57 research outputs found
The importance of glucocorticoid delivery in the development of atherosclerosis
No full textElevated circulating glucocorticoids are causally linked to increased cardiovascular
disease. Atherosclerosis is a key cardiovascular pathology, yet the contribution of
glucocorticoids to underlying atherogenesis is unclear. Corticosteroid binding globulin
(CBG) regulates glucocorticoid action by controlling systemic bioavailability, with only
unbound ‘free’ glucocorticoids able to enter tissue and exert biological effects.
Cleavage of CBG by neutrophil elastase (NE) is proposed to enhance bioavailability
by reducing its affinity for glucocorticoids, elevating local ‘free’ glucocorticoid levels.
Strikingly, Apoe-/- mice lacking NE display reduced atherosclerosis. This thesis
addresses the hypothesis that CBG mediates atherogenesis by facilitating
glucocorticoid action at sites of plaque development.
First, the role of glucocorticoids and their regulation by CBG in atherogenesis was
established in an adenovirus-induced murine model of atherosclerosis. Male C57Bl6/J
mice were injected with recombinant adeno-associated viral vector serotype 8
expressing the gain-of-function mutation of mouse proprotein convertase
subtilisin/kexin type 9 (AAV8-PCSK9) and fed a Western diet (21% fat; 0.21%
cholesterol) for up to 18 weeks. Control mice were fed the same Western diet in the
absence of AAV8-PCSK9. Brachiocephalic lesion development in AAV8-PCSK9-
treated mice was accompanied by reduced hepatic Serpina6 (CBG) mRNA levels, in
parallel with reduced circulating CBG binding capacity. Moreover, AAV8-PCSK9
treatment increased circulating NE protein levels, suggesting the ability of CBG to bind
glucocorticoids may also be reduced in this model because of increased cleavage.
Surprisingly, AAV8-PCSK9 treatment did not alter circulating total glucocorticoid levels.
These data provide the first evidence of CBG dysregulation in this model of
atherosclerosis.
The increase in circulating NE levels in the AAV8-PCSK9 overexpression model of
atherosclerosis resembles that seen in Apoe⁻/⁻ mice, which are resistant to
atherosclerotic plaque development in the absence of NE. This suggests that NE may
also play a role in atherogenesis in the PCSK9 overexpression model. To test this,
male NE deficient mice (Ela⁻/⁻) and WT littermates were administered AAV8-PCSK9
and fed Western diet for 18 weeks. In contrast to published data from Apoe⁻/⁻/Ela-⁻/⁻
double-knockout mice, NE deficiency did not alter brachiocephalic lesion incidence,
size, or composition (collagen, smooth muscle cell, and macrophage content),
suggesting that NE does not play an important role in the development of lesions in
this model of atherosclerosis. Additionally, deletion of NE did not alter total or free
glucocorticoid levels in the circulation, indicating that NE-mediated CBG cleavage
does not occur in the systemic circulation.
The importance of CBG itself in regulating atherogenesis was explored next. To test
the hypothesis that deletion of CBG will reduce lesion size through reduced
glucocorticoid action, AAV8-PCSK9 was administered to mice lacking CBG (Cbg⁻/⁻)
mice and wild type littermates which then received Western diet for 18 weeks. Total
and free glucocorticoid levels were decreased in Cbg-/- mice. There was an apparent
decrease in incidence of lesion development in the brachiocephalic artery of Cbg⁻/⁻
mice compared with WT controls; however, this did not achieve statistical significance
(p=0.1312). This suggests that although CBG regulates the bioavailability of
glucocorticoids this does not play an important role in the development of
brachiocephalic lesions in the AAV8-PCSK9 model. Furthermore, method
development of mass-spectrometry imaging (MSI) was able to detect and quantify
local corticosterone in an aortic root lesion (n=1).
In conclusion, this thesis presents data demonstrating the first glucocorticoid profiling
of mice who develop inducible-atherosclerosis via AAV8-induced over-expression of
PCSK9. Despite the reported role of NE in lesion development in the Apoe⁻/⁻ model,
this thesis provides evidence that manipulation of CBG, either genetically or indirectly
via NE, does not alter the development of lesions in this model. This suggests that the
choice of model when investigating factors that influence atherogenesis should be
given careful consideration. Future work may involve investigating the effect of NE and
CBG deletion on alternative sites of lesion development, e.g., aortic root. Additionally,
while this work does not support a role for changes in systemic glucocorticoid
bioavailability in mediating plaque developing, advancement in steroid quantification
by MSI may allow assessment of local glucocorticoid levels within various regions of
atherosclerotic plaques to determine local glucocorticoid action
Vascular lesion development: influence of endogenous and exogenous glucocorticoids
No full textAtherosclerotic and restenotic lesions develop as a result of an excess inflammatory
response to vascular injury. Glucocorticoid hormones have widely-recognised
anti-inflammatory and anti-proliferative properties which appear to make them ideal
candidates for inhibition of vascular lesion development. Indeed, administration of
glucocorticoids to experimental animals does inhibit the growth of vascular lesions in
some models. In addition, glucocorticoids are currently being trialled clinically as
anti-restenotic agents. However, glucocorticoid excess in patients, either as a result
of Cushing’s syndrome or chronic steroid therapy, is associated with enhanced CVD
risk. Therefore, the effects of glucocorticoids on vascular lesion development remain
imperfectly understood.
The overall objective of these studies was to explore the influence of endogenous and
exogenous glucocorticoids on vascular lesion development using murine models of
atherosclerosis (ApoE-/- mice fed a “western” diet) and neointimal hyperplasia (wireinduced
femoral artery injury). The work described in this thesis addresses the
hypothesis that glucocorticoids are pro-atherogenic, yet anti-restenotic.
Mice were bilaterally adrenalectomised to investigate the role of endogenous
glucocorticoids on vascular lesion development. Removal of the adrenal glands had
no influence on atherogenesis or neointima development. The influence of
exogenous glucocorticoids on lesion development was assessed by orally
administering dexamethasone (0.1 or 0.8mg/kg/day). Atherosclerotic lesion burden
was augmented by dexamethasone administration. Conversely, fibro-proliferative
neointimal proliferation was inhibited by dexamethasone. However, this effect was
obscured by thrombotic lesion development. It was proposed that this thrombotic
effect is attributable to increased plasminogen activator inhibitor-1 (PAI-1), which
was tested using PAI-1 deficient mice. Although PAI-1 was found to mediate the
systemic pro-thrombotic effect of dexamethasone, it is not required for the enhanced
development of thrombotic lesions at the site of intra-luminal injury. These results suggest that physiological levels of endogenous glucocorticoids play a
limited role in vascular lesion development. Conversely, although exogenous
glucocorticoids inhibit fibro-proliferative intimal hyperplasia, they appear to have
significant detrimental influences on lesion development, augmenting atherosclerosis
and inducing thrombotic neointimal lesion formation following vascular injury.
Further research is therefore required to improve the cardiovascular outcome of
patients requiring glucocorticoid therapy and for the use of glucocorticoids as antirestenotic
agents
Non-invasive imaging of fibrosis with positron emission tomography in a rat model with systemic hypertension and myocardial fibrosis
No full textHeart failure is one of the leading causes of death worldwide. Hypertension can initiate myocardial remodelling processes which, often via fibrotic triggers through the renin-angiotensin-aldosterone system, can lead to the development of heart failure. A main contributor of these pathways is angiotensin II, increased levels of which can induce volume and pressure overload in the cardiovascular system, making it an important factor in both hypertension and associated cardiovascular disease. A main process during cardiac remodelling is fibrosis which can be divided into two types: reactive and replacement fibrosis. The latter refers to the changes via scar formation at an injury site while the former (interstitial or perivascular fibrosis) can happen as a response to changes in the physical or chemical environment within the tissue such as hypertension or inflammation. Fibrillary collagen is an important extracellular matrix component and abundantly deposited during fibrosis. Collagen can have various subtypes based on its structure which can add different characteristics to the tissue. During collagen biosynthesis, cis- or trans-proline containing pro-α chains can be integrated into the protein, where chains containing cis isomer are associated with more distensible and abnormal collagen and those with trans isomer with more rigid triple helix collagen. Other factors can also influence the development of heart failure via the myocardial remodelling processes, such as inflammatory and angiogenic pathways.
Heart failure can be diagnosed and assessed in the clinic via blood tests and imaging techniques such as ultrasound, magnetic resonance imaging (MRI), computerised tomography (CT), and single-photon emission computed tomography (SPECT) / positron emission tomography (PET). This thesis aimed to investigate the effect of increased angiotensin II and subsequent hypertension on the levels of myocardial collagen synthesis and to test PET radiotracers cis-4-18F-fluoro-L-proline and trans-4-18F-fluoro-L-proline for the detection of myocardial fibrosis and potential differentiation of the types of collagen fibers. The overarching hypothesis of the project was that myocardial fibrosis can be imaged non-invasively with PET in a rat pressure overload model with via persistent hypertension resulting in end-organ damage.
A hypertensive rat model with myocardial remodelling was established via angiotensin II infusion using osmotic mini-pumps. Treatment length and dosage were tested and the optimal protocol was chosen to induce myocardial fibrosis. The model was assessed for myocardial collagen content as well as markers of inflammation and vasculature. On a separate set of experiments, the performance of PET radiotracers, cis-4-18F-fluoro-L-proline and trans-4-18F-fluoro-L-proline, was assessed in naïve rats to understand their in vivo metabolism and kinetics. Then, the optimised animal model of hypertensive heart failure and PET imaging protocols were used to investigate whether the new imaging probes could visualise areas if increased collagen synthesis and whether the uptake was related to the type of collagen involved.
Using 500 ng/kg/min angiotensin II dose for 4 weeks duration was adequate to induce myocardial fibrosis and hypertension in the rat model. The fibrosis pattern was mainly perivascular in nature. Immunostaining also showed increased CD68 in the myocardium of rats on 250 ng/kg/min but not with the higher dose. The highest percentage of cells stained positive for all three of CD68, TSPO and isolectin B4 was found in the atria of animals on the higher angiotensin II dose, which area also showed the most fibrosis overall.
Both radiotracers were successfully assessed in naïve rats, showing no metabolism and favourable kinetics in vivo, allowing for simplified quantification of radiotracer uptake. Myocardial radiotracer uptake in the angiotensin II treated cohort showed increased myocardial signal with the trans-4-18F-fluoro-L-proline radiotracer but no significant differences with cis-4-18F-fluoro-L-proline compared to vehicle treated animals. Animals undergoing the imaging experiment showed increased myocardial fibrosis similarly to rats in the model development experiments.
Myocardial fibrosis develops during hypertension induced via angiotensin II treatment, and this change can be measured with PET imaging targeting collagen biosynthesis. Further investigation into the types of collagens involved and how they contribute to pathology needs to be carried out to characterise the underlying biological processes in more detail. Fluoroproline radiotracer PET imaging can become a valuable tool for the assessment of fibrosis pathology both in terms of early detection and disease progression
Vascular dysfunction in horses with endocrinopathic laminitis
No full text\ua9 2016 Morgan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Endocrinopathic laminitis (EL) is a vascular condition of the equine hoof resulting in severe lameness with both welfare and economic implications. EL occurs in association with equine metabolic syndrome and equine Cushing\u27s disease. Vascular dysfunction, most commonly due to endothelial dysfunction, is associated with cardiovascular risk in people with metabolic syndrome and Cushing\u27s syndrome. We tested the hypothesis that horses with EL have vascular, specifically endothelial, dysfunction. Healthy horses (n = 6) and horses with EL (n = 6) destined for euthanasia were recruited. We studied vessels from the hooves (laminar artery, laminar vein) and the facial skin (facial skin arteries) by small vessel wire myography. The response to vasoconstrictors phenylephrine (10-9-10-5M) and 5-hydroxytryptamine (5HT; 10-9-10-5M) and the vasodilator acetylcholine (10-9-10-5M) was determined. In comparison with healthy controls, acetylcholine-induced relaxation was dramatically reduced in all intact vessels from horses with EL (% relaxation of healthy laminar arteries 323.5 \ub1 94.1% v EL 90.8 \ub1 4.4%, P = 0.01, laminar veins 129.4 \ub1 14.8% v EL 71.2 \ub1 4.1%, P = 0.005 and facial skin arteries 182.0 \ub1 40.7% v EL 91.4 \ub1 4.5%, P = 0.01). In addition, contractile responses to phenylephrine and 5HT were increased in intact laminar veins from horses with EL compared with healthy horses; these differences were endothelium-independent. Sensitivity to phenylephrine was reduced in intact laminar arteries (P = 0.006) and veins (P = 0.009) from horses with EL. Horses with EL exhibit significant vascular dysfunction in laminar vessels and in facial skin arteries. The systemic nature of the abnormalities suggest this dysfunction is associated with the underlying endocrinopathy and not local changes to the hoof
The role of androgen receptors in atherosclerosis
No full textMale disadvantage in cardiovascular health is well recognised. However, the influence of androgens on atherosclerosis, one of the major causes of many life-threatening cardiovascular events, is not well understood. With the dramatic increase in clinical prescription of testosterone in the past decade, concerns about the cardiovascular side-effects of androgen supplementation or androgen deprivation therapy are increasing. Potential atheroprotective effects of testosterone could be secondary to (aromatase-mediated) conversion into oestradiol or, alternatively, to direct activation of androgen receptors (AR). Recent development of animal models with cell-specific AR knockout has indicated a complex role for androgen action in atherosclerosis. Most studies suggest androgens are atheroprotective but the precise role of AR remains unclear. Increased use of AR knockout models should clarify the role of AR in atherogenesis and, thus, lead to exploitation of this pathway as a therapeutic target
Sertoli cells modulate testicular vascular network development, structure, and function to influence circulating testosterone concentrations in adult male mice
No full textThe testicular vasculature forms a complex network, providing oxygenation, micronutrients, and waste clearance from the testis. The vasculature is also instrumental to testis function because it is both the route by which gonadotropins are delivered to the testis and by which T is transported away to target organs. Whether Sertoli cells play a role in regulating the testicular vasculature in postnatal life has never been unequivocally demonstrated. In this study we used models of acute Sertoli cell ablation and acute germ cell ablation to address whether Sertoli cells actively influence vascular structure and function in the adult testis. Our findings suggest that Sertoli cells play a key role in supporting the structure of the testicular vasculature. Ablating Sertoli cells (and germ cells) or germ cells alone results in a similar reduction in testis size, yet only the specific loss of Sertoli cells leads to a reduction in total intratesticular vascular volume, the number of vascular branches, and the numbers of small microvessels; loss of germ cells alone has no effect on the testicular vasculature. These perturbations to the testicular vasculature leads to a reduction in fluid exchange between the vasculature and testicular interstitium, which reduces gonadotropin-stimulated circulating T concentrations, indicative of reduced Leydig cell stimulation and/or reduced secretion of T into the vasculature. These findings describe a new paradigm by which the transport of hormones and other factors into and out of the testis may be influenced by Sertoli cells and highlights these cells as potential targets for enhancing this endocrine relationship.Full Tex
In Vivo Assays for Assessing the Role of the Wilms’ Tumor Suppressor 1 (Wt1) in Angiogenesis
No full textEstrogens protect male mice from obesity complications and influence glucocorticoid metabolism
No full text\ua9 2015 Macmillan Publishers Limited. Background:Although the prevalence of obesity is higher among women than men, they are somewhat protected from the associated cardiometabolic consequences. The increase in cardiovascular disease risk seen after the menopause suggests a role for estrogens. There is also growing evidence for the importance of estrogen on body fat and metabolism in males. We hypothesized that that estrogen administration would ameliorate the adverse effects of obesity on metabolic parameters in males.Methods:Male and female C57Bl/6 mice were fed control or obesogenic (DIO) diets from 5 weeks of age until adulthood. Glucose tolerance testing was performed at 13 weeks of age. Mice were killed at 15 weeks of age and liver and adipose tissue were collected for analysis of gene expression. A second cohort of male mice underwent the same experimental design with the addition of estradiol pellet implantation or sham surgery at 6 weeks.Results:DIO males had greater mesenteric adipose deposition and more severe increases in plasma glucose, insulin and lipids than females. Treatment of males with estradiol from 6 weeks of age prevented DIO-induced increases in adipose tissue mass and alterations in glucose-insulin homeostasis. We also identified sex differences in the transcript levels and activity of hepatic and adipose glucocorticoid metabolizing enzymes. Estrogen treatment feminized the pattern of DIO-induced changes in glucocorticoid metabolism, rendering males similar to females.Conclusions:Thus, DIO induces sex-specific changes in glucose-insulin homeostasis, which are ameliorated in males treated with estrogen, highlighting the importance of sex steroids in metabolism. Given that altered peripheral glucocorticoid metabolism has been observed in rodent and human obesity, our results also suggest that sexually dimorphic expression and activity of glucocorticoid metabolizing enzymes may have a role in the differential metabolic responses to obesity in males and females
Altered vascular contractility in adult female rats with hypertension programmed by prenatal glucocorticoid exposure
No full textSafer topical treatment for inflammation using 5α-tetrahydrocorticosterone in mouse models
No full text\ua9 2017 The Author(s) Use of topical glucocorticoid for inflammatory skin conditions is limited by systemic and local side-effects. This investigation addressed the hypothesis that topical 5α-tetrahydrocorticosterone (5αTHB, a corticosterone metabolite) inhibits dermal inflammation without affecting processes responsible for skin thinning and impaired wound healing. The topical anti-inflammatory properties of 5αTHB were compared with those of corticosterone in C57Bl/6 male mice with irritant dermatitis induced by croton oil, whereas its effects on angiogenesis, inflammation, and collagen deposition were investigated by subcutaneous sponge implantation. 5αTHB decreased dermal swelling and total cell infiltration associated with dermatitis similarly to corticosterone after 24 h, although at a five fold higher dose, but in contrast did not have any effects after 6 h. Pre-treatment with the glucocorticoid receptor antagonist RU486 attenuated the effect of corticosterone on swelling at 24 h, but not that of 5αTHB. After 24 h 5αTHB reduced myeloperoxidase activity (representative of neutrophil infiltration) to a greater extent than corticosterone. At equipotent anti-inflammatory doses 5αTHB suppressed angiogenesis to a limited extent, unlike corticosterone which substantially decreased angiogenesis compared to vehicle. Furthermore, 5αTHB reduced only endothelial cell recruitment in sponges whereas corticosterone also inhibited smooth muscle cell recruitment and decreased transcripts of angiogenic and inflammatory genes. Strikingly, corticosterone, but not 5αTHB, reduced collagen deposition. However, both 5αTHB and corticosterone attenuated macrophage infiltration into sponges. In conclusion, 5αTHB displays the profile of a safer topical anti-inflammatory compound. With limited effects on angiogenesis and extracellular matrix, it is less likely to impair wound healing or cause skin thinning
- …
