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.

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.

Glucose metabolism and insulin sensitivity in Dahl hypertensive rats.

There is increasing recognition that hypertension is only one facet of a metabolic syndrome that represents a cluster of risk factors including insulin resistance evident during long-term development of cardiovascular disease. The objectives of the present study were to evaluate glucose metabolism and insulin sensitivity in the Dahl genetic salt-sensitive rat model of hypertension and to find out whether there is a correlation between high blood pressure, salt sensitivity and insulin sensitivity. The experiments were performed in Wistar and Dahl salt-resistant (DSR) and salt-sensitive (DSS) rats given a normal or salt-loaded (2% NaCl in the drinking water) diet for 2 months. Glucose turnover, metabolic clearance of glucose and insulin sensitivity were determined using the euglycemic clamp technique in anesthetized animals. Four different concentrations of insulin were used (2.1, 4.2, 8.4 and 16.8 mg/kg/min) with results showing that there were no significant differences in serum glucose and insulin levels, glucose utilization and clearance and insulin sensitivity between Wistar and DSR; these groups remained normotensive throughout the 2-month experiment. However, DSS rats, even on a normal diet, developed hypertension by the end of the experiment and consistent with their hypertensive condition, significantly decreased glucose utilization and clearance and decreased insulin sensitivity were observed. The changes were more marked at higher insulin infusion rates (8.4 and 16.8 mg/kg/min). These results demonstrate that DSS rats with fully developed hypertension were insulin resistant. In contrast, DSR rats remained normotensive despite sodium loading. Sodium loading significantly exaggerated the hypertensive state of DSS rats but did not further increase insulin resistance. The results from respective weanling rats showed that even at this early stage before development of hypertension, DSS rats already had significantly increased serum insulin suggesting insulin resistance. In conclusion, the present results suggest that DSR genetically hypertensive rats were insulin resistant although insulin resistance was unrelated to high blood pressure or NaCl intake.

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.

An experimental rat model of salt-sensitive hypertension; biochemical and morphological parameters and sympathetic nervous system.

The objective of the study was to outline the characteristics of the development of hypertension and some neurohumoral, haematological and morphological factors contributing to development of high blood pressure in a genetic model of salt-sensitive rat. Characteristics of Dahl salt-sensitive (DS) rats, as compared to their Dahl salt-resistant (DR) controls were as follows: 1) DS rats display higher blood pressure and lower heart rate compared to DR rats as early as 1 month of age at weaning). They gradually develop hypertension at 2 months of age, irrespective of diet. Low-Na diet (0.5% NaCl) does not prevent hypertension but delays its development and ameliorates it. High Na-diet (8% NaCl) exacerbates hypertension. 2) DS rats have retardation in body weight gain. They develop mild hypochromic anaemia. 3) After 2 months of Na loading (3 months of age), DS rats express significantly increased Na and water retention and increased plasma volume by 15% compared to 2.8% increase in DR rats on high-Na diet. 4) DS rats showed renal parenchymal lesions, more pronounced after Na-loading, focal atrophy of cortical tubules, mesangial matrix expansion and glomerulosclerosis. Consistent with high blood pressure were changes in renal arterioles, fibromuscular proliferation, deposition of fibrinoid material in intima. 5) Sodium loading produced increased activity of the sympathetic nervous system (SNS), and sodium restriction reduced SNS responsiveness.

Zinc, copper and iron levels in tissues of the vitamin B6 deficient rat.

Tissue zinc (Zn), copper (Cu) and iron (Fe) were determined in three groups of young male Wistar rats that received a daily pyridoxine hydrochloride (PN.HCl) intake of 45, 23 and 0 micrograms respectively in their diets over 8 weeks. No significant differences were found in the Zn and Cu levels in the liver, kidney, skeletal and cardiac tissue of all 3 groups. The Fe levels were significantly higher in the heart and liver and significantly lower in the skeletal muscle of the group receiving no PN.HCl in the diet (P < 0.05). This study indicates that the increased fecal excretion of Zn and Cu observed during a previous balance study on the above vitamin B6 deficient group of animals may be due to a decreased absorption of these elements from the diet rather than their excretion from tissue stores. The changes in Fe levels in the heart, liver and skeletal muscle points towards some alteration in tissue stores of this element during a vitamin B6 deficiency.

Zinc, copper and iron balance in the vitamin B-6-deficient rat.

Zinc (Zn), copper (Cu) and iron (Fe) balance was assessed over 8 weeks in 3 groups of young male Wistar rats receiving a daily pyridoxine hydrochloride (PN-HCl) intake of 45, 23 and 0 micrograms respectively. Although all 3 groups remained in a positive balance throughout the study the Zn and Cu balance, growth and food intake was significantly lower in the group receiving no PN-HCl (p < 0.05). The lowered Zn and Cu balance in this group resulted from a decreased food intake coupled with an increased excretion of Zn and Cu. The group receiving 23 micrograms/d of PN-HCl showed no significant difference in growth, food intake and Zn and Cu balance when compared to the group receiving 45 micrograms/d, indicating that subminimal levels of vitamin may still be exerting beneficial effects on the balance of these trace elements.