ATPase mRNA Research Articles

Noradrenaline enhances Na-K ATPase subunit expression by HuR-induced mRNA stabilization and their transportation to the cell surface through PLC and PKC mediated pathway: Implications with REMS-loss associated disorders.

Rapid eye movement sleep (REMS) maintains brain excitability at least by regulating Na-K ATPase activity. Although REMS deprivation (REMSD)-associated elevated noradrenaline (NA) increases Na-K ATPase protein expression, its mRNA transcription did not increase. We hypothesized and confirmed both invivo as well as invitro that elevated mRNA stability explains the apparent puzzle. The mRNA stability was measured in control and REMSD rat brain with or without invivo treatment with α1-adrenoceptor (AR) antagonist, prazosin (PRZ). Upon REMSD, Na-K ATPase α1-, and α2-mRNA stability increased significantly, which was prevented by PRZ. To decipher the molecular mechanism of action, we estimated NA-induced Na-K ATPase mRNA stability in Neuro-2a cells under controlled conditions and by transcription blockage using Actinomycin D (Act-D). NA increased Na-K ATPase mRNA stability, which was prevented by PRZ and propranolol (PRP, β-AR antagonist). The knockdown assay confirmed that the increased mRNA stabilization was induced by elevated cytoplasmic abundance of Human antigen R (HuR) and involving (Phospholipase C) PLC-mediated activation of Protein Kinase C (PKC). Additionally, using cell-impermeable Enz-link sulfo NHS-SS-Biotin, we observed that NA increased Na-K ATPase α1-subunits on the Neuro-2a cell surface. We conclude that REMSD-associated elevated NA, acting on α1- and β-AR, increases nucleocytoplasmic translocation of HuR and increases Na-K ATPase mRNA stability, resulting in increased Na-K ATPase protein expression. The latter then gets translocated to the neuronal membrane surface involving both PKC and (Protein Kinase A) PKA-mediated pathways. These findings may be exploited for the amelioration of REMSD-associated chronic disorders and symptoms.

Activation of farnesoid X receptor retards expansion of renal collecting duct cell-derived cysts via inhibition of CFTR-mediated Cl- secretion.

The transcription factor farnesoid X receptor (FXR) regulates energy metabolism. Specifically, FXR functions to regulate cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl- secretion in intestinal epithelial cells. Therefore, this study aimed to investigate the role of FXR in CFTR-mediated Cl- secretion in renal tubular cells and to further elucidate its effects on renal cyst formation and growth. CFTR-mediated Cl- transport was evaluated via short-circuit current (ISC) measurements in Madin-Darby canine kidney (MDCK) cell monolayers and primary rat inner medullary collecting duct cells. The role of FXR in renal cyst formation and growth was determined by the MDCK cell-derived cyst model. Incubation with synthesized (GW4064) and endogenous (CDCA) FXR ligands reduced CFTR-mediated Cl- secretion in a concentration- and time-dependent manner. The inhibitory effect of FXR ligands was not due to the result of reduced cell viability and was attenuated by cotreatment with an FXR antagonist. FXR activation significantly decreased CFTR protein but not its mRNA. In addition, FXR activation inhibited CFTR-mediated Cl- secretion in primary renal collecting duct cells. FXR activation decreased ouabain-sensitive ISC without altering Na+-K+-ATPase mRNA and protein levels. Furthermore, FXR activation significantly reduced the number of cysts and renal cyst expansion. These inhibitory effects were correlated with a decrease in the expression of protein synthesis regulators mammalian target of rapamycin/S6 kinase. This study shows that FXR activation inhibits Cl- secretion in renal cells via inhibition of CFTR expression and retards renal cyst formation and growth. The discoveries point to a physiological role of FXR in the regulation of CFTR and a potential therapeutic application in polycystic kidney disease treatment.NEW & NOTEWORTHY The present study reveals that farnesoid X receptor (FXR) activation reduces microcyst formation and enlargement. This inhibitory effect of FXR activation is involved with decreased cell proliferation and cystic fibrosis transmembrane conductance regulator-mediated Cl- secretion in renal collecting duct cells. FXR might represent a novel target for the treatment of autosomal dominant polycystic kidney disease.

Th2 cytokine signaling through IL-4Rα increases eotaxin-3 secretion and tension in human esophageal smooth muscle.

In esophageal epithelial cells in eosinophilic esophagitis (EoE), Th2 cytokines (IL-4, IL-13) signal through IL-4Rα, activating JAK to increase eotaxin-3 secretion, which draws eosinophils into the mucosa. We explored whether Th2 cytokines also might stimulate eotaxin-3 secretion and increase tension in esophageal smooth muscle (ESM), which might impair esophageal distensibility, and whether those events could be blocked by proton pump inhibitors (PPIs) or agents that disrupt IL-4Rα signaling. We established human ESM cell cultures from organ donors, characterizing Th2 cytokine receptor and P-type ATPase expression by qPCR. We measured Th2 cytokine-stimulated eotaxin-3 secretion by enzyme-linked immunosorbent assay (ELISA) and ESM cell tension by gel contraction assay, before and after treatment with omeprazole, ruxolitinib (JAK inhibitor), or IL-4Rα blocking antibody. CPI-17 (inhibitor of a muscle-relaxing enzyme) effects were studied with CPI-17 knockdown by siRNA or CPI-17 phospho(T38A)-mutant overexpression. ESM cells expressed IL-4Rα and IL-13Rα1 but only minimal H+-K+-ATPase mRNA. Th2 cytokines increased ESM eotaxin-3 secretion and tension, effects blocked by ruxolitinib and IL-4Rα blocking antibody but not consistently blocked by omeprazole. IL-13 increased ESM tension by increasing CPI-17 expression and phosphorylation, effects blocked by CPI-17 knockdown. Blocking IL-4Rα decreased IL-13-stimulated eotaxin-3 secretion, CPI-17 expression, and tension in ESM. Th2 cytokines increase ESM eotaxin-3 secretion and tension via IL-4Rα signaling that activates CPI-17. Omeprazole does not reliably inhibit this process, but IL-4Rα blocking antibody does. This suggests that ESM eosinophilia and impaired esophageal distensibility might persist despite elimination of mucosal eosinophils by PPIs, and IL-4Rα blocking agents might be especially useful in this circumstance.NEW & NOTEWORTHY We have found that Th2 cytokines increase eotaxin-3 secretion and tension in esophageal smooth muscle (ESM) cells via IL-4Rα signaling. Unlike esophageal epithelial cells, ESM cells do not express H+-K+-ATPase, and omeprazole does not inhibit their cytokine-stimulated eotaxin-3 secretion or tension. An IL-4Rα blocking antibody reduces both eotaxin-3 secretion and tension induced by Th2 cytokines in ESM cells, suggesting that an agent such as dupilumab might be preferred for patients with EoE with esophageal muscle involvement.

Dietary magnesium chelate alleviates oxidative stress and improves growth in white-leg shrimp, Penaeus vannamei (Boone, 1931), reared in inland saline water

Nine semi-purified diets containing either magnesium citrate (MgC) levels of 0.0, 2.24, 4.48, 6.72, 8.96 g Kg-1 or magnesium amino acid complex (MgAA) levels of 2.17, 4.35, 6.52, 8.7 g Kg-1 were fed to P. vannamei juveniles thrice daily and reared for 60 days in sub-optimum inland saline ground water (ISGW) of 10‰. Five hundred sixty-seven healthy juvenile shrimps (3.51 ± 0.03 g) were randomly stocked in triplicate into 350 L tanks at a density of 60 m-3. The weight gain (WG %) and specific growth rate were highest in the shrimps fed with 4.48 g Kg-1 MgC and 2.17 g Kg-1 MgAA. Further increase in the dietary magnesium did not show any additional growth, relatively inhibited it. The positive correlations (p < 0.01) between WG % and magnesium contents of serum, body and hepatopancreas suggests indirect involvement of tissue magnesium in growth performance. Conversely, negative correlations (p < 0.01) of WG % with superoxide dismutase and heat shock protein 70 (HSP 70) in gill and hepatopancreas indicate stress-induced inhibition of growth. Higher inclusion of dietary magnesium altered sodium-potassium ATPase (NKA) mRNA expression in hepatopancreas. The minimum dietary MgC and MgAA supplementations of P. vannamei juveniles for growth improvement were determined to be 4.25 g Kg-1 and 2.38 g Kg-1 feed, respectively, which was equivalent to 2.86 g Kg-1 and 1.64 g Kg-1 feed respectively, using broken-line regression analysis based on the weight gain (%). Overall results reveal that dietary MgAA was better than MgC in alleviating ionic stress and improving growth performance and survival rate in shrimp reared in 10‰ ISGW of the sub-optimum magnesium suite.

Cardiotoxicity of (-)-borneol, (+)-borneol, and isoborneol in zebrafish embryos is associated with Na+ /K+ -ATPase and Ca2+ -ATPase inhibition.

Borneol is an example of traditional Chinese medicine widely used in Asia. There are different isomers of chiral borneol in the market, but its toxicity and effects need further study. In this study, we used zebrafish embryos to examine the effects of exposure to three isomers of borneol [(-)-borneol, (+)-borneol, and isoborneol] on heart development and the association with Na+ /K+ -ATPase from 4 h post-fertilization (4 hpf). The results showed that the three isomers of borneol increased mortality and decreased hatching rate when the zebrafish embryo developed to 72 hpf. All three isomers of borneol (0.01-1.0 mM) significantly reduced heart rate from 48 to 120 hpf and reduced the expression of genes related to Ca2+ -ATPase (cacna1ab and cacna1da) and Na+ /K+ -ATPase (atp1b2b, atp1a3b, and atp1a2). At the same time, the three isomers of borneol significantly reduced the activities of Ca2+ -ATPase and Na+ /K+ -ATPase at 0.1 to 1.0 mM. (+)-Borneol caused the most significant reduction (p < 0.05), followed by isoborneol and (-)-borneol. Na+ /K+ -ATPase was mainly expressed in otic vesicles and protonephridium. All three isomers of borneol reduced Na+ /K+ -ATPase mRNA expression, but isoborneol was the most significant (p < 0.01). Our results indicated that (+)-borneol was the least toxic of the three isomers while the isoborneol showed the most substantial toxic effect, closely related to effects on Na+ /K+ -ATPase.

Pharmacological Basis of Rumex hastatus D. Don in Gastrointestinal Diseases with Focusing Effects on H+/K+-ATPase, Calcium Channels Inhibition and PDE Mediated Signaling: Toxicological Evaluation on Vital Organs.

This present study aimed to delineate Rumex hastatus D. Don crude extract (Rh.Cr), n-Hexane, ethyl acetate, aqueous fractions (Rh.n-Hex, Rh.ETAC, Rh.Aq) and rutin for antidiarrheal, antisecretory effects, anti-spasmodic, gastrointestinal transient time, anti H. pylori, antiulcer effects, and toxicology. The preliminary phytochemical analysis of Rumex hastatus showed different phytoconstituents and shows different peaks in GC-MC chromatogram. Rumex hastatus crude extract (Rh.Cr), fractions, and rutin attributed dose-dependent (50–300 mg/kg) protection (0–100%) against castor oil-induced diarrhea and dose-dependently inhibited intestinal fluid secretions in mice. They decreased the distance traversed by charcoal in the gastrointestinal transit model in rats. In rabbit jejunum preparations, Rh.Cr and Rh.ETAC caused a concentration-dependent relaxation of both spontaneous and K+ (80 mM)-induced contractions at a similar concentration range, whereas Rh.n-Hex, rutin, and verapamil were relatively potent against K+-induced contractions and shifted the Ca2+ concentration–response curves (CRCs) to the right, Rh.Cr (0.3–1 mg/mL) and Rh.ETAC (0.1–0.3 mg/mL) shifted the isoprenaline-induced inhibitory CRCs to the left. Rh.n-Hex, Rh.ETAC and rutin showed anti-H. pylori effect, also shows an inhibitory effect against H+/K+-ATPase. Rumex hastatus showed gastroprotective and antioxidant effects. Histopathological evaluation showed improvement in cellular architecture and a decrease in the expression of inflammatory markers such as, cyclooxygenase (COX-2), tumor necrosis factor (TN,F-α) and phosphorylated nuclear factor kappa B (p-NFƙB), validated through immunohistochemistry and ELISA techniques. In RT-PCR it decreases H+/K+-ATPase mRNA levels. Rumex hastatus was found to be safe to consume up to a dose of 2000 mg/kg in a comprehensive toxicity profile. Docking studies revealed that rutin against H+/K+-ATPase pump and voltage-gated L-type calcium channel showed E-values of −8.7 and −9.4 Kcal/mol, respectively. MD simulations Molecular Mechanics Poisson Boltzmann surface area and molecular mechanics Generalized Born surface area (MMPBSA/GBSA) findings are consistent with the in-vitro, in-vivo and docking results.

Pharmacological investigation of brucine anti-ulcer potential.

Gastric ulcer is one of the most common chronic gastrointestinal diseases characterized by a significant defect in the mucosal barrier. The current study has been conducted to evaluate the brucine anti-ulcer effect. Brucine has binding energy values ranging from −2.99 to −8.11 kcal/mol against chosen targets, according to in silico research. Brucine exhibits an inhibitory effect against Helicobacter pylori. In vivo findings revealed that brucine (3 mg/kg) showed effective results in healing ethanol-induced ulcer lesions of the gastric region in rats. Brucine showed an inhibitory effect against H+/K+-ATPase. Levels of glutathione, glutathione-s-transferase, and catalase were enhanced in the gastric rat tissue with the use of brucine, while a significant decrease in lipid peroxide levels was seen. Histopathological evaluation showed improvement in cellular architecture and a decrease in inflammatory indicators like cyclooxygenase, tumor necrosis factor, and nuclear factor kappa B expression, validated through immunohistochemistry, enzyme-linked immunosorbent assay, and Western blot techniques. In the reverse transcription–polymerase chain reaction, brucine decreased H+/K+-ATPase mRNA levels. This study reveals that brucine possesses stable binding affinities against selected targets. Brucine exhibits an anti-ulcer effect, mediated via anti-H. pylori, H+/K+-ATPase inhibition, and antioxidant and anti-inflammatory pathways.

Effect of Rumex dentatus on Gastrointestinal Protection and Toxicology in Rodents via Investigating H+/K+-ATPase, Calcium Channels, and PDE Mediated Signaling

This present study aims to delineate Rumex dentatus crude extract (Rd.Cr), n-Hexane, ethyl acetate, aqueous fractions (Rd.n-Hex, Rd.ETAC, and Rd.Aq), and emodin for antidiarrheal, antisecretory effects, anti-spasmodic, gastrointestinal transient time, anti-H. pylori, antiulcer effects, and toxicology. Plant extracts attributed dose-dependent protection against castor oil-induced diarrhea and dose-dependently inhibited intestinal fluid secretions in mice. They decreased the distance transverse by charcoal in the gastrointestinal transit model in rats. In rabbit jejunum preparations, it causes a concentration-dependent relaxation of both spontaneous and K+ (80 mM)-induced contraction, Rd.n-Hex and verapamil were relatively potent against K+-induced contractions and shifted the Ca2+ concentration-response curves (CRCs) to the right, Rd.Cr and Rd.ETAC shifted the isoprenaline-induced inhibitory CRCs to the left, showing potentiating effect similar to papaverine. Rd.n-Hex showed anti-H. pylori effect. Extracts and emodin also show an inhibitory effect against H+/K+-ATPase. Rumex dentatus showed a gastroprotective and antioxidant effect. Histopathological evaluation showed improvement in cellular architecture and decrease in the expression of inflammatory markers such as cyclooxygenase (COX2), tumor necrosis factor (TNF-α), and phosphorylated nuclear factor kappa B (p-NFƙB), validated through immunohistochemistry, ELISA, and western blot techniques. In RT-PCR, it decreases H+/K+-ATPase mRNA levels. Rumex dentatus was analyzed for certain safety aspects and exhibited a relative safety profile as no impairment was observed in kidneys, heart, liver, and brain further assisted by biochemical and hematological analysis. Docking studies revealed that emodin against H+/K+-ATPase pump and voltage gated L-type calcium channel showed E-value of −7.9 and −7.4 kcal/mol, respectively. MD simulations and molecular mechanics Poisson Boltzmann surface area and molecular mechanics Generalized Born surface area MMPBSA/GBSA findings are consistent with the in-vitro, in-vivo, and docking results. In conclusion, Rumex dentatus extracts and its phytoconstituent could be considered a potent antioxidant and anti-inflammatory drug candidates that possess anti-diarrheal, anti-secretary, antispasmodic, anti-H. pylori, and anti-ulcer potential. Toxicity studies were done according to OECD standards 425. It belongs to group 5 (LD50 > 2000 mg/kg), which suggests that it is in the lower toxicity class.

Potential mechanisms of Na+/K+-ATPase attenuation by heat and pesticides co-exposure in goldfish: role of cellular apoptosis, oxidative/nitrative stress, and antioxidants in gills.

In this study, we examined the dose-dependent effects of an environmentally relevant pesticide cocktail (metalachlor, linuron, isoproturon, tebucanazole, aclonifen, atrazine, pendimethalin, and azinphos-methyl) and temperature change (22 vs. 32°C for 4-week exposure) on Na+/K+-ATPase, 3-nitrotyrosine protein (NTP), dinitrophenyl protein (DNP), catalase (CAT), and superoxide dismutase (SOD) expressions in gills of goldfish (Carassius auratus). Histopathological analysis showed widespread damage to gill in elevated temperature (32°C) and pesticide co-exposure groups, including fusion of secondary lamellae, club-shaped primary lamellae, rupture of epithelial layer, loss of normal architecture, and hemorrhaging. Immunohistochemical and qRT-PCR analyses showed significant decreases in Na+/K+-ATPase protein and mRNA expressions in gills exposed to higher temperature and pesticides; however, combined exposure to heat and pesticides significantly increases NTP, DNP, CAT, and SOD expressions. In situ TUNEL assay revealed elevated levels of apoptotic cells in response to combined exposure. Collectively, our results suggest the combined effects of heat and pesticide stress cause cellular damage, upregulate oxidative/nitrative stress biomarkers, and increase apoptotic cells, downregulate Na+/K+-ATPase expression in gills. This provides new evidence for oxidant/antioxidant-dependent mechanisms for downregulation of Na+/K+-ATPase expression in gills during combined exposure.

Effect of high salinity acclimation on glucose homeostasis in Mozambique tilapia (Oreochromis mossambicus).

During salinity stress, osmoregulatory processes in euryhaline fish need to modify for their survival, and glucose is the preferred mode of extra energy during such conditions. These organisms must have a proper mechanism to maintain glucose homeostasis during such modified osmoregulatory process across different body fluids. Hence, we studied high salinity effect on regulation of glucose homeostasis in Mozambique tilapia. The fish were induced to 15‰ salinity for 21 days. Glucose, glycogen, ion concentrations, Na+-K+-ATPase, pyruvate kinase, γ-amylase activities and GLUT mRNA expressions were investigated in liver, intestine, gill and white muscle tissues. At the end of experiment, Na+ ion concentrations, glucose content and activity of Na+-K+-ATPase especially in the gill and intestine were increased, while decrease in liver and gill glycogen content was seen. Lower concentration of glycogen decrease was observed in the intestine and white muscle of the treated group. High pyruvate kinase activity was noticed in liver and gill tissues that correlates with high Na+-K+-ATPase activity. Elevated γ-amylase activity was observed in the liver and intestine suggesting breakdown of glycogen; however, gill and white muscle did not show any increased activity. Increase in GLUT1 and GLUT4 mRNA expressions was observed especially in the gill and intestine, while increase in GLUT2 mRNA expressions was observed in the liver. Upregulations of GLUTs suggest higher influx of glucose into the cell for catabolism to provide energy and further to drive the enhanced osmoregulatory process. These findings suggest glucose homeostasis being regulated in Mozambique tilapia during salinity acclimation.

Soybean Bioactive Peptides Improves The Energy Metabolism Of Muscle In Mice

INTRODUCTION AND PURPOSE: Soybean bioactive peptides (SBP), a plant-derived nutrition supplement, regulates various metabolism and improves health. However, which dose of SBP best enhances energy metabolism is unclear. This study explored different doses of SBP related to energy metabolism of muscle in mice. METHODS: C57BL/6 mice were randomly divided into four groups (n = 6 per group): Control (G0), Low Dose (G1), Medium Dose (G2), and High Dose (G3). All mice fed by oral gavage for 30 consecutive days (once per day): G0 fed pure water, G1, G2, and G3 fed SBP 110 mg/kg.BW, 220 mg/kg.BW, and 440 mg/kg.BW. Then all mice took exhaustive exercise and tested respiratory chain complex II and IV (RCCII, RCCIV), Mn superoxide dismutase (Mn-SOD), ATPase, and AMPK mRNA expression in mitochondria of quadriceps femoris. RESULTS: Compared to G0: (1) In RCCIV, G3 increased 134.5% (460.936 ± 171.14 vs. 196.514 ± 37.47 pg/mL, p < 0.05); (2) In Mn-SOD, G3 increased 134.4% (769.79 ± 285.67 vs. 328.41 ± 62.54 pg/mL, p < 0.05); (3) In AMPK mRNA expression, G3 increased 81.5% (1.48 ± 0.43 vs. 0.81 ± 0.09, p < 0.05). Besides, there was no statistical significance in RCCII and ATPase. CONCLUSION: Our finding demonstrated that the high dose of SBP has a greater effect on energy metabolism of muscle.

The levels of follicular fluid cell-free mitochondrial DNA could serve as a biomarker for pregnancy success in patients with small ovarian endometriosis cysts: A case-control study.

Ovarian endometriosis cyst (OEC) is caused by the growth of ectopic endometrium into the ovarian cortex, leading to disrupted ovarian cortical structures and infertility. Large OECs are usually surgically removed, and assisted reproductive technology (ART) is required for future pregnancy. The oocyte reserve and development of patients with small non-surgical OECs are unknown. In this study, we compared mitochondrial abnormality, ATPase and IF1 mRNA expression levels, and OXPHO complex proteins between OEC vs control mural granulosa cells (mGCs).OEC mGCs show fewer mitochondria per cell, a higher proportion of aberrant morphology, lower ATPase mRNA levels, higher IF1 mRNA levels, and impaired expression of 3 of the 5 critical proteins involved in the OXPHOS complex, compared with control mGCs. Cell-free mitochondrial DNA (cfmtDNA) levels are higher in the follicular fluid of patients with OEC and were inversely associated with the expression of mtDNA in mGCs and cumulus granulosa cells (cGCs).Taken together, this study indicates that small non-surgical OECs lead to poor quality of oocytes and subsequent embryos during ART compared with control, which was accompanied by mGC mitochondrial dysfunction. mGC and cGC mtDNA and FF cfmtDNA might serve as efficient biomarkers for the non-invasive prediction of pregnancy outcomes in patients with OEC undergoing ART.

Expression of ion transport genes in ionocytes isolated from larval zebrafish (Danio rerio) exposed to acidic or Na+-deficient water.

In zebrafish (Danio rerio), a specific ionocyte subtype, the H+-ATPase-rich (HR) cell, is presumed to be a significant site of transepithelial Na+ uptake/acid secretion. During acclimation to environments differing in ionic composition or pH, ionic and acid-base regulations are achieved by adjustments to the activity level of HR cell ion transport proteins. In previous studies, the quantitative assessment of mRNA levels for genes involved in ionic and acid-base regulations relied on measurements using homogenates derived from the whole body (larvae) or the gill (adult). Such studies cannot distinguish whether any differences in gene expression arise from adjustments of ionocyte subtype numbers or transcriptional regulation specifically within individual ionocytes. The goal of the present study was to use fluorescence-activated cell sorting to separate the HR cells from other cellular subpopulations to facilitate the measurement of gene expression of HR cell-specific transporters and enzymes from larvae exposed to low pH (pH 4.0) or low Na+ (5 μM) conditions. The data demonstrate that treatment of larvae with acidic water for 4 days postfertilization caused cell-specific increases in H+-ATPase (atp6v1aa), ca17a, ca15a, nhe3b, and rhcgb mRNA in addition to increases in mRNA linked to cell proliferation. In fish exposed to low Na+, expression of nhe3b and rhcgb was increased owing to HR cell-specific regulation and elevated numbers of HR cells. Thus, the results of this study demonstrate that acclimation to low pH or low Na+ environmental conditions is facilitated by HR cell-specific transcriptional control and by HR cell proliferation.

Assessment of ionic homeostasis imbalance and cytochrome P450 system disturbance in mice during fumonisin B1 (FB1) exposure

Fumonisin B1 (FB1) is a mycotoxin frequently found in agricultural commodities, and poses a considerable risk for human and animal health. The aim of this study was to investigate the toxic effect of FB1 in mice intestine. Male Kunming mice (n = 40) were treated with FB1 diet for 42 days. Histopathological and biochemical analyses, including ion concentrations, transcription of ATPase subunits and mRNA expression of cytochrome P450s (CYP450s) analyses were performed on duodenum, cecum and colon of mice. The results revealed that FB1 caused histological alterations, including partial shedding of villous epithelial cells and inflammatory cell infiltration. Furthermore, a significant change in Na+, K+ and Ca2+ in serum, and the mRNA expression of ATPase subunits and CYP450s in intestinal tracts were observed in FB1-exposed mice. Our results suggested that FB1 exposure induce histopathological injury via disrupting CYP isoforms transcription and triggering ion homeostasis imbalance in mice intestinal tracts.

Comparison of the efficacy of a Tabernaemontana divaricata extract and of biosynthesized silver nanoparticles in preventing cataract formation in an in-vivo system of selenite-induced cataractogenesis

Abstract The putative anticataractogenic efficacy of silver nanoparticles (AgNPs), biosynthesized with a Tabernaemontana divaricata (T. divaricata) extract, was evaluated in-vivo in four groups of Wistar rat pups: Group I (normal [control]); Group II (selenite-challenged [19 μmol/kg body weight], untreated); Group III (selenite-challenged, T. divaricata extract-treated [350 mg/kg body weight]); and Group IV (selenite-challenged, biosynthesized AgNPs-treated [200 mg/kg body weight]). Mean levels/activities of lenticular reduced glutathione (GSH), malondialdehyde (MDA), calcium, Ca2+ and Na+/K+-ATPase and total calpain were measured. The expression pattern of genes at mRNA transcript (by reverse transcription-PCR) and protein (by immunoblot analysis) levels for Ca2+ transporters, calpain isoforms and lenticular proteins (αA- and βB1-crystallins, and vimentin) were evaluated. In extract-treated and AgNPs-treated rats, relatively higher mean lenticular levels/activities of GSH, Ca2+ and Na+/K+-ATPase, total calpain, and higher mRNA transcript and protein levels of calpain isoforms and lenticular proteins were observed. However, relatively lower mean lenticular levels of MDA, calcium and also mRNA transcript and protein levels of plasma membrane Ca2+-ATPase-1 (PMCA-1) and sarco/endoplasmic reticulum Ca2+-ATPase-2 (SERCA-2) were recorded. The prevention of such lenticular alterations was more pronounced in AgNPs-treated rats than in plain extract-treated rats. These observations suggest that AgNPs biosynthesized with T. divaricata extract have the potential to prevent selenite-induced cataractogenesis in-vivo by maintaining normal lenticular calcium homeostasis and calpain cascade activities and by preventing alterations in key lenticular proteins.

Energy Metabolism and Redox State in Brains of Wistar Audiogenic Rats, a Genetic Model of Epilepsy.

The Wistar Audiogenic Rat (WAR) strain is a genetic model of epilepsy, specifically brainstem-dependent tonic-clonic seizures, triggered by acute auditory stimulation. Chronic audiogenic seizures (audiogenic kindling) mimic temporal lobe epilepsy, with significant participation of the hippocampus, amygdala, and cortex. The objective of the present study was to characterize the mitochondrial energy metabolism in hippocampus and cortex of WAR and verify its relationship with seizure severity. Hippocampus of WAR naïve (no seizures) presented higher oxygen consumption in respiratory states related to the maximum capacities of phosphorylation and electron transfer system, elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl and lactate contents, compared with their Wistar counterparts. Audiogenic kindling had no adding functional effect in WAR, but in Wistar, it induced the same alterations observed in the audiogenic strain. In the cortex, WAR naïve presented elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl levels. Chronic acoustic stimulation in Wistar induced the same alterations in cortex and hippocampus. Mainly in the hippocampus, WAR naïve presented elevated mRNA expression of glucose, lactate and excitatory amino acids transporters, several glycolytic enzymes, lactate dehydrogenase, and Na+/K+ ATPase in neurons and in astrocytes. In vivo treatment with mitochondrial uncoupler 2,4-dinitrophenol (DNP) or N-acetylcysteine (NAC) in WAR had no effect on mitochondrial metabolism, but lowered oxidative stress. Unlike DNP, NAC downregulated all enzyme genes involved in glucose and lactate uptake, and metabolism in neurons and astrocytes. Additionally, it was able to reduce brainstem seizure severity in WAR. In conclusion, in WAR naïve animals, both cerebral cortex and hippocampus display elevated mitochondrial density and/or activity associated with oxidative damage, glucose and lactate metabolism pathways upregulation, and increased Na+/K+ ATPase mRNA expression. Only in vivo treatment with NAC was able to reduce seizure severity of kindled WARs, possibly via down regulation of glucose/lactate metabolism. Taken together, our results are a clear contribution to the field of mitochondrial metabolism associated to epileptic seizures.

Long-term Follow-up of Interleukin-1B and H+,K+-ATPase mRNA Expression After Helicobacter Pylori Eradication

Long-term Follow-up of Interleukin-1B and H+,K+-ATPase mRNA Expression After Helicobacter Pylori Eradication

Modulation of exercise training related adaptation of body composition and regulatory pathways by anabolic steroids

Anabolic steroids have a long history of abuse in amateur and professional athletics. However, their interaction with training and the resulting effects on body composition and tissue adaptation, relying on a concert of factors and pathways, remain under investigation. This study aims at investigating the changes of body composition and the expression of selected genes and pathways essential for this adaptation process. Therefore, male wistar rats were treated with the anabolic steroid metandienone in two groups (n = 16; metandienone, metandienone + exercise) alongside with control groups (n = 16; control, exercise). Following a 6-week steep-angle treadmill training protocol, weight of organs, visceral fat and muscles was determined. M. gastrocnemius was histologically assessed by ATPase staining, mRNA and protein levels of factors of regeneration, hypertrophy and myogenesis and selected master regulators and markers were determined. Results show additive effects of anabolic steroids and exercise on body, tibia and reproductive organs weight. Mm. gastrocnemius and soleus weight was increased by training but not anabolic steroids. Muscle fiber diameter and composition remained unchanged. Visceral fat mass and fat cell size was affected by training and anabolic steroids but no additive effects could be observed. Exercise and anabolic steroids result in a complex regulation of the expression of genes in M. Gastrocnemius involved in skeletal muscle metabolism, hypertrophy, inflammation and regeneration. In summary, our data suggests distinct molecular mechanisms involved in the adaptation of the skeletal muscle to anabolic androgenic steroids and exercise. Metandienone treatment neither results in skeletal muscle hypertrophy nor liver-toxic effects but in an induction of skeletal muscle regeneration and an activation of endocrine negative feedback. Moreover our study demonstrates that visceral fat and bone responds with higher sensitivity to ASS and exercise than the skeletal muscle. This apparent plasticity of adipose and bone tissue rather than skeletal muscle could indicate a potentially superior future role of fat rather than muscle related parameters to detect and AAS abuse in a biologic passport strategy in professional athletes.

A Novel Synthetic Dihydroindeno[1,2-b] Indole Derivative (LS-2-3j) Reverses ABCB1- and ABCG2-Mediated Multidrug Resistance in Cancer Cells.

10-oxo-5-(3-(pyrrolidin-1-yl) propyl)-5,10-dihydroindeno [1,2-b] indol-9-yl propionate (LS-2-3j) is a new chemically synthesized indole compound and some related analogues are known to be inhibitors (such as alectinib and Ko143) of ATP-binding cassette (ABC) transporters, especially the ABC transporter subfamily B member 1 (ABCB1) and the ABC transporter subfamily G member 2 (ABCG2). This study aimed to evaluate the multidrug resistance (MDR) reversal effects and associated mechanisms of LS-2-3j in drug-resistant cancer cells. The inhibition of cell proliferation in tested agents was evaluated by the 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide (MTT) assay. Accumulation or efflux of chemotherapy drugs was analyzed by flow cytometry. The ATPase activity was measured using an ATPase activity assay kit. The mRNA transcripts and protein expression levels were detected by real-time PCR and Western blot, respectively. In this connection, LS-2-3j significantly enhanced the activity of chemotherapeutic drugs in MDR cells and could significantly increase the intracellular accumulation of doxorubicin (DOX) and mitoxantrone (MITX) by inhibiting the function of the efflux pumps in ABCB1- or ABCG2-overexpressing cells. Furthermore, reduced ATPase activity, mRNA transcription, and protein expression levels of ABCB1 and ABCG2 were observed in a concentration dependent manner in MDR cancer cells.

Caste-Specific Expression of Na+/K+-ATPase in the Asian Weaver Ant, Oecophylla smaragdina (Fabricius, 1775).

Social insect colonies adopt different levels of survival strategies and exhibit well-defined reproductive division of labour. Oecophylla smaragdina (Fabricius, 1775) has physiological and behavioral adaptations that enable them to forage at extreme environmental conditions and are lethal to most other insects. Ion homeostasis is the key process in an organism's survival mechanism. Among ion pumps, the ATP-dependent sodium-potassium ion pump is essential for maintaining the Na+ and K+ ionic balance and is well known as the primary consumer of energy. Oecophylla smaragdina plays pivotal role as a model among social insects for understanding ion homeostasis at the organization level of the castes. We have evaluated the expression and activity of Na+/K+-ATPase among various castes of O. smaragdina (worker subcastes, queen and male). Real-time PCR and immunoblotting analyses revealed the differential expression of Na+/K+-ATPase in the castes. Significantly higher expression of Na+/K+-ATPase mRNA and protein were observed in the minor workers, queen, major workers and males respectively. These results suggest that in the weaver ant colony, the castes might have variously adapted and evolved with a well-developed ion transport mechanism which allows them to perform allocated tasks within the nest and could be a key to their adaptive benefits towards division of labour.