Engineering allorejection-resistant CAR-NKT cells from hematopoietic stem cells for off-the-shelf cancer immunotherapy.

The clinical potential of current FDA-approved chimeric antigen receptor (CAR)-engineered T (CAR-T) cell therapy is encumbered by its autologous nature, which presents notable challenges related to manufacturing complexities, heightened costs, and limitations in patient selection. Therefore, there is a growing demand for off-the-shelf universal cell therapies. In this study, we have generated universal CAR-engineered NKT (UCAR-NKT) cells by integrating iNKT TCR engineering and HLA gene editing on hematopoietic stem cells (HSCs), along with an ex vivo, feeder-free HSC differentiation culture. The UCAR-NKT cells are produced with high yield, purity, and robustness, and they display a stable HLA-ablated phenotype that enables resistance to host cell-mediated allorejection. These UCAR-NKT cells exhibit potent antitumor efficacy to blood cancers and solid tumors, both in vitro and in vivo, employing a multifaceted array of tumor-targeting mechanisms. These cells are further capable of altering the tumor microenvironment by selectively depleting immunosuppressive tumor-associated macrophages and myeloid-derived suppressor cells. In addition, UCAR-NKT cells demonstrate a favorable safety profile with low risks of graft-versus-host disease and cytokine release syndrome. Collectively, these preclinical studies underscore the feasibility and significant therapeutic potential of UCAR-NKT cell products and lay a foundation for their translational and clinical development.

The effect of renal function on the pharmacokinetics and pharmacodynamics of enavogliflozin, a potent and selective sodium-glucose cotransporter-2 inhibitor, in type 2 diabetes.

AIMS: To explore the effect of renal function on the pharmacokinetic (PK) and pharmacodynamic (PD) profile and safety of enavogliflozin, a selective sodium-glucose cotransporter 2 (SGLT2) inhibitor, in patients with type 2 diabetes mellitus (T2DM). METHODS: An open-label, two-part clinical trial was conducted in T2DM patients, stratified by renal function: Group 1, normal renal function; Group 2, mild renal impairment (RI); Group 3, moderate RI; and Group 4, severe RI. In Part A, Groups 2 and 4 received enavogliflozin 0.5 mg once. In Part B, Groups 1 and 3 received enavogliflozin 0.5 mg once daily for 7 days. Serial blood and timed urine samples were collected to analyse the PK and PD characteristics of enavogliflozin. Pearson's correlation coefficients were calculated to assess the correlations between PK or PD parameters and creatinine clearance (CrCL). RESULTS: A total of 21 patients completed the study as planned. The area under the curve (AUC) for enavogliflozin was not significantly correlated with CrCL, although the maximum concentration slightly decreased as renal function decreased. By contrast, daily urinary glucose excretion (UGE) was positively correlated with CrCL after both single- (r = 0.7866, p < 0.0001) and multiple-dose administration (r = 0.6606, p = 0.0438). CONCLUSIONS: Systemic exposure to oral enavogliflozin 0.5 mg was similar among the patients with T2DM regardless of their renal function levels. However, the glucosuric effect of enavogliflozin decreased with RI. Considering the UGE observed and approved therapeutic use of other SGLT2 inhibitors, the efficacy of enavogliflozin with regard to glycaemic control could be explored in patients with mild and moderate RI (estimated glomerular filtration rate ≥30 or ≥45 mL/min/1.73 m2) in a subsequent larger study.

A 52-week efficacy and safety study of enavogliflozin versus dapagliflozin as an add-on to metformin in patients with type 2 diabetes mellitus: ENHANCE-M extension study.

AIM: To evaluate the long-term safety and efficacy of enavogliflozin 0.3 mg/day added to metformin in patients with type 2 diabetes mellitus. MATERIALS AND METHODS: After 24 weeks of a randomized, double-blind treatment period with enavogliflozin 0.3 mg/day (n = 101) or dapagliflozin 10 mg/day (n = 99) added to metformin, all patients received enavogliflozin 0.3 mg/day plus metformin for an additional 28 weeks during the open-label extension period. RESULTS: Eighty-two patients continued enavogliflozin (maintenance group), and 77 were switched from dapagliflozin to enavogliflozin (switch group). All adverse drug reactions (ADR) were mild in severity. In the maintenance group, ADRs (cystitis and vaginal infection) were reported in two patients (2.44%) during 52 weeks. In the switch group, ADR (hypoglycaemia) was reported in one patient (1.30%) during a 28-week open-label extension period. At week 52, glycated haemoglobin and fasting plasma glucose were significantly lower than at the baseline, by 0.85% and 29.08 mg/dl, respectively, in the maintenance group (p < .0001 for both), and by 0.81% and 32.77 mg/dl, respectively, in the switch group (p < .0001 for both). At week 52, 68.92% of patients from the maintenance group and 64.29% from the switch group achieved glycated haemoglobin <7%. A significant increase in the urine glucose-creatinine ratio was observed at week 52, by 58.81 g/g and 63.77 g/g in the maintenance and switch groups, respectively (p < .0001). CONCLUSIONS: Enavogliflozin added to metformin was tolerated well for up to 52 weeks and provided continual glycaemic control in type 2 diabetes mellitus, along with a significant increase in the urine glucose-creatinine ratio.

Exercise mitigates flow recirculation and activates metabolic transducer SCD1 to catalyze vascular protective metabolites.

Exercise promotes pulsatile shear stress in the arterial circulation and ameliorates cardiometabolic diseases. However, exercise-mediated metabolic transducers for vascular protection remain under-investigated. Untargeted metabolomic analysis demonstrated that wild-type mice undergoing voluntary wheel running exercise expressed increased endothelial stearoyl-CoA desaturase 1 (SCD1) that catalyzes anti-inflammatory lipid metabolites, namely, oleic (OA) and palmitoleic acids (PA), to mitigate NF-κB-mediated inflammatory responses. In silico analysis revealed that exercise augmented time-averaged wall shear stress but mitigated flow recirculation and oscillatory shear index in the lesser curvature of the mouse aortic arch. Following exercise, endothelial Scd1-deleted mice (Ldlr-/- Scd1EC-/-) on high-fat diet developed persistent VCAM1-positive endothelium in the lesser curvature and the descending aorta, whereas SCD1 overexpression via adenovirus transfection mitigated endoplasmic reticulum stress and inflammatory biomarkers. Single-cell transcriptomics of the aorta identified Scd1-positive and Vcam1-negative endothelial subclusters interacting with other candidate genes. Thus, exercise mitigates flow recirculation and activates endothelial SCD1 to catalyze OA and PA for vascular endothelial protection.

Subchronic inhalation exposure to ultrafine particulate matter alters the intestinal microbiome in various mouse models.

Exposure to ultrafine particles (UFPs) has been associated with multiple adverse health effects. Inhaled UFPs could reach the gastrointestinal tract and influence the composition of the gut microbiome. We have previously shown that oral ingestion of UFPs alters the gut microbiome and promotes intestinal inflammation in hyperlipidemic Ldlr-/- mice. Particulate matter (PM)2.5 inhalation studies have also demonstrated microbiome shifts in normolipidemic C57BL/6 mice. However, it is not known whether changes in microbiome precede or follow inflammatory effects in the intestinal mucosa. We hypothesized that inhaled UFPs modulate the gut microbiome prior to the development of intestinal inflammation. We studied the effects of UFP inhalation on the gut microbiome and intestinal mucosa in two hyperlipidemic mouse models (ApoE-/- mice and Ldlr-/- mice) and normolipidemic C57BL/6 mice. Mice were exposed to PM in the ultrafine-size range by inhalation for 6 h a day, 3 times a week for 10 weeks at a concentration of 300-350 µg/m3.16S rRNA gene sequencing was performed to characterize sequential changes in the fecal microbiome during exposures, and changes in the intestinal microbiome at the end. PM exposure led to progressive differentiation of the microbiota over time, associated with increased fecal microbial richness and evenness, altered microbial composition, and differentially abundant microbes by week 10 depending on the mouse model. Cross-sectional analysis of the small intestinal microbiome at week 10 showed significant changes in α-diversity, ß-diversity, and abundances of individual microbial taxa in the two hyperlipidemic models. These alterations of the intestinal microbiome were not accompanied, and therefore could not be caused, by increased intestinal inflammation as determined by histological analysis of small and large intestine, cytokine gene expression, and levels of fecal lipocalin. In conclusion, 10-week inhalation exposures to UFPs induced taxonomic changes in the microbiome of various animal models in the absence of intestinal inflammation.

Phenolic changes in a combined herbal extract of Lithospermum erythrorhizon, Houttuynia cordata, and Spirodela polyrhiza and alleviation of DNCB-induced atopic dermatitis in BALB/c mice.

Atopic dermatitis (AD) is an inflammatory skin disease showing skin barrier dysfunction, eczematous lesions, severe itching, and abnormal immune responses. The aim of this study was to determine whether an herb combination of Lithospermum erythrorhizon (LE), Houttuynia cordata (HC), and Spirodela polyrhiza (SP) has a superior anti-AD effect. Forty-two compounds were identified in LE, HC, SP, and a combined herb extract of LE, HC, and SP (LHS) using ultra-high-pressure liquid chromatography (UHPLC)-Orbitrap mass spectrometer (MS). The concentration of flavonoid glycosides including orientin (luteolin-8-C-glucoside), quercetin-3-O-rhamnoside, and luteolin-7-O-glucoside in the LHS was increased than in individual extracts. Furthermore, the treatment of LHS most effectively inhibited the increase of epidermal thickness, the number of mast cells, and the release of immunoglobulin E compared with that with each extract. These results suggest that the potential anti-AD effects of the LHS are due to the changes of bioactive compounds by the combination of herbs. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-023-01329-7.

Integrating 4-D light-sheet fluorescence microscopy and genetic zebrafish system to investigate ambient pollutants-mediated toxicity.

Ambient air pollutants, including PM2.5 (aerodynamic diameter d ~2.5 µm), PM10 (d ~10 µm), and ultrafine particles (UFP: d < 0.1 µm) impart both short- and long-term toxicity to various organs, including cardiopulmonary, central nervous, and gastrointestinal systems. While rodents have been the principal animal model to elucidate air pollution-mediated organ dysfunction, zebrafish (Danio rerio) is genetically tractable for its short husbandry and life cycle to study ambient pollutants. Its electrocardiogram (ECG) resembles that of humans, and the fluorescent reporter-labeled tissues in the zebrafish system allow for screening a host of ambient pollutants that impair cardiovascular development, organ regeneration, and gut-vascular barriers. In parallel, the high spatiotemporal resolution of light-sheet fluorescence microscopy (LSFM) enables investigators to take advantage of the transparent zebrafish embryos and genetically labeled fluorescent reporters for imaging the dynamic cardiac structure and function at a single-cell resolution. In this context, our review highlights the integrated strengths of the genetic zebrafish system and LSFM for high-resolution and high-throughput investigation of ambient pollutants-mediated cardiac and intestinal toxicity.

Gemcitabine and rapamycin-loaded mixed polymeric thermogel for metastatic pancreatic cancer therapy.

Pancreatic ductal adenocarcinoma (PDAC) is the 4th leading cause of cancer-related death and has a poor 5-year overall survival. The superior therapeutic benefits of combination or co-administration of drugs as intraperitoneal chemotherapy have increased interest in developing strategies to deliver chemotherapeutic agents to patients safely. In this study, we prepared a gel comprising the thermosensitive poly(lactide-co-glycolide)-b-poly(ethylene glycol)-b-poly(lactide-co-glycolide) (PLGA-PEG-PLGA) polymer and gemcitabine (GEM), which is currently used as the primary chemotherapy for PDAC and rapamycin (RAPA), a mammalian TOR (mTOR) inhibitor, to deliver the drug through intraperitoneal injection. We performed in vitro cytotoxicity experiments to verify the synergistic effects of the two drugs at different molar ratios and characterized the physicochemical properties of the GEM, RAPA, and GEM/RAPA-loaded thermosensitive PLGA-PEG-PLGA gels, hereafter referred to as (g(G), g(R), and g(GR)), respectively. The g(GR) comprising PLGA-PEG-PLGA polymer (25% w/v) and GEM and RAPA at a molar ratio of 11:1 showed synergism and was optimized. An in vitro cytotoxicity assay was performed by treating Panc-1-luc2 tumor spheroids with g(G), g(R), or g(GR). The g(GR) treatment group showed a 2.75-fold higher inhibition rate than the non-treated (NT) and vehicle-treated groups. Furthermore, in vivo drug release assay in mice by intraperitoneal injection of g(G), g(R), or g(GR) showed a more rapid release rate of GEM than RAPA, similar to the in vitro release pattern. The drugs in the gel were released faster in vivo than in vitro and degraded in 48 h. In addition, g(GR) showed the highest anti-tumor efficacy with no toxicity to mice. These results provide evidence for the safety and efficacy of g(GR) for intraperitoneal drug delivery. This study will assist in developing and clinically administering topical anti-cancer formulations.

Quantitative 4D imaging of biomechanical regulation of ventricular growth and maturation.

Abnormal cardiac development is intimately associated with congenital heart disease. During development, a sponge-like network of muscle fibers in the endocardium, known as trabeculation, becomes compacted. Biomechanical forces regulate myocardial differentiation and proliferation to form trabeculation, while the molecular mechanism is still enigmatic. Biomechanical forces, including intracardiac hemodynamic flow and myocardial contractile force, activate a host of molecular signaling pathways to mediate cardiac morphogenesis. While mechanotransduction pathways to initiate ventricular trabeculation is well studied, deciphering the relative importance of hemodynamic shear vs. mechanical contractile forces to modulate the transition from trabeculation to compaction requires advanced imaging tools and genetically tractable animal models. For these reasons, the advent of 4-D multi-scale light-sheet imaging and complementary multiplex live imaging via micro-CT in the beating zebrafish heart and live chick embryos respectively. Thus, this review highlights the complementary animal models and advanced imaging needed to elucidate the mechanotransduction underlying cardiac ventricular development.

Umbelliferone Ameliorates Memory Impairment and Enhances Hippocampal Synaptic Plasticity in Scopolamine-Induced Rat Model.

Alzheimer's disease (AD) is a neurodegenerative disorder, characterized by memory loss and cognitive decline. Among the suggested pathogenic mechanisms of AD, the cholinergic hypothesis proposes that AD symptoms are a result of reduced synthesis of acetylcholine (ACh). A non-selective antagonist of the muscarinic ACh receptor, scopolamine (SCOP) induced cognitive impairment in rodents. Umbelliferone (UMB) is a Apiaceae-family-derived 7-hydeoxycoumarin known for its antioxidant, anti-tumor, anticancer, anti-inflammatory, antibacterial, antimicrobial, and antidiabetic properties. However, the effects of UMB on the electrophysiological and ultrastructure morphological aspects of learning and memory are still not well-established. Thus, we investigated the effect of UMB treatment on cognitive behaviors and used organotypic hippocampal slice cultures for long-term potentiation (LTP) and the hippocampal synaptic ultrastructure. A hippocampal tissue analysis revealed that UMB attenuated a SCOP-induced blockade of field excitatory post-synaptic potential (fEPSP) activity and ameliorated the impairment of LTP by the NMDA and AMPA receptor antagonists. UMB also enhanced the hippocampal synaptic vesicle density on the synaptic ultrastructure. Furthermore, behavioral tests on male SD rats (7-8 weeks old) using the Y-maze test, passive avoidance test (PA), and Morris water maze test (MWM) showed that UMB recovered learning and memory deficits by SCOP. These cognitive improvements were in association with the enhanced expression of BDNF, TrkB, and the pCREB/CREB ratio and the suppression of acetylcholinesterase activity. The current findings indicate that UMB may be an effective neuroprotective reagent applicable for improving learning and memory against AD.

Exercise Mitigates Flow Recirculation and Activates Mechanosensitive Transcriptome to Uncover Endothelial SCD1-Catalyzed Anti-Inflammatory Metabolites.

Exercise modulates vascular plasticity in multiple organ systems; however, the metabolomic transducers underlying exercise and vascular protection in the disturbed flow-prone vasculature remain under-investigated. We simulated exercise-augmented pulsatile shear stress (PSS) to mitigate flow recirculation in the lesser curvature of the aortic arch. When human aortic endothelial cells (HAECs) were subjected to PSS ( τ ave = 50 dyne·cm -2 , ∂τ/∂t = 71 dyne·cm -2 ·s -1 , 1 Hz), untargeted metabolomic analysis revealed that Stearoyl-CoA Desaturase (SCD1) in the endoplasmic reticulum (ER) catalyzed the fatty acid metabolite, oleic acid (OA), to mitigate inflammatory mediators. Following 24 hours of exercise, wild-type C57BL/6J mice developed elevated SCD1-catalyzed lipid metabolites in the plasma, including OA and palmitoleic acid (PA). Exercise over a 2-week period increased endothelial SCD1 in the ER. Exercise further modulated the time-averaged wall shear stress (TAWSS or τ ave) and oscillatory shear index (OSI ave ), upregulated Scd1 and attenuated VCAM1 expression in the disturbed flow-prone aortic arch in Ldlr -/- mice on high-fat diet but not in Ldlr -/- Scd1 EC-/- mice. Scd1 overexpression via recombinant adenovirus also mitigated ER stress. Single cell transcriptomic analysis of the mouse aorta revealed interconnection of Scd1 with mechanosensitive genes, namely Irs2 , Acox1 and Adipor2 that modulate lipid metabolism pathways. Taken together, exercise modulates PSS ( τ ave and OSI ave ) to activate SCD1 as a metabolomic transducer to ameliorate inflammation in the disturbed flow-prone vasculature.

Ductal delivery of extracellular vesicles promote the recovery from salivary gland inflammation.

Salivary gland dysfunction worsens the quality of life, but treatment for restoration of salivary gland function is limited. Although previous reports have demonstrated the therapeutic potentials of extracellular vesicles (EVs) in different preclinical models, the role of EVs in salivary glands remains elusive. Furthermore, little is known about the roles of salivary gland-derived EVs in tissue repair or regeneration compared to other EVs. In this study, EVs secreted from salivary gland-derived mesenchymal stem cells (sgMSCs) were comparatively analyzed with those from Wharton's jelly-derived MSC (wjMSCs). sgMSCs secreted more significant amounts of EVs than wjMSCs, and salivary gland epithelial cells showed a more efficient uptake of sgMSC-EVs than wjMSC-EVs. The possibility of immune regulation was tested via macrophage polarization and LPS-induced epithelial inflammation, resulting in an M1-to-M2 shift and reversal of acinar-to-ductal metaplasia by sgMSC-EV. Furthermore, the roles of sgMSC-EV-mediated immune regulation and tissue repair were clarified in vivo via retroductal delivery of sgMSC-EVs in a mouse model of obstructive sialadenitis. Collectively, our data demonstrate the superior role of sgMSC-EVs in the recovery from salivary gland inflammation and injury and suggest EVs as therapeutic tools for salivary gland dysfunction.

Pharmacokinetic and pharmacodynamic interaction between DWP16001, an sodium-glucose cotransporter 2 inhibitor and metformin in healthy subjects.

AIMS: DWP16001 is a novel sodium-glucose cotransporter 2 inhibitor for the treatment of type 2 diabetes with selective and sustained sodium-glucose cotransporter 2 inhibition. We aimed to evaluate whether the coadministration of DWP16001 and metformin causes any changes in pharmacokinetics (PK) or pharmacodynamics (PD). METHODS: A randomized, open-label, single- and multiple-dose, 2-sequence, crossover study was conducted in healthy male subjects. Subjects received the following treatments: a single oral dose of DWP16001 (DWP) 2 mg, metformin immediate release 1000 mg (MET) twice daily for 7 days and a single oral dose of DWP and MET at steady-state for metformin (DWP+MET). Serial blood and interval urine were collected for PK and PD analyses. Safety and tolerability profiles were assessed throughout the study. RESULTS: DWP+MET displayed increased peak concentration and area under the concentration-time curve from time 0 to time of the last quantifiable concentration compared with DWP (per standard bioequivalence boundaries, 0.8-1.25); the geometric mean ratios (GMRs) and their 90% confidence intervals (CIs) were 1.22 (1.13-1.31) and 1.09 (1.05-1.14), respectively. DWP+MET and MET showed similar peak concentration and area under the concentration-time curve within a dosing interval at steady state for metformin; the GMRs and 90% CIs were 0.98 (0.90-1.06) and 1.05 (0.98-1.13), respectively. The amount of urinary glucose excretion from time 0 to 144 h was also comparable between DWP+MET and DWP (GMR and 90% CI; 0.99, 0.94-1.05). CONCLUSION: The results suggest that DWP16001 and metformin could be coadministered without clinically relevant PK and PD interactions.

Naringin enhances long-term potentiation and recovers learning and memory deficits of amyloid-beta induced Alzheimer's disease-like behavioral rat model.

Alzheimer's disease (AD), as the most typical type of dementia, is a chronic neurodegenerative disorder characterized by progressive learning and memory impairment. It is known that the main causes of AD are the accumulation of ß-amyloid (Aß) plaques and neurofibrillary tangles (NFT) containing hyperphosphorylated tau protein. Naringin is a flavonoid from citrus fruits, especially in grapefruit, which has anti-inflammatory, antioxidant, anti-apoptotic, and neuroprotective activities. However, the effect of naringin in AD caused by Aß has not been clearly studied, and there are few studies on the electrophysiological aspect. Thus, we investigated the ex vivo neuroprotective effect of naringin through the long-term potentiation (LTP) on organotypic hippocampal slice cultures. We evaluated the in vivo effects of naringin (100 mg/kg/day) orally treated for 20 days on learning, memory, and cognition which was impaired by bilateral CA1 subregion injection of Aß. Cognitive behaviors were measured 2 weeks after Aß injection using behavioral tests and the hippocampal expression of apoptotic and neurotrophic regulators were measured by immunoblotting. In hippocampal tissue slices, naringin dose-dependently increased the field excitatory postsynaptic potential (fEPSP) after theta burst stimulation and attenuated Aß-induced blockade of fEPSP in the hippocampal CA1 area. In Aß injected rats, naringin improved object recognition memory in the novel object test, avoidance memory in the passive avoidance test and spatial recognition memory in the Morris water maze test. In the hippocampus, naringin attenuated the Aß-induced cyclooxygenase-2, Bax activation and Bcl-2, CREB, BDNF and TrkB inhibition. These results suggest that naringin has therapeutic potential to reduce neuronal inflammation and apoptosis induced by Aß related with the BDNF/TrkB/CREB signaling.

Treatment Patterns, Clinical Outcomes and Health Care Resource Utilisation in Patients with EGFR-mutated Metastatic Non-Small Cell Lung Cancer: A Real-World Study in South Korea.

BACKGROUND: Despite the dynamic treatment landscape for EGFR mutant-positive metastatic non-small cell lung cancer (EGFRm+ mNSCLC), most of the earlier studies have focused on US or Western populations. OBJECTIVE: The objective of this study was to explore real-world treatment patterns and outcomes of South Korean patients with EGFRm+ mNSCLC. METHODS: Retrospective chart review of adult patients with EGFRm+ mNSCLC who received systemic treatment between January-2019 and June-2019. RESULTS: A total of 162 patients were included from 21 hospitals, with a median follow-up of 15.6 months. Median age was 65.0 years, 22% had central nervous system metastasis, and 57% and 38% had exon 19 deletion and exon 21 L858R, respectively. Among 144 patients (89%) who received first-line EGFR-tyrosine kinase inhibitor, afatinib was most the common (44%), followed by gefitinib (28%) and erlotinib (13%). First-line chemotherapy was more common when an EGFR-mutation was detected after versus before first-line treatment initiation (31% vs 5%). Discontinuation of first-line treatment was mostly due to disease-progression (81%) and toxicity (7%). Among 58 (78%) patients who received second-line treatment, osimertinib was the most common (40%). Most (60%) patients reported ≥1 Grade ≥3 adverse event during first-line treatment. Following initiation of first-line treatment, physician visits and chest X-rays were the most frequent healthcare utilisation events. Rates of emergency-room visits and hospitalization were 12% and 16%, respectively, with a mean length-of-stay of 10.4 days. At 12 months, overall survival rate was 95%, and numerically worse for patients with exon 21 versus 19 mutations. CONCLUSIONS: Characteristics and clinical outcomes of Korean patients with EGFRm+ mNSCLC in real-world practice were comparable to those observed in clinical trials. As osimertinib was not reimbursed for first-line treatment before study completion, further investigation is warranted to explore evolving treatment practice.

Activating P2Y1 receptors improves function in arteries with repressed autophagy.

AIM: The importance of endothelial cell (EC) autophagy to vascular homeostasis in the context of health and disease is evolving. Earlier, we reported that intact EC autophagy is requisite to maintain shear-stress-induced nitric oxide (NO) generation via glycolysis-dependent purinergic signalling to endothelial NO synthase (eNOS). Here, we illustrate the translational and functional significance of these findings. METHODS AND RESULTS: First, we assessed translational relevance using older male humans and mice that exhibit blunted EC autophagy and impaired arterial function vs. adult controls. Active hyperaemia evoked by rhythmic handgrip exercise-elevated radial artery shear-rate similarly from baseline in adult and older subjects for 60 min. Compared with baseline, indexes of autophagy initiation, p-eNOSS1177 activation, and NO generation, occurred in radial artery ECs obtained from adult but not older volunteers. Regarding mice, indexes of autophagy and p-eNOSS1177 activation were robust in ECs from adult but not older animals that completed 60-min treadmill-running. Furthermore, 20 dyne • cm2 laminar shear stress × 45-min increased autophagic flux, glycolysis, ATP production, and p-eNOSS1177 in primary arterial ECs obtained from adult but not older mice. Concerning functional relevance, we next questioned whether the inability to initiate EC autophagy, glycolysis, and p-eNOSS1177in vitro precipitates arterial dysfunction ex vivo. Compromised intraluminal flow-mediated vasodilation displayed by arteries from older vs. adult mice was recapitulated in vessels from adult mice by (i) NO synthase inhibition; (ii) acute autophagy impairment using 3-methyladenine (3-MA); (iii) EC Atg3 depletion (iecAtg3KO mice); (iv) purinergic 2Y1-receptor (P2Y1-R) blockade; and (v) germline depletion of P2Y1-Rs. Importantly, P2Y1-R activation using 2-methylthio-ADP (2-Me-ADP) improved vasodilatory capacity in arteries from (i) adult mice treated with 3-MA; (ii) adult iecAtg3KO mice; and (iii) older animals with repressed EC autophagy. CONCLUSIONS: Arterial dysfunction concurrent with pharmacological, genetic, and age-associated EC autophagy compromise is improved by activating P2Y1-Rs.

Reduce, Reuse, Recycle, Run ! : 4 Rs to improve cardiac health in advanced age.

During the aging process damaged/dysfunctional proteins and organelles accumulate and contribute to organ dysfunction. Luckily, there is a conserved intracellular process to reuse and recycle these dysregulated cellular components termed macroautophagy (autophagy). Unfortunately, strong evidence indicates autophagy is compromised with aging, protein quality control is jeopardized, and resultant proteotoxicity can contribute significantly to age-associated organ dysfunction. Are there interventions that can re-establish autophagic flux that is otherwise impaired with aging? With particular regard to the heart, here we review evidence that caloric-restriction, the polyamine spermidine, and the mTOR inhibitor rapamycin, even when initiated late-in-life, restore cardiomyocyte autophagy to an extent that lessens age-associated cardiac dysfunction. Cho et al. provide a physiological intervention to this list i.e., regular physical exercise initiated late-in-life boosts cardiomyocyte autophagic flux and rejuvenates cardiac function in male mice. While this study provides strong evidence for a mechanism whereby heightened physical activity can lead to improved heart health in the context of aging, (i) only male mice were studied; (ii) the intensity of exercise-training might not be suitable for all; and (iii) mice with aging-associated comorbidities were not investigated. Nonetheless, Cho et al. provide robust evidence that a low-cost and simple behavioral intervention initiated late-in-life improves cardiomyocyte autophagic flux and rejuvenates cardiac function.

Gut Microbes Are Associated with the Vascular Beneficial Effects of Dietary Strawberry on Metabolic Syndrome-Induced Vascular Inflammation.

SCOPE: Metabolic syndrome (MetS) alters the gut microbial ecology and increases the risk of cardiovascular disease. This study investigates whether strawberry consumption reduces vascular complications in an animal model of MetS and identifies whether this effect is associated with changes in the composition of gut microbes. METHODS AND RESULTS: Seven-week-old male mice consume diets with 10% (C) or 60% kcal from fat (high-fat diet fed mice; HF) for 12 weeks and subgroups are fed a 2.35% freeze-dried strawberry supplemented diet (C+SB or HF+SB). This nutritional dose is equivalent to ≈160 g of strawberry. After 12 weeks treatment, vascular inflammation is enhanced in HF versus C mice as shown by an increased monocyte binding to vasculature, elevated serum chemokines, and increased mRNA expression of inflammatory molecules. However, strawberry supplementation suppresses vascular inflammation in HF+SB versus HF mice. Metabolic variables, blood pressure, and indices of vascular function were similar among the groups. Further, the abundance of opportunistic microbe is decreased in HF+SB. Importantly, circulating chemokines are positively associated with opportunistic microbes and negatively associated with the commensal microbes (Bifidobacterium and Facalibaculum). CONCLUSION: Dietary strawberry decreases the abundance of opportunistic microbe and this is associated with a decrease in vascular inflammation resulting from MetS.

Mucoepidermoid carcinoma of the breast: A case report and literature review focused on radiological findings.

RATIONALE: Mucoepidermoid carcinoma (MEC) of the breast is a rare entity, with an estimated incidence of only 0.2% to 0.3% of all primary breast tumors. The radiological features of breast MEC have scarcely been investigated mainly because of its rarity. In this article, we present a case of breast MEC diagnosed at our hospital and review the literature, focusing on radiological findings and radiologic-pathologic correlations that could improve clinical management of this entity. To the best of our knowledge, our study is the first review of the literature that focuses on the radiological features of breast MEC. PATIENT CONCERNS: A 47-year-old premenopausal woman presented with a painless palpable mass in the right breast. DIAGNOSIS: Mammography and ultrasonography revealed a mass with suspicious malignant features, which was categorized as Breast Imaging Reporting and Data System category 4c. A 14-gauge core-needle biopsy revealed an intermediate-grade MEC of the breast. The patient underwent breast magnetic resonance imaging and chest computed tomography for preoperative evaluation. Postoperative histopathological examination confirmed a diagnosis of intermediate-grade MEC. The clinical staging was T2N0M0. INTERVENTIONS: The patient underwent breast-conserving surgery, adjuvant chemotherapy, radiotherapy, and hormonal therapy. OUTCOMES: No evidence of recurrence has been reported over 37 months. LESSONS: The imaging characteristics of breast MEC were variable, and there were no specific radiological features for diagnosis. The presence of cystic components on radiological imaging is likely to be an indicator of a low-grade tumor and better prognosis, although the number of reported cases is limited.

Salivary gland organoid culture maintains distinct glandular properties of murine and human major salivary glands.

Salivary glands that produce and secrete saliva, which is essential for lubrication, digestion, immunity, and oral homeostasis, consist of diverse cells. The long-term maintenance of diverse salivary gland cells in organoids remains problematic. Here, we establish long-term murine and human salivary gland organoid cultures. Murine and human salivary gland organoids express gland-specific genes and proteins of acinar, myoepithelial, and duct cells, and exhibit gland functions when stimulated with neurotransmitters. Furthermore, human salivary gland organoids are established from isolated basal or luminal cells, retaining their characteristics. Single-cell RNA sequencing also indicates that human salivary gland organoids contain heterogeneous cell types and replicate glandular diversity. Our protocol also enables the generation of tumoroid cultures from benign and malignant salivary gland tumor types, in which tumor-specific gene signatures are well-conserved. In this study, we provide an experimental platform for the exploration of precision medicine in the era of tissue regeneration and anticancer treatment.