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Reactive oxygen species comprise oxygen-based free radicals and non-radical species such as peroxynitrite and electronically excited (singlet) oxygen. These reactive species often have short lifetimes, and much of our understanding of their formation and reactivity in biological and especially medical environments has come from complimentary fast reaction methods involving pulsed lasers and high-energy radiation techniques. These and related methods, such as EPR, are discussed with particular reference to singlet oxygen, hydroxy radicals, the superoxide radical anion, and their roles in medical aspects, such as cancer, vision and skin disorders, and especially pro- and anti-oxidative processes.

International audience; Aims: In the EMPA-REG OUTCOME trial, empagliflozin reduced risk of death from heart failure (HF) or hospitalization for heart failure (HHF) versus placebo in patients with type 2 diabetes mellitus (T2DM) and established cardiovascular (CV) disease. We evaluated post hoc the degree to which covariates mediated the effects of empagliflozin on HHF or HF death.Methods and results: A mediator had to fulfil the following criteria: (i) affected by active treatment, (ii) associated with the outcome, and finally (iii) adjustment for it results in a reduced treatment effect compared with unadjusted analysis. Potential mediators were calculated as change from baseline or updated mean and evaluated in univariable analyses as time-dependent covariates in Cox regression of time to HHF or HF death; those with the largest mediating effects were then included in a multivariable analysis. Increases in heart rate, log urine albumin-to-creatinine ratio (UACR), waist circumference, and uric acid were associated with increased risk of HHF or HF death; increases in high-density lipoprotein cholesterol, estimated glomerular filtration rate, haematocrit, haemoglobin, and albumin were associated with reduced risk of HHF or HF death. In univariable analyses, change from baseline in haematocrit, haemoglobin, albumin, uric acid, and logUACR mediated 51%, 54%, 23%, 24%, and 27% of the risk reduction with empagliflozin versus placebo, respectively. Multivariable analysis including haemoglobin, logUACR, and uric acid mediated 85% of risk reduction with similar results when updated means were evaluated.Conclusions: Changes in haematocrit and haemoglobin were the most important mediators of the reduction in HHF and death from HF in patients with T2DM and established CV disease treated with empagliflozin. Albumin, uric acid, and logUACR had smaller mediating effects in this population.

Background Nephrotic-range proteinuria (NRP) is associated with rapid kidney function loss and increased cardiovascular (CV) disease risk. We assessed the effects of linagliptin (LINA) on CV and kidney outcomes in people with Type 2 diabetes (T2D) with or without NRP. Methods Cardiovascular and renal microvascular outcome study with LINA randomized participants with T2D and CV disease and/or kidney disease to LINA 5 mg or placebo (PBO). The primary endpoint [time to first occurrence of 3-point major adverse cardiac events (3P-MACE)], and kidney outcomes, were evaluated by NRP status [urinary albumin:creatinine ratio (UACR) ≥2200 mg/g] at baseline (BL) in participants treated with one or more dose of study medication. Results NRP was present in 646/6979 [9.3% (LINA/PBO n = 317/n = 329); median UACR 3486 (Q1: 2746/Q3: 4941) mg/g] participants, who compared with no-NRP were younger (62.3/66.1 years) and had lower estimated glomerular filtration rate (eGFR) (39.9/56.1 mL/min/1.73 m2). Over a median of 2.2 years, 3P-MACE occurred with a 2.0-fold higher rate in NRP versus no-NRP (PBO group), with a neutral LINA effect, regardless of NRP. The composite of time to renal death, end-stage kidney disease (ESKD) or decrease of ≥40 or ≥50% in eGFR, occurred with 12.3- and 13.6-fold higher rate with NRP (PBO group); evidence of heterogeneity of effects with LINA was observed for the former [NRP yes/no: hazard ratio 0.80 (0.63–1.01)/1.25 (1.02–1.54); P-interaction 0.005], but not the latter [0.83 (0.64–1.09)/1.17 (0.91–1.51), P-interaction 0.07]. No heterogeneity was observed for renal death or ESKD [0.88 (0.64–1.21)/0.94 (0.67–1.31), P-interaction 0.79]. Glycated haemoglobin A1c (HbA1c) was significantly reduced regardless of NRP, without increasing hypoglycaemia risk. Regression to normoalbuminuria [1.20 (1.07–1.34)] and reduction of UACR ≥50% [1.15 (1.07–1.25)] from BL, occurred more frequently with LINA, regardless of NRP status (P-interactions >0.05). Conclusions Individuals with T2D and NRP have a high disease burden. LINA reduces their albuminuria burden and HbA1c, without affecting CV or kidney risk., Graphical Abstract

A hypothesis is proposed to explain the increased detrimental effect of COVID-19 for Black, Asian and Minority Ethnic (BAME) men and women compared to Caucasian individuals. This is based on the differing photochemistry of phaeomelanin in fair skin and eumelanin in dark/black skin. It is suggested that a range of reactive oxygen species, including, singlet oxygen and the superoxide radical anion, derived via direct photolysis of phaeomelanin, may escape the melanocyte and cause subsequent damage to the SARS-CoV-2 virus. It is further suggested that (large) carbon and sulphur peroxy radicals, from oxygen addition to radicals formed by carbon–sulphur bond cleavage, may assist via damage to the cell membranes. It is also speculated that light absorption by phaeomelanin and the subsequent C-S bond cleavage, leads to release of pre-absorbed reactive oxygen species, such as singlet oxygen and free radicals, which may also contribute to an enhanced protective effect for fair-skinned people.

The interactions of dietary carotenoids, and particularly the xanthophylls in the macula, with singlet oxygen and three different oxy-radicals, (hydroxyl radical, nitrogen dioxide and the superoxide radical anion) are compared using pulsed laser and γ-techniques. The results give possible molecular mechanisms for the switch from anti-oxidant (protection) by carotenoids to pro-oxidant (damage) by carotenoids. The participation of oxygen in radical mechanisms in the presence of different carotenoids is compared for the different radicals. It is shown that the mechanistic role of oxygen differs very significantly for anti-/pro-oxidation by hydroxyl radicals when compared to nitrogen dioxide. Lutein was found to be an extremely good cell protector against hydroxyl radicals at all oxygen concentrations, including under physiological conditions.

Context Control of multiple cardiovascular (CV) risk factors reduces CV events in individuals with type 2 diabetes. Objective To investigate this association in a contemporary clinical trial population, including how CV risk factor control affects the CV benefits of empagliflozin, a sodium-glucose cotransporter-2 inhibitor. Design Post hoc analysis. Setting Randomized CV outcome trial (EMPA-REG OUTCOME). Participants Type 2 diabetes patients with established CV disease. Intervention Empagliflozin or placebo. Main Outcome Measures Risk of CV outcomes—including the treatment effect of empagliflozin—by achieving 7 goals for CV risk factor control at baseline: (1) glycated hemoglobin Results In the placebo group, the hazard ratio (HR) for CV death was 4.00 (95% CI, 2.26–7.11) and 2.48 (95% CI, 1.52–4.06) for patients achieving only 0–3 or 4–5 risk factor goals at baseline, respectively, compared with those achieving 6–7 goals. Participants achieving 0–3 or 4–5 goals also had increased risk for the composite outcome of hospitalization for heart failure or CV death (excluding fatal stroke) (HR 2.89 [1.82–4.57] and 1.90 [1.31–2.78], respectively) and 3-point major adverse CV events (HR 2.21 [1.53–3.19] and 1.42 [1.06–1.89]). Empagliflozin significantly reduced these outcomes across all risk factor control categories (P > 0.05 for treatment-by-subgroup interactions). Conclusions Cardiovascular risk in EMPA-REG OUTCOME was inversely associated with baseline CV risk factor control. Empagliflozin’s cardioprotective effect was consistent regardless of multiple baseline risk factor control.

Responding to concerns about the potential for increased risk of adverse cardiovascular outcomes, specifically myocardial infarction, associated with certain glucose-lowering therapies, the US Food and Drug Administration and the Committee for Medicinal Products for Human Use of the European Medicines Agency issued guidance to the pharmaceutical industry in 2008. Glucose-lowering therapies were granted regulatory approval primarily from smaller studies that have demonstrated reductions in glycated hemoglobin concentration. Such studies were overall underpowered and of insufficient duration to show any effect on cardiovascular outcomes. The 2008 guidance aimed to ensure the cardiovascular safety of new glucose-lowering therapies to treat patients with type 2 diabetes mellitus. This resulted in a plethora of new cardiovascular outcome trials, most designed primarily as placebo-controlled noninferiority trials, but with many also powered for superiority. Several of these outcome trials demonstrated cardiovascular benefits of the newer agents, resulting in the first-ever cardiovascular protection indications for glucose-lowering therapies. Determining whether the guidance continues to have value in its current form is critically important as we move forward after the first decade of implementation. In February 2018, a think tank comprising representatives from academia, industry, and regulatory agencies convened to consider the guidance in light of the findings of the completed cardiovascular outcome trials. The group made several recommendations for future regulatory guidance and for cardiovascular outcome trials of glucose-lowering therapies. These recommendations include requiring only the 1.3 noninferiority margin for regulatory approval, conducting trials for longer durations, considering studying glucose-lowering therapies as first-line management of type 2 diabetes mellitus, considering heart failure or kidney outcomes within the primary outcome, considering head-to-head active comparator trials, increasing the diversity of patients enrolled, evaluating strategies to streamline registries and the study of unselected populations, and identifying ways to improve translation of trial results to general practice.

Objective: Functional HDL (high-density lipoprotein) particles that facilitate cholesterol efflux may be cardioprotective. EL (endothelial lipase) hydrolyzes phospholipids promoting catabolism of HDL and subsequent renal excretion. MEDI5884 is a selective, humanized, monoclonal, EL-neutralizing antibody. We sought to determine the safety, pharmacokinetics, and pharmacodynamic effects of multiple doses of MEDI5884 in patients with stable coronary artery disease. Approach and Results: LEGACY was a phase 2a, double-blind, placebo-controlled, parallel-design trial that randomized 132 patients with stable coronary artery disease receiving high-intensity statin therapy to 3 monthly doses of 1 of 5 dose levels of MEDI5884 (50, 100, 200, 350, or 500 mg SC) or matching placebo. The primary end point was the safety and tolerability of MEDI5884 through the end of the study (day 151). Additional end points included change in HDL cholesterol and cholesterol efflux from baseline to day 91, hepatic uptake of cholesterol at day 91, changes in various other lipid parameters. The incidence of adverse events was similar between the placebo and MEDI5884 groups. In a dose-dependent manner, MEDI5884 increased HDL cholesterol up to 51.4% ( P P P P =0.04) was observed with MEDI5884. However, at the potential target doses for future studies, there was no meaningful increase in LDL cholesterol or apoB. Conclusions: Inhibition of EL by MEDI5884 increases the quantity and quality of functional HDL in patients with stable coronary artery disease on high-intensity statin therapy without an adverse safety signal at the likely dose to be used. These data support further clinical investigation. Registration: URL: https://www.clinicaltrials.gov ; Unique identifier: NCT03351738.

Background MEDI6012 is recombinant human lecithin cholesterol acyltransferase, the rate‐limiting enzyme in reverse cholesterol transport. Infusions of lecithin cholesterol acyltransferase have the potential to enhance reverse cholesterol transport and benefit patients with coronary heart disease. The purpose of this study was to test the safety, pharmacokinetic, and pharmacodynamic profile of MEDI6012. Methods and Results This phase 2a double‐blind study randomized 48 subjects with stable coronary heart disease on a statin to a single dose of MEDI6012 or placebo (6:2) (NCT02601560) with ascending doses administered intravenously (24, 80, 240, and 800 mg) and subcutaneously (80 and 600 mg). MEDI6012 demonstrated rates of treatment‐emergent adverse events that were similar to those of placebo. Dose‐dependent increases in high‐density lipoprotein cholesterol were observed with area under the concentration‐time curves from 0 to 96 hours of 728, 1640, 3035, and 5318 should be: mg·h/mL in the intravenous dose groups and 422 and 2845 mg·h/mL in the subcutaneous dose groups. Peak mean high‐density lipoprotein cholesterol percent change was 31.4%, 71.4%, 125%, and 177.8% in the intravenous dose groups and 18.3% and 111.2% in the subcutaneous dose groups, and was accompanied by increases in endogenous apoA1 (apolipoprotein A1) and non‐ATP‐binding cassette transporter A1 cholesterol efflux capacity. Decreases in apoB (apolipoprotein B) were observed across all dose levels and decreases in atherogenic small low‐density lipoprotein particles by 41%, 88%, and 79% at the 80‐, 240‐, and 800‐mg IV doses, respectively. Conclusions MEDI6012 demonstrated an acceptable safety profile and increased high‐density lipoprotein cholesterol, endogenous apoA1, and non‐ATP‐binding cassette transporter A1 cholesterol efflux capacity while reducing the number of atherogenic low‐density lipoprotein particles. These findings are supportive of enhanced reverse cholesterol transport and a functional high‐density lipoprotein phenotype. Registration URL: https://www.clinicaltrials.gov ; Unique identifier: NCT02601560.

Background Cardio/kidney composite end points are clinically relevant but rarely analyzed in cardiovascular trials. This post hoc analysis of the EMPA‐REG OUTCOME (Empagliflozin Cardiovascular Outcome Event Trial in Type 2 Diabetes Mellitus Patients) trial evaluated cardio/kidney composite end points by 2 statistical approaches. Methods and Results A total of 7020 patients with type 2 diabetes mellitus and established cardiovascular disease were treated with empagliflozin 10 or 25 mg (n=4687) or placebo (n=2333) on top of standard care. Cardio/kidney composite end points studied were: (1) cardiac or kidney death, kidney failure, hospitalization for heart failure, sustained decline in estimated glomerular filtration rate ≥40% from baseline, or sustained progression to macroalbuminuria; (2) cardiac or kidney death, kidney failure, hospitalization for heart failure, or sustained estimated glomerular filtration rate decline ≥40% from baseline; and (3) cardiac or kidney death, kidney failure, hospitalization for heart failure, or sustained doubling in serum creatinine from baseline. Cox regression using time‐to‐first‐event analysis and win ratio (WR) using hierarchical order of events were applied. Empagliflozin reduced the risk of all cardio/kidney composites. The results varied only slightly between Cox and WR (eg, composite 1: hazard ratio, 0.56 [95% CI, 0.49–0.64]; WR, 1.76 [95% CI, 1.53–2.02]. WR prioritizes events by clinical importance; in particular, all fatal events are evaluated, whereas Cox regression ignores deaths when preceded by nonfatal events. Of the 285 cardio/kidney deaths in the analysis, 44 to 56 (15%–20%), depending on the composite, occurred after a nonfatal event and were not evaluated in Cox regression but evaluated by the WR. Conclusions By considering the clinical relevance of different event types, the WR represents an appropriate method to complement the traditional time‐to‐first‐event analysis in cardio/kidney outcomes. Registration URL: https://www.clinicaltrials.gov ; Unique identifier: NCT01131676.

Thermoresponsive poly(N-isopropylacrylamide) (PNIPAAm) hydrogels are widely studied smart materials, particularly for biomedical applications, but are limited by their mechanical strength. In this study, double network (DN) hydrogels were prepared with an asymmetric crosslink design and inclusion of an electrostatic co-monomer, 2-acrylamido-2-methylpropane sulfonic acid (AMPS). These P(NIPAAm-co-AMPS)/PNIPAAm DN hydrogels were sequentially formed with a tightly crosslinked 1(st) network comprised of variable levels of AMPS (100 : 0 to 25 : 75 wt% ratio of NIPAAm:AMPS) and a loosely crosslinked 2(nd) network comprised of PNIPAAm. The impact of AMPS content in the 1(st) network on the volume phase transition temperature (VPTT), morphology, deswelling–reswelling kinetics and mechanical properties was evaluated. Without substantially altering the VPTT of conventional PNIPAAm hydrogels but with improving thermosensitivity, the DN hydrogel formed with 25 : 75 wt% of NIPAAm:AMPS achieved exceptional strength, high modulus and high %strain at break.

Introduction: In EMPA-REG OUTCOME, empagliflozin (EMPA) reduced the risk of major adverse cardiovascular (CV) events (MACE), CV mortality and hospitalization for heart failure (HHF) in analyses of first events in patients with type 2 diabetes (T2D) and atherosclerotic CV disease (ASCVD). We assessed the effect of EMPA on the total burden of CV events. Methods: Patients were randomized to EMPA 10 mg, EMPA 25 mg, or placebo. We assessed the effects of EMPA pooled vs placebo on any (first plus recurrent) adjudicated CV event (composite of myocardial infarction (MI), stroke, coronary revascularization (CR), hospitalization for unstable angina, transient ischemic attack, HHF, and CV death) using a negative binomial model. Results: Among 7,020 treated patients (mean [SD] age 63 [9] years), there were 2,142 total adjudicated CV events, most frequently CR (585), MI (421), and HHF (321). EMPA reduced the risk of total adjudicated CV events by 24% vs placebo (event rate ratio (95% CI): 0.76 (0.67, 0.87), p Conclusions: EMPA produced a sizeable risk reduction in the total burden of any adjudicated CV outcome, including HHF, MI and CV death, in patients with T2D and ASCVD.

Introduction: In the REDUCE-IT trial, icosapent ethyl (IPE) was shown to reduce major adverse cardiac events (MACE) including cardiovascular (CV) death in patients with elevated triglycerides (TG) and atherosclerotic CV disease (ASCVD) and/or diabetes. There are limited data evaluating the external applicability of REDUCE-IT inclusion criteria in contemporary diabetes trials. In EMPA-REG OUTCOME, empagliflozin (EMPA) reduced the risk of CV death and hospitalization for HF (HHF). We sought to evaluate the benefit of EMPA on CV outcomes across IPE eligibility in the EMPA-REG OUTCOME trial. Methods: In total, 7020 patients with type 2 diabetes and ASCVD were treated with EMPA 10mg, 25 mg, or placebo (PBO). We examined the proportion of patients that had baseline (BL) TG 135 to 499 mg/dl, LDL-C 41 to 100 mg/dl and using a statin (“REDUCE-IT-like patients”). We evaluated the effect of pooled EMPA vs. PBO on CV death, HHF, HHF or CV death (excluding fatal stroke), all-cause death, and 3-point-MACE across the subgroups of patients fulfilling vs not fulfilling the REDUCE-IT criteria at BL using Cox regression. Results: A total of 6935 patients had TG and 6932 had LDL-C available. 1810 patients (25.8%) were REDUCE-IT like with TG 206±69 vs. 158±140 mg/dl, and LDL-C 71±15 vs. 91±39 mg/dl in those who did not meet the criteria. At BL, the former had more often coronary artery disease (84 vs. 73%) and higher BMI (31.7±5.1 kg/m 2 vs. 30.3±5.3) vs. the latter group. CV event rates in PBO were similar in patients who met/did not meet the criteria. Treatment effects of EMPA vs. PBO were consistent in patients who met/did not meet the REDUCE-IT criteria (Fig). Conclusions: In patients with T2D and established ASCVD treated in EMPA-REG OUTCOME, about one-quarter of patients would qualify for IPE according to the REDUCE-IT inclusion criteria. Empagliflozin consistently reduced CV outcomes and mortality in this sub-group, suggesting that IPE and SGLT2 inhibition may be complementary.

Introduction: In EMPA-REG OUTCOME®, the sodium-glucose transporter inhibitor, empagliflozin (EMPA), reduced the risk of first cardiovascular (CV) events primarily CV mortality and hospitalization for heart failure in patients with type 2 diabetes (T2D) and established atherosclerotic cardiovascular disease. In analyses including total events, EMPA also reduced the risk of myocardial infarction (MI). Here we report the effects of EMPA on the total number of MIs by subtype. Methods: Patients were randomized to receive EMPA 10 mg, EMPA 25 mg, or placebo. We assessed the effect of pooled EMPA versus placebo on total (first plus recurrent) events of fatal and non-fatal MI using a negative binomial model with robust confidence intervals (CI) that preserves randomization and accounts for the within-patient correlation of multiple events. Post-hoc, we analyzed established subtypes of MI: Type 1: Typically related to plaque-rupture or thrombus; Type 2: Supply-demand; Type 3: Sudden-death related (i.e., fatal MI); Type 4: Percutaneous coronary intervention related; and Type 5: Coronary artery bypass graft related. Except type 3 MIs, MIs were non-fatal. The MIs, including subtypes, were centrally adjudicated and could be assigned to more than one subtype. Results: 7,020 patients were treated. Overall, there was 421 total events of MIs. Specifically, 299, 86, 26, 19, and 1 of the MIs were classified as type 1, 2, 3, 4, and 5 MIs, respectively (not mutually exclusive). Overall, EMPA reduced the risk of total events of MI by 21% versus placebo (rate ratio [95% CI], EMPA versus placebo: 0.79 [0.620; 0.998], p=0.0486; each MI only counted once). The overall reduction in total events of MIs by EMPA was driven by the effect on type 1 and 2 MIs, as well as type 3 MIs, with limited number of type 4 and 5 MIs (Figure). Conclusions: EMPA reduced the risk of total MI events with consistent effects across the most common etiologies, the type 1 (plaque-rupture or thrombus) and 2 (supply-demand) MIs.

Introduction: Type 2 diabetes (T2D) and chronic obstructive pulmonary disease (COPD) often co-exist yielding increased risk of cardiovascular (CV) complications, including heart failure (HF). In the EMPA-REG OUTCOME trial, empagliflozin (EMPA) reduced the risk of CV death and hospitalization for HF (HHF) in patients with T2D and CV disease (CVD). We hypothesized that patients with COPD had a higher risk of CV outcomes than those without COPD, but similar treatment benefits on CV outcomes from EMPA as the overall population. Methods: In total, 7020 patients were treated with EMPA 10mg, 25mg, or placebo (PBO) with median follow-up of 3.1 years. We defined COPD at baseline (BL) by investigator reported presence of COPD or emphysema, or the use of medications for obstructive airways disease. We explored the effect of pooled EMPA groups vs. PBO within the subgroups of patients with vs. without COPD on CV death, HHF, HHF or CV death (excluding fatal stroke), all-cause death and 3-point-MACE by Cox regression adjusted for baseline risk factors. Results: The 707 (10.1%) patients with COPD at BL were slightly older (65±8 vs. 63±9 yrs), had more often HF at BL (17 vs. 9%) and coronary artery disease (84 vs. 75%), used insulin (56 vs. 47%) and diuretics (57 vs. 42%,) more frequently, but had similar beta-blocker use (64 vs. 65%) as compared to those without COPD. During follow-up, those with COPD in the placebo group had increased risk of HHF (HR 2.15 [95%CI 1.32-3.49], p=0.002), HHF or CV death (HR 1.60 [1.10-2.33], p Conclusions: In EMPA-REG OUTCOME, 10% of patients had concomitant COPD. COPD patients were at increased risk of HHF, HHF or CV death, as well as all-cause death. EMPA consistently reduced these outcomes vs PBO among patients with and without COPD. These data suggest that EMPA’s benefits in patients with T2D and CVD are not attenuated by presence of COPD.

Aims Reverse cholesterol transport (RCT) removes cholesterol and stabilizes vulnerable plaques. In addition, high-density lipoprotein (HDL) may be cardioprotective in acute myocardial infarction (MI). Lecithin-cholesterol acyltransferase (LCAT) may enhance RCT. The objective of this study was to investigate the pharmacokinetics, pharmacodynamics, and safety of multiple ascending doses of recombinant human LCAT (MEDI6012) to inform a Phase 2b programme. Methods and results This was a randomized, blinded, placebo-controlled, dose-escalation Phase 2a study of MEDI6012. Patients were randomized into one of four cohorts (40, 120, 300 mg IV weekly ×3 doses, or 300 mg IV-push, 150 mg at 48 h and 100 mg at 7 days). All cohorts were planned to randomize 6:2 (MEDI6012 vs. placebo). The primary endpoints were baseline-adjusted area under the curve (AUC) from 0 to 96 h post dose 3 (AUC 0–96 h) for HDL-C, HDL cholesteryl ester (HDL-CE), and total cholesteryl ester (CE). The primary safety endpoints were treatment-emergent adverse events. A total of 32 patients were randomized. MEDI6012 significantly increased AUC 0–96 h for HDL-C, HDL-CE and CE in a graded fashion with increasing doses. Relative to placebo, MEDI6012 increased HDL-C at Day 19 by 66% (95% CI 33–99, P = 0.014) with 120 mg and 144% (95% CI 108–181, P Conclusions Multiple ascending doses of MEDI6012 were safe and well tolerated and significantly increased HDL-C, HDL-CE and CE in a dose-related manner. These data support the ongoing Phase 2b programme investigating MEDI6012 in ST-elevation MI.

Background Individuals with polyvascular disease and impaired renal function are at high risk of cardiovascular (CV) events, but this relationship is not well investigated in people with type 2 diabetes (T2D). Furthermore, the impact of polyvascular disease plus renal dysfunction on the risk for hospitalisation for heart failure (HHF) remains unclear. Purpose We investigated this in a post hoc analysis of the EMPA-REG OUTCOME trial in which empagliflozin reduced risk of CV death and HHF versus placebo in people with T2D and vascular disease. In addition, we explored the treatment effect of empagliflozin on CV, HF and mortality outcomes across the spectrum of baseline polyvascular disease and impaired renal function. Methods Patients with T2D, CV disease and estimated glomerular filtration rate (eGFR) of ≥30 ml/min/1.73 m2 received empagliflozin 10 mg, 25 mg, or placebo. Vascular beds (VBs) were defined as coronary artery disease, peripheral artery disease, and cerebrovascular disease (Fig). By use of Cox regression, we explored the association between baseline eGFR < or ≥60 ml/min/1.73 m2, with or without polyvascular disease (1 vs ≥2 VBs involved), and CV death, HHF, CV death (excl. fatal stroke)/HHF, and all-cause mortality (ACM), as well as the treatment effect of empagliflozin versus placebo on these outcomes. Results Patients with ≥2 VBs involved and eGFR 10 years more often (73.4% vs. 63.2% or 54.9%), and a higher HF prevalence at baseline (19.4% vs. 11.1% or 9.2%) versus those with ≥2 VBs involved and eGFR ≥60 ml/min/1.73 m2 [n=866], or those with only 1 VB involved regardless of eGFR [n=5630], respectively. However, characteristics were generally balanced between treatment groups. Notably, co-existing polyvascular disease and eGFR Conclusions Co-existing polyvascular disease and eGFR Funding Acknowledgement Type of funding source: Private company. Main funding source(s): Boehringer Ingelheim and Eli Lilly and Company Diabetes Alliance

Objective Interindividual variation in responses to alcohol is substantial, posing challenges for medical management and for understanding the biological underpinnings of alcohol use disorders (AUD). It is important to understand whether diverse alcohol responses such as sedation, which is predictive of risk and partly heritable, occur concurrently or independently from responses such as blackouts and withdrawal. We hypothesized that latent factors accounting for sources of variance in diverse alcohol response phenotypes could be identified in a large, deeply phenotyped sample of patients with AUD. Methods We factor analyzed 17 alcohol response related items from the Alcohol Dependence Scale (ADS) in 938 individuals diagnosed with AUD via structured clinical interviews. Demographic, genetic, and clinical characteristics were tested as predictors of the latent factors by multiple indicators, multiple causes analysis. Results The final factor solution included three alcohol response factors: Physical Symptoms, Perceptual Disturbances, and Neurobiological Effects. Both gender and genetic ancestry were identified as variables influencing alcohol response. Major depressive disorder positively predicted physical symptoms and aggression negatively predicted physical symptoms. Barratt's Impulsivity Scale total score predicted the Physical and Perceptual domains. Family history, average drinks per drinking day, and negative urgency (an impulsivity measure) predicted all three domains. Conclusions Diverse items from the ADS concurrently load onto three correlated alcohol response factors rather than loading independently. Genetic ancestry and clinical characteristics predicted the severity of items that define the alcohol response factors even after accounting for degree of alcohol consumption. Co‐occurring phenotypes point towards an underlying shared physiology of diverse alcohol responses., HIGHLIGHTS Three common factors relevant for diverse alcohol responses are identified: Physical Symptoms, Perceptual Disturbances, and Neurobiological EffectsAlcohol response items from the Alcohol Dependence Scale concurrently load onto these three factors rather than loading independentlyThe three factors are correlated; patients presenting to clinical settings with a problem such as blackout are likely to experience several other problems either acutely or in the future

Background: Glucose variability has been associated with cardiovascular (CV) outcomes in type 2 diabetes, however, the interplay between glucose variability, empagliflozin and CV death has not been explored. In the EMPA-REG OUTCOME trial, empagliflozin reduced the risk of CV death by 38%. We explore post-hoc the association between HbA1c variability and CV death, and the potential mediating effects of HbA1c variability on empagliflozin’s CV death reductions.Methods: In total, 7,020 patients with type 2 diabetes and established CV disease received placebo, empagliflozin 10 mg or 25 mg. We defined within-patient HbA1c variability as standard deviation, coefficient of variation and range of HbA1c measurements (%) post-baseline. First, we compared HbA1c variability until week 28 and 52 by Wilcoxon tests. We explored the association between CV death and HbA1c variability in placebo and pooled empagliflozin arms separately with landmark analyses at week 28 and 52, and additionally with HbA1c variability as a time-dependent co-variate. We used Cox regression models adjusted for baseline risk factors including changes in HbA1c from baseline to week 12, and the interaction term HbA1c variability* treatment.Results: HbA1c variability was lower with empagliflozin compared to placebo. In all Cox analyses, high HbA1c variability increased the risk for CV death in both treatment arms with no interaction with treatment.Conclusions: HbA1c variability was reduced by empagliflozin and high values of HbA1c variability were associated with an increased risk of CV death. Empagliflozin’s reduction in CV death did not appear to be mediated by reductions in HbA1c variability.ClinicalTrials.gov number, NCT01131676. Registered May 27 2010. https://clinicaltrials.gov/ct2/show/NCT01131676

OBJECTIVE To evaluate the safety and efficacy of empagliflozin 10- and 25-mg doses plus a unique lower dose (2.5 mg) as adjunct to intensified insulin in patients with type 1 diabetes (T1D). RESEARCH DESIGN AND METHODS The EASE (Empagliflozin as Adjunctive to inSulin thErapy) program (N = 1,707) included two double-blind, placebo-controlled phase 3 trials: EASE-2 with empagliflozin 10 mg (n = 243), 25 mg (n = 244), and placebo (n = 243), 52-week treatment; and EASE-3 with empagliflozin 2.5 mg (n = 241), 10 mg (n = 248), 25 mg (n = 245), and placebo (n = 241), 26-week treatment. Together they evaluated empagliflozin 10 mg and 25 mg, doses currently approved in treatment of type 2 diabetes, and additionally 2.5 mg on 26-week change in glycated hemoglobin (primary end point) and weight, glucose time-in-range (>70 to ≤180 mg/dL), insulin dose, blood pressure, and hypoglycemia. RESULTS The observed largest mean placebo-subtracted glycated hemoglobin reductions were −0.28% (95% CI −0.42, −0.15) for 2.5 mg, −0.54% (−0.65, −0.42) for 10 mg, and −0.53% (−0.65, −0.42) for 25 mg (all P < 0.0001). Empagliflozin 2.5/10/25 mg doses, respectively, reduced mean weight by −1.8/−3.0/−3.4 kg (all P < 0.0001); increased glucose time-in-range by +1.0/+2.9/+3.1 h/day (P < 0.0001 for 10 and 25 mg); lowered total daily insulin dose by −6.4/−13.3/−12.7% (all P < 0.0001); and decreased systolic blood pressure by −2.1/−3.9/−3.7 mmHg (all P < 0.05). Genital infections occurred more frequently on empagliflozin. Adjudicated diabetic ketoacidosis occurred more with empagliflozin 10 mg (4.3%) and 25 mg (3.3%) but was comparable between empagliflozin 2.5 mg (0.8%) and placebo (1.2%). Severe hypoglycemia was rare and frequency was similar between empagliflozin and placebo. CONCLUSIONS Empagliflozin improved glycemic control and weight in T1D without increasing hypoglycemia. Ketoacidosis rate was comparable between empagliflozin 2.5 mg and placebo but increased with 10 mg and 25 mg. Ketone monitoring for early ketoacidosis detection and intervention and lower empagliflozin doses may help to reduce this risk.

Zinc deficiency is a frequent complication of alcohol abuse for multiple reasons including poor intake, increased excretion, internal redistribution and altered transporters. Zinc deficiency has been postulated to play a role in the development/progression of alcoholic liver disease (ALD). This study aimed to relate serum zinc levels with alcohol intake, serum albumin concentration and markers of inflammation and liver injury. One hundred and eight male and female very heavy drinking (≥10 drinks/day) individuals without clinical evidence of ALD were grouped by serum zinc concentration: normal-zinc group (zinc level≥71 μg/dl) included 67 patients, and low-zinc group (zinc level71 μg/dl) included 41 patients. Data were collected on demographics, drinking history in last 90 days (heavy drinking days, HDD90 and total drinks, TD90), lifetime drinking history (LTDH) and clinical/ laboratory assessments. Our data show that in a very well-characterized, chronically heavy-drinking population without clinical evidence of liver disease, about 40% of subjects had low serum zinc levels. Frequency of heavy drinking days (HDD90) was significantly higher in the low-zinc group. Total drinks in past 90 days, LTDH and HDD90 showed significant associations with low zinc levels. The group with the low serum zinc had a higher aspartate aminotransferase/alanine aminotransferase ratio (good marker of alcoholic liver disease). Those in the low-zinc group had the lower albumin levels, a marker of hepatic synthetic function, and the highest C-reactive protein level, a biomarker of inflammation.

// Federico Bocci 1, 2 , Mohit Kumar Jolly 1 , Jason Thomas George 1, 3, 6 , Herbert Levine 1, 2, 3, 4 and Jose Nelson Onuchic 1, 2, 4, 5 1 Center for Theoretical Biological Physics, Rice University, Houston, TX 77005, USA 2 Department of Chemistry, Rice University, Houston, TX 77005, USA 3 Department of Bioengineering, Rice University, Houston, TX 77005, USA 4 Department of Physics and Astronomy, Rice University, Houston, TX 77005, USA 5 Department of Biosciences, Rice University, Houston, TX 77005, USA 6 Medical Scientist Training Program, Baylor College of Medicine, Houston, TX 77030, USA Correspondence to: Jose Nelson Onuchic, email: jonuchic@rice.edu Herbert Levine, email: herbert.levine@rice.edu Keywords: epithelial-mesenchymal transition (EMT); cancer stem cells (CSCs); Notch signaling; hybrid epithelial/mesenchymal (E/M) phenotype; stemness window Received: May 05, 2018 Accepted: June 13, 2018 Published: July 06, 2018 ABSTRACT Epithelial-mesenchymal transition (EMT) and cancer stem cell (CSCs) formation are two fundamental and well-studied processes contributing to cancer metastasis and tumor relapse. Cells can undergo a partial EMT to attain a hybrid epithelial/mesenchymal (E/M) phenotype or a complete EMT to attain a mesenchymal one. Similarly, cells can reversibly gain or lose 'stemness'. This plasticity in cell states is modulated by signaling pathways such as Notch. However, the interconnections among the cell states enabled by EMT, CSCs and Notch signaling remain elusive. Here, we devise a computational model to investigate the coupling among the core decision-making circuits for EMT, CSCs and Notch. Our model predicts that hybrid E/M cells are most likely to associate with stem-like traits and enhanced Notch-Jagged signaling – a pathway implicated in therapeutic resistance. Further, we show that the position of the 'stemness window' on the 'EMT axis' is varied by altering the coupling strength between EMT and CSC circuits, and/or modulating Notch signaling. Finally, we analyze the gene expression profile of CSCs from several cancer types and observe a heterogeneous distribution along the 'EMT axis', suggesting that different subsets of CSCs may exist with varying phenotypes along the epithelial-mesenchymal axis. We further investigate therapeutic perturbations such as treatment with metformin, a drug associated with decreased cancer incidence and increased lifespan of patients. Our mechanism-based model explains how metformin can both inhibit EMT and blunt the aggressive potential of CSCs simultaneously, by driving the cells out of a hybrid E/M stem-like state with enhanced Notch-Jagged signaling.

Plasmodium vivax invasion into human reticulocytes is a complex process. The Duffy binding protein (DBP) dimerization with its cognate receptor is vital for junction formation in the invasion process. Due to its functional importance, DBP is considered a prime vaccine candidate, but variation in B-cell epitopes at the dimer interface of DBP leads to induction of strain-limited immunity. We believe that the polymorphic residues tend to divert immune responses away from functionally conserved epitopes important for receptor binding or DBP dimerization. As a proof of concept, we engineered the vaccine DEKnull to ablate the dominant Bc epitope to partially overcome strain-specific immune antibody responses. Additional surface engineering on the next generation immunogen, DEKnull-2, provides an immunogenicity breakthrough to conserved protective epitopes. DEKnull-2 elicits a stronger broadly neutralizing response and reactivity with long-term persistent antibody responses of acquired natural immunity. By using novel engineered DBP immunogens, we validate that the prime targets of protective immunity are conformational epitopes at the dimer interface. These successful results indicate a potential approach that can be used generally to improve efficacy of other malaria vaccine candidates.

Carotenoid pigments, particularly β-carotene and lycopene, are consumed in human foodstuffs and play a vital role in maintaining health. β-carotene is known to quench singlet oxygen and can have strong antioxidant activity. As such, it was proposed that β-carotene might reduce the risk of cancer. Epidemiological studies found inverse relationships between cancer risk and β-carotene intake or blood levels. However, clinical trials failed to support those findings and β-carotene supplementation actually increased lung cancer incidence in male smokers. Early experimental animal studies found dietary β-carotene inhibited UV-induced skin cancers. Later studies found that β-carotene supplementation exacerbated UV-carcinogenic expression. The discrepancies of these results were related to the type of diet the animals consumed. Lycopene has been associated with reduced risk of lethal stage prostate cancer. Other carotenoids, e.g., lutein and zeaxanthin, play a vital role in visual health. Numerous studies of molecular mechanisms to explain the carotenoids’ mode of action have centered on singlet oxygen, as well as radical reactions. In cellular systems, singlet oxygen quenching by carotenoids has been reported but is more complex than in organic solvents. In dietary β-carotene supplement studies, damaging pro-oxidant reactivity can also arise. Reasons for this switch are likely due to the properties of the carotenoid radicals themselves. Understanding singlet oxygen reactions and the anti-/pro-oxidant roles of carotenoids are of importance to photosynthesis, vision and cancer.

Despite the harm caused by binge drinking, the neural mechanisms leading to risky and disinhibited intoxication-related behaviors are not well understood. Evidence suggests that the globus pallidus externus (GPe), a substructure within the basal ganglia, participates in inhibitory control processes, as examined in stop-signaling tasks. In fact, studies in rodents have revealed that alcohol can change GPe activity by decreasing neuronal firing rates, suggesting that the GPe may have a central role in explaining impulsive behaviors and failures of inhibition that occur during binge drinking. In this study, twenty-five healthy volunteers underwent intravenous alcohol infusion to achieve a blood alcohol level of 0.08 g/dl, which is equivalent to a binge drinking episode. A resting state functional magnetic resonance imaging scan was collected prior to the infusion and at binge-level exposure. Functional connectivity analysis was used to investigate the association between alcohol-induced changes in GPe connectivity, drinking behaviors, and impulsivity traits. We found that individuals with greater number of drinks or heavy drinking days in the recent past had greater alcohol-induced deficits in GPe connectivity, particularly to the striatum. Our data also indicated an association between impulsivity and alcohol-induced deficits in GPe – frontal/precentral connectivity. Moreover, alcohol induced changes in GPe-amygdala circuitry suggested greater vulnerabilities to stress-related drinking in some individuals. Taken together, these findings suggest that alcohol may interact with impulsive personality traits and drinking patterns to drive alterations in GPe circuitry associated with behavioral inhibition, possibly indicating a neural mechanism by which binge drinking could lead to impulsive behaviors.

Background/Introduction CARMELINA was a randomized placebo-controlled clinical trial designed to demonstrate the cardiovascular (CV) safety of linagliptin in patients with type 2 diabetes (T2D) and concomitant cardiorenal disease. Despite a particularly elevated CV risk, only limited long-term evidence from randomized controlled trials for safety and efficacy of glucose lowering medications is available for this population. Purpose To characterize the effects of linagliptin on net CV disease and hospitalization burden in this population. Methods People with T2D and either i) urine albumin creatinine ratio (UACR) >30 mg/g with concomitant CV disease, or ii) estimated glomerular filtration rate (eGFR) 200 mg/g, were randomized to linagliptin 5 mg or placebo q.d. in a double-blind fashion in addition to standard of care. We assessed the effects of linagliptin versus placebo on all first plus recurrent CV events and all-cause hospitalizations using a using a negative binomial model to account for within-subject correlation. Results A total of 6979 participants were enrolled (mean age 66 years, 63% male, eGFR 54.6 ml/min/1.73m2, median UACR 162 mg/g, 59% with history of ischemic heart disease, 27% with history of heart failure (HF)) and followed for a median of 2.2 years. Adding recurrent events increased the number of events for analysis from 5.3–57.5% across CV/HF outcomes and 112.4% for hospitalizations. In analyses of first plus recurrent events, the event rate ratio (95% CI) with linagliptin versus placebo was 0.98 (0.82, 1.16; p=0.78) for 3-point MACE, 1.03 (0.79, 1.35; p=0.83) for myocardial infarction 1.03 (0.83, 1.29; p=0.77) for myocardial infarction plus revascularization, 0.89 (0.65, 1.22; p=0.48) for stroke, 0.94 (0.70, 1.27; p=0.69) for stroke plus TIA, 0.94 (0.75, 1.20; p=0.63) for hospitalized HF, 0.92 (0.77, 1.11; p=0.40) for the composite of CV death or hospitalized HF, and 0.96 (0.87, 1.06; p=0.40) for all-cause hospitalization (Figure). Conclusion Linagliptin showed similar risk of either first or recurrent CV or hospitalization events compared with placebo in patients with T2D and cardiorenal disease. These data support the CV safety of linagliptin and, considering the high volume of recurrent events, underscores the significant CV disease burden experienced by patients with T2D and cardiorenal disease Acknowledgement/Funding Boehringer Ingelheim and Eli Lilly

Background The cardiovascular (CV) benefits of sodium-glucose co-transporter-2 (SGLT2) inhibitors in patients with type 2 diabetes (T2D) have been demonstrated in long-term clinical trials. In the EMPA-REG OUTCOME trial, the SGLT2 inhibitor empagliflozin (EMPA), compared with placebo (PBO), significantly reduced the risk of CV death and hospitalisation for heart failure (HHF) in patients with T2D and established CV disease, with a median follow-up time of 3.1 years. Purpose To investigate the early benefits and safety associated with use of EMPA in patients enrolled in the EMPA-REG OUTCOME trial according to heart failure (HF) status at baseline. Methods We evaluated the effects of treatments on glycated haemoglobin (HbA1c) levels and on the clinical endpoints of HHF, HHF or CV death, and HHF or all-cause mortality (ACM), as well as the occurrence of adverse events (AEs), at 12 weeks, 6 months, and 1 year after randomisation. Outcomes data were explored descriptively at 12 weeks, and assessed by Cox regression models adjusting for baseline risk factors at 6 months, and 1 year, whereas safety data were explored descriptively. Effects on HbA1c were evaluated using a Mixed Model Repeated Measures (MMRM) model. Results A total of 7020 participants, 706 (10%) with investigator-reported HF at baseline, were randomised to PBO, or two different doses of EMPA (10 mg or 25 mg once daily). In patients with HF at baseline, the adjusted mean differences in HbA1c between pooled EMPA and PBO at 12 weeks, 6 months, and 1 year after randomisation were −0.55, −0.54 and −0.53%-point, respectively, p Conclusions In the EMPA-REG OUTCOME trial, EMPA led to early beneficial effects on morbidity and mortality outcomes in patients with T2D with or without HF, which were not offset by an increased risk of AEs.

Background In type 2 diabetes, the temporal proximity of an atherosclerotic cardiovascular (CV) event can impact prognosis, but whether timing influences sodium glucose co-transporter 2 inhibitor effects is unknown. We explored the association of time from last qualifying CV event before randomisation (myocardial infarction [MI], stroke, coronary artery disease or peripheral arterial disease) with CV outcomes and benefit of empagliflozin (EMPA) in EMPA-REG OUTCOME. Methods Patients (pts) were randomised to EMPA 10 mg, 25 mg or placebo and followed for 3.1 years (median). Risk of major adverse CV events (3P MACE: CV death, MI, stroke), CV death or hospitalisation for heart failure (HHF) were evaluated using Cox regression in subgroups of ≤1/>1 year since last qualifying CV event. Qualifying event stratification was possible in 6796 (97%) pts. Results In the overall population, N=6796 (4547 EMPA and 2249 placebo pts), median (Q1, Q3) time from last CV event was 3.8 (1.5–7.6) years. Overall, 1214 (EMPA 841; placebo 373) and 5582 (EMPA 3706; placebo 1876) pts had a last qualifying CV event ≤1 and >1 year, respectively. Pts with more recent events had similar risk for CV outcomes compared with pts >1 year from qualifying event (Figure). Moreover, the benefit of EMPA on CV outcomes was consistent between pts enrolled ≤1 or >1 year from the qualifying CV event (all p-interaction >0.05; Figure). Conclusion Although most pts had a qualifying CV event >1 year before randomisation in EMPA-REG OUTCOME, the benefits of EMPA appear to extend to pts with more recent CV events. Acknowledgement/Funding Boehringer Ingelheim & Eli Lilly and Company Diabetes Alliance

In the EMPA‐REG OUTCOME trial, we explored the association between pre‐randomization uric acid level tertile (0.05. Further investigation of these relationships is required.

A new catalytic radical-polar crossover annulation between two unsaturated carbonyl compounds is described. The annulation proceeds under exceptionally mild conditions and provides direct and expedient access to complex terpenoid motifs. Application of this chemistry allows for synthesis of forskolin, a densely functionalized terpenoid, in 14 steps from commercially available material.

Markers of GV have been associated with CV outcomes in type 2 diabetes (T2D), however, the association of GV with HF outcomes remains unclear. In EMPA-REG OUTCOME, empagliflozin reduced the risk of CV death by 38% and HF hospitalizations (HHF) by 35%. Here we explore the association between GV and CV death/HHF outcomes. In EMPA-REG OUTCOME, 7,020 patients with T2D and established CV disease received placebo (PBO), empagliflozin (EMPA) 10 mg or 25 mg and were observed for a median period of 3.1 years. We defined within-patient GV as range and standard deviation of HbA1c measurements on-treatment until week 28. The associations with CV death, first HHF, and HHF or CV death were explored in PBO and pooled EMPA arms separately with landmark (LM) analyses at week 28 by Cox regression models adjusted for baseline risk factors including changes in HbA1c from baseline to week 12. GV levels were lower with EMPA (Table). In the LM analysis, higher GV increased the risk for CV death and CV death or HHF in both treatment arms. For HHF, no significant association could be shown (Table). In EMPA-REG OUTCOME, EMPA reduced GV. An increase in GV correlated with increased risk of subsequent CV death and CV death or HHF. The association with HHF alone was not significant maybe due to a lower numbers of HHF events, or the assumption that the effects of EMPA on HHF are largely glucose-independent. Disclosure A. Ceriello: Advisory Panel; Self; Abbott Laboratories, Eli Lilly and Company, Janssen Pharmaceuticals, Inc., Novo Nordisk A/S, Vifor Pharma. Research Support; Self; Mitsubishi Tanabe Pharma Corporation. Speaker's Bureau; Self; A. Menarini Diagnostics, AstraZeneca, Berlin-Chemie AG, Boehringer Ingelheim Pharmaceuticals, Inc. A. Ofstad: Employee; Self; Boehringer Ingelheim International GmbH. I. Zwiener: Employee; Self; Boehringer Ingelheim Pharma GmbH & Co KG, Merck KGaA. S. Kaspers: Employee; Self; Boehringer Ingelheim International GmbH. J. George: Employee; Self; Boehringer Ingelheim International GmbH. A. Nicolucci: Consultant; Self; AstraZeneca, Eli Lilly and Company, Medtronic, Novo Nordisk A/S. Research Support; Self; Artsana S.p.A., Dexcom, Inc., Novo Nordisk A/S, Sanofi-Aventis. Funding Boehringer Ingelheim; Eli Lilly and Company

Spoilage of cassava root begins immediately after harvest, but its shelf-life could be enhanced by adopting freezing as a storage method. This study investigated the physicochemical properties and morphology of starch isolated from cassava roots frozen for 0, 7, 14, 21 and 28 days. Extracted starches can be categorized as compound starches with most granules irregularly shaped, with some oval, round and truncated. The amylose contents (22.05–26.41%) decreased with an increase in the freezing time, but the starches showed similar crystallinity pattern (Type A). Fourier infrared transform spectroscopy showed a reduction in double-helical order structure of starches from frozen cassava roots. Starches from the stored roots were generally less firm, less sticky, more cohesive and had higher peak and trough viscosities compared to starch from freshly harvested roots. Starch from frozen cassava starch may be suited for use in certain types of noodles, such as Japanese noodles due to low amylose content. Future studies are however, required to explore the starches in food applications.

AE4 is an anion exchanger known to counter transport Cl− and HCO3− across secretory cell membranes. Peña-Münzenmayer et al. show that AE4 is an electroneutral exchanger and that it also transports monovalent cations in the same direction as HCO3−., Ae4 (Slc4a9) belongs to the Slc4a family of Cl−/HCO3− exchangers and Na+-HCO3− cotransporters, but its ion transport cycle is poorly understood. In this study, we find that native Ae4 activity in mouse salivary gland acinar cells supports Na+-dependent Cl−/HCO3− exchange that is comparable with that obtained upon heterologous expression of mouse Ae4 and human AE4 in CHO-K1 cells. Additionally, whole cell recordings and ion concentration measurements demonstrate that Na+ is transported by Ae4 in the same direction as HCO3− (and opposite to that of Cl−) and that ion transport is not associated with changes in membrane potential. We also find that Ae4 can mediate Na+-HCO3− cotransport–like activity under Cl−-free conditions. However, whole cell recordings show that this apparent Na+-HCO3− cotransport activity is in fact electroneutral HCO3−/Na+-HCO3− exchange. Although the Ae4 anion exchanger is thought to regulate intracellular Cl− concentration in exocrine gland acinar cells, our thermodynamic calculations predict that the intracellular Na+, Cl−, and HCO3− concentrations required for Ae4-mediated Cl− influx differ markedly from those reported for acinar secretory cells at rest or under sustained stimulation. Given that K+ ions share many properties with Na+ ions and reach intracellular concentrations of 140–150 mM (essentially the same as extracellular [Na+]), we hypothesize that Ae4 could mediate K+-dependent Cl−/HCO3− exchange. Indeed, we find that Ae4 mediates Cl−/HCO3− exchange activity in the presence of K+ as well as Cs+, Li+, and Rb+. In summary, our results strongly suggest that Ae4 is an electroneutral Cl−/nonselective cation–HCO3− exchanger. We postulate that the physiological role of Ae4 in secretory cells is to promote Cl− influx in exchange for K+(Na+) and HCO3− ions.

Tumor microenvironments contain multiple cell types interacting among one another via different signaling pathways. Furthermore, both cancer cells and different immune cells can display phenotypic plasticity in response to these communicating signals, thereby leading to complex spatiotemporal patterns that can impact therapeutic response. Here, we investigate the crosstalk between cancer cells and macrophages in a tumor microenvironment through in silico (computational) co-culture models. In particular, we investigate how macrophages of different polarization (M1 vs. M2) can interact with epithelial-mesenchymal plasticity of cancer cells, and conversely, how cancer cells exhibiting different phenotypes (epithelial vs. mesenchymal) can influence the polarization of macrophages. Based on interactions documented in the literature, an interaction network of cancer cells and macrophages is constructed. The steady states of the network are then analyzed. Various interactions were removed or added into the constructed-network to test the functions of those interactions. Also, parameters in the mathematical models were varied to explore their effects on the steady states of the network. In general, the interactions between cancer cells and macrophages can give rise to multiple stable steady-states for a given set of parameters and each steady state is stable against perturbations. Importantly, we show that the system can often reach one type of stable steady states where cancer cells go extinct. Our results may help inform efficient therapeutic strategies.

Polyvascular disease increases the risk for atherosclerotic cardiovascular (CV) events, but this is not well investigated in people with type 2 diabetes (T2D). Moreover, as it is unknown whether the number of vascular beds (VBs) involved is a risk marker also for hospitalization for heart failure (

Teduglutide, a glucagon-like peptide 2 analog, has demonstrated efficacy in treating adult patients with short bowel syndrome (SBS) and dependence on parenteral nutrition (PN), but its role in chronic malabsorptive states that do not necessitate PN remains uncertain. To evaluate teduglutide use beyond its approved indications and to discuss the results of this adjunctive treatment in patients resistant to established therapy. This series reports four patients treated with teduglutide off-label. The first case had Crohn’s disease (CD) with persistent colocutaneous fistulae that demonstrated complete closure after 8 months of teduglutide therapy. The second case involved a PN-dependent CD patient with persistent fistulae and intra-abdominal abscesses who weaned off PN and had a significant improvement in her nutritional status after 3 months of teduglutide therapy. The third case had CD complicated by severe malnutrition and previous PN-associated line infections, but by 9 months of teduglutide therapy, she gained 5 kg and no longer required re-initiation of PN. The fourth case had a high-output diverting ileostomy with resultant impaired healing of a stage IV decubitus ulcer, and after 2 months of therapy, the patient’s pre-albumin increased by 250% and the ulcer had decreased by 40% in size. The use of teduglutide might be broadened to include patients with functional SBS not meeting strict criteria for intestinal failure. Further studies should evaluate the efficacy of teduglutide in patients who may require short-term small intestine rehabilitation or who have chronically impaired absorptive capacity not yet requiring PN.