Machine-learning algorithm incorporating capacitated sperm intracellular pH predicts conventional in vitro fertilization success in normospermic patients.

OBJECTIVE: To measure human sperm intracellular pH (pHi) and develop a machine-learning algorithm to predict successful conventional in vitro fertilization (IVF) in normospermic patients. DESIGN: Spermatozoa from 76 IVF patients were capacitated in vitro. Flow cytometry was used to measure sperm pHi, and computer-assisted semen analysis was used to measure hyperactivated motility. A gradient-boosted machine-learning algorithm was trained on clinical data and sperm pHi and membrane potential from 58 patients to predict successful conventional IVF, defined as a fertilization ratio (number of fertilized oocytes [2 pronuclei]/number of mature oocytes) greater than 0.66. The algorithm was validated on an independent set of data from 18 patients. SETTING: Academic medical center. PATIENT(S): Normospermic men undergoing IVF. Patients were excluded if they used frozen sperm, had known male factor infertility, or used intracytoplasmic sperm injection only. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Successful conventional IVF. RESULT(S): Sperm pHi positively correlated with hyperactivated motility and with conventional IVF ratio (n = 76) but not with intracytoplasmic sperm injection fertilization ratio (n = 38). In receiver operating curve analysis of data from the test set (n = 58), the machine-learning algorithm predicted successful conventional IVF with a mean accuracy of 0.72 (n = 18), a mean area under the curve of 0.81, a mean sensitivity of 0.65, and a mean specificity of 0.80. CONCLUSION(S): Sperm pHi correlates with conventional fertilization outcomes in normospermic patients undergoing IVF. A machine-learning algorithm can use clinical parameters and markers of capacitation to accurately predict successful fertilization in normospermic men undergoing conventional IVF.

Association of bioimpedance-derived 50-kHz phase angle as marker of body composition with electrical parameters regarding the Cole-Cole model.

Our aim is to clarify the association of the bioelectrical impedance-derived 50-kHz phase angle (φ50 ) with electrical parameters regarding the Cole-Cole model and clinical parameters. A total of 440 sets of bioelectrical impedance data from pre- and post-hemodialysis from 157 patients were used. Resistance at infinite frequency (Rinf ), resistance at 0 frequency (R0 ), capacitance of the cell membrane (Cm), and a parameter for the distribution of the time constant (α) were examined as electrical parameters. Normally hydrated lean tissue mass as a percentage of the dry weight (PNHLT ), excess fluid mass as a percentage of the dry weight (PExF ), body mass index (BMI), age, and sex (Nsex , 0 for male, 1 for female) were examined as clinical parameters. φ50 increased with the decrease in Rinf /R0 and α and also with the increase in Cm (multiple regression coefficients [ß]: pre/post, -0.886/-0.936, -0.175/-0.212, and 0.167/0.141), which determined the ratio of intra- to extracellular fluid volume (ICV/ECV), tissue homogeneity, and total cell mass. φ50 increased with an increase in PNHLT and BMI and decrease in PExF , Nsex , and age (ß: pre/post, 0.654/0.581, 0.466/0.412, -0.483/-0.473, -0.216/-0.154, and -0.145/-0.127). The concordance correlation coefficient between φ50 for pre- and post-hemodialysis (ρ = 0.772) may be improved (ρ = 0.950) by adding a multiplication of 0.2 and PExF to φ50 for correction. φ50 may be used to estimate body composition through the association with ICV/ECV, tissue homogeneity, and total cell mass. The correction for excess fluid is essential in order to use φ50 as a marker of body composition related to nutrition.

ANALYSIS OF FLUID VOLUME AND ITS IMPACT ON VISUAL ACUITY IN THE FLUID STUDY AS QUANTIFIED WITH DEEP LEARNING.

PURPOSE: To investigate quantitative differences in fluid volumes between subretinal fluid (SRF)-tolerant and SRF-intolerant treat-and-extend regimens for neovascular age-related macular degeneration and analyze the association with best-corrected visual acuity. METHODS: Macular fluid (SRF and intraretinal fluid) was quantified on optical coherence tomography volumetric scans using a trained and validated deep learning algorithm. Fluid volumes and complete resolution was automatically assessed throughout the study. The impact of fluid location and volumes on best-corrected visual acuity was computed using mixed-effects regression models. RESULTS: Baseline fluid quantifications for 348 eyes from 348 patients were balanced (all P > 0.05). No quantitative differences in SRF/intraretinal fluid between the treatment arms was found at any study-specific time point (all P > 0.05). Compared with qualitative assessment, the proportion of eyes without SRF/intraretinal fluid did not differ between the groups at any time point (all P > 0.05). Intraretinal fluid in the central 1 mm and SRF in the 1-mm to 6-mm macular area were negatively associated with best-corrected visual acuity (-2.8 letters/100 nL intraretinal fluid, P = 0.007 and -0.20 letters/100 nL SRF, P = 0.005, respectively). CONCLUSION: Automated fluid quantification using artificial intelligence allows objective and precise assessment of macular fluid volume and location. Precise determination of fluid parameters will help improve therapeutic efficacy of treatment in neovascular age-related macular degeneration.

Transcriptome Landscape of Intracellular Brucella ovis Surviving in RAW264.7 Macrophage Immune System.

Brucella ovis infection results in genital damage and epididymitis in rams, placental inflammation and rare abortion in ewes, and neonatal mortality in lambs. However, the mechanism underlying B. ovis infection remains unclear. In the present study, we used prokaryotic transcriptome sequencing to identify the differentially expressed genes (DEGs) between wild-type B. ovis and intracellular B. ovis in RAW264.7 macrophages. Gene ontology (GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed, and quantitative reverse transcriptase PCR (qRT-PCR) was used to validate the top 10 upregulated and downregulated DEGs. The results showed that 212 genes were differentially expressed, including 68 upregulated and 144 downregulated genes, which were mainly enriched in 30 GO terms linked to biological process, cellular component, and molecular function. KEGG analysis showed that the DEGs were enriched in the hypoxia-inducible factor 1 (HIF-1) signaling pathway, mitogen-activated protein kinase (MAPK) signaling pathway, beta-alanine metabolism, and quorum sensing pathway. BME_RS01160, BME_RS04270, BME_RS08185, BME_RS12880, BME_RS25875, predicted_RNA865, and predicted_RNA953 were confirmed with the transcriptome sequencing data. Hence, our findings not only reveal the intracellular parasitism of B. ovis in the macrophage immune system, but also help to understand the mechanism of chronic B. ovis infection.

Markovian approaches to modeling intracellular reaction processes with molecular memory.

Many cellular processes are governed by stochastic reaction events. These events do not necessarily occur in single steps of individual molecules, and, conversely, each birth or death of a macromolecule (e.g., protein) could involve several small reaction steps, creating a memory between individual events and thus leading to nonmarkovian reaction kinetics. Characterizing this kinetics is challenging. Here, we develop a systematic approach for a general reaction network with arbitrary intrinsic waiting-time distributions, which includes the stationary generalized chemical-master equation (sgCME), the stationary generalized Fokker-Planck equation, and the generalized linear-noise approximation. The first formulation converts a nonmarkovian issue into a markovian one by introducing effective transition rates (that explicitly decode the effect of molecular memory) for the reactions in an equivalent reaction network with the same substrates but without molecular memory. Nonmarkovian features of the reaction kinetics can be revealed by solving the sgCME. The latter 2 formulations can be used in the fast evaluation of fluctuations. These formulations can have broad applications, and, in particular, they may help us discover new biological knowledge underlying memory effects. When they are applied to generalized stochastic models of gene-expression regulation, we find that molecular memory is in effect equivalent to a feedback and can induce bimodality, fine-tune the expression noise, and induce switch.

Acid-Base Basics.

Although students initially learn of ionic buffering in basic chemistry, buffering and acid-base transport in biology often is relegated to specialized classes, discussions, or situations. That said, for physiology, nephrology, pulmonology, and anesthesiology, these basic principles often are critically important for mechanistic understanding, medical treatments, and assessing therapy effectiveness. This short introductory perspective focuses on basic chemistry and transport of buffers and acid-base equivalents, provides an outline of basic science acid-base concepts, tools used to monitor intracellular pH, model cellular responses to pH buffer changes, and the more recent development and use of genetically encoded pH-indicators. Examples of newer genetically encoded pH-indicators (pHerry and pHire) are provided, and their use for in vitro, ex vivo, and in vivo experiments are described. The continued use and development of these basic tools provide increasing opportunities for both basic and potentially clinical investigations.

Extracellular-to-Intracellular Fluid Volume Ratio as a Prognostic Factor for Survival in Patients With Metastatic Cancer.

PURPOSE: This study aimed to investigate whether the extracellular-to-intracellular fluid volume (E/I) ratio can predict survival in patients with metastatic cancer. METHODS: Clinical data were collected from April 2016 to March 2018. Patients aged ≥19 years with metastatic solid tumor were eligible. Bioimpedance analysis was used to assess body fluid distribution and the E/I ratio. Clinical characteristics, including laboratory test results and nutrition status according to the E/I ratio, were analyzed. Cox proportional hazards models and Kaplan-Meier analysis were used to identify risk factors for mortality. RESULTS: In total, 87 patients were included in the study. The 87 patients were divided into 2 groups according to the median E/I ratio: a high E/I group (E/I ratio ≥1.0, n = 43) and a low E/I group (E/I ratio <1.0, n = 44). Poor performance status, fluid retention, malnutrition, elevation of C-reactive protein levels, and decreases in hemoglobin, albumin, and protein levels were significantly associated with the high E/I group. The median overall survival time was 1.6 and 12.5 months in the high E/I and low E/I groups, respectively ( P < .001). In the multivariate analysis, poor performance status, leukocytosis, fluid retention, and E/I ratio were independent prognostic factors, and the E/I ratio was the strongest prognostic factor predicting overall survival (hazard ratio = 3.49, 95% confidence interval = 1.75-6.96, P < .001). CONCLUSIONS: The E/I ratio can predict survival time in patients with metastatic cancer. More rigorous research is required to confirm this result.

Analysis of Ca(2+) signaling mechanisms - our experience on the intercellular communication in muscle remodeling.

The aim of this study was to evaluate cell diversity by considering how Ca(2+) signaling has been adapted in skeletal muscle cell function. We characterized single C2C12 myoblasts through intracellular Ca(2+) signaling kinetics after exposure to specific drugs and calcium blockers using fast fluorescence microspectrofluorimetry followed by ATP effect analysis, which confirmed the expression of functional purinergic adenosine and P2 receptors. Further, we found that glutamate sensitivity of C2C12 cells was mediated by ionotropic glutamate receptors; on the other hand, most cells were responsive to cyclopiazonic acid, which inhibits the sarco-endoplasmic reticulum Ca(2+)-ATPase pump. These results suggest that C2C12 cells possess functional L- and P/Q-type voltage-operated Ca(2+) channels, ryanodine receptors and functional sarcoplasmic reticulumCa(2+) stores (typical for muscle cells), adenosine and P2 purinergic receptors, as well as ionotropic glutamate receptors. The evaluation of intracellular Ca(2+) signaling is a promising approach towards a better understanding and control of the physiopathological properties of myogenic cells that could be used as a predictive factor in the selection of optimal cells for scaffold recellularization or for tissue engineered constructs used in stem cell therapy.

Synaptic Potentiation at Basal and Apical Dendrites of Hippocampal Pyramidal Neurons Involves Activation of a Distinct Set of Extracellular and Intracellular Molecular Cues.

In the central nervous system, several forms of experience-dependent plasticity, learning and memory require the activity-dependent control of synaptic efficacy. Despite substantial progress in describing synaptic plasticity, mechanisms related to heterogeneity of synaptic functions at local circuits remain elusive. Here we studied the functional and molecular aspects of hippocampal circuit plasticity by analyzing excitatory synapses at basal and apical dendrites of mouse hippocampal pyramidal cells (CA1 region) in acute brain slices. In the past decade, activity of metalloproteinases (MMPs) has been implicated as a widespread and critical factor in plasticity mechanisms at various projections in the CNS. However, in the present study we discovered that in striking contrast to apical dendrites, synapses located within basal dendrites undergo MMP-independent synaptic potentiation. We demonstrate that synapse-specific molecular pathway allowing MMPs to rapidly upregulate function of NMDARs in stratum radiatum involved protease activated receptor 1 and intracellular kinases and GTPases activity. In contrast, MMP-independent scaling of synaptic strength in stratum oriens involved dopamine D1/D5 receptors and Src kinases. Results of this study reveal that 2 neighboring synaptic systems differ significantly in extracellular and intracellular cascades that control synaptic gain and provide long-searched transduction pathways relevant for MMP-dependent synaptic plasticity.

Comparison of the Body Composition of Caucasian Young Normal Body Mass Women, Measured in the Follicular Phase, Depending on the Carbohydrate Diet Level.

Background and objectives: Some publications indicate the possibility of the influence of meal nutritional value on results of bioelectrical impedance, and of the relation between the long-term carbohydrate intake and body composition. The aim of the presented study was to evaluate the influence of long-term intake of carbohydrates on body composition results assessed using the bioelectrical impedance of Caucasian young women with normal body mass, who were in the follicular phase of their menstrual cycle. Materials and Methods: Body composition was assessed in 100 women (18⁻30 years), according to strict rules, to minimize the influence of disturbing factors and by using two types of bioelectrical impedance device of the same operator to eliminate the influence of measurement (BIA 101/SC and BIA 101/ASE by Akern Srl, Firenze, Italy with the Bodygram 1.31 software and its equations by Akern Srl, Firenze, Italy). The analysis included validation of reproducibility of body composition assessment (fat, fat-free, body cell and muscle mass, water, extracellular water, and intracellular water content), and comparison of body composition for groups characterized by carbohydrate content <50% (n = 55) and >50% of the energy value of the diet (n = 45). Results: Analysis conducted using Bland⁻Altman method, analysis of correlation, analysis of quartile distribution, and weighted κ statistic revealed a positively validated reproducibility, but extracellular water associations were the weakest. Depending on the device, participants characterized by higher carbohydrate intake had significantly higher intracellular water content (p = 0.0448), or close to significantly higher (p = 0.0851) than those characterized by lower carbohydrate intake, whose extracellular water content was close to significantly lower (p = 0.0638) or did not differ. Conclusions: The long-term, moderately reduced, carbohydrate intake may cause the shift of intracellular water to the extracellular space and, as a result, influence the body composition results.

A swine model of intracellular cerebral edema - Cerebral physiology and intracranial compliance.

Cerebral edema leading to elevated intracranial pressure (ICP) is a fundamental concern after severe traumatic brain injury (TBI), stroke, and severe acute hyponatremia. We describe a swine model of water intoxication and its cerebral histological and physiological sequela. We studied female swine weighing 35-45 kg. Four serum sodium intervals were designated: baseline, mild, moderate, and severe hyponatremia attained by infusing hypotonic saline. Intracranial fluid injections were performed to assess intracranial compliance. At baseline and following water intoxication wedge biopsy was obtained for pathological examination and electron microscopy. We studied 8 swine and found an increase in ICP that was strongly related to the decrease in serum sodium level. Mean ICP rose from a baseline of 6 ±â€¯2 to 28 ±â€¯6 mm Hg during severe hyponatremia, while cerebral perfusion pressure (CPP) decreased from 72 ±â€¯10 to 46 ±â€¯11 mm Hg. Brain tissue oxygen tension (PbtO2) decreased from 18.4 ±â€¯8.9 to 5.3 ±â€¯3.0 mm Hg. Electron microscopy demonstrated intracellular edema and astrocytic foot process swelling following water intoxication. With severe hyponatremia, 2 cc intracranial fluid injection resulted in progressively greater ICP dose, indicating a worsening intracranial compliance. Our model leads to graded and sustained elevation of ICP, lower CPP, and decreased PbtO2, all of which cross clinically relevant thresholds. Intracranial compliance worsens with increased cerebral swelling. This model may serve as a platform to study which therapeutic interventions best improve the cerebral physiological profile in the face of severe brain edema.

Apoptosis: Mediator Molecules, Interplay with Other Cell Death Processes and Therapeutic Potentials.

Apoptosis, a form of programmed cell death, plays a very crucial role in various physiological processes for maintaining cell homeostasis. This process has several characteristic features like membrane blebbing, nuclear condensation, DNA fragmentation and cell shrinkage. Any defect in this highly regulated process eventually leads to extended cell survival and could result in neoplastic cell expansion followed by genetic instability. The apoptotic machinery is mainly processed and regulated by various caspases, a family of cysteine proteases. Significant advancement has been made towards understanding the molecular mechanisms of apoptosis which provides new insights in modulating the life or death of a cell. The main goal of this review is to highlight recent updates on apoptosis, the cross-talk with other cellular death processes and its therapeutic potentials.

In vitro assessment of the differences in retinal ganglion cell responses to intra- and extracellular electrical stimulation.

OBJECTIVE: To compare responses of retinal ganglion cells (RGCs) to intracellular and extracellular electrical stimulation of varying frequency and amplitude. APPROACH: In vitro patch clamp was used to record the responses of RGCs to sinusoidal current stimulation of varying frequency and amplitude. The results were simulated using the Neuron software package. MAIN RESULTS: The stimulation frequency yielding the greatest response was higher for extracellular stimulation compared to intracellular stimulation in the same cells (256 Hz versus 64 Hz). In fact, at the high end of the frequency range, where extracellular stimulation was highly efficacious, no responses could be generated using intracellular stimulation. A region in the amplitude-frequency stimulation space was identified where OFF-RGCs could be preferentially stimulated over ON-RGCs. We found that the inability of RGCs to respond at high frequencies of intracellular stimulation is likely the result of the axon acting as a low pass filter. SIGNIFICANCE: There is no direct translation of the results obtained with intracellular stimulation to those that employ extracellular stimulation.

Enhanced late sodium current underlies pro-arrhythmic intracellular sodium and calcium dysregulation in murine sodium channelopathy.

BACKGROUND: Long QT syndrome mutations in the SCN5A gene are associated with an enhanced late sodium current (INa,L) which may lead to pro-arrhythmic action potential prolongation and intracellular calcium dysregulation. We here investigated the dynamic relation between INa,L, intracellular sodium ([Na+]i) and calcium ([Ca2+]i) homeostasis and pro-arrhythmic events in the setting of a SCN5A mutation. METHODS AND RESULTS: Wild-type (WT) and Scn5a1798insD/+ (MUT) mice (age 3-5 months) carrying the murine homolog of the SCN5A-1795insD mutation on two distinct genetic backgrounds (FVB/N and 129P2) were studied. [Na+]i, [Ca2+]i and Ca2+ transient amplitude were significantly increased in 129P2-MUT myocytes as compared to WT, but not in FVB/N-MUT. Accordingly, INa,L wassignificantly more enhanced in 129P2-MUT than in FVB/N-MUT myocytes, consistent with a dose-dependent correlation. Quantitative RT-PCR analysis revealed intrinsic differences in mRNA expression levels of the sodium/potassium pump, the sodium/hydrogen exchanger, and sodium­calcium exchanger between the two mouse strains. The rate of increase in [Na+]i, [Ca2+]i and Ca2+ transient amplitude following the application of the Na+/K+-ATPase inhibitor ouabain was significantly greater in 129P2-MUT than in 129P2-WT myocytes and was normalized by the INa,L inhibitor ranolazine. Furthermore, ranolazine decreased the incidence of pro-arrhythmic calcium after-transients elicited in 129P2-MUT myocytes. CONCLUSIONS: In this study we established a causal link between the magnitude of INa,L, extent of Na+ and Ca2+ dysregulation, and incidence of pro-arrhythmic events in murine Scn5a1798insD/+ myocytes. Furthermore, our findings provide mechanistic insight into the anti-arrhythmic potential of pharmacological inhibition of INa,L in patients with LQT3 syndrome.

Effect of Andrographolide on Gene Expression Profile and Intracellular Calcium in Primary Rat Myocardium Microvascular Endothelial Cells.

Andrographolide (ANDRO) is a diterpene lactone compound with extensive biological effects, such as antibacterial, antitumor and treatment of cardiovascular diseases. Until now, studies on the pharmacological functions of ANDRO are still in progress. However, little is known about the gene expression profile and calcium response of endothelial cells to ANDRO. In this study, we used a microarray technology to investigate the gene expression responses in primary rat myocardium microvascular endothelial cells treated with 10 µg/mL ANDRO. The expression of caveolin-1 and 1-phosphatidylinositol 4, 5-bisphosphate phosphodiesterase δ3 was verified by RT-PCR and western blot. In addition, we investigated the effect of ANDRO on intracellular calcium induced by exogenous adenosine triphosphate and on inflammatory response induced by lipopolysaccharide. Results showed that ANDRO treatment induced an abundance of differential expressed genes, exhibiting a multitarget regulatory effect. ANDRO significantly decreased caveolin-1 and phosphodiesterase δ3 expression, lipopolysaccharide-induced IL-6 and TNF-α levels and expression of several chemokine genes, which are associated with reducing inflammation response and decreasing calcium release without affecting normal endothelia cell function, suggesting that ANDRO may be a potential candidate to treat cardiovascular diseases with less toxicity.

LPA1 is a key mediator of intracellular signalling and neuroprotection triggered by tetracyclic antidepressants in hippocampal neurons.

Both lysophosphatidic acid (LPA) and antidepressants have been shown to affect neuronal survival and differentiation, but whether LPA signalling participates in the action of antidepressants is still unknown. In this study, we examined the role of LPA receptors in the regulation of extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) activity and neuronal survival by the tetracyclic antidepressants, mianserin and mirtazapine in hippocampal neurons. In HT22 immortalized hippocampal cells, antidepressants and LPA induced a time- and concentration-dependent stimulation of ERK1/2 phosphorylation. This response was inhibited by either LPA1 and LPA1/3 selective antagonists or siRNA-induced LPA1 down-regulation, and enhanced by LPA1 over-expression. Conversely, the selective LPA2 antagonist H2L5186303 had no effect. Antidepressants induced cyclic AMP response element binding protein phosphorylation and this response was prevented by LPA1 blockade. ERK1/2 stimulation involved pertussis toxin-sensitive G proteins, Src tyrosine kinases and fibroblast growth factor receptor (FGF-R) activity. Tyrosine phosphorylation of FGF-R was enhanced by antidepressants through LPA1 . Serum withdrawal induced apoptotic death, as indicated by increased annexin V staining, caspase activation and cleavage of poly-ADP-ribose polymerase. Antidepressants inhibited the apoptotic cascade and this protective effect was curtailed by blockade of either LPA1 , ERK1/2 or FGF-R activity. Moreover, in primary mouse hippocampal neurons, mianserin acting through LPA1 increased phospho-ERK1/2 and protected from apoptosis induced by removal of growth supplement. These data indicate that in neurons endogenously expressed LPA1 receptors mediate intracellular signalling and neuroprotection by tetracyclic antidepressants.

Calcium and Excitation-Contraction Coupling in the Heart.

Cardiac contractility is regulated by changes in intracellular Ca concentration ([Ca2+]i). Normal function requires that [Ca2+]i be sufficiently high in systole and low in diastole. Much of the Ca needed for contraction comes from the sarcoplasmic reticulum and is released by the process of calcium-induced calcium release. The factors that regulate and fine-tune the initiation and termination of release are reviewed. The precise control of intracellular Ca cycling depends on the relationships between the various channels and pumps that are involved. We consider 2 aspects: (1) structural coupling: the transporters are organized within the dyad, linking the transverse tubule and sarcoplasmic reticulum and ensuring close proximity of Ca entry to sites of release. (2) Functional coupling: where the fluxes across all membranes must be balanced such that, in the steady state, Ca influx equals Ca efflux on every beat. The remainder of the review considers specific aspects of Ca signaling, including the role of Ca buffers, mitochondria, Ca leak, and regulation of diastolic [Ca2+]i.

Does lymphadenectomy have influence on postoperative body fluid distribution?

OBJECTIVE: We compared the fluid volume parameters in women undergoing gynaecological surgery for benign and malignant conditions before and after surgery using bioelectrical impedance vectors. STUDY DESIGN: A total of 181 patients were enrolled. In all, 89 patients had surgery for benign conditions and 92 patients underwent oncological procedures, including lymph node dissection, for malignant diseases. Bioelectrical impedance analysis (BIA) parameters were measured on the day of hospitalisation before any treatment and at 24h and 1 month after the surgical intervention. The BIA parameters measured included extracellular water (ECW), intracellular water (ICW), and total body water (TBW). RESULTS: TBW increased significantly 1 month after surgery in all cases (p<0,05 in both group). ECW was significantly higher (p<0.05) and ICW was significantly lower (p<0,05) in the malignant group than the benign group. CONCLUSION: Radical gynaecological surgeries, including lymph node dissection, have a greater effect on body water distribution than surgeries performed for benign conditions.

Roselle Polyphenols Exert Potent Negative Inotropic Effects via Modulation of Intracellular Calcium Regulatory Channels in Isolated Rat Heart.

Roselle (Hibiscus sabdariffa Linn.) calyces have demonstrated propitious cardioprotective effects in animal and clinical studies; however, little is known about its action on cardiac mechanical function. This study was undertaken to investigate direct action of roselle polyphenols (RP) on cardiac function in Langendorff-perfused rat hearts. We utilized RP extract which consists of 12 flavonoids and seven phenolic acids (as shown by HPLC profiling) and has a safe concentration range between 125 and 500 µg/ml in this study. Direct perfusion of RP in concentration-dependent manner lowered systolic function of the heart as shown by lowered LVDP and dP/dt max, suggesting a negative inotropic effect. RP also reduced heart rate (negative chronotropic action) while simultaneously increasing maximal velocity of relaxation (positive lusitropic action). Conversely, RP perfusion increased coronary pressure, an indicator for improvement in coronary blood flow. Inotropic responses elicited by pharmacological agonists for L-type Ca2+ channel [(±)-Bay K 8644], ryanodine receptor (4-chloro-m-cresol), ß-adrenergic receptor (isoproterenol) and SERCA blocker (thapsigargin) were all abolished by RP. In conclusion, RP elicits negative inotropic, negative chronotropic and positive lusitropic responses by possibly modulating calcium entry, release and reuptake in the heart. Our findings have shown the potential use of RP as a therapeutic agent to treat conditions like arrhythmia.

Acute neuroinflammation provokes intracellular acidification in mouse hippocampus.

BACKGROUND: Maintaining pH levels within the physiological norm is an important component of brain homeostasis. However, in some pathological or physiological conditions, the capacity of the pH regulatory system could be overpowered by various factors resulting in a transient or permanent alteration in pH levels. Such changes are often observed in pathological conditions associated with neuroinflammation. We hypothesized that neuroinflammation itself is a factor affecting pH levels in neural tissue. To assess this hypothesis, we examined the effects of acute LPS-induced neuroinflammation on intra- and extracellular pH (pHi and pHo) levels in the CA1 region of mouse hippocampus. METHODS: Acute neuroinflammation was induced using two approaches: (1) in vivo by i.p. injections of LPS (5 mg/kg) and (2) in vitro by incubating hippocampal slices of naïve animals in the LPS-containing media (1 µg/mL, 1 h at 35 °C). Standard techniques were used to prepare hippocampal slices. pHi was measured using ratiometric pH-sensitive fluorescent dye BCECF-AM. pHo was assessed using calibrated pH-sensitive micropipettes. The presence of neuroinflammation was verified with immunohistochemistry (IL-1ß and Iba1) and ELISA (IL-1ß and TNF-α). RESULTS: A significant reduction of pHi was observed in the slices of the LPS-injected 3-month-old (LPS 7.13 ± 0.03; Sal 7.22 ± 0.03; p = 0.043, r = 0.43) and 19-month-old (LPS 6.78 ± 0.08; Sal 7.13 ± 0.03; p = 0.0001, r = 0.32) mice. In contrast, the levels of pHo within the slice, measured in 19-month-old animals, were not affected (LPS 7.27 ± 0.02; Sal 7.26 ± 0.02; p = 0.6, r = 0.13). A reduction of pHi was also observed in the LPS-treated slices during the interval 3.5-7 h after the LPS exposure (LPS 6.92 ± 0.07; Veh 7.28 ± 0.05; p = 0.0001, r = 0.46). CONCLUSIONS: Acute LPS-induced neuroinflammation results in a significant intracellular acidification of the CA1 neurons in mouse hippocampus, while the pHo remains largely unchanged. Such changes may represent a specific protective reaction of neural tissue in unfavorable external conditions or be a part of the pathological process.