Christine Clerici

Comparison of nutritional consequences of conventional therapy of obesity, adjustable gastric banding, and gastric bypass

Background: Roux-en-Y gastric bypass (RYGBP) is more efficient than adjustable gastric banding (AGB) in weight loss and relieving co-morbidities, but nutritional complications of each surgical procedure have been poorly evaluated. Methods: A cross-sectional study was performed to compare nutritional parameters in 201 consecutive obese patients, who had been treated either by conventional behavioral and dietary therapy (CT, n=110) or by bariatric surgery, including 51 AGB and 40 RYGBP. Results: BMI was similar after AGB (36.6 ± 5.3 kg/m2) and RYGBP (35.4 ± 6.3 kg/m2), but patients in the RYGBP group had lost more weight and had less metabolic disturbances than those in the AGB group. On the other hand, the prevalence of nutritional deficits was significantly higher in the RYGBP group than in the 2 other groups (P <0.01), whereas the AGB group did not differ from CT. Particularly, the RYGBP group presented an unexpected high frequency of deficiencies in fat-soluble vitamins. Moreover, vitamin B12, hemoglobin, plasma prealbumin and creatinine concentrations were low in the RYGBP group. Conclusion: RYGBP is more efficient than AGB in correcting obesity, but this operation is associated with a higher frequency of nutritional deficits that should be carefully monitored.

ENaC-mediated alveolar fluid clearance and lung fluid balance depend on the channel-activating protease 1

Sodium transport via epithelial sodium channels (ENaC) expressed in alveolar epithelial cells (AEC) provides the driving force for removal of fluid from the alveolar space. The membrane‐bound channel‐activating protease 1 (CAP1/Prss8) activates ENaC in vitro in various expression systems. To study the role of CAP1/Prss8 in alveolar sodium transport and lung fluid balance in vivo, we generated mice lacking CAP1/Prss8 in the alveolar epithelium using conditional Cre‐loxP‐mediated recombination. Deficiency of CAP1/Prss8 in AEC induced in vitro a 40% decrease in ENaC‐mediated sodium currents. Sodium‐driven alveolar fluid clearance (AFC) was reduced in CAP1/Prss8‐deficient mice, due to a 48% decrease in amiloride‐sensitive clearance, and was less sensitive to β2‐agonist treatment. Intra‐alveolar treatment with neutrophil elastase, a soluble serine protease activating ENaC at the cell surface, fully restored basal AFC and the stimulation by β2‐agonists. Finally, acute volume‐overload increased alveolar lining fluid volume in CAP1/Prss8‐deficient mice. This study reveals that CAP1 plays a crucial role in the regulation of ENaC‐mediated alveolar sodium and water transport and in mouse lung fluid balance.

Gene regulation in the adaptive process to hypoxia in lung epithelial cells

Lung alveolar epithelial cells are normally very well oxygenated but may be exposed to hypoxia in many pathological conditions such as pulmonary edema, acute respiratory distress syndrome, chronic obstructive pulmonary diseases, or in some environmental conditions such ascent to high altitude. The ability of alveolar epithelial cells to cope with low oxygen tensions is crucial to maintain the structural and functional integrity of the alveolar epithelium. Alveolar epithelial cells appear to be remarkably tolerant to oxygen deprivation as they are able to maintain adequate cellular ATP content during prolonged hypoxic exposure when mitochondrial oxidative phosphorylation is limited. This property mostly relies on the ability of the cells to rapidly modify their gene expression program, stimulating the expression of genes involved in anaerobic energy supply and repressing expression of genes involved in some ATP-consuming cellular processes. This adaptive strategy of the cells is mostly, but not entirely, dependent on the expression of hypoxia-inducible factors (HIFs), known to be responsible for orchestrating a large number of hypoxia-sensitive genes. This review focuses on the role of HIF isoforms expressed in alveolar epithelial cells exposed to hypoxia and on the specific hypoxic gene regulation that takes place in alveolar epithelial cells either through HIF-dependent or -independent pathways.

GFR Estimation Using the Cockcroft-Gault, MDRD Study, and CKD-EPI Equations in the Elderly

RESEARCH LETTER GFR Estimation Using the Cockcroft-Gault, MDRD Study, and CKD-EPI Equations in the Elderly To the Editor: Kidney function loss in the elderly results from physiologic aging of the kidney and lifelong pathologic insults; this translates into a high incidence of chronic kidney disease (CKD) and susceptibility to ischemic or toxic renal insults. Determining glomerular filtration rate (GFR) is necessary for diagnosing and classifying CKD and adjusting drug dosage. The Cockcroft-Gault (CG), 4-variable Modification of Diet in Renal Disease (MDRD) Study, and CKD Epidemiology Collaboration (CKD-EPI) equations are the most widely used GFR estimates. They incorporate serum creatinine (SCr) concentration with determinants of its muscular production, including age. Many studies have shown that GFR estimates gave very different results in the elderly, raising concerns about which equation should be used in this population, especially for therapeutic adaptation. The proportion of very old patients in the populations that were used to establish the estimation formulas is very low, especially for the CG equation. Thus, the validity of GFR estimates in the elderly remains a matter of debate. We evaluated and compared performances of the CG, MDRD Study, and CKD-EPI equations in a large cohort of old patients with CKD. Between January 2007 and September 2010, data from all patients 65 years and older referred for GFR measurement to Bichat and Tenon hospitals (Paris, France) were collected. Patients gave their consent for scientific use of anonymous data. Urinary clearance of Cr-EDTA was determined on 6 consecutive 30-

Conditioned media from mesenchymal stromal cells restore sodium transport and preserve epithelial permeability in an in vitro model of acute alveolar injury

Mesenchymal stromal cells (MSCs) or their media (MSC-M) were reported to reverse acute lung injury (ALI)-induced decrease of alveolar fluid clearance. To determine the mechanisms by which MSC-M exert their beneficial effects, an in vitro model of alveolar epithelial injury was created by exposing primary rat alveolar epithelial cells (AECs) to hypoxia (3% O2) plus cytomix, a combination of IL-1β, TNF-α, and IFN-γ. MSC-M were collected from human MSCs exposed for 12 h to either normoxia (MSC-M) or to hypoxia plus cytomix (HCYT-MSC-M). This latter condition was used to model the effect of alveolar inflammation and hypoxia on paracrine secretion of MSCs in the injured lung. Comparison of paracrine soluble factors in MSC media showed that the IL-1 receptor antagonist and prostaglandin E2 were markedly increased while keratinocyte growth factor (KGF) was twofold lower in HCYT-MSC-M compared with MSC-M. In AECs, hypoxia plus cytomix increased protein permeability, reduced amiloride-sensitive short-circuit current (AS-Isc), and also decreased the number of α-epithelial sodium channel (α-ENaC) subunits in the apical membrane. To test the effects of MSC media, MSC-M and HCYT-MSC-M were added for an additional 12 h to AECs exposed to hypoxia plus cytomix. MSC-M and HCYT-MSC-M completely restored epithelial permeability to normal. MSC-M, but not HCYT-MSC-M, significantly prevented the hypoxia plus cytomix-induced decrease of ENaC activity and restored apical α-ENaC channels. Interestingly, KGF-deprived MSC-M were unable to restore amiloride-sensitive sodium transport, indicating a possible role for KGF in the beneficial effect of MSC-M. These results indicate that MSC-M may be a preferable therapeutic option for ALI.

Sodium-dependent phosphate and alanine transports but sodium-independent hexose transport in type II alveolar epithelial cells in primary culture.

Inorganic phosphate, amino acids and sugars are of obvious importance in lung metabolism. We investigated sodium-coupled transports with these organic and inorganic substrates in type II alveolar epithelial cells from adult rat after one day in culture. Alveolar type II cells actively transported inorganic phosphate and alanine, a neutral amino acid, by sodium-dependent processes. Cellular uptakes of phosphate and alanine were decreased by about 80% by external sodium substitution, inhibited by ouabain (30 and 41%, respectively) and displayed saturable kinetics. Two sodium-phosphate cotransport systems were characterized: a high-affinity one (apparent Km = 18 microM) with a Vmax of 13.5 nmol/mg protein per 10 min and a low-affinity one (apparent Km = 126 microM) with a Vmax of 22.5 nmol/mg protein per 10 min. Alanine transport had an apparent Km of 87.9 microM and a Vmax of 43.5 nmol/mg protein per 10 min. By contrast, cultured alveolar type II cells did not express sodium-dependent hexose transport. Increasing time in culture decreased Vmax values of the two phosphate transport systems on day 4 while sodium-dependent alanine uptake was unchanged. This study demonstrated the existence of sodium-dependent phosphate and amino acid transports in alveolar type II cells similar to those documented in other epithelial cell types. These sodium-coupled transports provide a potent mechanism for phosphate and amino acid absorption and are likely to play a role in substrate availability for cellular metabolism and in regulating the composition of the alveolar subphase. The decrease in phosphate uptake with time in culture is parallel to decrease in surfactant synthesis reported in cultured alveolar type II cells, suggesting that phosphate availability for surfactant synthesis may be accomplished by a sodium-dependent phosphate uptake.

β‐Liddle mutation of the epithelial sodium channel increases alveolar fluid clearance and reduces the severity of hydrostatic pulmonary oedema in mice

Transepithelial sodium transport via alveolar epithelial Na+ channels and Na+,K+‐ATPase constitutes the driving force for removal of alveolar oedema fluid. Decreased activity of the amiloride‐sensitive epithelial Na+ channel (ENaC) in the apical membrane of alveolar epithelial cells impairs sodium‐driven alveolar fluid clearance (AFC) and predisposes to pulmonary oedema. We hypothesized that hyperactivity of ENaC in the distal lung could improve AFC and facilitate the resolution of pulmonary oedema. AFC and lung fluid balance were studied at baseline and under conditions of hydrostatic pulmonary oedema in the β‐Liddle (L) mouse strain harbouring a gain‐of‐function mutation (R566stop) within the Scnn1b gene. As compared with wild‐type (+/+), baseline AFC was increased by 2‐ and 3‐fold in heterozygous (+/L) and homozygous mutated (L/L) mice, respectively, mainly due to increased amiloride‐sensitive AFC. The β2‐agonist terbutaline stimulated AFC in +/+ and +/L mice, but not in L/L mice. Acute volume overload induced by saline infusion (40% of body weight over 2 h) significantly increased extravascular (i.e. interstitial and alveolar) lung water as assessed by the bloodless wet‐to‐dry lung weight ratio in +/+ and L/L mice, as compared with baseline. However, the increase was significantly larger in +/+ than in L/L groups (P= 0.01). Volume overload also increased the volume of the alveolar epithelial lining fluid in +/+ mice, indicating the presence of alveolar oedema, but not in L/L mice. Cardiac function as evaluated by echocardiography was comparable in both groups. These data show that constitutive ENaC activation improved sodium‐driven AFC in the mouse lung, and attenuated the severity of hydrostatic pulmonary oedema.

Lidocaine induces a reversible decrease in alveolar epithelial fluid clearance in rats.

Background Lidocaine is widely used in patients with acute cardiac disorders and has also been recently implicated as a possible cause of pulmonary edema after liposuction. The objective of this study was to assess the effect of lidocaine on alveolar fluid clearance, the primary mechanism responsible for the resolution of alveolar edema. Methods Alveolar fluid clearance was measured in 29 ventilated rats using our well-validated method over 1 h using a 5% albumin solution instilled into the distal air spaces of the lung. Lidocaine was added to the instilled albumin solution (10−5 m) or administered intravenously at a dose estimated to achieve a clinically relevant plasma concentration of 10−5 m. Standard agonists and antagonists were used to determine the effect of lidocaine on alveolar fluid clearance. To determine whether lidocaine acted predominantly on the apical or basal surface, we also used QX314, lidocaine n-ethyl bromide quaternary salt, an analog of lidocaine, which is unable to cross the alveolar epithelium. The effect of lidocaine on the apical epithelial sodium channel transfected in Xenopus oocytes was also studied. Results Alveolar or intravenous lidocaine decreased alveolar fluid clearance by 50%, an effect that was reversible with the &bgr;2 agonist, terbutaline. Lidocaine acted predominantly on the basal surface of the epithelium because n-ethyl bromide quaternary salt decreased alveolar fluid clearance only when it was given intravenously and because lidocaine did not inhibit the apical epithelial sodium channel when expressed in oocytes. Conclusions Lidocaine decreased alveolar fluid clearance by 50%, an effect that may have major clinical implications in the care of patients with cardiac disease or during the perioperative period in some patients. Importantly, the effect of lidocaine was completely reversible with &bgr;2-adrenergic therapy.

Evidence for Na-K-Cl cotransport in alveolar epithelial cells: effect of phorbol ester and osmotic stress.

We have investigated the presence of Na-K-Cl cotransport in alveolar type II cells using uptake of 86Rb. Several data support the presence of a Na-K-Cl cotransport in these cells. First, a large fraction of ouabain-resistant 86Rb uptake was inhibited by bumetanide and furosemide. Second, bumetanide-sensitive 86Rb up-take required the presence of Na+ and Cl− in the incubation medium; dependency on extracellular Na+ and K+ was hyperbolic, with a Km of 14.6 μm and 8.3 μm, respectively, while dependency on extracellular Cl− was sigmoidal, which suggests a 1∶1∶2 stoichiometry. Third, a fraction of amiloride-insensitive 22Na influx was deeply inhibited by bumetanide. 22Na influx was dependent on the presence of extracellular K+ and Cl−. Since Na-K-Cl activity dramatically decreased with time in culture, further characterization of the cotransport on polarized cells could not be performed. The phorbol ester PMA inhibited Na-K-Cl cotransport in a time-and concentration-dependent manner. This inhibition was mimicked by oleoylacetylglycerol, dioctanoylglycerol, and the diacylglycerol kinase inhibitor R59022, and was reversed by an antagonist of PKC, staurosporine. Since the Na-K-Cl cotransport has been reported to be involved in cell volume regulation, we investigated its modulation by changes in extracellular osmolarity. Na-K-Cl activity was increased after a two-step procedure: swelling in hypotonic medium followed by shrinking in hypertonic medium. Under these conditions, cotransport activity increased whenever PKC activity was up-or downregulated, which suggests that the cell volume-induced modulation of the cotransport is independent from the PKC activity. Though we were not able to determine the polarity of the cotransport, it may also be involved in the absorptive function of alveolar type II cells, and would provide an alternate pathway for sodium entry.

Hypoxia-Induced Inhibition of Epithelial Na+ Channels in the Lung. Role of Nedd4-2 and the Ubiquitin-Proteasome Pathway

Transepithelial sodium transport via alveolar epithelial Na(+) channels (ENaC) and Na(+),K(+)-ATPase constitutes the driving force for removal of alveolar edema fluid. Alveolar hypoxia associated with pulmonary edema may impair ENaC activity and alveolar Na(+) absorption through a decrease of ENaC subunit expression at the apical membrane of alveolar epithelial cells (AECs). Here, we investigated the mechanism(s) involved in this process in vivo in the β-Liddle mouse strain mice carrying a truncation of β-ENaC C-terminus abolishing the interaction between β-ENaC and the ubiquitin protein-ligase Nedd4-2 that targets the channel for endocytosis and degradation and in vitro in rat AECs. Hypoxia (8% O2 for 24 h) reduced amiloride-sensitive alveolar fluid clearance by 69% in wild-type mice but had no effect in homozygous mutated β-Liddle littermates. In vitro, acute exposure of AECs to hypoxia (0.5-3% O2 for 1-6 h) rapidly decreased transepithelial Na(+) transport as assessed by equivalent short-circuit current Ieq and the amiloride-sensitive component of Na(+) current across the apical membrane, reflecting ENaC activity. Hypoxia induced a decrease of ENaC subunit expression in the apical membrane of AECs with no change in intracellular expression and induced a 2-fold increase in α-ENaC polyubiquitination. Hypoxic inhibition of amiloride-sensitive Ieq was fully prevented by preincubation with the proteasome inhibitors MG132 and lactacystin or with the antioxidant N-acetyl-cysteine. Our data strongly suggest that Nedd4-2-mediated ubiquitination of ENaC leading to endocytosis and degradation of apical Na(+) channels is a key feature of hypoxia-induced inhibition of transepithelial alveolar Na(+) transport.