[How to assess the impact of morbimortality conferences on healthcare quality and safety in ICU ?]. / Comment évaluer l'impact des revues de morbi-mortalité sur la qualité et la sécurité des soins en réanimation ?

OBJECTIVE: To estimate the morbidity and mortality conferences (MMC) impact in intensive care unit (ICU) setting on quality of care and patients' safety. DATA SOURCES: A review of English and French articles in Medline database (1990-2011) related to MMC in the ICU. Keywords used: "morbidity (and) mortality conference(s)", "intensive care unit", "intensive/critical care medicine". Additional studies identified by hand search in French national guidelines about MMCs and in the Annales Françaises d'Anesthésie Réanimation and Réanimation journals index. Identification and preliminary analysis performed using title and abstract, for every study related to MMC in the ICU. STUDY SELECTION: Only original studies about MMC in the ICU setting that reported an assessment were included. Papers reporting guidelines and methods for MMC implementation were excluded. DATA EXTRACTION: Extraction used predefined data fields, including study design, MMC characteristics, assessment methods and results. DATA SYNTHESIS: Studies about MMC in the ICU are recent and scarce. Results comparison and synthesis are impaired by discrepancies in study designs. Although the effectiveness of MMC is not evidence-based, data are consistent for their positive impact on quality of care and patient safety in the ICU. CONCLUSION: Further studies are required to assess the impact of MMC in the ICU. Based on this literature review, a 4-level evaluation scheme can be suggested: 1) evaluation of MMC implementation in care units and facilities; 2) evaluation of MMC organization; 3) evaluation of MMC on quality of care; 4) evaluation of MMC impact on patients' mortality and morbidity.

Is phosphorylation of the alpha1 subunit at Ser-16 involved in the control of Na,K-ATPase activity by phorbol ester-activated protein kinase C?

The alpha1 subunit of Na,K-ATPase is phosphorylated at Ser-16 by phorbol ester-sensitive protein kinase(s) C (PKC). The role of Ser-16 phosphorylation was analyzed in COS-7 cells stably expressing wild-type or mutant (T15A/S16A and S16D-E) ouabain-resistant Bufo alpha1 subunits. In cells incubated at 37 degrees C, phorbol 12, 13-dibutyrate (PDBu) inhibited the transport activity and decreased the cell surface expression of wild-type and mutant Na,K-pumps equally ( approximately 20-30%). This effect of PDBu was mimicked by arachidonic acid and was dependent on PKC, phospholipase A(2), and cytochrome P450-dependent monooxygenase. In contrast, incubation of cells at 18 degrees C suppressed the down-regulation of Na,K-pumps and revealed a phosphorylation-dependent stimulation of the transport activity of Na,K-ATPase. Na,K-ATPase from cells expressing alpha1-mutants mimicking Ser-16 phosphorylation (S16D or S16E) exhibited an increase in the apparent Na affinity. This finding was confirmed by the PDBu-induced increase in Na sensitivity of the activity of Na,K-ATPase measured in permeabilized nontransfected COS-7 cells. These results illustrate the complexity of the regulation of Na,K-ATPase alpha1 isozymes by phorbol ester-sensitive PKCs and reveal 1) a phosphorylation-independent decrease in cell surface expression and 2) a phosphorylation-dependent stimulation of the transport activity attributable to an increase in the apparent Na affinity.

Identification of plasma proteins adsorbed on hemodialysis tubing that promote Staphylococcus aureus adhesion.

Risk factors for Staphylococcus aureus infections in patients undergoing hemodialysis include underlying disease, material-induced host defects, and the presence of vascular access catheters. To determine the specific contribution of various potentially adsorbed plasma components in promoting S aureus adhesion to shunt tubing during chronic hemodialysis, we quantified their respective amounts by Western immunoblotting and densitometry and estimated their individual adhesion-promoting activities with specific adhesion-modified bacterial mutants. Fibrinogen, which was the only component consistently present in tubing protein extracts from all patients, was adsorbed in significantly higher amounts on predialyzer than on postdialyzer tubing segments. In contrast, fibronectin and von Willebrand factor were irregularly present in patients' tubing, whereas vitronectin or thrombospondin remained undetectable in all samples. The contribution of fibrinogen in promoting S aureus attachment to hemodialysis tubing was demonstrated by (1) the significantly lower adhesion of a cIfA mutant of strain Newman compared with its parent; (2) the increased attachment of strain 8325-4 after complementation with the cloned cIfA gene on the multicopy plasmid pCF4; and (3) the general tendency for strains Newman and 8325-4(pCF4) to express higher attachment on predialyzer compared with postdialyzer tubing segments in relationship with the higher content of fibrinogen on the former material. However, the specific S aureus attachment-promoting activity of both prefilter and postfilter tubing-adsorbed fibrinogen were much lower than that of the native in vitro-adsorbed protein and may reflect masking or inactivation of the in vivo-adsorbed protein by unknown mechanisms.

From the molecular biology of prolactin and its receptor to the lessons learned from knockout mice models.

Prolactin (PRL), a polypeptide hormone secreted mainly by the pituitary and, to a lesser extent, by peripheral tissues, affects more physiological processes than all other pituitary hormones combined since it is involved in > 300 separate functions in vertebrates. Its main actions are related to lactation and reproduction. The initial step of PRL action is the binding to a specific membrane receptor, the PRLR, which belongs to the class 1 cytokine receptor superfamily. PRL-binding sites have been identified in a number of tissues and cell types in adult animals. Signal transduction by this receptor is mediated, at least in part, by two families of signaling molecules: Janus tyrosine kinases and signal transducers and activators of transcription (STATs). Disruption of the PRLR gene has provided a new mouse model with which to identify actions directly associated with PRL or any other PRLR ligands, such as placental lactogens. To date, several different phenotypes have been analyzed and are briefly described in this review. Coupled with the SAGE technique, this PRLR knockout model is being used to qualitatively and quantitatively evaluate the expression pattern of hepatic genes in two physiological situations: transcriptomes corresponding to livers from both wild type and PRLR KO mice are being compared, and following statistical analyses, candidate genes presenting a differential profile will be further characterized. Such a new approach will undoubtedly open future avenues of research for PRL targets. To date, no pathology linked to any mutation in the genes encoding PRL or its receptor have been identified. The development of genetic models provides new opportunities to understand how PRL can participate to the development of pathologies throughout life, as for example the initiation and progression of breast cancer.

Cell shrinkage triggers the activation of mitogen-activated protein kinases by hypertonicity in the rat kidney medullary thick ascending limb of the Henle's loop. Requirement of p38 kinase for the regulatory volume increase response.

The kidney medulla is exposed to very high interstitial osmolarity leading to the activation of mitogen-activated protein kinases (MAPK). However, the respective roles of increased intracellular osmolality and of cell shrinkage in MAPK activation are not known. Similarly, the participation of MAPK in the regulatory volume increase (RVI) following cell shrinkage remains to be investigated. In the rat medullary thick ascending limb of Henle (MTAL), extracellular hypertonicity produced by addition of NaCl or sucrose increased the phosphorylation level of extracellular signal-regulated kinase (ERK) and p38 kinase and to a lesser extent c-Jun NH(2)-terminal kinase with sucrose only. Both hypertonic solutions decreased the MTAL cellular volume in a dose- and time-dependent manner. In contrast, hypertonic urea had no effect. The extent of MAPK activation was correlated with the extent of MTAL cellular volume decrease. Increasing intracellular osmolality without modifying cellular volume did not activate MAPK, whereas cell shrinkage without variation in osmolality activated both ERK and p38. In the presence of 600 mosmol/liter NaCl, the maximal cell shrinkage was observed after 10 min at 37 degrees C and the MTAL cellular volume was reduced to 70% of its initial value. Then, RVI occurred and the cellular volume progressively recovered to reach about 90% of its initial value after 30 min. SB203580, a specific inhibitor of p38, almost completely inhibited the cellular volume recovery, whereas inhibition of ERK did not alter RVI. In conclusion, in rat MTAL: 1) cell shrinkage, but not intracellular hyperosmolality, triggers the activation of both ERK and p38 kinase in response to extracellular hypertonicity; and 2) RVI is dependent on p38 kinase activation.

Glomerulonephritis and sodium retention: enhancement of Na+/K+-ATPase activity in the collecting duct is shared by rats with puromycin induced nephrotic syndrome and mice with spontaneous lupus-like glomerulonephritis.

BACKGROUND: In rats with puromycin aminoglucoside-induced (PAN) nephrotic syndrome, micropuncture studies have localized the site of sodium retention to the collecting duct. We have confirmed this finding by demonstrating a two-fold increase in Na+/K+-ATPase activity specifically limited to the cortical collecting duct in PAN rats. To further define whether this phenomenon was dependent on the chemical induction of the nephrotic syndrome or was a general phenomenon observed in glomerulonephritis, we measured Na+/K+-ATPase activity in nephron segments from mice with spontaneous lupus-like nephritis. METHODS: Hydrolytic activity of Na+/K+-ATPase was measured in three isolated nephron segments: proximal convoluted tubule, thick ascending limb and cortical collecting duct. The Na+/K+-ATPase activities were measured in PAN rats, sham-injected controls, and in (MRL x BXSB) F1 male mice which develop a well established spontaneous lupus-like glomerulonephritis by 4 months of age and their controls. Control mice have the same genetic background, but lack the Yaa mutant gene responsible for autoimmune acceleration and are free of glomerular lesions at 4 months of age. RESULTS: In (MRL x BXSB) F1 male mice, Na+/K+-ATPase was similar to control mice in the proximal convoluted tubule and the thick ascending limb. In contrast, cortical collecting duct Na+/K+-ATPase activity was two times higher in (MRL x BXSB) F1 mice than controls. These results were identical to those observed in PAN rats compared to their sham-injected controls studied 7 days after an intraperitoneal injection of puromycin or isotonic saline, respectively. CONCLUSIONS: Enhancement of Na+/K+-ATPase activity localized to the cortical collecting duct is a general characteristic of glomerulonephritis independent of its mode of induction, i.e. chemical versus autoimmune. Therefore, the experimental model of PAN is suitable to study the underlying mechanisms leading to Na+/K+-ATPase dysfunction.

Insulin-induced stimulation of Na+,K(+)-ATPase activity in kidney proximal tubule cells depends on phosphorylation of the alpha-subunit at Tyr-10.

Phosphorylation of the alpha-subunit of Na+,K(+)-ATPase plays an important role in the regulation of this pump. Recent studies suggest that insulin, known to increase solute and fluid reabsorption in mammalian proximal convoluted tubule (PCT), is stimulating Na+,K(+)-ATPase activity through the tyrosine phosphorylation process. This study was therefore undertaken to evaluate the role of tyrosine phosphorylation of the Na+,K(+)-ATPase alpha-subunit in the action of insulin. In rat PCT, insulin and orthovanadate (a tyrosine phosphatase inhibitor) increased tyrosine phosphorylation level of the alpha-subunit more than twofold. Their effects were not additive, suggesting a common mechanism of action. Insulin-induced tyrosine phosphorylation was prevented by genistein, a tyrosine kinase inhibitor. The site of tyrosine phosphorylation was identified on Tyr-10 by controlled trypsinolysis in rat PCTs and by site-directed mutagenesis in opossum kidney cells transfected with rat alpha-subunit. The functional relevance of Tyr-10 phosphorylation was assessed by 1) the abolition of insulin-induced stimulation of the ouabain-sensitive (86)Rb uptake in opossum kidney cells expressing mutant rat alpha1-subunits wherein tyrosine was replaced by alanine or glutamine; and 2) the similarity of the time course and dose dependency of the insulin-induced increase in ouabain-sensitive (86)Rb uptake and tyrosine phosphorylation. These findings indicate that phosphorylation of the Na+,K(+)-ATPase alpha-subunit at Tyr-10 likely participates in the physiological control of sodium reabsorption in PCT.

Inhibition and restoration of prolactin signal transduction by suppressors of cytokine signaling.

Prolactin (PRL) has been shown to activate the cytoplasmic tyrosine kinase Janus kinase 2 (Jak2) and the subsequent recruitment of various signaling molecules including members of the signal transducer and activator of transcription family of transcription factors. Recently, an expanding family of cytokine-inducible inhibitors of signaling has been identified that initially included four members: suppressor of cytokine signaling (SOCS)-1, SOCS-2, SOCS-3, and cytokine-inducible src homology domain 2 (SH-2) proteins. The present study analyzes the role of these members in PRL signaling. Constitutive expression of SOCS-1 and SOCS-3 suppressed PRL-induced signal transducer and activator of transcription 5-dependent gene transcription, and Jak2 tyrosine kinase activity was greatly reduced in the presence of SOCS-1 or SOCS-3. SOCS-1 was shown to associate with Jak2, whereas SOCS-2 was associated with the prolactin receptor. Co-transfection studies were conducted to further analyze the interactions of SOCS proteins. SOCS-2 was shown to suppress the inhibitory effect of SOCS-1 by restoring Jak2 kinase activity but did not affect the inhibitory effect of SOCS-3 on PRL signaling. Northern blot analysis revealed that SOCS-3 and SOCS-1 genes were transiently expressed in response to PRL, both in vivo and in vitro, whereas the expression of SOCS-2 and CIS genes was still elevated 24 h after hormonal stimulation. We thus propose that the early expressed SOCS genes (SOCS-1 and SOCS-3) switch off PRL signaling and that the later expressed SOCS-2 gene can restore the sensitivity of cells to PRL, partly by suppressing the SOCS-1 inhibitory effect.

Improvement of drug-induced chronic renal failure in lung transplantation.

BACKGROUND: Nephrotoxicity is a frequently encountered adverse effect of calcineurin inhibitors (cyclosporine and tacrolimus)-combined immunosuppressive regimens. METHODS: We have compared the glomerular filtration rate in 14 patients who underwent lung transplantation, before and after replacement of azathioprine by mycophenolate mofetil and reduction of associated calcineurin inhibitors doses. RESULTS: After a mean follow-up of 16+/-4 months with the modified immunosuppressive regimen, the mean glomerular filtration rate increased by 20% with no change in lung function. CONCLUSION: By its strong immunosuppressive effect, mycophenolate mofetil allows the decrease of associated calcineurin inhibitor doses, with subsequent improvement of renal function without jeopardizing the transplanted lung.

Dual effects of suppressor of cytokine signaling (SOCS-2) on growth hormone signal transduction.

A family of suppressors of cytokine signaling (SOCS) has recently been identified of which two members have been shown to block growth hormone (GH) signaling. Dose-response experiments were conducted in 293 cells and SOCS-1 and SOCS-3 were shown to inhibit the transcriptional activation of a GH-responsive element and suppressed Jak2 tyrosine kinase activity. SOCS-2 had two opposite effects: at low concentrations it inhibited GH-induced STAT5-dependent gene transcription, but restoration of GH signaling was observed at higher concentrations. In cotransfection studies, SOCS-2 was able to block the inhibitory effect of SOCS-1 but not that of SOCS-3 on GH signaling. These findings suggest that a major function for SOCS-2 is to restore the sensitivity to GH by overcoming the initial inhibitory effects of other endogenous SOCS molecules.

Effect of cAMP on the activity and the phosphorylation of Na+,K(+)-ATPase in rat thick ascending limb of Henle.

BACKGROUND: In rat kidney medullary thick ascending limb of Henle's loop (MTAL), activation of protein kinase A (PKA) was previously reported to inhibit Na+,K(+)-ATPase activity. This is paradoxical with the known stimulatory effect of cAMP on sodium reabsorption. Because this inhibition was mediated by phospholipase A2 (PLA2) activation, a pathway stimulated by hypoxia, we evaluated the influence of oxygen supply on cAMP action on Na+,K(+)-ATPase in MTAL. METHODS: Ouabain-sensitive 86Rb uptake and Na+,K(+)-ATPase activity were measured in isolated MTALs. Cellular ATP content and the phosphorylation level of Na+,K(+)-ATPase were determined in suspensions of outer medullary tubules. Experiments were carried out under nonoxygenated or oxygenated conditions in the absence or presence of PKA activators. RESULTS: cAMP analogues or forskolin associated with 3-isobutyl-1-methylxanthine (IBMX) inhibited ouabain-sensitive 86Rb uptake in nonoxygenated MTALs. In contrast, when oxygen supply was increased, cAMP stimulated ouabain-sensitive 86Rb uptake and Na+,K(+)-ATPase activity. Improved oxygen supply was associated with increased intracellular ATP content. The phosphorylation level of the Na+,K(+)-ATPase alpha subunit was increased by cAMP analogues or forskolin associated with IBMX in oxygenated as well as in nonoxygenated tubules. Under nonoxygenated conditions, the inhibition of Na+,K(+)-ATPase was dissociated from its cAMP-dependent phosphorylation, whereas under oxygenated conditions, the stimulatory effect of cAMP analogues on ouabain-sensitive 86Rb uptake was linearly related and cosaturated with the level of phosphorylation of the Na+,K(+)-ATPase alpha subunit. CONCLUSION: In oxygenated MTALs, PKA-mediated stimulation of Na+,K(+)-ATPase likely participates in the cAMP-dependent stimulation of sodium reabsorption. Under nonoxygenated conditions, this stimulatory pathway is likely overridden by the PLA2-mediated inhibitory pathway, a possible adaptation to protect the cells against hypoxic damage.

Protein kinase A induces recruitment of active Na+,K+-ATPase units to the plasma membrane of rat proximal convoluted tubule cells.

1. The aim of this study was to investigate the mechanism of control of Na+,K+-ATPase activity by the cAMP-protein kinase A (PKA) pathway in rat proximal convoluted tubules. For this purpose, we studied the in vitro action of exogenous cAMP (10-3 M dibutyryl-cAMP (db-cAMP) or 8-bromo-cAMP) and endogenous cAMP (direct activation of adenylyl cyclases by 10-5 M forskolin) on Na+,K+-ATPase activity and membrane trafficking. 2. PKA activation stimulated both the cation transport and hydrolytic activity of Na+,K+-ATPase by about 40%. Transport activity stimulation was specific to the PKA signalling pathway since (1) db-cAMP stimulated the ouabain-sensitive 86Rb+ uptake in a time- and dose-dependent fashion; (2) this effect was abolished by addition of H-89 or Rp-cAMPS, two structurally different PKA inhibitors; and (3) this stimulation was not affected by inhibition of protein kinase C (PKC) by GF109203X. The stimulatory effect of db-cAMP on the hydrolytic activity of Na+,K+-ATPase was accounted for by an increased maximal ATPase rate (Vmax) without alteration of the efficiency of the pump, suggesting that cAMP-PKA pathway was implicated in membrane redistribution control. 3. To test this hypothesis, we used two different approaches: (1) cell surface protein biotinylation and (2) subcellular fractionation. Both approaches confirmed that the cAMP-PKA pathway was implicated in membrane trafficking regulation. The stimulation of Na+,K+-ATPase activity by db-cAMP was associated with an increase (+40%) in Na+, K+-ATPase units expressed at the cell surface which was assessed by Western blotting after streptavidin precipitation of biotinylated cell surface proteins. Subcellular fractionation confirmed the increased expression in pump units at the cell surface which was accompanied by a decrease (-30%) in pump units located in the subcellular fraction corresponding to early endosomes. 4. In conclusion, PKA stimulates Na+,K+-ATPase activity, at least in part, by increasing the number of Na+-K+ pumps in the plasma membrane in proximal convoluted tubule cells.

[Choice of renal replacement therapy]. / Choix de la technique d'épuration extra-rénale.

Renal replacement therapies (RRT) for acute renal failure patients consist of continuous or intermittent methods. Continuous renal replacement therapies (CRRT) are used in 54% of centers. Determinant for the choice of the continuous versus intermittent method is the clinical status of the patients. Those who are hemodynamically unstable, who receive a large fluid intake, who are anuric, who present with severe and persistent metabolic disorders such as acidosis or hyperkaliemia, and who are in respiratory failure, are preferably treated with CRRT. Once the patient has improved, he can be switched to intermittent RRT (IRRT). Thus CRRT and IRRT are complementary approaches. Dialysis material should be biocompatible; the RRT method used should guarantee stable hemodynamics. Coping with these two requirements reduces the time duration of renal recovery, diminishes the need for catecholamines and may improve the survival rate.

Mitochondrial anomalies in a Swiss family with autosomal dominant myoglobinuria.

We report on a Swiss family in which 10 individuals of both sexes in 4 successive generations suffered from myoglobinuria, precipitated by febrile illness. It is the second family described with autosomal dominant inheritance of myoglobinuria. Four individuals suffered acute renal failure, which in two was reversible only after dialysis. In a recent case, a mitochondrial disorder was suspected because of an abnormal increase in lactate levels during an exercise test and because of a subsarcolemmal accumulation of mitochondria in a muscle biopsy, associated with a lack of cytochrome C oxidase in some muscle fibers. No mutation in the mitochondrial DNA was identified. Along with the inheritance pattern, these findings suggest that the myoglobinuria in this family is caused by a nuclear-encoded mutation affecting the respiratory chain.

Modulation of Na+,K(+)-ATPase activity by a tyrosine phosphorylation process in rat proximal convoluted tubule.

1. In the rat kidney proximal convoluted tubule, epidermal growth factor and insulin have been reported to stimulate Na+ reabsorption. Because most of the effects of these growth factors are mediated by a process of tyrosine phosphorylation and Na+,K(+)-ATPase drives Na+ reabsorption, the influence of tyrosine kinases and tyrosine phosphatases on Na+,K(+)-ATPase activity located in the proximal convoluted tubule was evaluated. 2. Activation of receptor tyrosine kinases by epidermal growth factor and insulin stimulated ouabain-sensitive 86Rb+ uptake. The effects of epidermal growth factor and insulin were prevented by genistein, a tyrosine kinase inhibitor, but were unaffected by GF109203X, a protein kinase C inhibitor. 3. Inhibition of tyrosine phosphatases by orthovanadate (10(-7) and 10(-6)M) mimicked the effects of activation of receptor tyrosine kinases: stimulation of the ouabain-sensitive 86Rb+ uptake and of the hydrolytic activity of Na+,K(+)-ATPase under rate-limiting Na+ concentration, and absence of modification of the maximal activity (Vmax) of the enzyme. The effects of orthovanadate and insulin on the ouabain-sensitive 86Rb+ uptake were not additive. 4. The present results show that both activation of receptor tyrosine kinases and inhibition of tyrosine phosphatases stimulate the Na+,K(+)-ATPase activity through a common mechanism. Thus, a tyrosine phosphorylation process directly controls the Na+,K(+)-ATPase activity and contributes to the physiological control of water and solute reabsorption in the proximal convoluted tubule.

Haemodynamic and endocrinological effects of noninvasive mechanical ventilation in respiratory failure.

The aim of this study was to investigate the haemodynamic and endocrinological effects of noninvasive positive pressure ventilation (NIPPV). Eleven patients with oedema and recent hypercapnic and hypoxaemic worsening of a chronic respiratory insufficiency were included. Echocardiography, cardiac radionuclide assessment, blood catecholamines, salt and water handling hormones were measured at admission and discharge (long study (LS)). To discriminate between the action of NIPPV and other treatments, measurements were performed on the fourth day, for 4 h without NIPPV and 4 h with NIPPV (short study (SS)). NIPPV entailed a correction of P(a,CO2) and an increase of P(a,O2) in LS and SS. Oedema disappeared. Body weight decreased (from 85+/-42 to 81+/-40 kg) during LS. Systolic and mean pulmonary arterial pressure decreased in LS and SS. Right ventricular ejection fraction increased in LS. Left ventricular ejection fraction did not change. Cardiac index was normal on admission and then decreased. Natriuretic peptides and catecholamines were increased on admission, whereas plasma renin activity, aldosterone and vasopressin were normal. We suggest that in these patients, oedema can occur independently of renin-angiotensin-aldosterone-vasopressin and with a normal cardiac output. Noninvasive positive pressure ventilation allowed a correction of blood gases, associated with the resolution of oedema, a decrease in pulmonary arterial pressures and an increase in right ventricular ejection fraction.

Stimulation of ouabain-sensitive 86Rb+ uptake and Na+,K+-ATPase alpha-subunit phosphorylation by a cAMP-dependent signalling pathway in intact cells from rat kidney cortex.

We investigated in intact cortical kidney tubules the role of PKA-mediated phosphorylation in the short-term control of Na+,K+-ATPase activity. The phosphorylation level of Na+,K+-ATPase was evaluated after immunoprecipitation of the enzyme from 32P-labelled cortical tubules and the cation transport activity of Na+,K+-ATPase was measured by ouabain-sensitive 86Rb+ uptake. Incubation of cells with cAMP analogues (8-bromo-cAMP, dibutyryl-cAMP) or with forskolin plus 3-isobutyl-1-methylxanthine increased the phosphorylation level of the Na+,K+-ATPase alpha-subunit and stimulated ouabain-sensitive 86Rb+ uptake. Inhibition of PKA by H-89 blocked the effects of dibutyryl-cAMP on both phosphorylation and 86Rb+ uptake processes. The results suggest that phosphorylation by PKA stimulates the Na+,K+-ATPase activity.

Kinetics of cellular viability in warm versus cold ischemia conditions of kidney preservation. A biometric study.

We have determined the kinetics of the cellular viability ratio (CVR), defined as the number of living cells over the total cell count, in pig kidneys using propidium iodide and fluorescein diacetate staining, as a function of time and preservation conditions. The kidneys were preserved in warm or cold ischemia in order to mimic the conditions of transplantation from non-heart-beating donors or multiple removal with optimal preservation of the graft, respectively. To determine the CVR, the cells were obtained by a fine-needle aspiration biopsy, which minimizes the damage to the graft. A biometric analysis by regression enabled the determination of the time dependence for warm ischemia (CVR(t) = 80.0 x e(-0.733-t)(+2.7/-0.36)) and for cold ischemia (CVR(t) = 80.0 x e(-0.022-t)(+1.57/-0.64)) with a confidence interval of 95%. These master curves allow us to predict, under the described conditions, the CVR after a given ischemia time. The half-life of the cells can be deduced from the time-dependent CVR(t), and is 0.64 hr (38 min) for warm ischemia and 21.4 hr for cold ischemia. Further, the CVR for a given kidney can be used to assess its condition at removal: if the CVR is below 48% at 2 hr after removal, one can conclude that the organ has suffered a period of warm ischemia.