Neuroprotective and antioxidative effects of pioglitazone in brain tissue adjacent to the ischemic core are mediated by PI3K/Akt and Nrf2/ARE pathways.

The present study elucidates the neuroprotective mechanisms of the PPARγ (peroxisome proliferator-activated receptor γ) agonist pioglitazone in survival of ischemic neurons following middle cerebral artery occlusion with reperfusion (MCAO). Intracerebroventricular infusion of pioglitazone over 5 days before and 24 or 48 h after MCAO alleviated neurological impairments, inhibited apoptosis 24 h, and activated the PI3K/Akt pathway along with increased phosphorylation of Akt (ser473) and GSK-3ß (ser9) in the peri-infarct cortical areas 48 h after MCAO. In primary cortical neurons, pioglitazone suppressed the glutamate-induced release of lactate dehydrogenase by a PPARγ-dependent mechanism. This protective effect was reversed after co-treatment with PI3K and Akt inhibitors, LY294002 and SH-6, respectively. Pioglitazone enhanced the expression of the antioxidative transcription factor Nrf2 and its target gene protein, heme oxidase-1, in the peri-infarct area. Pioglitazone also increased activation of the antioxidant response element (ARE) in neuronal PC12 cells transfected with the pNQO1-rARE plasmid. We demonstrate in primary cortical neurons from Nrf2 knockout mice that the lack of Nrf2 completely abolished the neuroprotective effects of pioglitazone against oxidative and excitotoxic damage. Our results strongly suggest that the neuroprotective effects of PPARγ in peri-infarct brain tissues comprise the concomitant activation of the PI3K/Akt and Nrf2/ARE pathways. KEY MESSAGES: Pioglitazone inhibits apoptosis in ischemic brain tissue.  Pioglitazone acting on PPARγ activates PI3K/Akt pathway in ischemic brain tissue. Pioglitazone activates via Nrf2 the antioxidant defense pathway in injured neurons. Pioglitazone activates the antioxidant response element in neuronal PC12 cells. Pioglitazone fails to protect primary neurons lacking Nrf2 against oxidative damage. Activation of PPARγ supports the survival of viable neurons in peri-infarct regions.

Neuroprotective effects of AT1 receptor antagonists after experimental ischemic stroke: what is important?

The present study conducted in rats defines the requirements for neuroprotective effects of systemically administered AT1 receptor blockers (ARBs) in acute ischaemic stroke. The inhibition of central effects to angiotensin II (ANG II) after intravenous (i.v.) treatment with candesartan (0.3 and 3 mg/kg) or irbesartan and losartan (3 and 30 mg/kg) was employed to study the penetration of these ARBs across the blood-brain barrier. Verapamil and probenecid were used to assess the role of the transporters, P-glycoprotein and the multidrug resistance-related protein 2, in the entry of losartan and irbesartan into the brain. Neuroprotective effects of i.v. treatment with the ARBs were investigated after transient middle cerebral artery occlusion (MCAO) for 90 min. The treatment with the ARBs was initiated 3 h after the onset of MCAO and continued for two consecutive days. Blood pressure was continuously recorded before and during MCAO until 5.5 h after the onset of reperfusion. The higher dose of candesartan completely abolished, and the lower dose of candesartan and higher doses of irbesartan and losartan partially inhibited the drinking response to intracerebroventricular ANG II. Only 0.3 mg/kg candesartan improved the recovery from ischaemic stroke, and 3 mg/kg candesartan did not exert neuroprotective effects due to marked blood pressure reduction during reperfusion. Both doses of irbesartan and losartan had not any effect on the stroke outcome. An effective, long-lasting blockade of brain AT1 receptors after systemic treatment with ARBs without extensive blood pressure reductions is the prerequisite for neuroprotective effects in ischaemic stroke.

Mitochondrial uncoupling proteins regulate angiotensin-converting enzyme expression: crosstalk between cellular and endocrine metabolic regulators suggested by RNA interference and genetic studies.

Uncoupling proteins (UCPs) regulate mitochondrial function, and thus cellular metabolism. Angiotensin-converting enzyme (ACE) is the central component of endocrine and local tissue renin-angiotensin systems (RAS), which also regulate diverse aspects of whole-body metabolism and mitochondrial function (partly through altering mitochondrial UCP expression). We show that ACE expression also appears to be regulated by mitochondrial UCPs. In genetic analysis of two unrelated populations (healthy young UK men and Scandinavian diabetic patients) serum ACE (sACE) activity was significantly higher amongst UCP3-55C (rather than T) and UCP2 I (rather than D) allele carriers. RNA interference against UCP2 in human umbilical vein endothelial cells reduced UCP2 mRNA sixfold (P < 0·01) whilst increasing ACE expression within a physiological range (<1·8-fold at 48 h; P < 0·01). Our findings suggest novel hypotheses. Firstly, cellular feedback regulation may occur between UCPs and ACE. Secondly, cellular UCP regulation of sACE suggests a novel means of crosstalk between (and mutual regulation of) cellular and endocrine metabolism. This might partly explain the reduced risk of developing diabetes and metabolic syndrome with RAS antagonists and offer insight into the origins of cardiovascular disease in which UCPs and ACE both play a role.

Mitochondrial uncoupling proteins regulate angiotensin-converting enzyme expression: crosstalk between cellular and endocrine metabolic regulators suggested by RNA interference and genetic studies.

Uncoupling proteins (UCPs) regulate mitochondrial function, and thus cellular metabolism. Angiotensin-converting enzyme (ACE) is the central component of endocrine and local tissue renin-angiotensin systems (RAS), which also regulate diverse aspects of whole-body metabolism and mitochondrial function (partly through altering mitochondrial UCP expression). We show that ACE expression also appears to be regulated by mitochondrial UCPs. In genetic analysis of two unrelated populations (healthy young UK men and Scandinavian diabetic patients) serum ACE (sACE) activity was significantly higher amongst UCP3-55C (rather than T) and UCP2 I (rather than D) allele carriers. RNA interference against UCP2 in human umbilical vein endothelial cells reduced UCP2 mRNA sixfold (P < 0·01) whilst increasing ACE expression within a physiological range (<1·8-fold at 48 h; P < 0·01). Our findings suggest novel hypotheses. Firstly, cellular feedback regulation may occur between UCPs and ACE. Secondly, cellular UCP regulation of sACE suggests a novel means of crosstalk between (and mutual regulation of) cellular and endocrine metabolism. This might partly explain the reduced risk of developing diabetes and metabolic syndrome with RAS antagonists and offer insight into the origins of cardiovascular disease in which UCPs and ACE both play a role.

Omapatrilat: penetration across the blood-brain barrier and effects on ischaemic stroke in rats.

Omapatrilat (OMA), which simultaneously inhibits the angiotensin-converting enzyme (ACE) and the neutral endopeptidase (neprilysin (NEP)), is widely used in experimental protocols related to hypertension and heart failure. The penetration of OMA across the blood-brain barrier (BBB) and the effects of ACE/NEP inhibition on the recovery from ischaemic stroke have not yet been investigated. Angiotensin (Ang) I injected intracerebroventricularly (ICV) or intravenously (IV) is converted to Ang II by ACE and induces an immediate increase in blood pressure. The pressor responses to OMA administered ICV, orally or IV were studied in male Wistar rats instrumented with an ICV and arterial and venous catheters. OMA infused ICV rapidly appeared in the systemic circulation and more effectively attenuated the systemic than the central pressor responses to Ang I. OMA administered orally (5, 25, 100 µmol/kg body weight) or IV (0.5, 1, 5, 25 µmol/kg body weight) completely abolished increases in blood pressure to IV Ang I up to 2 h after treatment. The pressor responses to ICV Ang I were not altered, indicating that systemically administered OMA does not cross the BBB. To study the effects of ACE and NEP inhibition in the brain on the recovery from ischaemic stroke, OMA was infused ICV over a 5-day period before and 24 h after the occlusion of the middle cerebral artery (MCAO) for 90 min. ICV application of OMA had no effect on infarction volume and marginally improved neurological outcome. We demonstrate for the first time that simultaneous inhibition of ACE and NEP in the brain tissue does not alter the recovery from ischaemic stroke.

Treatment of rats with pioglitazone in the reperfusion phase of focal cerebral ischemia: a preclinical stroke trial.

Thiazolidinediones (TZDs), pioglitazone, rosiglitazone and troglitazone, the synthetic agonists for the PPARγ, administered prior or during ischemic insult improve stroke outcome in rodents, post-occlusion treatments yielded inconsistent results. In the present experiments carried out according to the Stroke Therapy Academic Industry Roundtable (STAIR) guidelines, we studied the effects of post-ischemic pioglitazone treatment on the outcome of focal cerebral ischemia, inflammatory and apoptotic processes, neuronal degeneration and regeneration, blood pressure, heart rate and physiological variables in blood. Male Wistar rats were subjected to a 90 min middle cerebral artery occlusion (MCAO). Subcutaneous (SC) treatment with vehicle or pioglitazone was initiated 90 min after MCAO, i.e. in the post-ischemic, reperfusion phase and continued on 2 (2 day-experiment, protocol 1) or 5 (5-day experiment, protocol 2) consecutive days. In the 2-day experiment, pioglitazone at a dose of 2.5 mg/kg body weight (bw) reduced infarct volume by 31% and oedema by 43% on day 2 after MCAO and attenuated the infiltration of ischemic cortical tissue with activated microglia and macrophages. The slight reduction in infarct volume by approximately 18%, detected in rats treated with 10 mg/kg bw pioglitazone did not reach statistical significance. The neurological scores of sham-operated rats treated with vehicle or 10 mg/kg bw pioglitazone were not significantly different. In rats subjected to cerebral ischemia, post-ischemic treatment with either dose of pioglitazone alleviated particular motor deficits and sensory impairments on day 2 after MCAO. A single injection of 10 mg/kg bw pioglitazone in the reperfusion phase (90 min after the onset of reperfusion) did not modify systolic and diastolic blood pressure, heart rate and physiological variables compared to vehicle-treated rats at any time point after MCAO. In the 5-day experiment, continuous post-occlusion treatment with 2.5 mg/kg body weight pioglitazone significantly reduced cerebral infarction by 29% and improved the partial paralysis of the forelimb and alleviated sensory deficits. In the peri-infarct cortex, pioglitazone effectively suppressed the accumulation of activated microglia/macrophages, inhibited neuronal degeneration and promoted neuroregeneration and formation of neuronal networks. The current results provide evidence that pioglitazone treatment in the post-ischemic, reperfusion phase improves the recovery from ischemic stroke. Neuroprotective effects of pioglitazone are mediated by inhibition of post-ischemic inflammation and neuronal degeneration, protection of neurones against ischemic injury and by promoting of neuronal regeneration. Our data together with previous findings favour the view that pioglitazone is a promising candidate for clinical stroke trials.

Peroxisome-proliferator-activated receptors gamma and peroxisome-proliferator-activated receptors beta/delta and the regulation of interleukin 1 receptor antagonist expression by pioglitazone in ischaemic brain.

OBJECTIVE: The imbalance between the production and release of interleukin-1 (IL-1) ligands, IL-1alpha, IL-1beta and IL-1 receptor antagonist (IL-1ra) in ischaemic brain exaggerates inflammatory responses and contributes to neuronal death. Cerebral ischaemia also upregulates the peroxisome-proliferator-activated receptor (PPAR) gamma. We studied in rats the effects of the PPARgamma agonist, pioglitazone, on the regulation of IL-1beta, IL-1ra and IL-1 receptor I (IL-1RI) expression in ischaemic brain after occlusion of the middle cerebral artery for 90 min. METHODS: Pioglitazone or vehicle was infused intracerebroventricularly over a 5-day period before, during and 24 or 48 h after middle cerebral artery occlusion. The expression of IL-1beta, IL-1ra and IL-1RI in the peri-infarct cortex was investigated by immunohistochemistry, Western blotting and immunofluorescence staining. The mechanisms of the IL-1ra regulation by pioglitazone and the neuroprotection under excitotoxic neuronal injury were studied in primary cortical neurones expressing PPARgamma and PPAR beta/delta. RESULTS: Cerebral ischaemia increased the expression of IL-1beta, IL-1RI and IL-1ra in the ischaemic cortex. Pioglitazone reduced IL-1beta, but upregulated IL-1ra and increased the number of IL-1ra immunoreactive cells. In primary cortical neurones, pioglitazone stimulated the IL-1ra production via activation of the PPARbeta/delta, but prevented excitotoxic neuronal injury and death by a PPARgamma-dependent mechanism. CONCLUSION: Our data demonstrate that activation of PPARgamma and PPAR beta/delta by proglitazone in neurones triggers diverse neuroprotective mechanisms. The restoration of the equilibrium between I1-1beta and IL-1ra in ischaemic brain tissue limits IL-1beta signalling, reduces inflammatory responses and is an important mechanism by which thiazolidinediones improve the recovery from ischaemic stroke.

Peroxisome proliferator-activated receptorsgamma (PPARgamma) differently modulate the interleukin-6 expression in the peri-infarct cortical tissue in the acute and delayed phases of cerebral ischaemia.

Interleukin-6 (IL-6) exerts neuroprotective effects after cerebral ischaemia but can also exacerbate inflammation and induce neuronal death. The current study investigates the role of cerebral peroxisome proliferator-activated receptor(s) gamma (PPARgamma) in the regulation of IL-6 expression in the peri-infarct cortical tissue in rats exposed to focal cerebral ischaemia. Pioglitazone, a high-affinity PPARgamma ligand, was infused intracerebroventricularly (i.c.v.) via osmotic minipumps over a 5-day period before, during and 24 h or 48 h after middle cerebral artery occlusion (MCAO) for 90 min followed by reperfusion. The expression of PPARgamma and IL-6 in cortical tissue adjacent to the ischaemic core was studied 24 h and 48 h after MCAO. Pioglitazone augmented the ischaemia-induced upregulation of PPARgamma at both time points. Cerebral ischaemia substantially increased IL-6 expression in the peri-infarct cortical tissue. Twenty-four hours after MCAO, the majority of microglial cells/macrophages showed an intense IL-6 immunoreactivity. IL-6 was also localized in neurons, but the distribution of neurons positively stained for IL-6 at the border of the infarct was very heterogeneous. Pioglitazone effectively decreased the number of IL-6-immunoreactive cells and IL-6 protein levels at 24 h but not at 48 h after MCAO. Pioglitazone treatment reduced the infarct size and improved neurological functions. The present study demonstrates that cerebral PPARgamma suppresses the expression of IL-6 in ischaemic brain tissue during the initial phase of ischaemic stroke, in which the overproduction of IL-6 may aggravate neuronal damage, but not at later time points, when IL-6 promotes neuroprotection and inhibits neuronal death.

Haplotype of growth hormone and angiotensin I-converting enzyme genes, serum angiotensin I-converting enzyme and ventricular growth: pathway inference in pharmacogenetics.

OBJECTIVES: An insertion/deletion (I/D) polymorphism in the angiotensin I-converting enzyme (ACE) gene is associated with variations in circulating and tissue angiotensin I-converting enzyme activity, and differences in exercise-induced left ventricular hypertrophic response. A genetic marker (CSH1.01) in the syntenic GH-CSH gene cluster correlates with metabolic syndrome in adult life in males. Approximately 24% linkage disequilibrium between CSH1.01T and D alleles of ACE I/D has also been reported. The objective was to examine the hypothesis that effects ascribed to ACE genotype may reflect causality in the GH-CSH cluster. METHODS: The ACE I/D polymorphism and GH-CSH BglII-B single nucleotide polymorphism (in strong linkage disequilibrium with CSH1.01) were determined in 847 British Army recruits. Serum angiotensin I-converting enzyme activity and left ventricular mass were measured before and after a 12-week physical training program. Genotype and haplotype analyses of both markers were performed in relation to these phenotypes. RESULTS: The ACE I/D polymorphism was in linkage disequilibrium with BglII-B (D'=0.3). Strong association was seen between ACE I/D genotypes and serum angiotensin I-converting enzyme activity (P<0.0001), but not left ventricular mass change. BglII-B genotypes also associated significantly with serum angiotensin I-converting enzyme level (P<0.0001). Haplotype analysis, however, showed that most of this association resulted from linkage disequilibrium between BglII-B and ACE I/D. BglII-B did not associate with left ventricular mass change. CONCLUSIONS: GH-CSH BglII-B genotype associates significantly with angiotensin I-converting enzyme levels, but only through linkage disequilibrium with ACE I/D. Every phenotype with which ACE I/D has been associated merits investigation of potential causal effects originating in the GH-CSH cluster (and vice versa), otherwise the chain of causality could be misinterpreted.

PPAR-gamma: therapeutic target for ischemic stroke.

The peroxisome proliferator activated receptors (PPARs), which belong to the nuclear receptor superfamily, are key regulators of glucose and fat metabolism. The PPAR-gamma isoform is involved in the regulation of cellular glucose uptake, protection against atherosclerosis and control of immune reactions. In addition, the activation of PPAR-gamma effectively attenuates neurodegenerative and inflammatory processes in the brain. Here, we review a novel aspect of beneficial and clinically relevant PPAR-gamma actions: neuroprotection against ischemic injury mediated by intracerebral PPAR-gamma, which is expressed in neurons and microglia. Together with the recent observation that the PPAR-gamma ligand pioglitazone reduces the incidence of stroke in patients with type 2 diabetes, this review supports the concept that activators of PPAR-gamma are effective drugs against ischemic injury.

No correlation between circulating ACE activity and VO2max or mechanical efficiency in women.

The insertion (I) variant of the angiotensin-1 converting enzyme (ACE) I/D genetic polymorphism is associated with lower circulating and tissue ACE activity. Some studies have also suggested associations of ACE I/D genotype with endurance phenotypes. This study assessed the relationships between circulating ACE activity, ACE I/D genotype, mechanical efficiency and the maximal rate of oxygen uptake in sedentary individuals. Sixty-two untrained women were tested for mechanical efficiency during submaximal cycle ergometry (delta and gross efficiencies during exercise between 40 and 80 W) and the maximal rate of oxygen uptake during incremental treadmill running. Respiratory variables were measured using indirect calorimetry. Venous blood was obtained for direct assay of circulating ACE activity, allowing for the assessment of correlations between two continuous variables, rather than a categorical analysis of endurance phenotype by genotype alone. ACE I/D genotype was also determined, and was strongly associated with circulating ACE activity (P < 0.0005). Neither circulating ACE activity (27.4 +/- 8.4 nM His-Leu-ml(-1)) nor ACE genotype showed a statistically significant association with any of the endurance phenotypes measured. The weak correlations observed included r = -0.122 (P = 0.229) for the relationship between delta efficiency (23.9 +/- 2.5%) and circulating ACE activity and r = 0.134 (P > 0.6) for the relationship between maximal aerobic power (149.1 +/- 22.9 ml kg(-2/3) min(-1)) and circulating ACE activity. The data do not support a role for systemic ACE activity in the regulation of endurance performance in sedentary individuals, extending this observation to a large female cohort.

Comparison between early and delayed systemic treatment with candesartan of rats after ischaemic stroke.

OBJECTIVE: The effects of candesartan treatment starting early (3 h) and delayed (24 h) after middle cerebral artery occlusion (MCAO) with reperfusion was investigated in normotensive rats. METHODS: Subcutaneous treatment with candesartan (0.3 and 3 mg/kg) or vehicle was initiated 3 or 24 h after the onset of MCAO and continued for seven consecutive days (n=20 per group and timepoint). Neurological outcome was evaluated daily using two different scoring systems. Infarct and oedema volumes were determined in rats 2 or 7 days after MCAO. Mean arterial, systolic and diastolic blood pressures were recorded before and after the application of candesartan. RESULTS: Mean arterial, systolic and diastolic blood pressures were markedly decreased with the high dose, but only moderately decreased with the low dose of candesartan. Vehicle-treated rats showed marked neurological deficits 24 h after MCAO, which gradually improved with time. Candesartan improved neurological outcomes at all timepoints only when treatment was started 3, but not 24 h after MCAO. The infarct volume was reduced on days 2 and 7 after MCAO in rats treated with the low but not the high dose of candesartan. CONCLUSION: The present study demonstrates that only an early but not a delayed onset of treatment with candesartan exerts neuroprotection after focal ischaemia. The degree of neurological impairments did not correlate with the infarct volume, which was reduced only after the low dose of candesartan. The high dose of candesartan failed to reduce the infarct volume, probably because of an excessive blood pressure decrease.

Activation of cerebral peroxisome proliferator-activated receptors gamma promotes neuroprotection by attenuation of neuronal cyclooxygenase-2 overexpression after focal cerebral ischemia in rats.

Up-regulation of cyclooxygenase (COX)-2 exacerbates neuronal injury after cerebral ischemia and contributes to neuronal cell death. The present study clarifies the function of cerebral peroxisome-proliferator-activated receptor(s) gamma (PPARgamma) in the expression of COX-2 in neurons of the rat brain after middle cerebral artery occlusion (MCAO) with reperfusion by immunohistochemistry, Western blot, and immunofluorescence staining. In peri-infarct cortical areas the PPARgamma was located in both microglia and neurons, whereas COX-2 was almost exclusively expressed in neurons. PPARgamma immunolabeling reached the peak 12 h after MCAO, whereas the number of COX-2 immunostained cells gradually rose and reached its peak at 48 h. Intracerebroventricular infusion of pioglitazone, an agonist of the PPARgamma, over a 5-day period before and 2 days after MCAO, reduced the infarct size, the expression of tumor necrosis factor alpha (TNF-alpha), COX-2, and the number of cells positively stained for COX-1 and COX-2 in the peri-infarct cortical regions. COX-2 induction was also attenuated in the ipsilateral but not in the contralateral hippocampus. In primary cortical neurons expressing the PPARgamma, pioglitazone suppressed COX-2 expression in response to oxidative stress. This protective effect was reversed after cotreatment with GW 9662, a selective antagonist of the PPARgamma, clearly demonstrating a PPARgamma-dependent mechanism. Our data provide evidence that activation of neuronal PPARgamma considerably contributes to neuroprotection by prevention of COX-2 up-regulation in vitro and in peri-infarct brain areas.

Acute blood pressure effects and chronic hypotensive action of calcimimetics in uremic rats.

A previous study in subtotally nephrectomized (SNX) rats suggested beneficial effects of the calcimimetic R-568 beyond the control of mineral metabolism. This study analyzed potential blood pressure (BP)-lowering effects of R-568. Male Sprague-Dawley rats received two-stage subtotal nephrectomy or sham operation. Telemetry devices were inserted into the abdominal aorta, and BP was measured every 5 min. R-568 (20 mg/kg per d) or solvent was infused for 4 wk followed by once-daily subcutaneous injections for 2 wk. Total body sodium was measured by neutron activation analysis. The uremia-induced increase of mean arterial pressure from baseline to day 42 in SNX solvent rats (103+/-5 to 128+/-14 mmHg, P=0.006) was attenuated by R-568 (104+/-5 to 111+/-8 mmHg; P<0.0001 for difference of slopes). The circadian rhythm was abrogated in SNX rats and not restored by R-568. In sham-operated rats, R-568 had only a minor transient antihypertensive effect. R-568 injection induced a transient rise of mean arterial pressure by 23+/-4 and 26+/-10 mmHg in sham and SNX rats but only by 9+/-3 and 10+/-5 mmHg in solvent-treated rats (P<0.01 versus baseline and solvent versus R-568). Plasma angiotensin-converting enzyme activity and aldosterone levels were similar; food intake and physical activity did not differ throughout the study. In healthy rats, total body sodium was higher after 14 d of R-568 compared with solvent infusion (37.1+/-4 versus 32.5+/-1.4 mmol/kg; P=0.01). The calcimimetic R-568 causes an initial BP increase in sham-operated and uremic rats, which in uremic rats is followed by a marked and sustained antihypertensive effect.

Lack of change in serum angiotensin-converting enzyme activity during the menstrual cycle.

INTRODUCTION: The Deletion (D) rather than Insertion (I) variant of the angiotensin-converting enzyme (ACE) gene is associated with higher circulating ACE activity. Meanwhile, coronary risk rises with the menstrual nadir in oestrogen levels, exogenous oestrogen reduces serum ACE activity (with a greater reduction the higher the baseline ACE activity), and pharmacological reduction in ACE activity is cardioprotective. Alterations in coronary risk associated with the menstrual cycle may thus be mediated through (genotype-dependent) changes in ACE activity. We have examined this hypothesis. MATERIALS AND METHODS: Twenty-three healthy female subjects (12 II, 11 DD genotype) were studied. None were taking oral contraceptive agents. Blood was assayed for oestrogen, follicle stimulating hormone (FSH), luteinising hormone (LH), progesterone and ACE activity every three days throughout their menstrual cycle. RESULTS: ACE activity was unrelated to oestrogen, FSH or LH during the menstrual cycle, irrespective of ACE genotype. CONCLUSIONS: The increase in myocardial ischaemia during low oestrogen phases of the menstrual cycle does not appear mediated through a fall in serum ACE activity.

Low-dose lithium combined with captopril prevents stroke and improves survival in salt-loaded, stroke-prone spontaneously hypertensive rats.

OBJECTIVE: A number of potential interactions between angiotensin-converting enzyme inhibitors and lithium have been described in the literature. In the present study, we investigated the effects of a low-dose combination treatment with lithium and captopril on survival and stroke prevention in salt-loaded, stroke-prone spontaneously hypertensive rats (SHRSP). METHODS: Eight-week-old saline-drinking SHRSP (n = 21 per group) were treated with vehicle, LiCl (1 mmol/kg per day), captopril (25 mg/kg per day) and captopril plus LiCl for up to 37 weeks. Body weight, salt water intake blood pressure and mortality were recorded throughout the experimental period. Plasma renin activity, plasma lithium concentration and urinary excretion of albumin, sodium and potassium were measured at different time points. RESULTS: Captopril treatment doubled the life expectancy when compared with vehicle-treated rats. Lithium alone had minor effects on survival but led to a dramatic increase in survival when added to captopril (mean survival time > 237 versus 147 days, P < 0.001). Systolic blood pressure increased with age in all treatment groups but was comparable in the captopril-treated and the captopril-plus-lithium-treated groups. Plasma renin activity as well as urinary sodium and potassium excretion did not differ between both groups. In the captopril group a striking fivefold increase of albuminuria occurred between 14 and 26 weeks of age, while this progression was completely abolished by the addition of lithium. CONCLUSIONS: Our results demonstrate that the addition of lithium to captopril dramatically prolong the effects of the angiotensin-converting enzyme inhibitor on survival in salt-loaded SHRSP. This effect was independent of a reduction in blood pressure.

The intracerebral application of the PPARgamma-ligand pioglitazone confers neuroprotection against focal ischaemia in the rat brain.

The present study addresses the neuroprotective function of intracerebroventricular (i.c.v.) application of pioglitazone, a selective ligand of the peroxisome proliferator-activated receptor gamma (PPARgamma) in the rat brain after ischaemia. Pioglitazone or vehicle were i.c.v. infused via osmotic minipumps over a 5-day period before, and 2 days after transient middle cerebral artery occlusion (MCAO) for 90 min. This i.c.v. application of pioglitazone in the brain significantly reduced the infarct size and brain oedema, and attenuated in the peri-infarct cortical regions the invasion of activated microglia and macrophages. Moreover, pioglitazone improved the recovery of sensory deficits 48 h after MCAO. Our data demonstrate for the first time that it is the activation of intracerebral PPARgamma that can confer neuroprotection, anti-inflammatory and neurological improvement following ischaemic injury.

No association between Angiotensin Converting Enzyme (ACE) gene variation and endurance athlete status in Kenyans.

East African runners are continually successful in international distance running. The extent to which genetic factors influence this phenomenon is unknown. The insertion (I) rather than deletion (D) of a 287 bp fragment in the human angiotensin converting enzyme (ACE) gene is associated with lower circulating and tissue ACE activity and with endurance performance amongst Caucasians. To assess the association between ACE gene variation and elite endurance athlete status in an African population successful in distance running, DNA samples were obtained from 221 national Kenyan athletes (N), 70 international Kenyan athletes (I), and 85 members of the general Kenyan population (C). Blood samples were obtained from C and assayed for circulating ACE activity. ACE I/D (rs????--from NCBI SNPdb first time poly mentioned) genotype was determined, as was genotype at A22982GD (rs????--from NCBI SNPdb first time poly mentioned) which has been shown to associate more closely with ACE levels in African subjects than the I/D polymorphism. ACE I/D and A22982G genotypes explained 13 and 24% of variation in circulating ACE activity levels (P = 0.034 and <0.001 respectively). I/D genotype was not associated with elite endurance athlete status (df = 4, chi(2) = 4.1, P=0.39). In addition, genotype at 22982 was not associated with elite endurance athlete status (df = 4, chi(2) = 5.7, P = 0.23). Nor was the A allele at 22982, which is associated with lower ACE activity, more prevalent in N (0.52) or I (0.41) relative to C (0.53). We conclude that ACE I/D and A22982G polymorphisms are not strongly associated with elite endurance athlete status amongst Kenyans.

Circulating angiotensin converting enzyme activity is correlated with muscle strength.

PURPOSE: The D-variant of the angiotensin-1 converting enzyme (ACE) gene is associated with higher circulating and tissue ACE activity. Some studies have suggested a similar association of genotype with muscle strength or the gain in strength in response to training. This study has assessed the relationship between circulating ACE activity, strength, and the response to training. METHODS: Eighty-one untrained men were tested for quadriceps muscle strength, and 44 of these performed an 8-wk program of dynamic strength training of the quadriceps muscle group. Venous blood was obtained for assessment of circulating ACE activity before and after the training program. ACE genotype was also determined. RESULTS: At baseline, circulating ACE activity was significantly correlated with isometric (r = 0.25-0.29, P < 0.02) and isokinetic (r = 0.38, P < 0.0005) quadriceps muscle strength. ACE genotype also seemed to be related to pretraining muscle strength. However, circulating ACE activity showed no significant association with the 9-14% mean increases of muscle strength in response to the training intervention. ACE genotype also showed no association with the training-induced change in muscle strength. Circulating ACE activity did not change significantly after the training program. CONCLUSIONS: The data support a role for ACE in the regulation of human skeletal muscle strength, but do not confirm a role in altering the response to short-term training.

Sustained blockade of brain AT1 receptors before and after focal cerebral ischemia alleviates neurologic deficits and reduces neuronal injury, apoptosis, and inflammatory responses in the rat.

In the present study, we investigate whether a long-term blockade of brain AT1 receptors in male Wistar rats before and after ischemic injury exerts neuroprotective effects and modulates apoptosis and inflammatory responses, which are associated with the post-ischemic progression of brain damage. The AT1 receptor antagonist irbesartan was continuously infused intracerebroventricularly using osmotic minipumps over a 5-day period before and for 3 or 7 days after middle cerebral artery occlusion (MCAO) for 90 minutes. Neurologic status was evaluated daily, starting 24 hours after MCAO. After MCAO (3 and 7 days), brains were removed for the measurement of infarct size and immunohistochemical evaluation of apoptosis and accumulation of reactive microglia and macrophages. Treatment with irbesartan before ischemia improved motor functions, whereas post-ischemic treatment improved sensory functions. Blockade of brain AT1 receptors reduced the infarct size on days 3 and 7 after MCAO. In the peri-infarct cortex, irbesartan treatment decreased the number of apoptotic cells on day 3 and attenuated the invasion of activated microg-lia and macrophages on days 3 and 7 after ischemia. Long-term blockade of brain AT1 receptors improves the recovery from cerebral ischemia. Antiapoptotic mechanisms and inhibition of post-ischemic inflammation are involved in the AT1 receptor blockade-induced neuroprotective effects in ischemic brain tissue.