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Frontiers in Physiology 2024Intestinal organoids are stem cell-derived, 3D "mini-guts" with similar functions as the native intestinal epithelium such as electrolyte transport or establishment of...
BACKGROUND
Intestinal organoids are stem cell-derived, 3D "mini-guts" with similar functions as the native intestinal epithelium such as electrolyte transport or establishment of an epithelial barrier. During intestinal inflammation, epithelial functions are dysregulated by proinflammatory cytokines like tumor necrosis factor α (TNFα) and other messengers from the immune system resulting in a loss of electrolytes and water due to an impaired epithelial barrier and higher net secretion.
METHODS
A murine small intestinal organoid model was established to study (long-term) effects of TNFα on the intestinal epithelium using live imaging, immunohistochemical staining and qPCR.
RESULTS
TNFα induced apoptosis in intestinal organoids as indicated by an increased number of cells with immunoreactivity for cleaved caspase 3. Furthermore, TNFα exposure led to swelling of the organoids which was inhibited by bumetanide and was concomitant with an upregulation of the bumetanide-sensitive Na-K-2Cl symporter 1 (NKCC1) as shown by qPCR. Fura-2 imaging experiments revealed time-dependent changes in Ca signaling consisting of a rise in the basal cytosolic Ca concentration at day 1 and an increase of the carbachol-induced Ca response after 3 days TNFα exposure. This was prevented by preincubation with La, an inhibitor of non-selective cation channels, or by using a Ca-free buffer indicating an enhancement of the Ca influx from the extracellular side by the cytokine. No significant changes in cDNA levels of epithelial barrier proteins could be observed in the presence of TNFα.
CONCLUSION
Intestinal organoids are a useful tool to study the mechanism underlying the TNFα-induced secretion on enterocytes such as the regulation of NKCC1 expression or the modulation of cellular Ca signaling.
PubMed: 38737827
DOI: 10.3389/fphys.2024.1382238 -
Journal of Medicine and Life Jan 2024Heart failure (HF) remains a significant problem for healthcare systems, requiring the use of intervention and multimodal management strategies. We aimed to assess the...
Heart failure (HF) remains a significant problem for healthcare systems, requiring the use of intervention and multimodal management strategies. We aimed to assess the short-term effect of empagliflozin (EMPA) and metformin on cardiac function parameters, including ventricular dimension-hypertrophy, septal thickness, ejection fraction (EF), and N-terminal pro-brain natriuretic peptide (NT-proBNP) levels in patients with HF and mildly reduced EF. A case-control study included 60 newly diagnosed patients with HF. Patients were divided into two groups: Group E received standard HF treatment (carvedilol, bumetanide, sacubitril-valsartan, spironolactone) plus EMPA 10 mg daily, and Group M received standard HF treatment plus metformin 500 mg daily. After three months of treatment, Group E had a significantly higher EF than Group M compared to initial measurements (a change of 9.2% versus 6.1%, respectively). We found similar results in the left ventricular end-systolic dimension (LVESD), with mean reductions of 0.72 mm for Group E and 0.23 mm for Group M. Regarding cardiac indicators, the level of NT-proBNP was considerably decreased in both groups. However, the reduction was significantly greater in group E than in group M compared to the initial level (mean reduction: 719.9 vs. 973.6, respectively). When combined with quadruple anti-heart failure therapy, metformin enhanced several echocardiographic parameters, showing effects similar to those of EMPA when used in the same treatment regimen. However, the benefits of EMPA were more pronounced, particularly regarding improvements in EF and LVESD.
Topics: Humans; Heart Failure; Benzhydryl Compounds; Glucosides; Metformin; Stroke Volume; Male; Female; Case-Control Studies; Middle Aged; Aged; Natriuretic Peptide, Brain; Peptide Fragments; Echocardiography; Sodium-Glucose Transporter 2 Inhibitors
PubMed: 38737651
DOI: 10.25122/jml-2023-0340 -
Experimental Neurobiology Apr 2024In the auditory system, the spontaneous activity of cochlear inner hair cells (IHCs) is initiated by the release of ATP from inner supporting cells (ISCs). This ATP...
In the auditory system, the spontaneous activity of cochlear inner hair cells (IHCs) is initiated by the release of ATP from inner supporting cells (ISCs). This ATP release sets off a cascade, activating purinergic autoreceptors, opening of Ca-activated Cl channel TMEM16A, Cl efflux and osmotic cell shrinkage. Then, the shrunken ISCs efficiently regain their original volume, suggesting the existence of mechanisms for refilling Cland K, priming them for subsequent activity. This study explores the potential involvement of NKCCs (Na-K-Cl cotransporters) and KCCs (K-Cl cotransporters) in ISC spontaneous activity, considering their capability to transport both Cl and K ions across the cell membrane. Employing a combination of immunohistochemistry, pharmacological interventions, and shRNA experiment, we unveiled the pivotal role of NKCC1 in cochlear spontaneous activity. Immunohistochemistry revealed robust NKCC1 expression in ISCs, persisting until the 2nd postnatal week. Intriguingly, we observed a developmental shift in NKCC1 expression from ISCs to synaptophysin-positive efferent terminals at postnatal day 18, hinting at its potential involvement in modulating synaptic transmission during the post-hearing period. Experiments using bumetanide, a well-known NKCC inhibitor, supported the functional significance of NKCC1 in ISC spontaneous activity. Bumetanide significantly reduced the frequency of spontaneous extracellular potentials (sEP) and spontaneous optical changes (sOCs) in ISCs. NKCC1-shRNA experiments conducted in cultured cochlear tissues further supported these findings, demonstrating a substantial decrease in event frequency and area. Taken together, we revealed the role of NKCC1 in shaping the ISC spontaneous activity that govern auditory pathway development.
PubMed: 38724477
DOI: 10.5607/en24003 -
Current Issues in Molecular Biology Feb 2024Autism spectrum disorder (ASD) is thought to result from susceptibility genotypes and environmental risk factors. The offspring of women who experience pregnancy...
Autism spectrum disorder (ASD) is thought to result from susceptibility genotypes and environmental risk factors. The offspring of women who experience pregnancy infection have an increased risk for autism. Maternal immune activation (MIA) in pregnant animals produces offspring with autistic behaviors, making MIA a useful model for autism. However, how MIA causes autistic behaviors in offspring is not fully understood. Here, we show that NKCC1 is critical for mediating autistic behaviors in MIA offspring. We confirmed that MIA induced by poly(I:C) infection during pregnancy leads to autistic behaviors in offspring. We further demonstrated that MIA offspring showed significant microglia activation, excessive dendritic spines, and narrow postsynaptic density (PSD) in their prefrontal cortex (PFC). Then, we discovered that these abnormalities may be caused by overexpression of NKCC1 in MIA offspring's PFCs. Finally, we ameliorated the autistic behaviors using PFC microinjection of NKCC1 inhibitor bumetanide (BTN) in MIA offspring. Our findings may shed new light on the pathological mechanisms for autism caused by pregnancy infection.
PubMed: 38534737
DOI: 10.3390/cimb46030121 -
Journal of Alzheimer's Disease : JAD 2024Alzheimer's disease (AD) is a chronic neurodegenerative disease needing effective therapeutics urgently. Sildenafil, one of the approved phosphodiesterase-5 inhibitors,...
Sildenafil as a Candidate Drug for Alzheimer's Disease: Real-World Patient Data Observation and Mechanistic Observations from Patient-Induced Pluripotent Stem Cell-Derived Neurons.
BACKGROUND
Alzheimer's disease (AD) is a chronic neurodegenerative disease needing effective therapeutics urgently. Sildenafil, one of the approved phosphodiesterase-5 inhibitors, has been implicated as having potential effect in AD.
OBJECTIVE
To investigate the potential therapeutic benefit of sildenafil on AD.
METHODS
We performed real-world patient data analysis using the MarketScan® Medicare Supplemental and the Clinformatics® databases. We conducted propensity score-stratified analyses after adjusting confounding factors (i.e., sex, age, race, and comorbidities). We used both familial and sporadic AD patient induced pluripotent stem cells (iPSC) derived neurons to evaluate the sildenafil's mechanism-of-action.
RESULTS
We showed that sildenafil usage is associated with reduced likelihood of AD across four new drug compactor cohorts, including bumetanide, furosemide, spironolactone, and nifedipine. For instance, sildenafil usage is associated with a 54% reduced incidence of AD in MarketScan® (hazard ratio [HR] = 0.46, 95% CI 0.32- 0.66) and a 30% reduced prevalence of AD in Clinformatics® (HR = 0.70, 95% CI 0.49- 1.00) compared to spironolactone. We found that sildenafil treatment reduced tau hyperphosphorylation (pTau181 and pTau205) in a dose-dependent manner in both familial and sporadic AD patient iPSC-derived neurons. RNA-sequencing data analysis of sildenafil-treated AD patient iPSC-derived neurons reveals that sildenafil specifically target AD related genes and pathobiological pathways, mechanistically supporting the beneficial effect of sildenafil in AD.
CONCLUSIONS
These real-world patient data validation and mechanistic observations from patient iPSC-derived neurons further suggested that sildenafil is a potential repurposable drug for AD. Yet, randomized clinical trials are warranted to validate the causal treatment effects of sildenafil in AD.
Topics: Aged; United States; Humans; Alzheimer Disease; Induced Pluripotent Stem Cells; Sildenafil Citrate; Neurodegenerative Diseases; Spironolactone; tau Proteins; Medicare; Neurons
PubMed: 38427489
DOI: 10.3233/JAD-231391 -
Pflugers Archiv : European Journal of... May 2024Increases in the current threshold occur in optic nerve axons with the application of infra-red laser light, whose mechanism is only partly understood. In isolated rat...
Increases in the current threshold occur in optic nerve axons with the application of infra-red laser light, whose mechanism is only partly understood. In isolated rat optic nerve, laser light was applied near the site of electrical stimulation, via a flexible fibre optic. Paired applications of light produced increases in threshold that were reduced on the second application, the response recovering with increasing delays, with a time constant of 24 s. 3-min duration single applications of laser light gave rise to a rapid increase in threshold followed by a fade, whose time-constant was between 40 and 50 s. After-effects were sometimes apparent following the light application, where the resting threshold was reduced. The increase in threshold was partially blocked by 38.6 mM Li in combination with 5 M bumetanide, a manoeuvre increasing refractoriness and consistent with axonal depolarization. Assessing the effect of laser light on the nerve input resistance ruled out a previously suggested fall in myelin resistance as contributing to threshold changes. These data appear consistent with an axonal membrane potential that partly relies on temperature-dependent electroneutral Na influx, and where fade in the response to the laser may be caused by a gradually diminishing Na pump-induced hyperpolarization, in response to falling intracellular [Na].
Topics: Animals; Rats; Optic Nerve; Lasers; Sodium; Axons; Membrane Potentials; Male; Bumetanide; Rats, Sprague-Dawley
PubMed: 38421407
DOI: 10.1007/s00424-024-02932-1 -
Frontiers in Neuroscience 2024Equipped with an early social predisposition immediately post-birth, humans typically form associations with mothers and other family members through exposure learning,... (Review)
Review
Equipped with an early social predisposition immediately post-birth, humans typically form associations with mothers and other family members through exposure learning, canalized by a prenatally formed predisposition of visual preference to biological motion, face configuration, and other cues of animacy. If impaired, reduced preferences can lead to social interaction impairments such as autism spectrum disorder (ASD) via misguided canalization. Despite being taxonomically distant, domestic chicks could also follow a homologous developmental trajectory toward adaptive socialization through imprinting, which is guided via predisposed preferences similar to those of humans, thereby suggesting that chicks are a valid animal model of ASD. In addition to the phenotypic similarities in predisposition with human newborns, accumulating evidence on the responsible molecular mechanisms suggests the construct validity of the chick model. Considering the recent progress in the evo-devo studies in vertebrates, we reviewed the advantages and limitations of the chick model of developmental mental diseases in humans.
PubMed: 38356650
DOI: 10.3389/fnins.2024.1279947 -
Nanomaterials (Basel, Switzerland) Jan 2024Poorly water-soluble drugs represent a challenge for the pharmaceutical industry because it is necessary to find properly tuned and efficient systems for their release....
Poorly water-soluble drugs represent a challenge for the pharmaceutical industry because it is necessary to find properly tuned and efficient systems for their release. In this framework, organic-inorganic hybrid systems could represent a promising strategy. A largely diffused inorganic host is hydroxyapatite (HAP, Ca(PO)(OH)), which is easily synthesized with different external forms and can adsorb different kinds of molecules, thereby allowing rapid drug release. Hybrid nanocomposites of HAP nanorods, obtained through hydrothermal synthesis, were prepared with two model pharmaceutical molecules characterized by low and pH-dependent solubility: meloxicam, a non-steroidal anti-inflammatory drug, and bumetanide, a diuretic drug. Both hybrids were physically and chemically characterized through the combined use of X-ray powder diffraction, scanning electron microscopy with energy-dispersive spectroscopy, differential scanning calorimetry, and infrared spectroscopy measurements. Then, their dissolution profiles and hydrophilicity (contact angles) in different media as well as their solubility were determined and compared to the pure drugs. This hybrid system seems particularly suitable as a drug carrier for bumetanide, as it shows higher drug loading and good dissolution profiles, while is less suitable for meloxicam, an acid molecule.
PubMed: 38202568
DOI: 10.3390/nano14010113 -
Frontiers in Aging 2023To investigate whether exposure history to two common loop diuretics, bumetanide and furosemide, affects the risk of developing Alzheimer's disease (AD) after...
To investigate whether exposure history to two common loop diuretics, bumetanide and furosemide, affects the risk of developing Alzheimer's disease (AD) after accounting for socioeconomic status and congestive heart failure. Individuals exposed to bumetanide or furosemide were identified in the Stanford University electronic health record using the de-identified Observational Medical Outcomes Partnership platform. We matched the AD case cohort to a control cohort (1:20 case:control) on gender, race, ethnicity, and hypertension, and controlled for variables that could potentially be collinear with bumetanide exposure and/or AD diagnosis. Among individuals older than 65 years, 5,839 AD cases and 116,103 matched controls were included. A total of 1,759 patients (54 cases and 1,705 controls) were exposed to bumetanide. After adjusting for socioeconomic status and other confounders, the exposure of bumetanide and furosemide was significantly associated with reduced AD risk (respectively, bumetanide odds ratio [OR] = 0.23; 95% confidence interval [CI], 0.15-0.36; = 4.0 × 10; furosemide OR = 0.42; 95% CI, 0.38-0.47; < 2.0 × 10). Our study replicates in an independent sample that a history of bumetanide exposure is associated with reduced AD risk while also highlighting an association of the most common loop diuretic (furosemide) with reduced AD risk. These associations need to be additionally replicated, and the mechanism of action remains to be investigated.
PubMed: 37822457
DOI: 10.3389/fragi.2023.1211571 -
Cerebral Cortex Communications 2022Several environmental chemicals are suspected risk factors for autism spectrum disorder (ASD), including valproic acid (VPA) and pesticides acting on nicotinic...
Several environmental chemicals are suspected risk factors for autism spectrum disorder (ASD), including valproic acid (VPA) and pesticides acting on nicotinic acetylcholine receptors (nAChRs), if administered during pregnancy. However, their target processes in fetal neuro-development are unknown. We report that the injection of VPA into the fetus impaired imprinting to an artificial object in neonatal chicks, while a predisposed preference for biological motion (BM) remained intact. Blockade of nAChRs acted oppositely, sparing imprinting and impairing BM preference. Beside ketamine and tubocurarine, significant effects of imidacloprid (a neonicotinoid insecticide) appeared at a dose ≤1 ppm. In accord with the behavioral dissociations, VPA enhanced histone acetylation in the primary cell culture of fetal telencephalon, whereas ketamine did not. VPA reduced the brain weight and the ratio of NeuN-positive cells (matured neurons) in the telencephalon of hatchlings, whereas ketamine/tubocurarine did not. Despite the distinct underlying mechanisms, both VPA and nAChR blockade similarly impaired imprinting to biological image composed of point-light animations. Furthermore, both impairments were abolished by postnatal bumetanide treatment, suggesting a common pathology underlying the social attachment malformation. Neurotransmission via nAChR is thus critical for the early social bond formation, which is hindered by ambient neonicotinoids through impaired visual predispositions for animate objects.
PubMed: 37674673
DOI: 10.1093/texcom/tgac041