Inhibition Of Calcium Influx Research Articles

8538 Pancreatic Islet Adaptations to Hyperglycemia Are Prevented by Glycolytic Inhibition and Amplified by Inhibiting Membrane Depolarization

Abstract Disclosure: N.B. Whitticar: None. B.P. List: None. N. Bruce: None. R. Bertram: None. C.S. Nunemaker: None. Long before type 2 diabetes ensues, pancreatic islets undergo functional changes to compensate for increasing insulin resistance. Primarily, the islets increase their ability to secrete insulin by augmenting the glucose-stimulated insulin secretion (GSIS) pathway. As metabolic syndrome worsens, islets become functionally exhausted, leading to cellular dysfunction and apoptosis. To avoid the negative consequences of this compensatory response, various steps of the GSIS pathway can be attenuated to restore normal functional levels before the islets fail. Determining which inhibitors can prevent the earliest adaptations to hyperglycemia may reveal the processes that play a causal role in islet failure. The chemicals mannoheptulose (MH), diazoxide (DZ), and nifedipine (NP) were chosen to inhibit the glucokinase enzyme, KATP-channel closure, and voltage-gated calcium channels, respectively. CD-1 mouse islets were placed in 20 mM glucose to simulate hyperglycemia along with a concentration of each inhibitory drug that reduced insulin secretion by 30-40%. After 48-hour treatment, glucose-stimulated calcium tests were performed to determine if the treatments could prevent the negative adaptation to hyperglycemia. Only islets treated with MH to reduce glycolytic activity maintained a normal calcium response. Islets in the MH group also retained a strong insulin response to glucose and were not depleted of insulin content. Unexpectedly, inhibition of calcium influx and insulin secretion with DZ or NF caused a substantial increase in basal insulin secretion compared to islets in standard culture (P<0.01, n=6; P=0.06, n=6, respectively). These results indicate that the signaling pathway that causes the adaptation to hyperglycemia lies upstream of KATP channels and downstream of glucokinase. Mathematical modeling using the Integrated Oscillator Model was able to replicate our results and suggests that high rates of glycolytic and mitochondrial metabolism are associated with the signal that adapts islets to hyperglycemia, and that reducing intracellular calcium in the face of high glucose further increases basal insulin secretion. Ongoing studies will determine if metabolic acceleration or altered membrane conductance are responsible for the earliest islet adaptations to hyperglycemia. Our results suggest that reducing metabolic activity, but not calcium influx, prevents adaptations to hyperglycemia in mouse islets. These findings can be applied to potentially prevent the progression of obesity and hyperinsulinemia into type 2 diabetes. Presentation: 6/2/2024

Solubility determination and thermodynamic modelling of verapamil hydrochloride in methanol-water binary solvent systems from 293.15 K to 323.15 K

Verapamil Hydrochloride is a key calcium channel blocker used to treat cardiovascular disorders and prevent cluster headaches by inhibiting calcium influx, thus improving cardiovascular health. The solubility of Verapamil Hydrochloride (VH) in methanol + water binary solvent systems were studied using a gravimetric method at temperatures from 293.15 K to 323.15 K under atmospheric pressure. The solubility increased with temperature and methanol content, with the highest mole fraction solubility at a 0.90 methanol mole fraction and the lowest at 0.10. Various models, including Apelblat, Van't Hoff, modified Jouyban–Acree, and CNIBS/R-K, were used to calculate theoretical solubility, with RAD and RMSD assessing the model accuracy. Thermodynamic analysis indicated that VH dissolution is endothermic and entropy-driven. This data is valuable for optimizing VH purification and crystallization processes in the pharmaceutical industry.

Nitazoxanide synergizes polymyxin B against Escherichia coli by depleting cellular energy.

The rapid expansion of antibiotic-resistant bacterial diseases is a global burden on public health. It makes sense to repurpose and reposition already-approved medications for use as supplementary agents in synergistic combinations with existing antibiotics. Here, we demonstrate that the anthelmintic drug nitazoxanide (NTZ) synergistically enhances the effectiveness of the lipopeptide antibiotic polymyxin B in inhibiting gram-negative bacteria, including those resistant to polymyxin B. Mechanistic investigations revealed that nitazoxanide inhibited calcium influx and cell membrane depolarization, enhanced the affinity between polymyxin B and the extracellular membrane, and promoted intracellular ATP depletion and an increase in reactive oxygen species (ROS), thus enhancing the penetration and disruption of the Escherichia coli cell membrane by polymyxin B. The transcriptomic analysis revealed that the combination resulted in energy depletion by inhibiting both aerobic and anaerobic respiration patterns in bacterial cells. The increased bactericidal effect of polymyxin B on the E. coli ∆nuoC strain further indicates that NuoC could be a promising target for nitazoxanide. Furthermore, the combination of nitazoxanide and polymyxin B showed promising therapeutic effects in a mouse infection model infected with E. coli. Taken together, these results demonstrate the potential of nitazoxanide as a novel adjuvant to polymyxin B, to overcome antibiotic resistance and improve therapeutic outcomes in refractory infections.IMPORTANCEThe rapid spread of antibiotic-resistant bacteria poses a serious threat to public health. The search for potential compounds that can increase the antibacterial activity of existing antibiotics is a promising strategy for addressing this issue. Here, the synergistic activity of the FDA-approved agent nitazoxanide (NTZ) combined with polymyxin B was investigated in vitro using checkerboard assays and time-kill curves. The synergistic mechanisms of the combination of nitazoxanide and polymyxin B were explored by fluorescent dye, transmission electron microscopy (TEM), and transcriptomic analysis. The synergistic efficacy was evaluated in vivo by the Escherichia coli and mouse sepsis models. These results suggested that nitazoxanide, as a promising antibiotic adjuvant, can effectively enhance polymyxin B activity, providing a potential strategy for treating multidrug-resistant bacteria.

Antispasmodic Activity of Light-Roasted Coffee Extract and Its Potential Use in Gastrointestinal Motility Disorders.

Antispasmodic agents are crucial in managing gastrointestinal motility disorders by modulating muscle contractions and reducing symptoms like cramping and diarrhea. This study investigated the antispasmodic potential of different coffee bean extracts, including light coffee (LC), medium coffee (MC), and dark coffee (DC), on ileum contractions induced by potassium chloride (KCl), and elucidated their mechanisms of action using in vitro isolated tissue techniques. The results demonstrated that all coffee extracts reduced spontaneous contractions of rat ileum tissue in a dose-dependent manner. Among these, LC showed the most significant reduction in ileum contractions, particularly at higher concentrations. The key findings reveal that LC at 5 mg/mL significantly reduced CaCl2-induced contractions in isolated rat ileum tissue, indicating that LC may inhibit calcium influx or interfere with calcium signaling pathways. The presence of nifedipine, propranolol, and N-nitro-L-arginine methyl ester (L-NAME) have been confirmed in their involvement; they block calcium influx and calcium channels and activate β-adrenergic pathways as part of LC's mechanism of action. The presence of their active compounds, particularly chlorogenic acid and caffeine, likely contributes to the observed antispasmodic effects. These findings suggest that LC exerts its antispasmodic effects by targeting key mechanisms involved in muscle spasms and intestinal motility, providing a potential for managing such conditions.

Sertraline Alleviates Chronic Prostatitis by Regulating the TRPV1 Channel.

Although sertraline has been widely used for chronic prostatitis (CP), the mechanisms are unclear. Herein, we explored the mechanisms of sertraline in treating CP. Network pharmacology methods were used to explore the potential targets and molecular mechanisms. LPS was used to stimulate RWPE-1 cells to construct an in vitro model of CP. An experimental autoimmune prostatitis (EAP) mice model was built. CCK-8 assay, EdU assay, BrdU detection, and Tunel assay were performed to evaluate the proliferation and apoptosis process of cells or tissues, respectively. DCFH-DA and Fluo-4 fluorescence probes were used to detect intracellular ROS and calcium concentrations. Von Frey filaments and open-field tests were utilized to evaluate pain response and depressive-like behavior of mice. Histopathology was evaluated through hematoxylin and eosin staining. RT-qPCR, Western blot, immunofluorescence, and immunohistochemistry were utilized to evaluate the transcription, expression, and location of related proteins. Molecular dynamics (MD) simulation and surface plasmon resonance (SPR) assay were performed to measure the binding capacity of sertraline and related proteins. Through a network pharmacology analysis, 27 potential targets of sertraline for CP were obtained, and 5 key targets (CHRM1, ADRA1B, HTR2B, HTR2A, and TRPV1) were finally identified. Functional experiments suggested that TRPV1 was involved in the proliferation, apoptosis inhibition, and ROS production of LPS-induced RWPE-1 cells. In vitro experiments showed that sertraline significantly inhibited cell proliferation, ROS generation, and transcription of inflammation cytokines of LPS-induced RWPE-1 cells. Additionally, sertraline markedly promoted the apoptosis level of LPS-stimulated RWPE-1 cells and elevated the expression level of BAX while reducing the expression levels of Bcl2 and Caspase-3. MD simulation and SPR assay confirmed the direct binding of sertraline to TRPV1. Moreover, sertraline significantly down-regulated the expression level of TRPV1 and inhibited calcium influx of LPS-induced RWPE-1 cells. TRPV1 agonist (Capsaicin) significantly restored the effects on proliferation, apoptosis, ROS production, and calcium influx of sertraline on LPS-induced RWPE-1 cells. Mice experiments demonstrated that sertraline treatment could reduce pain response, improve depression-like symptoms, and relieve local prostate inflammation of EAP mice, as well as down-regulated the expression level of TRPV1, inhibit the proliferation, and promote apoptosis of prostate tissues in EAP mice. The results revealed the anti-inflammatory effect of sertraline for RWPE-1 cells and EAP mice, and the potential mechanism was regulating the TRPV1 channel. It indicated that sertraline might serve as a complementary anti-inflammatory agent for CP.

Vascular endothelial-cadherin is involved in endothelial cell detachment during thrombotic thrombocytopenic purpura

BackgroundImmune thrombotic thrombocytopenic purpura (i-TTP) is a life-threatening thrombotic microangiopathy linked to ADAMTS-13 deficiency. It has long been assumed that the activation of endothelial cells is the triggering factor for the thrombotic thrombocytopenic purpura crisis. Circulating endothelial cells (CECs) have been shown to be a biomarker of vascular damage and are associated with the clinical severity of i-TTP. However, the mechanisms leading to endothelial cell detachment remain unclear. ObjectivesWe investigated junctional destabilization the mechanisms underlying cell detachment in thrombotic thrombocytopenic purpura. MethodsWe quantified CECs in i-TTP patients and investigated the effect of plasmas in vitro by measuring phosphorylation and internalization of vascular endothelial (VE)-Cadherin and in vivo in a vascular permeability model. ResultsIn plasma from i-TTP patients, we show that CEC count is associated with severity and correlated to intracellular calcium influx (P < .01). In vitro, serum from i-TTP patients induced stronger detachment of human umbilical vein endothelial cells than serum from control patients (P < .001). Plasma from i-TTP patients induced a higher calcium-dependent phosphorylation (P < .05) and internalization (P < .05) of VE-cadherin compared with plasma from control patients. This effect could be reproduced by immunoglobulin (Ig)G fraction isolated from patient plasma and, in particular, by the F(ab)’2 fragments of the corresponding IgG. In addition, subcutaneous injection of i-TTP plasma into mice resulted in higher vascular permeability than plasma from control patients. An inhibitor of endothelial calcium influx, ITF1697, normalized this increase in permeability. ConclusionOur results suggest that plasma-induced endothelial activation also leads to an increase in vascular permeability. They contribute to the understanding of the mechanisms behind the presence of elevated CECs in patients' blood by linking endothelial activation to endothelial injury.

COMPARISON BETWEEN NEBULISATION OF MGSO4 20 MG/KGBW WITH KETAMINE 0.5 MG/KGBW ON THE INCIDENT OF POST-OPERATIVE SORE THROAT DUE TO ENDOTRACHEAL INTUBATION IN PATIENTS UNDERGOING GENERAL ANESTHESIA IN DR. HASAN SADIKIN HOSPITALS BANDUNG

Post Operative Sore Throat (POST) is a Symptoms often experienced by patients after surgery under general anesthesia and endotracheal intubation. The incidence of POST is between 21-71.8. POST causes discomfort to patients which can prolong hospital stay. In this study, a comparison was made between MgSO4 nebulization 20 mg/ KgBW and ketamine nebulization 0.5 mg/ KgBW on the incidence of POST after general anesthesia at RSUP Dr. Hasan Sadikin Bandung . This study used a double blind randomized controlled trial design. 72 elective surgery patients were randomly divided into 2 groups. Group M was given MgSO4 nebulization while group K was given ketamine nebulization. Nebulization was performed 15 minutes before induction of general anesthesia. POST monitoring was performed at 1, 4 and 24 hours after extubation using POST scoring. In this study, it was found that the incidence of POST in group M, namely the 1st hour of 13.5, the 4th hour of 10.8 and the 24th hour of 0 was lower than that in group K, namely the 1st hour of 28.5, the 4th hour of 32.5 and the 24th hour of 0. Statistical test results p0.033 showed a significant difference in group M better than group K at the 4th hour in reducing the incidence of POST. MgSO4 and ketamine have antinociceptive and anti-inflammatory effects with the mechanism of action of blocking NMDA receptors that inhibit calcium influx that avoids central sensitization. MgSO4 also has a muscle relaxing effect and reduces airway mucus secretion, thus reducing the risk of trauma during endotracheal intubation. The conclusion of this study is that MgSO4 nebulization shows a lower incidence of POST than ketamine nebulization.

Galectin-3 impairs calcium transients and β-cell function

In diabetes, macrophages and inflammation are increased in the islets, along with β-cell dysfunction. Here, we demonstrate that galectin-3 (Gal3), mainly produced and secreted by macrophages, is elevated in islets from both high-fat diet (HFD)-fed and diabetic db/db mice. Gal3 acutely reduces glucose-stimulated insulin secretion (GSIS) in β-cell lines and primary islets in mice and humans. Importantly, Gal3 binds to calcium voltage-gated channel auxiliary subunit gamma 1 (CACNG1) and inhibits calcium influx via the cytomembrane and subsequent GSIS. β-Cell CACNG1 deficiency phenocopies Gal3 treatment. Inhibition of Gal3 through either genetic or pharmacologic loss of function improves GSIS and glucose homeostasis in both HFD-fed and db/db mice. All animal findings are applicable to male mice. Here we show a role of Gal3 in pancreatic β-cell dysfunction, and Gal3 could be a therapeutic target for the treatment of type 2 diabetes.

Effects of Chinese ginger Alpinia officinarum-based phytopharmaceutical compound on gastric motility

BackgroundAlpinia officinarum, better known as Chinese ginger, is used for the treatment of gastrointestinal disorders in traditional Chinese and Ayurveda medicine. However, scientific evidence of an effect on gastrointestinal motility is missing. PurposeThis study aimed to estimate the effect of a phytopharmaceutical compound (PPC) based on Alpinia officinarum on gastric motility in vitro in order to assess its potential use in the treatment of functional dyspepsia. Study DesignWe investigated to which extent the compound affects the activity of gastric smooth muscle and we identified involved mediating cellular mechanisms. MethodsBasal and stimulated muscle activity were recorded from muscle-myenteric plexus preparations from three different regions of the guinea pig stomach before and after incubation with different concentrations of the PPC in an organ bath setup. Effect-mediating mechanisms were investigated by pharmacology. ResultsPPC exhibited a smooth muscle relaxation in a concentration-dependent manner. This was region- and muscle type specific as it was greater in fundus and corpus than in antrum and more pronounced in the circular than in the longitudinal muscle layer. The effect was primarily myogenic and at least partly mediated by TRPA1. ConclusionThe investigated PPC reduces basal tone particularly of fundus smooth muscle in a non-neuronal manner by inhibiting calcium influx via TRPA1, without affecting antral motility. Based on these data, Alpinia officinarum could be considered for phytopharmaceutical treatment of functional dyspepsia.

Sakuranetin as a Potential Regulator of Blood Pressure in Spontaneously Hypertensive Rats by Promoting Vasorelaxation through Calcium Channel Blockade.

Natural compounds, known for diverse pharmacological properties, have attracted attention as potential sources for hypertension treatment. Previous studies have revealed the hypotensive effect and vascular relaxation of prunetin, a natural compound derived from Prunus yedoensis. However, the potential blood pressure-lowering and vasorelaxant effects of sakuranetin, another representative compound found in plants belonging to the genus Prunus, have remained unexplored. We aimed to fill this gap by investigating the hypotensive and vasorelaxant effects of sakuranetin in rats. Results indicated that sakuranetin, particularly in the sakuranetin 20 mg/kg group, led to significant reductions in systolic blood pressure (SBP) and diastolic blood pressure (DBP) by -14.53 ± 5.64% and -19.83 ± 6.56% at 4 h after administration. In the sakuranetin 50 mg/kg group, the SBP and DBP decreased by -13.27 ± 6.86% and -16.62 ± 10.01% at 2 h and by -21.61 ± 4.49% and -30.45 ± 5.21% at 4 h after administration. In addition, we identified the vasorelaxant effects of sakuranetin, attributing its mechanisms to the inhibition of calcium influx and the modulation of angiotensin II. Considering its hypotensive and vasorelaxant effects, sakuranetin could potentially serve as an antihypertensive agent. However, further research is required to evaluate the safety and long-term efficacy.

Pathological effects of pregabalin toxicity in rats

Abstract Background: Pregabalin (PGB) is one kind of gabapentinoid. The main mechanism of action is binding at the alpha-2-delta site, which inhibits calcium influx in response to depolarization at nerve terminals and, in turn, suppresses the release of glutamate, noradrenaline, and substance P. Objectives: This study aimed to study the pathological effects of PGB toxicity on brain and liver of rats. Materials and Methods: We chose 20 mature albino rats, both sexes, averaging 220 g in weight; these were divided into two groups (10 rats per group): the toxic group and the control group. Tablets of Lyrica (Pfizer) may be purchased over the counter. The 300 mg of PGB in each tablet was dissolved in 3 mL of 0.9% normal saline. The dosage was determined using the maximum lethal oral dose in rats (5000 mg/kg) (Pfizer, 2017). Based on the rats’ weights, a toxic dosage of 1000 mg of PGB was determined and reconstituted in normal saline (0.9% in 3 mL). Each animal in the acute toxicity group received a single dose of the produced medication orally. After 24 h, all of the animals in both groups were euthanized. The brain and liver were quickly dissected and removed, washed in saline solution, and then processed for histopathological study. Results: Focal regions of hemorrhage and congestion were seen in H&amp;E-stained sections from the acute toxicity group, and most pyramidal cells were degraded, pyknotic, and exhibited karyolysis. Cerebellar cortical layers were preserved; however, Purkinje cells were destroyed in the acute toxicity group, which also exhibited an increase in pyknotic cells, hemorrhage, vascular congestion, and localized loss of tissue. Hemorrhages, congestion of portal region blood vessels, and central veins and hepatic sinusoids were some of the most notable pathological abnormalities seen in the livers of those using PGB. Hepatocytes show nuclear pyknosis and a homogeneously acidophilic cytoplasm as a result of severe degenerative alterations, such as vacuolar degeneration and severe necrotic changes. Conclusion: PGB can cause pathological lesions in the brain and liver at a single toxic dose.

A peptidomimetic modulator of the CaV2.2 N-type calcium channel for chronic pain.

Transmembrane Cav2.2 (N-type) voltage-gated calcium channels are genetically and pharmacologically validated, clinically relevant pain targets. Clinical block of Cav2.2 (e.g., with Prialt/Ziconotide) or indirect modulation [e.g., with gabapentinoids such as Gabapentin (GBP)] mitigates chronic pain but is encumbered by side effects and abuse liability. The cytosolic auxiliary subunit collapsin response mediator protein 2 (CRMP2) targets Cav2.2 to the sensory neuron membrane and regulates their function via an intrinsically disordered motif. A CRMP2-derived peptide (CBD3) uncouples the Cav2.2-CRMP2 interaction to inhibit calcium influx, transmitter release, and pain. We developed and applied a molecular dynamics approach to identify the A1R2 dipeptide in CBD3 as the anchoring Cav2.2 motif and designed pharmacophore models to screen 27 million compounds on the open-access server ZincPharmer. Of 200 curated hits, 77 compounds were assessed using depolarization-evoked calcium influx in rat dorsal root ganglion neurons. Nine small molecules were tested electrophysiologically, while one (CBD3063) was also evaluated biochemically and behaviorally. CBD3063 uncoupled Cav2.2 from CRMP2, reduced membrane Cav2.2 expression and Ca2+ currents, decreased neurotransmission, reduced fiber photometry-based calcium responses in response to mechanical stimulation, and reversed neuropathic and inflammatory pain across sexes in two different species without changes in sensory, sedative, depressive, and cognitive behaviors. CBD3063 is a selective, first-in-class, CRMP2-based peptidomimetic small molecule, which allosterically regulates Cav2.2 to achieve analgesia and pain relief without negative side effect profiles. In summary, CBD3063 could potentially be a more effective alternative to GBP for pain relief.

Ocular Pharmacology and Toxicology of TRPV1 Antagonist SAF312 (Libvatrep).

To evaluate the pharmacology and toxicology of SAF312, a transient receptor potential vanilloid 1 (TRPV1) antagonist. TRPV1 expression in human ocular tissues was evaluated with immunohistochemistry. Inhibition of calcium influx in Chinese hamster ovary (CHO) cells expressing human TRPV1 (hTRPV1) and selectivity of SAF312 were assessed by a fluorescent imaging plate reader assay. Ocular tissue and plasma pharmacokinetics (PK) were assessed following a single topical ocular dose of SAF312 (0.5%, 1.0%, 1.5%, 2.5%) in rabbits. Safety and tolerability of SAF312 were evaluated in rabbits and dogs. Effects of SAF312 on corneal wound healing after photorefractive keratectomy (PRK) surgery were assessed in rabbits. TRPV1 expression was noted in human cornea and conjunctiva. SAF312 inhibited calcium influx in CHO-hTRPV1 cells induced by pH 5.5 (2-[N-morpholino] ethanesulfonic acid), N-arachidonoylethanolamine, capsaicin, and N-arachidonoyl dopamine, with IC50 values of 5, 10, 12, and 27 nM, respectively, and inhibition appeared noncompetitive. SAF312 demonstrated high selectivity for TRPV1 (>149-fold) over other TRP channels. PK analysis showed highest concentrations of SAF312 in cornea and conjunctiva. SAF312 was found to be safe and well tolerated in rabbits and dogs up to the highest feasible concentration of 2.5%. No delay in wound healing after PRK was observed. SAF312 is a potent, selective, and noncompetitive antagonist of hTRPV1 with an acceptable preclinical safety profile for use in future clinical trials. SAF312, which was safe and well tolerated without causing delay in wound healing after PRK in rabbits, may be a potential therapeutic agent for ocular surface pain.

Vasodilator activity of Poecilotheria ornata venom involves activation of the NO/cGMP pathway and inhibition of calcium influx to vascular smooth muscle cells.

Tarantula venoms may be a natural source of new vasodilator components useful in pharmacological research. Moreover, biological function data of the venoms are important to enhance the knowledge about the biodiversity and evolution of these species. The present study aims to describe the vasodilatory activity induced by the venom of Poecilotheria ornata on isolated rat aortic rings. This venom induced a vasodilator activity that was significantly reduced after incubation with L-NAME or ODQ. Measurements of nitrite concentrations on rat aorta homogenates showed that the venom significantly increased the basal levels. Moreover, the venom attenuates the contraction induced by calcium. These results suggest that P. ornata venom contains a mixture of vasodilator components that act through the activation of the nitric oxide/cGMP pathway, as well as, through an endothelium-independent mechanism that involves the calcium influx into vascular smooth muscle cells.

Abstract PR010: First-in-class TRPV6 inhibitors for the treatment of prostate cancer

Abstract TRPV6, a calcium channel, is an oncochannel that is overexpressed in epithelial cancers including prostate cancer. We have developed a novel series of first-in-class small molecule TRPV6 inhibitors for the treatment of advanced prostate cancer. Our preclinical candidate QED-203 has been shown to inhibit calcium influx at low nM potency in TRPV6 cellular assays (FLIPR, electrophysiology), and has low nM potency in a TRPV6-driven NFAT assay (NFAT luciferase reporter). Inhibition of TRPV6 with our TRPV6 inhibitors or by siRNA knockdown causes changes in genes related to NFAT and WNT signalling, ER stress and the cell cycle (RNAseq and qPCR analysis), and causes cell cycle arrest, decreased proliferation and apoptosis in prostate cancer cells (demonstrated via imaging and FACS analysis). Importantly, QED-203 demonstrates potency in enzalutamide resistant cell lines (in vitro proliferation inhibition) and has superior potency over enzalutamide and darolutamide [androgen receptor targeting agents (ARTA)] in prostate cancer cell lines with clinically relevant AR mutations or the ARV7 splice variant. The ARV7 variant in particular is represented in a significant portion of patients no longer responding to ARTA SoC interventions. QED-203 has high bioavailability with a pharmacokinetic profile amenable to once-a-day oral dosing. QED-203 is well tolerated in rodents and has similar in vivo efficacy (tumour growth and PSA inhibition) to enzalutamide in a castrated LNCaP mouse model of prostate cancer. We have also shown QED-203 target engagement in rodents by demonstrating a change in calcium levels in the urine, and have observed changes in genes consistent with a TRPV6-specific mode of action in xenograft tumours (demonstrated via qPCR). QED-203 targets a novel, non-hormonal mechanism in prostate cancer, and could be used to treat prostate cancer patients who have developed resistance to AR therapies, where there is a large unmet need. Citation Format: Kimberley Beaumont, Rebecca Pouwer, Raphael Rahmani, Matthew McLachlan, Claire Levrier, Rebecca Farrow, Mei Yeh, Therese Johnson, Malika Kumarasiri, Hasanthi Wijesekera, Grant Stuchbury, Terrie-Anne Cock, Andrew Harvey, Gregory Monteith, Brian Dymock. First-in-class TRPV6 inhibitors for the treatment of prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr PR010.

Vasorelaxant Properties of Moringa oleiferaleaf extract: An in-vitro study on mice blood vessels

Background: Moringa oleifera is a plant rich in bioactive compounds, including flavonoids, polyphenols, and alkaloids, which have been shown to have numerous health benefits. In recent years, several studies have investigated the potential of Moringa oleifera in the treatment ofhypertension and diseases of vascular dysfunction. Aim: To observe the effect of M. oleifera leaf extract on contractility of aortic tissue of mice, in vitro. Methods: An animal-based in-vitro experimental study was carried out in Pharmacology Department of CMH Lahore Medical College and Institute of Dentistry from February 2022 to March 2023. Mice were euthanized with ketamine. Aortic tissue was dissected and mounted in the organ bath. For evaluation of vasodilatory effect, cumulative dose-response curve of M. oleifera leaf extract was obtained using increasing doses of the extract against1 µM adrenaline and 80 mM KCl, in group I and group II, respectively.Response was obtained through Powe rLab (AD Instruments, Australia). Results: A consistent decrease was recorded against adrenaline and KCl-induced contractions in aorta, resulting in vasodilation. IC50 was 0.068 mg and 0.161 mg, in group I and II, respectively. Difference between IC50 of both groups was statistically significant (p-value = 0.0008). Conclusion: Leaf extract of Moringa oleifera exerts smooth muscle relaxant effectin blood vessels, most likely through inhibition of calcium influx, mediated through Gq-signalling pathway as well as depolarization-dependent voltage operated calcium channels. Keywords: Moringa oleifera, dose-response relationship, trachea, aorta, calcium channels

A New Lignan from Annona squamosa L. (Annonaceae) Demonstrates Vasorelaxant Effects In Vitro.

Esquamosan, a new furofuran lignan, has been isolated by bio-guided assays from the methanolic extract of the leaves of Annona squamosa L., and its structure was elucidated by spectroscopic methods. Esquamosan inhibited the rat aortic ring contraction evoked by phenylephrine in a concentration-dependent manner and showed an inhibitory effect on vasocontraction of the depolarized aorta with high-concentration potassium. The vasorelaxant effect by esquamosan could be attributed mainly to the inhibition of calcium influx from extracellular space through voltage-dependent calcium channels or receptor-operated Ca2+ channels and also partly mediated through the increased release of NO from endothelial cells. The ability of esquamosan to modify the vascular reactivity of rat aortic rings incubated with high glucose (D-glucose 55 mM) was then evaluated, and this furofuran lignan reverted the endothelium-dependent impairment effect of high glucose in rat aortic rings. The antioxidant capacity of esquamosan was assessed using DPPH and FRAP assays. Esquamosan exhibited a similar antioxidant capacity compared to ascorbic acid, which was used as a positive control. In conclusion, this lignan showed a vasorelaxant effect, free radical scavenging capacity, and potential reductive power, suggesting its potential beneficial use to treat complex cardiometabolic diseases due to free radical-mediated diseases and its calcium antagonist effect.

Effects of Shiwei Longdanhua formula on LPS induced airway mucus hypersecretion, cough hypersensitivity, oxidative stress and pulmonary inflammation

Shiwei Longdanhua Granule (SWLDH) is a classic Tibetan medicine (TM) ranking in the top 20 Chinese patent medicines in prescription rate to treat respiratory diseases like pneumonia, acute and chronic tracheobronchitis, acute exacerbation of COPD and bronchial asthma in solution of inflammation, cough and phlegm obstruction in clinical practice. However, its systematic pharmacological mechanisms have not been elucidated yet. Here, we studied the therapeutic efficacy of SWLDH in treatment of acute respiratory diseases in BALB/c mice by comprehensive analysis of airway inflammation, oxidative stress, mucus hypersecretion, cough hypersensitivities and indicators associated with the development of chronic diseases. Our results show that SWLDH might exhibit its inhibitory effects on pulmonary inflammation by interference with arachidonic acid (AA) metabolism pathways. Oxidative stress that highly related to the degree of tissue injury could be alleviated by enhancing the reductive activities of glutathione redox system, thioredoxin system and the catalytic activities of catalase and superoxide dismutase (SOD) after SWLDH treatment. In addition, SWLDH could significantly abrogate the mucus hypersecretion induced bronchiole obstruction by inactivate the globlet cells and decrease the secretion of gel-forming mucins (MUC5AC and MUC5B) under pathological condition, demonstrating its mucoactive potency. SWLDH also showed reversed effects on the release of neuropeptides that are responsible for airway sensory hypersensitivity. Simultaneously observed inhibition of calcium influx, reduction in in vivo biosynthesis of acetylcholine and the recovery of the content of cyclic adenosine monophosphate (cAMP) might collaboratively contribute to cause airway smooth muscle cells (ASMCs) relexation. These findings indicated that SWLDH might exhibited antitussive potency via suppression of the urge to cough and ASMCs contraction. Moreover, SWLDH might affect airway remodeling. We found SWLDH could retard the elevation of TGF-β1 and α-SMA, which are important indicators for hyperplasia and contraction during the progression of the chronic airway inflammatory diseases like COPD and asthma.

Taurine and Astrocytes: A Homeostatic and Neuroprotective Relationship (P9-4.003)

<h3>Objective:</h3> This review provides an overview of the significant relationship between taurine and astrocytes, as well as its homeostatic and neuroprotective role in the nervous system. <h3>Background:</h3> Taurine is considered the most abundant free amino acid present in the brain, with multiple functions including antioxidative and anti-inflammatory effects, gene transcription regulation and maintenance of cellular energy processes, among others. Despite previous research regarding taurine’s effect in different tissues, its role in the nervous system, particularly the functional relationship with astrocytes, remains to be further elucidated. Understanding taurine’s effect in astrocytes can contribute to a better insight of its physiological effects, which could potentially be used to ameliorate the course of several nervous system pathologies. <h3>Design/Methods:</h3> We conducted a literature review concerning the relationship between taurine and astrocytes and its neuroprotective effects in the nervous system. We searched in different databases using search criteria such as “taurine AND neuroprotection”, “taurine AND glia”, “taurine AND brain”, “taurine AND astrocyte”, among others. There was no chosen range for year of publication. <h3>Results:</h3> There is a metabolic coupling between astrocytes and neurons in which astrocytes provide neurons with hypotaurine as a substrate for taurine production. Taurinergic gliotransmission can enhance post-synaptic neuronal inhibition mediated by GABA-AR and GlyR, thus protecting the brain towards glutamate-induced excitotoxicity. Furthermore, taurine prevents mitochondrial stress, and activation of apoptotic pathways by inhibiting calcium influx from VGCC. <h3>Conclusions:</h3> There is compelling evidence of taurine’s neuroprotective and homeostatic effects in the CNS. The metabolic coupling for taurine synthesis and degradation between astrocytes and neurons, evidence neuronal dependence on astrocytes for the adequate functioning of the mentioned effects in the brain. Therefore, the astrocyte’s role in taurine-induced neuroprotective functions should be considered as a promising therapeutic target in the management of several neurodegenerative and neuropsychiatric diseases in the near future. <b>Disclosure:</b> Ms. Ramirez Guerrero has nothing to disclose. Mr. Guardo-Maya has received personal compensation in the range of $10,000-$49,999 for serving as a Pre-doctoral fellow with NIH. Mr. Medina-Rincón has nothing to disclose. Dr. Cabezas has nothing to disclose. The institution of an immediate family member of Dr. Gonzalez has received research support from Minciencias. Dr. Gonzalez has received publishing royalties from a publication relating to health care. Dr. Gonzalez has received personal compensation in the range of $0-$499 for serving as a Speaker with Mitotherapies.

DNA tension assays reveal that force-dependent integrin activation regulates neurite outgrowth in primary cortical neurons

Biomechanical inputs are ubiquitously present in biological systems and are known to regulate various cell functions. In particular, neural cell development is sensitive to mechanical regulation, as these cells reside in one of the softest microenvironments in the body. To fully characterize and comprehend how mechanical force modulates early neuronal processes, we prepared substrates functionalized with DNA probes displaying integrin ligands, including cRGD and laminin, to quantify integrin-mediated molecular tension during neurite initiation in primary cortical neurons. Our live-cell imaging analysis reveals that integrin-mediated tension force is highly dynamic and distributed across the cell body, with the overall tension signal gradually increasing during neurite outgrowth. Notably, we detected a consistent level of mechanical force (amplitude = 4.7–12 piconewtons, pN) for cell integrin-ligand interactions. Further quantifications reveal that neurons exhibit faster cell spreading and neurite outgrowth upon interacting with ligands functionalized with 4.7 pN relative to 12 pN probes. These findings indicate that the magnitude of integrin-mediated mechanical feedback regulates neuronal activity during early neuritogenesis. Additionally, we observed that mechanical tension is correlated with calcium signaling, since inhibiting calcium influx substantially reduced mechanical tension. Thus, our findings support that the magnitude of integrin-mediated mechanical feedback regulates neuronal activity during early neuritogenesis and that mechanical force is an essential element complementing well-known biochemical regulatory mechanisms orchestrating the integrin activation machinery and controlled neurite outgrowth in cortical neurons.