Up-regulation of caveolin expression by cytotoxic agents in drug-sensitive cancer cells

Caveolin 1 expression is down-regulated in various cancer cell lines. Interestingly, in several drug-resistant cancer cells, a strong induction of caveolin 1 expression has been reported, suggesting a role for caveolin 1 in the acquisition and/or the maintenance of the multidrug-resistance phenotype. Here, we show, in drug-sensitive lung cancer cells (A549, Calu-6 or NCI-H69), that exposure to cytotoxic drugs (taxol, doxorubicin or etoposide) is sufficient to strongly up-regulate caveolin 1 and 2 protein levels. This up-regulation is sustained even 1 week after drug removal. Our results suggest that caveolin up-regulation is an early cellular response to a cytotoxic stress taking place before drug resistance

G2/M blockade by paclitaxel induces caveolin-1 expression in A549 lung cancer cells : caveolin-1 as a marker of cytotoxicity.

Caveolins are highly expressed in terminally differentiated cells, but this expression is down-regulated in various cancer cell lines. Exposure to low doses of paclitaxel (taxol) is sufficient to up-regulate caveolin-1, suggesting that a mild cytotoxic stress induces a response implying caveolin and caveolae. Here we show that this up-regulation is sustained even after the cessation of paclitaxel treatment. After exposure to a cytostatic dose of paclitaxel (50 nM), A549 lung cancer cells are blocked in the G2/M cell cycle phase. After removal of paclitaxel, cell death occurs, accompanied with an increase in caveolin expression, suggesting an effect of caveolin in this process. Three days post-paclitaxel treatment, surviving A549 cells were passaged and only a half of them adhered to the culture dish. Adhering cells (still mainly in the G2/M cell cycle phase) were still unable to grow and progressively entered in an apoptotic state. This study suggests that effects of a low dose of paclitaxel were still present even 1 week after drug removal and that caveolin-1 is a good marker of cytotoxicity

Aortic valve sclerosis in rabbits

BACKGROUND AND AIM OF THE STUDY: Aortic valve sclerosis is fairly common and is currently seen as a marker of systemic atherosclerosis. For unclear reasons only a minority of those sclerotic valves will evolve to become stenotic suggesting that atherogenic factors alone are insufficient to explain the development of valve stenosis. We had reported in a model of cholesterol fed rabbits that a combination of high cholesterol with vitamin D supplementation was necessary to induce valve stenosis and significant calcium deposition whereas high cholesterol alone only induced a sclerosis of the valve. In this study, we further evaluated the role of vitamin D treatment in the development of aortic valve disease (sclerosis or stenosis) in this rabbit model. METHODS: Rabbits were divided in 4 groups followed for 12 weeks: 1) no treatment; 2) cholesterol-enriched diet, 3) cholesterol-enriched diet + vitamin D2 (VD; 50000IU, daily) 4) VD alone for 12 weeks. Echocardiographic assessment of the aortic valve was done at baseline, and every 4 weeks thereafter. Aortic valve area, maximal and mean transvalvular gradients were recorded and compared over time. Immunohistological study of the valves of AS rabbits was also realized for several classical atherosclerosis markers. RESULTS: Vitamin D2 treated animal did not develop any stenosis of the valve despite increased echogenicity due to diffuse calcium deposits on the leaflets without any atherosclerotic lesions. Only the combination of high cholesterol with VD resulted in a decrease of aortic valve area. Immunohistological analysis of aortic valves from VD rabbits showed the presence of calcium deposits, T-cell infiltration in addition to positive labeling for alpha-smooth muscle cell actin. We did not observe macrophage infiltration in aortic valve leaflets of VD rabbits. CONCLUSION: Hypercholesterolemia or vitamin D supplements alone could not induce aortic valve stenosis in our animal model whereas the combination resulted in a decreased aortic valve area. These findings support the hypothesis that a combination of atherosclerotic and calcifying factors is necessary to induce aortic valve stenosis in this model

Carvedilol treatment for aortic valve regurgitation

Background — Aortic regurgitation (AR) is a chronic disease for which there is currently no approved medical treatment. We previously reported in an animal model that β-blockade with metoprolol exerted beneficial effects on left ventricular remodeling and survival. Despite the recent publication of promising human data, β-blockade in chronic AR remains controversial. More data are needed to support this potentially new treatment strategy. We hypothesized that carvedilol might be another safe treatment option in chronic AR, considering its combined β-blocking and α-blocking effects and proven efficacy in patients with established heart failure. Methods and Results — The effects of a 6-month treatment with carvedilol 30 mg/kg/d orally were evaluated in adult Wistar rats with severe AR. Sham-operated and untreated AR animals were used as controls. Carvedilol treatment resulted in less left ventricular hypertrophy and dilatation. Ejection fraction was improved and filling pressures were reduced by carvedilol. β1-Receptor expression was also improved as well as myocardial capillary density. Those beneficial effects were noted despite the presence of drug-induced bradycardia. Conclusions — Carvedilol exerted protective effects against volume-overload cardiomyopathy in this model of aortic valve regurgitation with preserved ejection fraction. These results suggest a protective class effect of β-blockers. Combined with the recent publication of promising human data, our findings support the need to carefully design a prospective study in humans to evaluate the effects of β-blockers in chronic aortic valve regurgitation

Spironolactone treatment for aortic valve regurgitation

BACKGROUND AND AIM OF THE STUDY : Aortic regurgitation (AR) is a disease for which there is currently no effective medical treatment. It has been shown previously in an experimental model of AR that the renin-angiotensin-aldosterone system (RAAS) plays a major role, and that medications blocking the RAAS are effective to protect against left ventricular (LV) hypertrophy and also help to maintain a normal systolic function. The role of aldosterone receptor blockers in this disease has never been evaluated. Thus, the effects were studied of the aldosterone receptor blocking agent spironolactone in a model of chronic AR in rats. METHODS : The effects of a six-month treatment with spironolactone were evaluated in adult Wistar rats with severe AR, compared to sham-operated and untreated AR animals. RESULTS : Spironolactone treatment decreased the total heart weight. In addition, the LV expression of atrial natriuretic peptide mRNA was decreased by spironolactone treatment, as was the expression of collagen 1 and LOX1 mRNAs. Left ventricular fibrosis was decreased by spironolactone treatment. CONCLUSION : Spironolactone protected against volume-overload cardiomyopathy in this model of aortic valve regurgitation. The predominant protective effect was a decrease in myocardial fibrosis

A Western high fat/high carbohydrate diet induces aortic valve disease in C57BL/6J mice

Objectives : The purpose of this study was to compare aortic valve function and morphology in adult wild-type (WT) mice and in low-density lipoprotein receptor-deficient (LDLr−/−) mice fed or not fed a high-fat/high-carbohydrate (HF/HC) diet. Background : Observations suggest a link between degenerative aortic valve stenosis (AS) and atherosclerosis. Aortic valve stenosis has been successfully induced in animal models of extreme hypercholesterolemia, but these models are less relevant to humans. It is not known if a proatherogenic HF/HC diet without added cholesterol could have the same negative impacts. Methods : Forty C57BL/6J mice were divided into four groups: WT + normal diet, WT + HF/HC diet, LDLr−/− with a normal diet, and LDLr−/− with a HF/HC diet. Aortic valve function and histology were evaluated by echocardiography after four months. Results : Wild-type mice on a HF/HC diet became mildly hypercholesterolemic, obese, and hyperglycemic. As expected, LDLr−/− mice became severely hypercholesterolemic. Both WT and LDLr−/− mice on a HF/HC diet displayed smaller valve areas and higher transvalvular velocities (p < 0.01) after four months. Aortic valve leaflets were thicker and infiltrated with lipids and macrophages in both HF/HC groups. Conclusions : A HF/HC diet in mice results in significant aortic valve abnormalities. Putting WT mice on a HF/HC diet reproduced a combination of atherogenic factors (obesity, mild dyslipidemia, and hyperglycemia) more commonly encountered in humans than isolated severe hypercholesterolemia. Severe hypercholesterolemia was not a prerequisite in our model. This experimental model suggests that AS development is multifactorial and that hypercholesterolemia should not be the only target in this disease

Chronic high-fat diet-induced obesity decreased survival and increased hypertrophy of 2 rats with experimental eccentric hypertrophy from chronic aortic regurgitation.

Background : The composition of a diet can influence myocardial metabolism and development of left ventricular hypertrophy (LVH). The impact of a high-fat diet in chronic left ventricular volume overload (VO) causing eccentric LVH is unknown. This study examined the effects of chronic ingestion of a high-fat diet in rats with chronic VO caused by severe aortic valve regurgitation (AR) on LVH, function and on myocardial energetics and survival. Methods : Male Wistar rats were divided in four groups: Shams on control or high-fat (HF) diet (15 rats/group) and AR rats fed with the same diets (ARC (n = 56) and ARHF (n = 32)). HF diet was started one week before AR induction and the protocol was stopped 30 weeks later. Results : As expected, AR caused significant LV dilation and hypertrophy and this was exacerbated in the ARHF group. Moreover, survival in the ARHF group was significantly decreased compared the ARC group. Although the sham animals on HF also developed significant obesity compared to those on control diet, this was not associated with heart hypertrophy. The HF diet in AR rats partially countered the expected shift in myocardial energy substrate preference usually observed in heart hypertrophy (from fatty acids towards glucose). Systolic function was decreased in AR rats but HF diet had no impact on this parameter. The response to HF diet of different fatty acid oxidation markers as well as the increase in glucose transporter-4 translocation to the plasma membrane compared to ARC was blunted in AR animals compared to those on control diet. Conclusions : HF diet for 30 weeks decreased survival of AR rats and worsened eccentric hypertrophy without affecting systolic function. The expected adaptation of myocardial energetics to volume-overload left ventricle hypertrophy in AR animals seemed to be impaired by the high-fat diet suggesting less metabolic flexibility

Fenofibrate and left ventricle remodeling in volume overload

Aims : Fenofibrate is a peroxisome proliferator-associated receptor alpha agonist (PPARα) used clinically for the management of dyslipidemia and is a myocardial fatty acid oxidation stimulator. It has also been shown to have cardiac anti-hypertrophic properties but the effects of fenofibrate on the development of eccentric LVH and ventricular function in chronic left ventricular (LV) volume overload (VO) are unknown. This study was therefore designed to explore the effects of fenofibrate treatment in a VO rat model caused by severe aortic valve regurgitation (AR) with a focus on cardiac remodeling and myocardial metabolism. Main methods : Male Wistar rats were divided in four groups (13–15 animals/group): Shams (S) treated with fenofibrate (F; 100 mg/kg/d PO) or not (C) and severe AR receiving or not fenofibrate. Treatment was started one week before surgery and the animals were sacrificed 9 weeks later. Key findings : AR rats developed severe LVH (increased LV weight) during the course of the protocol. Fenofibrate did not reduce LV weight. However, eccentric LV remodeling was strongly reduced by fenofibrate in AR animals. Fractional shortening was significantly less affected in ARF compared to ARC group. Fenofibrate also increased the myocardial enzymatic activity of enzymes associated with fatty acid oxidation while inhibiting glycolytic enzyme phosphofructokinase. Significance : Fenofibrate decreased LV eccentric remodeling associated with severe VO and helped maintain systolic function. Studies with a longer follow-up will be needed to assess the long-term effects of fenofibrate in chronic volume overload caused by aortic regurgitation

Interstitial cells from left-sided heart valves display more calcification potential than right-sided ones : an in vitro study of porcine valves

BACKGROUND AND AIM OF THE STUDY: The calcification of cardiac valves is more frequently observed on left-sided (aortic or mitral) than right-sided (pulmonic or tricuspid) valves. The cause of this preferential left-sided calcification remains relatively unknown. The study aim was to evaluate the capacity of interstitial cells isolated from the four cardiac valves of healthy adult pigs to calcify in culture. METHODS: Interstitial cells were isolated from the valve leaflets of three healthy young pigs and cultured in DMEM/fetal bovine serum (10%) in the presence or absence of osteogenic additives (ascorbic acid, dexamethasone, beta-glycerophosphate). RESULTS: The proliferation rate was similar for cells from each of the four valves. After longer periods of culture (> 10 days), cells from each valve spontaneously formed several calcification nodules, the process being accelerated in the presence of osteogenic additives (to 4-7 days). Alkaline phosphatase (AP) activity was highest in cells originating from the aortic and mitral valves, respectively, and least in those from the pulmonic and tricuspid valves. Culture with the osteogenic additives increased the AP activity by at least 50% for each valve, but the relative AP activity between cells from each valve origin tended to remain similar (aortic > mitral > pulmonic > tricuspid). Interestingly, the levels of matrix Gla-protein mRNA (an endogenous calcification inhibitor) followed an opposite trend of expression for each valve. CONCLUSION: Interstitial cells from porcine cardiac valves share similarities, although the capacity to calcify is more evident in cells from valves of the left side of the heart. Interstitial cells from the aortic valve displayed the greatest potential for calcification

Impact of anesthesia on echocardiographic evaluation of systolic and diastolic function in rats

Background : Echocardiography is used on rats but general anesthesia is usually necessary to be able to obtain a good quality echocardiogram. Each type of anesthetic agent has specific impacts on hemodynamics and, therefore, may affect differentially the echocardiographic measurements. Objectives : We sought to compare the echocardiograms of normal rats and rats with chronic aortic regurgitation under anesthesia using ketamine-xylazine or isoflurane. Methods : Animals underwent an echocardiogram with both drugs sequentially. Echocardiographic measurements were compared. Results : Mitral diastolic Doppler measurements (early diastolic filling wave [E] and late atrial diastolic filling wave [A] velocities) were significantly affected by the type of anesthesia in the normal group but not left ventricular dimensions or ejection fraction. Left ventricular dimensions were affected by the type of anesthesia in the aortic regurgitation group and diastolic Doppler flow. Conclusion : The anesthetic agent has significant specific impacts on many echocardiographic measurements. Investigators working with rat models should be aware of those potential effects