Search

In this study, the effect of cumulative ACEs experienced on human maternal DNA methylation (DNAm) was estimated while accounting for interaction with domains of ACEs in prenatal peripheral blood mononuclear cell samples from the Maternal and Developmental Risks from Environmental Stressors (MADRES) pregnancy cohort. The intergenerational transmission of ACE-associated DNAm was also explored used paired maternal (N = 120) and neonatal cord blood (N = 69) samples. Replication in buccal samples was explored in the Childrens Health Study (CHS) among adult parental (N = 31) and pediatric (N = 114) samples. We used a four-level categorical indicator variable for ACEs exposure: none (0 ACEs), low (1-3 ACEs), moderate (4-6 ACEs), and high (>6 ACEs). Effects of ACEs on maternal DNAm (N = 240) were estimated using linear models. To evaluate evidence for intergenerational transmission, mediation analysis (N = 60 mother-child pairs) was used. Analysis of maternal samples displayed some shared but mostly distinct effects of ACEs on DNAm across low, moderate, and high ACEs categories. CLCN7 and PTPRN2 was associated with maternal DNAm in the low ACE group and this association replicated in the CHS. CLCN7 was also nominally significant in the gene expression correlation analysis among maternal profiles (N = 35), along with 11 other genes. ACE-associated methylation was observed in maternal and neonatal profiles in the COMT promoter region, with some evidence of mediation by maternal COMT methylation. Specific genomic loci exhibited mutually exclusive maternal ACE effects on DNAm in either maternal or neonatal population. There is some evidence for an intergenerational effect of ACEs, supported by shared DNAm signatures in the COMT gene across maternal-neonatal paired samples.

Cholera is a global health problem with no targeted therapies. The Ca2+-sensing receptor (CaSR) is a regulator of intestinal ion transport and a therapeutic target for diarrhea, and Ca2+ is considered its main agonist. We found that increasing extracellular Ca2+ had a minimal effect on forskolin-induced Cl- secretion in human intestinal epithelial T84 cells. However, extracellular Mg2+, an often-neglected CaSR agonist, suppressed forskolin-induced Cl- secretion in T84 cells by 65% at physiological levels seen in stool (10 mM). The effect of Mg2+ occurred via the CaSR/Gq signaling that led to cAMP hydrolysis. Mg2+ (10 mM) also suppressed Cl- secretion induced by cholera toxin, heat-stable E. coli enterotoxin, and vasoactive intestinal peptide by 50%. In mouse intestinal closed loops, luminal Mg2+ treatment (20 mM) inhibited cholera toxin-induced fluid accumulation by 40%. In a mouse intestinal perfusion model of cholera, addition of 10 mM Mg2+ to the perfusate reversed net fluid transport from secretion to absorption. These results suggest that Mg2+ is the key CaSR activator in mouse and human intestinal epithelia at physiological levels in stool. Since stool Mg2+ concentrations in patients with cholera are essentially zero, oral Mg2+ supplementation, alone or in an oral rehydration solution, could be a potential therapy for cholera and other cyclic nucleotide-mediated secretory diarrheas.

Background Hypothesis/Objectives Animals Methods Results Conclusions and Clinical Importance Gallbladder mucocele (GBM) is a common disease in the canine gallbladder. Although the pathogenesis of GBM remains unclear, we recently reported that the excessive accumulation of mucin in the gallbladder is not a result of overproduction by gallbladder epithelial cells (GBECs).Changes in the function of GBECs other than the production of mucin are associated with the pathogenesis of GBM. We performed an RNA sequencing (RNA‐seq) analysis to comprehensively search for abnormalities in gene expression profiles of GBECs in dogs with GBM.Fifteen dogs with GBM and 8 dogs euthanized for reasons other than gallbladder disease were included.The GBECs were isolated from gallbladder tissues to extract RNA. The RNA‐seq analysis was performed using the samples from 3 GBM cases and 3 dogs with normal gallbladders, and the gene expression profiles were compared between the 2 groups. Differences in mRNA expression levels of the extracted differentially expressed genes (DEGs) were validated by quantitative reverse transcription polymerase chain reaction (RT‐qPCR) using samples of 15 GBM cases and 8 dogs with normal gallbladders.Comparison of gene expression profiles by RNA‐seq extracted 367 DEGs, including ANO1, a chloride channel associated with changes in mucin morphology, and HTR4, which regulates the function of chloride channels. The ANO1 and HTR4 genes were confirmed to be downregulated in the GBM group by RT‐qPCR.Our results suggest that GBM may be associated with decreased function of chloride channels expressed in GBECs. [ABSTRACT FROM AUTHOR]

The four cystic fibrosis (CF) transmembrane conductance regulator (CFTR) modulators, ivacaftor, lumacaftor, tezacaftor, and elexacaftor, have revolutionised the treatment of CF by direct action on the protein target behind the disease's development. The aim was to develop and validate a quantification method for these CFTR modulators in plasma and breast milk to better understand inter-patient variability in pharmacokinetics and treatment outcome, including the risk of adverse drug reactions. The ability to monitor CFTR modulators in breast milk enables the estimation of the exposure of breastfed infant, with a potential concern for CFTR modulator-induced liver injury. The analysis was performed on a Thermo Vanquish Flex Binary UHPLC system coupled to a high-resolution mass spectrometer (HRMS), Thermo Q Exactive. The analytes were detected using positive electrospray ionisation in full scan mode. After sample preparation by protein precipitation, the supernatant was injected onto the LC system and the analytes were separated using a Zorbax SB-C18 Rapid Res HPLC column (3.5 µm, 4.6 × 75 mm). This is the first published method for CFTR modulators in breast milk. The validated quantification range for ivacaftor is 0.0050–10 µg/mL with a coefficient of variation < 6% and a mean accuracy of 97–106%; for lumacaftor, tezacaftor, and elexacaftor, the validated quantification range is 0.050–100 µg/mL with a coefficient of variation < 8% and a mean accuracy 93–106%. A simple and sensitive quantification method for CFTR modulators has been developed and used for routine analysis of human plasma and breast milk samples since 2022. [ABSTRACT FROM AUTHOR]

To maintain internal ion balance in marine environments, teleost fishes leverage seawater (SW)-type ionocytes to actively secrete Na+ and Cl− into the environment. It is well established that SW-type ionocytes use apically expressed cystic fibrosis transmembrane conductance regulator 1 (Cftr1) as a conduit for Cl− to exit the gill. Here, we investigated whether the Ca2+-activated Cl− channel, anoctamin 1 (Ano1), provides an additional path for Cl−-secretion in euryhaline mummichogs (Fundulus heteroclitus). Two ano1 gene isoforms, denoted ano1.1a and -1.1b, exhibited higher expression in the gill and opercular epithelium of mummichogs long-term acclimated to SW versus fresh water (FW). Branchial ano1.1b and cftr1 expression was increased in mummichogs sampled 24 h after transfer from FW to SW; ano1.1a and -1.1b were upregulated in the gill and opercular epithelium following transfer from SW to hypersaline SW. Alternatively, the expression of ano1.1a, -1.1b, and cftr1 in the gill and opercular epithelium was markedly decreased after transfer from SW to FW. Given its role in attenuating ion secretion, we probed whether prolactin downregulates ano1 isoforms. In addition to attenuating cftr1 expression, a prolactin injection reduced branchial ano1.1a and -1.1b levels. Given how Ano1 mediates Cl− secretion by mammalian epithelial cells, the salinity- and prolactin-sensitive nature of ano1 expression reported here indicates that Ano1 may constitute a novel Cl−-secretion pathway in ionocytes. This study encourages a wider evaluation of this putative Cl−-secretion pathway and its regulation by hormones in teleost fishes. NEW & NOTEWORTHY: In this study, we provide evidence in a teleost fish that the Ca2+-activated Cl− channel, anoctamin 1 may provide an additional path for Cl− secretion by seawater-type ionocytes. Not only is this the first report of a Cftr-independent Cl−-secreting pathway conferring survival in seawater but also the first description of its regulation by the pituitary hormone prolactin. [ABSTRACT FROM AUTHOR]

Patients with myotonia congenita suffer from slowed relaxation of muscle (myotonia), due to hyperexcitability caused by loss-of-function mutations in the ClC-1 chloride channel. A recent study suggested that block of large-conductance voltage- and Ca2+- activated K+ channels (BK) may be effective as therapy. The mechanism underlying efficacy was suggested to be lessening of the depolarizing effect of build-up of K+ in t-tubules of muscle during repetitive firing. BK channels are widely expressed in the nervous system and have been shown to play a central role in regulation of excitability, but their contribution to muscle excitability has not been determined. We performed intracellular recordings as well as force measurements in both wild type and BK−/− mouse extensor digitorum longus muscles. Action potential width was increased in BK−/− muscle due to slowing of repolarization, consistent with the possibility K+ build-up in t-tubules is lessened by block of BK channels in myotonic muscle. However, there was no difference in the severity of myotonia triggered by block of muscle Cl− channels with 9-anthracenecarboxylic acid (9AC) in wild type and BK−/− muscle fibers. Further study revealed no difference in the interspike membrane potential during repetitive firing suggesting there was no reduction in K+ build-up in t-tubules of BK−/− muscle. Force recordings following block of muscle Cl− channels demonstrated little reduction in myotonia in BK−/− muscle. In contrast, the current standard of care, mexiletine, significantly reduced myotonia. Our data suggest BK channels regulate muscle excitability, but are not an attractive target for therapy of myotonia. [ABSTRACT FROM AUTHOR]

Aim Methods Results Conclusions Calcium‐activated ligand‐gated chloride channels, beyond their role in maintaining anion homeostasis, modulate neuronal excitability by facilitating nonvesicular neurotransmitter release. BEST1, a key member of this family, is permeable to γ‐aminobutyric acid (GABA) and glutamate. While astrocytic BEST1 is well‐studied and known to regulate neurotransmitter levels, its distribution and role in other brain cell types remain unclear. This study aimed to reassess the localization of BEST1 in the mouse cerebral cortex.We examined the localization and distribution of BEST1 in the mouse parietal cortex using light microscopy, confocal double‐labeling with markers for astrocytes, neurons, microglia, and oligodendrocyte precursor cells, and 3D reconstruction techniques.In the cerebral cortex, BEST1 is more broadly distributed than previously thought. Neurons are the second most abundant BEST1+ cell type in the cerebral cortex, following astrocytes. BEST1 is diffusely expressed in neuronal somatic and neuropilar domains and is present at glutamatergic and GABAergic terminals, with a prevalence at GABAergic terminals. We also confirmed that BEST1 is expressed in cortical microglia and identified it in oligodendrocyte precursor cells, albeit to a lesser extent.Together, these findings suggest that BEST1's role in controlling neurotransmission may extend beyond astrocytes to include other brain cells. Understanding BEST1's function in these cells could offer new insights into the molecular mechanisms shaping cortical circuitry. Further research is needed to clarify the diverse roles of BEST1 in both normal and pathophysiological conditions. [ABSTRACT FROM AUTHOR]

Neuromuscular blocking agents are used commonly to induce skeletal muscle relaxation during surgery. While muscle relaxation facilitates surgical procedures and tracheal intubation, adequate recovery of muscle function after surgery is required to support pulmonary function, and even mild residual neuromuscular block increases the risk of severe postoperative pulmonary complications. While recovery of muscle function after surgery involving neuromuscular blocking agents can be monitored and, in addition, be accelerated by use of current antagonists (reversal agents), there is a clear clinical need for a safe drug to antagonize all types of neuromuscular blocking agents. Here, we show that inhibition of the skeletal muscle-specific chloride ion (Cl-) channel, the ClC-1 channel, markedly accelerates recovery of both single contraction (twitch) and, important physiologically, sustained (tetanic) contractions in a rat model mimicking neuromuscular blocking agent-induced muscle block used during surgery. This suggests ClC-1 inhibition as a mechanism for fast and efficacious recovery of neuromuscular function induced by any neuromuscular blocking agents. Reversal agents are intravenous drugs used to accelerate recovery from neuromuscular block post-surgery. We report that inhibition of ClC-1, a skeletal muscle chloride channel, reverses any nondepolarizing blocking agent, providing a novel reversal (antagonism) mechanism. [ABSTRACT FROM AUTHOR]

Background: CLCN2-related leukoencephalopathy (CC2L) is a rare autosomal recessive disorder caused by biallelic variants of CLCN2, which encodes chloride channel 2. Although CC2L is associated with distinct radiological features, it presents with a wide range of clinical features. Case presentation: A 34-year-old woman presented to our hospital with a sudden onset of vertigo with headache. The patient reported intermittent headaches and tingling in both arms since the age of 31 years. On the first visit, the patient was alert and neurologically intact, except for slight hyperreflexia of the limbs without laterality. Head magnetic resonance imaging (MRI) showed high-intensity signals on axial T2-weighted fluid-attenuated inversion recovery and diffusion-weighted images bilaterally in the posterior limbs of the internal capsules, cerebral peduncles, superior and middle cerebellar peduncles, decussation of superior cerebellar peduncles, and central tegmental tract. All the patient's symptoms were resolved or eased following supportive care. The patient stopped attending our hospital at the age of 46 years. At 51 years of age, the patient revisited our hospital because of the recurrence of vertigo, headache, and nausea. She did not present with any abnormalities by neurological examination. Head MRI showed widespread high-intensity signals similar to those 17 years ago. Genetic testing revealed compound heterozygous variants in CLCN2 (NM_004366.6): a novel variant c.1828 C > T, p.(Arg 610*) from her father and c.61dup, p.(Leu21Profs*27) from her mother. The patient was finally diagnosed with CC2L. She received supportive treatment, which made her symptoms manageable. Conclusions: This is a detailed report of a patient with adult-onset CC2L who was successfully diagnosed and followed up with head MRI. This report provides new insights into CC2L and highlights its persistent, distinct, and stable characteristics observed in head MRI over one decade and the difficulty in forming a diagnosis without MRI when patients have minimal and common symptoms, such as in the present case. [ABSTRACT FROM AUTHOR]

In this work, we cloned and functionally expressed two novel GABAA receptor subunits from Procambarus clarkii crayfish. These two new subunits, PcGABAA‐α and PcGABAA‐β2, revealed significant sequence homology with the PcGABAA‐β subunit, previously identified in our laboratory. In addition, PcGABAA‐α subunit also shared a significant degree of identity with the Drosophila melanogaster genes DmGRD (GABA and glycine‐like receptor subunits of Drosophila) as well as PcGABAA‐β2 subunit with DmLCCH3 (ligand‐gated chloride channel homolog 3). Electrophysiological recordings showed that the expression in HEK cells of the novel subunits, either alone or in combination, failed to form functional homo‐ or heteromeric receptors. However, the co‐expression of PcGABAA‐α with PcGABAA‐β evoked sodium‐ or chloride‐dependent currents that accurately reproduced the time course of the GABA‐evoked currents in the X‐organ neurons from crayfish, suggesting that these GABA subunits combine to form two types of GABA receptors, one with cationic selectivity filter and the other preferentially permeates anions. On the other hand, PcGABAA‐β2 and PcGABAA‐β co‐expression generated a chloride current that does not show desensitization. Muscimol reproduced the time course of GABA‐evoked currents in all functional receptors, and picrotoxin blocked these currents; bicuculline did not block any of the recorded currents. Reverse transcription polymerae chain reaction (RT‐PCR) amplifications and FISH revealed that PcGABAA‐α and PcGABAA‐β2 are predominantly expressed in the crayfish nervous system. Altogether, these findings provide the first evidence of a neural GABA‐gated cationic channel in the crayfish, increasing our understanding of the role of these new GABAA receptor subunits in native heteromeric receptors. [ABSTRACT FROM AUTHOR]

In cystic fibrosis (CF), there is abnormal translocation and function of the cystic fibrosis transmembrane conductance regulator (CFTR) and an upregulation of the epithelial sodium channel (ENaC). This leads to hyperabsorption of sodium and fluid from the airway, dehydrated mucus, and an increased risk of respiratory infections. In this study, we performed a proteomic assessment of differentially regulated proteins from CF and non-CF small airway epithelial cells (SAEC) that are sensitive to Mycobacterium avium. CF SAEC and normal non-CF SAEC were infected with M. avium before the cells were harvested for protein. Protein kinase C (PKC) activity was greater in the CF cells compared to the non-CF cells, but the activity was significantly attenuated in both cell types after infection with M. avium compared to vehicle. Western blot and densitometric analysis showed a significant increase in cathepsin B protein expression in M. avium infected CF cells. Myristoylated alanine rich C-kinase substrate (MARCKS) protein was one of several differentially expressed proteins between the groups that was identified by mass spectrometry-based proteomics. Total MARCKS protein expression was greater in CF cells compared to non-CF cells. Phosphorylation of MARCKS at serine 163 was also greater in CF cells compared to non-CF cells after treating both groups of cells with M. avium. Taken together, MARCKS protein is upregulated in CF cells and there is decreased phosphorylation of the protein due to a decrease in PKC activity and presumably increased cathepsin B mediated proteolysis of the protein after M. avium infection. [ABSTRACT FROM AUTHOR]

Chronic low back pain (LBP) can severely affect daily physical activity. Aberrant osteoclast-mediated resorption leads to porous endplates, which allow the sensory innervation that causes LBP. Here, we report that expression of the proton-activated chloride (PAC) channel was induced during osteoclast differentiation in the porous endplates via a RANKL/ NFATc1 signaling pathway. Extracellular acidosis evoked robust PAC currents in osteoclasts. An acidic environment of porous endplates and elevated PAC activation-enhanced osteoclast fusion provoked LBP. Furthermore, we found that genetic knockout of the PAC gene Pacc1 significantly reduced endplate porosity and spinal pain in a mouse LBP model, but it did not affect bone development or homeostasis of bone mass in adult mice. Moreover, both the osteoclast bone-resorptive compartment environment and PAC traffic from the plasma membrane to endosomes to form an intracellular organelle Cl channel had a low pH of approximately 5.0. The low pH environment activated the PAC channel to increase sialyltransferase St3gal1 expression and sialylation of TLR2 in the initiation of osteoclast fusion. Aberrant osteoclast-mediated resorption is also found in most skeletal disorders, including osteoarthritis, ankylosing spondylitis, rheumatoid arthritis, heterotopic ossification, and enthesopathy. Thus, elevated Pacc1 expression and PAC activity could be a potential therapeutic target for the treatment of LBP and osteoclast-associated pain. [ABSTRACT FROM AUTHOR]

Myoblast differentiation plays a vital role in skeletal muscle regeneration. However, the protein-coding genes controlling this process remain incompletely understood. Here, we showed that chloride intracellular channel 5 (CLIC5) exerts a critical role in mediating myogenesis and skeletal muscle regeneration. Deletion of CLIC5 in skeletal muscle leads to reduced muscle weight and decreases the number and differentiation potential of satellite cells. In vitro, CLIC5 consistently inhibits myoblast proliferation while promoting myotube formation. CLIC5 promotes myogenic differentiation by activating the canonical Wnt/ß-catenin signaling pathway in a biglycan (BGN)-dependent manner. CLIC5 deletion impairs muscle regeneration. Paired box gene 7 (Pax7) expression and the activity of BGN-mediated canonical Wnt/ß-catenin signaling are reduced in CLIC5-deficient mice. Conversely, increasing CLIC5 levels in skeletal muscles enhances muscle regeneration capacity. In conclusion, our findings underscore CLIC5 as a pivotal regulator of myogenesis and skeletal muscle regeneration, functioning through interaction with BGN to activate the canonical Wnt/ß-catenin signaling pathway. [ABSTRACT FROM AUTHOR]

Background: miR-210 is one of the most evolutionarily conserved microRNAs. It is known to be involved in several physiological and pathological processes, including response to hypoxia, angiogenesis, cardiovascular diseases and cancer. Recently, new roles of this microRNA are emerging in the context of eye and visual system homeostasis. Recent studies in Drosophila melanogaster unveiled that the absence of miR-210 leads to a progressive retinal degeneration characterized by the accumulation of lipid droplets and disruptions in lipid metabolism. However, the possible conservation of miR-210 knock-out effect in the mammalian retina has yet to be explored. Results: We further investigated lipid anabolism and catabolism in miR-210 knock-out (KO) flies, uncovering significant alterations in gene expression within these pathways. Additionally, we characterized the retinal morphology of flies overexpressing (OE) miR-210, which was not affected by the increased levels of the microRNA. For the first time, we also characterized the retinal morphology of miR-210 KO and OE mice. Similar to flies, miR-210 OE did not affect retinal homeostasis, whereas miR-210 KO mice exhibited photoreceptor degeneration. To explore other potential parallels between miR-210 KO models in flies and mice, we examined lipid metabolism, circadian behaviour, and retinal transcriptome in mice, but found no similarities. Specifically, RNA-seq confirmed the lack of involvement of lipid metabolism in the mice's pathological phenotype, revealing that the differentially expressed genes were predominantly associated with chloride channel activity and extracellular matrix homeostasis. Simultaneously, transcriptome analysis of miR-210 KO fly brains indicated that the observed alterations extend beyond the eye and may be linked to neuronal deficiencies in signal detection and transduction. Conclusions: We provide the first morphological characterization of the retina of miR-210 KO and OE mice, investigating the role of this microRNA in mammalian retinal physiology and exploring potential parallels with phenotypes observed in fly models. Although the lack of similarities in lipid metabolism, circadian behaviour, and retinal transcriptome in mice suggests divergent mechanisms of retinal degeneration between the two species, transcriptome analysis of miR-210 KO fly brains indicates the potential existence of a shared upstream mechanism contributing to retinal degeneration in both flies and mammals. [ABSTRACT FROM AUTHOR]

Although salt is an essential substance vital to life, excessive salt intake could cause various health issues. Therefore, new technologies and strategies should be developed to reduce salt intake without compromising taste. However, the underlying physiological mechanisms of salt taste reception is complex and not completely understood. Sodium chloride is a typical salty substance. It is widely believed that only sodium is important for the generation of salty taste. On the other hand, from a psychophysical perspective, the importance of chloride in salty taste has been indicated. Thus, understanding the mechanisms of both sodiumand chloride-tastes generation is necessary to completely comprehended the fundamentals of salt taste reception. However, the mechanism for detecting chloride taste has remained unclear for many years. Recently, we have identified transmembrane channel-like 4 (TMC4) as the first molecule that mediates the reception of chloride taste. TMC4 functions as a voltage-dependent chloride channel and plays an important role in the reception of the chloride taste by detecting chloride ions. In this mini-review, we first introduce the known reception mechanism of salty taste, and then discuss the roles of TMC4 in the salt taste reception. The finding of TMC4 may serve as a basis for developing new technologies and formulating strategies to reduce salt intake without compromising taste. [ABSTRACT FROM AUTHOR]

The major female ovarian hormone, 17ß-estradiol (E2), can alter neuronal excitability within milliseconds to regulate a variety of physiological processes. Estrogen receptor-a (ERa), classically known as a nuclear receptor, exists as a membrane-bound receptor to mediate this rapid action of E2, but the ionic mechanisms remain unclear. Here, we show that a membrane channel protein, chloride intracellular channel protein-1 (Clic1), can physically interact with ERa with a preference to the membrane-bound ERa. Clic1-mediated currents can be enhanced by E2 and reduced by its depletion. In addition, Clic1 currents are required to mediate the E2-induced rapid excitations in multiple brain ERa populations. Further, genetic disruption of Clic1 in hypothalamic ERa neurons blunts the regulations of E2 on female body weight balance. In conclusion, we identified the Clic1 chloride channel as a key mediator for E2-induced rapid neuronal excitation, which may have a broad impact on multiple neurobiological processes regulated by E2. [ABSTRACT FROM AUTHOR]

AIM: To investigate the activation of Cl- channels by sodium taurocholate （NaTC） in mouse pancreatic acinar cells. METHODS: The single isolated pancreatic acinar cells from FVB/N mice were prepared using collagenase digestion method. Whole-cell patch clamp technique was performed to record the currents. Intracellular adenosine triphosphate （ATP） dependence of the channels was examined via eliminating ATP from the pipette solution. Anion permeability of the channels was investigated with ion-exchange method. The pharmacological characteristics of the channels was confirmed by two Cl- channel blockers. The volume sensitivity of the channels was detected using 47% hypertonic bathing solution. Extracellular Ca2+ dependence of activating the channels was examined through eliminating Ca2+ from the bathing solution. Intracellular reactive oxygen species （ROS） level was detected by an oxidation-sensitive fluorescent probe, 2', 7'-dichlorofluorescin diacetate. The experiment was repeated 6 times in each group. RESULTS: Extracellular application of 5 mmol/L sodium taurocholate induced a Cl- current, exhibiting the properties of outward-rectification, a selectivity sequence of I- > Br- ≥ Cl- > gluconate- and intracellularATP dependence（ P<0. 01）. The currents were inhibited by chloride channel blocker 4, 4'-diisothiocyanatostilbene-2, 2'-disulfonic acid disodium salt hydrate （DIDS） and tamoxifen and by 47% hypertonicity stimulation （P<0. 01）. When ROS production was scavenged by N-acetyl-L-cysteine, the sodium taurocholate-induced Cl- currents were unaffected. The effect of sodium taurocholate on ROS production did not alter with the treatment with DIDS. Sodium taurocholate failed to induce Cl- currents when Ca2+ was absent in extracellular bathing solution （P>0. 05）. CONCLUSION: Sodium taurocholate activates Cl- channels in mouse pancreatic acinar cells, which is dependent on extracellular Ca2+ but not ROS pathway. [ABSTRACT FROM AUTHOR]

Soil salinization has become a global problem and high salt concentration in soil negatively affects plant growth. In our previous study, we found that overexpression of PsAMT1.2 from Populus simonii could improve the salt tolerance of poplar, but the physiological and molecular mechanism was not well understood. To explore the regulation pathway of PsAMT1.2 in salt tolerance, we investigated the morphological, physiological and transcriptome differences between the PsAMT1.2 overexpression transgenic poplar and the wild type under salt stress. The PsAMT1.2 overexpression transgenic poplar showed better growth with increased net photosynthetic rate and higher chlorophyll content compared with wild type under salt stress. The overexpression of PsAMT1.2 increased the catalase, superoxide dismutase, peroxidase and ascorbate peroxidase activities, and therefore probably enhanced the reactive oxygen species clearance ability, which also reduced the degree of membrane lipid peroxidation under salt stress. Meanwhile, the PsAMT1.2 overexpression transgenic poplar maintained a relatively high K+/Na+ ratio under salt stress. RNA-seq analysis indicated that PsAMT1.2 might improve plant salt tolerance by regulating pathways related to the photosynthetic system, chloroplast structure, antioxidant activity and anion transport. Among the 1056 differentially expressed genes, genes related to photosystem I and photosystem II were up-regulated and genes related to chloride channel protein-related were down-regulated. The result of the present study would provide new insight into regulation mechanism of PsAMT1.2 in improving salt tolerance of poplar. [ABSTRACT FROM AUTHOR]

Alterations in anion balance potential, along with the involvement of cation-chloride cotransporters, play pivotal roles in the development of hyperalgesia after peripheral nerve injury. Chloride voltage-gated channel seven (CLCN7) is the predominant member of the CLC protein family. Investigations on CLCN7 have focused primarily on its involvement in osteosclerosis and lysosomal storage disorders; nevertheless, its contribution to neuropathic pain has not been determined. In this investigation, we noted high expression of CLCN7 in neurons situated within the spinal dorsal horns and dorsal root ganglions (DRGs). Immunofluorescence analysis revealed that CLCN7 was predominantly distributed among IB4-positive and CGRP-positive neurons. Furthermore, the expression of CLCN7 was observed to be mainly reduced in neurons within the spinal dorsal horns and in small- and medium-sized neurons located in the DRGs of spared nerve injury mice. Knockdown of CLCN7 via siRNA in the DRGs resulted in increased mechanical and thermal hyperalgesia in naïve mice. Furthermore, the excitability of cultured DRG neurons in vitro was augmented upon treatment with CLCN7 siRNA. These findings suggested that CLCN7 downregulation following SNI was crucial for the manifestation of mechanical and thermal hyperalgesia, highlighting potential targeting strategies for treating neuropathic pain. [ABSTRACT FROM AUTHOR]

Background: Prokinetic agents are effective in increasing gastrointestinal (GI) contractility and alleviating constipation, often caused by slow intestinal motility. Lubiprostone (LUB), known for activating CLC-2 chloride channels, increases the chloride ion concentration in the GI tract, supporting water retention and stool movement. Despite its therapeutic efficacy, the exact mechanisms underlying its pharmacological action are poorly understood. Here, we investigated whether LUB activates the canonical transient receptor potential cation channel type 4 (TRPC4) through stimulation with E-type prostaglandin receptor (EP) type 3. Methods: Using isotonic tension recordings on mouse colon strips, we examined LUB-induced contractility in both proximal and distal colon segments. Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to determine mRNA levels of EP1-4 receptor subtypes in distal colonic muscular strips and isolated myocytes. The effects of a TRPC4 blocker and EP3 antagonist on LUB-stimulated contractions were also evaluated. Results: LUB showed significant contraction in the distal segment compared to the proximal segment. EP3 receptor mRNA levels were highly expressed in the distal colon tissue, which correlated with the observed enhanced contraction. Furthermore, LUB-induced spontaneous contractions in distal colon muscles were reduced by a TRPC4 blocker or EP3 antagonist, indicating that LUB-stimulated EP3 receptor activation may lead to TRPC4 activation and increased intracellular calcium in colonic smooth muscle. Conclusions: These findings suggest that LUB improves mass movement through indirect activation of the TRPC4 channel in the distal colon. The segment-specific action of prokinetic agents like LUB provides compelling evidence for a personalized approach to symptom management, supporting the defecation reflex. [ABSTRACT FROM AUTHOR]

The epsilon toxin (Etx) from Clostridium perfringens has been identified as a potential trigger of multiple sclerosis, functioning as a pore‐forming toxin that selectively targets cells expressing the plasma membrane (PM) myelin and lymphocyte protein (MAL). Previously, we observed that Etx induces the release of intracellular ATP in sensitive cell lines. Here, we aimed to re‐examine the mechanism of action of the toxin and investigate the connection between pore formation and ATP release. We examined the impact of Etx on Xenopus laevis oocytes expressing human MAL. Extracellular ATP was assessed using the luciferin‐luciferase reaction. Activation of calcium‐activated chloride channels (CaCCs) and a decrease in the PM surface were recorded using the two‐electrode voltage‐clamp technique. To evaluate intracellular Ca2+ levels and scramblase activity, fluorescent dyes were employed. Extracellular vesicles were imaged using light and electron microscopy, while toxin oligomers were identified through western blots. Etx triggered intracellular Ca2+ mobilization in the Xenopus oocytes expressing hMAL, leading to the activation of CaCCs, ATP release, and a reduction in PM capacitance. The toxin induced the activation of scramblase and, thus, translocated phospholipids from the inner to the outer leaflet of the PM, exposing phosphatidylserine outside in Xenopus oocytes and in an Etx‐sensitive cell line. Moreover, Etx caused the formation of extracellular vesicles, not derived from apoptotic bodies, through PM fission. These vesicles carried toxin heptamers and doughnut‐like structures in the nanometer size range. In conclusion, ATP release was not directly attributed to the formation of pores in the PM, but to scramblase activity and the formation of extracellular vesicles. [ABSTRACT FROM AUTHOR]

SO2/H2SO3 can damage plants. However, its toxic mechanism has still been controversial. Two models have been proposed, cytosolic acidification model and cellular oxidation model. Here, we assessed the toxic mechanism of H2SO3 in three cell types of Arabidopsis thaliana , mesophyll cells, guard cells (GCs), and petal cells. The sensitivity of GCs of Chloride channel a (CLC a)-knockout mutants to H2SO3 was significantly lower than those of wildtype plants. Expression of other CLC genes in mesophyll cells and petal cells were different from GCs. Treatment with antioxidant, disodium 4,5-dihydroxy-1,3-benzenedisulfonate (tiron), increased the median lethal concentration (LC50) of H2SO3 in GCs indicating the involvement of cellular oxidation, while the effect was negligible in mesophyll cells and petal cells. These results indicate that there are two toxic mechanisms of SO2 to Arabidopsis cells: cytosolic acidification and cellular oxidation, and the toxic mechanism may vary among cell types. [ABSTRACT FROM AUTHOR]

Cystic fibrosis (CF) is a genetic disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, resulting in defective chloride ion channels. This leads to thick, dehydrated mucus that severely disrupts mucociliary clearance in the respiratory system and triggers infection that eventually is the cause of death of CF patients. Current therapeutic strategies primarily focus on restoring CFTR function, blocking epithelial sodium channels to prevent mucus dehydration, or directly targeting mucus to reduce its viscosity. Among the ion channels expressed in ciliated bronchial epithelial cells, the transient receptor potential vanilloid 4 (TRPV4) channel emerges as a significant channel in CF pathogenesis. Activation of TRPV4 channels affects the regulation of airway surface liquid by modulating sodium absorption and intracellular calcium levels, which indirectly influences CFTR activity. TRPV4 is also involved in the regulatory volume decrease (RVD) process and enhances inflammatory responses in CF patients. Here, we combine current findings on TRPV4 channel modulation as a promising therapeutic approach for CF. Although limited studies have directly explored TRPV4 in CF, emerging evidence indicates that TRPV4 activation can significantly impact key pathological processes in the disease. Further investigation into TRPV4 modulators could lead to innovative treatments that alleviate severe respiratory complications and improve outcomes for CF patients. [ABSTRACT FROM AUTHOR]

Complex alleles of the CFTR gene complicate the diagnosis of cystic fibrosis (CF), the classification of its pathogenic variants, affect the clinical picture of the disease and can affect the efficiency of targeted drugs. The total frequency of complex allele [L467F;F508del] in the Russian population of patients with CF is 0.74%, and in patients with the F508del/F508del genotype, its frequency reaches 8%. This article presents multi-faceted study of the complex allele [L467F;F508del] in a cohort of patients with genotypes [L467F;F508del]/class I (c.3532_3535dup, c.1766+2T>C, W1310X, 712-1G>T), and data for a unique patient with the genotype [L467F;F508del]/[L467F;F508del]. Using the intestinal current measurement method, it was demonstrated the absence of CFTR function for [L467F;F508del]/class I and [L467F;F508del]/[L467F;F508del] genotypes. In intestinal organoids, it was shown that [L467F;F508del] in combination with class I variants and in the homozygotes abolishes the efficacy of both two-component (ivacaftor+lumacaftor; ivacaftor+tezacaftor) and three-component (ivacaftor+tezacaftor+elexacaftor) targeted drugs. When prescribing ivacaftor+tezacaftor+elexacaftor to three patients, they did not have a clinical effect after 6–12 months. [ABSTRACT FROM AUTHOR]

The triple combination therapy for cystic fibrosis (CF), including elexacaftor, tezacaftor and ivacaftor (ETI or Trikafta), has been shown to improve lung function and reduce pulmonary exacerbations, thereby enhancing the quality of life for most CF patients. Recent findings suggest that both the individual components and ETI may have potential off-target effects, highlighting the need to understand how these modulators impact cellular physiology, particularly in cells that do not express CF transmembrane conductance regulator (CFTR). We used HEK293 cells, as a cell model not expressing the CFTR protein, to evaluate the effect of ETI and each of its components on autophagic machinery and on the Rab5/7 components of the Rab pathway. We firstly demonstrate that the single modulators Teza and Iva, and the combinations ET and ETI, increased ROS production in the absence of their target while decreasing it in cells expressing the CFTR ∆F508del. This increase in cellular stress was followed by an increase in the total level of polyubiquitinated proteins as well as the p62 level and LC3II/LC3I ratio. Furthermore, we found that ETI had the opposite effect on Rabs by increasing Rab5 levels while decreasing Rab7. Interestingly, these changes were abolished by the expression of mutated CFTR. Overall, our data suggest that in the absence of their target, both the individual modulators and ETI increased ROS production and halted both autophagic flux and plasma membrane protein recycling. [ABSTRACT FROM AUTHOR]

It has long been an aspirational goal to create artificial channel structures that replicate the feat achieved by ion channel proteins. Biological ion channels occasionally demonstrate multiple conductance states (known as subconductance), remaining a challenging property to achieve in artificial channel molecules. We report a funnel‐shaped single‐molecule channel constructed by an electron‐deficient macrocycle and two electron‐deficient aromatic imide arms. Planar lipid bilayer measurements reveal distinct current recordings, including a closed state, two conducting states, and spontaneous transitions between the three states, resembling the events seen in biological ion channels. The transitions result from conformational changes induced by chloride transport in the channel molecule. Both opening states show a non‐linear and rectifying I–V relationship, indicating voltage‐dependent transport due to the asymmetrical channel structure. This work could enhance our understanding of ion permeation and channel opening mechanism. [ABSTRACT FROM AUTHOR]

Natural scenes are highly dynamic, challenging the reliability of visual processing. Yet, humans and many animals perform accurate visual behaviors, whereas computer vision devices struggle with rapidly changing background luminance. How does animal vision achieve this? Here, we reveal the algorithms and mechanisms of rapid luminance gain control in Drosophila, resulting in stable visual processing. We identify specific transmedullary neurons as the site of luminance gain control, which pass this property to direction-selective cells. The circuitry further involves wide-field neurons, matching computational predictions that local spatial pooling drive optimal contrast processing in natural scenes when light conditions change rapidly. Experiments and theory argue that a spatially pooled luminance signal achieves luminance gain control via divisive normalization. This process relies on shunting inhibition using the glutamate-gated chloride channel GluClα. Our work describes how the fly robustly processes visual information in dynamically changing natural scenes, a common challenge of all visual systems. Stable vision in quickly changing environments requires a mechanism that ensures that contrast is computed correctly when light conditions change rapidly. Here authors identify the circuits and algorithm of this mechanism in the fruit fly Drosophila. [ABSTRACT FROM AUTHOR]

It has been documented that caspase 3 activity is necessary for skeletal muscle regeneration, but how its activity is regulated is largely unknown. Our previous report shows that intracellular TMEM16A, a calcium activated chloride channel, significantly regulates caspase 3 activity in myoblasts during skeletal muscle development. By using a mouse line with satellite cell (SC)-specific deletion of TMEM16A, we examined the role of TMEM16A in regulating caspase 3 activity in SC (or SC-derived myoblast) as well as skeletal muscle regeneration. The mutant animals displayed apparently impaired regeneration capacity in adult muscle along with enhanced ER stress and elevated caspase 3 activity in Tmem16a−/− SC derived myoblasts. Blockade of either excessive ER stress or caspase 3 activity by small molecules significantly restored the inhibited myogenic differentiation of Tmem16a−/− SCs, indicating that excessive caspase 3 activity resulted from TMEM16A deletion contributes to the impaired muscle regeneration and the upstream regulator of caspase 3 was ER stress. Our results revealed an essential role of TMEM16A in satellite cell-mediated skeletal muscle regeneration by ensuring a moderate level of caspase 3 activity. [ABSTRACT FROM AUTHOR]

Populations of various economic species of wireworms are increasing in the key cereal crop production areas of Canada and the United States. To address this problem, seed treatments are under development that both provide crop protection and significantly reduce populations equivalent in effectiveness to the formerly used but now deregistered organochlorine lindane. Herein, we evaluated isocycloseram (PLINAZOLIN technology), the first isoxazoline (GABA-gated Chloride Channel Allosteric Modulator) agricultural insecticide, as a seed treatment for the protection of cereal crops from the sugarbeet wireworm, Limonius californicus (Mannerheim). In wheat and barley field trials conducted over 4 years under extreme wireworm pressure, isocycloseram applied as a seed treatment at 5.0–7.5 g AI/100 kg seed was as effective as or more effective than the current industry standard thiamethoxam at 20.0 g AI/100 kg seed in protecting crop stand and yield. Isocycloseram also reduced neonate wireworms (produced from eggs during the growing season) and resident wireworms (in the field at the time of planting) to levels expected from the formerly used seed treatment lindane. [ABSTRACT FROM AUTHOR]

This document summarizes a study on the effects of CFTR modulators on patients with cystic fibrosis (CF) and leukocyte adhesion deficiency type IV (LAD-IV). The study found that CFTR modulators improved certain cellular functions in CF patients, suggesting that they may correct both CF and LAD-IV phenotypes. However, there were some patients who showed improvement in CF symptoms but not in LAD-IV characteristics, indicating the need for further research. The study also suggests that measuring certain cellular markers could be a useful method for monitoring patients in the long term. [Extracted from the article]

Background: Cystic Fibrosis (CF) is a monogenic disease caused by mutations in the gene coding the Cystic Fibrosis Transmembrane Regulator (CFTR) protein, but its overall physio-pathology cannot be solely explained by the loss of the CFTR chloride channel function. Indeed, CFTR belongs to a yet not fully deciphered network of proteins participating in various signalling pathways. Methods: We propose a systems biology approach to study how the absence of the CFTR protein at the membrane leads to perturbation of these pathways, resulting in a panel of deleterious CF cellular phenotypes. Results: Based on publicly available transcriptomic datasets, we built and analyzed a CF network that recapitulates signalling dysregulations. The CF network topology and its resulting phenotypes were found to be consistent with CF pathology. Conclusion: Analysis of the network topology highlighted a few proteins that may initiate the propagation of dysregulations, those that trigger CF cellular phenotypes, and suggested several candidate therapeutic targets. Although our research is focused on CF, the global approach proposed in the present paper could also be followed to study other rare monogenic diseases. [ABSTRACT FROM AUTHOR]

The cystic fibrosis transmembrane conductance regulator (CFTR) is a cyclic adenosine monophosphate (cAMP)‐regulated chloride and bicarbonate ion channel found in many human cells. Its unique biochemical characteristics and role as a member of the adenosine triphosphate (ATP)‐binding cassette transporters superfamily are pivotal for the transport of several substrates across cellular membranes. CFTR is known to interact, physically and functionally, with several other cellular proteins. Hence, its properties are essential for moving various substances across cell membranes and ensuring correct cell functioning. Genetic mutations or environmental factors may disrupt CFTR's function resulting in different possible phenotypes due to gene variations that affect not only CFTR's function, localization, and processing within cells, but also those of its interactors. This has been reported as an underlying cause of various diseases, including cystic fibrosis. The severe clinical implications of cystic fibrosis have driven intense research into the role of CFTR in lung function but its significance to fertility, particularly in men, has been comparatively understudied. However, ongoing and more recent research into CFTR and its interacting proteins in the testis or specific testicular cells is beginning to shed light on this field. Herein, we provide a comprehensive and up‐to‐date overview of the CFTR, its interactome, and its crucial role in male reproduction, highlighting recent discoveries and advancements in understanding the molecular mechanisms involved. The comprehension of these complex interactions may pave the way for potential therapeutic approaches to improve fertility of men suffering from alterations in the function of CFTR. [ABSTRACT FROM AUTHOR]

Background Pseudomonas aeruginosa is a multidrug-resistant pathogen causing recalcitrant pulmonary infections in people with cystic fibrosis (pwCF). Cystic fibrosis transmembrane conductance regulator (CFTR) modulators have been developed that partially correct the defective chloride channel driving disease. Despite the many clinical benefits, studies in adults have demonstrated that while P. aeruginosa sputum load decreases, chronic infection persists. Here, we investigate how P. aeruginosa in pwCF may change in the altered lung environment after CFTR modulation. Methods P. aeruginosa strains (n = 105) were isolated from the sputum of 11 chronically colonized pwCF at baseline and up to 21 months posttreatment with elexacaftor-tezacaftor-ivacaftor or tezacaftor-ivacaftor. Phenotypic characterization and comparative genomics were performed. Results Clonal lineages of P. aeruginosa persisted after therapy, with no evidence of displacement by alternative strains. We identified commonly mutated genes among patient isolates that may be positively selected for in the CFTR-modulated lung. However, classic chronic P. aeruginosa phenotypes such as mucoid morphology were sustained, and isolates remained just as resistant to clinically relevant antibiotics. Conclusions Despite the clinical benefits of CFTR modulators, clonal lineages of P. aeruginosa persist that may prove just as difficult to manage in the future, especially in pwCF with advanced lung disease. [ABSTRACT FROM AUTHOR]

Cystic fibrosis transmembrane conductance regulator (CFTR) protein is an ion channel found in numerous epithelia and controls the flow of water and salt across the epithelium. The aim of our study to find natural compounds that can improve lung function for people with cystic fibrosis (CF) caused by the p.Gly628Arg (rs397508316) mutation of CFTR protein. The sequence of CFTR protein as a target structure was retrieved from UniProt and PDB database. The ligands that included Armepavine, Osthole, Curcumin, Plumbagine, Quercetin, and one Trikafta (R*) reference drug were screened out from PubChem database. Autodock vina software carried out docking, and binding energies between the drug and the target were included using docking-score. The following tools examined binding energy, interaction, stability, toxicity, and visualize protein-ligand complexes. The compounds having binding energies of −6.4, −5.1, −6.6, −5.1, and − 6.5 kcal/mol for Armepavine, Osthole, Curcumin, Plumbagine, Quercetin, and R*-drug, respectively with mutated CFTR (Gly628Arg) structure were chosen as the most promising ligands. The ligands bind to the mutated CFTR protein structure active sites in hydrophobic bonds, hydrogen bonds, and electrostatic interactions. According to ADMET analyses, the ligands Armepavine and Quercetin also displayed good pharmacokinetic and toxicity characteristics. An MD simulation for 200 ns was also established to ensure that Armepavine and Quercetin ligands attached to the target protein favorably and dynamically, and that protein-ligand complex stability was maintained. It is concluded that Armepavine and Quercetin have stronger capacity to inhibit the effect of mutated CFTR protein through improved trafficking and restoration of original function. [ABSTRACT FROM AUTHOR]

A-to-I RNA editing is a cellular mechanism that generates transcriptomic and proteomic diversity, which is essential for neuronal and immune functions. It involves the conversion of specific adenosines in RNA molecules to inosines, which are recognized as guanosines by cellular machinery. Despite the vast number of editing sites observed across the animal kingdom, pinpointing critical sites and understanding their in vivo functions remains challenging. Here, we study the function of an evolutionary conserved editing site in Drosophila, located in glutamate-gated chloride channel (GluCla). Our findings reveal that flies lacking editing at this site exhibit reduced olfactory responses to odors and impaired pheromone-dependent social interactions. Moreover, we demonstrate that editing of this site is crucial for the proper processing of olfactory information in projection neurons. Our results highlight the value of using evolutionary conservation as a criterion for identifying editing events with potential functional significance and paves the way for elucidating the intricate link between RNA modification, neuronal physiology, and behavior. [ABSTRACT FROM AUTHOR]

Secretory proteins, including plasma membrane proteins, are generally known to be transported to the plasma membrane through the endoplasmic reticulum- to-Golgi pathway. However, recent studies have revealed that several plasma membrane proteins and cytosolic proteins lacking a signal peptide are released via an unconventional protein secretion (UcPS) route, bypassing the Golgi during their journey to the cell surface. For instance, transmembrane proteins such as the misfolded cystic fibrosis transmembrane conductance regulator (CFTR) protein and the Spike protein of coronaviruses have been observed to reach the cell surface through a UcPS pathway under cell stress conditions. Nevertheless, the precise mechanisms of the UcPS pathway, particularly the molecular machineries involving cytosolic motor proteins, remain largely unknown. In this study, we identified specific kinesins, namely KIF1A and KIF5A, along with cytoplasmic dynein, as critical players in the unconventional trafficking of CFTR and the SARS-CoV-2 Spike protein. Gene silencing results demonstrated that knockdown of KIF1A, KIF5A, and the KIF-associated adaptor protein SKIP, FYCO1 significantly reduced the UcPS of ΔF508-CFTR. Moreover, gene silencing of these motor proteins impeded the UcPS of the SARS-CoV-2 Spike protein. However, the same gene silencing did not affect the conventional Golgimediated cell surface trafficking of wild-type CFTR and Spike protein. These findings suggest that specific motor proteins, distinct from those involved in conventional trafficking, are implicated in the stress-induced UcPS of transmembrane proteins. [ABSTRACT FROM AUTHOR]

Scorpions, an ancient group of venomous invertebrates, have existed for over 430 million years. Their toxins, important for predation and defense, exhibit a variety of biological and pharmacological activities. Research on scorpion toxins has spanned decades. Notably, the toxin genes of Mesobuthus martensii (Scorpiones: Buthidae), a well-known Chinese herbal medicine, have been described at genomic and proteomic levels. However, previous studies primarily focused on the toxin genes expressed in the venom glands, overlooking their expression in multiple tissues. This study analyzed transcriptomes from 14 tissues of M. martensii. Gene annotation revealed 83 toxin and toxin-like genes, including those affecting sodium, potassium, calcium, and chloride ion channels. Approximately 70% of toxin genes were highly expressed in the vesicle; additionally, some exhibited low or no expression in the vesicle while showing high expression in other tissues. Beyond the vesicle, high expression levels of toxin genes were observed in metasoma segments II-V, blood, lateral eyes, chelicerae, legs, pedipalp chelae, femurs, and patellae. This expression pattern suggests that toxin genes are recruited from multiple tissues and may help prevent intraspecific harm during courtship and competition for prey. These findings inspire further research into the evolutionary recruitment process of scorpion toxins. [ABSTRACT FROM AUTHOR]

Chloride channels (ClCs) have received global interest due to their significant role in the regulation of ion homeostasis, fluid transport, and electrical excitability of tissues and organs in different mammals and contributing to various functions, such as neuronal signaling, muscle contraction, and regulating the electrolytes' balance in kidneys and other organs. In order to define the chloride voltage-gated channel (CLCN) gene family in buffalo, this study used in silico analyses to examine physicochemical properties, evolutionary patterns, and genome-wide identification. We identified eight CLCN genes in buffalo. The ProtParam tool analysis identified a number of important physicochemical properties of these proteins, including hydrophilicity, thermostability, in vitro instability, and basic nature. Based on their evolutionary relationships, a phylogenetic analysis divided the eight discovered genes into three subfamilies. Furthermore, a gene structure analysis, motif patterns, and conserved domains using TBtool demonstrated the significant conservation of this gene family among selected species over the course of evolution. A comparative amino acid analysis using ClustalW revealed similarities and differences between buffalo and cattle CLCN proteins. Three duplicated gene pairs were identified, all of which were segmental duplications except for CLCN4-CLCN5, which was a tandem duplication in buffalo. For each gene pair, the Ka/Ks test ratio findings showed that none of the ratios was more than one, indicating that these proteins were likely subject to positive selection. A synteny analysis confirmed a conserved pattern of genomic blocks between buffalo and cattle. Transcriptional control in cells relies on the binding of transcription factors to specific sites in the genome. The number of transcription factor binding sites (TFBSs) was higher in cattle compared to buffalo. Five main recombination breakpoints were identified at various places in the recombination analysis. The outcomes of our study provide new knowledge about the CLCN gene family in buffalo and open the door for further research on candidate genes in vertebrates through genome-wide studies. [ABSTRACT FROM AUTHOR]

A thin film of pulmonary surfactant lines the surface of the airways and alveoli, where it lowers the surface tension in the peripheral lungs, preventing collapse of the bronchioles and alveoli and reducing the work of breathing. It also possesses a barrier function for maintaining the blood-gas interface of the lungs and plays an important role in innate immunity. The surfactant film covers the epithelium lining both large and small airways, forming the first line of defense between toxic airborne particles/pathogens and the lungs. Furthermore, surfactant has been shown to relax airway smooth muscle (ASM) after exposure to ASM agonists, suggesting a more subtle function. Whether surfactant masks irritant sensory receptors or interacts with one of them is not known. The relaxant effect of surfactant on ASM is absent in bronchial tissues denuded of an epithelial layer. Blocking of prostanoid synthesis inhibits the relaxant function of surfactant, indicating that prostanoids might be involved. Another possibility for surfactant to be active, namely through ATP-dependent potassium channels and the cAMP-regulated epithelial chloride channels [cystic fibrosis transmembrane conductance regulators (CFTRs)], was tested but could not be confirmed. Hence, this review discusses the mechanisms of known and potential relaxant effects of pulmonary surfactant on ASM. This review summarizes what is known about the role of surfactant in smooth muscle physiology and explores the scientific questions and studies needed to fully understand how surfactant helps maintain the delicate balance between relaxant and constrictor needs. [ABSTRACT FROM AUTHOR]

Just over thirty years is the span of a generation. It is also the time that has passed since the discovery of the gene responsible for cystic fibrosis. Today, it is safe to say that this discovery has revolutionized our understanding, research perspectives, and management of this disease, which was, thirty years ago, a pediatric condition with a grim prognosis. The aim of this review is to present the advances that science and medicine have brought to our understanding of the pathophysiology of the disease and its management, which in many ways, epitomizes modern molecular genetic research. Since the discovery of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in 1989, modeling the CFTR protein, deciphering its function as an ion channel, and identifying its molecular partners have led to numerous therapeutic advances. The most significant advancement in this field has been the discovery of protein modulators that can target its membrane localization and chloride channel activity. However, further progress is needed to ensure that all patients can benefit from a therapy tailored to their mutations, with the primary challenge being the development of treatments for mutations leading to a complete absence of the protein. The present review delves into the history of the multifaceted world of CF, covering main historical facts, current landscape, clinical management, emerging therapies, patient perspectives, and the importance of ongoing research, bridging science and medicine in the fight against the disease. [ABSTRACT FROM AUTHOR]

Cystic fibrosis (CF) is a serious genetic disease caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Approved small molecule therapies benefit the majority of people with CF (pwCF), but unfortunately not all. Gene addition offers a mutation agnostic treatment option for all pwCF. SP-101 is an adeno-associated virus gene therapy vector (AAV2.5T) that has been optimized for efficient human airway cell transduction, and that contains a functional and regulated shortened human CFTR minigene (hCFTRΔR) with a small synthetic promoter/enhancer. To understand SP-101 airway distribution, activity, and the associated immune response, in vivo studies were performed in wild-type and CF ferrets. After single dose inhaled delivery of SP-101, followed by single dose inhaled doxorubicin (an AAV transduction augmenter) or saline, SP-101 vector genomes were detected throughout the respiratory tract. hCFTRΔR mRNA expression was highest in ferrets also receiving doxorubicin and persisted for the duration of the study (13 weeks). Pre-existing mucus in the CF ferrets did not present a barrier to effective transduction. Binding and neutralizing antibodies to the AAV2.5T capsid were observed regardless of doxorubicin exposure. Only a portion of ferrets exhibited a weak T-cell response to AAV2.5T and no T-cell response was seen against hCFTRΔR. These data strongly support the continued development of inhaled SP-101, followed by inhaled doxorubicin, for the treatment of CF. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a highly prevalent joint disease that causes substantial disability, yet effective approaches to disease prevention or to the delay of OA progression are lacking. Emerging evidence has pinpointed ion channels as pivotal mediators in OA pathogenesis and as promising targets for disease-modifying treatments. Preclinical studies have assessed the potential of a variety of ion channel modulators to modify disease pathways involved in cartilage degeneration, synovial inflammation, bone hyperplasia and pain, and to provide symptomatic relief in models of OA. Some of these modulators are currently being evaluated in clinical trials. This review explores the structures and functions of ion channels, including transient receptor potential channels, Piezo channels, voltage-gated sodium channels, voltage-dependent calcium channels, potassium channels, acid-sensing ion channels, chloride channels and the ATP-dependent P2XR channels in the osteoarthritic joint. The discussion spans channel-targeting drug discovery and potential clinical applications, emphasizing opportunities for further research, and underscoring the growing clinical impact of ion channel biology in OA. Ion channels have key functions in chondrocytes, bone cells, immune cells and neurons. Liu and colleagues discuss how these functions might contribute to cartilage degeneration, bone formation inflammation and pain in osteoarthritis, and highlight the therapeutic potential of ion channel modulators. Key points: Ion channels have key functions in the joints and emerge as important contributors to pathogenic processes in osteoarthritis (OA). Dysregulation of mechanical and biochemical stimuli activates ion channels such as TRPV4 and Piezo channels, leading to Ca2+ and Na+ influx, which contributes to cartilage destruction, inflammation and pain in OA. Some ion channels typically associated with peripheral neurons are also expressed in chondrocytes and immune cells, and have a crucial role in the pathophysiology of OA. Exploration of ion channel expression, interactions, and properties in joints, along with development of specific channel agonists and antagonists, has helped to accelerate research on their potential roles in OA. Several ion channel modulators have been recently tested in animal models and patients with OA, with encouraging results. Targeting of ion channels holds promise for the development of new and more effective OA treatment strategies. [ABSTRACT FROM AUTHOR]

ClC‐K/barttin channels are involved in the transepithelial transport of chloride in the kidney and inner ear. Their physiological role is crucial in humans because mutations in CLCNKB or BSND, encoding ClC‐Kb and barttin, cause Bartter's syndrome types III and IV, respectively. In vitro experiments have shown that an amino acid change in a proline‐tyrosine motif in the C‐terminus of barttin stimulates ClC‐K currents. The molecular mechanism of this enhancement and whether this potentiation has any in vivo relevance remains unknown. We performed electrophysiological and biochemical experiments in Xenopus oocytes and kidney cells co‐expressing ClC‐K and barttin constructs. We demonstrated that barttin possesses a YxxØ motif and, when mutated, increases ClC‐K plasma membrane stability, resulting in larger currents. To address the impact of mutating this motif in kidney physiology, we generated a knock‐in mouse. Comparing wild‐type (WT) and knock‐in mice under a standard diet, we could not observe any difference in ClC‐K and barttin protein levels or localization, either in urinary or plasma parameters. However, under a high‐sodium low‐potassium diet, known to induce hyperplasia of distal convoluted tubules, knock‐in mice exhibit reduced hyperplasia compared to WT mice. In summary, our in vitro and in vivo studies demonstrate that the previously identified PY motif is indeed an endocytic YxxØ motif in which mutations cause a gain of function of the channel. Key points: It is revealed by mutagenesis and functional experiments that a previously identified proline‐tyrosine motif regulating ClC‐K plasma membrane levels is indeed an endocytic YxxØ motif.Biochemical characterization of mutants in the YxxØ motif in Xenopus oocytes and human embryonic kidney cells indicates that mutants showed increased plasma membrane levels as a result of an increased stability, resulting in higher function of ClC‐K channels.Mutation of this motif does not affect barttin protein expression and subcellular localization in vivo.Knock‐in mice with a mutation in this motif, under conditions of a high‐sodium low‐potassium diet, exhibit less hyperplasia in the distal convoluted tubule than wild‐type animals, indicating a gain of function of the channel in vivo. [ABSTRACT FROM AUTHOR]

ObjectiveDespite great advances in obesity therapeutics in recent years, there is still a need to identify additional therapeutic targets for the treatment of this disease. We previously discovered a signature of genes, including Chloride intracellular channel 1 (Clic1), whose expression was associated with drug-induced weight gain, and in these studies, we assess the effect of Clic1 inhibition on food intake and body weight in mice.MethodsWe studied the impact of Clic1 inhibition in mouse models of binge-eating, diet-induced obese mice and genetic models of obesity (Magel2 KO mice).ResultsClic1 knockout (KO) mice ate significantly less and had a lower body weight than WT littermates when either fed chow or high fat diet. Furthermore, pharmacological inhibition of Clic1 in diet-induced obese mice resulted in suppression of food intake and promoted highly efficacious weight loss. Clic1 inhibition also reduced food intake in binge-eating models and hyperphagic Magel2 KO mice. We observed that chronic obesity resulted in a significant change in subcellular localization of Clic1 with an increased ratio of Clic1 in the membrane in the obese state. These observations provide a novel therapeutic strategy to block Clic1 translocation as a potential mechanism to reduce food intake and lower body weight.ConclusionsThese studies attribute a novel role of Clic1 as a driver of food intake and overconsumption. In summary, we have identified hypothalamic expression of Clic1 plays a key role in food intake, providing a novel therapeutic target to treat overconsumption that is the root cause of modern obesity.

Nature performs critical physiological functions using a series of structurally and functionally diverse membrane proteins embedded in cell membranes, in which native ion protein channels modify the electrical potential inside and outside the cell membrane through charged ion movements. Consequently, the cell responds to external stimuli, playing an essential role in various life activities, such as nerve excitation conduction, neurotransmitter release, muscle movement, and control of cell differentiation. Supramolecular artificial channels, which mimic native protein channels in structure and function, adopt unimolecular or self‐assembled structures, such as crown ethers, cyclodextrins, cucurbiturils, column arenes, cyclic peptide nanotubes, and metal‐organic artificial channels, in channel construction strategies. Owing to the various driving forces involved, artificial synthetic ion channels can be divided into artificial cation and anion channels in terms of ion selectivity. Cation selectivity usually originates from ion coordination, whereas anion selectivity is related to hydrogen bonding, ion pairing, and anion‐dipole interactions. Several studies have been conducted on artificial cation channels, and several reviews have summarized them in detail; however, the research on anions is still in the initial stages, and related reviews have rarely been reported. Hence, this article primarily focuses on the recent research on anion channels. [ABSTRACT FROM AUTHOR]

Cystic fibrosis (CF) is the most common life-threatening genetic disease in the United States and among people of European descent. Despite the widespread distribution of the cystic fibrosis transmembrane conductance regulator (CFTR) along kidney tubules, specific renal phenotypes attributable to CF have not been well documented. Recent studies have demonstrated the downregulation of the apical Cl-/HCO3 - exchanger pendrin (Slc26a4) in kidney B-intercalated cells of CF mouse models. These studies have shown that kidneys of both mice and humans with CF have an impaired ability to excrete excess HCO3 -, thus developing metabolic alkalosis when subjected to excess HCO3 - intake. The purpose of this minireview is to discuss the latest advances on the role of pendrin as a molecule with dual critical roles in acid base regulation and systemic vascular volume homeostasis, specifically in CF. Given the immense prevalence of vascular volume depletion, which is primarily precipitated via enhanced chloride loss through perspiration, we suggest that the dominant presentation of metabolic alkalosis in CF is due to the impaired function of pendrin, which plays a critical role in systemic vascular volume and acid base homeostasis. [ABSTRACT FROM AUTHOR]

The function of the chloride channel ClC‐1 is crucial for the control of muscle excitability. Thus, reduction of ClC‐1 functions by CLCN1 mutations leads to myotonia congenita. Many different animal models have contributed to understanding the myotonia pathophysiology. However, these models do not allow in vivo screening of potentially therapeutic drugs, as the zebrafish model does. In this work, we identified and characterized the two zebrafish orthologues (clc‐1a and clc‐1b) of the ClC‐1 channel. Both channels are mostly expressed in the skeletal muscle as revealed by RT‐PCR, western blot, and electrophysiological recordings of myotubes, and clc‐1a is predominantly expressed in adult stages. Characterization in Xenopus oocytes shows that the zebrafish channels display similar anion selectivity and voltage dependence to their human counterparts. However, they show reduced sensitivity to the inhibitor 9‐anthracenecarboxylic acid (9‐AC), and acidic pH inverts the voltage dependence of activation. Reduction of clc‐1a/b expression hampers spontaneous and mechanically stimulated movement, which could be reverted by expression of human ClC‐1 but not by some ClC‐1 containing myotonia mutations. Treatment of clc‐1‐depleted zebrafish with mexiletine, a typical drug used in human myotonia, improves the motor behaviour. Our work extends the repertoire of ClC channels to evolutionary structure–function studies and proposes the zebrafish clcn1 crispant model as a simple tool to find novel therapies for myotonia. Key points: We have identified two orthologues of ClC‐1 in zebrafish (clc‐1a and clc‐1b) which are mostly expressed in skeletal muscle at different developmental stages.Functional characterization of the activity of these channels reveals many similitudes with their mammalian counterparts, although they are less sensitive to 9‐AC and acidic pH inverts their voltage dependence of gating.Reduction of clc‐1a/b expression hampers spontaneous and mechanically stimulated movement which could be reverted by expression of human ClC‐1.Myotonia‐like symptoms caused by clc‐1a/b depletion can be reverted by mexiletine, suggesting that this model could be used to find novel therapies for myotonia. [ABSTRACT FROM AUTHOR]

Many years ago, asbestos fibers were banned and replaced by synthetic vitreous fibers because of their carcinogenicity. However, the toxicity of the latter fibers is still under debate, especially when it concerns the early fiber interactions with biological cell membranes. Here, we aimed to investigate the effects of a synthetic vitreous fiber named FAV173 on the Xenopus laevis oocyte membrane, the cell model we have already used to characterize the effect of crocidolite asbestos fiber exposure. Using an electrophysiological approach, we found that, similarly to crocidolite asbestos, FAV173 was able to stimulate a chloride outward current evoked by step membrane depolarizations, that was blocked by the potent and specific TMEM16A channel antagonist Ani9. Exposure to FAV173 fibers also altered the oocyte cell membrane microvilli morphology similarly to crocidolite fibers, most likely as a consequence of the TMEM16A protein interaction with actin. However, FAV173 only partially mimicked the crocidolite fibers effects, even at higher fiber suspension concentrations. As expected, the crocidolite fibers' effect was more similar to that induced by the co-treatment with (Fe3+ + H2O2), since the iron content of asbestos fibers is known to trigger reactive oxygen species (ROS) production. Taken together, our findings suggest that FAV173 may be less harmful that crocidolite but not ineffective in altering cell membrane properties. [ABSTRACT FROM AUTHOR]

The article discusses the seventh cholera pandemic, which began in 1961 and highlights how cholera has historically linked different parts of the world and affected both rich and poor communities. Topics include the pathophysiology of cholera and the discovery of intestinal secretion, secretory diarrheas other than cholera, and evidence that chloride secretion was at the center of the pathophysiology of cystic fibrosis.

This article discusses the mechanisms by which enteric viruses, such as rotavirus, norovirus, sapovirus, astrovirus, adenovirus, and poliovirus, induce diarrhea. The viruses disrupt intestinal homeostasis by infecting specific cells and causing structural changes, impairing nutrient absorption and leading to malabsorptive diarrhea. The enteric nervous system is also involved in diarrhea development, as viral infections activate serotonin secretion, leading to increased intestinal motility and fluid secretion. Viroporins, which are virally encoded proteins, play a role in manipulating cellular calcium signaling and facilitating viral replication, contributing to diarrhea. The article concludes by highlighting the importance of understanding these mechanisms for the development of antivirals and vaccines to reduce infection-associated diarrheal illness. [Extracted from the article]