bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1124: CD40L Induces Inflammation in Adipose Tissue - a Potential Link Between the Metabolic Syndrome and Atherosclerosis
Authors: Missiou Anna , Platzer Isabel , Ernst Sandra , Schonbeck Uwe , Libby Peter , Bode Christoph , Zirlik Andreas
Published: July 2021

Background: CD40L figures prominently as marker and mediator of atherosclerosis and its clinical complications. Recent data also demonstrate an association between CD40L and the metabolic syndrome. In the light of these data we hypothesized a functional pro-inflammatory role of CD40L in adipose tissue that provokes systemic pro-inflammatory responses and potentially mediates some of the high cardiovascular risk associated with the metabolic syndrome.

Methods and Results: Adipocytes and preadipocytes isolated from adipose tissue obtained from bariatic surgery did not express CD40L but its receptor CD40 as assessed by PCR and immunohistochemistry. Furthermore, sections of human adipose tissue from obese donors contained more macrophages and T cells colocalizing with CD40L than those from lean controls. Stimulation of both adipocytes and preadipocytes with recombinant CD40L resulted in a concentration- and time-dependent release of the pro-inflammatory cytokine IL-6, the chemokines MCP -1 and IL-8 as well as the inhibitor of fibrinolysis PAI-1 into the supernatant as quantified by ELISA. Membranes isolated from a Murine cell line overexpressing human CD40L reproduced these pro-inflammatory effects compared to respective controls. Interestingly, fenofibrate but neither rosiglitazone, metformin, nor atorvastatin attenuated the CD40L-inducible expression of these pro-atherogenic mediators. Finally, supernatants from adipocytes and preadipocytes stimulated with CD40L activated endothelial cells and macrophages, typical cell types resident in atherosclerotic lesions, as assessed by FACS for tissue factor, Mac-1, and ICAM-1, respectively.

Conclusions: Our data suggest that CD40L induces inflammatory cytokine production in adipose tissue resulting in activation of endothelial cells and macrophages. This new mechanism may contribute to the high cardiovascular risk of patients suffering from the metabolic syndrome.
Abstract 1130: Role Of PI3Kγ For Integrin-dependent Adhesive And Migratory Functions Of Progenitor Cells In Vitro And In Vivo
Authors: Chavakis Emmanouil , Carmona Guillaume , Chavakis Triantafyllos , Zeiher Andreas M , Dimmeler Stefanie
Published: July 2021

Endothelial progenitor cells (EPC) are recruited to ischemic regions to improve neovascularization. β1- and β2-integrins play a crucial role for progenitor cell homing to ischemic tissues. Chemokines and their respective G-protein coupled receptors (GPCRs) are involved in the EPC homing to ischemic tissues. The phosphatidylinositol-3-kinase catalytic subunit gamma (PI3Kγ) is the PI3K isoform, which selectively transduces signals from GPCRs. Here, we investigated the role of PI3Kγ for integrin-dependent homing functions of progenitor cells. As assessed by western blot, EPC express the catalytic subunit PI3Kγ. We then studied the role of PI3Kγ for EPC migration. AS-605240 (100 nM), a selective PI3Kγ-inhibitor (Camps M, Nat. Med., 2005), significantly reduced the SDF1- and the IL-8-induced migration and the SDF1-induced transendothelial migration of human EPC. Adhesion is a further essential step during EPC homing to ischemic tissues. In this regard, the PI3Kγ-inhibitor significantly reduced the SDF1-induced adhesion of EPC on HUVEC monolayers by 69 ± 8 % and on ICAM-1, a β2-integrin ligand. However, the PI3Kγ-inhibitor did not affect the SDF1-induced adhesion of EPC on fibronectin, a β1-integrin ligand, suggesting that PI3Kγ in EPC is involved in the regulation of β2-, but not of β 1-integrin-dependent adhesion. In line with these results, inhibition PI3Kγ blocked the SDF1-induced increase of β2-, but not of β1-integrin-affinity in EPC. Beside EPC, the SDF1-induced migration and adhesion on ICAM-1 of murine bone marrow (BM)-derived Lin
−
progenitor cells from PI3Kγ-deficient mice (PI3Kγ
−
/
−
) were reduced in comparison to wild type (WT) cells. In addition, PI3Kγ-deficiency led to a significant reduction of homing of murine BM-Lin
−
progenitor cells to ischemic muscles after intravenous infusion in the model of hind limb ischemia in comparison to WT cells (48 ± 8 % inhibition). In conclusion, these data demonstrate that PI3Kγ plays an essential role for the integrin-dependent homing of progenitor cells
in vitro
and
in vivo.
The understanding of the molecular mechanisms of progenitor cell homing is essential for the development of new therapeutic strategies in order to improve the efficacy of cell-based therapies in patients with ischemic disorders.
Abstract 1144: Successful Surface-Aminated Nanofiber Expansion of Human Umbilical Cord-Derived CD133+ Cells Leads to Augmentation of Angiogenic Functionality
In Vitro
and
In Vivo
Authors: Das Hiranmoy , Joseph Matthew , Abdulhameed Nasreen , Mao Hai-Quan , Pompili Vincent J
Published: July 2021

Background:
Umbilical cord blood (UCB) and marrow-derived CD133+ cells have been shown to mediate encouraging effects on therapeutic angiogenesis in both animal models and early clinical trials. Low numbers of CD133+ cells derived from a single donor have been a limitation of use of these cells in cardiovascular therapy. We hypothesized that an
ex vivo
aminated nanofiber system combined with cytokine supplementation would provide optimized topographical and biochemical signals to allow the expansion and potential functional augmentation of CD133+ cells without promoting terminal differentiation.

Methods and Results:
Human UCB derived CD133+ progenitor cells were isolated by MACS sorting and
ex vivo
expanded on aminated nanofiber plates with cytokine rich media. Cells harvested 10 days after expansion demonstrated a 225X increase in total number. Flow cytometric analysis demonstrated CD133–24%, CD34–93%, CXCR4–97%, LFA-97% surface expression. The expanded cells can uptake AcLDL efficiently and demonstrate a 2.3X increase in transwell migration to SDF-1 as compared to fresh UCB CD133+ cells.
In vitro
analysis revealed that expanded cells have potential to differentiate into endothelial or smooth muscle phenotype as demonstrated with CD31, vWF, VCAM-1 and F-pholloidin, α-actin, and SM myosin heavy chain immunocytochemistry when re-cultured for 14d in EGM2 or SMBM respectively. RT-QPCR analysis of 1% O
2
exposed (hypoxic) cells demonstrated a 2X increase in VEGF and 3X increase in IL-8 gene expression compared to normoxic control.
In vivo
functionality in a NOD/SCID mouse hind limb ischemic model demonstrated that mice treated with 5 x 10
6
expanded cells (n=7) augmented blood flow ratio (ischemic/control limb) as compared to mice treated with CD133+ cells (n=7) and control (n=7) at 28d. (control 0.32±.02 vs. UCB133+ 0.37±.02 vs. expanded cells 0.50±.04 p<0.01) Capillary density in ischemic hind-limb was increased at 28d (control 62.5±5.4 vs. expanded cell 97.6±2.5 p< 0.01)

Conclusions:
These studies demonstrate successful high level expansion of UCB derived CD133+ cells into functionally potent stem cells which have the capacity to differentiate into vascular cells and promote
in vivo
neovascularization.
Abstract 1180: Protection Against Lipopolysaccharide-Induced Myocardial Dysfunction in Mice by Cardiac-Specific Expression of Soluble Fas
Authors: Niu Jianli , Azfer Asim , Kolattukudy Pappachan E
Published: July 2021

The mechanisms responsible for myocardial dysfunction in the setting of sepsis remain undefined. Fas ligation with its cognate ligand (FasL) induces apoptosis and activates cellular inflammatory responses associated with tissue injury. We determined whether interruption of Fas/FasL interaction by cardiac-specific expression of soluble Fas (sFas), a competitive inhibitor of FasL, would rescue myocardial dysfunction and inflammation in a lipopolysaccharide (LPS)-induced mouse model of sepsis. Wild-type (WT) and sFas transgenic mice were injected intraperitoneally with 10mg/kg LPS or with an equivalent volume of saline. At 18 hours after LPS administration, echocardiographic evaluation revealed a significant decrease in left ventricular fractional shortening in the WT mice, whereas it was preserved in the sFas mice (WT versus sFas, 27.5 ± 4.2% versus 43.3 ± 3.8%,
P
< 0.05). The increase in the transcript levels of proinflammatory cytokines, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 resulting from LPS treatment were attenuated in the myocardium of sFas mice. sFas expression also caused a decrease in myocardial Toll-like receptor 4 (TLR-4) mRNA and protein expression. Myocardial inducible nitric oxide synthase (iNOS) expression and formation of peroxynitrite as well as apoptotic cell death were suppressed in sFas mice compared with WT mice after LPS administration. LPS-induced lung injury and increase in lung water content were also significantly reduced in sFas mice compared with WT mice (4.3 ± 0.15 versus 4.7 ± 0.88,
P
< 0.05). These observations indicate that neutralization of FasL by expression of sFas significantly preserved cardiac function and reduced inflammatory responses in the heart, suggesting that Fas/FasL signaling pathway is important in mediating the deleterious effects of LPS on myocardial function associated with sepsis.
Abstract 225: Role Of PEDF In Regulation Of Skeletal Blood Flow And Neovascularization In Ischemic Limb Via Interaction With VEGF
Authors: Arima Ken , Katsuda Yousuke , Saito Yutaka , Katsuki Yoshio , Sasaki Kenichiro , Yamagishi Shoichi , Ikeda Hisao , Kai Hisashi , Imaizumi Tsutomu
Published: July 2021

[Introduction] We reported that pigment epithelium-derived factor (PEDF) plays an important role in the angiogenesis, vascular permeability and regulation of blood flow in the eye and brain via interaction with VEGF.

[Objective] We examined the role of PEDF in ischemia or VEGF-related angiogenesis or regulation of blood flow in severe limb ischemia.

[Methods] Unilateral hindlimb ischemia was performed in klotho-mice by binding the unilateral femoral artery. After hindlimb ischemia was surgically induced, we injected saline (control group) or VEGF (3 μg/day × 7, VEGF group) to ischemic limb from day 1 to 7. The expression level of PEDF in the ischemic tissue was examined by RT-PCR method. Furthermore, we injected recombinant mouse PEDF (4 μg/day × 7) or anti-PEDF antibody (60 μg/day × 7) in addition to VEGF (VEGF + PEDF group and VEGF + anti-PEDF group, respectively) to ischemic limb from day 1 to 7. We evaluated limb blood flow by laser doppler blood-flow (LDBF) at day 0, 3, 7, and 14, and number of CD34 positive cells in the ischemic tissue by histology as an index of ischemic angiogenesis at day 7 in all groups.

[Results] Expression levels of PEDF in ischemic tissue were significantly elevated in VEGF group. In VEGF group, the ischemic/normal limb blood flow ratio by LDBF analysis and number of CD34 positive cells were significantly increased compared with control group (p < 0.05). The effects of VEGF on blood flow or number of CD34 positive cells were completely abolished by administration of PEDF. Furthermore, anti-PEDF group, accelerated recover of blood flow in ischemic limb, but not number of CD34 positive cells compared in VEGF administration.

[Conclusion] VEGF augmented the expression level of PEDF in the ischemic limb. VEGF-induced ischemic angiogenesis was abolished by exogenous administration of PEDF. Furthermore, anti-PEDF antibody augmented VEGF-induced increase of blood flow. These results suggest that PEDF may play an important role in regulation of skeletal blood flow and neovascularization in ischemic limb via interaction with VEGF.
Abstract 1220: Thioredoxin 1 Negatively Regulates Angiotensin II-Induced Cardiac Hypertrophy through Upregulation of miR-98
Authors: Yang Yanfei , Ago Tetsuro , Sayed Danish , Abdellatif Maha , Sadoshima Junichi
Published: July 2021

Thioredoxin 1 (Trx1) is a 12 kD antioxidant, which protects the heart through multiple mechanisms, including suppression of hypertrophy. We examined the effect of Trx1 on angiotensin II (AII)-induced cardiac hypertrophy (CH). Continuous infusion of a subpressor dose (200 ng/kg/min) of AII by osmotic pump for 2 weeks induced CH (+10% in left ventricular weight/body weight, p<0.05) in mice. AII-induced CH was significantly attenuated in cardiac specific Trx1 transgenic mice (Tg-Trx1) (+1%, ns), whereas it was significantly enhanced in dominant negative Trx1 mice (Tg-DN-Trx1) (+26%, p<0.05). These results suggest that Trx1 is a negative regulator of AII-induced CH. MicroRNAs are naturally existing small noncoding RNA molecules that downregulate posttranscriptional gene expression. In order to examine the involvement of microRNA in the anti-hypertrophic effects of Trx1, total RNA was extracted from Tg-Trx1 hearts, enriched for small RNA, and analyzed by microarrays of mature rodent microRNA. Both miR1 and miR98 are upregulated in the Tg-Trx1 heart. Since miR1 negatively regulates CH, we here examined the role of miR98 in regulation of CH. qRT-PCR showed that miR98 is upregulated in both Tg-Trx1 and Trx1 adenovirus transduced cardiac myocytes (CM) (+15 fold, 3.2 fold). Adenovirus-mediated expression of miR98 in CM reduced cell size both at basal levels (−14%, p<0.05) and in response to AII (−20%, p<0.05), suggesting that miR98 negatively regulates CH. Cyclin D2 (ClnD2) is one of the predicted targets of miR98 and has been shown to play an essential role in mediating AII-induced CH. AII significantly upregulated ClnD2 protein (+34%, p<0.05) at 3 to 6 h in CM. Overexpression of Trx1 downregulated, while expression of DN-Trx1 augmented the AII-induced increase in ClnD2 in vitro (+17.4%, +45.3 %). Similarly ClnD2 was downregulated in Tg-Trx1 and upregulated in Tg-DN-Trx1 mouse hearts (−70.2%, +86.7%). Expression of miR98 in CM decreased expression of ClnD2 by 27% as determined by immunoblotting and reduced nuclear immunostaining of ClnD2, suggesting that both Trx1 and miR98 negatively regulate ClnD2 in CM. These results suggest that Trx1 upregulates miR98, which in turn inhibits CH, possibly through negative regulation of ClnD2 in CM.
Abstract 1254: Novel Insights In Arrhytmogenesis Of Catecholaminergic Ventricular Tachycardia From The First Knock In Model Of Homozygous Calsequestrin Mutation
Authors: Rizzi Nicoletta , Liu Nian , Arcelli Diego , Rossenbacker Tom , Nori Alessandra , Scelsi Mario , Napolitano Carlo , Volpe Pompeo , Priori Silvia G
Published: July 2021

Cardiac Calsequestrin (CASQ2) is a high affinity low capacity calcium binding protein essential in the regulation of intracellular Ca storage and release. Mutations in CASQ2 have been linked to the recessive form of catecholaminergic polymorphic ventricular tachycardia (CPVTr). To elucidate the mechanisms of arrhythmogensis in CPVTr, we generated a knock-in mouse model carrier of the R33Q mutation identified in one of our patients (pts) with a severe CPVT phenotype. Continuous ECG recording (DSI implantable monitors) showed that homozygous (CASQ2
R33Q/R33Q
) mice develop bidirectional and polymorphic VT upon exposure to adrenergic triggers (noise, physical contact). CASQ2
R33Q/R33Q
mice have no sign of structural cardiac abnormality (normal heart/body weight ratio, histology no fibrosis nor myofibrillar disarray). Calcium-binding affinity of CASQ2
R33Q/R33Q
is identical to that of wild type (WT) CASQ2, its localization evaluated with confocal microscopy is superimposable to that of WT. Western blot analysis shows that CASQ2
R33Q/R33Q
content in myocytes is reduced by 40% but Real Time PCR showed that levels of mRNA are not reduced suggesting an abnormal protein turn-over. Evaluation of electrophysiological properties of isolated CASQ2
R33Q/R33Q
myocytes revealed that pacing (1–5 Hz) induces delayed afterdepolarizations (DADs; 18/32 cells 56%) and triggered activity (TA; 9/31 29%) addition of adrenalin (200 nM) enhances DADs and TA (DADs 15/16 cells 93%; TA 8/17 cells 47%) and also elicits (25% of myocytes) early after depolarizations (EADs) and EADs-mediated TA. Our data demonstrate that in analogy with our knock-in model of the dominant form of CPVT due to mutations in the cardiac ryanodine receptor (RyR2) the CASQ2
R33Q/R33Q
mice 1) do not present signs of cardiomyopathy, 2) develop DADs and TA in vitro and bidirectional and polymorphic VT in vivo. At variance with RyR2 mice, CASQ2
R33Q/R33Q
mice 1) present a more malignant phenotype, lower threshold for DADs, TA and VT; 2) EADs concur to arrhythmogenesis 3) overcoming with a knock-in the limitations of previous in vitro and in vivo models overexpressing mutant CASQ2 on the background of the WT protein we demonstrated that a reduction in CASQ2 is implicated in the arrhythmogenesis of recessive CPVT.
Abstract 1274: Syndecan-4 Pathway Interception Reduces Infarct Size After Myocardial Ischemia And Impacts On Remodelling
Authors: Echtermeyer Frank , Hubrich Svenja , Herzog Christine , Hahnenkamp Anke , Stypmann JÃ¶rg M , Theilmeier Gregor
Published: July 2021

Objectives:
Reperfusion injury after myocardial ischemia is governed by leukocyte recruitment. Syndecan-4 (SDC4) has been shown to be a major regulator of cell-cell and cell-matrix interactions. We tested if the SDC4-pathway impacts on myocardial necrosis, tissue loss and remodeling in mouse models of myocardial ischemia/infarction. In addition we analyzed the role of SDC4 in leukocyte recruitment in vitro and in mouse peritonitis.

Methods:
Syndecan4-deficient (SDC4
−/−
) or wildtype (WT) mice were subjected to transient (30 min; MI/R) or permanent (MI) LAD-ligation. In MI/R, after reperfusion for 24h, infarct size was measured by lack of TTC-staining. The extracellular SDC4-domain (ED-SDC4) was systemically overexpressed by hydrodynamic transfection (HDTF). Echocardiography revealed LV-function 4 days after MI(/R). Neutrophil extravasation was tested by lavaging mice 4h after IP-injection of thioglycollate. Adhesion of SDC4
−/−
- or WT-PMN to resting or TNFα-activated, SDC4-siRNA-treated mouse endothelioma cells (EC) was tested in an adhesion assay.

Results:
Infarctions after MI/R were 43±7% larger in SDC4
−/−
compared to WT (p<0.01, n=8). ED-SDC4-HDTF reduced infarct size by 27±4% (p<0.05, n=6). LV-function after MI was significantly worse in WT than SDC-4-KO (EF
pre-post
: −45±7% and −55±8%; p<0.05). In SDC4
−/−
LV-dilation after MI was significantly ameliorated (EDV
post-pre
: 28±7μl vs. 39±8μl, SDC4
−/−
vs WT, p<0.05). Emigration of GR-1
+
-PMN of SDC4
−/−
was increased by 2±0.5-fold in peritoneal lavages (n=6, p<0.05). SDC4 knock down (−80% in RT-PCR) in EC augmented adhesion of PMNs to resting EC by 41±8% and TNFα-activated EC by 20±5%. Adhesion of SDC-4-KO-PMN to resting and TNFα-activated EC was increased by 22±5% and 16±5%. SDC-4-KO-PMN adhesion to activated SDC4 knock down-EC was increased by 40±5% (p<0.05, n=3).

Conclusion:
SDC-4 is a powerful regulator of leukocyte-endothelial interactions and directly links molecular events driving myocardial necrosis and dilatative remodelling. Lack of SDC-4 increases, soluble SDC4 decreases infarct size. SDC4-deficiency reduces dilatative remodelling after MI. Therefore SDC4 beneficially influences early tissue necrosis after ischemia but adversely influences cardiac remodelling.
Abstract 1280: Transcriptional Profiling Of Young And Old Mesenchymal Stem Cells For Their Response To Oxygen Deprivation And Myocardial Reparability
Authors: Jiang Shujia , Haider Husnain K , Idris Niagara Muhammad , Ahmed Rafeeq , Salim Asmat , Ashraf Muhammad
Published: July 2021

Background:
Most clinical studies used autologous bone marrow stem cells (BMSCs) for myocardial repair in elderly patients. Accounting for the limited success in clinical studies, we hypothesized that aging impairs the reparability of BMSCs which limits their therapeutic efficacy.

Methods and Results
: We compared MSCs from young (
Yng
MSCs; 8–12 weeks) and old (
Old
MSCs; 24–26 weeks) rats for their responsiveness to anoxia and therapeutic potential. A significantly higher expression of angiogenic growth factors was observed by
Yng
MSCs under anoxia as compared with
Old
MSCs, cultured either alone or in co-culture (
Co-old
MSCs) with
Yng
MSCs. Likewise,
Yng
MSCs showed significantly higher ability to form tubular structures during Matrigel assay. LDH release assay showed that
Yng
MSCs were more tolerant to apoptotic stimuli (glucose and serum starvation under anoxia) as compared with
Old
MSCs and
Co-old
MSCs, with a possible role for pro-survival proteins
p21
and
p27
which showed increased gene expression (>8-fold and 13-fold respectively) in
Yng
MSCs under anoxia as compared with
Old
MSCs. For
in vivo
studies, acute myocardial infarction model was developed in Fisher-344 rats (n=38). The animals were grouped to receive 70μl DMEM without cells (group-1) or containing 3×10
6 Yng
MSCs (PKH67 labeled; group-2), or
Old
MSCs (PKH26 labeled; group-3) and mixture of
Yng
MSCs+
Old
MSCs (1.5×10
6
cells each, group-4). Histological studies revealed that by day-7,
Yng
MSCs were already showing elongated morphology with orientation similar to the host muscle architecture. Transmission electron microscopy and confocal imaging after immuno-staining for vonWillebrand Factor-VIII, myosin heavy chain, cardiac actinin and connexin-43, showed superior angiomyogenic potential of
Yng
MSCs. Echocardiography showed significantly improved indices of left ventricle contractile function in group-4.

Conclusions:
Aging significantly weakened the responsiveness of
Old
MSCs to anoxia and their differentiation potential. Their reparability of infarcted heart was also impaired with aging which was not improved by the presence of
Yng
MSCs. Hence, transplantation of stem cells from young donors should be considered for heart cell therapy in elderly patients for better prognosis.
Abstract 1298: Adipocytes Produce The Angiogenesis Inhibitor, Soluble Fms-like Tyrosine Kinase (sflt-1)
Authors: Herse Florian , Hering Lydia , Janke Juergen , Gorzcelniak Kerstin , Engeli Stefan , Harsem Nina K , Luft Friedrich C , Staff Anne C , Mueller Dominik N , Dechend Ralf
Published: July 2021

Vascular endothelial growth factor (VEGF) exerts its effects via receptors, including Flt-1. sFLT-1 is a soluble splice variant that binds VEGF locally so that the molecule cannot signal; sFLT-1 is implicated in preeclampsia, a condition associated with obesity. Adipose tissue has a dense vascular network regulated in part by LYVE-1-expressing macrophages and VEGF. The role of sFLT-1 in adipose tissue angiogenesis has not been investigated. We used RT-PCR and found that sFlt-1 was 2.6-fold upregulated in adipose tissue from preeclamptic women, 2.5-fold in decidua, and 6.9-fold in placenta. Circulating sFlt-1 was 2.4-fold elevated (all compared to pregnant nonpreeclamptic controls; p<0.05). We next cultured human preadi-pocytes and adipocytes to examine sFlt-1 and VEGF. sFlt-1 expression increased 9-fold as preadipocytes matured to adipocytes. Adipocytes actively secreted sFlt-1 into the supernatant. Hypoxia (O2 tension reduced to 3%) did not alter sFlt-1 mRNA, while VEGF expression increased 4-fold. Tumor necrosis factor (TNF)- α (100 ng/ml, 48 h) induced VEGF 2-fold, but reduced sFlt-1 production 4-fold. Angiotensin II, insulin, hydrocortisone, pioglitazone, and 17β-estradiol all had no influcence on adipocyte sFlt-1 expression. In fat tissue biopsies from 69 postmenopausal women, sFlt-1 correlated inversely with BMI (r=-0.36, p=0.002), but directly with adiponectin expression (r=0.33, p=0.01). No significant correlation was found to blood pressure, insulin, or microalbuminuria. Our study shows that sFlt-1 is elevated in the circulation, placenta, and is upregulated in adipose tissue from preeclamptic women. sFlt-1 is expressed and secreted by adipocytes. sFlt-1 is elevated during fat cell differentiation and downregulated by TNF-α. In healthy postmeonopausal women, adipose tissue sFlt-1 correlates with low BMI and high adiponectin expression. We speculate that secreted sFlt-1 from adipocytes may contribute to the vascular damage during preeclampsia. Furthermore, sFlt-1 may play a regulatory role in adipose tissue under other circumstances, perhaps as an angiogenesis regulating factor.
Abstract 1306: Late-outgrowth Endothelial Cells Attenuate Intimal Hyperplasia Contributed By Mesenchymal Stem Cells After Vascular Injury
Authors: Wang Chao Hung , Cherng Wen-Jin , Hsieh I-Chang , Yang Ning-I , Yeh Chi-Hsiao
Published: July 2021

Objectives:
Mesenchymal stem cells (MSCs) from the bone marrow (BM) are pluripotent and have the capacity to differentiate into cardiomyocytes, endothelial cells (ECs), and smooth muscle cells (SMCs). Currently, MSCs are one of a number of cell types undergoing extensive investigation for cardiac regeneration therapy. However, it has not yet been determined whether this cell therapy also substantially contributes to vascular remodeling of diseased vessels, such as in intimal hyperplasia.

Methods and Results:
Human MSCs and a variety of progenitor and vascular cells were used for
in vitro
and
in vivo
adhesion experiments. To test the contribution of MSCs to intimal hyperplasia, MSCs from eGFP mice were injected via the tail vein of wild-type littermates after femoral artery wire injury. A model of direct BM transplantation of eGFP MSCs into the tibias of irradiated wild-type littermates was also conducted. Wire-induced vascular injury mobilized MSCs into the circulation. Compared to human aortic SMCs, MSCs exhibited a 2.8-fold increase in the adhesion capacity
in vitro
(
p
< 0.001), and a 6.3-fold increase
in vivo
(
p
< 0.001). In all animal models, immunostaining showed that a significant amount of eGFP MSCs contributed to intimal hyperplasia after vascular injury. Furthermore, MSCs were able to differentiate into cells of endothelial or smooth muscle lineage on the injured vessel wall. Co-culture experiments demonstrated that late-outgrowth ECs guided MSCs to differentiate towards an endothelial lineage through a paracrine effects rather than direct cell-cell interactions.
In vivo
, cell therapy with late-outgrowth ECs significantly attenuated the thickness of the neointima contributed by MSCs (intima/media ratio, from 3.2 ± 0.4 to 0.4 ± 0.1,
p
< 0.001).

Conclusions:
This study raises concerns about the detrimental effects of stem cell therapy on injured vessel remodeling. Tissue regeneration therapy with MSCs or cell populations containing MSCs requires a strategy to attenuate the high potential of MSCs to develop intimal hyperplasia on diseased vessels.
Abstract 1308: Mobilization Of Murine Endothelial Progenitor Cells Requires Medullary Fibroblast Growth Factor-2
Authors: Werner Nikos , 1/4aine Vincent , Preiss Tilman , Arnal Jean Francois , Nickenig Georg
Published: July 2021

Background
Fibroblast growth factor-2 (FGF2) regulates proliferation of bone marrow (BM) stromal cells and has pro-angiogenic effects regulating endothelial cell growth, migration, and reendothelialization. We have previously demonstrated that estrogen-mediated endothelial progenitor cell (EPC) mobilization from BM into peripheral blood is inhibited in FGF2 knock-out (−/−) mice indicating a central role of medullary FGF2 in progenitor cell mobilization. Here we determine the underlying molecular mechanisms of FGF2-dependent statin-mediated progenitor cell mobilization.

Methods and Results
To determine the molecular mechanisms of FGF2-mediated progenitor cell mobilization, FGF2−/− and wild type (wt) mice were treated with the HMG-CoA reductase-inhibitor rosuvastatin (10mg/kg body weight s.c. per day) and placebo. In contrast to wt mice, the number of Sca1/flk-1 positive EPC in peripheral blood of rosuvastatin treated FGF2−/− mice did not increase. This was due to an accumulation of Sca1/flk-1 positive cells within the BM. Inhibition of EPC mobilization in FGF2−/− mice was associated with a delay in reendothelialization in a mouse model of focal endothelial cell denudation and a severe reduction in neoangiogenesis despite statin treatment. BM transplantation experiments demonstrated that reconstitution of FGF2−/− mice with wt stem cells completely restored statin-mediated EPC mobilization while wt mice reconstituted with FGF2−/− stem cells developed a mobilization defect associated with an impaired endothelial-dependent vasore-laxation and enhanced neointima formation. In wt mice, statin treatment was associated with an increased number of activated osteoclasts compared to placebo treated mice which are known to play a pivotal role in stem cell mobilization.

Conclusion
Medullary FGF2 is essential in the statin-mediated mobilization of EPC from BM into peripheral blood while it seems not to influence EPC proliferation. The mobilization defect in FGF2−/− mice is associated with a significant impairment of vascular endothelial function and repair which underlines the importance of EPC in endothelial cell regeneration.
Abstract 1364: Not All Refractory Period Abbreviation Promotes Atrial Fibrillation Equally: A Comparison Between Vagal and Atrial Tachycardia Remodeled Substrates
Authors: Katsouras Grigorios , Sakabe Masao , Lemola Kristina , Comtois Philippe , Ting Michael , Maguy Ange , Burstein Brett , Talajic Mario , Guerra Peter , Nattel Stanley
Published: July 2021

Background:
Vagal (VG) and atrial tachycardia remodeled (ATR) AF substrates share many features: reduced effective refractory period (ERP), increased ERP heterogeneity and some common molecular mechanisms (I
KACh
enhancement by acetylcholine release in VG, constitutive I
KACh
enhancement in ATR). This study compared VG and ATR substrates at comparable ERP abbreviation.

Methods:
In each of 5 VG dogs, bilateral cervical VG stimulation parameters were adjusted (mean±SD: 3.6±1.7 V and 12.2±1.5 Hz; 0.2 ms) to produce the same mean ERP (at 4 RA and 4 LA sites) as a sex and weight matched ATR dog (RA paced 400 bpm × 7 days). Mean duration of burst pacing induced AF (DAF) and local dominant frequencies (DFs, analyzed by FFT at 240 bipolar electrodes, Fig A

) were determined.

Results:
Mean ERP was 79±13 ms in VG and 78±13 ms in ATR dogs. DAF was greater in VG than ATR dogs (1056±323 vs 289±510 s *P<0.01; both significantly > control, 43±61 s). Despite matched ERPs, there were significant differences in DF distribution (Fig B

): DF was faster (mean DF: 11.8±1.1 Hz VG vs 9.7±1.3 Hz ATR*) and DF variability greater (indicated by SD: 1.8±0.6 Hz VG vs 0.8±0.5 Hz ATR*) in VG dogs. AF drivers reflected by maximum DF zones were adjacent to autonomic ganglia (over RA in 4/5) for VG dogs; in ATR dogs driver zones were less clear and showed variable location.

Conclusions:
For a comparable atrial ERP, VG AF is faster and more persistent than AF with an ATR substrate. These results are consist with modeling work suggesting that VG-induced hyperpolarization is an important contributor to AF-maintaining rotor stabilization and acceleration, and indicate important differences between these superficially similar AF substrates.
Abstract 1372: Characterization Of The Rejuvenation Of Cardiac Sympathetic Nerves In Cardiac Hypertrophy
Authors: Kimura Kensuke , Ieda Masaki , Kanazawa Hideaki , Arai Takahide , Kawakami Takashi , Onizuka Takeshi , Kageyama Toshimi , Ohno Yohei , Yoon Sung Han , Makino Shinji , Sano Motoaki , Ogawa Satoshi , Fukuda Keiichi
Published: July 2021

Background
: Cardiac hypertrophy induces the fetal isoform of genes (rejuvenation), including contractile proteins, ion channels, and natriuretic peptides. Cardiac sympathetic nerve function is known to be altered in cardiac hypertrophy and congestive heart failure. We recently reported that alteration of cardiac sympathetic nerves (CSN) was caused by their rejuvenation (Circ Res, 2007). The present study was designed to examine the precise characterization of the rejuvenation of CSN in cardiac hypertrophy.

Methods and Results
:

RV hypertrophy was produced by consistent hypoxia (10% O
2
) in C57/BL6 mice. RV pressure increased to 47 mmHg, and RV/(body weight) ratio increased by 1.6 fold.

Nerve growth factor protein was augmented in hypertrophic RV, but was unchanged in LV.

Double-transgenic mice, which specifically express eGFP (enhanced green fluorescent protein) in the sympathetic neurons, was generated by crossing dopamine β-hydroxylase (DBH)-Cre mice with Floxed-eGFP mice.

The eGFP-positive CSN were markedly increased in hypertrophic RV, but not in LV. Nerve density, quantitated by immunostained area with eGFP and GAP43 (growth-associated corn marker), increased by 8.1 and 9.3 fold, respectively, in RV, but not in LV. (4) Catecholamine content was attenuated in RV. (5) Western blot revealed that tyrosine hydroxylase was markedly down-regulated in RV. (6) Immunostaining clearly demonstrated that the immature neuron markers, PSA-NCAM (highly polysialylated neural cell adhesion molecule) and Ulip-1 (Unc-33-like phosphoprotein 1), were expressed in CSN in hypertrophic RV and stellate ganglia. Basic helix-loop-helix transcription factor, Mash-1 (mammalian achaete-scute complex homolog 1) was strongly expressed in the stellate ganglia. (7) Immature neuron marker-immunopositive cells in stellate ganglia had a markedly decreased TH expression.

Conclusion
: The rejuvenated CSN showed various immature and fetal neuron marker genes at not only the peripheral axons but also the cellular bodies at the stellate ganglia. Rejuvenation of CSN might be critically involved in the alteration of sympathetic neuronal function in cardiac hypertrophy, including depressed norepinephrine synthesis and hyperinnervation.
Abstract 1398: Effects Of Cardioprotective Drugs On Mitochondrial Functions During Myocardial Ischemia/reperfusion Revealed By Real-time Two-photon Imaging Of Perfused Rat Hearts.
Authors: Akao Masaharu , Matsumoto-Ida Madoka , Kato Masashi , Iguchi Moritake , Takeda Toshihiro , Kita Toru
Published: July 2021

Background:

We recently established a real-time imaging system to monitor the mitochondrial functions in perfused hearts subjected to ischemia/reperfusion, using two-photon laser scanning microscopy. The opening of the mitochondrial permeability transition pore (MPTP), which leads to loss of mitochondrial membrane potential (ΔΨ
m
) and mitochondrial dysfunction, is a critical step in cardiomyocyte death during ischemia/reperfusion. We tested the effects of various drugs on Δ Ψ
m
loss in perfused rat hearts using this novel system.

Methods and Results:

Langendorff-perfused rat hearts were loaded with a fluorescent indicator of ΔΨ
m
, tetramethylrhodamine ethyl-ester, and was pretreated with the drugs for 30 minutes. The spatio-temporal changes of ΔΨ
m
in response to ischemia/reperfusion were monitored at a subcellular level, under excitation with a two-photon laser. Ischemia/reperfusion elicits stereotyped responses in cardiomyocytes; cells maintained a constant ΔΨ
m
for the cell-to-cell specific period of latency, followed by a rapid and irreversible ΔΨ
m
depolarization and subsequent cytolysis. The mitochondrial ATP-sensitive potassium channel opener diazoxide (100 μmol/L) and the mitochondrial peripheral benzodiazepine receptor ligand 4′-chlorodiazepam (50 μmol/L) markedly suppressed the latency and thus decreased the likelihood that cells would undergo ΔΨ
m
depolarization; they not only decreased the number of cells undergoing ΔΨ
m
loss but also delayed the onset of ΔΨ
m
loss, whereas it did not change the duration of depolarization in unprotected cells. While this mimicked the effects of ischemic preconditioning to suppress latency, it did not reflect that of cyclosporin A, a MPTP blocker which rather slowed the process of depolarization. In addition, neither a free radical scavenger 2-mercaptopropionyl glycine nor an iron chelator desferroxamine, which reduces the generation of hydroxyl radical through inhibiting the iron-mediated Fenton reaction, was protective. Simply scavenging oxygen radicals or reducing radical generation may not be a feasible strategy to achieve cardioprotection.

Conclusions:

This novel two-photon imaging provides deeper insight into anti-ischemia/reperfusion therapy targeting mitochondria.
Abstract 1422: Candesartan-induced Upregulation Of Transcriptional Regulator Yin Yang 1 Blocks Leukocyte-endothelial Interaction Under Physiological Flow.
Authors: Ishii Hideto , Mori Daisuke , Sato Mayuko , Yoshida Masayuki
Published: July 2021

[Objective]
Transcriptional control of inflammation-relevant genes plays an important role in atherosclerosis. We have reported a dominant role of Yin Yang 1 (YY1), a transcriptional regulator, in proliferation of vascular smooth muscle cells (Santiago FS, Ishii H, et al. Circ Res in press). However, its effect in vascular endothelium remains unclear. Recent observation points a novel “genomic” effect of Angiotensin II to influence inflammation. In this study, we examined a potential contribution of YY1 in angiotensin type II receptor blocker (ARB)-mediated modulation of leukocyte-endothelial adhesion under flow condition.

[Methods and Results]
Human umbilical vein endothelial cells (HUVEC) were co-incubated in the presence or absence of candesartan, an ARB (1 uM) for 4 hours with TNFα (5 ng/ml). Treatment with candesartan dramatically inhibited THP-1 cell adhesion to TNFα-activated HUVEC under static (p <0.01) and flow conditions (p <0.05). We documented that upregulation of YY1 mRNA after candesartan treatment. To examine a role for YY1 in leukocyte-endothelial interaction, YY1 was overexpressed via recombinant adenovirus. The expression levels of VCAM-1 was significantly decreased in HUVEC treated with adenovirus YY1. In addition, adenovirus YY1 blocked THP-1 adhesion to TNFα-activated HUVEC under flow conditions (TNFα, 8.40±0.64/HPF; TNFα+candesartan, 1.93±0.54/HPF; p <0.0001). The expression level of phosphorylated-JNK, as determined by western blotting analysis, was reduced following YY1 transfection. In contrast, YY1 inhibition by YY1 siRNA did not influence leukocyte-endothelial adhesion and the expression of adhesion molecules.

[Conclusion]
We demonstrated that candesartan treatment upregulated YY1 in HUVEC and inhibited leukocyte-endothelial adhesion by inhibition of VCAM-1 expression. Our findings suggest a novel anti-inflammatory effect of ARB via YY1-dependent mechanisms.
Abstract 1449: A Soluble Form Of Ldl Receptor Gene Family Member, Lr11, Is Essential For Angiotensin Ii -induced Smc Migration
Authors: jiang meizi , bujo hideaki , ohwaki kenji , takahashi mao , shirai kohji , Schneider Wolfgang J , saito yasushi
Published: July 2021

Migration of vascular smooth muscle cells (SMCs) from the media to the intima plays a key role in the development of atherosclerosis. Considerable amounts of the soluble form of LR11 (sLR11) are produced by intimal SMCs and enhance migration of SMCs in vitro via upregulation of urokinase-type plasminogen activator receptor (uPAR) expression. Circulating levels of sLR11 are increased in subjects with coronary artery diseases. Here, we show that targeted disruption of the LR11 gene greatly reduces intimal thickening of arteries through attenuation of angiotensin II (Ang II)-induced migration of SMCs. The intimal thickness of femoral arteries after cuff placement was decreased in
Lr11
−/−
mice. In cultured
Lr11
−/−
SMCs, AngII-stimulated migration and particularly, attachment, were almost completely abolished. In SMCs, sLR11 caused membrane ruffle formation via activation of focal adhesion kinase (FAK)/extracellular signal-regulated protein kinase (ERK)/Rac1, accompanied by formation of a complex between uPAR and integrin αvβ3. AngII accelerated membrane ruffling through an increase in sLR11-mediated formation of this complex. Overproduction of sLR11 resulted in the reduced sensitivity of AngII-induced activation pathways to inhibition by an AngII type 1 receptor blocker in mice. These results show that AngII-induced SMC migration is mediated by sLR11- induced signal activation for membrane ruffle formation. Our discovery of involvement of LR11 in AngII-induced SMC migration possibly leads to an establishment of novel approach for regulating plaque formation.
Abstract 1516: Does A Normal Coronary Angiography Really Mean A Good Prognosis In Patients With Acute Coronary Syndromes?
Authors: Dudek Dariusz , Depukat Rafal , Chyrchel Bernadeta , Siudak Zbigniew , Dziewierz Artur , Rzeszutko Lukasz , Legutko Jacek , Dubiel Jacek S
Published: July 2021

Introduction: Coronary angiography remains gold standard for diagnosis of coronary artery disease (CAD) and acute coronary syndrome (ACS). Among patients (pts) referred to cath-lab with diagnosis of ACS exists a subgrup presenting symptoms of myocardial ischaemia and no criticial coronary lesions in angiography. The long-term outcome is not well established and managing treatment for such patients is still challenging.

Aim: Evaluation of clinical outcome and received pharmacotherapy in pts with ACS and no significant coronary lesions.

Methods: We collected data of consecutive pts admitted to cath-lab between July 2004 and June 2006 with diagnosis of ACS and coronary angiography considered as normal or near-normal (lesions under 50% of stenosis assessed visually). We analyzed demographic data, angiographic, electrocardiographic and laboratory tests results. During long-term follow-up received treatment, major adverse cardiac events were assessed.

Results: One hundred eight pts (56 females; mean age 58.5 +/−13.5 years) admitted to cath-lab with diagnosis of acute myocardial infarction (38%) or unstable angina (62%) had normal or near normal coronary angiography. Mean LVEF was 52.6 +/− 12.9%, prevalence of CAD risk factors was: hypertension 72.2%, hypercholesterolemia 51.8%, diabetes mellitus 13.0%. The treatment administered during the hospitalization and follow-up is shown in table

. During the mean follow-up of 16.5 months (range 6–30 months) all-cause mortality rate was 9.3%, cardiovascular mortality rate was 7.4%, repeat hospitalization for cardiovascular reasons 20.4%.

Conclusions: Long-term outcome of patients with ACS and nonobstructive coronary angiography is not as benign as it is commonly thought. These patients have high rate of major adverse cardiac events and rehospitalization for cardiovascular reasons. They also are not receiving therapy directed at aggressive antiatherosclerotic therapy with statins, ACE-I and b-blockers.

Treatment administered during the hospitalization, at discharge and during the follow-up
Abstract 1519: Atrial Fibrillation Is Independently Associated With A High Risk of Death in Patients With Acute Coronary Syndromes
Authors: Lopes Renato D , Mahaffey Kenneth , Harrington Robert A , Horton John , Armstrong Paul , Newby Kristin , Ohman Magnus , Van de Werf Frans , Al-Khatib Sana M , Wallentin Lars , Granger Christopher B
Published: July 2021

Purpose:
To assess whether atrial fibrillation (AF) is associated with 1-year mortality among patients with acute coronary syndromes (ACS).

Methods:
We analyzed the 1-year mortality of 110,588 ACS patients using a pooled database of clinical trials. Cox proportional hazards modeling was used to adjust for baseline characteristics including AF that may be associated with the risk of death within 7 days, and death from day 7 to 1 year. We examined the effect of AF in patients with ST-segment elevation versus those with non ST-segment elevation.

Results:
The prevalence of in-hospital AF was 8.0%. Baseline characteristics according to AF or not were: median age (70 vs 62 years), systolic blood pressure (130 vs 130mmHg), Killip class III and IV (4.2% vs 1.6%), history of hypertension (41% vs 48%), diabetes mellitus (19% vs 16%), prior congestive heart failure (7% vs 3.5%), and prior myocardial infarction (23% vs 20%). The figure

shows the significant relationship of AF with mortality in the overall population. AF was significantly associated with an increased risk of in-hospital mortality in the non ST elevation ACS patients (adjusted odds ratio (OR) of 1.9 95% CI 1.5 to 2.5),

but not

in the ST elevation ACS patients (adjusted OR 1.0 95% CI 0.9 to 1.1). However, AF was significantly associated with an increased risk of death from day 7 to 1 year for both ST and non ST elevation ACS patients (adjusted OR 3.8 95% CI 3.5 to 4.1, and 2.2 95% CI 1.7 to 2.7, respectively).

Conclusion:
Among patients with ACS, AF is independently associated with a higher risk of 1-year mortality for both ST and non ST elevation ACS patients. Understanding the reasons for this increased risk may provide opportunity for improving care.
Abstract 1520: Are Contemporary Patients Hospitalized with Acute Coronary Syndrome Being Discharged on Intensive Lipid Lowering Therapy?: An Analysis Of 22, 807 Hospitalizations In GWTG-CAD
Authors: Deedwania Prakash C , Fonarow Gregg C , Cannon Christopher P , Dai David
Published: July 2021

Background: Based on the documented benefits of intensive lipid lowering therapy (LLT) with statins in patients with acute coronary syndrome (ACS) guidelines recommend intensive LLT in ACS patients. However, little information is available regarding application of these guidelines in hospitalized ACS patients.

Methods: The Get With The Guidelines database was analyzed for all ACS hospitalizations from 07/2005 to 04/2007 at 329 hospitals across the USA of which 277 sites reported the dose of LLT. Intensive LLT was defined as that expected to provide a >50% reduction in LDL: atorvastatin 40/80 mg, rosuvastatin 20/40 mg, simvastatin 80 mg, and any statin combined with ezetimibe. All other LLTs were considered less intensive therapy.

Results: Of 60,453 ACS hospitalizations, 84.4% eligible were discharged on LLT. LLT dosing was available in 22,807 (37.7%). Of these patients only 8400 (36.8%) of ACS patients were treated at time of discharge with intensive LLT, whereas 63.2% were discharged on less intensive LLT. Comparison of demographic characteristic and clinical features revealed that older age and female gender were associated with use of less intensive LLT whereas patients undergoing PCI with a stent and history of smoking were more likely to receive intensive LLT. Admission LDL level was modestly predictive of more intensive LLT.

Conclusions: In this large cohort of contemporary ACS patients, close to two-thirds of eligible patients were not discharged on intensive LLT. Age and gender appear to be significant contributors to less intensive LLT. These findings emphasize the ongoing need for implementation of current guidelines for intensive LLT in ACS patients.