Sorbs2 Deficiency and Vascular BK Channelopathy in Diabetes.

BACKGROUND: Vascular large conductance Ca2+-activated K+ (BK) channel, composed of the α-subunit (BK-α) and the ß1-subunit (BK-ß1), is a key determinant of coronary vasorelaxation and its function is impaired in diabetic vessels. However, our knowledge of diabetic BK channel dysregulation is incomplete. The Sorbs2 (Sorbin homology [SoHo] and Src homology 3 [SH3] domains-containing protein 2), is ubiquitously expressed in arteries, but its role in vascular pathophysiology is unknown. METHODS: The role of Sorbs2 in regulating vascular BK channel activity was determined using patch-clamp recordings, molecular biological techniques, and in silico analysis. RESULTS: Sorbs2 is not only a cytoskeletal protein but also an RNA-binding protein that binds to BK channel proteins and BK-α mRNA, regulating BK channel expression and function in coronary smooth muscle cells. Molecular biological studies reveal that the SH3 domain of Sorbs2 is necessary for Sorbs2 interaction with BK-α subunits, while both the SH3 and SoHo domains of Sorbs2 interact with BK-ß1 subunits. Deletion of the SH3 or SoHo domains abolishes the Sorbs2 effect on the BK-α/BK-ß1 channel current density. Additionally, Sorbs2 is a target gene of the Nrf2 (nuclear factor erythroid-2-related factor 2), which binds to the promoter of Sorbs2 and regulates Sorbs2 expression in coronary smooth muscle cells. In vivo studies demonstrate that Sorbs2 knockout mice at 4 months of age display a significant decrease in BK channel expression and function, accompanied by impaired BK channel Ca2+-sensitivity and BK channel-mediated vasodilation in coronary arteries, without altering their body weights and blood glucose levels. Importantly, Sorbs2 expression is significantly downregulated in the coronary arteries of db/db type 2 diabetic mice. CONCLUSIONS: Sorbs2, a downstream target of Nrf2, plays an important role in regulating BK channel expression and function in vascular smooth muscle cells. Vascular Sorbs2 is downregulated in diabetes. Genetic knockout of Sorbs2 manifests coronary BK channelopathy and vasculopathy observed in diabetic mice, independent of obesity and glucotoxicity.

A real-world experience of atrioventricular synchronous pacing with leadless ventricular pacemakers.

AIMS: The MicraTM transcatheter pacing system (TPS) (Medtronic) is the only leadless pacemaker that promotes atrioventricular (AV) synchrony via accelerometer-based atrial sensing. Data regarding the real-world experience with this novel system are scarce. We sought to characterize patients undergoing MicraTM -AV implants, describe percentage AV synchrony achieved, and analyze the causes for suboptimal AV synchrony. METHODS: In this retrospective cohort study, electronic medical records from 56 consecutive patients undergoing MicraTM -AV implants at the Mayo Clinic sites in Minnesota, Florida, and Arizona with a minimum follow-up of 3 months were reviewed. Demographic data, comorbidities, echocardiographic data, and clinical outcomes were compared among patients with and without atrial synchronous ventricular pacing (AsVP) ≥ 70%. RESULTS: Sixty-five percent of patients achieved AsVP ≥ 70%. Patients with adequate AsVP had smaller body mass indices, a lower proportion of congestive heart failure, and prior cardiac surgery. Echocardiographic parameters and procedural characteristics were similar across the two groups. Active device troubleshooting was associated with higher AsVP. The likely reasons for low AsVP were small A4-wave amplitude, high ventricular pacing burden, and inadequate device reprogramming. Importantly, in patients with low AsVP, subjective clinical worsening was not noted during follow-up. CONCLUSION: With the increasing popularity of leadless pacemakers, it is paramount for device implanting teams to be familiar with common predictors of AV synchrony and troubleshooting with MicraTM -AV devices.

CRRL269.

RATIONALE: Acute kidney injury (AKI) has a high prevalence and mortality in critically ill patients. It is also a powerful risk factor for heart failure incidence driven by hemodynamic changes and neurohormonal activation. However, no drugs have been approved by the Food and Drug Administration. Endogenous pGC-A (particulate guanylyl cyclase A receptor) activators were reported to preserve renal function and improve mortality in AKI patients, although hypotension accompanied by pGC-A activators have limited their therapeutic potential. OBJECTIVE: We investigated the therapeutic potential of a nonhypotensive pGC-A activator/designer natriuretic peptide, CRRL269, in a short-term, large animal model of ischemia-induced AKI and also investigated the potential of uCNP (urinary C-type natriuretic peptide) as a biomarker for AKI. METHODS AND RESULTS: We first showed that CRRL269 stimulated cGMP generation, suppressed plasma angiotensin II, and reduced cardiac filling pressures without lowering blood pressure in the AKI canine model. We also demonstrated that CRRL269 preserved glomerular filtration rate, increased renal blood flow, and promoted diuresis and natriuresis. Further, CRRL269 reduced kidney injury and apoptosis as evidenced by ex vivo histology and tissue apoptosis analysis. We also showed, compared with native pGC-A activators, that CRRL269 is a more potent inhibitor of apoptosis in renal cells and induced less decreases in intracellular Ca2+ concentration in vascular smooth muscle cells. The renal antiapoptotic effects were at least mediated by cGMP/PKG pathway. Further, CRRL269 inhibited proapoptotic genes expression using a polymerase chain reaction gene array. Additionally, we demonstrated that AKI increased uCNP levels. CONCLUSIONS: Our study supports developing CRRL269 as a novel renocardiac protective agent for AKI treatment.

Regulation of KCNMA1 transcription by Nrf2 in coronary arterial smooth muscle cells.

The large conductance Ca2+-activated K+ (BK) channels, composed of the pore-forming α subunits (BK-α, encoded by KCNMA1 gene) and the regulatory ß1 subunits (BK-ß1, encoded by KCNMB1 gene), play a unique role in the regulation of coronary vascular tone and myocardial perfusion by linking intracellular Ca2+ homeostasis with excitation-contraction coupling in coronary arterial smooth muscle cells (SMCs). The nuclear factor erythroid 2-related factor 2 (Nrf2) belongs to a member of basic leucine zipper transcription factor family that regulates the expression of antioxidant and detoxification enzymes by binding to the antioxidant response elements (AREs) of these target genes. We have previously reported that vascular BK-ß1 protein expression was tightly regulated by Nrf2. However, the molecular mechanism underlying the regulation of BK channel expression by Nrf2, particularly at transcription level, is unknown. In this study, we hypothesized that KCNMA1 and KCNMB1 are the target genes of Nrf2 transcriptional regulation. We found that BK channel protein expression and current density were diminished in freshly isolated coronary arterial SMCs of Nrf2 knockout (KO) mice. However, BK-α mRNA expression was reduced, but not that of BK-ß1 mRNA expression, in the arteries of Nrf2 KO mice. Promoter-Nrf2 luciferase reporter assay confirmed that Nrf2 binds to the ARE of KCNMA1 promoter, but not that of KCNMB1. Adenoviral expression and pharmacological activation of Nrf2 increased BK-α and BK-ß1 protein levels and enhanced BK channel activity in coronary arterial SMCs. Hence, our results indicate that Nrf2 is a key determinant of BK channel expression and function in vascular SMCs. Nrf2 facilitates BK-α expression through a direct increase in gene transcription, whereas that on BK-ß1 is through a different mechanism.

Resolving the topological enigma in Ca2+ signaling by cyclic ADP-ribose and NAADP.

Cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) are two structurally distinct messengers that mobilize the endoplasmic and endolysosomal Ca2+ stores, respectively. Both are synthesized by the CD38 molecule (CD38), which has long been thought to be a type II membrane protein whose catalytic domain, intriguingly, faces to the outside of the cell. Accordingly, for more than 20 years, it has remained unresolved how CD38 can use cytosolic substrates such as NAD and NADP to produce messengers that target intracellular Ca2+ stores. The discovery of type III CD38, whose catalytic domain faces the cytosol, has now begun to clarify this topological conundrum. This article reviews the ideas and clues leading to the discovery of the type III CD38; highlights an innovative approach for uncovering its natural existence; and discusses the regulators of its activity, folding, and degradation. We also review the compartmentalization of cADPR and NAADP biogenesis. We further discuss the possible mechanisms that promote type III CD38 expression and appraise a proposal of a Ca2+-signaling mechanism based on substrate limitation and product translocation. The surprising finding of another enzyme that produces cADPR and NAADP, sterile α and TIR motif-containing 1 (SARM1), is described. SARM1 regulates axonal degeneration and has no sequence similarity with CD38 but can catalyze the same set of multireactions and has the same cytosolic orientation as the type III CD38. The intriguing finding that SARM1 is activated by nicotinamide mononucleotide to produce cADPR and NAADP suggests that it may function as a regulated Ca2+-signaling enzyme like CD38.

A cytosolic chaperone complex controls folding and degradation of type III CD38.

Cluster of differentiation 38 (CD38) is the best-studied enzyme catalyzing the synthesis of the Ca2+ messenger cyclic ADP-ribose. It is a single-pass transmembrane protein, but possesses dual orientations. We have documented the natural existence of type III CD38 in cells and shown that it is regulated by a cytosolic activator, calcium- and integrin-binding 1 (CIB1). However, how type III CD38 can be folded correctly in the reductive cytosol has not been addressed. Using the yeast two-hybrid technique with CD38's catalytic domain (sCD38) as bait, here we identified a chaperone, Hsp70-interacting protein (Hip), that specifically interacts with both the type III CD38 and sCD38. Immunoprecipitation coupled with MS identified a chaperone complex associated specifically with sCD38. Pharmacological and siRNA-mediated knockdown of Hsp90 chaperones decreased the expression levels of both sCD38 and type III CD38, suggesting that these chaperones facilitate their folding. Moreover, knockdown of Hsc70 or DNAJA2 increased the levels of both CD38 types, consistent with the roles of these proteins in mediating CD38 degradation. Notably, Hip knockdown decreased type III CD38 substantially, but only marginally affected sCD38, indicating that Hip was selective for the former. More remarkably, DNAJA1 knockdown decreased sCD38 but increased type III CD38 levels. Mechanistically, we show that Hsc70 mediates lysosomal degradation of type III CD38, requiring the lysosomal receptor Lamp2A and the C19-motif in the C terminus of CD38. Our results indicate that folding and degradation of type III CD38 is effectively controlled in cells, providing further strong support of its physiological relevance.

The transferrin receptor CD71 regulates type II CD38, revealing tight topological compartmentalization of intracellular cyclic ADP-ribose production.

The CD38 molecule (CD38) catalyzes biogenesis of the calcium-mobilizing messenger cyclic ADP-ribose (cADPR). CD38 has dual membrane orientations, and type III CD38, with its catalytic domain facing the cytosol, has low abundance but is efficient in cyclizing cytosolic NAD to produce cADPR. The role of cell surface type II CD38 in cellular cADPR production is unknown. Here we modulated type II CD38 expression and assessed the effects of this modulation on cADPR levels. We developed a photoactivatable cross-linking probe based on a CD38 nanobody, and, combining it with MS analysis, we discovered that cell surface CD38 interacts with CD71. CD71 knockdown increased CD38 levels, and CD38 knockout reciprocally increased CD71, and both could be cocapped and coimmunoprecipitated. We constructed a chimera comprising the N-terminal segment of CD71 and a CD38 nanobody to mimic CD71's ligand property. Overexpression of this chimera induced a dramatically large decrease in CD38 via lysosomes. Remarkably, cellular cADPR levels did not decrease correspondingly. Bafilomycin-mediated blockade of lysosomal degradation greatly elevated active type II CD38 by trapping it in the lysosomes but also did not increase cADPR levels. Retention of type II CD38 in the endoplasmic reticulum (ER) by expressing an ER construct that prevented its transport to the cell surface likewise did not change cADPR levels. These results provide first and direct evidence that cADPR biogenesis occurs in the cytosol and is catalyzed mainly by type III CD38 and that type II CD38, compartmentalized in the ER or lysosomes or on the cell surface, contributes only minimally to cADPR biogenesis.

F-box protein-32 down-regulates small-conductance calcium-activated potassium channel 2 in diabetic mouse atria.

Diabetes mellitus (DM) is an independent risk factor for atrial fibrillation, but the underlying ionic mechanism for this association remains unclear. We recently reported that expression of the small-conductance calcium-activated potassium channel 2 (SK2, encoded by KCCN2) in atria from diabetic mice is significantly down-regulated, resulting in reduced SK currents in atrial myocytes from these mice. We also reported that the level of SK2 mRNA expression is not reduced in DM atria but that the ubiquitin-proteasome system (UPS), a major mechanism of intracellular protein degradation, is activated in vascular smooth muscle cells in DM. This suggests a possible role of the UPS in reduced SK currents. To test this possibility, we examined the role of the UPS in atrial SK2 down-regulation in DM. We found that a muscle-specific E3 ligase, F-box protein 32 (FBXO-32, also called atrogin-1), was significantly up-regulated in diabetic mouse atria. Enhanced FBXO-32 expression in atrial cells significantly reduced SK2 protein expression, and siRNA-mediated FBXO-32 knockdown increased SK2 protein expression. Furthermore, co-transfection of SK2 with FBXO-32 complementary DNA in HEK293 cells significantly reduced SK2 expression, whereas co-transfection with atrogin-1ΔF complementary DNA (a nonfunctional FBXO-32 variant in which the F-box domain is deleted) did not have any effects on SK2. These results indicate that FBXO-32 contributes to SK2 down-regulation and that the F-box domain is essential for FBXO-32 function. In conclusion, DM-induced SK2 channel down-regulation appears to be due to an FBXO-32-dependent increase in UPS-mediated SK2 protein degradation.

Cytosolic interaction of type III human CD38 with CIB1 modulates cellular cyclic ADP-ribose levels.

CD38 catalyzes the synthesis of the Ca2+ messenger, cyclic ADP-ribose (cADPR). It is generally considered to be a type II protein with the catalytic domain facing outside. How it can catalyze the synthesis of intracellular cADPR that targets the endoplasmic Ca2+ stores has not been resolved. We have proposed that CD38 can also exist in an opposite type III orientation with its catalytic domain facing the cytosol. Here, we developed a method using specific nanobodies to immunotarget two different epitopes simultaneously on the catalytic domain of the type III CD38 and firmly established that it is naturally occurring in human multiple myeloma cells. Because type III CD38 is topologically amenable to cytosolic regulation, we used yeast-two-hybrid screening to identify cytosolic Ca2+ and integrin-binding protein 1 (CIB1), as its interacting partner. The results from immunoprecipitation, ELISA, and bimolecular fluorescence complementation confirmed that CIB1 binds specifically to the catalytic domain of CD38, in vivo and in vitro. Mutational studies established that the N terminus of CIB1 is the interacting domain. Using shRNA to knock down and Cas9/guide RNA to knock out CIB1, a direct correlation between the cellular cADPR and CIB1 levels was demonstrated. The results indicate that the type III CD38 is functionally active in producing cellular cADPR and that the activity is specifically modulated through interaction with cytosolic CIB1.

CD38 produces nicotinic acid adenosine dinucleotide phosphate in the lysosome.

Nicotinic acid adenosine dinucleotide phosphate (NAADP) is a Ca2+-mobilizing second messenger that regulates a wide range of biological activities. However, the mechanism of its biogenesis remains controversial. CD38 is the only enzyme known to catalyze NAADP synthesis from NADP and nicotinic acid. CD38-mediated catalysis requires an acidic pH, suggesting that NAADP may be produced in acidic endolysosomes, but this hypothesis is untested. In this study, using human cell lines, we specifically directed CD38 to the endolysosomal system and assessed cellular NAADP production. First, we found that nanobodies targeting various epitopes on the C-terminal domain of CD38 could bind to cell surface-localized CD38 and induce its endocytosis. We also found that CD38 internalization occurred via a clathrin-dependent pathway, delivered CD38 to the endolysosome, and elevated intracellular NAADP levels. We also created a CD38 variant for lysosome-specific expression, which not only withstood the degradative environment in the lysosome, but was also much more active than WT CD38 in elevating cellular NAADP levels. Supplementing CD38-expressing cells with nicotinic acid substantially increased cellular NAADP levels. These results demonstrate that endolysosomal CD38 can produce NAADP in human cells. They further suggest that CD38's compartmentalization to the lysosome may allow for its regulation via substrate access, rather than enzyme activation, thereby providing a reliable mechanism for regulating cellular NAADP production.

Rational Design and Identification of Small-Molecule Allosteric Inhibitors of CD38.

CD38 is a multi-functional signaling enzyme that catalyzes the biosynthesis of two calcium-mobilizing second messengers: cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate. It also regulates intracellular nicotinamide adenine dinucleotide (NAD) contents, associated with multiple pathophysiological processes such as aging and cancer. As such, enzymatic inhibitors of CD38 offer great potential in drug development. Here, through virtual screening and enzymatic assays, we discovered compound LX-102, which targets CD38 on the side opposite its enzymatic pocket with a binding affinity of 7.7 µm. It inhibits the NADase activity of CD38 with an IC50 of 14.9 µm. Surface plasmon resonance (SPR) and hydrogen/deuterium exchange and mass spectrometry experiments verified that LX-102 competitively binds to the epitope of the therapeutic SAR 650984 antibody in an allosteric manner. Molecular dynamics simulation was performed to demonstrate the binding dynamics of CD38 with the allosteric ligand. In summary, we established that the cavity to which SAR 650984 binds was an allosteric site and was accessible for the rational design of small chemical modulators of CD38. The lead compound LX-102 that we identified in this study could also be a useful tool for probing CD38 functions and promoting drug discovery.

Electrophysiologic effects and outcomes of sympatholysis in patients with recurrent ventricular arrhythmia and structural heart disease.

INTRODUCTION: Autonomic modulation has been used as a therapy to control recurrent ventricular arrhythmia (VA). This study was to explore stellate ganglion block (SGB) effect on cardiac electrophysiologic properties and evaluate the long-term outcome of cardiac sympathetic denervation (CSD) for patients with recurrent VA and structural heart disease (SHD). MATERIALS AND METHODS: Patients who had recurrent VA due to SHD were enrolled prospectively. Electrophysiologic study and ventricular tachycardia (VT) induction were performed before and after left and right SGB. VA burden and long-term outcomes were assessed for a separate patient group who underwent left or bilateral CSD for drug-refractory VA due to SHD. RESULTS: Electrophysiologic study of nine patients showed that baseline mean (SD) corrected sinus node recovery time (cSNRT) increased from 320.4 (73.3) ms to 402.9 (114.2) ms after left and 482.4 (95.7) ms after bilateral SGB (P = .03). SGB did not significantly change P-R, QRS, and Q-T intervals and ventricular effective refractory period, nor did the inducibility of VA. Nineteen patients underwent left (n = 14) or bilateral (n = 5) CSD. CSD reduced VA burden and appropriate ICD therapies from a median (interquartile range) of 2.5 (0.4-11.6) episodes weekly to 0.1 (0.0-2.4) episodes weekly at 6-month follow-up (P = .002). Three-year freedom from orthotopic heart transplant (OHT) and death was 52.6%. New York Heart Association functional class III/IV and VT rate less than 160 beats per minute were predictors of recurrent VA, OHT, and death. CONCLUSION: SGB increased cSNRT without changing heart rate. CSD was more beneficial for patients with mild-to-moderate heart failure and faster VA.

Stellate ganglion block and cardiac sympathetic denervation in patients with inappropriate sinus tachycardia.

BACKGROUND: Inappropriate sinus tachycardia (IST) remains a clinical challenge because patients often are highly symptomatic and not responsive to medical therapy. OBJECTIVE: To study the safety and efficacy of stellate ganglion (SG) block and cardiac sympathetic denervation (CSD) in patients with IST. METHODS: Twelve consecutive patients who had drug-refractory IST (10 women) were studied. According to a prospectively initiated protocol, five patients underwent an electrophysiologic study before and after SG block (electrophysiology study group). The subsequent seven patients had ambulatory Holter monitoring before and after SG block (ambulatory group). All patients underwent SG block on the right side first, and then on the left side. Selected patients who had heart rate reduction ≥15 beats per minute (bpm) were recommended to consider CSD. RESULTS: The mean (SD) baseline heart rate (HR) was 106 (21) bpm. The HR significantly decreased to 93 (20) bpm (P = .02) at 10 minutes after right SG block and remained significantly slower at 97(19) bpm at 60 minutes. Left SG block reduced HR from 99 (21) to 87(16) bpm (P = .02) at 60 minutes. SG block had no significant effect on blood pressure or HR response to isoproterenol or exercise (all P > .05). Five patients underwent right (n = 4) or bilateral (n = 1) CSD. The clinical outcomes were heterogeneous: one patient had complete and two had partial symptomatic relief, and two did not have improvement. CONCLUSION: SG blockade modestly reduces resting HR but has no significant effect on HR during exercise. Permanent CSD may have a modest role in alleviating symptoms in selected patients with IST.

Atrial Fibrillation: Community Screening Events Improve Awareness in Older Adults.

Educating older adults during atrial fibrillation (AF) screening events to recognize signs and symptoms and seek evaluation may promote detection of AF that occurs between screenings. The authors evaluated learning outcomes of AF awareness education provided during AF screening using a single-arm, pre/posttest design. Participants completed the Knowledge, Attitudes, Beliefs about Atrial Fibrillation Self-Monitoring and Treatment-Seeking (KABAF-SMTS) survey, participated in AF awareness education, and completed a KABAF-SMTS survey 2 weeks after education. Paired t tests revealed that knowledge of AF symptoms increased (p = 0.007). Scores for recognizing the seriousness of AF (p = 0.003), benefits of self-monitoring (p < 0.001), perception of barriers to self-monitoring (p = 0.002), and confidence (p < 0.001) to recognize AF and seek treatment improved. AF awareness education strengthened knowledge, beliefs, and attitudes that may be conducive to recognition and treatment-seeking for AF. [Journal of Gerontological Nursing, 45(9), 31-38.].

Role of the endothelial caveolae microdomain in shear stress-mediated coronary vasorelaxation.

In this study, we determined the role of caveolae and the ionic mechanisms that mediate shear stress-mediated vasodilation (SSD). We found that both TRPV4 and SK channels are targeted to caveolae in freshly isolated bovine coronary endothelial cells (BCECs) and that TRPV4 and KCa2.3 (SK3) channels are co-immunoprecipitated by anti-caveolin-1 antibodies. Acute exposure of BCECs seeded in a capillary tube to 10 dynes/cm2 of shear stress (SS) resulted in activation of TRPV4 and SK currents. However, after incubation with HC067047 (TRPV4 inhibitor), SK currents could no longer be activated by SS, suggesting SK channel activation by SS was mediated through TRPV4. SK currents in BCECs were also activated by isoproterenol or by GSK1016790A (TRPV4 activator). In addition, preincubation of isolated coronary arterioles with apamin (SK inhibitor) resulted in a significant diminution of SSD whereas preincubation with HC067047 produced vasoconstriction by SS. Exposure of BCECs to SS (15 dynes/cm2 16 h) enhanced the production of nitric oxide and prostacyclin (PGI2) and facilitated the translocation of TRPV4 to the caveolae. Inhibition of TRPV4 abolished the SS-mediated intracellular Ca2+ ([Ca2+] i ) increase in BCECs. These results indicate a dynamic interaction in the vascular endothelium among caveolae TRPV4 and SK3 channels. This caveolae-TRPV4-SK3 channel complex underlies the molecular and ionic mechanisms that modulate SSD in the coronary circulation.

Impact of Left Atrial Appendage Closure During Cardiac Surgery on the Occurrence of Early Postoperative Atrial Fibrillation, Stroke, and Mortality: A Propensity Score-Matched Analysis of 10 633 Patients.

BACKGROUND: Prophylactic exclusion of the left atrial appendage (LAA) is often performed during cardiac surgery ostensibly to reduce the risk of stroke. However, the clinical impact of LAA closure in humans remains inconclusive. METHODS: Of 10 633 adults who underwent coronary artery bypass grafting and valve surgery between January 2000 and December 2005, 9792 patients with complete baseline characteristics, surgery procedure, and follow-up data were included in this analysis. A propensity score-matching analysis based on 28 pretreatment covariates was performed and 461 matching pairs were derived and analyzed to estimate the association of LAA closure with early postoperative atrial fibrillation (POAF) (atrial fibrillation ≤30 days of surgery), ischemic stroke, and mortality. RESULTS: In the propensity-matched cohort, the overall incidence of POAF was 53.9%. In this group, the rate of early POAF among the patients who underwent LAA closure was 68.6% versus 31.9% for those who did not undergo the procedure (P<0.001). LAA closure was independently associated with an increased risk of early POAF (adjusted odds ratio, 3.88; 95% confidence interval, 2.89-5.20), but did not significantly influence the risk of stroke (adjusted hazard ratio, 1.07; 95% confidence interval, 0.72-1.58) or mortality (adjusted hazard ratio, 0.92; 95% confidence interval, 0.75-1.13). CONCLUSIONS: After adjustment for treatment allocation bias, LAA closure during routine cardiac surgery was significantly associated with an increased risk of early POAF, but it did not influence the risk of stroke or mortality. It remains uncertain whether prophylactic exclusion of the LAA is warranted for stroke prevention during non-atrial fibrillation-related cardiac surgery.

Anti-Multiple Myeloma Activity of Nanobody-Based Anti-CD38 Chimeric Antigen Receptor T Cells.

Chimeric antigen receptor T cells (CAR-Ts) are a promising strategy for the treatment of many cancers, including multiple myeloma (MM), a hematological malignancy characterized by the high expression of CD38. To broaden the applications of using CD38 as a therapeutic target for the disease, we developed a new nanobody against CD38 and constructed a CD38-CAR that was composed of this nanobody as the targeting domain, and 4-1BB and CD3ζ as the costimulatory and activating domains, in a lentiviral vector. CD3+ T cells from healthy individuals were transduced with the CD38-CAR at an efficiency higher than 60%, as determined by CD38-CAR expression using flow cytometry. The CD38-CAR-Ts proliferated efficiently and produced more inflammatory cytokines, such as IL-2, IFN-γ, and TNF-α, when activated. The CD38-CAR-Ts effectively lysed CD38+ MM cell lines, including LP-1, RPMI 8226, OPM2, and MOLP8, and primary MM cells from multiple myeloma patients. The specificity was demonstrated by the fact that CD38-CAR-Ts showed little cytotoxicity on LP-1 cells with CD38 knocked out or on K562 cells, which do not express CD38. CD38-CAR-Ts appeared to have a very slight cytotoxicity against CD38+ fractions of T cells, B cells, and natural killer cells. In addition, the lysis of CD34+ hematopoietic progenitor cells did not completely inhibit the development of colony-forming units. In vivo, CD38-CAR-Ts inhibited tumor growth in NOD/SCID mice that were subcutaneously inoculated with RPMI 8226 cells. These results demonstrate that the CD38-CAR-Ts constructed with the anti-CD38 nanobody are a promising approach for the treatment of multiple myeloma.

Luteolinidin Protects the Postischemic Heart through CD38 Inhibition with Preservation of NAD(P)(H).

We recently showed that ischemia/reperfusion (I/R) of the heart causes CD38 activation with resultant depletion of the cardiac NADP(H) pool, which is most marked in the endothelium. This NADP(H) depletion was shown to limit the production of nitric oxide by endothelial nitric oxide synthase (eNOS), which requires NADPH for nitric oxide production, resulting in greatly altered endothelial function. Therefore, intervention with CD38 inhibitors could reverse postischemic eNOS-mediated endothelial dysfunction. Here, we evaluated the potency of the CD38 inhibitor luteolinidin, an anthocyanidin, at blocking CD38 activity and preserving endothelial and myocardial function in the postischemic heart. Initially, we characterized luteolinidin as a CD38 inhibitor in vitro to determine its potency and mechanism of inhibition. We then tested luteolinidin in the ex vivo isolated heart model, where we determined luteolinidin uptake with aqueous and liposomal delivery methods. Optimal delivery methods were then further tested to determine the effect of luteolinidin on postischemic NAD(P)(H) and tetrahydrobiopterin levels. Finally, through nitric oxide synthase-dependent coronary flow and left ventricular functional measurements, we evaluated the efficacy of luteolinidin to protect vascular and contractile function, respectively, after I/R. With enhanced postischemic preservation of NADPH and tetrahydrobiopterin, there was a dose-dependent effect of luteolinidin on increasing recovery of endothelium-dependent vasodilatory function, as well as enhancing the recovery of left ventricular contractile function with increased myocardial salvage. Thus, luteolinidin is a potent CD38 inhibitor that protects the heart against I/R injury with preservation of eNOS function and prevention of endothelial dysfunction.

Cryoballoon ablation in Chinese patients with paroxysmal atrial fibrillation: 1-year follow-up.

OBJECTIVES: We assessed the effectiveness and safety of cryoballoon ablation (CBA) in the Chinese population with paroxysmal atrial fibrillation (AF) with a 1-year follow-up and determined the association of early recurrence of atrial tachyarrhythmias (ERAT) with late recurrence (LR). METHODS: A total of 114 patients (age 61 ± 10 years, 78 males) with paroxysmal AF who underwent CBA were consecutively enrolled. After procedures, patients were observed for 3 days with continuous electrocardiogram monitoring in the hospital with routine follow-up visits at 3 months, 6 months, and 1 year. Documented atrial tachyarrhythmia >30 seconds was defined as recurrence. ERAT was defined as any recurrence during the first 3 months, and LR was recurrence between 3 and 12 months. RESULTS: With the first 3 months as blanking period, 76% of patients were free of LR at 12 months. Five patients (4%) experienced complications, including phrenic nerve palsy, stroke, and groin complications. Forty-five percent of patients had ERAT in the first 3 months and 31% of patients had ERAT in the first 3 days. Patients with ERAT had higher LR rate (LRR) than those without ERAT (43% vs 8%, P < 0.001). The LRR of patients with ERAT only in the first 3 days was lower than those with ERAT both in the first 3 days and in 4-90 days (29% vs 64%, P = 0.036). CONCLUSIONS: CBA was an effective and safe treatment option for paroxysmal AF. Patients with ERAT had higher LRR after CBA of AF. The time when ERAT occurred had an impact on LRR.

Down-regulation of the small conductance calcium-activated potassium channels in diabetic mouse atria.

The small conductance Ca(2+)-activated K(+) (SK) channels have recently been found to be expressed in the heart, and genome-wide association studies have shown that they are implicated in atrial fibrillation. Diabetes mellitus is an independent risk factor of atrial fibrillation, but the ionic mechanism underlying this relationship remains unclear. We hypothesized that SK channel function is abnormal in diabetes mellitus, leading to altered cardiac electrophysiology. We found that in streptozotocin-induced diabetic mice, the expression of SK2 and SK3 isoforms was down-regulated by 85 and 92%, respectively, whereas that of SK1 was not changed. SK currents from isolated diabetic mouse atrial myocytes were significantly reduced compared with controls. The resting potentials of isolated atrial preparations were similar between control and diabetic mice, but action potential durations were significantly prolonged in the diabetic atria. Exposure to apamin significantly prolonged action potential durations in control but not in diabetic atria. Production of reactive oxygen species was significantly increased in diabetic atria and in high glucose-cultured HL-1 cells, whereas exposure of HL-1 cells in normal glucose culture to H2O2 reduced the expression of SK2 and SK3. Tyrosine nitration in SK2 and SK3 was significantly increased by high glucose culture, leading to accelerated channel turnover. Treatment with Tiron prevented these changes. Our results suggest that increased oxidative stress in diabetes results in SK channel-associated electrical remodeling in diabetic atria and may promote arrhythmogenesis.