Targeting Apoptosis Signal-Regulating Kinase-1 (ASK-1) As a Biomarker of Monocrotaline-Induced Pulmonary Hypertension following Administration of Antiretroviral Medications in Rat Model.

BACKGROUND: Apoptosis resistance is a recognized pathogenetic mechanism in pulmonary hypertension. However, the link between apoptosis signal-regulating kinase-1 (ASK-1) and pulmonary hypertension (PH) is unclear. This study was conducted to elucidate ASK-1 as a potential biomarker in PH. The study aimed to identify the role of ASK-1 in the mechanism of monocrotaline-induced PH in rats. METHODS: Forty adult male Sprague-Dawley rats (body weight: 200-250 g) were randomly divided into five groups (n=8 per group). The four treatment groups received a single intraperitoneal injection of monocrotaline (MCT) at a dose of 60 mg. kg-1 while the control group received an equivalent volume of intraperitoneal saline injection. Zidovudine (100mg. kg-1), ritonavir (30mg. kg-1), or combination of both drugs (zidovudine 100mg. kg-1 and ritonavir 30mg. kg-1) were administrated daily for the study period of 28 days to the rats in three of the four treatment groups with MCT for 28 days. On the twenty-eighth day of the study, rats were sacrificed, and organ harvested with the heart analyzed using RT-PCR for ASK-1. Antioxidant enzyme activities were determined using the colorimetric method. RESULTS: Animal survival rate was one hundred percent in the treated and control groups while the untreated group recorded 62% survival rate. There was significantly lower mRNA gene expression of ASK-1 in the heart tissues of the treated rats with zidovudine (2.67 ± 0.09, p < 0.0001), ritonavir (2.57 ±0.11, p < 0.0001) and a combination of both (2.75 ± 0.06, p < 0.0001) when compared to rats in the untreated group. An overexpressed mRNA gene of ASK-1 in the untreated rats was observed (12.0 ± 0.90, p < 0.0001) when compared to the controls. CONCLUSION: ASK-1 is a veritable biomarker for anti-apoptotic characteristics of PH. Our findings will spur new investigations on the role of ASK-1 in PH and the potential therapeutic benefits of antiretroviral medications in the prevention of PH. CONTEXTE: La résistance à l'apoptose est une pathogénétique reconnue mécanisme dans l'hypertension pulmonaire. Cependant, le lien entrekinase-1 régulatrice du signal d'apoptose (ASK-1) et pulmonaire l'hypertension (HTP) n'est pas claire. La présente étude a été menée pour :élucider ASK-1 comme biomarqueur potentiel de l'HTP. L'étude visait à :identifier le rôle de l'ASK-1 dans le mécanisme induit par la monocrotalinePH chez le rat. MÉTHODES: Quarante rats Sprague-Dawley mâles adultes (poids corporel:200 à 250 g) ont été divisés au hasard en cinq groupes (n = 8 par groupe).Les quatre groupes de traitement ont reçu une seule injection intrapéritonéalede monocrotaline (TCM) à une dose de 60 mg. kg­1 pendant que le témoina reçu un volume équivalent d'injection intrapéritonéale de solution saline.Zidovudine (100 mg kg­1), ritonavir (30 mg kg­1) ou combinaison deles deux médicaments (zidovudine 100 mg. Kg­1 et ritonavir 30 mg. kg­1) étaient administré quotidiennement pendant la période d'étude de 28 jours aux rats dans trois des quatre groupes de traitement avec MCT pendant 28 jours. Sur levingt-huitième jour de l'étude, des rats ont été sacrifiés et des organesrécolté avec le cœur analysé à l'aide de rt-PCR pour ASK-1. Les activités enzymatiques antioxydantes ont été déterminées à l'aide de la colorimétrieméthode. RÉSULTATS: Le taux de survie des animaux était de cent pour cent dans les groupes traités et témoins tandis que le groupe non traité a enregistré 62 %taux de survie. L'expression des gènes de l'ARNm était significativement plus faible d'ASK-1 dans les tissus cardiaques des rats traités par la zidovudine (2.67 ± 0.09, p < 0.0001), ritonavir (2.57 ±0.11, p < 0.0001) et acombinaison des deux (2.75 ± 0.06, p < 0.0001) par rapport aux rats dans le groupe non traité. Un gène d'ARNm surexprimé d'ASK-1 dans les rats non traités ont été observés (12.0 ± 0.90, p < 0.0001) lorsque par rapport aux contrôles. CONCLUSION: ASK-1 est un véritable biomarqueur antiapoptotique caractéristiques du pH. Nos conclusions donneront lieu à de nouvelles enquêtes sur le rôle de l'ASK-1 dans l'HTP et les avantages thérapeutiques potentiels demédicaments antirétroviraux dans la prévention de l'HTP. Mots-clés: Hypertension pulmonaire, régulation du signal d'apoptosekinase 1 (ASK-1), zidovudine, ritonavir, VIH/SIDA.

Trematodal granulomatous uveitis in paediatric Egyptian patients: a case series.

PURPOSE: To describe the clinical presentations and results of laboratory analysis of waterborne ophthalmic granulomas of the anterior chamber (AC) in Egyptian patients. PARTICIPANTS: 110 patients with granulomatous anterior uveitis and distinctive AC nodules. DESIGN: Prospective, non-comparative, case series. METHODS: Demographic data including age, gender and place of residence were recorded. A full ophthalmic examination with emphasis on the inflammatory characteristics and systemic workup was performed. The nodules were surgically removed in selected cases and molecular and histopathological analyses were performed. RESULTS: 102 boys and 8 girls were recruited (mean age 11.5 years). All children came from villages along the basin of the River Nile in Egypt and were engaged in swimming in the local fresh water repertoires just prior to the development of the ocular lesions. 99 patients (103 eyes) showed active granulomatous anterior uveitis with distinct pearl-like white nodules in the AC measuring between 2 and 7 mm in diameter. Inactive scarred lesions were noted in 11 patients (12 eyes). Structural complications including cataract, corectopia and phthisis were documented in 29 eyes. PCR detected digenic trematode DNA in 6 out of 14 excised nodules. Histopathological examination showed aggregates of eosinophils and epithelioid cell granulomas. CONCLUSIONS: In Egypt, a unique pattern of granulomatous anterior uveitis in rural children attributable to a waterborne helminthic infection is reported. The River Nile and its fresh water fauna are implicated in our series, and the need for an environmental investigation to further outline best management options in the given endemic areas is highlighted.

Recent advances in the management of liposarcoma.

Liposarcoma is the most common soft tissue sarcoma. With its various subtypes, the natural history of this disease can vary significantly from a locally recurrent tumor to a highly malignant one carrying a poor prognosis. Progress in the understanding of the specific molecular abnormalities in liposarcoma provides greater opportunity for new treatment modalities. Although surgical resection and radiation therapy remain the keystones for the management of primary liposarcoma, the inclusion of novel agents that target known abnormalities in advanced liposarcoma enhances the potential for improved outcomes.

A subpressor dose of angiotensin II elevates blood pressure in a normotensive rat model by oxidative stress.

Oxidative stress is an imbalance between free radicals and antioxidants, and is an important etiological factor in the development of hypertension. Recent experimental evidence suggests that subpressor doses of angiotensin II elevate oxidative stress and blood pressure. We aimed to investigate the oxidative stress related mechanism by which a subpressor dose of angiotensin II induces hypertension in a normotensive rat model. Normotensive male Wistar rats were infused with a subpressor dose of angiotensin II for 28 days. The control group was sham operated and infused with saline only. Plasma angiotensin II and H2O2 levels, whole-blood glutathione peroxidase, and AT-1a, Cu/Zn SOD, and p22phox mRNA expression in the aorta was assessed. Systolic and diastolic blood pressures were elevated in the experimental group. There was no change in angiotensin II levels, but a significant increase in AT-1a mRNA expression was found in the experimental group. mRNA expression of p22phox was increased significantly and Cu/Zn SOD decreased significantly in the experimental group. There was no significant change to the H2O2 and GPx levels. Angiotensin II manipulates the free radical-antioxidant balance in the vasculature by selectively increasing O2(-) production and decreasing SOD activity and causes an oxidative stress induced elevation in blood pressure in the Wistar rat.

C-reactive protein in acute and delayed preconditioning of the rat heart.

UNLABELLED: The involvement of C-reactive protein (CRP) in early (acute) and delayed ischaemic (IPC) and pharmacological (chemical) (CPC) preconditioning in an in vivo model of rat myocardial infarction is presented. Acute IPC was produced by three 5-min occlusion (ischaemia) periods interspersed with 5 min reperfusion, followed by 30-min occlusion of the left coronary artery and 2 h reperfusion injury. Acute CPC was produced by a Kappa-opioid receptor agonist U50488H (5 mg/kg) applied i.v. 15 min before 30-min ischaemia/2-h reperfusion. Delayed preconditioning was produced by 30-min ischaemia/2-h reperfusion, induced 24 h after either ischaemic or pharmacological preconditioning. The myocardial ischaemia/reperfusion injury was evaluated on the basis of total and cardiac creatine kinase isoenzyme activity, functional recovery of the heart (ECG), infarct size (% IS/RA) and mortality at the end of the experiments. The results obtained showed that: . The Kappa-opioid receptor agonist U50488H mimics both the acute and delayed IPC in the above experimental protocol. .Both acute IPC and CPC produce effects by opening of the KATP channels (the effects were blocked by nonspecific ATP-sensitive K channel blocker glybenclamide), and via activation of protein kinase C (a selective protein kinase C inhibitor chelerythrine blocked the effects). .C-reactive protein was significantly elevated by 54% in non-preconditioned acute ischaemia/reperfusion injury. The elevation was more pronounced (82% increase) 24 h after non-preconditioned ischaemia/ reperfusion injury. It reflected very well the increase in cardiac isoenzymes, infarct size and mortality of the rats, and can be used as a marker of the severity of myocardial injury in this model. . The increase of CRP was prevented by both IPC and CPC in early, and especially in late preconditioning. This shows the involvement of CRP, not only as a marker, but as a causative factor in cardiac ischaemic/reperfusion injury. CONCLUSION: In addition to the established involvement of adenosine, bradykinin, opioid and other receptors, a suppression of myocardial CRP/complement production might be involved in the biological mechanism of preconditioning. This could be a promising perspective in clinical interventions against ischaemia/reperfusion injuries of the heart.

The role of C-reactive protein in ischemia/reperfusion injury and preconditioning in a rat model of myocardial infarction.

For the first time the involvement of C-Reactive protein (CRP) in early (acute) and delayed ischemic (IPC) and pharmacological (chemical) preconditioning (CPC) in an in vivo model of rat myocardial infarction was presented. Acute IPC was produced by three 5 minute occlusion (ischemia) periods interspersed with 5 minute reperfusion, followed by 30 minute occlusion of the left coronary artery and 2 hour reperfusion injury. Acute CPC was produced by a k-opioid receptor agonist U50488H (5 mg/kg) applied i.v. 15 minutes before 30 minute ischemia/ 2 hour reperfusion. Delayed preconditioning was produced by 30 minute ischemia/ 2 hour reperfusion, induced 24 hour after either ischemic or pharmacological preconditioning. The myocardial ischemia/reperfusion injury was evaluated on the basis of total and cardiac creatine kinase isoenzyme activity, functional recovery of the heart (ECG), infarct size (% IS/RA) and mortality at the end of the experiments. The results obtained showed that: k-opioid receptor agonist U50488H mimics both the acute and delayed IPC in the above experimental protocol; Both acute IPC and most probably CPC act by opening of K(ATP) channels (the effects were blocked by nonspecific ATP-sensitive K channel blocker glybenclamide), and via activation of protein kinase C (a selective protein kinase C inhibitor chelerythrine blocked the efects); C-reactive protein (CRP) was significantly elevated by 54% in non-preconditioned acute ischemia/reperfusion injury. The elevation was more pronounced (82% increase) 24 hour after non-preconditioned ischemia/reperfusion injury. It reflected very well the increase in cardiac isoenzymes, infarct size and mortality of the rats, and can be used as a marker of the severity of myocardial injury in this model; The increase of CRP was prevented by both IPC and CPC in early, and especially in late preconditioning. This confirms the involvement of CRP as a marker in cardiac ischemic/reperfusion injury. It was concluded that in addition to the established involvement of adenosine, bradykinin, opioid and other receptors, a suppression of myocardial CRP/complement production might be involved in the biological mechanism of preconditioning. This could be a promising perspective in clinical interventions against ischemia/reperfusion injuries of the heart.

Facets of the metabolic syndrome in Dahl hypertensive rats.

We recently established that the Dahl salt-sensitive rat, a model for genetic salt-sensitive hypertension, was insulin resistant. This study was undertaken to evaluate whether other features of the metabolic syndrome developed in this animal model. Two groups of 16 Dahl salt-sensitive (DSS) rats and their controls, Dahl salt-resistant (DSR) rats were used. For eight weeks, half of each group was fed a standard diet with low sodium content (85 mmol Na/kg diet) while the remainder was fed a high-sodium diet (340 mmol Na/kg diet). Weekly systolic and diastolic blood pressures were measured for all animals. At the end of eight weeks, the urinary Na(+)/K(+) ratio, fasting blood glucose, plasma uric acid and blood lipids were determined for all animals. The same parameters were measured in two additional matched weanling DSS and DSR groups of eight animals each. Adult DSS rats became hypertensive, with the DSS high-salt group exhibiting both genetic hypertension and the pressor effects of a high-salt diet. The DSS high-salt and weanling groups exhibited a lowered urinary Na(+)/K(+) ratio, indicative of greater sodium retention, when compared to their respective DSR groups (p < 0.05). No strain differences were observed in the uric acid levels. However a high-salt diet in both DSS and DSR groups elevated uric acid levels. Weanling and DSS high-salt groups showed increased total plasma cholesterol when compared to their corresponding DSR groups (p < 0.05). In addition, the DSS high-salt group also had both increased total plasma cholesterol and high-density lipoprotein (HDL) cholesterol when compared to the DSS low-salt group (p < 0.05). No significant differences in blood glucose and plasma insulin were observed in the adult groups. The weanling DSS group showed a marked hyperinsulinaemia, suggesting that DSS rats were possibly insulin resistant even before hypertension was fully established. This could indicate that insulin resistance and hypertension may be inherited as separate traits that develop in a parallel but independent manner.

Cardiovascular, antihyperlipidemic and antioxidant effects of oleanolic and ursolic acids in experimental hypertension.

Cardiovascular (systolic and diastolic blood pressure, heart rate), antihyperlipidemic (tryglycerides, total cholesterol and lipoprotein fractions), antioxidant (glutathione peroxidase--GPx, and superoxide dismutase--SOD), diuretic/saluretic and hypoglycemic activity of 98% pure oleanolic (OA) and ursolic (UA) acid were studied in Dahl salt-sensitive (DSS), insulin resistant rat model of genetic hypertension. Both OA and UA displayed low toxicity, with LC50 0.10 and 0.95 mg/ml, respectively. Although both triterpenoids did not have direct hypotensive effect, after 6-week application in a daily dose 60 mg/kg b.w., i.p., they prevented the development of severe hypertension. The antihypertensive effect was attributed to their potent diuretic-natriuretic-saluretic activity; direct cardiac effect (heart rate decrease by 34% and 32%, respectively); antihyperlipidemic (more than two times decrease of LDL and triglycerides); antioxidant (GPx increase by 12% and 10%, respectively; SOD increase by 12% and 22%, respectively), and hypoglycemic (blood glucose decrease by 20% and 50%, respectively) effects on the DSS rats. Except for the antihyperlipidemic effects, the other described above in vivo antihypertensive effects of OA and UA are reported for the first time and the underlying mechanisms are currently under investigation.

Antihypertensive, antiatherosclerotic and antioxidant activity of triterpenoids isolated from Olea europaea, subspecies africana leaves.

For the first time a biossay-directed study of triterpenoids isolated from the leaves of Olea europaea from Greece, from wild African olive and from a cultivar of O. europaea grown in Cape Town was reported. The experiment was undertaken since our preliminary analyses showed that the African wild olive leave is rich in triterpenoids and contain only traces of the glycoside oleuropein, which is typical for the European olive leaves. The isolate of the African wild olive leaves (AO) used in the experiments was found to contain 0.27% 1:1 mixture of oleanolic acid and ursolic acid, named oleuafricein. The isolate of Greek olive leaves (GO) was found to contain 0.71% oleanolic acid, and the Cape Town cultivar (CT) contained 2.47% oleanolic acid. No ursolic acid was found in either GO or CT. The antihypertensive, diuretic, antiatherosclerotic, antioxidant and hypoglycemic effects of authentic oleanolic and ursolic acid and the three isolates (GO, AO and CT) were studied on Dahl salt-sensitive (DSS), insulin-resistant rat genetic model of hypertension. All three isolates, in a dose 60 mg/kg b.w. for 6 weeks treatment, prevented the development of severe hypertension and atherosclerosis and improved the insulin resistance of the experimental animals. GO, OA and CT isolates could provide an effective and cheap treatment of this particular, most common type of salt-sensitive hypertension in the African population.

Antioxidant status of the hypertrophic heart of Dahl hypertensive rat as a model for evaluation of antioxidants.

Development of hypertension, myocardial hypertrophy and the cardiac antioxidant status of male Dahl salt-sensitive (DSS) genetically hypertensive rats was evaluated and compared to that of normotensive Dahl salt-resistant (DSR) controls. In order to obtain exaggerated and more severe hypertension, half of the animals (10 per group) were Na loaded (8% NaCl diet) for 6 weeks. The myocardial antioxidant status was estimated in tissue homogenates on the basis of tissue glutathione (GSH), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). The results showed that 6 weeks of hypertension resulted in left ventricle myocardial hypertrophy, documented by weight, morphometry and morphological changes. The compromised myocardial antioxidant status of the DSS rats was defined by significantly decreased GSH-Px and glutathione activity (13% and 41%, respectively) as compared to DSR rats. SOD in DSS myocardium was increased by 47% compared to that in DSR myocardium, an effect that is considered a compensatory mechanism to the oxidative stress. All of the above changes were exaggerated by NaCl loading. It was concluded that DSS rats, on either a normal or high NaCl diet, displayed decreased antioxidant capacity, which is most likely genetically determined. Before the Dahl rat can be considered as a suitable model for testing new cardiac antioxidants, a full characterization of the level of cardiac oxygen free radicals is required.

Immunolocalisation and endothelin-1 values In pre-eclampsia: an immunocytochemical study.

Maternal plasma ET-1 levels and the immunolocalisation of ET1 in the fetal membranes of pre-eclamptic primigravidae at >/=28 weeks, gestation were studied. The levels of maternal plasma ET1 and immunoreactive ET-1 were increased in pre-eclampsia. Immunoreactive ET-1 was localised in the amnion, chorion and decidua of normal pregnant women as well as those with preeclampsia-eclampsia. Intense labelling was observed in moderate pre-eclampsia (BP 140/90 - 170/110 mmHg) with very intense labelling in severe pre-eclampsia (BP >170/110 mmHg), especially in the amniotic epithelium, chorionic villi, maternal blood vessels, cytotrophoblasts and giant cells of the decidua. The increased ET-1 levels demonstrated in fetal membranes of pre-eclamptic women are probably produced in a paracrine and/or autocrine manner, contributing to the hypertension, vasospasm and fetal growth restriction characteristic of the syndrome. A larger study would be required to show significant change in endothelin production in pre-eclampsia.

Endothelin-1 and endothelin receptor status in kidney transplants undergoing acute rejection.

Endothelin-1 (ET-1) is a potent vasoconstrictor with vasopressor and mitogenic effects. Blood samples were collected from 21 renal transplant patients undergoing acute rejection at the time of diagnostic kidney biopsy: there were 20 men and one woman, mean age 35.6 years. All patients were on triple immunosuppressive therapy with cyclosporine A, azathioprine and methylprednisolone. Twenty living kidney donors pre-uninephrectomy (11 men and nine women, mean age 34 years) served as controls. Control kidney was obtained from fresh autopsy material and normal kidney tissue from nephrectomies for malignancy. Mean plasma ET-1 was significantly increased at 1.56 +/- 0.2 pg ml(-1) during acute rejection compared to 0.74 +/- 0.06 pg ml(-1) in donors (p = 0.0009 unpaired t-test). ET(A) receptor immunolabelling was visualised in distal tubules and collecting ducts with minimal labelling in the glomeruli and blood vessels of control kidney tissue ET(A) receptor labelling was similar in kidney biopsies with acute rejection. ET(B) receptor immunolabelling was significantly increased in glomeruli (p = 0.002) and decreased in distal tubules (p = 0.004) in kidneys with acute rejection compared to control kidney tissue. While these findings may account for the oedema and hypertension observed during acute rejection, the exact significance needs to be studied further.

Dose-dependent effect of dietary fish-oil (n-3) polyunsaturated fatty acids on in vivo insulin sensitivity in rat.

The objective of this study was to assess the effect of dietary fish oil (n-3) polyunsaturated fatty acids (PUFA) on in vivo glucose metabolism and insulin sensitivity of rats. The experiments were carried out on 18 Wistar male rats divided into three groups: control rats given a normal diet containing corn oil, and two experimental groups given 1% or 5% fish oil supplemented diets. Glucose metabolism and insulin sensitivity were assessed by euglycemic clamp technique on anesthetized animals. The results showed: 1) despite the normotensive state of the animals, a significant decrease in blood pressure, especially systolic, and decreased heart rate in both experimental groups; 2) significantly decreased plasma total and LDL cholesterol and nonsignificantly decreased plasma triglycerides; 3) significantly increased bleeding time; 4) normoglycemia and dose-dependent hypoinsulinemia; and 5) significantly increased and dose-dependent in vivo glucose utilization and glucose clearance, significantly increased insulin sensitivity. The above results suggesting that insulin production is suppressed, provide another possible explanation for the beneficial effects of fish oil via modulation of the well-known damaging cardiovascular effects of insulin.