The role of phospholipase D in modulating the MTOR signaling pathway in polycystic kidney disease

The mammalian target of rapamycin (mTOR) signaling pathway is aberrantly activated in polycystic kidney disease (PKD). Emerging evidence suggests that phospholipase D (PLD) and its product phosphatidic acid (PA) regulate mTOR activity. In this study, we assessed in vitro the regulatory function of PLD and PA on the mTOR signaling pathway in PKD. We found that the basal level of PLD activity was elevated in PKD cells. Targeting PLD by small molecule inhibitors reduced cell proliferation and blocked mTOR signaling, whereas exogenous PA stimulated mTOR signaling and abolished the inhibitory effect of PLD on PKD cell proliferation. We also show that blocking PLD activity enhanced the sensitivity of PKD cells to rapamycin and that combining PLD inhibitors and rapamycin synergistically inhibited PKD cell proliferation. Furthermore, we demonstrate that targeting mTOR did not induce autophagy, whereas targeting PLD induced autophagosome formation. Taken together, our findings suggest that deregulated mTOR pathway activation is mediated partly by increased PLD signaling in PKD cells. Targeting PLD isoforms with pharmacological inhibitors may represent a new therapeutic strategy in PKD.

Re-thinking resuscitation: leaving blood pressure cosmetics behind and moving forward to permissive hypotension and a tissue perfusion-based approach

Definitions of shock and resuscitation endpoints traditionally focus on blood pressures and cardiac output. This carries a high risk of overemphasizing systemic hemodynamics at the cost of tissue perfusion. In line with novel shock definitions and evidence of the lack of a correlation between macro- and microcirculation in shock, we recommend that macrocirculatory resuscitation endpoints, particularly arterial and central venous pressure as well as cardiac output, be reconsidered. In this viewpoint article, we propose a three-step approach of resuscitation endpoints in shock of all origins. This approach targets only a minimum individual and context-sensitive mean arterial blood pressure (for example, 45 to 50 mm Hg) to preserve heart and brain perfusion. Further resuscitation is exclusively guided by endpoints of tissue perfusion irrespectively of the presence of arterial hypotension ('permissive hypotension'). Finally, optimization of individual tissue (for example, renal) perfusion is targeted. Prospective clinical studies are necessary to confirm the postulated benefits of targeting these resuscitation endpoints.

Contrast-enhanced ultrasound for myocardial perfusion imaging.

Ultrasound contrast agents are gas-filled microbubbles that enhance visualization of cardiac structures, function and blood flow during contrast-enhanced ultrasound (CEUS). An interesting cardiovascular application of CEUS is myocardial contrast echocardiography, which allows real-time myocardial perfusion imaging. The intraoperative use of this technically challenging imaging method is limited at present, although several studies have examined its clinical utility during cardiac surgery in the past. In the present review we provide general information on the basic principles of CEUS and discuss the methodology and technical aspects of myocardial perfusion imaging.

HDL Cholesterol Is Not Associated with Lower Mortality in Patients with Kidney Dysfunction

In the general population, HDL cholesterol (HDL-C) is associated with reduced cardiovascular events. However, recent experimental data suggest that the vascular effects of HDL can be heterogeneous. We examined the association of HDL-C with all-cause and cardiovascular mortality in the Ludwigshafen Risk and Cardiovascular Health study comprising 3307 patients undergoing coronary angiography. Patients were followed for a median of 9.9 years. Estimated GFR (eGFR) was calculated using the Chronic Kidney Disease Epidemiology Collaboration eGFR creatinine-cystatin C (eGFRcreat-cys) equation. The effect of increasing HDL-C serum levels was assessed using Cox proportional hazard models. In participants with normal kidney function (eGFR>90 ml/min per 1.73 m(2)), higher HDL-C was associated with reduced risk of all-cause and cardiovascular mortality and coronary artery disease severity (hazard ratio [HR], 0.51, 95% confidence interval [95% CI], 0.26-0.92 [P=0.03]; HR, 0.30, 95% CI, 0.13-0.73 [P=0.01]). Conversely, in patients with mild (eGFR=60-89 ml/min per 1.73 m(2)) and more advanced reduced kidney function (eGFR<60 ml/min per 1.73 m(2)), higher HDL-C did not associate with lower risk for mortality (eGFR=60-89 ml/min per 1.73 m(2): HR, 0.68, 95% CI, 0.45-1.04 [P=0.07]; HR, 0.84, 95% CI, 0.50-1.40 [P=0.50]; eGFR<60 ml/min per 1.73 m(2): HR, 1.18, 95% CI, 0.60-1.81 [P=0.88]; HR, 0.82, 95% CI, 0.40-1.69 [P=0.60]). Moreover, Cox regression analyses revealed interaction between HDL-C and eGFR in predicting all-cause and cardiovascular mortality (P=0.04 and P=0.02, respectively). We confirmed a lack of association between higher HDL-C and lower mortality in an independent cohort of patients with definite CKD (P=0.63). In summary, higher HDL-C levels did not associate with reduced mortality risk and coronary artery disease severity in patients with reduced kidney function. Indeed, abnormal HDL function might confound the outcome of HDL-targeted therapies in these patients.

The effects of endothelin-1 on coronary perfusion pressure during cardiopulmonary resuscitation in a canine model

OBJECTIVE To study the hemodynamic effects of exogenously administered endothelin-1 (ET-1), a peptide produced by endothelial cells with potent non-adrenergically mediated vasoconstrictor properties. METHODS A prospective drug intervention study was carried out in a resuscitation research laboratory. Fifteen mixed-breed dogs were anesthetized and instrumented for hemodynamic monitoring. Asphyxia arrest was produced by clamping the endotracheal tube. Hemodynamic data were collected continuously. Following loss of aortic fluctuations monitored by thoracic aortic catheter, the animals remained in pulseless electrical activity (PEA) for 10 minutes. After 10 minutes of no-flow PEA, closed-chest CPR was begun and the animals were randomized to one of three treatment groups (EPI, 0.02 mg/kg epinephrine IV every 3 minutes; ENDO, 100 micrograms ET-1 IV at 0 minutes; and EPI/ENDO, a combination of the EPI and ENDO treatments). RESULTS ENDO and EPI alone produced similar coronary perfusion pressures (CPPs). The EPI/ENDO combination produced significantly improved CPP compared with that of either EPI or ENDO alone. In the EPI group, the best mean CPP was 16 +/- 14 mm Hg and occurred 7 minutes after drug administration. In the ENDO group, the best mean CPP was 28 +/- 7 mm Hg and occurred 13 minutes after drug administration. In the EPI/ENDO combination group, the best mean CPP was 61 +/- 37 mm Hg and occurred 7 minutes after drug administration (p < 0.05 compared with the EPI and ENDO groups alone). CONCLUSION ET-1 is a potent vasoconstrictor. The combination of EPI and ENDO significantly improved CPP compared with that for either agent alone. ET-1 should be investigated further as a vasoconstrictor in cardiac arrest.

Plasma and urine neutrophil gelatinase-associated lipocalin (NGAL) in dogs with acute kidney injury or chronic kidney disease.

BACKGROUND Neutrophil gelatinase-associated lipocalin (NGAL) is a protein that is used in human medicine as a real-time indicator of acute kidney injury (AKI). HYPOTHESIS Dogs with AKI have significantly higher plasma NGAL concentration and urine NGAL-to-creatinine ratio (UNCR) compared with healthy dogs and dogs with chronic kidney disease (CKD). ANIMALS 18 healthy control dogs, 17 dogs with CKD, and 48 dogs with AKI. METHODS Over a period of 1 year, all dogs with renal azotemia were prospectively included. Urine and plasma samples were collected during the first 24 hours after presentation or after development of renal azotemia. Plasma and urine NGAL concentrations were measured with a commercially available canine NGAL Elisa Kit (Bioporto® Diagnostic) and UNCR was calculated. A single-injection plasma inulin clearance was performed in the healthy dogs. RESULTS Median (range) NGAL plasma concentration in healthy dogs, dogs with CKD, and AKI were 10.7 ng/mL (2.5-21.2), 22.0 ng/mL (7.7-62.3), and 48.3 ng/mL (5.7-469.0), respectively. UNCR was 2 × 10(-8) (0-46), 1,424 × 10(-8) (385-18,347), and 2,366 × 10(-8) (36-994,669), respectively. Dogs with renal azotemia had significantly higher NGAL concentrations and UNCR than did healthy dogs (P < .0001 for both). Plasma NGAL concentration was significantly higher in dogs with AKI compared with dogs with CKD (P = .027). CONCLUSIONS AND CLINICAL IMPORTANCE Plasma NGAL could be helpful to differentiate AKI from CKD in dogs with renal azotemia.

A novel clinical scoring system for outcome prediction in dogs with acute kidney injury managed by hemodialysis.

BACKGROUND No reliable tool to predict outcome of acute kidney injury (AKI) exists. HYPOTHESIS A statistically derived scoring system can accurately predict outcome in dogs with AKI managed with hemodialysis. ANIMALS One hundred and eighty-two client-owned dogs with AKI. METHODS Logistic regression analyses were performed initially on clinical variables available on the 1st day of hospitalization for relevance to outcome. Variables with P< or = .1 were considered for further analyses. Continuous variables outside the reference range were divided into quartiles to yield quartile-specific odds ratios (ORs) for survival. Models were developed by incorporating weighting factors assigned to each quartile based on the OR, using either the integer value of the OR (Model A) or the exact OR (Models B or C, when the etiology was known). A predictive score for each model was calculated for each dog by summing all weighting factors. In Model D, actual values for continuous variables were used in a logistic regression model. Receiver-operating curve analyses were performed to assess sensitivities, specificities, and optimal cutoff points for all models. RESULTS Higher scores were associated with decreased probability of survival (P < .001). Models A, B, C, and D correctly classified outcomes in 81, 83, 87, and 76% of cases, respectively, and optimal sensitivities/specificities were 77/85, 81/85, 83/90 and 92/61%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE The models allowed outcome prediction that corresponded with actual outcome in our cohort. However, each model should be validated further in independent cohorts. The models may also be useful to assess AKI severity.

Congenital Hepatic Fibrosis in the Franches-Montagnes Horse Is Associated with the Polycystic Kidney and Hepatic Disease 1 (PKHD1) Gene

Congenital hepatic fibrosis has been described as a lethal disease with monogenic autosomal recessive inheritance in the Swiss Franches-Montagnes horse breed. We performed a genome-wide association study with 5 cases and 12 controls and detected an association on chromosome 20. Subsequent homozygosity mapping defined a critical interval of 952 kb harboring 10 annotated genes and loci including the polycystic kidney and hepatic disease 1 (autosomal recessive) gene (PKHD1). PKHD1 represents an excellent functional candidate as variants in this gene were identified in human patients with autosomal recessive polycystic kidney and hepatic disease (ARPKD) as well as several mouse and rat mutants. Whereas most pathogenic PKHD1 variants lead to polycystic defects in kidney and liver, a small subset of the human ARPKD patients have only liver symptoms, similar to our horses with congenital hepatic fibrosis. The PKHD1 gene is one of the largest genes in the genome with multiple alternative transcripts that have not yet been fully characterized. We sequenced the genomes of an affected foal and 46 control horses to establish a comprehensive list of variants in the critical interval. We identified two missense variants in the PKHD1 gene which were strongly, but not perfectly associated with congenital hepatic fibrosis. We speculate that reduced penetrance and/or potential epistatic interactions with hypothetical modifier genes may explain the imperfect association of the detected PKHD1 variants. Our data thus indicate that horses with congenital hepatic fibrosis represent an interesting large animal model for the liver-restricted subtype of human ARPKD.

Interhemispheric cerebral blood flow balance during recovery of motor hand function after ischemic stroke - a longitudinal MRI study using arterial spin labeling perfusion

BACKGROUND Unilateral ischemic stroke disrupts the well balanced interactions within bilateral cortical networks. Restitution of interhemispheric balance is thought to contribute to post-stroke recovery. Longitudinal measurements of cerebral blood flow (CBF) changes might act as surrogate marker for this process. OBJECTIVE To quantify longitudinal CBF changes using arterial spin labeling MRI (ASL) and interhemispheric balance within the cortical sensorimotor network and to assess their relationship with motor hand function recovery. METHODS Longitudinal CBF data were acquired in 23 patients at 3 and 9 months after cortical sensorimotor stroke and in 20 healthy controls using pulsed ASL. Recovery of grip force and manual dexterity was assessed with tasks requiring power and precision grips. Voxel-based analysis was performed to identify areas of significant CBF change. Region-of-interest analyses were used to quantify the interhemispheric balance across nodes of the cortical sensorimotor network. RESULTS Dexterity was more affected, and recovered at a slower pace than grip force. In patients with successful recovery of dexterous hand function, CBF decreased over time in the contralesional supplementary motor area, paralimbic anterior cingulate cortex and superior precuneus, and interhemispheric balance returned to healthy control levels. In contrast, patients with poor recovery presented with sustained hypoperfusion in the sensorimotor cortices encompassing the ischemic tissue, and CBF remained lateralized to the contralesional hemisphere. CONCLUSIONS Sustained perfusion imbalance within the cortical sensorimotor network, as measured with task-unrelated ASL, is associated with poor recovery of dexterous hand function after stroke. CBF at rest might be used to monitor recovery and gain prognostic information.

Relative indexes of cutaneous blood perfusion measured by real-time laser Doppler imaging (LDI) in healthy volunteers.

We used real-time LDI to study regional variations in microcirculatory perfusion in healthy candidates to establish a new methodology for global perfusion body mapping that is based on intra-individual perfusion index ratios. Our study included 74 (37 female) healthy volunteers aged between 22 and 30 years (mean 24.49). Imaging was performed using a recent microcirculation-imaging camera (EasyLDI) for different body regions of each volunteer. The perfusion values were reported in Arbitrary Perfusion Units (APU). The relative perfusion indexes for each candidate's body region were then obtained by normalization with the perfusion value of the forehead. Basic parameters such as weight, height, and blood pressure were also measured and analyzed. The highest mean perfusion value was reported in the forehead area (259.21APU). Mean perfusion in the measured parts of the body correlated positively with mean forehead value, while there was no significant correlation between forehead blood perfusion values and room temperature, BMI, systolic blood pressure and diastolic blood pressure (p=0.420, 0.623, 0.488, 0.099, respectively). Analysis of the data showed that perfusion indexes were not significantly different between male and female volunteers except for the ventral upper arm area (p=.001). LDI is a non-invasive, fast technique that opens several avenues for clinical applications. The mean perfusion indexes are useful in clinical practice for monitoring patients before and after surgical interventions. Perfusion values can be predicted for different body parts for patients only by taking the forehead perfusion value and using the perfusion index ratios to obtain expected normative perfusion values.

Glomerular filtration rate, urine production, and fractional clearance of electrolytes in acute kidney injury in dogs and their association with survival.

BACKGROUND Acute kidney injury (AKI) is common in dogs. Few studies have assessed sequential changes in indices of kidney function in dogs with naturally occurring AKI. OBJECTIVE To document sequential changes of conventional indices of renal function, to better define the course of AKI, and to identify a candidate marker for recovery. ANIMALS Ten dogs with AKI. METHODS Dogs were prospectively enrolled and divided into surviving and nonsurviving dogs. Urine production was measured with a closed system for 7 days. One and 24-hour urinary clearances were performed daily to estimate solute excretion and glomerular filtration rate (GFR). Solute excretion was calculated as an excretion ratio (ER) and fractional clearance (FC) based on both the 1- and 24-hour urine collections. RESULTS Four dogs survived and 6 died. At presentation, GFR was not significantly different between the outcome groups, but significantly (P = .03) increased over time in the surviving, but not in the nonsurviving dogs. Fractional clearance of Na decreased significantly over time (20.2-9.4%, P < .0001) in the surviving, but not in the nonsurviving dogs. The ER and FC of solutes were highly correlated (r, 0.70-0.95). CONCLUSION AND CLINICAL IMPACT Excretion ratio might be used in the clinical setting as a surrogate marker to follow trends in solute excretion. Increased GFR, urine production, and decreased FC of Na were markers of renal recovery. The FC of Na is a simple, noninvasive, and cost-effective method that can be used to evaluate recovery of renal function.