ist.
Für weiterführende Untersuchungen wurden menschliche strukturierte multizelluläre intestinal Sphäroide (SMIS) entwickelt. Dieses 3D-Zellkulturmodell besteht aus vier relevanten Zelllinien mit einem Fibroblastenkern, der von einer Einzelschicht aus Enterozyten, Becherzellen und aktivierten Monozyten umgeben ist. Damit repliziert SMIS die Mikroarchitektur der luminalen Oberfläche der menschlichen intestinalen Mukosa. Zusätzlich weisen die Sphäroide bereits nach zwei Tagen in Kultur differenzierte morphologische Merkmale wie Mikrovilli auf. SMIS wurden speziell dafür entwickelt, molekulare
3
Mechanismen nach Infektionen mit Darmpathogenen zu untersuchen. Nach C. jejuni Infektion zeigen sie regulatorische Muster immunologischer Marker, die die von 2D-Monolayern übertreffen und die den in vivo Reaktionen auf die Infektion stark ähneln. Das menschliche SMIS-Protokoll ist auf die Spezies Maus und Schwein übertragbar, was dessen Vielseitigkeit demonstriert
Nanoscale Tools for Biosensing and Treatment of Bacterial Biofilms and Eukaryotic Cells
No full textNanoparticles are well-suited for developing diagnostic and therapeutic tools due to their
small size and favorable physicochemical properties. They provide unique features that many conventional approaches cannot, such as high-resolution imaging utilizing nanosensors, improved bioavailability of nanoformulated therapeutics, and targeted delivery. Polymeric nanoparticles can be easily structurally modified to tailor specific properties, making them suitable for a wide range of applications. Three studies were carried out as part of this thesis, demonstrating the diverse applications of polystyrene nanoparticles as nanosensors for biosensing in biofilms and eukaryotic cells, and as nanocarriers for the treatment of pathogenic biofilms.
In the first study, a pH nanosensor based on biocompatible polystyrene nanoparticles was
developed to determine and visualize the pH in biofilms. The nanosensor employs a ratiometric principle to determine pH, based on the fluorescence intensity ratio of the pH insensitive dye nile red and the pH-sensitive dye fluorescein isothiocyanate (FITC). The fluorescence is acquired by confocal laser scanning microscopy. This method allows for threedimensional measurement of pH over extended time periods, enabling detailed studies of dynamic processes in biofilms. The study demonstrated the functionality of the pH nanosensor by imaging the time-dependent pH changes induced by the metabolic activity of Escherichia coli biofilms. The nanosensor is easy to use, no special equipment is required, yet the measurements are precise, and the sensor is very robust. This is achieved by the smart design concept with its ratiometric working principle, making it a valuable tool for characterizing the chemical microenvironment of biofilms. The pH nanosensor can improve the understanding of biofilm dynamics and enable the development of improved strategies to combat biofilmassociated health problems in industry and for clinical settings.
The second study describes a nanosensor for the determination of extracellular pH and the
extracellular pH microenvironment of eukaryotic cells. The nanosensor operates on the same principle as the sensor in the first study, but a significant addition enables the direct
measurement of pH at the cell surface. The nanosensor is conjugated to a lectin, which binds to the cell membrane and anchors the nanosensor to the cell surface. This method enables a precise and spatially resolved measurement of extracellular pH at the cell surface of individual cells. The study demonstrates the versatility and compatibility of this pH nanosensor with different cell lines from various organs, combined with effective targeting. It has great potential for studying the cellular microenvironment and gaining a deeper understanding of cellular processes based on these microenvironments. Its applications are found in biomedical research, particularly in cancer research, for understanding and studying metabolic disorders, and for diagnostic or therapeutic purposes.
In the third study, a novel tool for the photodynamic eradication of biofilms was developed
and applied. Polystyrene nanoparticles were used as carriers to embed the lipophilic
photosensitizer (a boron-dipyrromethene derivative) and deliver it to the biofilm for
activation at the target site. The study demonstrated that the photosensitizer-loaded
nanoparticles were highly effective against planktonic bacteria and bacterial biofilms of
pathogenic bacteria such as Escherichia coli, Staphylococcus aureus, and Streptococcus
mutans. Furthermore, the study aimed to characterize the interactions between nanoparticles
and biofilms to enhance the understanding of the mechanisms behind antimicrobial
photodynamic therapy using nanoscale treatment agents against biofilms. The
photosensitizer-loaded nanoparticles were found to be a highly effective tool in the
prevention and removal of biofilms. They showed even higher efficacy than many tools in
previously published studies about antimicrobial photodynamic therapy, both with and
without nanoparticles. The nanoparticles presented in this study have great potential to be
used as effective tools in the fight against biofilms. They offer a practical and straightforward
alternative to existing methods, with a lower risk of bacterial resistance developing in the
future.
In summary, these studies highlight the potential of polymeric nanoparticles as carriers for
effective antimicrobial treatment and as sensors for providing valuable insights into biofilm
microenvironments. Furthermore, they enable precise extracellular pH measurements in
diverse cell lines. These advancements hold promise for future research and applications in
fields ranging from biofilm characterization to biomedical research and antimicrobial therapy
Väterliche Programmierung von Nieren- und Stoffwechselerkrankungen bei Nachkommen
No full textPaternal programming explores how a father's experiences, exposures, and epigenetic changes impact the health of his offspring and future generations. This research has broad implications in genetics, epigenetics, and developmental biology, shedding light on how environmental factors experienced by males, such as diet, can influence offspring health. These insights are critical for informing public health policies and lifestyle recommendations.
We focused on the effects of a paternal pre-conception diet high in fat, sucrose, and salt—similar to a fast food diet—on offspring kidney health. Using a rat model, we found that such a diet could induce chronic kidney disease (CKD) in the F1 and F2 generations. The most significant effects were observed in female F2 offspring, who exhibited decreased glomerular filtration rate (GFR), increased urinary albumin excretion, glomerulosclerosis, and interstitial fibrosis. We analyzed the underlying epigenetic mechanisms through RNA sequencing, qPCR, DNA methylation, and systems biology approaches, making our study the first to establish a paternal diet-induced CKD model in rats.
In a subsequent study, we explored an advanced fetal programming hypothesis by examining the effects of paternal eNOS deficiency on offspring health without transmitting the gene defect to the next generation. By crossbreeding male mice with a heterozygous eNOS genotype with wild-type females, we found that wild type male offspring born to eNOS-deficient fathers showed elevated fasting insulin levels, increased insulin responses, and higher liver glycogen content. These changes were linked to altered gene expression and DNA methylation in the offspring, particularly in genes such as Pgc1a and Gr, with effects more pronounced in male offspring. Further, we revealed that hepatic fat accumulation in female offspring of eNOS+/- mothers was associated with increased liver Fitm1 expression. In male offspring of eNOS-deficient fathers, elevated Tfb2m expression correlated with increased insulin levels post-glucose load. Additionally, seven genes were significantly linked to enhanced liver glycogen levels in these male offspring. We also found a connection between altered gene expression, changes in the metabolite fumarate, and increased liver glycogen.
These findings highlight the significant influence of paternal factors on the health of future generations, emphasizing the complex interactions between genetics, epigenetics, and the environment. Understanding the mechanisms of paternal programming provides valuable insights for developing intervention strategies to mitigate adverse health outcomes in offspring, with important implications for public health policies and lifestyle modifications aimed at improving the well-being of future generations.Die väterliche Programmierung untersucht, wie die Erfahrungen, Expositionen und epigenetischen Veränderungen eines Vaters die Gesundheit seiner Nachkommen und zukünftiger Generationen beeinflussen. Diese Forschung hat weitreichende Implikationen in den Bereichen Genetik, Epigenetik und Entwicklungsbiologie und beleuchtet, wie Umweltfaktoren, denen Männer ausgesetzt sind, wie etwa die Ernährung, die Gesundheit ihrer Nachkommen beeinflussen können. Diese Erkenntnisse sind entscheidend für die Gestaltung von Gesundheitspolitiken und Lebensstilempfehlungen.
Wir konzentrierten uns auf die Auswirkungen einer väterlichen Vorkonzeptionsdiät, die reich an Fett, Zucker und Salz ist – ähnlich einer Fast-Food-Diät – auf die Nierengesundheit der Nachkommen. Anhand eines Rattenmodells fanden wir heraus, dass eine solche Ernährung chronische Nierenerkrankungen (CKD) in der F1- und F2-Generation verursachen kann. Die bedeutendsten Effekte wurden bei weiblichen F2-Nachkommen beobachtet, die eine verringerte glomeruläre Filtrationsrate (GFR), erhöhte Albuminausscheidung im Urin, Glomerulosklerose und interstitielle Fibrose aufwiesen. Die zugrunde liegenden epigenetischen Mechanismen analysierten wir mittels RNA-Sequenzierung, qPCR, DNA-Methylierung und systembiologischen Ansätzen, wodurch unsere Studie die erste war, die ein durch väterliche Ernährung induziertes CKD-Modell bei Ratten etablierte.
In einer nachfolgenden Studie untersuchten wir eine erweiterte Hypothese zur fetalen Programmierung, indem wir die Auswirkungen eines väterlichen eNOS-Mangels auf die Gesundheit der Nachkommen untersuchten, ohne den Gendefekt an die nächste Generation weiterzugeben. Durch die Kreuzung von männlichen Mäusen mit einem heterozygoten eNOS-Genotyp mit wildtypischen Weibchen fanden wir heraus, dass wildtypische männliche Nachkommen von eNOS-defizienten Vätern erhöhte Nüchterninsulinspiegel, gesteigerte Insulinreaktionen und einen höheren Leberglykogengehalt aufwiesen. Diese Veränderungen standen im Zusammenhang mit veränderter Genexpression und DNA-Methylierung bei den Nachkommen, insbesondere in Genen wie Pgc1a und Gr, wobei die Effekte bei männlichen Nachkommen ausgeprägter waren. Darüber hinaus zeigten wir, dass die Anreicherung von Leberfett bei weiblichen Nachkommen von eNOS+/- Müttern mit einer erhöhten Expression von Fitm1 in der Leber verbunden war. Bei männlichen Nachkommen von eNOS-defizienten Vätern korrelierte eine erhöhte Tfb2m-Expression mit erhöhten Insulinspiegeln nach einer Glukosebelastung. Zusätzlich wurden bei diesen männlichen Nachkommen sieben Gene identifiziert, die signifikant mit erhöhten Leberglykogenspiegeln in Verbindung standen. Wir fanden auch einen Zusammenhang zwischen veränderter Genexpression, Veränderungen des Metaboliten Fumarat und erhöhtem Leberglykogen.
Diese Erkenntnisse unterstreichen den erheblichen Einfluss väterlicher Faktoren auf die Gesundheit zukünftiger Generationen und betonen die komplexen Wechselwirkungen zwischen Genetik, Epigenetik und Umwelt. Das Verständnis der Mechanismen der väterlichen Programmierung liefert wertvolle Erkenntnisse für die Entwicklung von Interventionsstrategien zur Minderung negativer gesundheitlicher Auswirkungen bei Nachkommen, mit wichtigen Implikationen für Gesundheitspolitiken und Lebensstiländerungen, die darauf abzielen, das Wohlbefinden zukünftiger Generationen zu verbessern
- …
