18 research outputs found
Interactions of Gastrointestinal Peptides: Ghrelin and Its Anorexigenic Antagonists
Food intake behaviour and energy homeostasis are strongly regulated by a complex system of humoral factors and nerval structures constituting the brain-gut-axis. To date the only known peripherally produced and centrally acting peptide that stimulates food intake is ghrelin, which is mainly synthesized in the stomach. Recent data indicate that the orexigenic effect of ghrelin might be influenced by other gastrointestinal peptides such as cholecystokinin (CCK), bombesin, desacyl ghrelin, peptide YY (PYY), as well as glucagon-like peptide (GLP). Therefore, we will review on the interactions of ghrelin with several gastrointestinal factors known to be involved in appetite regulation in order to elucidate the interdependency of peripheral orexigenic and anorexigenic peptides in the control of appetite
Expression of Toll-Like Receptors in the Developing Brain
Toll-like receptors (TLR) are key players of the innate and adaptive immune response in vertebrates. The original protein Toll in Drosophila melanogaster regulates both host defense and morphogenesis during development. Making use of real-time PCR, in situ hybridization, and immunohistochemistry we systematically examined the expression of TLR1–9 and the intracellular adaptor molecules MyD88 and TRIF during development of the mouse brain. Expression of TLR7 and TLR9 in the brain was strongly regulated during different embryonic, postnatal, and adult stages. In contrast, expression of TLR1–6, TLR8, MyD88, and TRIF mRNA displayed no significant changes in the different phases of brain development. Neurons of various brain regions including the neocortex and the hippocampus were identified as the main cell type expressing both TLR7 and TLR9 in the developing brain. Taken together, our data reveal specific expression patterns of distinct TLRs in the developing mouse brain and lay the foundation for further investigation of the pathophysiological significance of these receptors for developmental processes in the central nervous system of vertebrates
Docosahexaensäure unterdrückt die Arachidonsäure-induzierte Proliferation der humanen Kolonkarzinomzelllinie LS 174T
Background: Colorectal cancer (CRC) is one of the leading causes of death in
Western countries. A potential role of omega-3 and omega-6 polyunsaturated
fatty acids (n-3 and n-6 PUFAs) in the context of CRC has been implicated by
numerous epidemiological studies, suggesting a protective role of n-3 PUFAs.
On the other hand it is well-established that prostaglandin E2 (PGE2),
generated from n-6 PUFA arachidonic acid (AA), is an important promoting
factor in the tumorigenesis of the colorectum. In this context it has been
proposed that the human genetic profile was originally established on a n-6 to
n-3 PUFA ratio of approximately 1:1 as found in “ancient” diets, whereas
today’s Western diet has been estimated to provide n-6 to n-3 PUFAs in a ratio
of 15:1–20:1. It has been hypothesized that this may contribute to many
serious health issues typically found in Western societies, including CRC.
However, previous in vitro observations have led to some uncertainty regarding
differential roles of n-3 and n-6 PUFAs in CRC cells. While the majority of
investigations conducted in this field addressed neither the effects of n-6
PUFAs nor the impact of a balanced n-6 to n-3 PUFA ratio, several other
studies reported n-3 and n-6 PUFAs to exert anti-cancerous effects in vitro.
Hence, it was the aim of the present study to investigate the impact of n-3
PUFA docosahexaenoic acid (DHA) and n-6 PUFA AA and their combination on CRC
cell line LS 174T in vitro. Methods: In order to study the effects of n-3 PUFA
DHA, n-6 PUFA AA, PGE2 and several combinations on cellular viability of LS
174T CRC cells, XTT assays were applied. Cell cycle and cell death were
assessed via flow cytometry experiments and DAPI stainings. Moreover, mRNA
expressions of cyclooxygenase-2 (COX-2), B-cell lymphoma-2 (Bcl-2) and cyclin-
dependent kinase inhibitor 1A (p21WAF1) in cells incubated with AA, DHA or
vehicle were examined by semiquantitive real-time PCR (qRT-PCR). Biosynthesis
of PGE2 in the presence of AA and DHA was measured in a PGE2-ELISA. Results:
While incubation with n-6 PUFA AA increased cellular proliferation,
supplementation of n-3 PUFA DHA significantly reduced cellular viability and
induced apoptosis in a time- and dose-dependent manner. Moreover, DHA
incubation potently decreased mRNA expression of anti-apoptotic Bcl-2 and
induced the expression of cell cycle inhibitor p21WAF1. In contrast,
incubation with AA resulted in a significantly increased expression of Bcl-2
mRNA, whereas amounts of p21WAF1 mRNA remained unaltered compared to control
cells. In addition, AA incubation increased mRNA levels of COX-2, the key
enzyme in AA-dependent PGE2 formation, and also resulted in a significant
biosynthesis of PGE2, an important inductor of cellular proliferation in LS
174T cells. In contrast, DHA incubation decreased COX-2 mRNA expression and
significantly diminished AA-induced PGE2 formation in co-incubation
experiments. Importantly, results from co-incubations furthermore indicated
DHA to potently reverse and suppress AA- as well as PGE2-induced CRC cell
growth. Conclusion and impact: Results presented here clearly demonstrate
diametrically opposing effects of n-3 PUFA DHA and n-6 PUFA AA in regard of
cellular viability and mRNA regulation, thus helping to clarify the
aforementioned uncertainty concerning differential effects of n-3 and n-6
PUFAs in vitro. As novel aspect in this context, the present investigation
addresses the interplay between AA and DHA in the context of CRC cells in
vitro. AA/DHA co-incubations, leading to an (almost) balanced n-6 to n-3
ratio, thus mimicking conditions found in “ancient” diets, showed DHA to
potently reverse pro-proliferatory effects of n-6 PUFA AA and of AA-derived
PGE2 towards growth inhibition. Thus, our data add evidence to the argument
that the ratio of n-6/n-3 PUFA (particularly the ratio of AA/DHA) may be a
critical determinant of proliferation and tumor growth in CRC. However,
further in vitro and in vivo studies are required in order to fully elucidate
this interplay and its clinical impact and relevance. As another important
aspect, DHA potently diminished PGE2 formation from exogenous AA and decreased
mRNA expression of COX-2. This may be of special interest in regard of CRC
prevention as pharmacological inhibition of COXs (via ASS and other
nonsteroidal anti-inflammatory drugs), subsequently reducing PGE2 synthesis,
has been found to be protective against CRC development. Nevertheless, long
term use of these drugs is associated with severe adverse effects and is a
matter of ongoing debate. Thus, based upon the present data and previous
studies, n-3 PUFAs including DHA may be hypothesized as an alternative
preventive agent against CRC, also interfering with the AA-associated
metabolism. However, besides interaction with the COX-dependent AA metabolism,
data of the present study additionally suggest DHA to exert its anti-cancerous
actions also via COX- and AA-independent pathways, which yet remain to be
fully understood by future investigations. In conclusion, although deduced
from a limited in vitro setup, the present study provides significant evidence
concerning beneficial effects of DHA as well as for the pivotal importance of
n-6 to n-3 ratio in the context of CRC.Hintergrund: Das kolorektale Karzinom stellt eine der häufigsten Todesursachen
in den westlichen Industrienationen dar. Zahlreiche epidemiologische Studien
weisen in diesem Zusammenhang auf einen Einfluss mehrfach ungesättigter
Omega-3- und Omega-6-Fettsäuren (n-3 PUFAs und n-6 PUFAs) hin, wobei n-3 PUFAs
im Allgemeinen eine protektive Wirkung zugeschrieben wird. Andererseits ist
bekannt, dass Prostaglandin E2 (PGE2), ein aus der n-6 PUFA Arachidonsäure
(AA) synthetisierter Lipidmediator, die kolorektale Karzinogenese fördert. In
diesem Kontext wurde postuliert, dass sich das menschliche Genprofil
ursprünglich auf Basis einer Ernährung mit ausgeglichenem n-6 zu n-3 PUFA-
Verhältnis entwickelte, während das n-6 zu n-3 PUFA-Verhältnis heutiger
westlicher Nahrung bei etwa 15:1-20:1 liegt. Diese veränderte
Nahrungszusammensetzung wird von vielen Experten für die hohe Prävalenz vieler
sogenannter „Wohlstandskrankheiten“, zu denen auch das kolorektale Karzinom
zählt, mitverantwortlich gemacht. In bisherigen in vitro-Untersuchungen zeigte
sich jedoch kein einheitliches Bild bezüglich unterschiedlicher Wirkungen von
n-3 und n-6 PUFAs auf Kolonkarzinomzellen. Während die meisten Studien in
diesem Gebiet weder die Effekte von n-6 PUFAs noch den Einfluss des n-3 zu n-6
PUFA-Verhältnisses untersuchten, zeigten andere Veröffentlichungen
antikanzerogene Wirkungen beider Fettsäureklassen. Das Ziel der vorliegenden
Arbeit war es daher, die Wirkung der n-3 PUFA Docosahexaensäure (DHA), der n-6
PUFA AA und des AA-basierten Lipidmediators PGE2 sowie deren Kombination auf
die Kolonkarzinom-Zelllinie LS 174T zu untersuchen. Methoden: Der Einfluss von
DHA, AA, des Lipidmediators PGE2 und deren Kombination auf Überleben und
Wachstum der Kolonkarzinom-Zelllinie LS 174T wurden mittels XTT Assays
untersucht. Mit Hilfe von Durchflusszytometrie und DAPI-Färbungen wurden die
Effekte der Fettsäuren auf Zellzyklus und Zelltod evaluiert, während der
Einfluss beider Fettsäuren auf die mRNA-Expression von Cyclooxygenase-2
(COX-2), B-cell lymphoma-2 (Bcl-2) und Cyclin-dependent kinase inhibitor 1A
(p21WAF1) durch semiquantitive real-time PCR ermittelt wurde. Zusätzlich wurde
der Einfluss von AA und DHA auf die PGE2-Synthese durch ELISA-Untersuchungen
quantifiziert. Ergebnisse: Während die Inkubation mit der n-6 PUFA AA die
Proliferation der LS 174T Zellen anregte, reduzierte DHA zeit- und
dosisabhängig das Überleben der Karzinomzellen und induzierte Apoptose.
Weiterhin verminderte die Inkubation mit DHA die mRNA-Expression des
antiapoptotischen Proteins Bcl-2 und erhöhte dan mRNA-Level des
Zellzyklusinhibitors p21WAF1. Im Gegensatz hierzu führte die Zugabe von AA zur
vermehrten Expression von Bcl-2-mRNA, während sich die Menge an p21WAF1-mRNA
unverändert zeigte. Daneben induzierte die Inkubation mit AA die mRNA-
Expression von COX-2 (dem Schlüsselenzym der AA-abhängigen PGE2-Synthese) und
resultierte außerdem in einer gesteigerten PGE2-Biosynthese. Demgegenüber
verminderte die Inkubation mit DHA sowohl die Expression von COX-2-mRNA als
auch die AA-induzierte PGE2-Synthese. Weitere Koinkubationsexperimente
zeigten, dass DHA dem AA- und PGE2-induzierten Wachstum in LS 174T
entgegenwirkt. Diskussion und Fazit: Die vorliegende Studie zeigt
gegensätzliche Effekte der n-3 PUFA DHA und der n-6 PUFA AA bezüglich
Proliferation, Apoptose und mRNA-Regulation in der kolorektalen
Karzinomzelllinie LS 174T und trägt somit dazu bei, die Wirkung dieser
Fettsäuren weiter zu klären. Als neuer Aspekt wurde die Interaktion zwischen
AA und DHA in vitro untersucht. AA/DHA-Koinkubationsexperimente mit einem
(fast) ausgeglichenen Verhältnis von n-6 zu n-3 PUFAs (entsprechend der
Zusammensetzung der Nahrung vor der Industrialisierung) zeigten, dass DHA
sowohl dem AA- als auch dem PGE2-induzierten Wachstum entgegenwirkt. Diese
Ergebnisse unterstreichen den Einfluss des n-6 zu n-3 Verhältnisses (im
Besonderen AA zu DHA) im Kontext der kolorektalen Karzinogenese, wobei weitere
in vitro- und in vivo-Studien nötig sind, um die Interaktion der
unterschiedlichen Fettsäuren sowie deren klinische Relevanz vollständig zu
erfassen. Daneben zeigen die vorliegenden Experimente, dass DHA sowohl die
PGE2-Synthese aus exogen zugeführter AA als auch die mRNA-Expression des
Schlüsselenzyms COX-2 reduziert. Dies ist insofern von besonderem Interesse,
als dass die pharmakologische Inhibition der COXs (beispielsweise durch
Acetylsalicylsäure oder andere nicht-steroidale Antiphlogistika) mit der
hieraus resultierenden Reduktion der PGE2-Synthese präventiv gegen das
kolorektale Karzinom wirkt, jedoch aufgrund der assoziierten Nebenwirkungen
nicht allgemein empfohlen wird. In diesem Zusammenhang legen die vorliegenden
Daten in Zusammenschau mit den Ergebnissen anderer Studien nahe, dass
alternativ n-3 PUFAs wie DHA als präventive, mit dem AA-Metabolismus
interferierende Substanzen eingesetzt werden könnten. Darüber hinaus wird aber
aus den vorliegenden Daten zusätzlich deutlich, dass die anti-kanzerogenen
Effekte von DHA zum Teil auch auf COX- und AA-unabhängigen Mechanismen
basieren, die in zukünftigen Studien weiter erforscht werden sollten.
Zusammenfassend liefert die vorliegende Studie neue und signifikante Hinweise
bezüglich protektiver Effekte der n-3 PUFA DHA und hebt die Relevanz eines n-6
zu n-3 PUFA-Verhältnisses im Kontext des kolorektalen Karzinoms hervor
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Prostate Cancer Diagnosis and Characterization with Mass Spectrometry Imaging
Background: Prostate cancer (PCa), the most common cancer and second leading cause of cancer death in American men, presents the clinical challenge of distinguishing between indolent and aggressive tumors for proper treatment. PCa presents significant alterations in metabolic pathways that can potentially be measured using techniques like mass spectrometry (MS) or mass spectrometry imaging (MSI) and used to characterize PCa aggressiveness. MS quantifies metabolomic, proteomic, and lipidomic profiles of biological systems that can be further visualized for their spatial distributions through MSI. Methods: PubMed was queried for all publications relating to MS and MSI in human prostate cancer from April 2007 to April 2017. With the goal of reviewing the utility of MSI in diagnosis and prognostication of human PCa, MSI articles that reported investigations of PCa-specific metabolites or metabolites indicating PCa aggressiveness were selected for inclusion. Articles were included that covered MS and MSI principles, limitations, and applications in PCa. Results: We identified nine key studies on MSI in intact human prostate tissue specimens that determined metabolites which could either differentiate between benign and malignant prostate tissue or indicate prostate cancer aggressiveness. These MSI-detected biomarkers show promise in reliably identifying PCa and determining disease aggressiveness. Conclusions: MSI represents an innovative technique with the ability to interrogate cancer biomarkers in relation to tissue pathologies and investigate tumor aggressiveness. We propose MSI as a powerful adjuvant histopathology imaging tool for prostate tissue evaluations, where clinical translation of this ex vivo technique could make possible the use of MSI for personalized medicine in diagnosis and prognosis of prostate cancer. Moreover, the knowledge provided from this technique can majorly contribute to the understanding of molecular pathogenesis of PCa and other malignant diseases
High Resolution Magic Angle Spinning Proton NMR Study of Alzheimer’s Disease with Mouse Models
Alzheimer’s disease (AD) is a crippling condition that affects millions of elderly adults each year, yet there remains a serious need for improved methods of diagnosis. Metabolomic analysis has been proposed as a potential methodology to better investigate and understand the progression of this disease; however, studies of human brain tissue metabolomics are challenging, due to sample limitations and ethical considerations. Comprehensive comparisons of imaging measurements in animal models to identify similarities and differences between aging- and AD-associated metabolic changes should thus be tested and validated for future human non-invasive studies. In this paper, we present the results of our highresolution magic angle spinning (HRMAS) nuclear magnetic resonance (NMR) studies of AD and wild-type (WT) mouse models, based on animal age, brain regions, including cortex vs. hippocampus, and disease status. Our findings suggest the ability of HRMAS NMR to differentiate between AD and WT mice using brain metabolomics, which potentially can be implemented in in vivo evaluations
Docosahexaenoic acid suppresses arachidonic acid-induced proliferation of LS-174T human colon carcinoma cells
AIM: To investigate the impact of arachidonic acid (AA) and docosahexaenoic acid (DHA) and their combination on colon cancer cell growth