bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 508: Peripheral Artery Disease is Associated With Frailty in Patients With Chorionic Hemodialysis
Authors: Takeuchi Hidemi , Okuyama Michihiro , Uchida Haruhito A , Otaka Nozomu , Kakio Yuki , Okuyama Yuka , Umebayashi Ryoko , Sugiyama Hitoshi , Wada Jun
Published: July 2021

Objective:
The clinical condition of frailty is a common problem in the elderly population. Chronic hemodialysis (HD) patients often have peripheral artery disease (PAD) as a vascular complication. However, the relationship between PAD and frailty in Japanese HD patients remains unknown. The aim of this study was to identify the relationships among PAD and risk factors in Japanese chronic HD patients.

Method:
This study was a multi-center, cross-sectional and observational investigation which was conducted at 6 institutions, including 5 general hospitals and 1 private clinic. Subjects were all chronic HD patients who regularly visited the institutions. To evaluate frailty, we used the modified Fried’s frailty phenotype adjusted for Japanese as the self-reported questionnaire, and measured each physical domain. Furthermore, we calculated ankle-brachial index (ABI) to define PAD. PAD was defined as ABI < 0.9 in our study.

Result:
Of the 542 patients in all institutions, 362 were enrolled in this study. Sixty-two patients (17.1%) were categorized as PAD group and 300 patients (82.9%) as non-PAD group. In the PAD group, the prevalence of frailty was significantly higher than in the non-PAD group (34% vs 18%). Non-shunt side grip strength was significantly stronger in the non-PAD group (23.6 kg vs 17.0 kg,
P
<0.0001). Thigh circumferences (the mean of both sides) were also significantly larger in the non-PAD group (41.7 cm vs 39.7 cm,
P
=0.0054). Univariate regression analyses showed that frailty, age, number of oral medicine, and history of myocardial infarction (MI) had significant correlations with PAD. Multivariate logistic regression analysis demonstrated that the factors independently associated with PAD were as follows: frailty (OR = 2.061, 95% C.I. 1.091-3.894) and MI (OR = 3.742, 95% C.I. 2.051-6.831).

Conclusion:
PAD is associated with frailty in HD patients.
Abstract 51: A Critical Role for Outside-In Signaling of Integrin AlphalIIIb/Beta3 in Shear-Dependent Platelet Microvesicle Formation and Phosphatidylerine Exposure
Authors: Pang Aiming , Cui Yujie , Delaney Michael K , Stojanovic-Terpo Aleksandra , Du Xiaoping
Published: July 2021

Platelets promote coagulation mainly by exposing membrane phosphatidylserine (PS) and releasing PS-expressing microvesicles (MV). We have recently shown that PS exposure and MV release induced by platelet agonists requires shear stress. To identify the receptor responsible for the shear-dependent signaling leading to PS exposure and MV release, we compared platelets from β
3
-/-
mice and wild-type mice in MV release and PS exposure under defined shear stress introduced using a cone-plate rheometer. MV release and PS exposure were determined using flow cytometry. Shear-dependent PS exposure and MV release were significantly suppressed in β
3
-/- platelets. Similarly, Wild type platelets treated with integrin antagonists also showed defective PS exposure and MV release. These data indicate an important role for the ligand binding function of integrin αιιb/β3 in shear-dependent MV release and PS exposure. To determine whether the role of integrin αιιb/β3 is due to its outside-in signaling, β
3
-/- platelets were transplanted with wild type β
3
or a mutant β
3
with the critical Gα13 binding site of the β
3
cytoplasmic domain (EEE) changed to alanines (AAA), which was previously shown to selectively abolish outside-in signaling of αIIb/β
3
. Transplantation of wild type β
3
rescued the defective MVs release and PS exposure of β
3
-/- platelets. In contrast, AAA mutant failed to rescue these defects. Consistently, wild type platelets treated with the selective inhibitor of Gα13-integrin interaction, inhibited integrin outside-in signaling and also PS exposure and MV release under shear stress. Furthermore, we also showed that the inhibition of Src, which is important in outside-in signaling downtream of Gα13, also abolished shear-dependent MV release and PS exposure. These data suggest that integrin outside-in signaling mediated by the Gα13-β
3
interaction and Src-dependent signaling pathway plays an important role in transmitting shear-induced mechanical signals leading to MV release and PS exposure in activated platelets.
Abstract 511: High Signal Intensity on Diffusion Weighted Magnetic Resonance Imaging Reflects Acuity of Deep Vein Thrombus
Authors: Gi Toshihiro , Yamashita Atsushi , Kuroiwa Yasuyoshi , Konishi Yuko , Asanuma Taketoshi , Imamura Takuroh , Asada Yujiro
Published: July 2021

Objective:
Acuity of deep vein thrombus/ thrombosis (DVT) may affect effectiveness of anti-thrombotic therapy. However, the acuity of DVT is not reliably detected by current noninvasive imaging techniques. This study investigated whether diffusion weighted magnetic resonance (MR) imaging can detect DVT and define the acuity of thrombus in patients with DVT and a rabbit model of venous thrombus.

Methods:
Diffusion weighted MR imaging was performed with a 1.5-T MR system in 8 patients with DVT. Venous thrombus was induced in rabbit jugular vein by endothelial denudation and 10 minutes blood stasis with a balloon catheter. The thrombus was imaged with a 3.0-T MR system at 4 hours and at 1, 2 and 3 weeks, and the jugular veins were histologically assessed.

Results:
All patients were detected DVT with diffusion weighted MR imaging, and the DVT showed high or mixed high and iso signal intensity on the diffusion sequence. The rabbit venous thrombi were rich in erythrocyte and fibrin at 4 hours, and showed focal organizing reaction at 1 and 2 weeks, and was replaced by fibrous tissue at 3 weeks. The rabbit thrombi showed high signal intensity on diffusion weighted MR imaging at 4 hours, mixed high and iso signal intensity at 1 and 2 weeks, or mixed iso and low signal intensity at 3 weeks. The signal intensity was positively correlated with erythrocyte and fibrin contents, and negatively correlated with macrophage and collagen contents.

Conclusions:
Diffusion weighted MR imaging can detect DVT and high signal intensity on the sequence may reflect acuity of DVT.
Abstract 514: Sex-Related Disparities in Outcomes After Myocardial Infarction Among Patients With Atrial Fibrillation: Evidence From a Nationwide Study
Authors: Akinjero Akintunde M , Adegbala Oluwole , Akinyemiju Tomi
Published: July 2021

Background:
The overall mortality rate after acute myocardial infarction (AMI) is falling in the United States. However, outcomes remain unacceptably worse in females compared to males. It is not known how coexisting atrial fibrillation (AF) modify outcomes among the sexes. We sought to examine the association of sex with clinical characteristics and outcomes after AMI among patients with AF.

Methods:
We accessed the Healthcare Cost and Utilization Project (HCUP) Nationwide Inpatient Sample (NIS), to extract all hospitalizations between 2007 and 2011 for patients above 18yrs with principal diagnosis of AMI and coexisting diagnosis of AF using ICD 9-CM codes. The NIS represents the largest all-payer hospitalization database in the United States, sampling approximately 8 million hospitalizations per year. We also extracted outcomes data (length of stay (LOS), stroke and in-hospital mortality) after AMI among Patients with AF. We then compared sex differences. Univariate and Multivariate analysis were conducted to determine the presence of statistically significant difference in outcomes between men and women.

Results:
A total of 184,584 AF patients with AMI were sampled, consisting of 46.82% (86,420) women and 53.13% (98,164) men. Compared with men, women with AF and AMI had a greater multivariate-adjusted risk for increased stroke rate (aOR=1.51, 95% CI=1.45-1.59), and higher in-hospital mortality (aOR=1.12, 95% CI=1.09-1.15). However, female gender was not significantly associated with longer LOS (aOR=-0.22, 95% CI= -0.29-(-0.14).

Conclusion:
In this large nationwide study of a population-based cohort, women experienced worse outcomes after AMI among patients with AF. They had higher in-hospital mortality and increased stroke rates. Our findings highlight the need for targeted interventions to improve these disparities in outcomes.
Abstract 518: Coronary Thrombus Leukocytes of Acute Coronary Syndrome Patients Trans-Differentiate Into Endothelial Cell-Like Angiogenic Cells
Authors: Fu Hangfei , Vadalia Nish , Xue Eric R , Johnson Candice , Yu Jun , Yang Xiaofeng , Wang Hong
Published: July 2021

Current angiogenic therapies for cancers and cardiovascular diseases have not yet achieved expected benefits, which reflects the need for improved understanding of angiogenesis. In this study, we focused on solving the problem of whether tissues have different angiogenic potentials (APs) in physiological conditions; and how angiogenesis is regulated in various disease conditions. In healthy and diseased human and mouse tissues, we profiled the expression of 163 angiogenic genes, including transcription regulators (TRs), growth factors and receptors (GF/Rs), cytokines and chemokines (C/Cs), and proteases and inhibitors (P/Is). TRs were categorized as inflammatory, homeostatic, and endothelial cell-specific TRs, and C/Cs were categorized as pro-angiogenic, anti-angiogenic, and bi-functional C/Cs. We made the following findings:
1)
human heart, muscle, eye, pancreas, and lymph node are among the tissues with the highest APs;
2)
tissues with high APs have more active angiogenic pathways and angiogenic C/C responses; 3
)
inflammatory TRs dominate regulation of all angiogenic C/Cs; homeostatic TRs regulate all to a lower extent, while endothelial cell-specific TRs mainly regulate pro-angiogenic and bi-functional C/Cs;
4)
tissue AP is positively correlated with the expression of oxygen sensors PHD2 and HIF1B, VEGF pathway gene VEGFB, and stem cell gene SOX2;
5)
cancers of the digestive system tend to have increased angiogenesis dominated by endothelial cell-specific pro-angiogenic pathways, while lung cancer and prostate cancer have significantly decreased angiogenesis;
6)
endothelial cell-specific pro-angiogenic pathways are significantly increased in thrombus-derived leukocytes in patients with acute coronary artery disease. Our results demonstrate that thrombus-derived leukocytes are partially “hijacked” to become endothelial cell-like angiogenic cells that directly promote angiogenesis after myocardial infarction; and that certain solid tumors may be more sensitive to anti-angiogenic therapies than others.
Abstract 519: Inhibition of von Willebrand Factor Activity Does Not Delay Cutaneous Wound Healing
Authors: Gorman Kristen , Sommerville Laura , Pitoc George , Monroe Dougald M , Hoffman Maureane
Published: July 2021

Introduction:
Physiologically, von Willebrand Factor (VWF) recruits platelets to sites of injury by binding to exposed collagen and platelet GPIbα, particularly at sites of high shear. VWF is critical to hemostasis, since complete deficiency leads to bleeding as severe as hemophilia. Conversely, VWF plays a key role in arterial thrombosis, as in myocardial infarction and stroke. Despite being an attractive therapeutic target, no agents directed against VWF are currently available. Our group is testing an RNA aptamer that blocks VWF binding to GPIbα as an antithrombotic. We have previously shown that leukocyte influx and cutaneous healing are impaired in a mouse model of hemophilia. We have also found that wound healing is delayed in mice given target specific oral anticoagulants. The purpose of the current study was to determine whether inhibiting VWF would similarly impair wound healing.

Methods:
We used C57BL/6J mice administered vehicle or VWF aptamer. At the dose selected, aptamer prolonged the saphenous vein bleeding time for the expected 10 day healing period. Healing was assessed in a skin punch biopsy model. Wound size was measured daily, and following sacrifice the wound area harvested for histology. Inflammation (in the absence of tissue injury) was induced by topical application of cantharidin for 24 hours, followed by tissue harvesting.

Results:
The in vivo efficacy of the aptamer was verified, since bleeding remained prolonged to a level slightly less severe than hemophilia at the end of the experiment (10 days after wounding). There was no difference in time to healing between control and aptamer-treated mice, with all wounds healed at 10 days. There was also no statistically significant difference in wound size at any time. Cantharidin induced a similar level of cellular influx in treated and control mice. Aptamer-treated mice exhibited a slight amount of inflammation-induced hemorrhage, though not the gross hemorrhage observed in thrombocytopenic mice.

Conclusion:
Inhibition of VWF activity does not delay cutaneous wound closure, despite inducing a severe hemostatic defect. Compared to other antithrombotic strategies, an aptamer against VWF could avert some undesirable side effects on host defense and tissue repair.
Abstract 52: IkB Kinase beta is a Key Modulator of Platelet Function, and the Remodeling of CARMA1-Bcl10-MALT1 Complex Formation
Authors: Karim Zubair A
Published: July 2021

Secretion plays an important role in platelet function, and hence the secretory machinery offers a unique target to modulate thrombogenesis. We previously established that IκB kinase (IKK)-β is phosphorylated upon platelet activation, regulates their SNARE machinery, and involves CBM (CARMA1, Bcl10, and MALT1) complex formation. However, the detailed role of IKKβ in platelet function, the mechanism by which it regulates CBM complex formation and downstream effector activation remain elusive. Using a knockout mouse model system (
IKK
β
flox/flox
-PF4Cre), we first showed that IKKβ plays a vital role in thrombus formation, and that it does so, in part, by regulating platelet functional responses, including dense and alpha granule release, αIIbβ3 activation, and PS exposure. Furthermore, we observed defects in compound fusion, platelet spreading, actin remodeling, and clot retraction. To this end, under clot retraction conditions in the knockout platelets, 7S complex formation was found to be inhibited. In terms of its signaling, using knockout platelets, we observed that IKKβ is required for the recruitment of Bcl10/MALT1 to CARMA1 upon platelet activation, i.e., CBM complex formation, and that this was due to the defective phosphorylation of Bcl10 and the IKKγ polyubiquitination. In conclusion, our data shows that IKKβ is a key regulator of platelet activation,
in vitro
and
in vivo
, and the remodeling of the CBM complex. Furthermore, our findings indicate that inducible clustering of signaling mediators and the formation of higher-order multi-protein complexes (i.e., the CBM complex) is a dynamic process, that supports nonlinear signaling networks and is important for platelet activation.
Abstract 520: Acoustic Tweezing Thromboelastometry
Authors: Luo Daishen , Khismatullin Damir , Holt R. Glynn
Published: July 2021

Background:
Critical care patients such as trauma and major surgery patients often develop coagulopathy due to depletion of both pro- and anti-coagulants. They are at high risk of both bleeding and thrombotic complications and require monitoring of their coagulation status. The contact of a blood sample with artificial surfaces and its exposure to clot activators, which happen in all commercially available coagulation analyzers, may lead to improper assessment of blood coagulation and thus errors in predicting bleeding/thrombosis risks.

Objective:
Real-time assessment of whole blood or blood plasma coagulation by novel non-contact acoustic tweezing technology.

Method:
4-5 microliter drops of whole blood collected from healthy volunteers or commercial control plasma were levitated in air by acoustic radiation forces. Their coagulation kinetics including reaction time, fibrin network formation time (FNFT), clot formation time and maximum clot strength was assessed from mechanical (drop shape) and photo-optical (light intensity) data. FNFT was determined as a difference between mechanical and photo-optical reaction times.

Results:
Whole blood samples were exposed to pro- or anti-coagulants during levitation in the acoustic tweezing device. Changes in the coagulation status between different experimental groups were detected within 10 minutes. Similarly, less than 7 minutes was required to detect significant changes in reaction time, clot formation time and maximum clot strength between low, normal, and high fibrinogen level control plasma samples. FNFT was shown to be significantly reduced in plasma samples with a higher level of Factor XIII.

Conclusions:
The acoustic tweezing technology integrates photo-optical tests used in plasma coagulation assays with viscoelastic tests used in whole blood analysis. Its key disruptive features are the increased reliability and accuracy due to non-contact measurement, small sample volume requirement, relatively short procedure time (<10 minutes), and the ability to assess the level of Factor XIII function from FNFT measurements. Our technology addresses a current lack of reliable methods to measure blood coagulation in patients with coagulopathy.
Abstract 521: A Pilot Study Comparing Anti-Inflammatory Effects of Tranexamic Acid and Epsilon Aminocaproic Acid in Pediatric Congenital Heart Surgery
Authors: Parashar Nirbhay , Nafee Tarek , Lefaiver Cheryl , Steffensen Christine , Rizzo Vince , Kumar Vibhash , ElZein Chawki , Van Bergen Andrew
Published: July 2021

Background:
Antifibrinolytic agents are frequently used during pediatric heart surgery with cardiopulmonary bypass (CPB) to reduce transfusions. There are no studies comparing anti-inflammatory effects of antifibrinolytic agents, tranexamic acid (TXA) and Epsilon Aminocaproic acid (EACA). We compared the two agents in pediatric patients undergoing redo sternotomy with CPB.

Aim:
To compare anti-inflammatory effects of tranexamic acid versus aminocaproic acid in pediatric patients undergoing redo sternotomy and cardiopulmonary bypass.

Methods:
We conducted a randomized, double blind pilot study, comparing 10 subjects in each group receiving EACA and TXA. A cytokine panel was used to measure 13 inflammatory markers in pre, immediate post and 24 hours post-CPB period. Between group comparisons were tested with Mann-Whitney U tests and within group comparisons with Friedman tests.

Results:
Sample characteristics were comparable in both groups. Post CPB, plasma levels of 7 markers increased significantly (p<0.05) in both groups, including MCP-1; 3 increased significantly (p<0.03) in the EACA group alone, including GM-CSF; and 3 did not change over time (Table 1). No difference was found between groups for markers except for IL-10, which was significantly higher in EACA group post CPB. While absolute values of markers, chest tube output and volume of blood product needs were lower in TXA group, the differences were not statistically significant.

Conclusion:
There was no significant difference in anti-inflammatory profiles between EACA and TXA in this pilot study. GM-CSF and MCP-1 were increased in our study post CBP which has not been described in previous studies.
Abstract 525: Integration of Platelet Agonist Signaling During the Hemostatic Response in vivo
Authors: Shen Jian , Sampietro Sara , Wu Jie , Tang Juan , Matzko Chelsea N , Tang Chaojun , Yu Ying , Brass Lawrence F , Zhu Li , Stalker Timothy J
Published: July 2021

The local microenvironment within an evolving hemostatic plug shapes the distribution of soluble platelet agonists, resulting in a gradient of platelet activation emanating from the site of injury. While thrombin and ADP mediated platelet activation are clearly critical for establishment of this gradient, it remains unclear to what extent there is overlap of these and other platelet signaling pathways in time and space. In other words, does a single agonist dominate the platelet activation state within specific regions of a hemostatic plug, or do multiple agonist pathways need to be integrated to produce the gradient of platelet activation observed? Further, how do the relationships among different agonists change in the face of anti-platelet or anti-coagulant therapy and what are the consequences for hemostatic plug organization? Here, we used a combination of genetic and pharmacologic approaches coupled with real-time intravital imaging to examine how thrombin, thromboxane A
2
, P2Y
12
and epinephrine signaling are coordinated in time and space to regulate the development of platelet activation gradients at a site of vascular injury in vivo. We found that both thromboxane A
2
/TP signaling and ADP/P2Y
12
signaling are required for accumulation of minimally activated platelets in the outer shell region of hemostatic plugs. Interestingly, dual inhibition of both thromboxane and P2Y
12
signaling did not have an additive effect, but rather was similar to inhibition of either pathway alone. Epinephrine, which activates a Gi signaling pathway similar to ADP/P2Y
12
, was completely dispensable for hemostatic plug formation, even in the absence of P2Y
12
signaling. Finally, inhibition of P2Y
12
in the setting of sub-maximal thrombin activity revealed no role for P2Y
12
signaling in the development of robust platelet activation within the hemostatic plug core region. Taken together, these data shed new light on the way multiple platelet signaling pathways are integrated during the hemostatic response in vivo, and predict the outcome of therapeutically targeting specific platelet signaling pathways alone and in combination on hemostatic plug organization.
Abstract 528: Selectin-Binding Peptide Conjugate Molecule Decreases Murine Deep Vein Thrombosis Formation
Authors: Chambers Andrea M , Wodicka James R , Sangha Gurneet S , Panitch Alyssa , Goergen Craig
Published: July 2021

Introduction:
Deep vein thrombosis (DVT) affects 300,000 to 600,000 people annually with a recurrence in 20-50% after 10 years. Treatment is managed through anticoagulant drugs, but side effects occur due to systemic administration. The venous endothelium is thought to perpetrate thrombosis, especially through endothelial cell surface adhesion receptors P-selectin and E-selectin. In this study, a selectin receptor-binding molecule (EC-SEAL), made with a selectin-binding peptide and dermatan sulfate backbone was used to evaluate thrombus prevention in the infrarenal vena cava (IVC) of a mouse over 24 hours using high frequency ultrasound.

Purpose:
P-selectin and E-selectin have exemplified a link between DVT, leukocyte accumulation and rolling, and platelet binding. Thus, the purpose was to evaluate the ability of EC-SEAL to reduce leukocyte and platelet binding, and subsequently DVT, in a mouse model by blocking the corresponding endothelial cell receptors.

Methods:
A surgical murine model of thrombosis was induced by IVC stenosis with non-absorbable 6-0 silk suture and a 30-gauge needle (male C57BL/6; 10.8 ± 0.2 weeks old). The stenosis led to a 90% reduction in blood flow through the vessel. Within 10 minutes of suture placement around the IVC, 100 μl of saline (n = 6), EC-SEAL (n = 4), or heparin (n = 6) were systemically injected via the tail vein. Thrombus was observed before surgery, 6 hours and 24 hours after surgery using the Vevo 2100 high frequency ultrasound (FUJIFILM VisualSonics Inc.). Thrombus volume and IVC volume were analyzed using 3D ultrasound images, while thrombus weight was measured
ex vivo
.

Results and Conclusions:
Six hours after post-ligation, the mean thrombus percentage (thrombus volume/total IVC volume) was 63.1±5.3% for saline, 25.8±11.2% for EC-SEAL, and 26.1±11.4% for heparin. EC-SEAL and heparin groups had significantly lower 6-hour mean thrombus percentage (p < 0.05; ANOVA with a Tukey HSD post-hoc). The 24-hour mean thrombus percentage and thrombus weight were not statistically significant. The data suggest EC-SEAL reduces thrombus at 6 hours while avoiding reduced clotting time side effects associated with heparin. Further work will be needed to determine the true potential for EC-SEAL as a DVT treatment.
Abstract 538: Gum Arabic Acutely Inhibit PCSK9 and Upregulate LDLr in HepG2
Authors: Abulola Hatim , Dybel Stacey , Ochin Chinedu , Ali Mohamed , Garelnabi Mahdi
Published: July 2021

Introduction:
Gum arabic (GA) (Acacia Senegal) is widely used agent in both the pharmaceutical and food industries as an emulsifier and stabilizer. It is a branched complex polysaccharide molecule rich in calcium, potassium, and magnesium in addition to other minerals. Its aqueous solution is either neutral or slightly acidic. Human intake of GA with atorvastatin was reported to significantly reduce plasma low density lipoprotein, and triglyceride. This study is designed to test mechanism (s) of its action on HepG2 cell.

Hypothesis:
The plasma lipids reduction associated with GA intake is likely associated with PCSK9/LDLr mechanism.

Methods:
Gum Arabic was dissolved in water and irradiated under UV light for 5 hours. Three final concentrations: 5ng/ml, 100ng/ml and 500 ng/ml were used for treatment of HepG2 cells at 6 hours, 12 hours and 24 hours using 12 well plates. Following treatment cells were collected into Trizol® solution; RNA was extracted for gene expression following Trizol® supplier protocol. cDNA was synthesized and the expression of PSCK9, LDLR and other related genes were tested using real time PCR.

Results:
The use of viscous (soluble) fiber (e.g., oats, guar, pectin, and psyllium) as therapeutic dietary options to enhance lowering of LDL cholesterol in primary and secondary prevention of CHD was well known, however there is no study to date that have shown the GA cholesterol lowering mechanism(s). In this study cell treated with GA for six hours has resulted in approximately 50 fold reductions of PCSK9 gene expression across the various concentrations, and approximately 25 fold up regulation of LDLr gene. Prolonged treatments beyond the six hours have shown differential responses.

Conclusions:
This study demonstrates that Gum Arabic has acute anti PCSK9 properties, as well as beneficial LDLr modulation effects. Our data explain the modulatory mechanism of Gum Arabic plasma LDL lowering effects associated with its intake in humans.
Abstract 539: Lipogenesis Regulates the Response of Cardiac Muscle to Ischemic Stress Through Sarcoplasmic Reticulum Calcium Atpase
Authors: Spears Larry D , Razani Babak , Funai Katsuhiko , Feng Chu , Song Haowei , Semenkovich Clay F
Published: July 2021

Dysfunctional calcium homeostasis is a hallmark of heart failure, a sequela of myocardial infarction (MI). Previous research has shown that mice deficient in cardiac fatty acid synthase (FAS), the rate-limiting enzyme for de novo lipogenesis, have dysfunctional calcium signaling and a predisposition to heart failure. Yet the mechanisms linking endogenous lipid synthesis, myocardial calcium homeostasis, and cardiac function remain poorly understood. A major step in calcium handling is the transport of cytosolic calcium back into the sarcoplasmic reticulum (SR) by sarcoplasmic reticulum calcium ATPase (SERCA), and recent evidence suggests that SR lipid composition can affect SERCA function. To determine whether FAS plays a role in SERCA activity in the heart, we evaluated FASKard (
FAS K
nockout in the Myoc
ard
ium) mice in the setting of experimental myocardial infarction (MI). Deletion of heart FAS caused a decrease in SERCA activity in younger adult (4 months) and aged (12 months) mice when compared to control (Ctrl) mice of the same age. Lipidomic analysis of the SR showed the reduced SERCA activity in FASKard mice was due to an altered SR phospholipid composition, leading to an imbalance of SR phosphatidylcholine to phosphatidylethanolamine phospholipid ratios. This altered ratio of SR phospholipids was due to a decrease in the abundance of specific phosphatidylethanolamine phospholipid species. MI caused a decrease in SERCA activity in both Ctrl and FASKard mice hearts. FASKard mice after MI had impaired survival with concomitantly decreased SERCA activity compared to Ctrl mice after MI. FASKard MI mice also had a perturbed ratio of SR phosphatidylcholine to phosphatidylethanolamine phospholipids compared to Ctrl MI mice, due to a decrease in certain phosphatidylethanolamine phospholipids. Additionally, MI induced FAS expression in the myocardium. These findings suggest that myocardial de novo lipogenesis is critical to the maintenance of the SR phospholipid composition, SR-mediated calcium homeostasis, and blunting the development of heart failure, particularly ischemic cardiomyopathy. Furthermore, FAS regulation of the SR lipid composition represents a novel therapeutic strategy in the treatment of heart failure.
Abstract 546: Metformin Significantly Increased Survival Rate in Patients With Diabetes and Comorbid Heart Disease
Authors: Liu Longjian , Brown Darryl , Simon Barbara , Eisen Howard J
Published: July 2021

Objective:
We assessed the hypothesis that metformin use significantly decreases risk of all-cause and heart disease (HD) mortality in patients with diabetes mellitus (DM).

Methods:
Subjects with DM aged ≥30 (n=3612) participating in 1999 -2010 National Health and Nutrition Examination Surveys, who had anti-DM drug therapy information, and were followed up by December 2011 were analyzed. Baseline DM, coronary heart disease (CHD) and heart failure (HF) were defined as a physician-diagnosis of the disease(s). Death from HD was classified using ICD10:I20-I25).

Results:
Mean (SD) age of 3612 participants was 63.1 (13.02), 83.4% (3056/3612) were under anti-DM therapies, and 20.3% was in metformin monotherapy, 4.9% in metformin combination (MCO), 24.8% in insulin, 45.4% in thiazolidinedione, sulfonylurea, and combinations, and 4.7% in any other antidiabetic drugs therapies. Baseline CHD and HF rates were 11.38% and 9.13%. At the end of follow-up, 1024 died from all-cause (21.86%), and 222 died from HD. Multivariate Cox’s proportional hazard regression analysis indicated that diabetic patients with comorbid HD (CHD and/or HF) had significant higher all-cause mortality than those without. However, these excess mortalities were significantly reduced in patients with metformin therapies than their counterparts without metformin. The hazard ratios (HR, 95%CI) of metformin use versus those without for all-cause mortality was 0.53 (0.38-0.76, p=0.001). Diabetic patients with CHD and with metformin therapies had a significantly lower 10-year all-cause mortality than those without metformin (33% vs. 64%, p<0.001). Similarly, diabetic patients with HF and with metformin therapies had a significantly lower 10-year all-cause mortality than their counterparts (42.8% vs. 73.1%, p<0.001). The effects of metformin on all-cause mortality were positively modified after adjusting dyslipidemia and insulin resistance.

In conclusion
, metformin significantly increased survival rates from all-cause mortality in diabetic patients with comorbid CHD and/or HF. The mortality risk reduction may be partially explained by an effect of metformin on a reduction of dyslipidemia and insulin resistance.
Abstract 548: Molecular Mechanism Behind Protein S Deficiency in Obese Patients
Authors: Pilli Vijaya S
Published: July 2021

Introduction:
Protein S is a vitamin K-dependent plasma protein, produced mainly in the liver; Protein S circulates in the blood at a concentration of 450 nM. Protein S is an anticoagulant, serving as a cofactor for APC and TFPI, and as an inhibitor of Factor IX (FIXa). Protein S deficiency causes deep vein thrombosis (DVT), increased risk of inflammation and, because DVT is a complication commonly observed in obese individuals, Protein S deficiency might be associated with obesity.

Aim:
To identify a correlation between Protein S deficiency and obesity, and identify the probable molecular mechanism behind the Protein S deficiency in the obese subjects.

Methods:
Immunoblots, ELISA, EMSA, CHIP, aPTT assay, and thrombin generation assay.

Results:
By ELISA, we measured a decrease in Protein S level in obese mice compared with wild type mice. In obesity, the liver becomes hypoxic, thus, we hypothesized that hypoxia and hypoxia inducible factor 1 alpha (HIF1α) may regulate Protein S expression in obesity. We found that a high fat diet induced HIF1α stability in mice. HIF1α levels were inversely proportional to Protein S levels, suggesting that HIF1α is a negative regulator of Protein S expression. We further identified a putative HIF1α binding site in the Protein S promoter, and, by using
in vitro
and
in vivo
assays, we demonstrated that HIF1α binds directly to the Protein S promoter and suppresses transcription. We further confirmed HIF1α-mediated Protein S transcriptional regulation
in vivo,
Plasma Protein S levels are increased in the liver-specific HIF1α knockout mouse whereas, liver-specific overexpression of HIF1α reduced the concentration of Protein S in the plasma.

Conclusion:
We conclude that HIF1α regulates Protein S expression in mouse liver and in obesity. Inhibition of HIF1α or intravenous injection of Protein S may reduce the occurrence of DVT in obese individuals.
Abstract 552: Aberrant Expression of MicroRNA-378a Induces Hepatic Insulin Resistance via Activation of ER Stress and the dsRNA Activated Protein Kinase PKR
Authors: Cheng Xiao , Song Yongyan , Su Qiaozhu
Published: July 2021

microRNAs(miRNAs) are noncoding RNAs with a length of 19 to 25 nt that are involved in posttranscriptional gene regulation by binding to the 3’-untranslated regions (3’-UTR) of target mRNA and impacting diverse cellular processes, including cell differentiation, energy metabolism and chronic inflammation. MicroRNA-378a (miR-378a) has been reported to be involved in adipose tissue browning and cancer development. However, its role in cellular stress signaling and hepatic insulin resistance has not yet been investigated. Here we reported that expression of hepatic miR-378a was upregulated by metabolic inflammatory inducers, such as high fructose feeding, bacterial lipopolysaccharide (LPS) and inflammatory cytokine TNFα. The elevated miR-378a subsequently targeted the 3’-UTR of PPARα which compromised mitochondrial fatty acid β-oxidation and induced mitochondrial and ER stress. miR-378a was further found to directly interacted with the dsRNA binding motifs within the dsRNA activated protein kinase PKR and activated the kinase to sustain the inflammatory stress and blunt the insulin signaling in hepatocytes. Genetic depletion of miR-378a rescued hepatocytes from mitochondrial and ER stress, systemic inflammation and insulin resistance induced by fructose and LPS.

Conclusion:
This study, for the first time, demonstrates that miR-378a is involved in mediating the metabolic inflammatory response in the onset of insulin resistance. This study further unveils a novel finding that miR-378a is capable of directly interacting with and activating a protein kinase PKR to sustain the stress signaling between mitochondria and ER. This discovery greatly broadens the physiological function of miR-378a by demonstrating that, in addition to regulate its target genes on the mRNA level, miRNA-378a is able to interact with RNA binding protein(s) and exerts its regulatory effect directly on the protein levels. Results from this study may provide rationale for using miR-378a as a pharmaceutical target in the treatment of insulin resistance.
Abstract 556: Lipoprotein Remodeling and Cholesterol Exchange Monitored Using Fluorescent Lipids and Proteins
Authors: Neufeld Edward B , Francone Nicolas O , Yilmaz Gizem , Gordon Scott M , Sviridov Denis O , Sampson Maureen , Demosky Stephen J , Pryor Milton , Amar Marcelo J , Remaley Alan T
Published: July 2021

We developed a sensitive and robust
in vitro
method to monitor lipoprotein cholesterol and protein exchange and lipoprotein remodeling, using non-exchangeable fluorescent phosphatidylethanolamine (PE) as a lipoprotein marker. We applied this method to monitor the exchange of unesterified cholesterol (FC) and apoA-I among isolated human lipoproteins and synthetic lipoprotein-X (LpX). Fluorescent FC, but not PE, rapidly equilibrated between VLDL and HDL, and transferred almost entirely from VLDL or HDL to LDL. Fluorescent apoA-I bound specifically to HDL and remodeled fluorescent PE and FC-labeled LpX into a new lipoprotein particle that contained both fluorescent lipids and apoA-I. LpX-derived fluorescent PE incorporated into plasma HDL only. The incorporation of LpX-derived fluorescent FC into plasma lipoproteins was similar to fluorescent FC alone, consistent with remodeling of LpX to HDL with concomitant exchange of FC between lipoproteins. LPL remodeled fluorescent PE and FC-tagged VLDL into a new particle containing both fluorescent lipids and apoA-I. We also developed a model system to study lipid transfer
in vitro
and
in vivo
by depositing lipids on calcium silicate hydrate crystals to form dense lipid coated donor particles that are readily separated from acceptor membranes and can be used as a surrogate for cell-dependent cholesterol efflux. These methodologies can readily be applied to study the other members of the vast lipoprotein proteome and the wide variety of remodeling events involved in lipoprotein-mediated lipid homeostasis in health and disease.
Abstract 561: GITR Activation Promotes Regulatory CD4+ T-cell Responses and Stimulates Leukocyte Recruitment in Atherosclerotic Plaques
Authors: Shami Annelie , Meiler Svenja , Winkels Holger , Gerdes Norbert , Lutgens Esther
Published: July 2021

Glucocorticoid-induced tumor necrosis factor receptor family-related protein (GITR) - a costimulatory molecule - is expressed on CD4(+) effector memory T cells and regulatory T cells as well as antigen-presenting cells and mast cells; while its ligand (GITRL) is mainly found on antigen-presenting cells and endothelial cells. However, the definitive role of GITR in atherosclerosis is not fully understood. Our hypothesis is that signaling through GITR plays a vital role in atherosclerosis progression.

Low-density lipoprotein receptor-deficient mice (Ldlr
-/-
) with B-cell-restricted overexpression of GITRL (
Gitrl
tg
) fed a high-cholesterol diet showed a profound increase in both CD4(+) effector memory T cells and regulatory T cells in secondary lymphoid organs in comparison to wild-type controls. Additionally, the number of regulatory T cells was significantly enhanced in the thymus and aorta of these mice along with increased GITRL and interleukin-2 transcript levels. Atherosclerotic lesions of Ldlr
-/-
Gitrl
tg
mice contained more total CD3
+
T cells as well as Foxp3
+
regulatory T cells overall, leading to significantly less severe atherosclerosis.

Conversely, atherosclerosis was found to be less severe in mice deficient in apolipoprotein E and GITR (ApoE
-/-
GITR
-/-
). Atherosclerotic lesions in these mice were found to contain less macrophages and CD3-positive T-cells. Perfusion assays using two-photon excitation microscopy revealed less wild type leukocyte adhesion on GITR-deficient endothelium, with a further reduction in adhesion by GITR-deficient leukocytes to both wild type and GITR-deficient endothelia. Finally, expression of GITR expression in human plaque tissue was significantly increased in ruptured plaques.

In conclusion, these data indicate that continuous GITR stimulation through B cell GITRL acts protective in a mouse model of atherosclerosis by regulating the balance between regulatory and effector memory CD4(+) T cells, while GITR activation on endothelial cells promotes atherogenesis by stimulating leukocyte recruitment into the plaque.
Abstract 563: Serum Amyloid A3 is a High Density Lipoprotein-associated Acute Phase Protein
Authors: De Beer Maria C , Ji Ailing , Noffsinger Victoria P , Schridas Preetha , De Beer Frederick C , Tannock Lisa R , Webb Nancy R
Published: July 2021

Acute phase serum amyloid (SAA) is a family of evolutionarily conserved, secreted proteins that exerts innate functions relevant to vascular disease. In humans, two SAA isoforms (SAA1 and SAA2) are highly induced in the liver and extrahepatic tissues under the regulation of inflammatory cytokines. During severe inflammation, SAA1/2 levels can increase ≥1000-fold in plasma, where it is found associated with HDL. Mice produce an additional acute phase SAA, SAA3, which is thought to be produced mainly by adipocytes and macrophages and has not previously been found circulating on HDL. The goal of this study was to investigate whether SAA3 serves as a third liver-derived, HDL-associated acute phase SAA in mice. Using isoform-specific oligonucleotide primers for qRT-PCR, we determined that SAA3 is transcriptionally induced to a similar extent (~2500-fold) compared to SAA1.1/2.1 (~6000-fold) in livers of C57BL/6 mice 19 hr after lipopolysaccharide (LPS) injection (100 μg/mouse). SAAs were also robustly induced in fat tissue (SAA1/2~100-fold; SAA3~400-fold). The analysis of primary mouse hepatocytes and
in situ
hybridization of mouse liver sections indicated that liver-derived SAAs are produced by hepatocytes and not other stromal cells, including Kupffer cells. All 3 SAA isoforms were detected in plasma of LPS-injected mice, although SAA3 levels were ~20% of SAA1/2. After separation by FPLC, virtually all of plasma SAA1/2 eluted with the HDL fraction, whereas ~15% of plasma SAA3 appeared to be lipid poor/free. HDL isolated from acute phase mouse plasma by density gradient ultracentrifugation was subjected to isoelectric focusing to determine the relative recovery of the various SAA isoforms. Whereas the bulk of plasma SAA1.1 was found in the d=1.063-1.21 fraction, only ~50% of SAA2.1 and ~10% of SAA3 was recovered after ultracentrifugation. These findings suggest that SAA3 may be more loosely associated with HDL compared to SAA1.1/2.1, which may give rise to lipid poor/free SAA3 that is susceptible to more rapid clearance in vivo. We conclude that SAA3 is a major hepatic acute phase SAA in mice that may produce systemic effects during inflammation. Future studies investigating SAA pathobiology in mice must take into account the previously under-studied SAA3.
Abstract 565: Ambient Ultrafine Particle Digestion Alters Gut Microbiota in Association With Increased Atherogenic Lipid Metabolites
Authors: Li Rongsong , Yang Jieping , Saffari Arian , Baek kyung , Jacobs Jonathan , Pisegna Joseph , Li Zhaoping , sioutas Constantinos , Navab Mohamad , Hsiai Tzung
Published: July 2021

Ambient particulate matter (PM) exposure is associated with atherosclerosis and inflammatory bowel disease. Ultrafine particles (UFP,
d
p
< 0.1-0.2 μm) are redox active components of PM. We hypothesized that orally ingested UFP promoted atherogenic lipid metabolites in both the intestine and plasma via altered gut microbiota composition. Low density lipoprotein receptor-null (
Ldlr
-/-
) mice on a high-fat diet were orally administered with vehicle control or UFP (40 μg/mouse/day) for 3 days a week. After 10 weeks, UFP ingested mice developed macrophage and neutrophil infiltration in the intestinal villi, accompanied by elevated cholesterol but reduced coprostanol levels in the cecum, as well as elevated atherogenic lysophosphatidylcholine (LPC 18:1) and lysophosphatidic acids (LPAs) in the intestine and plasma. At the phylum level, Principle Component Analysis revealed significant segregation of microbiota compositions which was validated by Beta diversity analysis. UFP-exposed mice developed increased abundance in Verrocomicrobia but decreased Actinobacteria, Cyanobacteria, and Firmicutes as well as a reduced diversity in microbiome. Spearman’s analysis negatively correlated Actinobacteria with cecal cholesterol, intestinal and plasma LPC18:1, and Firmicutes and Cyanobacteria with plasma LPC 18:1. Thus, ultrafine particles ingestion alters gut microbiota composition, accompanied by increased atherogenic lipid metabolites. These findings implicate the gut-vascular axis in a atherosclerosis model.
Abstract 569: Albuminuria, the HDL Proteome, and Prevalent Coronary Artery Calcium in Type 1 Diabetes
Authors: Shao Baohai , Zelnick Leila , Wimberger Jake , Brunzell John , Himmelfarb Jonathan , Bornfeldt Karin E , de Boer Ian H , Heinecke Jay W
Published: July 2021

Patients with type 1 diabetes (T1D) are at high risk of atherosclerotic cardiovascular disease (CVD). Albuminuria is strongly associated with CVD risk and fully accounts for the excess overall mortality risk in some T1D cohorts. One important contributor might be alterations of the HDL proteome. In the current study, we tested the hypotheses that albuminuria is associated with alterations in the HDL proteome in T1D, and that these alterations are associated with prevalent CVD. We performed a cross-sectional study of 191 Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications (DCCT/EDIC) participants selected according to levels of urine albumin excretion (67 persistent normoalbuminuria, 64 persistent microalbuminuria, and 60 persistent macroalbuminuria). We used targeted proteomics and isotope dilution tandem-mass spectrometry to quantify the concentration of 47 proteins in HDL. Adjusting for age, sex, DCCT treatment group, duration of diabetes, lipid-lowering medications, renin-angiotensin system inhibitors, smoking, BMI, and HbA1c, and after accounting for multiple comparisons, six proteins in HDL were significantly associated with albuminuria (2 increased and 4 decreased). For example, compared to normoalbuminuria, macroalbuminuria was associated with 57% and 177% higher AMBP (
P
=0.0003) and PTGDS (
P
=0.0006), respectively, and 28% and 27% lower PON1 (
P
=0.002) and PON3 (
P
=0.008), respectively. Furthermore, PON1 and PON3 in HDL were strongly and negatively associated with the presence of coronary artery calcium, with odds ratios per 1-SD difference of 0.63 (0.43-0.92, 95% CI,
P
=0.02) for PON1 and 0.59 (0.40-0.87, 95% CI,
P
=0.008) for PON3. Our observations indicate that the HDL proteome is remodeled in albuminuric patients with T1D, and that these alterations in HDL’s protein cargo may mediate, in part, the relationship of albuminuria with CVD. Because PON1 and PON3 are anti-atherogenic in hypercholesterolemic mice, our data suggest that low levels of PON1 and PON3 in HDL increase the risk of atherosclerosis in diabetic humans. In future studies, it will be important to determine whether reductions of PON1 and PON3 in HDL predict the risk of future CVD in subjects with T1D.