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Journal of Autism and Developmental... Mar 2024We present the secondary-analysis of neurocognitive tests in the 'Bumetanide in Autism Medication and Biomarker' (BAMBI;EUDRA-CT-2014-001560-35) study, a randomized... (Randomized Controlled Trial)
Randomized Controlled Trial
We present the secondary-analysis of neurocognitive tests in the 'Bumetanide in Autism Medication and Biomarker' (BAMBI;EUDRA-CT-2014-001560-35) study, a randomized double-blind placebo-controlled (1:1) trial testing 3-months bumetanide treatment (≤ 1 mg twice-daily) in unmedicated children 7-15 years with ASD. Children with IQ ≥ 70 were analyzed for baseline deficits and treatment-effects on the intention-to-treat-population with generalized-linear-models, principal component analysis and network analysis. Ninety-two children were allocated to treatment and 83 eligible for analyses. Heterogeneous neurocognitive impairments were found that were unaffected by bumetanide treatment. Network analysis showed higher modularity after treatment (mean difference:-0.165, 95%CI:-0.317 to - 0.013,p = .034) and changes in the relative importance of response inhibition in the neurocognitive network (mean difference:-0.037, 95%CI:-0.073 to - 0.001,p = .042). This study offers perspectives to include neurocognitive tests in ASD trials.
Topics: Child; Humans; Autism Spectrum Disorder; Bumetanide; Autistic Disorder; Intention; Linear Models
PubMed: 36626004
DOI: 10.1007/s10803-022-05841-3 -
Journal of Immunology Research 2022Interstitial cystitis/bladder pain syndrome (IC/BPS) is a chronic, severely distressing clinical syndrome characterized by bladder pain and pressure perceptions. The...
Identification of Immune-Related Genes and Small-Molecule Drugs in Interstitial Cystitis/Bladder Pain Syndrome Based on the Integrative Machine Learning Algorithms and Molecular Docking.
BACKGROUND
Interstitial cystitis/bladder pain syndrome (IC/BPS) is a chronic, severely distressing clinical syndrome characterized by bladder pain and pressure perceptions. The origin and pathophysiology of IC/BPS are currently unclear, making it difficult to diagnose and formulate successful treatments. Our study is aimed at investigating the role of immune-related genes in the diagnosis, progression, and therapy of IC/BPS.
METHOD
The gene expression datasets GSE11783, GSE11839, GSE28242, and GSE57560 were retrieved from the GEO database for further analysis. Immune-related IC/BPS differentially expressed genes (DEGs) were identified by limma. Three distinct machine learning approaches, least absolute shrinkage and selection operator (LASSO), support vector machine-recursive feature elimination (SVM-RFE), and random forest (RF), were used to find the immune-related IC characteristic genes. Nomogram and receiving operator curves (ROC) were plotted to measure characteristic effectiveness. Using the CMap database and the molecular docking approach, potential small-molecule medicines were found and verified. Consensus cluster analysis was also performed to separate the IC/BPS samples into immunological subtypes.
RESULTS
A total of 24 immune-related IC/BPS-DEGs were identified. When compared to the normal control group, the IC/BPS cohort had significantly more immune cell infiltration. Integrative machine learning methods discovered 5 IC/BPS characteristic genes (RASGRP1, PPBP, RBP4, CR2, and PROS2) that may predict IC/BPS diagnosis and immune cell infiltration. Furthermore, two immunological subgroups with substantial variations in immune cell infiltration across IC/BPS samples were identified, which were named cluster1 and cluster2, with the hallmark genes having greater expression in cluster2. Finally, bumetanide was shown to have the potential to be a medication for the treatment of IC/BPS, and it performed well in terms of its molecular binding with RASGRP1.
CONCLUSION
We found and validated 5 immune-related IC/BPS genes (RASGRP1, PPBP, RBP4, CR2, and PROS2) and 2 IC/BPS immune subtypes. In addition, bumetanide was discovered to be a potential drug for treating IC/BPS, which may provide new insight into the diagnosis and immune therapy of IC/BPS patients.
Topics: Humans; Cystitis, Interstitial; Molecular Docking Simulation; Bumetanide; Algorithms; Guanine Nucleotide Exchange Factors; Retinol-Binding Proteins, Plasma
PubMed: 36619718
DOI: 10.1155/2022/2069756 -
Cerebral Cortex (New York, N.Y. : 1991) May 2023The Na-K-2Cl cotransporter NKCC1 is widely expressed in cells within and outside the brain. However, our understanding of its roles in brain functions throughout...
The Na-K-2Cl cotransporter NKCC1 is widely expressed in cells within and outside the brain. However, our understanding of its roles in brain functions throughout development, as well as in neuropsychiatric and neurological disorders, has been severely hindered by the lack of reliable data on its developmental and (sub)cellular expression patterns. We provide here the first properly controlled analysis of NKCC1 protein expression in various cell types of the mouse brain using custom-made antibodies and an NKCC1 knock-out validated immunohistochemical procedure, with parallel data based on advanced mRNA approaches. NKCC1 protein and mRNA are expressed at remarkably high levels in oligodendrocytes. In immature neurons, NKCC1 protein was located in the somata, whereas in adult neurons, only NKCC1 mRNA could be clearly detected. NKCC1 immunoreactivity is also seen in microglia, astrocytes, developing pericytes, and in progenitor cells of the dentate gyrus. Finally, a differential expression of NKCC1 splice variants was observed, with NKCC1a predominating in non-neuronal cells and NKCC1b in neurons. Taken together, our data provide a cellular basis for understanding NKCC1 functions in the brain and enable the identification of major limitations and promises in the development of neuron-targeting NKCC1-blockers.
Topics: Mice; Animals; Solute Carrier Family 12, Member 2; Brain; Neurons; RNA, Messenger; Hippocampus
PubMed: 36573432
DOI: 10.1093/cercor/bhac470 -
Scientific Reports Dec 2022Na/K-ATPases (NKA) in the basolateral membrane of the intestinal enterocytes create a Na-gradient that drives both ion-coupled fluid uptake and nutrient transport. Being...
Na/K-ATPases (NKA) in the basolateral membrane of the intestinal enterocytes create a Na-gradient that drives both ion-coupled fluid uptake and nutrient transport. Being dependent on the same gradient as well as on the environmental salinity, these processes have the potential to affect each other. In salmonids, L-lysine absorption has been shown to be higher in freshwater (FW) than in seawater (SW) acclimated fish. Using electrophysiology (Ussing chamber technique), the aim was to explore if the decrease in L-lysine transport was due to allocation of the Na-gradient towards ion-driven fluid uptake in SW, at the cost of amino acid transport. Intestinal NKA activity was higher in SW compared to FW fish. Exposure to ouabain, an inhibitor of NKA, decreased L-lysine transport. However, exposure to bumetanide and hydrochlorothiazide, inhibitors of Na, K, 2Cl-co-transporter (NKCC) and Na, Cl-co-transporter (NCC) respectively, did not affect the rate of intestinal L-lysine transport. In conclusion, L-lysine transport is Na-dependent in rainbow trout and the NKA activity and thus the available Na-gradient increases after SW acclimation. This increased Na-gradient is most likely directed towards osmoregulation, as amino acid transport is not compromised in SW acclimated fish.
Topics: Animals; Oncorhynchus mykiss; Salinity; Lysine; Intestines; Sodium; Acclimatization; Symporters; Seawater; Gills; Sodium-Potassium-Exchanging ATPase
PubMed: 36564520
DOI: 10.1038/s41598-022-26904-6 -
Acta Bio-medica : Atenei Parmensis Dec 2022Neonatal stroke is the second cause of acute symptomatic neonatal seizures after hypoxic-ischemic encephalopathy. The aim of this systematic review is to determine which...
BACKGROUND AND AIM
Neonatal stroke is the second cause of acute symptomatic neonatal seizures after hypoxic-ischemic encephalopathy. The aim of this systematic review is to determine which drug among those available represents the best therapeutic choice for treatment of secondary seizures due to neonatal stroke.
METHODS
We performed a systematic review searching on PubMed the keywords "Neonatal", "Stroke", "Seizures" and "Treatment". Search was limited only to English language with no time limit. Last literature search was done on May 30, 2022.
RESULTS
We selected 5 articles involving a total of 52 full-term neonates. In 96.1% the first line treatment was phenobarbital and in 3.9% was used phenobarbital associated with midazolam from the seizure onset but in all of these cases it was necessary to introduce further medications for controlling the seizures. As second line treatment was used lidocaine (response rate of 53.3%), midazolam (response rate of 15.38%) bumetanide (response rate of 100%), and fosphenytoin (no response). As third line treatment was used lidocaine (response rate of 87.5%), Midazolam (response rate of 60%), levetiracetam and clonazepam (response rate of 100%).
CONCLUSIONS
Our review shows that the use of ASMs that act throughout a gabaergic mechanism are inadequate in controlling seizures secondary to neonatal stroke in full-term newborns. Very effective seems to be lidocaine and levetiracetam with an apparent safer profile in short and long term. Bumetanide shows promising results, but they need to be confirmed by phase 3 studies.
Topics: Infant, Newborn; Humans; Levetiracetam; Anticonvulsants; Bumetanide; Midazolam; Phenobarbital; Epilepsy; Lidocaine; Stroke; Infant, Newborn, Diseases
PubMed: 36533757
DOI: 10.23750/abm.v93i6.13440 -
International Journal of Molecular... Dec 2022The secondary active Na-K-Cl cotransporter 1 (NKCC1) promotes electroneutral uptake of two chloride ions, one sodium ion and one potassium ion. NKCC1 regulates Cl...
The secondary active Na-K-Cl cotransporter 1 (NKCC1) promotes electroneutral uptake of two chloride ions, one sodium ion and one potassium ion. NKCC1 regulates Cl homeostasis, thus being implicated in transepithelial water transport and in neuronal excitability. Aberrant NKCC1 transport is linked to a variety of human diseases. The loop diuretic drugs bumetanide, furosemide, azosemide and ethacrynic acid target NKCC1, but are characterized by poor selectivity leading to severe side effects. Despite its therapeutic importance, the molecular details of the NKCC1 inhibition mechanism remain unclear. Using all-atom simulations, we predict a putative binding mode of these drugs to the zebrafish (z) and human (h) NKCC1 orthologs. Although differing in their specific interactions with NKCC1 and/or monovalent ions, all drugs can fit within the same cavity and engage in hydrophobic interactions with M304/M382 in z/hNKCC1, a proposed ion gating residue demonstrated to be key for bumetanide binding. Consistent with experimental evidence, all drugs take advantage of the K/Na ions, which plastically respond to their binding. This study not only provides atomic-level insights useful for drug discovery campaigns of more selective/potent NKCC1 inhibitors aimed to tackle diseases related to deregulated Cl homeostasis, but it also supplies a paradigmatic example of the key importance of dynamical effects when drug binding is mediated by monovalent ions.
Topics: Animals; Humans; Zebrafish; Bumetanide; Solute Carrier Family 12, Member 2; Potassium; Sodium; Chlorides; K Cl- Cotransporters
PubMed: 36499764
DOI: 10.3390/ijms232315439 -
Hypertension (Dallas, Tex. : 1979) Feb 2023Previous studies showed that miR-195a-5p was among the most abundant microRNAs (miRNAs) expressed in the kidney.
BACKGROUND
Previous studies showed that miR-195a-5p was among the most abundant microRNAs (miRNAs) expressed in the kidney.
METHODS
Lentivirus silencing of tumor necrosis factor-α (TNF) was performed in vivo and in vitro. Luciferase reporter assays confirmed that bumetanide-sensitive Na-K-2Cl cotransporter isoform A (NKCC2A) mRNA is targeted and repressed by miR-195a-5p. Radiotelemetry was used to measure mean arterial pressure.
RESULTS
TNF upregulates mmu-miR-195a-5p, and -203 and downregulates mmu-miR-30c and -100 in the medullary thick ascending limb of male mice. miR-195a-5p was >3-fold higher in the renal outer medulla of mice given an intrarenal injection of murine recombinant TNF, whereas silencing TNF inhibited miR-195a-5p expression by ≈51%. Transient transfection of a miR-195a-5p mimic into medullary thick ascending limb cells suppressed NKCC2A mRNA by ≈83%, whereas transfection with Anti-miR-195a-5p increased NKCC2A mRNA. Silencing TNF in medullary thick ascending limb cells prevented increases in miR-195 induced by 400 mosmol/kg HO medium, an effect reversed by transfection with a miR-195a-5p mimic. Expression of phosphorylated NKCC2 increased 1.5-fold in medullary thick ascending limb cells transfected with Anti-miR-195a-5p and a miR-195a-5p mimic prevented the increase, which was induced by silencing TNF in cells exposed to 400 mosmol/kg HO medium after osmolality was increased by adding NaCl. Intrarenal injection of TNF suppressed NKCC2A mRNA, whereas injection of miR-195a-5p prevented the increase of NKCC2A mRNA abundance and phosphorylated NKCC2 expression when TNF was silenced. Intrarenal injection with miR-195a-5p markedly attenuated MAP after renal silencing of TNF in mice given 1% NaCl.
CONCLUSIONS
The study identifies miR-195a-5p as a salt-sensitive and TNF-inducible miRNA that attenuates NaCl-mediated increases in blood pressure by inhibiting NKCC2A.
Topics: Animals; Male; Mice; Antagomirs; Blood Pressure; MicroRNAs; RNA, Messenger; Sodium Chloride; Solute Carrier Family 12, Member 1
PubMed: 36448465
DOI: 10.1161/HYPERTENSIONAHA.122.19794 -
Advances in Clinical and Experimental... Apr 2023Serotonin is a substance with a propulsive effect on the gastrointestinal tract. It stimulates the intestinal secretion of water and electrolytes, and plays an important...
BACKGROUND
Serotonin is a substance with a propulsive effect on the gastrointestinal tract. It stimulates the intestinal secretion of water and electrolytes, and plays an important role in the pathophysiology of secretory diarrhea. However, the influence of serotonin on intestinal absorption is very poorly understood.
OBJECTIVES
This study aimed to evaluate the serotonin and selected antagonists of serotonin receptors, i.e., ondansetron (5-HT3) and GR113808 (5-HT4), on electrogenic sodium ion absorption in the colon.
MATERIAL AND METHODS
The electrophysiologic method developed by Ussing and modified with a stimulating function on the mucosal side of the isolated colon wall was used. The influence of selected serotonergic compounds on the electrogenic transport of sodium ions under stationary conditions and mechanical stimulation was investigated. For this purpose, experiments were performed on specimens of isolated rabbit colon. Amiloride and bumetanide were used as reagents directly controlling individual ion transport. The data were analyzed using tests for paired samples (paired sample t-test, Wilcoxon signed-rank test and one-sided sign test).
RESULTS
Serotonin reduced stationary and stimulated colonic sodium absorption. The 5-HT3 receptor antagonist did not influence the studied phenomenon, while 5-HT4 antagonists acted contrary to serotonin.
CONCLUSIONS
Serotonin reduces both stationary and stimulated sodium ion absorption, thus playing an important role in the pathophysiology of secretory diarrhea. The described phenomenon depends on serotonin's action on 5-HT4 receptors.
Topics: Animals; Rabbits; Serotonin; Colon; Ondansetron; Diarrhea; Sodium
PubMed: 36374545
DOI: 10.17219/acem/155111 -
International Journal of Molecular... Oct 2022Lens ion homeostasis depends on Na,K-ATPase and NKCC1. TRPV4 and TRPV1 channels, which are mechanosensitive, play important roles in mechanisms that regulate the...
Lens ion homeostasis depends on Na,K-ATPase and NKCC1. TRPV4 and TRPV1 channels, which are mechanosensitive, play important roles in mechanisms that regulate the activity of these transporters. Here, we examined another mechanosensitive channel, piezo1, which is also expressed in the lens. The purpose of the study was to examine piezo1 function. Recognizing that activation of TRPV4 and TRPV1 causes changes in lens ion transport mechanisms, we carried out studies to determine whether piezo1 activation changes either Na,K-ATPase-mediated or NKCC1-mediated ion transport. We also examined channel function of piezo1 by measuring calcium entry. Rb uptake was measured as an index of inwardly directed potassium transport by intact mouse lenses. Intracellular calcium concentration was measured in Fura-2 loaded cells by a ratiometric imaging technique. Piezo1 immunolocalization was most evident in the lens epithelium. Potassium (Rb) uptake was increased in intact lenses as well as in cultured lens epithelium exposed to Yoda1, a piezo1 agonist. The majority of Rb uptake is Na,K-ATPase-dependent, although there also is a significant NKCC-dependent component. In the presence of ouabain, an Na,K-ATPase inhibitor, Yoda1 did not increase Rb uptake. In contrast, Yoda1 increased Rb uptake to a similar degree in the presence or absence of 1 µM bumetanide, an NKCC inhibitor. The Rb uptake response to Yoda1 was inhibited by the selective piezo1 antagonist GsMTx4, and also by the nonselective antagonists ruthenium red and gadolinium. In parallel studies, Yoda1 was observed to increase cytoplasmic calcium concentration in cells loaded with Fura-2. The calcium response to Yoda1 was abolished by gadolinium or ruthenium red. The calcium and Rb uptake responses to Yoda1 were absent in calcium-free bathing solution, consistent with calcium entry when piezo1 is activated. Taken together, these findings point to stimulation of Na,K-ATPase, but not NKCC, when piezo1 is activated. Na,K-ATPase is the principal mechanism responsible for ion and water homeostasis in the lens. The functional role of lens piezo1 is a topic for further study.
Topics: Mice; Animals; TRPV Cation Channels; Sodium-Potassium-Exchanging ATPase; Ruthenium Red; Gadolinium; Fura-2; Potassium; Sodium; Ion Transport; Ion Channels
PubMed: 36361659
DOI: 10.3390/ijms232112870 -
Long-term administration of bumetanide improve functional recovery after spinal cord injury in rats.Frontiers in Pharmacology 2022Ion disturbances are among the most remarkable deficits in spinal cord injury (SCI). GABA is an integral part of neural interaction. Action of the GABA receptor depends...
Ion disturbances are among the most remarkable deficits in spinal cord injury (SCI). GABA is an integral part of neural interaction. Action of the GABA receptor depends on the amount of intracellular chloride. Homeostasis of chloride is controlled by two co-transporters, NKCC1 and KCC2. Previous studies revealed that NKCC1 are disturbed in SCI. In this study, NKCC1 is highly expressed in the epicenter of the lesioned spinal cord at 3 hours after induction of the lesion and reached the peak around 6 hours after SCI. Bumetanide (2 and 4 mg/day), as a specific NKCC1 inhibitor, was used at 3 hours post SCI for 28 days. The functional recovery outcomes were measured by the Basso-Beattie-Bresnahan (BBB) locomotor rating scale, ladder walking test, and hot plate test. The rats that received bumetanide 4 mg/day exhibited improved recovery of locomotor function, reduction of NKCC1 gene expression, and upregulation of GAP protein levels 28 days post SCI. Histological tissue evaluations confirmed bumetanide's neuroprotective and regenerative effects. This study provides novel evidence for the benefits of bumetanide in early administration after SCI.
PubMed: 36339604
DOI: 10.3389/fphar.2022.932487