Background and aims: Newly synthesised cholesterol contributes poorly to biliary lipid secretion but may assume greater importance when the rate limiting enzyme 3-hydroxy-3-methylglutaryl-CoA ...reductase (HMGR) is upregulated. As this occurs in the gall stone susceptible C57L/J inbred mouse, we employed two cholesterol biosynthesis inhibitors, Tu 2208 and Ro 48-8071, potent inhibitors of squalene epoxidase and oxidosqualene-lanosterol cyclase, respectively, to assess their potential in preventing cholesterol cholelithiasis in the C57L/J mouse strain. Mice were fed a lithogenic diet comprising a balanced nutrient intake with 15% dairy fat, 1% cholesterol, and 0.5% cholic acid added. Methods: We determined gall stone phenotype, HMGR activity, biliary lipid secretion rates, and counterregulatory events in male C57L/J mice and gall stone resistant AKR treated with Tu 2208 (30–60 mg/kg/day) or Ro 48-8071 (30–100 mg/kg/day), while ingesting chow or the lithogenic diet. Results: Both agents reduced the gall stone prevalence rate from 73% to 17% in C57L/J mice, inhibited HMGR activity, and decreased hepatic cholesterol concentrations without appreciably influencing biliary cholesterol secretion. In C57L as well as AKR mice, both agents increased biliary phospholipid (which is mostly phosphatidylcholine) secretion rates and at the highest doses effectively reduced the biliary cholesterol saturation index. Conclusions: Cholesterol biosynthesis inhibitors acting distally to squalene do not reduce biliary cholesterol secretion rates despite reductions in cholesterol biosynthesis and hepatocellular levels. However, they effectively prevent gall stone formation through stimulation of pathways that lead to enhanced biliary phospholipid secretion.
Objective - Angiotensin II plays an important role in the pathogenesis of atherosclerosis.This study was designed to examine the effect of valsartan, an angiotensin receptor blocker, on ...atherosclerotic lesions in pulmonary arteries of rabbits with atherosclerosis induced by a high-cholesterol (HC) diet.
Methods - Male New Zealand rabbits were randomly divided into four groups: control, HC without valsartan, HC with 3 mg/kg/d valsartan, and HC with 10 mg/kg/d valsartan. Following 12 weeks of treatment, serum lipid profiles were determined. Pulmonary arteries were harvested and stained with Sudan IV for evaluation of atherosclerotic lesions. The middle lobes of the rabbit lungs were isolated, embedded in paraffin, and sectioned and stained with haematoxylin and eosin. Nitric oxide (NO) and endothelin-1 (ET-1) levels were determined in pulmonary arteries by nitrate reductase assay and radioimmunoassay, respectively.
Results - HC feeding altered serum lipid levels and induced atherosclerotic lesions in pulmonary arteries. Although 12 weeks of valsartan treatment failed to alter serum lipid levels, it significantly ameliorated HC-induced atherosclerotic lesions. Lesion areas, inflammatory cell infiltration, and occlusions of small arteries of lungs were reduced. Moreover, the endothelium-derived NO levels in pulmonary arteries were increased by valsartan treatment (10 mg/kg/d) compared to levels in the HC group. ET-1 levels were decreased by valsartan treatment compared to levels in the HC group. NO and ET-1 levels were not altered by valsartan at 3 mg/kg/d.
Conclusion - Our data demonstrates that HC diet-induced atherosclerotic lesions in rabbit pulmonary arteries can be ameliorated by treatment with valsartan, possibly through a NO and ET-1-dependent mechanism.
Provider: - Institution: - Data provided by Europeana Collections- Ageing of the central nervous system is associated with a number of changes
that disturb nerve function, especially motor control ...and cognitive
abilities. It is believed that cognitive disorders in aging are consequence
of reduced synaptic plasticity, especially in the cortex and hippocampus,
brain regions extremely important for learning and memory processes. Dominant
factor that can influence aging in general, and therefore the aging of the
brain is reduced diet regime, without malnutrition. In addition to being
known for decades that reduced caloric intake prolongs life span of many
species, including mammals, it is shown that it delays and alleviates
age-dependent changes. Cholesterol is the most abundant lipid in mammalian
cells. By participating in the construction of phospholipid bilayer of
biological membranes, it regulates their fluidity, permeability and rigidity,
and consequently the functional properties of membrane proteins, such as ion
channels and transmmitters receptors. Cholesterol metabolism represents one
aspect of synaptic plasticity, as has been shown that cholesterol is
essential for biogenesis of synaptic vesicles, and vesicle transport, as well
as for proper functional organization of lipid rafts. The brain contains the
most cholesterol of all the organs in the body. Maintenance of cholesterol
homeostasis is essential for proper functioning of the central nervous
system. The aim of this study was to analyze the influence of aging and
long-term dietary restriction on cholesterol metabolism in the rat serum,
cortex and hippocampus. The experiments were performed on 3-, 12-, and
24-month-old male Wistar rats fed ad libitum (AL), or exposed to long term DR
(100% every other day-EOD) starting from the age of 3. At the indicated time
points, the rats were killed by decapitation. The expression of proteins
involved in cholesterol synthesis (HMGCR), transport (ApoE), catabolism
(CYP46) and regulation (LXRα) was determined using Western blot and
immunohistochemistry. The levels of cholesterol, its precursors (lanosterol,
lathosterol, desmosterol) and metabolites (24S-hydroxycholesterol,
27-hydroxycholesterol, cholestanol) were determined by gas
chromatography/mass spectrometry (GC/MS). The same method was used to analyze
the amount of phytosterols, given the increasing use of foods enriched with
plant analogues of cholesterol in human nutrition. Additionally, the link
between cholesterol content and neurotrophin signaling in the rat cortex and
hippocampus was analyzed at protein level. In rat serum cholesterol levels
were increased during ageing, regardless of diet regime. Long-term dietary
restriction counteracted age-dependent increase in the amount of cholesterol
precursors, maintaining their level at the one measured in 3 month old
animals. The same influence of dietary restrictions was detected on the
age-dependent increase in the cholestanol levels. The amount of
24S-hydroxycholesterol remained stable under experimental conditions, while
dietary restriction had no effect on elevated 27-hydroxycholesterol levels
during ageing. Ageing has led to increased phytosterols levels, while reduced
dietary intake maintained their content on control levels measured in
3-month-old animals. During aging, there was a reduction in HMGCR protein in
the rat cortex, regardless of the diet, and reduction in the levels of LXRα
protein, which has been reinforced by dietary restriction. The amount of ApoE
protein was increased during ageing. Dietary restriction in the group of
12-month-old animals acted contrary to aging, reducing the amount of this
protein in relation to age control. CYP46 protein levels were increased in
the group of 24-month-old animals exposed to diet. In the rat hippocampus
aging has caused decreased levels of HMGCR, CYP46 and LXRα proteins,
regardless of the diet. The amount of ApoE protein in the oldest group of
rats that were exposed to dietary restriction was increased in comparison to
age-control, and compared to 3 month old animals. Aging induced slight but
significant increase of cholesterol content in the rat cortex, while there
were no changes in the amount of cholesterol in the hippocampus. With the
exception of lanosterol at 12- month- old AL- fed animals, cholesterol
precursors were less common in aging in both brain regions examined. Dietary
restriction in the group 12mAL animals prevented the increase of lanosterol
amount, keeping it at the level detected in 3-month-old animals. In the
hippocampus of 24-month old animals, reduced diet intake increased
desmosterol amount in relation to age control, although the amount was still
less than that measured in 3-month-old controls. Cholestanol content was
increased during ageing, regardless of diet regime. The levels of
24S-hydroxycholesterol remained stable under experimental conditions,
indicating that experimental conditions had no influence on brain cholesterol
catabolism. Amount of plant sterols in both brain structures was increased
during aging, while the reduced diet intake acted contrary to the aging,
decreasing their levels in relation to age-appropriate controls. Only the
amount of brassicasterol in the cortex did not change in the analyzed
experimental conditions. In the rat cortex ageing influenced increased BDNF
protein levels, and decreased levels of activated receptor pTrkB. Dietary
restriction in the group of 12-month-old animals further increased the amount
of BDNF, while in the oldest group acted contrary to aging, keeping the
amount of this trephine on the control level. At 12-month-old animals dietary
restriction maintained levels of activated receptors at the one measured in
animals aged 3 months. The total receptor in the cortex did not change in the
analyzed conditions. Changes in trephine signaling in the hippocampus during
aging and under the influence of long-term dietary restriction were detected
only through the increase in BDNF protein in the group 12 months old animals
exposed to reduced feeding compared to 3 month old animals. The results
obtained in this thesis showed that the cholesterol metabolism in the rat
cortex and hippocampus have changed during ageing and under the influence of
long-term dietary restriction. The greatest changes were observed at the
level of cholesterol precursors, indicating a reduced rate of cholesterol
synthesis, while the elimination of cholesterol from the brain determined by
its derivate, 24S-hydroxycholesterol, was not altered under the experimental
conditions. Effect of long-term dietary restriction on cholesterol metabolism
was more pronounced in the hippocampus than in the cortex, and was similar to
the impact of reduced dietary intake manifested in the rat serum. The link
between cholesterol and neurotrophine signaling was more evident in the
cortex than in hippocampus. The results of this study showed the capacity of
long-term dietary restriction to influence cholesterol metabolism in the
brain by reducing its synthesis, whereas the amount of cholesterol remained
unchanged, and point to long-term DR as a possible alternative to statin use
for its ability to preserve cholesterol homeostasis in the brain.- Starenje centralnog nervnog sistema (CNS) praćeno je brojnim promenama koje
dovode do narušavanja nervnih funkcija, pre svega kontrole motorike i
kognitivnih sposobnosti. Smatra se da su kognitivni poremećaji tokom starenja
posledica smanjene sinaptičke plastičnosti, naročito u korteksu i
hipokampusu, regionima mozga izuzetno važnim za procese učenja i pamćenja.
Najvažniji faktor sredine kojim se može uticati na starenje uopšte, a samim
tim i na starenje mozga je režim redukovane ishrane, bez pothranjenosti.
Pored toga što se već nekoliko decenija zna da redukovana ishrana produžava
životni vek mnogih vrsta, uključujući i sisare, pokazano je da ona odlaže i
ublažava starosno zavisne promene u nervnom sistemu. Holesterol je
najzastupljeniji lipid u ćelijama sisara. Učestvujući u izgradnji
fosfolipidnog dvosloja bioloških mambrana, on reguliše njihovu fluidnost,
propustljivost i rigidnost, a samim tim i funkcionalna svojstva membranskih
proteina, poput jonskih kanala i transmiterskih receptora. Metabolizam
holesterola predstavlja važan aspekt sinaptičke plastičnosti, jer je pokazano
da je holesterol neophodan za biogenezu i transport sinaptičkih vezikula, kao
i za prvilnu funkcionalnu organizaciju lipidnih ostvaca. Od svih organa u
telu, mozak sadrži najviše holesterola. Održavanje homeostaze holesterola
neophodno je za normalno i neometano funkcionisanje CNS. Cilj ovog
istraživanja bio je da se vidi kako starenje i dugotrajna dijetalna
restrikcija (DR) utuču na metabolizam holesterola u mozgu. Mužjaci pacova
soja Wistar stari 3 meseca (m) bili su podeljeni u dve grupe. Prva grupa,
označena kao ad libitum (AL), dobijala je svakog dana 100% dnevnog unosa
hrane, dok je druga grupa (označena kao DR) stavljena na režim redukovane
ishrane koji je podrazumevao dobijanje 100% dnevnog unosa hrane svakog drugog
dana. Životinje su žrtvovane nakon dostizanja starosti od 12 i 24m. Životinje
stare 3m su predstavljale kontrolnu starosnu grupu. Metabolizam holesterola
određivan je na nivou proteina koji su uključeni u sintezu holesterola
(HMGCR), njegov transport (ApoE), katabolizam (CYP46) i regulaciju (LXRα)
metodom Western blota i imunohistohemijski. Zastupljenost samog holesterola,
njegovih prekursora (lanosterol, latosterol, dezmosterol) i metabolita
(24S-hidroksiholesterol, 27-hidroksiholesterol, holestanol) određena je
gasnom hromatografijom-masenom spektrometrijom (GC-MS). U primenjenim
eksperimentalnim uslovima istom metodom analizirana je i zastupljenost
biljnih sterola u svetlu sve veće upotrebe hrane obogaćene biljnim analozima
holesterola u ljudskoj ishrani. Pored toga, praćena je i veza između sadržaja
holesterola i neurotrofinske signalizacije. Praćenje promena u metabolizam
holesterola pokazalo je d
Provider: - Institution: - Data provided by Europeana Collections- Hypothalamic-pituitary-adrenal (HPA) axis represents a complex neuroendocrine system involved in maintaining optimal dynamic state ...of the organism as a whole (i.e. the state of homeostasis). Whether being in its basal state or in some of the perturbed stress states, this highly nonlinear system always functions under conditions far away from the thermodynamic equilibrium, possessing various feedback mechanisms. Consequently, hormones comprising the HPA axis can be found in different dynamic states. Under basal conditions, i.e. basal physiology, concentration of a hormone cortisol, the end-product and chief effector of human HPA axis, displays complex oscillatory time-evolution, with ultradian oscillations being superimposed on circadian oscillations. It has become evident from a growing body of evidence that adequate oscillatory dynamics of HPA axis is a necessary prerequisite for its normal functioning and health, given that alterations of HPA axis dynamics are associated with many metabolic and psychiatric disorders. Thus, more exhaustive understanding of dynamic properties of HPA axis is required, accounting for not only experimental studies but also modeling and numerical simulation predictions. To this end, many mathematical models emulating different dynamical properties of HPA axis, in its regular as well as in pathological states, have been recently developed. Modeling the mechanism of a process by employing jointly stoichiometric relations between the reaction species and basic principles of nonlinear dynamics has proven to constitute a useful approach in emulating various forms of complex dynamic states occurring in a variety of systems, including the oscillatory behavior within the HPA axis. The main goal of this dissertation is the modeling of dynamical states of human HPA axis and concentration of cortisol, as a representative hormone for describing these HPA axis states, occurring under different physiological conditions or due to the impact of the HPA axis perturbators, such as acute and chronic stress. This goal was accomplished by utilizing a stoichiometric model of HPA axis activity proposed by Jelić et al. (2005) as a starting ground. In this dissertation, the above-mentioned model was gradually developed by upgrading and adjusting its mechanism with known in vivo experimental data. During the course of these investigations, it was shown how predictive modeling based on stoichiometric relations between the model’s species possessed a strong potential for studying complex nonlinear reaction processes operating in states far from thermodynamic equilibrium, such as those appearing within the neuroendocrine HPA system. The initial model (Jelić et al., 2005) considers HPA axis as a nonlinear reaction network with positive feed-forward and negative feedback loops. This model takes into account only the most significant interactions among the four dynamical variable of the model (concentrations of key HPA axis hormones CRH, ACTH, cortisol and aldosterone), in accordance with established experimental findings. In numerical simulations, the model has demonstrated to be able to give rise to self-sustained ultradian oscillations. By coupling with a given circadian function, as a form of an external forcing, the model can simulate complex oscillatory dynamics, with ultradian oscillations being superimposed on circadian oscillations, as observed in vivo. Additionally, in previous studies the model has proven to possess strong predictive potential for describing dynamic response of the HPA axis under various acute and chronic stress conditions. In this doctoral thesis, the same model (basic model of the HPA system) was employed to investigate dynamic response of the HPA axis and cortisol concentration to acute perturbations with exogenous glucocorticoids, cortisol-alike drugs with strong anti-inflammatory and immunosuppressive effects, which are widely used for treating various medical conditions. Additionally, the model was employed to further characterize changes in dynamical states of the HPA occurring during chronic stress. The obtained results showed there was a strong dependence of the response of HPA axis and cortisol concentration response on intensity, ultradian and circadian timing of exogenous glucocorticoid perturbations. Furthermore, the simulation results indicated to the possibility of chronic stress to significantly modify important dynamic properties of dynamical states of cortisol and of other HPA axis hormones’ concentrations, such as mean level, shape, amplitude and frequency of their oscillations, and even the qualitative type of their dynamics. In that regard, bifurcation analysis has shown a nonlinear relationship between mean levels of CRH and the dynamic response of cortisol concentration. Such results might offer a possible theoretical interpretation in terms of nonlinear dynamics, for conflicting experimental findings regarding the ability of chronic stress to induce qualitatively opposite HPA axis dynamic behavior. In the following stage of investigation, the mechanism of the initial model was further extended and adjusted to the real, in vivo physiology of the HPA axis. Firstly, the basic model of the HPA system (Jelić et al., 2005) was extended by including reactions of eliminations of CRH and aldosterone. Subsequently, in thus obtained extended basic model of the HPA system, cholesterol, the initial precursor of cortisol, aldosterone and other steroid hormones, was introduced as a novel intermediary species, whose concentration represents an additional dynamical variable of the model. In order to give rise to oscillatory dynamics, each stoichiometric model must have at least one unstable nonequilibrium stationary (i.e. unstable steady) state. Determination whether the proposed reaction mechanism can at all be unstable, can be successfully carried out by employing stoichiometric network analysis (SNA), a mathematical method for stability analysis of models of arbitrary dimensionality, whose kinetics is based on the law of mass action. By utilizing an improved SNA methodology, it has been shown that each of derived models satisfies its own instability condition, thus being able to give rise to sustained oscillatory dynamics. This was further validated in numerical simulations. Furthermore, SNA has shown that all three investigated models of the HPA system (basic model, and both of the derived models – extended basic model and model with cholesterol) have common core of instability. Numerical simulations have shown that newly derived models retain many dynamic qualities of their predecessors, which is an important property and advantage of stoichiometric models. The HPA model with cholesterol has displayed several important improvements in comparison to the two previous ones. Shortcomings regarding CRH levels and aldosterone’s dynamics and concentration were corrected and adjusted to experimental observations. In simulations for all five dynamic variables (cholesterol, CRH, ACTH, cortisol and aldosterone), there was a good agreement with corresponding experimental concentration time-profiles of the species in terms of their oscillatory dynamical properties; concomitantly, concentration time-series for all species, except ACTH, were within their physiological levels. Additionally, model with cholesterol has shown that different types of dynamical states of cortisol concentration can arise by varying the values of cholesterol’s kinetic parameters (rate constants of its inflow or outflow). These results indicate to the significance of the role of cholesterol inflow and outflow rate, i.e. of cholesterol net reaction rate, on the dynamics of HPA axis as a whole, thus suggesting to their possible importance for maintaining HPA axis’ homeostatic capacity. Furthermore, the model with cholesterol predicts a negative correlation between mean cholesterol and mean cortisol concentrations, arising when the value of rate constant for cholesterol inflow into the system is changed. Taken altogether, these results indicate to the potential for further development of similar in silico experimental designs, which may be exploited as means towards more comprehensive understanding of intricate interplay between stress, HPA axis dynamics and cholesterol levels. In summary, in this doctoral thesis potentials and benefits of modeling of a complex nonlinear system, a neuroendocrine HPA system, were demonstrated. This goal was accomplished by employing stoichiometric models, which incorporate regulatory feedback loop mechanisms. The thesis also addressed possibilities for practical applications of such a modeling approach to predict dynamical states of HPA axis and cortisol concentration under different physiological conditions and due to influence of various perturbators, such as acute stress, chronic stress or glucocorticoid medications. The obtained results and subsequent conclusions indicate to the importance of certain dynamical states of HPA axis and cortisol concentration for maintaining homeostasis, as well as for the etiology of many metabolic and psychiatric dosorders (such as metabolic syndrome or major depression). The thesis also indicated to possibilities the modeling of neuroendocrine processes using stoichiometric models offers in order to aid the design of individually-tailored glucocorticoid therapy, thus contributing to further improvements in the field of personalized medicine.- Hipotalamo-hipofizno-adrenalna (eng. hypothalamic-pituitary-adrenal, HPA) osa predstavlja složen neuroendokrini sistem koji učestvuje u održavanju optimalnog dinamičkog stanja živog sistema (homeostaze). Bilo da je u osnovnom (bazalnom) ili nekom pobuđenom stanju pod dejstvom stresa, takav složeni nelinearni sistem je uvek daleko od termodinamičke ravnoteže i poseduje povratnu spregu. Zato hormone koji ga čine nalazimo u različitim dinamičkim
Inhibitors of cholesterol absorption have been sought for decades as a means to treat and prevent cardiovascular diseases associated with hypercholesterolemia. Ezetimibe is the one clear success ...story in this regard, and other compounds with similar efficacy continue to be sought. In the last decade, the laboratory mouse, with all its genetic power, has become the premier experimental model for discovering the mechanisms underlying cholesterol absorption and has become a critical tool for preclinical testing of potential pharmaceutical entities. This chapter briefly reviews the history of cholesterol absorption research and the various gene candidates that have come under consideration as drug targets. The most common and versatile method of measuring cholesterol absorption is described in detail along with important considerations when interpreting results, and an alternative method is also presented. In recent years, reverse cholesterol transport has become an area of intense new interest for drug discovery since this process is now considered another key to reducing cardiovascular disease risk. The ultimate measure of reverse cholesterol transport is sterol excretion and a detailed description is given for measuring neutral and acidic fecal sterols and interpreting the results.
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