Estimation of negative membrane tension in lipid bilayers and its effect on antimicrobial peptide magainin 2-induced pore formation.

Positive membrane tension in the stretched plasma membrane of cells and in the stretched lipid bilayer of vesicles has been well analyzed quantitatively, whereas there is limited quantitative information on negative membrane tension in compressed plasma membranes and lipid bilayers. Here, we examined negative membrane tension quantitatively. First, we developed a theory to describe negative membrane tension by analyzing the free energy of lipid bilayers to obtain a theoretical equation for negative membrane tension. This allowed us to obtain an equation describing the negative membrane tension (σosm) for giant unilamellar vesicles (GUVs) in hypertonic solutions due to negative osmotic pressure (Π). Then, we experimentally estimated the negative membrane tension for GUVs in hypertonic solutions by measuring the rate constant (kr) of rupture of the GUVs induced by the constant tension (σex) due to an external force as a function of σex. We found that larger σex values were required to induce the rupture of GUVs under negative Π compared with GUVs in isotonic solution and quantitatively determined the negative membrane tension induced by Π (σosm) by the difference between these σex values. At small negative Π, the experimental values of negative σosm agree with their theoretical values within experimental error, but as negative Π increases, the deviation increases. Negative tension increased the stability of GUVs because higher tensions were required for GUV rupture, and the rate constant of antimicrobial peptide magainin 2-induced pore formation decreased.

Role of Inhibiting Inflammation of LC3-Associated Phagocytosis in Dry Eye Disease.

PURPOSE: To confirm the expression and investigate the role of LC3-associated phagocytosis (LAP) in dry eye disease (DED). METHODS: The DED model of mice was established by scopolamine subcutaneous injection in a low-humidity environment chamber. Tear secretion test and corneal fluorescein sodium staining were used to evaluate the severity of DED. Expression levels of Rubicon, microtubule-associated protein light chain 3-II (LC3-II), Beclin-1 and autophagy-related gene-7 (Atg-7) in corneas of mice with DED were tested by western blot. Cell Counting Kit-8 (CCK-8) assay was used to detect the effects of different concentrations of hypertonic solutions on the proliferation activity of human corneal epithelial cells (HCECs). The expression levels of Dectin-1, IL-6 and IL-1ß in HCECs after stimulation with different concentrations of hypertonic solutions were tested. The expressions of Rubicon, LC3-II, Beclin-1 and ATG-7 in HCECs were detected by reverse transcription polymerase chain reaction (RT-PCR). After being pretreated with 10 µM si-Rubicon, the severity of the disease was documented by corneal fluorescein sodium staining. And the expression levels of IL-6 and IL-1ß were also tested by RT-PCR. RESULTS: Compared with the normal control group, the corneal fluorescein sodium staining scores and tear secretion were significantly reduced. Rubicon, LC3-II, Beclin-1 and ATG-7 were significantly elevated. CCK-8 showed that the 400 and 450 mOsM hypertonic solutions did not affect the proliferation activity of HCECs. The expression of Dectin-1, IL-1ß and IL-6 were elevated after stimulation with 450 mOsM solution. LC3-II, Rubicon, ATG-7 and Beclin-1 increased after stimulation with 450 mOsM hyperosmolar solution in HCECs. Corneal fluorescein staining showed that si-Rubicon increased the severity of DED in mice. Moreover, the mRNA expressions of inflammatory factors IL-1ß and IL-6 in the cornea of mice were significantly increased. CONCLUSION: DED increased the expression of proteins associated with LAP. LAP could play an anti-inflammatory effect in DED.

Effects of hypertonic solutions on two species of human intestinal parasites during fecal examination.

Aim: Ova and parasite examination by flotation requires hypertonic solutions, which can damage the egg and cyst membranes, leading to false negatives. The authors investigated the harmful effects of ZnSO4 and C12H22O11 solutions on the ova and parasite examination. Materials & methods: The authors processed samples using the Three Fecal Test technique. Aliquots were floated in different pH levels, temperatures and solution densities. Results: Densities above 1.12 g/ml led structures to collapse after 6-10 min. pH neutralization of the ZnSO4 solution did not prevent the parasites from changing. Conclusion: All structures were altered when standard methods were performed. To delay collapse, the parasite floating under 5 °C is highly desirable.

Fecal exams require solutions that can damage the intestinal parasite's shape. This is bad for diagnosis. The authors investigated the harmful effects of these solutions on fecal exams. The authors processed samples using a technique called the Three Fecal Test. Fecal samples were floated in different conditions, including neutral and acidic solutions, high and low temperatures and varying densities of chemical solutions. Densities above 1.12 g/ml altered the structures of parasites. Neutral solutions did not prevent the structures from changing. The structures of all parasites were altered when the usual techniques were performed. Thus, the techniques for diagnosing intestinal parasites in feces must be improved. Temperatures under 5 °C are the best for preventing the destruction of parasite membranes.

Influence of Na+ disorder on cytoplasmic conductivity and cellular electromagnetic (EM) energy absorption of human erythrocytes (PONE-D-21-36089).

Cytoplasmic conductivity of human erythrocytes may be significantly disturbed by the composition of the external suspending media. Effects of external NaCl on cytoplasmic conductivity of human erythrocyte (Human Red Blood Cells, HRBC) were investigated in a simple NaCl system. Using thermodynamic theory cytoplasmic conductivities could be calculated from internal [K+], [Na+], [Cl-] and [HCO3-]. Effect of cell volume and cell water changes were introduced and allowed for using the Debye-Hückel-Onsager relation and Walden's rule of viscosity. Cell volume and cell water change of HRBCs were measured in suspending isotonic solutions with conductivities from 0.50 S m-1 up to hypertonic solutions of conductivity of 2.02 S m-1 at selected temperatures of 25°C (standard benchmark temperature) and 37°C (physiological temperature). In isotonic solutions, cytoplasmic conductivity of human erythrocyte decreases with rise in the external media ionic concentration and vice versa for hypertonic solutions. The HRBC is capable of rapidly regulating its volume (and shape) over quite a wide range of osmolality. Specific Absorption Rate (SAR, 900 MHz) values (W kg-1) of electromagnetic radiation are below safe limits at non-physiological 25°C but above legal limits at 37°C [National Council on Radiation Protection and Measurements, NCRP]. However, at 37°C under both hypertonic [Na+] and isotonic but low [Na+], SAR increases further beyond legal limits.

Stability of benzylpenicillin for continuous intravenous infusions: An isotonic formulation for therapeutic use and a low-dose formulation for clinical trial.

INTRODUCTION: The objectives of this study were to develop a stability-indicating high performance liquid chromatography (HPLC) assay for benzylpenicillin (BPC) in pharmaceutical fluids, and to investigate the stability of (i) isotonic citrate-buffered BPC solutions at the clinically relevant concentration of 30 mg/mL, and (ii) low concentration citrate-buffered BPC intravenous infusions (5-30 µg/mL). METHODS: The stability of isotonic BPC solutions containing 3.4 or 7.2 mg/mL sodium citrate was compared against contemporary hypertonic solutions. The HPLC assay was shown to be stability-indicating following acidic, alkali, oxidative and elevated temperature stress testing. RESULTS: After 7 d storage at 4 °C and 24 h at 35 °C, the concentrations of isotonic BPC 30 mg/mL solutions containing 3.4 and 7.2 mg/mL sodium citrate were 96% and 95% respectively, compared to day 0. After 3 d at 4 °C and 24 h at room temperature (22 °C), the concentrations of isotonic BPC solutions with 3.4 and 7.2 mg/mL sodium citrate were 99% and 96% respectively, compared to day 0. These data were comparable to the hypertonic solutions and meet pharmacopeial stability requirements. Low concentration BPC infusions showed 0.5% and 2.5% degradation after 24 h storage at 22 °C and 35 °C, respectively. CONCLUSIONS: The isotonic BPC 30 mg/mL formulation is simple to prepare and may offer clinical benefits in settings where hypertonic solutions are problematic. This study provides assurance that high- and low-dose isotonic BPC infusions are stable at room temperature and our findings may be applicable to in vitro studies of BPC.

Determinants of hypokalemia following hypertonic sodium bicarbonate infusion.

The hypokalemic response to alkali infusion has been attributed to the resulting extracellular fluid (ECF) expansion, urinary potassium excretion, and internal potassium shifts, but the dominant mechanism remains uncertain. Hypertonic NaHCO3 infusion (1 N, 5 mmol/kg) to unanesthetized dogs with normal acid-base status or one of the four chronic acid-base disorders decreased plasma potassium concentration ([K+]p) at 30 min in all study groups (Δ[K+]p, - 0.16 to - 0.73 mmol/L), which remained essentially unaltered up to 90-min postinfusion. ECF expansion accounted for only a small fraction of the decrease in ECF potassium content, (K+)e. Urinary potassium losses were large in normals and chronic respiratory acid-base disorders, limited in chronic metabolic alkalosis, and minimal in chronic metabolic acidosis, yet, ongoing kaliuresis did not impact the stability of [K+]p. All five groups experienced a reduction in (K+)e at 30-min postinfusion, Δ(K+)e remaining unchanged thereafter. Intracellular fluid (ICF) potassium content, (K+)i, decreased progressively postinfusion in all groups excluding chronic metabolic acidosis, in which a reduction in (K+)e was accompanied by an increase in (K+)i. We demonstrate that hypokalemia following hypertonic NaHCO3 infusion in intact animals with acidemia, alkalemia, or normal acid-base status and intact or depleted potassium stores is critically dependent on mechanisms of internal potassium balance and not ECF volume expansion or kaliuresis. We envision that the acute NaHCO3 infusion elicits immediate ionic shifts between ECF and ICF leading to hypokalemia. Thereafter, maintenance of a relatively stable, although depressed, [K+]e requires that cells release potassium to counterbalance ongoing urinary potassium losses.

Hypertonic lactate for the treatment of intracranial hypertension in patients with acute brain injury.

Hypertonic lactate (HL) is emerging as alternative treatment of intracranial hypertension following acute brain injury (ABI), but comparative studies are limited. Here, we examined the effectiveness of HL on main cerebral and systemic physiologic variables, and further compared it to that of standard hypertonic saline (HS). Retrospective cohort analysis of ABI subjects who received sequential osmotherapy with 7.5% HS followed by HL-given at equi-osmolar (2400 mOsmol/L) and isovolumic (1.5 mL/kg) bolus doses-to reduce sustained elevations of ICP (> 20 mmHg). The effect of HL on brain (intracranial pressure [ICP], brain tissue PO2 [PbtO2], cerebral microdialysis [CMD] glucose and lactate/pyruvate ratio [LPR]) and blood (chloride, pH) variables was examined at different time-points (30, 60, 90, 120 min vs. baseline), and compared to that of HS. A total of 34 treatments among 17 consecutive subjects (13 traumatic brain injury [TBI], 4 non-TBI) were studied. Both agents significantly reduced ICP (p < 0.001, at all time-points tested): when comparing treatment effectiveness, absolute ICP decrease in mmHg and the duration of treatment effect (median time with ICP < 20 mmHg following osmotherapy 183 [108-257] vs. 150 [111-419] min) did not differ significantly between HL and HS (all p > 0.2). None of the treatment had statistically significant effects on PbtO2 and CMD biomarkers. Treatment with HL did not cause hyperchloremia and resulted in a more favourable systemic chloride balance than HS (Δ blood chloride - 1 ± 2.5 vs. + 4 ± 3 mmol/L; p < 0.001). This is the first clinical study showing that HL has comparative effectiveness than HS for the treatment of intracranial hypertension, while at the same time avoiding hyperchloremic acidosis. Both agents had no significant effect on cerebral oxygenation and metabolism.

Evidencia científica y recomendaciones sobre el manejo de la bronquiolitis en pediatria / Cientific evidence and recommendations on the bronchiolitis management in pediatry

En los ámbitos científico e institucional existe controversia sobre el manejo idóneo de la bron- quiolitis en pacientes pediátricos. El objetivo de este trabajo es valorar el nivel de evidencia cientí- fica que existe sobre el manejo de la bronquiolitis para determinar si las recomendaciones actuales son o no adecuadas. Se realizó una revisión sis- temática de artículos científicos consultando di- versas bases de datos, sin restricción de fecha, en los idiomas español e inglés. Se incluyó literatura gris mediante búsqueda manual. No se hicieron restricciones respecto al tipo de estudio. Se re- visaron los resúmenes y en los casos necesarios los artículos completos, teniéndose en cuenta fi- nalmente todos los artículos que incluían apor- tes sobre el manejo adecuado de la bronquiolitis. Como resultado la mayoría de las recomenda- ciones realizadas por las sociedades fueron a tra- vés de guías de práctica clínica o artículos de opinión, concluyendo que no se cuenta con un esquema de tratamiento adecuado para tratar la bronquiolitis aguda, existiendo un manejo erróneo con el uso de esteroides y antibióticos, mientras que uno de los tratamiento más viables y costo efectivos queda en el olvido como es la solución hipertónica al 3%, la cual ha demostra- do reducción de la estancia hospitalaria...(AU)

Hypoxic and osmotic expression of Kir2.1 potassium channels in retinal pigment epithelial cells: Contribution to vascular endothelial growth factor expression.

Retinal pigment epithelial (RPE) cells express different subtypes of inwardly rectifying potassium (Kir) channels. We investigated whether human and rat RPE cells express genes of strongly rectifying Kir2 channels. We also determined the hypoxic and hyperosmotic regulation of Kir2.1 gene expression in cultured human RPE cells and the effects of siRNA-mediated knockdown of Kir2.1 on VEGFA expression, VEGF secretion, proliferation, and viability of the cells. Extracellular hyperosmolarity was induced by addition of NaCl or sucrose. Hypoxia and chemical hypoxia were produced by cell culture in 0.25% O2 and addition of CoCl2, respectively. Gene expression levels were evaluated by real-time RT-PCR. Rat RPE cells contained Kir2.1, Kir2.2, Kir2.3, and Kir2.4 gene transcripts while human RPE cells contained Kir2.1, Kir2.2, and Kir2.4 transcripts. Immunocytochemical data may suggest that Kir2.1 protein in cultured human cells is expressed in both perinuclear and plasma membranes. Kir2.1 gene expression and Kir2.1 protein level in human cells increased under hypoxic and hyperosmotic conditions. The expression of the Kir2.1 gene was mediated in part by diverse intracellular signal transduction pathways and transcription factor activities under both conditions; the hyperosmotic, but not the CoCl2-induced Kir2.1 gene expression was dependent on intracellular calcium signaling. Autocrine/paracrine activation of purinergic receptors contributed to Kir2.1 gene expression under hyperosmotic (P2Y1, P2Y2, P2X7) and CoCl2-induced conditions (P2Y2, P2X7). Exogenous VEGF, TGF-ß1, and blood serum decreased Kir2.1 gene expression. Inhibition of VEGF receptor-2 increased the Kir2.1 gene expression under control conditions and in CoCl2-simulated hypoxia, and decreased it under high NaCl conditions. Knockdown of Kir2.1 by siRNA inhibited the CoCl2-induced and hyperosmotic transcription of the VEGFA gene and caused a delayed decrease of the constitutive VEGFA gene expression while VEGF protein secretion was not altered. Kir2.1 knockdown stimulated RPE cell proliferation under control and hyperosmotic conditions without affecting cell viability. The data indicate that Kir2.1 channel activity is required for the expression of the VEGFA gene and inhibits the proliferation of RPE cells. Under control and hypoxic conditions, the extracellular VEGF level may regulate the production of VEGF via its inhibitory effect on the Kir2.1 gene transcription; this feedback loop may prevent overproduction of VEGF.

Brain Protection after Anoxic Brain Injury: Is Lactate Supplementation Helpful?

While sudden loss of perfusion is responsible for ischemia, failure to supply the required amount of oxygen to the tissues is defined as hypoxia. Among several pathological conditions that can impair brain perfusion and oxygenation, cardiocirculatory arrest is characterized by a complete loss of perfusion to the brain, determining a whole brain ischemic-anoxic injury. Differently from other threatening situations of reduced cerebral perfusion, i.e., caused by increased intracranial pressure or circulatory shock, resuscitated patients after a cardiac arrest experience a sudden restoration of cerebral blood flow and are exposed to a massive reperfusion injury, which could significantly alter cellular metabolism. Current evidence suggests that cell populations in the central nervous system might use alternative metabolic pathways to glucose and that neurons may rely on a lactate-centered metabolism. Indeed, lactate does not require adenosine triphosphate (ATP) to be oxidated and it could therefore serve as an alternative substrate in condition of depleted energy reserves, i.e., reperfusion injury, even in presence of adequate tissue oxygen delivery. Lactate enriched solutions were studied in recent years in healthy subjects, acute heart failure, and severe traumatic brain injured patients, showing possible benefits that extend beyond the role as alternative energetic substrates. In this manuscript, we addressed some key aspects of the cellular metabolic derangements occurring after cerebral ischemia-reperfusion injury and examined the possible rationale for the administration of lactate enriched solutions in resuscitated patients after cardiac arrest.

Exposure to hypertonic solutions during pregnancy induces autism-like behaviors via the NFAT-5 pathway in offspring in a rat model.

OBJECTIVES: to investigate the effects of hyperosmolar state (HS) on immune response and inflammation via the NFAT5 pathway and examine whether immune-mediated conditions trigger autism-like behavior in offspring. METHODS: a pregnant rat model was performed by administering hyperosmotic solutions. Pregnant rats were divided into 2 main groups; control (group I) and hyperosmolar groups (group II). Control group rats were given % 0.25 NaCI (tap water) (n = 6), the Hyperosmolar (HO) group was further subdivided into 3 groups as; Group II a rats which were given % 3 hypertonic NaCl (n = 6), Group II b rats were given mineral water (% 3 NaHCO3+magnesium+calcium content) (n = 6), and Group II c rats were given Ayran (% 0.8 NaCl content) (n = 6). Their offspring were examined for behaviors, biochemical and histological abnormality. RESULTS: in offspring, TNF- α, IL-17, NFAT-5, and NGF levels in the brain were significantly higher in hyperosmotic solution groups than in control rats. Exposure of pregnant rats to hyperosmotic solution resulted in autism-like behaviors in their offspring. Through immunohistochemical methods, we found that CA1 and CA2 of the hippocampus indicated decreased number of neurons in hyperosmotic solution groups compared with the control group. CONCLUSIONS: our findings once again emphasized that the immune-mediated conditions involved in the pathophysiology of autism. NFAT5 pathway may be a key factor in the development of neuroinflammation by hyperosmotic solutions.

Hypertonic versus isotonic crystalloid infusion for cerebral perfusion pressure in a porcine experimental cardiac arrest model.

BACKGROUND: The effect of intravenous (IV) fluid administration type on cerebral perfusion pressure (CePP) during cardiopulmonary resuscitation (CPR) is controversial. The purpose of this study was to evaluate the association between IV fluid type and CePP in a porcine cardiac arrest model. METHODS: We randomly assigned 12 pigs to the hypertonic crystalloid, isotonic crystalloid and no-fluid groups. After 4 min of untreated ventricular fibrillation (VF), chest compression was conducted for 2 cycles (CC only). Chest compression with IV fluid infusion (CC + IV) was followed for 2 cycles. Advanced life support, including defibrillation and epinephrine, was added for 8 cycles (ALS phase). Mean arterial pressure (MAP), intracranial pressure (ICP) and CePP were measured. A paired t-test was used to measure the mean difference in CePP. RESULTS: Twelve pigs underwent the experiment. The hypertonic crystalloid group showed higher CePP values than those demonstrated by the isotonic crystalloid group from ALS cycles 2 to 8. The MAP values in the hypertonic group were higher than those in the isotonic group starting at ALS cycle 2. The ICP values in the hypertonic group were lower than those in the isotonic group starting at ALS cycle 4. From ALS cycles 2 to 8, the reduction in the mean difference in the isotonic group was larger than that in the other groups. CONCLUSION: In a VF cardiac arrest porcine study, the hypertonic crystalloid group showed higher CePP values by maintaining higher MAP values and lower ICP values than those of the isotonic crystalloid group.

Combined hyperosmolarity and inflammatory conditions in stressed human corneal epithelial cells and macrophages to evaluate osmoprotective agents as potential DED treatments.

PURPOSE: To develop an easy-to-perform combined model in human corneal epithelial cells (HCECs) and Balb/c mice macrophages J774.A1 (MP) for preliminary screening of potential ophthalmic therapeutic substances. METHODS: HCECs were exposed to different osmolarities (350-500 mOsm/L) and MTT assay was employed for cell survival and flow cytometry to assess apoptosis-necrosis and relative cell size (RCS) distribution. Effectiveness of Betaine, L-Carnitine, Taurine at different concentrations (ranging from 20 mM to 200 mM) was studied. Also, mucoadhesive polymers such as Hyaluronic acid (HA) and Hydroxypropylmethylcellulose (HPMC) (0.4 and 0.8%) were evaluated. Cells were pre-incubated with the compounds (8h) and then exposed to hyperosmotic stress (470 mOsm/L) for 16h. Moreover, anti-inflammatory activity was performed in LPS-stimulated MP. RESULTS: Exposure to hyperosmotic solutions between 450 and 500 mOsm/L promoted the highest cell death after 16h exposures (p < 0.0001) with a drop in viability to 34.96% ± 11.77 for 470 mOsm/L. Pre-incubation with Betaine at 150 mM and 200 mM provided the highest cell survival against hyperosmolarity (66.01% ± 3.65 and 65.90% ± 0.78 respectively) while HA 0.4% was the most effective polymer in preventing cell death (42.2% ± 3.60). Flow cytometry showed that Betaine and Taurine at concentrations between 150-200 mM and 20-80 mM respectively presented the highest anti-apoptotic activity. Also, HA and HPMC polymers reduced apoptotic-induced cell death. All osmoprotectants modified RCS, and polymers increased their value over 100%. L-Carnitine 50 mM, Taurine 40 mM and HA 0.4% presented the highest TNF-α inhibition activity (60%) albeit all of them showed anti-inflammatory inhibition percentages higher than 20% CONCLUSIONS: HCECs hyperosmolar model combined with inflammatory conditions in macrophages allows the screening of osmoprotectants by simulating chronic hyperosmolarity (16h) and inflammation (24h).

The Role of Nuclear Factor of Activated T Cells 5 in Hyperosmotic Stress-Exposed Human Lens Epithelial Cells.

We investigated the role of nuclear factor of activated T cells 5 (NFAT5) under hyperosmotic conditions in human lens epithelial cells (HLECs). Hyperosmotic stress decreased the viability of human lens epithelial B-3 cells and significantly increased NFAT5 expression. Hyperosmotic stress-induced cell death occurred to a greater extent in NFAT5-knockout (KO) cells than in NFAT5 wild-type (NFAT5 WT) cells. Bcl-2 and Bcl-xl expression was down-regulated in NFAT5 WT cells and NFAT5 KO cells under hyperosmotic stress. Pre-treatment with a necroptosis inhibitor (necrostatin-1) significantly blocked hyperosmotic stress-induced death of NFAT5 KO cells, but not of NFAT5 WT cells. The phosphorylation levels of receptor-interacting protein kinase 1 (RIP1) and RIP3, which indicate the occurrence of necroptosis, were up-regulated in NFAT5 KO cells, suggesting that death of these cells is predominantly related to the necroptosis pathway. This finding is the first to report that necroptosis occurs when lens epithelial cells are exposed to hyperosmolar conditions, and that NFAT5 is involved in this process.

A rapid and efficient method for the extraction and identification of menaquinones from Actinomycetes in wet biomass.

BACKGROUND: Menaquinones are constituents of prokaryote cell membranes where they play important functions during electron transport. Menaquinone profiles are strongly recommended for species classification when proposing a new Actinomycetes taxon. Presently, the most widely used methods to determine menaquinones are based on freeze-dried cells. Taxonomic research in our lab has revealed that menaquinone concentrations are low for some species of the genus Microbacterium, leading to difficulties in identifying menaquinones. RESULTS: Menaquinones extracted using the novel lysozyme-chloroform-methanol (LCM) method were comparable in quality to those obtained using the Collins method, the most widely used method. All tested strains extracted via the LCM method showed higher concentrations of menaquinones than those extracted via the Collins method. For some Microbacterium strains, the LCM method exhibited higher sensitivity than the Collins method, and more trace menaquinones were detected with the LCM method than the Collins method. In addition, LCM method is faster than the Collins method because it uses wet cells. CONCLUSION: The LCM method is a simple, rapid and efficient technique for the extraction and identification of menaquinones from Actinomycetes.

Uso de Solución Hipertónica en pacientes con insuficiencia cardiaca aguda como terapia adyuvante a altas dosis de diuréticos / Use of Hypertonic Solution in patients with acute heart failure as adjunctive therapy to high-dose diuretics

Resumen La insuficiencia cardíaca aguda descompensada (ICAD) es una causa común de hospitalización, con repercusiones significativas en los sistemas de salud. El manejo agudo se basa en la reducción de la volemia con diuréticos de asa, sin embargo, un porcentaje de pacientes presenta resistencia o no logra la respuesta clínica esperada con este tratamiento. Una de las medidas que ha comprobado ser efectiva en este contexto, es el uso de solución salina hipertónica (SSH) en conjunto con dosis altas de diuréticos de asa, como medida terapéutica temida por sus posibles repercusiones sobre la función renal y posible sobrecarga de sodio. Objetivos: Determinar si el uso de solución salina hipertónica en pacientes con falla cardiaca aguda e hipervolemia genera un deterioro de la función renal. Determinar la respuesta del Pro-BNP ante el uso de la solución salina hipertónica en pacientes con falla cardiaca aguda como marcador de respuesta terapéutica. Determinar si el uso de solución salina hipertónica aumenta la diuresis sin generar cambios importantes en el sodio. Se muestran datos de pacientes con insuficiencia cardiaca aguda descompensada, que tras no presentar mejoría con altas dosis de diurético de asa en bolo, se les aplicó la solución hipertónica como adyuvante a este tratamiento. Se toma un total de 26 pacientes analizando datos generales clínicos y de laboratorio, se valoran curvas con la respuesta diurética y por parámetros de laboratorio a las 48 y 72 horas. El uso de solución salina hipertónica consigue un aumento de más de un 200% de la diuresis en 24 horas, con un descenso del Pro BNP de más de un 60% a las 48 horas, sin mostrar un cambio importante en los niveles de creatinina, nitrógeno ureico y sodio. Se requirió reposición de potasio en la totalidad de los pacientes. Se concluye que la infusión de furosemida más solución hipertónica es efectiva tanto en disminuir niveles de NT Pro-BNP en los pacientes, como en generar un aumento en el volumen de diuresis. La principal complicación fue la hipokalemia, sin cambios considerables en el valor de sodio, creatinina y nitrógeno ureico séricos.

Abstract Uso de Solución Hipertónica en pacientes con insuficiencia cardiaca aguda como terapia adyuvante a altas dosis de diuréticos Acute decompensated heart failure (AHF) is a common cause of hospitalization, with significant repercussions on health systems. Acute management is based on the reduction of blood volume with loop diuretics; however, a percentage of patients show resistance or do not achieve the expected clinical response with this treatment. One of the measures that has proven to be effective in this context is the use of hypertonic saline (HSS) in conjunction with high doses of loop diuretics, as a therapeutic measure feared due to its possible repercussions on kidney function and possible sodium overload. Objetives: To determine if the use of hypertonic saline in patients with acute heart failure and hypervolemia leads to a deterioration in renal function. To determine the response of Pro-BNP to the use of hypertonic saline in patients with acute heart failure as a marker of therapeutic response. Determine if the use of hypertonic saline increases urine output without causing significant changes in sodium. Data are shown from patients with acute decompensated heart failure, who after not presenting improvement with high doses of bolus loop diuretic, the hypertonic solution was applied as an adjunct to this treatment. A total of 26 patients are taken analyzing general clinical and laboratory data, curves with the diuretic response and by laboratory parameters are evaluated at 48 and 72 hours. The use of hypertonic saline solution achieves an increase of more than 200% in diuresis in 24 hours, with a decrease in Pro BNP of more than 60% at 48 hours, without showing a significant change in creatinine levels, urea nitrogen and sodium. Potassium replacement was required in all patients. It is concluded that the infusion of furosemide plus hypertonic solution is effective both in reducing levels of NT Pro-BNP in patients, and in generating an increase in the volume of diuresis. The main complication was hypokalemia, without significant changes in serum sodium, creatinine, and urea nitrogen.

PLCδ1 Plays Central Roles in the Osmotic Activation of ΔN-TRPV1 Channels in Mouse Supraoptic Neurons and in Murine Osmoregulation.

The magnocellular neurosecretory cells (MNCs) of the hypothalamus play a vital role in osmoregulation, but the mechanisms underlying MNC osmosensitivity are not fully understood. We showed previously that high osmolality activates phospholipase C (PLC) in rat MNCs in a Ca2+-dependent manner and that PLC activation is necessary for full osmotic activation of an N-terminal variant of the TRPV1 (ΔN-TRPV1) channel. We therefore hypothesized that the Ca2+-dependent δ1 isoform of PLC contributes to ΔN-TRPV1 activation and tested whether MNC function is defective in a transgenic PLCδ1 KO mouse. Water deprivation for 24 h caused greater increases in serum osmolality and losses in body weight in PLCδ1 KO mice than it did in control mice. Action potentials and ΔN-TRPV1 currents were measured in acutely isolated mouse MNCs using whole-cell patch clamp before and after exposure to hypertonic solutions. This treatment elicited a significant activation of ΔN-TRPV1 currents and an increase in firing rate in MNCs isolated from control mice, but not from PLCδ1 KO mice. Submembranous filamentous actin was measured in isolated MNCs before and after treatment with angiotensin II and hypertonic solution. Both treatments caused an increase in filamentous actin fluorescence in MNCs isolated from control mice, but both responses were significantly attenuated in MNCs from PLCδ1 KO mice. Our data demonstrate that the PLCδ1 isoform plays a key role in the activation of ΔN-TRPV1 channels and in osmosensory transduction in MNCs. This study advances our understanding of the molecular mechanisms underlying mammalian osmoregulation.SIGNIFICANCE STATEMENT Magnocellular neurosecretory cells (MNCs) of the hypothalamus play a central role in osmoregulation. We have identified a key role for the PLCδ1 isoform in the activation of ΔN-TRPV1 channels and osmosensory transduction in MNCs. The data indicate that the PLCδ1 isoform is activated by the Ca2+ influx occurring during MNC action potentials and exerts a positive feedback on ΔN-TRPV1 channels to enhance MNC excitability. This study provides evidence that PLCδ1 is a key molecule underlying osmosensory transduction, the regulation of VP release, and osmoregulation.

Hypertonic Sodium Lactate to Alleviate Functional Deficits Following Diffuse Traumatic Brain Injury: An Osmotic or a Lactate-Related Effect?

BACKGROUND: There has been growing interest in the use of hypertonic sodium lactate (HSL) solution following traumatic brain injury (TBI) in humans. However, little is known about the effects of HSL on functional deficits with respect to the hyperosmotic nature of HSL. METHODS: We have compared the effects of HSL solution and isotonic saline solution using sensorimotor and cognitive tests for 14 days post-trauma in animals. Thirty minutes after trauma (impact-acceleration model), anesthetized rats were randomly allocated to receive a 2-h infusion of isotonic saline solution (TBI-saline group) or HSL (TBI-HSL group) (n = 10 rats per group). In another series of experiments using a similar protocol, the effects of equiosmolar doses of HSL and hypertonic saline solution (HSS) were compared in TBI rats (n = 10 rats per group). Blood lactate and ion concentrations were measured during the 2-h infusions. RESULTS: Compared to the TBI-saline group, the TBI-HSL group had a reduced latency to complete the adhesive removal test: 6 s (5-9) (median [25-75th centiles]) versus 13 s (8-17) on day 7, and 5 s (5-9) versus 11 s (8-26) on day 14 (P < 0.05), respectively, and a shorter delay to complete the radial arm maze test on day 7: 99 s (73-134) versus 176 s (127-300), respectively (P < 0.05). However, no differences were found between the TBI-HSL and TBI-HSS groups in neurocognitive tests performance. Compared to the TBI-saline group, the HSL and HSS groups had higher serum osmolality: 318 mOsm/Kg (315-321) and 315 mOsm/Kg (313-316) versus 307 mOsm/Kg (305-309), respectively (P < 0.05), and the HSL group had a higher serum lactate concentration: 6.4 mmol/L (5.3-7.2) versus 1.5 mmol/L (1.1-1.9) and 1.6 mmol/L (1.5-1.7), respectively (P < 0.05). CONCLUSIONS: These results indicate that improvements in cognitive and sensorimotor tests with HSL infusion post-TBI could be related to elevation of serum osmolality, not to exogenous administration of lactate.

A novel tonicity-responsive microRNA miR-23a-5p modulates renal cell survival under osmotic stress through targeting heat shock protein 70 HSPA1B.

There is growing evidence that microRNAs (miRNAs) are implicated in cellular adaptation to osmotic stress, but the underlying osmosignaling pathways are still not completely understood. In this study, we found that a passenger strand miRNA, miR-23a-5p, was significantly downregulated in response to high NaCl treatment in mouse inner medullary collecting duct cells (mIMCD3) through an miRNA profiling assay. The decrease of miR-23a-5p is hypertonicity-dependent and osmotolerant cell type-specific. Knockdown of miR-23a-5p increased cellular survival and proliferation in mIMCD3. In contrast, miR-23a-5p overexpression repressed cell viability and proliferation under hypertonic stress. RNA deep-sequencing revealed that a heat shock protein 70 (HSP70) isoform, HSP70 member 1B (HSPA1B), was significantly increased under hypertonic treatment. Based on the prediction analysis by Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and TargetScan, and a further validation via a dual-luciferase assay, HSPA1B was identified as a potential target of miR-23a-5p. Overexpressed miR-23a-5p suppressed HSPA1B, whereas downregulated miR-23a-5p promoted HSPA1B expression in mIMCD3. In addition, an in vivo study demonstrated that there is a reverse correlation between the levels of miR-23a-5p and HSPA1B in mouse renal inner medulla (papilla) that is exposed to extremely high osmolality. In summary, this study elucidates that passenger strand miR-23a-5p is a novel tonicity-responsive miRNA. The downregulation of miR-23a-5p facilitates cellular adaptation to hypertonic stress in mammalian renal cells through modulating HSPA1B.

Electrochemical Stability Investigations and Drug Toxicity Tests of Electrolyte-Gated Organic Field-Effect Transistors.

Electrolyte-gated organic field-effect transistors (EGOFETs) are emerging as a new frontier of organic bioelectronics, with promising applications in biosensing, pharmaceutical testing, and neuroscience. However, the limited charge carriers' mobility and well-known environmental instability of conjugated polymers constrain the real applications of organic bioelectronics. Here, we comparatively studied the electrochemical stability of p-type conjugated polymer films in the EGOFET configuration. By combining electrochemical stability tests, morphology characterization, and EQCM-D monitoring, we find that a donor-acceptor copolymer, poly(N-alkyldiketopyrrolo-pyrrole-dithienylthieno[3,2-b]thiophene) (DPP-DTT) shows improved mobility and electrochemical stability under an electrolyte, which may benefit from the ordered morphology and close alkyl side-chains' interdigitation preventing water diffusion and ion doping during long-term operation under an electrolyte. Based on the DPP-DTT EGOFETs, we have demonstrated a low-cost drug toxicity test platform that is sensitive enough to distinguish the cytotoxicity of different chemicals. This study overall pushes forward the development of organic bioelectronics with enhanced stability and sensitivity and presents successful exploitation of EGOFET in pharmaceutical research.