Protection against endotoxic shock as a consequence of reduced nitrosative stress in MLCK210-null mice.

This study investigated the consequences of deletion of the long isoform of myosin light chain kinase (MLCK210) on the cardiovascular changes induced by the bacterial endotoxin lipopolysaccharide (LPS) and cecal ligation puncture using MLCK210-/- mice. Here, we provide evidence that deletion of MLCK210 enhanced survival after intraperitoneal injection of LPS or cecal ligation puncture. LPS-induced vascular hyporeactivity to vasoconstrictor agents was completely prevented in aorta from MLCK210-/- mice. This was associated with a decreased up-regulation of nuclear facor-kappaB expression and activity, inducible nitric-oxide synthase, and level of oxidative stress in the vascular media. Furthermore, LPS-induced increase of nitric oxide production in the circulation and tissues (including heart, liver, and lung) that was correlated with an increased expression of inducible nitric-oxide synthase was also reduced in MLCK210-/- mice. These data demonstrate a role for MLCK210 in endotoxin shock injury associated with oxidative and nitrosative stresses and vascular hyporeactivity.

Cyclosporine monitoring in renal transplant recipients with induction therapy: C2 levels in patients monitored on C0.

The aim of this retrospective study was to compare the cyclosporine C(2) blood levels in renal transplant recipients with induction therapy, monitored on C(0) levels during the early and long-term post-transplantation periods in different French transplantation centres, to the target values recommended by the International Consensus on Neoral and used in the Mo2art study. A retrospective study was conducted by the therapeutic drug monitoring (TDM) committee of the French Pharmacological Society. Cyclosporine C(0) and C(2) concentrations from 168 renal transplant recipients were collected at different post-transplantation periods by six TDM laboratories of transplantation centres from April 2001 to April 2002. Cyclosporine blood levels were determined by fluorescence polarization immunoassay (mFPIA, AxSYM, Abbott) or enzyme immunoassay (EMIT, Dade Behring). Most patients had C(0) values in the recommended target ranges, with C(2) levels below the targets used in the Mo2art study or proposed by the International Consensus Conference, both during the early and long-term post-transplantation periods. Sixty-eight per cent of patients had C(2) below 1,500 microg/L +/- 20% in the first 2 months post-transplantation and 55% had C(2) below 800 microg/L +/- 20% in the late post-transplantation period (>1 year). Cyclosporine dose should be increased by 40% on average during the first week post-transplantation period and by 50% during the maintenance period to achieve the C(2) targets. In France, most renal transplant recipients receiving induction agents monitored on C(0) had C(2) levels below the targets recommended by the International Consensus Conference. In clinical practice, the optimal therapeutic windows for CsA monitoring based on C(2) needs to be more precisely defined, both during the early and long-term post-transplantation periods in renal transplant recipients receiving induction agents.

Deletion of peroxisome proliferator-activated receptor-alpha induces an alteration of cardiac functions.

The peroxisome proliferator-activated receptor-alpha (PPARalpha) plays a major role in the control of cardiac energy metabolism. The role of PPARalpha on cardiac functions was evaluated by using PPARalpha knockout (PPARalpha -/-) mice. Hemodynamic parameters by sphygmomanometric measurements show that deletion of PPARalpha did not affect systolic blood pressure and heart rate. Echocardiographic measurements demonstrated reduced systolic performance as shown by the decrease of left ventricular fractional shortening in PPARalpha -/- mice. Telemetric electrocardiography revealed neither atrio- nor intraventricular conduction defects in PPARalpha -/- mice. Also, heart rate, P-wave duration and amplitude, and QT interval were not affected. However, the amplitude of T wave from PPARalpha -/- mice was lower compared with wild-type (PPARalpha +/+) mice. When the myocardial function was measured by ex vivo Langendorff's heart preparation, basal and beta-adrenergic agonist-induced developed forces were significantly reduced in PPARalpha-null mice. In addition, Western blot analysis shows that the protein expression of beta1-adrenergic receptor is reduced in hearts from PPARalpha -/- mice. Histological analysis showed that hearts from PPARalpha -/- but not PPARalpha +/+ mice displayed myocardial fibrosis. These results suggest that PPARalpha-null mice have an alteration of cardiac contractile performance under basal and under stimulation of beta1-adrenergic receptors. These effects are associated with myocardial fibrosis. The data shed light on the role of PPARalpha in maintaining cardiac functions.

Deletion of MLCK210 induces subtle changes in vascular reactivity but does not affect cardiac function.

Myosin light chain kinase (MLCK) plays a key role in the regulation of actomyosin contraction in a large variety of cells. Two isoforms have been described: a short isoform, widely expressed in smooth muscle cells; and a long isoform (MLCK210), mainly localized in the endothelium. This study investigated the consequences on different cardiovascular parameters of MLCK210 gene deletion using MLCK210 knockout mice and of pharmacological inhibition of the kinase using a specific MLCK inhibitor. Deletion of MLCK210 did not affect systolic blood pressure and heart rate or echocardiographic measurements. Electrocardiographic analysis showed neither atrio- nor intraventricular conduction or repolarization defects. Ex vivo responses of aortic rings to vasoconstrictor and vasodilator agonists were not modified in MLCK210 null mice. However, deletion of MLCK210 attenuated shear stress-induced dilation and produced changes in the balance of endothelial-relaxing factors of small mesenteric arteries (SMA). In particular, a reduced flow-mediated NO-dependent dilation was observed. However, it was partially compensated by enhanced indomethacin-sensitive dilation. No significant changes were detected in the endothelium-derived hyperpolarizing component of the vasodilator response. The above effects of MLCK210 gene deletion were confirmed in SMA from wild-type mice by the use of the MLCK enzymatic inhibitor MMZ-10-057. In summary, deletion of MLCK210 was not associated with abnormalities of main in vivo cardiovascular parameters in mice. This study demonstrates a role for MLCK210 in the regulation of flow-dependent dilation in SMA.

Measurement by magnetic resonance imaging of the peritoneal membrane in contact with dialysate in rats.

To be optimal, a peritoneal dialysis prescription should consider the peritoneal surface area recruitment. In fact, as shown by computed tomography imaging, only a fraction of the available anatomic peritoneum is in contact with the dialysate (PDF). Various factors may dynamically affect the recruitment of the wetted membrane: posture, fill volume, PDF composition (biocompatibility), and pharmacologic agents (phospholipids). To precisely determine the peritoneal membrane recruitment capacity, we developed an animal model. In 5/6 bi-nephrectomized rats on peritoneal dialysis, between week 6 and week 8 post surgery, we used MRI to assess the contact area, with the dialysate acting as the contrast medium (fill volume: 10 mL per 100-g rat body weight). The MRI protocol consisted of axially oriented, turbo spin-echo, 3-mm slice, T2 weighted sequences. The contact area was measured using an adapted three-dimensional MRI reconstruction software based on DICOM (digital imaging and communications in medicine) images. The MRI studies (n=10) were successful. They showed that only a fraction of the presumed anatomic area (30% - 40%) was in contact with the PDF Peritoneal MRI in rats is a method that shows potential for assessing peritoneal contact area and its variation under experimental conditions.

Long daytime exchange in children on continuous cycling peritoneal dialysis: preservation of drained volume because of icodextrin use.

Daytime exchanges with glucose osmotic agents often lead to dialysate reabsorption, poor ultrafiltration (UF), positive sodium balance, and restricted purification of uremic toxins. We studied 5 anuric children on continuous cycling peritoneal dialysis (mean age: 10 years, 10 months), comparing icodextrin to a conventional glucose-based dialysate. The same fill volume (980 +/- 290 mL/m2) and the same dwell duration (720 minutes) were used with both solutions for the daytime exchange. In a crossover design, we compared 7.5% icodextrin with 1.36% glucose, and then 1.36% glucose with 7.5% icodextrin. Tolerance, net UF, sodium balance, and solute extraction were analyzed. The Student t-test for paired data was used for statistical analysis. The drained volume was 44% +/- 18% higher during icodextrin exchanges, allowing a mean enhanced sodium extraction of 44 +/- 15 mmol per daytime exchange. The uremic toxin extraction capacity was enhanced under icodextrin: weekly Kt/V urea increased by 0.41 +/- 0.1, weekly creatinine clearance increased by 8.4 +/- 3.6 L/1.73 m2, and phosphate removal increased by 23%. Similarly, beta2-microglobulin extraction increased with icodextrin use. Dialysate protein loss under icodextrin increased from 1.3 +/- 0.6 g to 1.9 +/- 0.96 g per daytime exchange. Icodextrin improved ultrafiltration and purification capacities (urea, creatinine, phosphate, beta2-microglobulin), but the large drained volume directly affected dialysate protein loss.

Oxytocin-induced renin secretion by denervated kidney in anaesthetized rat.

The effects of oxytocin on renin secretion by denervated kidney were investigated in vivo, by infusing the peptide directly into the renal artery of anaesthetized rats. Renin secretion was calculated by the renal veno-arterial difference in plasma renin activity multiplied by renal plasma flow. The intra-renal arterial (i.r.a.) infusion of oxytocin (1.5 or 15 ng/kg/min, 10 min) induced a sixfold increase in renin secretion as compared to vehicle-treated controls, without effects on renal blood flow, mean arterial blood pressure, glomerular filtration rate or natriuresis. The effect of oxytocin (1.5 ng/kg/min) was prevented by pretreatment with an oxytocin receptor antagonist, desGly-NH(2),d(CH(2))(5)[D-Tyr(2),Thr(4),Orn(8)]vasotocin] (5.6 microg/kg bolus i.v. 20 min before oxytocin infusion, followed by 2.8 microg/kg/min i.r.a.). Nadolol (2.5 mg/kg i.v.), a beta-adrenoceptor antagonist, also blocked the oxytocin-induced increase in renin secretion. These results show that oxytocin is able to stimulate renin release by activating oxytocin receptors but that beta-adrenoceptors also seem to be involved.

Shear stress modulates vasopressin-induced renal vasoconstriction in rats.

Vasopressin is a potent renal vasoconstrictor in vitro which elicits relatively minor renal vascular effects in vivo. Efficient modulation might occur through shear stress-elicited release of vasodilator compounds from endothelial cells. The aim of this study was to evaluate in vitro, in the isolated perfused kidney, the influence of shear stress and related nitric oxide (NO) release on basal renal vascular tone and on vasopressin-induced renal vasoconstriction. Rat kidneys were perfused at a constant flow rate of 8 ml/min with Tyrode's solution (relative viscosity eta=1) or, in order to increase shear stress, with Tyrode's solution supplemented with 4.7% Ficoll 400 (Ficoll 400; eta=2.3), which is representative of the relative viscosity found in small vessels. Renal shear stress was further elevated during vasoconstriction elicited by vasopressin. Basal renal true vascular conductance, which reflects mean blood vessel radius, was 2.5-fold higher and overall wall shear stress doubled in Ficoll 400 - as compared to Tyrode-perfused kidneys. The decrease in vascular conductance elicited by NO synthase inhibition with N(omega)-nitro-L-arginine (L-NNA) increased with the viscosity of the perfusate. Shear stress was elevated during vasopressin-induced renal vasoconstriction, all the more kidneys were Ficoll 400-perfused. In these kidneys, the concentration-response curve to vasopressin was shifted to the right, giving evidence of hyporeactivity to the peptide. L-NNA potentiated vasoconstriction to vasopressin particularly in Ficoll 400-perfused kidneys, although additional inhibition of cyclooxygenase and/or cytochrome P(450) was without effect. These results provide in vitro evidence that shear stress enhanced by perfusate viscosity increases basal renal vascular conductance by an NO-dependent mechanism. Together with shear stress enhanced during vasoconstriction, it blunts vasopressin-induced renal vasoconstriction.