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Neuroscience drug discovery is challenged by the brain’s structural and cell-type complexity, which is difficult to model in cellular systems compatible with high-throughput screening methods. Calcium oscillation assays, that harness neurons’ intrinsic capability to develop functional neural networks in cell culture, are currently the closest cellular models with a relevant functional endpoint to model human neuronal circuitry in a dish. Here we further developed this useful assay towards scalable drug discovery applications. We show the importance of defined neuron-to-astrocyte ratios for optimal cellular distribution and surface adherence in HTS-compatible cell culture vessels and how the cell type ratios affect network firing patterns. Increasing the neuron density resulted in decreased network spike frequencies, but increased network spike amplitudes. We identified DAPT, a molecule previously shown to promote neuronal maturation and synapse formation, as a negative regulator of astrocyte viability. Furthermore, inclusion of GABAergic neurons in the cocultures increased the network spike frequency while reducing network spike amplitudes. The GABAA receptor antagonist bicuculline did not affect network spike frequency, but increased network spike amplitudes. In order to access local field activity in an automated and scalable calcium imaging environment, we developed a pixel-based analysis for plate reader data. This method revealed that the effect of GABAergic neurons and bicuculline was restricted to local field calcium activity that coincided with synchronized network spikes. Our observations are consistent with previous findings suggesting that the presence of GABAergic neurons decreases synchronization and network spike participation of local neuronal activity, thus potentially echoing aspects of GABA action in vivo, and dysregulation thereof in pathological conditions.

The present guidelines recommend dual antiplatelet therapy (DAPT) for 6 to 12 months after percutaneous coronary intervention (PCI), with recent trials assessing the safety and efficacy of shortening DAPT duration to ≤3 months. A systematic search of PubMed, Scopus, and Cochrane Central databases identified studies comparing short DAPT, followed by P2Y12i monotherapy (78% ticagrelor) versus standard 12-month DAPT in patients who underwent PCI with a drug-eluting stent. A total of 9 randomized controlled trials, including 42,770 patients (short DAPT n = 21,370, 49.96%), of whom 28,307 (66.18%) presented with acute coronary syndrome (ACS). Short DAPT significantly reduced net adverse clinical events (NACEs) (risk ratio [RR] 0.78, 95% confidence interval [CI] 0.67 to 0.91, p = 0.001, I2 = 62%), major bleeding (RR 0.54, 95% CI 0.39 to 0.73, p <0.001, I2 = 63%), and any bleeding (RR 0.55, 95% CI 0.43 to 0.72, p <0.001, I2 = 77%) at 12 months compared with 1-year DAPT. No significant differences were observed in major adverse cardiovascular/cerebrovascular events, myocardial infarction, stroke, stent thrombosis, mortality, or revascularization. Ticagrelor monotherapy after short DAPT further reduced major adverse cardiovascular/cerebrovascular events (RR 0.85, 95% CI 0.73 to 0.99, p = 0.040, I² = 22%), NACE (RR 0.74, 95% CI 0.61 to 0.89, p = 0.001, I² = 68%), and major bleeding (RR 0.56, 95% CI 0.40 to 0.78, p <0.001, I² = 71%) compared with 1-year DAPT; however, the test for subgroup interaction (Pinteraction >0.05) for clopidogrel subgroup was not significant. P2Y12i monotherapy reduced the risk of NACEs (RR 0.77, 95%CI 0.66 to 0.90, p = 0.001, I2 = 52%, Pinteraction = 0.58) and major bleeding (RR 0.44, 95%CI 0.35 to 0.55, p <0.001, I2 = 0%, Pinteraction <0.01) in the ACS cohort but not in the chronic coronary syndrome cohort. In conclusion, short DAPT for ≤3 months followed by P2Y12i monotherapy (particularly, ticagrelor) was associated with decreased NACEs and bleeding without differences in other outcomes and should be considered a favorable option in patients with either ACS or chronic coronary syndrome after PCI with a drug-eluting stent.

Abstract Aims To characterize and follow patients with ST-segment elevation myocardial infarction (STEMI) at high bleeding risk (HBR) according to the predicting bleeding complications in patients undergoing stent implantation and subsequent dual antiplatelet therapy (PRECISE-DAPT) score, and to examine the use of P2Y12 inhibitors and the subsequent risk of major adverse cardiovascular events (MACE) and bleeding. Methods and results This single-centre cohort study included 6179 consecutive STEMI patients who underwent percutaneous coronary intervention (PCI) at Copenhagen University Hospital, Rigshospitalet, between 2009 and 2016. Individual linkage to nationwide registries was conducted to obtain information on diagnoses, claimed drugs, and vital status. Of the 5532 (89.5%) patients with available PRECISE-DAPT scores, 33.0% were at HBR and more often elderly and female with more comorbidities than non-HBR patients. One-year cumulative incidence rates per 100 person-years were 8.7 and 2.1 for major bleeding and 36.8 and 8.3 for MACE in HBR and non-HBR patients, respectively. Among the 4749 (85.8%) patients who survived and collected a P2Y12 inhibitor ≤7 days from discharge, 68.2% of HBR patients were treated with ticagrelor or prasugrel and 31.8% with clopidogrel, while 18.2% non-HBR patients were treated with clopidogrel. Adherence was high for all (>75% days coverage). The risk of MACE was lower in ticagrelor- and prasugrel-treated patients than in clopidogrel-treated patients without differences in major bleeding. Conclusion One-third of PCI-treated all-comer patients with STEMI were at HBR according to the PRECISE-DAPT score and were more often treated with potent P2Y12 inhibitors instead of clopidogrel. Thus, ischaemic risk may be weighted over bleeding risk in STEMI patients at HBR. Graphical Abstract Graphical Abstract PCI-treated STEMI patients at high bleeding risk and the use of P2Y12 inhibitors according to bleeding risk. PCI, percutaneous coronary intervention; PRECISE-DAPT, predicting bleeding complications in patients undergoing stent implantation and subsequent dual antiplatelet therapy; STEMI, ST-segment elevation myocardial infarction.

IntroductionCardiovascular diseases (CVD) accounted for an alarming 17.9 million deaths globally in 2019. Acute coronary syndrome (ACS) is oftenthe most common initial presentation of CVD. ACS encompasses of ST-elevation Myocardial Infarction (STEMI), Non-ST elevationMyocardial Infarction (NSTEMI) and unstable angina. It accounts for over 100,000 admissions to hospitals annually(1) and has a mortalityrate of around 15% pre-hospitalisation and 10% during a hospital admission.(2) European Society of Cardiology (ESC) guidelinesrecommend use of Dual Antiplatelet Therapy (DAPT) which comprises of aspirin and a P2Y12 as treatment for ACS. DAPT is estimated toincrease the risk of GI bleed by 1.2–2.4%. To reduce this risk of bleeding, ESC guideline until recently recommended Proton PumpInhibitor (PPI) in all patients receiving DAPT.(3) The aim of this quality improvement project was to assess current compliance rates andrecommend effective measures to improve compliance rates to 100%.MethodsData on all patients who presented to our cardiac centre with either a STEMI or NSTEMI over a 6-month period was collected. Weexcluded patients transferred from nearby district general hospitals for Percutaneous Coronary Intervention (PCI). We analysed electronicrecords, prescription charts and discharge summaries to assess the compliance rates on the use of PPI in patients receiving DAPT andincidence rates of GI bleed in this cohort of patients.ResultsThere were 543 patients who attended our hospital with either a STEMI or NSTEMI over the 6-months. STEMI was the cause of ACS in44.4% of our patients. Clinicians initiated DAPT in 486 patients. 98.1% of all patients had PPI prescribed alongside DAPT. Lansoprazolewas the most common PPI prescribed in 92% of patient’s, 5% omeprazole, 2% famotidine and 1% esomeprazole. There were threepatients on DAPT who had UGI bleed and two deaths as a result of this. All patients who had an incidence of UGI bleed had not receivedPPI therapy. There was no clear documentation of the risk of bleeding using scores such as CRUSADE or ACUITY in patients who did notreceive PPI therapy. There was a change in ESC guideline in 2023 that now recommends PPI use only in patients with a high risk of bleeding.(4)ConclusionThe incidence of UGI bleed was higher in patients who were not on PPI therapy in comparison to those who were. There was poordocumentation on bleeding risk in patients not started on PPI therapy. To improve this, we led a teaching session to doctors andpharmacists to raise awareness of the current ACS protocol on our prescribing system and importance of using risk stratification tools anddocumentation of scores in patient notes to ensure we are prescribing in line with the latest ESC guidelines.Conflict of Interestnil

Human epithelial stem cells (ESCs) are characterized by long‐term regenerative properties, much dependent on the tissue of origin and varying during their lifespan. We analysed such variables in cultures of ESCs isolated from the skin, conjunctiva, limbus and oral mucosa of healthy donors and patients affected by ectrodactyly‐ectodermal dysplasia‐clefting syndrome, a rare genetic disorder caused by mutations in the p63 gene. We cultured cells until exhaustion in the presence or in the absence of DAPT (γ‐secretase inhibitor; N‐[N‐(3, 5‐difluorophenacetyl)‐L‐alanyl]‐S‐phenylglycine T‐butyl ester). All cells were able to differentiate in vitro but exhibited variable self‐renewal potential. In particular, cells carrying p63 mutations stopped prematurely, compared with controls. Importantly, administration of DAPT significantly extended the replicative properties of all stem cells under examination. RNA sequencing analysis revealed that distinct sets of genes were up‐ or down‐regulated during their lifetime, thus allowing to identify druggable gene networks and off‐the‐shelf compounds potentially dealing with epithelial stem cell senescence. These data will expand our knowledge on the genetic bases of senescence and potentially pave the way to the pharmacological modulation of ageing in epithelial stem cells.

Curcumin, as a high effect and low toxicity anti-cancer drug and photosensitiser, has synergistic and complementary effects with photodynamic therapy (PDT). However, due to its unclear mechanism, PDT's application and efficacy were limited. Notch signaling pathway, which is highly correlates with carcinogenesis and development of cervical cancer, could be a potential therapeutic targets to improve the effectiveness of PDT. Therefore, in this study, we explored the effects of Notch signaling pathway in cervical cancer by curcumin mediated PDT with/without Notch receptor blocker (DAPT), and hope to elucidate its mechanism. Firstly, the effect on the proliferation of cervical cancer Me180 cells were detected with MTT assay, and apoptosis were detected with Annexin V-FITC/PI combined with flow cytometry. Secondly, after establishment of nude mice model, dividing the experimental animals into model group, curcumin PDT group, simple DAPT group, and curcumin-PDT+DAPT group, and analyzing tumor volume changes as well as HE staining in each group. mRNA and protein expression of gene Notch-1 and its downstream NF-κB and VEGF were observed with RT-PCR, immunohistochemical staining and Western-blot with/without inhibition of Notch signaling pathway by DAPT, both in vivo and in vitro experiments. We found both DAPT and curcumin-PDT can inhibit the proliferation and induce apoptosis of cervical cancer cell. The two have synergistic effect in vitro and in vivo. This effect can effectively block the conduction of Notch signaling pathway, which is associated with down-regulation of the expression of Notch1 and NF-κB. Notch signaling pathway could be one of the targets of curcumin-PDT photodynamic therapy.

Aims The PRECISE-DAPT (Predicting Bleeding Complication in Patients Undergoing Stent Implantation and Subsequent Dual Antiplatelet Therapy) score identifies patients at high risk of bleeding complications following percutaneous coronary intervention (PCI). International guidelines recommend the PRECISE-DAPT score to identify patients at high risk for bleeding, who may benefit from shortened dual antiplatelet therapy. The association of the PRECISE-DAPT score with ischaemic outcomes remains unclear. We performed a meta-analysis investigating the association between a high PRECISE-DAPT score and ischaemic outcomes. Methods and results A comprehensive literature search was conducted on articles published between 11 March 2017 and 5 June 2021. Two reviewers independently screened articles for inclusion using pre-defined criteria. The outcome measures extracted included composite ischaemic events, major bleeding events, and all-cause mortality. A random effects model was applied to obtain combined risk estimates for outcomes. From 12 included studies, there were 39 459 patients with PRECISE-DAPT <25 and 14 761 patients with PRECISE-DAPT ≥25. PRECISE-DAPT score ≥25 was associated with increased risk of composite ischaemic events [odds ratio (OR) 2.16; 95% confidence interval (CI) 1.77–2.65], myocardial infarction (OR 2.06; 95% CI 1.38–3.08), and ischaemic stroke (OR 2.90; 95% CI 1.76–4.78). Patients with a PRECISE-DAPT score ≥25 had increased risk of major bleeding (OR 3.62; 95% CI 2.62–4.99). Patients with a PRECISE-DAPT score ≥25 had higher risk of all-cause mortality (OR 5.83; 95% CI 5.37–6.33). Conclusion Patients with a PRECISE-DAPT score ≥25 are at increased risk for ischaemic events, bleeding, and all-cause mortality. Prospective evaluation of a PRECISE-DAPT guided approach to antiplatelet therapy is required to demonstrate benefit in this high-risk population.

Older patients have been remarkably underrepresented in bleeding risk cohorts. Thus, the PRECISE-DAPT (Derivation and validation of the predicting bleeding complications in patients undergoing stent implantation and subsequent dual antiplatelet therapy) and Academic Research Consortium for High Bleeding Risk (ARC-HBR) scores are not validated in older adults. Therefore, we sought to evaluate the PRECISE-DAPT and ARC-HBR scores in an exclusively older population and assess the prognostic value of a truly simplified clinical evaluation (SCE), consisting of only 3 binary clinical variables (hemoglobin <11 g/100 ml, previous bleeding, and anticipated use of anticoagulants). This is a retrospective analysis of the prospective single-center older-HCD registry. Consecutive patients aged ≥75 years who underwent percutaneous coronary intervention from 2012 to 2019 were included. The primary end point was postdischarge bleeding at 12 months of follow-up, defined according to the Bleeding Academic Research Consortium 3 or 5 criteria. A total of 693 patients with a mean age of 81 (±4.4) years were included in the study and 60 patients (6.8%) met the primary end point. The PRECISE-DAPT and ARC-HBR scores did not significantly predict postdischarge bleeding in the Cox regression models (hazard ratio 1.65 [0.78 to 3.42] and 1.46 [0.72 to 4.24], respectively), whereas the SCE outperformed both scores (hazard ratio 2.47, 1.34 to 4.49). All 3 scores exhibited a moderate discriminatory potential, as determined by a receiver-operating characteristic curve analysis (areas under the curve 0.601, 0.621, and 0.616, respectively), with no significant differences between them. The SCE showed an Integrated Discrimination Improvement of 0.25, p = 0.02 (SCE vs ARC-HBR) and 0.24, p = 0.01 (SCE vs PRECISE-DAPT), with an Net Reclassification Improvement of 6.54%, p = 0.37 and 7.12%, p = 0.43, respectively. In conclusion, the PRECISE-DAPT score and ARC-HBR criteria showed insufficient predictive value in older adults. A truly SCE consisting of 3 easily accessible variables not only provides equal discriminatory potential but also demonstrates superior predictive value, as determined by Cox regression models. This makes it a highly appealing tool for risk stratification, pending its evaluation in larger prospective studies.

Hypertrophic scar (HS) formation is a cutaneous fibroproliferative disease that occurs after skin injuries and results in severe functional and esthetic disability. To date, few drugs have shown satisfactory outcomes for the treatment of HS formation. Transforming growth factor-beta (TGF-β)/Notch interaction via small mothers against decapentaplegic 3 (Smad3) could facilitate HS formation; therefore, targeting TGF-β/ Notch interaction via Smad3 is a potential therapeutic strategy to attenuate HS formation. In addition, optic atrophy 1 (OPA1)-mediated mitochondrial fusion contributes to fibroblast proliferation, and TGF-β/Smad3 axis and the Notch1 pathway facilitate OPA1-mediated mitochondrial fusion. Thus, the aim of this study was to investigate whether drugs targeting TGF-β/Notch interaction via Smad3 suppressed fibroblast proliferation to attenuate HS formation through OPA1-mediated mitochondrial fusion. We found that the TGF-β pathway, Notch pathway, and TGF-β/Notch interaction via Smad3 were inhibited by pirfenidone, the gamma- secretase inhibitor DAPT, and SIS3 in human keloid fibroblasts (HKF) and an HS rat model, respectively. Protein interaction was detected by co-immunoprecipitation, and mitochondrial morphology was determined by electron microscopy. Our results indicated that pirfenidone, DAPT, and SIS3 suppressed the proliferation of HKFs and attenuated HS formation in the HS rat model by inhibiting TGF-β/Notch interaction via Smad3. Moreover, pirfenidone, DAPT, and SIS3 hindered OPA1-mediated mitochondrial fusion through inhibiting TGF-β/Notch interaction, thereby suppressing the proliferation of HS fibroblasts and HS formation. In summary, these findings investigating the effects of drugs targeting TGF-β/Notch interaction on HS formation might lead to novel drugs for the treatment of HS formation.