Veno-arterial extracorporeal membrane oxygenation (ECMO VA) as part of a multimodal approach for the protection of spinal cord ischemia in surgical repair of a thoracoabdominal aneurysm.

Spinal cord ischaemia leading to paraplegia or paraparesis is one of the most devastating complications of aortic surgery. The risk of ischaemia is particularly high in repairs involving both the thoracic and abdominal segments, because in these cases blood flow to the spinal arteries can be interrupted. Multimodal protocols have now been developed to reduce the incidence of this complication, and include measures such as cerebrospinal fluid (CSF) drainage, avoidance of hypotension and anaemia, systemic hypothermia, neuromonitoring, maintaining distal perfusion during proximal clamping of the aorta, and reimplantation of intercostal or lumbar arteries, whenever feasible. We describe a case in which, due to the special characteristics of the surgery, veno-arterial extracorporeal membrane oxygenation (VA ECMO) was used to maintain distal blood flow in the lumbar, inferior mesenteric, and hypogastric arteries during aortic clamping. This approach reduced the risk of spinal cord and visceral ischaemia, and also eliminated the need for thoracotomy because partial left bypass was not required.

Structure-Antitumor Activity Relationships of Aza- and Diaza-Anthracene-2,9,10-Triones and Their Partially Saturated Derivatives.

The 1,8-Diazaanthracene-2,9,10-triones, their 5,8-dihydro derivatives, and 1,8-diazaanthracene-2,7,9,10-tetraones, structurally related to the diazaquinomycin family of natural products, were synthesized in a regioselective fashion employing Diels-Alder strategies. These libraries were studied for their cytotoxicity in a variety of human cancer cell lines in order to establish structure-activity relationships. From the results obtained, we conclude that some representatives of the 1,8-diazaanthracene-2,9,10-trione framework show potent and selective cytotoxicity against solid tumors. Similar findings were made for the related 1-azaanthracene-2,9,10-trione derivatives, structurally similar to the marcanine natural products, which showed improved activity over their natural counterparts. An enantioselective protocol based on the use of a SAMP-related chiral auxiliary derived was developed for the case of chiral 5-substituted 1,8-diazaanthracene-2,9,10-triones, and showed that their cytotoxicity was not enantiospecific.

Cyclodextrin Inclusion Complexes for Improved Drug Bioavailability and Activity: Synthetic and Analytical Aspects.

Many active pharmaceutical ingredients show low oral bioavailability due to factors such as poor solubility and physical and chemical instability. The formation of inclusion complexes with cyclodextrins, as well as cyclodextrin-based polymers, nanosponges, and nanofibers, is a valuable tool to improve the oral bioavailability of many drugs. The microencapsulation process modifies key properties of the included drugs including volatility, dissolution rate, bioavailability, and bioactivity. In this context, we present relevant examples of the stabilization of labile drugs through the encapsulation in cyclodextrins. The formation of inclusion complexes with drugs belonging to class IV in the biopharmaceutical classification system as an effective solution to increase their bioavailability is also discussed. The stabilization and improvement in nutraceuticals used as food supplements, which often have low intestinal absorption due to their poor solubility, is also considered. Cyclodextrin-based nanofibers, which are polymer-free and can be generated using environmentally friendly technologies, lead to dramatic bioavailability enhancements. The synthesis of chemically modified cyclodextrins, polymers, and nanosponges based on cyclodextrins is discussed. Analytical techniques that allow the characterization and verification of the formation of true inclusion complexes are also considered, taking into account the differences in the procedures for the formation of inclusion complexes in solution and in the solid state.

Quinolinetrione-tacrine hybrids as multi-target-directed ligands against Alzheimer's disease.

Multi-target drug discovery is one of the most active fields in the search for new drugs against Alzheimer's disease (AD). This is because the complexity of AD pathological network might be adequately tackled by multi-target-directed ligands (MTDLs) aimed at modulating simultaneously multiple targets of such a network. In a continuation of our efforts to develop MTDLs for AD, we have been focusing on the molecular hybridization of the acetylcholinesterase inhibitor tacrine with the aim of expanding its anti-AD profile. Herein, we manipulated the structure of a previously developed tacrine-quinone hybrid (1). We designed and synthesized a novel set of MTDLs (2-6) by replacing the naphthoquinone scaffold of 1 with that of 2,5,8-quinolinetrione. The most interesting hybrid 3 inhibited cholinesterase enzymes at nanomolar concentrations. In addition, 3 exerted antioxidant effects in menadione-induced oxidative stress of SH-SY5Y cells. Importantly, 3 also showed low hepatotoxicity and good anti-amyloid aggregation properties. Remarkably, we uncovered the potential of the quinolinetrione scaffold, as a novel anti-amyloid aggregation and antioxidant motif to be used in further anti-AD MTDL drug discovery endeavors.

Efficacy and safety of adjustable suture in horizontal strabismus: Comparative study of children versus adults.

OBJECTIVE: Adjustable suture procedures allow addressing the unpredictability of some postoperative results in strabismus surgery. The purpose of the study was to compare the effectiveness of adjustable and non-adjustable suture in the treatment of horizontal strabismus in children and adults. METHODS: Prospective study including patients undergoing strabismus surgery to correct horizontal strabismus with fixed hanging suture (non-adjustable suture group) and adjustable suture. Visual acuity, amblyopia, deviation, oblique muscle involvement, previous surgeries, nystagmus, need for adjustment, and complications were recorded. The variables were recorded in the immediate postoperative period, at one week and at 3 and 6 months. RESULTS: 186 patients were included: 157 (84.4%) with adjustable suture and 29 (15.6%) with non-adjustable suture, of which 119 were children and 67 were adults. Postoperatively, 19 children (16.0%) and 19 adults (28.4%) required adjustment (p = 0.044). Of 157 patients with adjustable suture, it was adjusted in 20% (32/157). Success after adjustment was higher for adjustable suture (91.72% vs 79.31%; p = 0.043) and remained for 6 months (p < 0.05). Previous surgery (p = 0.004) and exotropia (p = 0.018) correlated with the need for adjustment. CONCLUSIONS: 20% of patients with horizontal strabismus can benefit from a postoperative adjustment to improve the surgical result. The adjustable suture was shown to be superior to the fixed hanging suture and is an excellent surgical option, both in children and adults.

Approaches to the Potential Therapy of COVID-19: A General Overview from the Medicinal Chemistry Perspective.

In spite of advances in vaccination, control of the COVID-19 pandemic will require the use of pharmacological treatments against SARS-CoV2. Their development needs to consider the existence of two phases in the disease, namely the viral infection and the inflammatory stages. The main targets for antiviral therapeutic intervention are: (a) viral proteins, including the spike (S) protein characteristic of the viral cover and the viral proteases in charge of processing the polyprotein arising from viral genome translation; (b) host proteins, such as those involved in the processes related to viral entry into the host cell and the release of the viral genome inside the cell, the elongation factor eEF1A and importins. The use of antivirals targeted at host proteins is less developed but it has the potential advantage of not being affected by mutations in the genome of the virus and therefore being active against all its variants. Regarding drugs that address the hyperinflammatory phase of the disease triggered by the so-called cytokine storm, the following strategies are particularly relevant: (a) drugs targeting JAK kinases; (b) sphingosine kinase 2 inhibitors; (c) antibodies against interleukin 6 or its receptor; (d) use of the traditional anti-inflammatory corticosteroids.

Multitarget Hybrid Fasudil Derivatives as a New Approach to the Potential Treatment of Amyotrophic Lateral Sclerosis.

Hybrid compounds containing structural fragments of the Rho kinase inhibitor fasudil and the NRF2 inducers caffeic and ferulic acids were designed with the aid of docking and molecular mechanics studies. Following the synthesis of the compounds using a peptide-coupling methodology, they were characterized for their ROCK2 inhibition, radical scavenging, effects on cell viability (MTT assay), and NRF2 induction (luciferase assay). One of the compounds (1d) was selected in view of its good multitarget profile and good tolerability. It was able to induce the NRF2 signature, promoting the expression of the antioxidant response enzymes HO-1 and NQO1, via a KEAP1-dependent mechanism. Analysis of mRNA and protein levels of the NRF2 pathway showed that 1d induced the NRF2 signature in control and SOD1-ALS lymphoblasts but not in sALS, where it was already increased in the basal state. These results show the therapeutic potential of this compound, especially for ALS patients with a SOD1 mutation.

Enhanced Stability and Bioactivity of Natural Anticancer Topoisomerase I Inhibitors through Cyclodextrin Complexation.

The use of cyclodextrins as drug nano-carrier systems for drug delivery is gaining importance in the pharmaceutical industry due to the interesting pharmacokinetic properties of the resulting inclusion complexes. In the present work, complexes of the anti-cancer alkaloids camptothecin and luotonin A have been prepared with ß-cyclodextrin and hydroxypropyl-ß-cyclodextrin. These cyclodextrin complexes were characterized by nuclear magnetic resonance spectroscopy (NMR). The variations in the 1H-NMR and 13C-NMR chemical shifts allowed to establish the inclusion modes of the compounds into the cyclodextrin cavities, which were supported by docking and molecular dynamics studies. The efficiency of the complexation was quantified by UV-Vis spectrophotometry and spectrofluorimetry, which showed that the protonation equilibria of camptothecin and luotonin A were drastically hampered upon formation of the inclusion complexes. The stabilization of camptothecin towards hydrolysis inside the cyclodextrin cavity was verified by the quantitation of the active lactone form by reverse phase liquid chromatography fluorimetric detection, both in basic conditions and in the presence of serum albumin. The antitumor activity of luotonin A and camptothecin complexes were studied in several cancer cell lines (breast, lung, hepatic carcinoma, ovarian carcinoma and human neuroblastoma) and an enhanced activity was found compared to the free alkaloids, particularly in the case of hydroxypropyl-ß-cyclodextrin derivatives. This result shows that the cyclodextrin inclusion strategy has much potential towards reaching the goal of employing luotonin A or its analogues as stable analogues of camptothecin.

Guidelines for enhanced recovery after cardiac surgery. Consensus document of Spanish Societies of Anesthesia (SEDAR), Cardiovascular Surgery (SECCE) and Perfusionists (AEP). / Vía clínica de recuperación intensificada en cirugía cardiaca. Documento de consenso de la Sociedad Española de Anestesiología, Reanimación y Terapéutica del Dolor (SEDAR), la Sociedad Española de Cirugía Cardiovascular y Endovascular (SECCE) y la Asociación Española de Perfusionistas (AEP).

The ERAS guidelines are intended to identify, disseminate and promote the implementation of the best, scientific evidence-based actions to decrease variability in clinical practice. The implementation of these practices in the global clinical process will promote better outcomes and the shortening of hospital and critical care unit stays, thereby resulting in a reduction in costs and in greater efficiency. After completing a systematic review at each of the points of the perioperative process in cardiac surgery, recommendations have been developed based on the best scientific evidence currently available with the consensus of the scientific societies involved.

Human amylin aggregates release within exosomes as a protective mechanism in pancreatic ß cells: Pancreatic ß-hippocampal cell communication.

Pancreatic ß cells are essential in the maintenance of glucose homeostasis during the progression to type 2 Diabetes Mellitus (T2DM), generating compensatory hyperinsulinemia to counteract insulin resistance. It is well known, that throughout the process there is an increased mTORC1 signaling pathway, with an impairment in different quality control systems including ubiquitin-proteasome system and autophagy. In addition, under this situation, pancreatic ß cells start to accumulate amylin protein (IAPP) in aggregates, and this accumulation contributes to the failure of autophagy, damaging different organelles such as plasma membrane, endoplasmic reticulum, mitochondria, and others. Here, we report that IAPP can be incorporated to multivesicular bodies (MVB) and secreted into exosomes, a mechanism responsible for the exportation of these toxic aggregates as vehicles of cell to cell communication. On this regard, we have demonstrated that the exosomes bearing toxic hIAPP released from pancreatic ß cells are capable to induce hyperactivation of mTORC1 signaling, a failure in the autophagic cellular quality control, and favor pro-fission status of the mitochondrial dynamics in hippocampal cells. In summary, our results show that harmful accumulation of hIAPP in pancreatic ß cells may be detoxified by the release of exosomes, which may be captured by endocytosis mechanism damaging neuronal hippocampal cells, which suggest an underlying molecular mechanism to the link between type 2 diabetes and neurodegenerative diseases.

Discovery of 7-aminophenanthridin-6-one as a new scaffold for matrix metalloproteinase inhibitors with multitarget neuroprotective activity.

Matrix metalloproteinases (MMPs) are zinc-dependent hydrolytic enzymes of great biological relevance, and some of them are key to the neuroinflammatory events and the brain damage associated to stroke. Non-zinc binding ligands are an emerging trend in drug discovery programs in this area due to their lower tendency to show off-target effects. 7-Amino-phenanthridin-6-one is disclosed as a new framework able to inhibit matrix metalloproteinases by binding to the distal part of the enzyme S1' site, as shown by computational studies. A kinetic study revealed inhibition to be noncompetitive. Some of the compounds showed some degree of selectivity for the MMP-2 and MMP-9 enzymes, which are crucial for brain damage associated to ischemic stroke. Furthermore, some compounds also had a high neuroprotective activity against oxidative stress, which is also very relevant aspect of ischaemic stroke pathogenesis, both decreasing lipid peroxidation and protecting against the oxidative stress-induced reduction in cell viability. One of the compounds, bearing a 2-thienyl substituent at C-9 and a 4-methoxyphenylamino at C-7, had the best-balanced multitarget profile and was selected as a lead on which to base future structural manipulation.

Estimación de costos unitarios directos de atención curativa de dengue en Ecuador 2020 / Estimate of direct unit costs of curative Dengue care in Ecuador 2020

El dengue es una enfermedad viral aguda transmitida a través de la picadura del mosquito (artrópodo) Aedes aegypti infectado con cualquiera de los serotipos de virus dengue. Son virus envueltos, de 40 a 50 nm de diámetro, con cápside icosaédrica y genoma de ácido ribonucleico (ARN) monocatenario, no segmentado, de polaridad positiva perteneciente al género flavivirus de la familia Flaviviridae. El análisis filogenético de las distintas cepas del virus dengue indica que la diseminación global ha dado lugar a distintos genotipos dentro de cada serotipo. En el quinquenio 2015-2020, en Ecuador, se notificaron en toda la región 76.085 casos de dengue. La atención unitaria hospitalaria para el tratamiento del dengue de las unidades de atención sanitaria hospitalarias (A, B, C), generaron un costo total promedio de 498,04 $ con una desviación estándar de ± 40,36, en el Ecuador para el año 2020. Cabe considerar, por otra parte, que el costo unitario directo promedio en ($) de la atención sanitaria en las entidades hospitalarias estudiadas en la presente investigación, generanron un gasto total para el tratamiento hospitalario de 9.585.114,02. Al mismo tiempo en los 2.135 pacientes diagnosticados con dengue que presentaron signos de alarma y requirieron cuatro días de hospitalización, señalando un gasto total para el tratamiento de 1.471.385,06. Por su parte los 51 pacientes que fueron diagnosticados con dengue grave, generaron un costo unitario total en las unidades hospitalarias de 134.152,54. Los resultados son conservadores, porque algunos componentes importantes no se incluyeron en los costos relacionados con el dengue.En ausencia de vacunación, y siendo los programas de control del vector la estrategia básica para mitigar la propagación del dengue, esta enfermedad seguirá produciendo una carga económica y social considerable en el Ecuador, lo que se refleja en el costo total de la enfermedad(AU)

Dengue is an acute viral disease transmitted through the bite of the mosquito (arthropod) Aedes aegypti infected with any of the dengue virus serotypes. They are enveloped viruses, 40 to 50 nm in diameter, with an icosahedral capsid and a single-stranded, non-segmented ribonucleic acid (RNA) genome of positive polarity belonging to the genus Flavivirus of the Flaviviridae family. Phylogenetic analysis of the different dengue virus strains indicates that global spread has given rise to different genotypes within each serotype. In the five-year period 2015-2020, in Ecuador, 76,085 cases of dengue were reported throughout the region. The hospital unit care for the treatment of dengue in the hospital health care units (A, B, C), generated an average total cost of $ 498.04 with a standard deviation of ± 40.36, in Ecuador for the year 2020. It should be considered, on the other hand, that the average direct unit cost in ($) of health care in the hospital entities studied in the present investigation, generated a total cost for hospital treatment of 9,585,114.02. At the same time, in the 2,135 patients diagnosed with dengue who presented alarm signs and required four days of hospitalization, indicating a total cost for treatment of 1,471,385.06. On the other hand, the 51 patients who were diagnosed with severe dengue, generated a total unit cost in the hospital units of 134,152.54. The results are conservative, because some important components were not included in the costs related to dengue. In the absence of vaccination, and with vector control programs the basic strategy to mitigate the spread of dengue, this disease will continue to produce an economic burden and considerable social in Ecuador, which is reflected in the total cost of the disease(AU)

[Development of a predictive model for hospital mortality and re-admission in a cohort of infected patients that require hospitalization]. / Desarrollo de un modelo predictivo para mortalidad y otro para reingreso hospitalario en una cohorte de paciente con infección que requieren hospitalización.

OBJECTIVE: The aims of the study were: to develop a predictive model for hospital mortality and another for hospital re-admission, to identify the impact of antibiotic delay in the mortality rate and, to report the rate of inappropriate antibiotic therapy. METHODS: A cohort and retrospective study was conducted at the HM Sanchinarro University Hospital during the period September 1st, 2012 to March 31th, 2013. The inclusion criteria were: age> 18 years, hospital admission from the ED with a diagnosis of bacterial infection. The exclusion criteria were: suspected viral infection, negative bacteriological cultures, life expectancy less than 6 months, lack of clinical information, assistance exclusively by the trauma emergency department. Two logistic models were made (hospital mortality and hospital re-admission). RESULTS: A total of 517 patients were included. The final mortality model (30 deaths) include the following variables: respiratory rate (OR 1.12; IC95% 1.02; 1.22), oxygen saturation (OR 0.92; IC95% 0.87; 0.98), creatinine (OR 2.33; IC95% 1.62; 3.36), COPD (OR 3.02; IC95% 1.06; 8.21), cancer (OR 3.34; IC95% 1.07; 9.98) and chemotherapy in the last 3 months (OR 4.83; IC95% 1.54; 16.41). The final model for hospital re-admission (28 re-admissions) include the following variables: hepatopathy (OR 5.51; IC95% 1.57; 16.88), GPT (OR 1.005; IC95% 1.003; 1.008), history of stroke (OR 5.06; IC95% 1.04; 18.80) and arterial hypertension (OR 3.15; IC95% 1.38; 7.56). The antibiotic therapy delays not influenced the mortality or re-admission rate. In 24.3% the causative microorganism was identified and antibiotic treatment was inappropriate 19.6%. CONCLUSIONS: Hospital mortality rate was 5.8% and readmission rate was 5.7%. Variables associated with mortality differ from those associated with re-admission. The delay in the antibiotic initiation was not associated with a deleterious effect. Antibiotic therapy was inadequate in almost 20% of patients.

First Glimpse of the N=82 Shell Closure below Z=50 from Masses of Neutron-Rich Cadmium Isotopes and Isomers.

We probe the N=82 nuclear shell closure by mass measurements of neutron-rich cadmium isotopes with the ISOLTRAP spectrometer at ISOLDE-CERN. The new mass of ^{132}Cd offers the first value of the N=82, two-neutron shell gap below Z=50 and confirms the phenomenon of mutually enhanced magicity at ^{132}Sn. Using the recently implemented phase-imaging ion-cyclotron-resonance method, the ordering of the low-lying isomers in ^{129}Cd and their energies are determined. The new experimental findings are used to test large-scale shell-model, mean-field, and beyond-mean-field calculations, as well as the ab initio valence-space in-medium similarity renormalization group.

D-Ring-Modified Analogues of Luotonin A with Reduced Planarity: Design, Synthesis, and Evaluation of Their Topoisomerase Inhibition-Associated Cytotoxicity.

A- and D-ring-modified luotonin-inspired heterocycles have been synthesized and were evaluated for their activity against the viability of four cancer cell lines in vitro, namely, MCF7, HCT116, JURKAT, and NCI-H460. The analysis of results indicated that two of the synthesized derivatives displayed good inhibition against the growth of the human colon cancer HCT116 cell line, with potencies lower than but in the same order of magnitude as camptothecin (CPT). These two luotonin analogues also showed an activity similar to that of the highly potent alkaloid CPT as inhibitors of topoisomerase I and also inhibited topoisomerase II. These results show that complete planarity is not a strict requirement for topoisomerase inhibition by luotonin-related compounds, paving the way to the design of analogues with improved solubility.

Progress in the Chemistry of Tetrahydroquinolines.

Tetrahydroquinoline is one of the most important simple nitrogen heterocycles, being widespread in nature and present in a broad variety of pharmacologically active compounds. This Review summarizes the progress achieved in the chemistry of tetrahydroquinolines, with emphasis on their synthesis, during the period from mid-2010 to early 2018.

Spirooxindole-pyrrolidine heterocyclic hybrids promotes apoptosis through activation of caspase-3.

A small library of spirooxindole-pyrrolidine hybrids have been synthesized for the first time in an ionic liquid, [bmim]Br in good to excellent yields employing a new class of non-stabilized azomethine ylides derived from isatin and tyrosine, a combination that has been rarely employed for the in situ generation of azomethine ylides using [3+2] cycloaddition strategy. Following the synthesis and characterization of the spirooxindole-pyrrolidine heterocyclic hybrids, they were tested for their anticancer activity as against the changes in the concentrations and time periods with different in vitro cell cultures containing cancer and non-cancer cells, where the results revealed for a potential therapeutic activity. Further analysis for the mechanism of cell death by the cancer cells indicated for the caspase-dependent apoptotic pathway, specifically mediated by caspase-3. Based on these results, it can be demonstrated that the synthesized spirooxindole-pyrrolidine hybrids may serve as one of the better therapeutic agents used for the treatment of malignant tumors.

Regio and stereoselective synthesis of anticancer spirooxindolopyrrolidine embedded piperidone heterocyclic hybrids derived from one-pot cascade protocol.

BACKGROUND: Spiropyrrolidine tethered piperidone heterocyclic hybrids were synthesized with complete regio- and stereoselectively in excellent yield via a tandem three-component 1,3-dipolar cycloaddition and subsequent enamine reaction in [bmim]Br. The synthesized compounds were evaluated for their anticancer activity against FaDu hypopharyngeal tumor cells. FINDINGS: Interestingly, most compounds displayed cytotoxicities similar to the standard anticancer agent bleomycin, with two of them (5a and 5g) being slightly more active than the reference drug. CONCLUSION: Synthesized compounds have also been evaluated for their apoptosis-inducing properties in a cancer cell model, finding that treatment with compounds 5a-e led to apoptotic cell death.

Tacripyrimidines, the first tacrine-dihydropyrimidine hybrids, as multi-target-directed ligands for Alzheimer's disease.

Notwithstanding the combination of cholinesterase (ChE) inhibition and calcium channel blockade within a multitarget therapeutic approach is envisaged as potentially beneficial to confront Alzheimer's disease (AD), this strategy has been scarcely investigated. To explore this promising line, a series of 5-amino-4-aryl-3,4,6,7,8,9-hexahydropyrimido [4,5-b]quinoline-2(1H)-thiones (tacripyrimidines) (4a-l) were designed by juxtaposition of tacrine, a ChE inhibitor (ChEI), and 3,4-dihydropyrimidin-2(1H)-thiones, as efficient calcium channel blockers (CCBs). In agreement with their design, all tacripyrimidines, except the unsubstituted parent compound and its p-methoxy derivative, acted as moderate to potent CCBs with activities generally similar or higher than the reference CCB drug nimodipine and were modest-to-good ChEIs. Most interestingly, the 3'-methoxy derivative (4e) emerged as the first well balanced ChEI/CCB agent, acting as low micromolar hChEI (3.05 µM and 3.19 µM on hAChE and hBuChE, respectively) and moderate CCB (30.4% at 1 µM) with no significant hepatotoxicity toward HepG2 cells and good predicted oral absorption and blood brain barrier permeability.

Multicomponent Domino Synthesis, Anticancer Activity and Molecular Modeling Simulation of Complex Dispirooxindolopyrrolidines.

A series of spirooxindolopyrrolidine fused N-styrylpiperidone heterocyclic hybrids has been synthesized in excellent yield via a domino multicomponent protocol that involves one-pot three component 1,3-dipolar cycloaddition and concomitant enamine reactions performed in an inexpensive ionic liquid, namely 1-butyl-3-methylimidazolium bromide ([bmim]Br). Compounds thus synthesized were evaluated for their cytotoxicity against U-937 tumor cells. Interestingly; compounds 5i and 5m exhibited a better cytotoxicity than the anticancer drug bleomycin. In addition; the effect of the synthesized compounds on the nuclear morphology of U937 FaDu cells revealed that treatment with compounds 5a⁻m led to their apoptotic cell death.