Application of HBOCs electrophoretic method to detect a new blood substitute derived from the giant extracellular haemoglobin of lugworm.

Manipulation of blood and blood components is prohibited in sports by the World Anti-Doping Agency (WADA). This includes the use of blood substitutes to increase oxygen transport, like haemoglobin-based oxygen carriers (HBOCs), which are compounds derived from haemoglobin. Despite their medical interest, the first generation of HBOCs had serious adverse effects and was abandoned. However, new studies are now exploiting the properties of marine worm haemoglobins, which circulate as giant extracellular complexes with high oxygen-binding capacities. HEMOXYCarrier® (HC), developed by Hemarina, is one of the most advanced and promising HBOCs, and HC may become a tempting doping tool for athletes in the future. Here, HC detection in plasma/serum was evaluated with the method used to detect the first HBOCs, based on electrophoresis and heme peroxidase properties. An HC-derived product was identified in human plasma up to 72 h after in vitro incubation at 37 °C. HC degradation also induced methemalbumin formation. After injecting HC at the effective dose of 200 mg/kg into mice, the HC-derived product was detected only for a few hours and no accumulation of methemalbumin was observed. Due to this limited detection window in vivo, measuring specific worm globin degradation products by mass spectrometry might be an alternative for future anti-doping analyses. Copyright © 2016 John Wiley & Sons, Ltd.

Beclin 1 and autophagy are required for the tumorigenicity of breast cancer stem-like/progenitor cells.

Malignant breast tissue contains a rare population of multi-potent cells with the capacity to self-renew; these cells are known as cancer stem-like cells (CSCs) or tumor-initiating cells. Primitive mammary CSCs/progenitor cells can be propagated in culture as floating spherical colonies termed 'mammospheres'. We show here that the expression of the autophagy protein Beclin 1 is higher in mammospheres established from human breast cancers or breast cancer cell lines (MCF-7 and BT474) than in the parental adherent cells. As a result, autophagic flux is more robust in mammospheres. We observed that basal and starvation-induced autophagy flux is also higher in aldehyde dehydrogenase 1-positive (ALDH1(+)) population derived from mammospheres than in the bulk population. Beclin 1 is critical for CSC maintenance and tumor development in nude mice, whereas its expression limits the development of tumors not enriched with breast CSCs/progenitor cells. We found that decreased survival in autophagy-deficient cells (MCF-7 Atg7 knockdown cells) during detachment does not contribute to an ultimate deficiency in mammosphere formation. This study demonstrates that a prosurvival autophagic pathway is critical for CSC maintenance, and that Beclin 1 plays a dual role in tumor development.

[Leprosy in French Guyana: a retrospective study from 1997 to 2006]. / La lèpre en Guyane française: étude rétrospective de 1997 à 2006.

French Guyana borders Brazil with the second highest number of cases of leprosy in the world. The purpose of this retrospective study of leprosy cases diagnosed in Guyana between January 1997 and December 2006 was to calculate the incidence of the disease and to identify any special clinical and epidemiological features. A total of 90 new cases were recorded during the study period for a mean incidence of 0.53 cases/10,000 inhabitants/year. Since this incidence is below the 1/10,000 threshold defined by the World Health Organization, leprosy is no longer considered as a major public health issue in French Guyana. However it must be noted that while the number of "native leprosy" cases has declined, the number of Brazilian cases has increased (p<0.01). Brazilian leprosy has different epidemiological features, i.e., dominance of multibacillary forms and high incidence in gold panning and western regions of the country where the incidence is over 1/10,000 inhabitants/year.

[Cutaneous, pulmonary and bone aspergillosis in a patient presumed immunocompetent presenting subacute cutaneous lupus erythematosus]. / Aspergillose cutanée, pulmonaire et osseuse chez un malade ayant un lupus érythémateux cutané subaigu sans déficit immunitaire associé.

INTRODUCTION: We report a case of cutaneous, pulmonary and bone aspergillosis successfully treated after many years of progression in a patient presumed immunocompetent presenting subacute cutaneous lupus erythematosus. CASE-REPORT: A 43-year-old man, treated with thalidomide for subacute cutaneous lupus erythematosus, presented chest pain with haemoptysis and dyspnea. A pulmonary nodule was detected but the microbiological investigation was negative. The histological examination showed granuloma with round structures. No cause was found. Three years later, skin lesions appeared on the patient's face concomitantly with a pulmonary relapse. Histopathological examination of these lesions demonstrated septate hyphae. Aspergillus fumigatus was isolated in skin and lung. Disseminated aspergillosis was then diagnosed as spondylodiscitis developed. Treatment with combined voriconazole and caspofungin produced significant and rapid improvement of lesions. DISCUSSION: While aspergillosis is commonly seen in immunocompetent patients, angiotropic dissemination points to cellular immunodepression. Our patient, however, was not presenting immunodepression. We discuss the possible contributory role of thalidomide in dissemination of aspergillosis given that the literature to date contains only one reported case of cutaneous aspergillosis secondary to A. fumigatus in an immunocompetent patient. We would also point out the specific histopathological pattern of this disseminated aspergillosis with both septate hyphae and round structures. Invasive aspergillosis is highly lethal but the chances of recovery are now greater thanks to new antifungal agents.

Combined angiotensin II AT1-receptor blockade and angiotensin I-converting enzyme inhibition on survival and cardiac remodeling in chronic heart failure in rats.

BACKGROUND: Angiotensin I-converting enzyme inhibition (ACEI) and angiotensin II AT(1)-receptor blockade are effective at improving survival and limiting cardiac remodeling in the rat model of postischemic heart failure. Whether their combination yields additive/synergistic effects is unknown. METHODS AND RESULTS: Rats underwent coronary artery ligation and 7 days later were treated orally for 9 months with placebo (controls), 5 mg/kg valsartan, 1 mg/kg enalapril (doses submaximally effective at reducing mortality in the experimental model used), or 5 mg/kg valsartan and 1 mg/kg enalapril combined. Compared with controls, valsartan, enalapril, and their combination decreased mortality by 40% (P =.006), 21% (P =.065), and 33% (P =.032), respectively, but there was no significant difference between the 3 treatments. At the doses used, valsartan, but neither enalapril nor the combination, slightly limited cardiac hypertrophy and fibrosis development and reduced left ventricular end-diastolic pressure as assessed in the surviving animals at 9 months. CONCLUSIONS: In experimental chronic heart failure in rats, valsartan reduces mortality similar to other AT(1)-receptor blockers and a combination of AT(1)-receptor blockade (valsartan) and ACEI (enalapril) at submaximal doses does not exert additive/synergistic beneficial effects on mortality.

Coronary dilatation reserve in experimental hypertension and chronic heart failure: effects of blockade of the renin-angiotensin system.

1. The aim of the present study was to investigate left and right ventricular (LV and RV, respectively) coronary vasodilatation reserve (CVR; fluorescent microsphere technique) in rats with hypertension (spontaneously hypertensive rats (SHR)) or congestive heart failure (CHF) and the effects of early and chronic renin-angiotensin system (RAS) blockade thereupon. 2. In adult SHR, both LV and RV CVR were impaired, especially in the non-hypertrophied RV, the main factor involved being coronary vascular remodelling. Blockade of the RAS normalized both LV and RV CVR, mainly through the prevention of hypertension and suppression of the resulting pericoronary fibrosis. 3. In postischaemic CHF rats, there was an early and severe degradation of LV and RV CVR that developed before any significant vascular remodelling and appeared to be linked to the deterioration of cardiac hypertrophy and haemodynamics. This degradation in CVR further worsened over the longer term due to late-developing pericoronary fibrosis and endothelial dysfunction. Blockade of the RAS had no early effects on LV and RV CVR, but improved RV CVR over the long term, mainly by limiting RV hypertrophy and by preventing the development of pericoronary fibrosis and coronary endothelial dysfunction. 4. In kallikrein-kinin system-deficient mice, CVR was not different from that of wild-type mice, suggesting that this system is not implicated in normal CVR regulation.

Fluospheres for cardiovascular phenotyping genetically modified mice.

Assessment of systemic and regional hemodynamic phenotypes in genetically engineered mice by nonradioactive methods is yet an unsolved problem. We therefore investigated whether the reference sample method using fluorescent microspheres (FMs), already validated in rats, might be used for this purpose in C57BL/6 and in apolipoprotein E (ApoE)-deficient mice. FMs were injected into the left ventricle of instrumented anesthetized mice. In 10-week-old C57BL/6, cardiac output was 18-19 ml/min, and its regional distribution under basal conditions was approximately 1.5% (brain), 3.5% (heart), 9. 1% (left kidney), 9.8% (right kidney), 1% (spleen), and 0.8% (stomach) (i.e., values similar to those previously reported with radioactive microspheres). Proper mixing of FMs was achieved as both kidneys had identical flows; distribution of two differently labeled FMs injected simultaneously was shown to be identical by an agreement study, and FM trapping in the capillary bed was demonstrated both histologically and by the recovery in the lungs of 90% of intravenously injected FMs. In addition, identical values for cardiac output and its distribution were obtained in different age-matched groups of C57BL/6. The FM technique also proved to be able to evidence angiotensin II and isoprenaline classic systemic and regional hemodynamic effects. Finally, applied to 30-week-old ApoE-deficient mice and age-matched C57BL/6, the FM technique showed no major hemodynamic difference between the two groups, except for coronary blood flow, which was significantly decreased in ApoE-deficient mice. In conclusion, we demonstrated for the first time the feasibility, accuracy, and reliability of the FM technique at characterizing the cardiovascular phenotype of genetically engineered mice.

Effects of long-term angiotensin II AT1 receptor blockade on survival, hemodynamics and cardiac remodeling in chronic heart failure in rats.

OBJECTIVE: The beneficial effect of chronic angiotensin I converting enzyme (ACE) inhibition on survival has for long been established in the rat post-infarction model of chronic heart failure (CHF) and has subsequently been confirmed in humans. This study investigates in rats whether chronic angiotensin II AT1 receptor blockade shares with ACE inhibition the same beneficial effect. METHODS: Rats we subjected to coronary artery ligation and, from 7 days later, orally treated for 7.5 months with placebo or irbesartan (5 or 50 mg/kg/day). RESULTS: Irbesartan dose-dependently increased survival (placebo: 27%, low dose: 52%, high dose: 82%, sham-ligated: 100%; high dose vs placebo: P < 0.001 and vs low dose: P < 0.05; low dose vs placebo: P = 0.11). Irbesartan also dose-dependently decreased urinary cyclic GMP excretion throughout the study. At 7.5 months, it dose-dependently decreased left ventricular (LV) end diastolic pressure. normalized LV pressure maximal rate of rise (dP/dt) and cardiac index values and improved LV and right ventricular regional blood flows (radioactive microspheres) and resistances. At 7.5 months, irbesartan markedly decreased myocardial hypertrophy but had almost no effect on LV dilatation and subendocardial fibrosis. CONCLUSIONS: Long-term angiotensin II AT1 receptor blockade with irbesartan strongly and dose-dependently increases survival in the rat model of coronary ligation-induced CHF. This effect is due to the combination of the beneficial effects that the drug exerts on systemic and coronary hemodynamics, on cardiac pump function and vs cardiac hypertrophy development. Long-term AT1 receptor blockade might thus prove useful and prolong survival in human CHF.

Systemic and regional hemodynamics assessment in rats with fluorescent microspheres.

The aim of this study was to validate in rats an alternative to the radioactive microspheres (RM) reference technique, the fluorescent microspheres (FM), for the simultaneous determination of cardiac index (CI) and regional blood flows (RBF). Validation of the FM method was performed in three steps: (a) comparison of CI and RBF values obtained simultaneously by FM and RM, (b) determination of the repeatability of the measurements by using two successive injections of FM, and (c) evaluation of the ability of the FM method to assess vasodilating effects (by using dipyridamole). CI values (range, 242-513 ml/min x kg; n = 20) obtained with FM correlated with those obtained with RM (r = 0.82: p<0.001), and agreement was found between FM and RM (error 95% confidence interval for one pair, +/-125 ml/min x kg). FM RBF values, although smaller than corresponding RM RBF values, were correlated with the latter (range, 0.1-7 ml/min x g; n = 71; r = 0.99; p< 0.001). Agreement was dependent on RBF values, e.g., error 95% confidence intervals for one pair were 0.08-0.13 and 3.86-6.48 for 0.1 and 5 ml/min x g, respectively. Two successive FM injections at a 10-min interval (conscious rats) provided similar values (n = 14) for CI (306+/-24 vs. 346+/- 18 ml/min x kg), and renal (5.1+/-0.2 vs. 6.2+/-0.3 ml/min x g), left (6.1+/-0.3 vs. 5.8+/-0.4 ml/min x g) and right (4.8+/-0.4 vs. 4.7+/-0.3 ml/min x g) myocardial RBF. Corresponding error 95% confidence intervals were +/-187 ml/min x kg, +/-2.8, +/-2.2, and +/-2.0 ml/min x g, respectively. Dipyridamole (2, 4, 6, and 8 mg/kg x min for 10 min, i.v.; n = 9-13 per group, conscious rats) significantly and dose dependently increased left and right myocardial blood flows, whereas renal blood flow was not affected. We conclude that the FM technique (a) is reliable and in agreement with the RM method, (b) provides repeatable measurements of systemic and regional hemodynamics, and (c) allows detection and quantification of vasodilating effects in rats.

Involvement of nitric oxide, but not prostaglandins, in the vascular sympathoinhibitory effects of losartan in the pithed spontaneously hypertensive rat.

1. The aim of this study was to investigate whether nitric oxide (NO) and/or vasodilator prostaglandins (PGs) are involved in the sympathoinhibitory effects exerted by losartan versus the vascular responses elicited by spinal cord electrical stimulation (SCS) in pithed spontaneously hypertensive rats (SHRs). 2. SHRs were given orally and for 8 days either losartan (10 mg kg-1 daily) or distilled water (controls). After pithing, blood pressure, heart rate, cardiac output, renal and muscular blood flows (pulsed Doppler technique) and the corresponding vascular resistance values were measured or calculated at baseline. Then, animals from both groups were given i.v. either saline, or NG-nitro-L-arginine methyl ester (L-NAME, 1 mg kg-1), or diclofenac (4 mg kg-1). Thereafter, haemodynamic parameters were determined in the six subgroups of animals in response (a) to SCS at increasing frequencies, and (b) to a noradrenaline bolus injection. 3. Losartan significantly decreased mean arterial pressure as well as renal and total peripheral resistances. In addition, losartan exhibited strong vascular sympathoinhibitory effects, significantly decreasing the systemic pressor and regional vasoconstrictor responses to SCS, but did not affect those to exogenous noradrenaline. In contrast, SCS-induced tachycardia was not modified by losartan. 4. L-NAME significantly increased total peripheral and regional vascular resistances but did not affect blood pressure and heart rate basal values. L-NAME potentiated the haemodynamic responses to SCS in control and, to a larger extent, in losartan-treated SHRs so that, with the exception of the renal vascular bed, the sympathoinhibitory effects of losartan were attenuated in all vascular beds studied. L-Arginine (300 mg kg-1) caused reversal of L-NAME effects in both control and losartan-treated SHRs. 5. Diclofenac did not affect the basal values of haemodynamic parameters in control and losartan-treated SHRs. Diclofenac potentiated the pressor and vasoconstrictor responses to SCS and to a similar extent, in both control and losartan-treated SHRs, so that the sympathoinhibitory effects of losartan were fully maintained. 6. These results demonstrate that in pithed SHRs: (a) NO but not PGs contribute to the basal vasomotor tone, (b) both NO and PGs attenuate the pressor and vasoconstrictor responses to SCS, (c) NO plays a major role in the vascular sympathoinhibitory effects of losartan, except at the renal level, and (d) endogenous PGs are not involved in these sympathoinhibitory effects.

Quinapril prevents stroke both during and after the treatment period in stroke-prone spontaneously hypertensive rats.

The effects of long-term oral administration of quinapril on the occurrence of stroke and on mortality were investigated in young salt-loaded stroke-prone spontaneously hypertensive rats (SHR-SPs) during the treatment period (8th-34th week of age) and up to 6 weeks thereafter. Simultaneously, blood pressure, saline intake, diuresis, and proteinuria were investigated at regular intervals, and cerebrovascular, renal, and cardiac lesions were assessed after death. Untreated SHR-SPs served as controls. Quinapril completely suppressed stroke and mortality, afforded only limited protection v blood pressure rise, and prevented any increase in saline intake, diuresis, and proteinuria both during and after the treatment period. Quinapril long-lastingly prevented vascular fibrinoid necrosis development at the cerebral, but also at the renal and cardiac levels. In the kidneys, vascular intimal and medial hyperplasia were strongly reduced, as were the glomerular and tubulo-interstitial lesions. At the cardiac level, intimal and medial hyperplasia were slightly reduced but infarction and fibrosis were hardly affected. As the renin-angiotensin system is highly stimulated in SHR-SPs and as angiotensin II (AII) is responsible for fibrinoid necrosis formation, vessel obstruction, and stroke in these animals, we conclude that the long-lasting protection afforded by quinapril v stroke and mortality in SHR-SPs both during and after the treatment period is mostly due to the drug-induced interruption of the renin-angiotensin system.(ABSTRACT TRUNCATED AT 250 WORDS)

Beneficial effects of trandolapril on experimentally induced congestive heart failure in rats.

Angiotensin-converting enzyme (ACE) inhibitors have been shown to prolong life expectancy in patients with congestive heart failure. In order to determine the relative contributions of the different factors involved in this beneficial effect, we investigated in an experimental model of postinfarction cardiac insufficiency in the rat over a 9-12-month period (1) the kinetics of the development of the hemodynamic, biologic, and morphologic alterations that accompany heart failure, and (2) the kinetics of the effects of a new, long-acting ACE inhibitor, trandolapril. Following induction of infarction, systolic blood pressure, left ventricular dP/dt, and end-diastolic pressure were immediately decreased, decreased, and increased, respectively, and these modifications persisted throughout the study. Cardiac index, on the other hand, was only initially and transiently decreased. Cardiac remodeling (left ventricular dilation, myocardial hypertrophy, and fibrosis) occurred as early as 7 days after infarction and worsened throughout the study. Plasma atrial natriuretic factor (ANF) and urinary cyclic guanosine monophosphate (cGMP) were also increased. In this model, a 1-year oral treatment with trandolapril resulted in early hemodynamic and biologic beneficial effects (reductions in pre- and afterload, increase in cardiac index, and decrease in plasma ANF), and in a delayed reversal of the infarction-induced cardiac morphologic alterations. Hence, the trandolapril-induced increase in survival rate is due initially to the drug's hemodynamic effects and over the long-term to both its hemodynamic and cardiac morphologic (limitation of remodeling) effects.

Long-term treatment with trandolapril opposes cardiac remodeling and prolongs survival after myocardial infarction in rats.

Long-term treatment with angiotensin I converting enzyme inhibitors (ACEIs) prolongs survival in rats developing congestive heart failure after myocardial infarction (MI). In this experimental model, we investigated at regular intervals the effects of a 1-year oral treatment with trandolapril, a new ACEI, on (a) survival rate and duration and (b) hemodynamic, biological, and cardiac and vascular histomorphological parameters. Control MI rats and sham-operated trandolapril-treated and control rats were simultaneously studied. In MI rats, trandolapril significantly increased the survival rate and duration, increasing the life expectancy by approximately 6 months. It also significantly decreased the arterial blood pressure and increased diuresis. These effects occurred as soon as the treatment was started and persisted throughout the study. Trandolapril significantly limited the development of myocardial hypertrophy, decreasing the heart weight and left ventricular hypertrophic area and increasing the left ventricular myocyte nuclear density, but these effects, starting after 3-6 months, were delayed compared to the hemodynamic ones. Finally, trandolapril limited the development of myocardial (both subendocardial and subepicardial) and aortic fibrosis. It is concluded that the early pre- and afterload effects of trandolapril are initially responsible for its beneficial action on survival, whereas later the drug's antihypertrophic and antifibrotic properties together with persistent hemodynamic effects account for the prolonged survival improvement.