Differential cardiomyocyte transcriptomic remodeling during in vitro Trypanosoma cruzi infection using laboratory strains provides implications on pathogenic host responses.

BACKGROUND: Chagas disease can lead to life-threatening cardiac manifestations. Regional factors, including genetic characteristics of circulating Trypanosoma cruzi (T. cruzi), have attracted attention as likely determinants of Chagas disease phenotypic expression and Chagas cardiomyopathy (CCM) progression. Our objective was to elucidate the differential transcriptomic signatures of cardiomyocytes resulting from infection with genetically discrete T. cruzi strains and explore their relationships with CCM pathogenesis and progression. METHODS: HL-1 rodent cardiomyocytes were infected with T. cruzi trypomastigotes of the Colombian, Y, or Tulahuen strain. RNA was serially isolated post-infection for microarray analysis. Enrichment analyses of differentially expressed genes (fold-change ≥ 2 or ≤ 0.5) highlighted over-represented biological pathways. Intracellular levels of reactive oxygen species (ROS) were compared between T. cruzi-infected and non-infected HL-1 cardiomyocytes. RESULTS: We found that oxidative stress-related gene ontology terms (GO terms), 'Hypertrophy model', 'Apoptosis', and 'MAPK signaling' pathways (all with P < 0.01) were upregulated. 'Glutathione and one-carbon metabolism' pathway, and 'Cellular nitrogen compound metabolic process' GO term (all with P < 0.001) were upregulated exclusively in the cardiomyocytes infected with the Colombian/Y strains. Mean intracellular levels of ROS were significantly higher in the T. cruzi-infected cardiomyocytes compared to the non-infected (P < 0.0001). CONCLUSIONS: The upregulation of oxidative stress-related and hypertrophic pathways constitutes the universal hallmarks of the cardiomyocyte response elicited by T. cruzi infection. Nitrogen metabolism upregulation and glutathione metabolism imbalance may implicate a relationship between nitrosative stress and poor oxygen radicals scavenging in the unique pathophysiology of Chagas cardiomyopathy.

Endoplasmic Reticulum Selective Autophagy Alleviates Anthracycline-Induced Cardiotoxicity.

Background: The administration of anthracycline drugs induces progressive and dose-related cardiac damage through several cytotoxic mechanisms, including endoplasmic reticulum (ER) stress. The unfolded protein response plays a crucial role for mitigating misfolded protein accumulation induced by excessive ER stress. Objectives: We aimed to clarify whether endoplasmic reticulum-selective autophagy machinery (ER-phagy) serves as an alternative system to protect cardiomyocytes from ER stress caused by anthracycline drugs. Methods: Primary cultured cardiomyocytes, H9c2 cell lines, and cardiomyocyte-specific transgenic mice, all expressing ss-RFP-GFP-KDEL proteins, were used as ER-phagy reporter models. We generated loss-of-function models using RNA interference or gene-trap mutagenesis techniques. We assessed phenotypes and molecular signaling pathways using immunoblotting, quantitative polymerase chain reaction, cell viability assays, immunocytochemical and histopathological analyses, and cardiac ultrasonography. Results: The administration of doxorubicin (Dox) activated ER-phagy in ss-RFP-GFP-KDEL-transduced cardiomyocytes. In addition, Dox-induced cardiomyopathy models of ER-phagy reporter mice showed marked activation of ER-phagy in the myocardium compared to those of saline-treated mice. Quantitative polymerase chain reaction analyses revealed that Dox enhanced the expression of cell-cycle progression gene 1 (CCPG1), one of the ER-phagy receptors, in H9c2 cells. Ablation of CCPG1 in H9c2 cells resulted in the reduced ER-phagy activity, accumulation of proapoptotic proteins, and deterioration of cell survival against Dox administration. CCPG1-hypomorphic mice developed more severe deterioration in systolic function in response to Dox compared to wild-type mice. Conclusions: Our findings highlight a compensatory role of CCPG1-driven ER-phagy in reducing Dox toxicity. With further study, ER-phagy may be a potential therapeutic target to mitigate Dox-induced cardiomyopathy.

Porphyromonas gingivalis, a periodontal pathogen, impairs post-infarcted myocardium by inhibiting autophagosome-lysosome fusion.

While several previous studies have indicated the link between periodontal disease (PD) and myocardial infarction (MI), the underlying mechanisms remain unclear. Autophagy, a cellular quality control process that is activated in several diseases, including heart failure, can be suppressed by Porphyromonas gingivalis (P.g.). However, it is uncertain whether autophagy impairment by periodontal pathogens stimulates the development of cardiac dysfunction after MI. Thus, this study aimed to investigate the relationship between PD and the development of MI while focusing on the role of autophagy. Neonatal rat cardiomyocytes (NRCMs) and MI model mice were inoculated with wild-type P.g. or gingipain-deficient P.g. to assess the effect of autophagy inhibition by P.g. Wild-type P.g.-inoculated NRCMs had lower cell viability than those inoculated with gingipain-deficient P.g. This study also revealed that gingipains can cleave vesicle-associated membrane protein 8 (VAMP8), a protein involved in lysosomal sensitive factor attachment protein receptors (SNAREs), at the 47th lysine residue, thereby inhibiting autophagy. Wild-type P.g.-inoculated MI model mice were more susceptible to cardiac rupture, with lower survival rates and autophagy activity than gingipain-deficient P.g.-inoculated MI model mice. After inoculating genetically modified MI model mice (VAMP8-K47A) with wild-type P.g., they exhibited significantly increased autophagy activation compared with the MI model mice inoculated with wild-type P.g., which suppressed cardiac rupture and enhanced overall survival rates. These findings suggest that gingipains, which are virulence factors of P.g., impair the infarcted myocardium by cleaving VAMP8 and disrupting autophagy. This study confirms the strong association between PD and MI and provides new insights into the potential role of autophagy in this relationship.

The Pathophysiological Significance of "Mitochondrial Ejection" from Cells.

Mitochondria have beneficial effects on cells by producing ATP and contributing to various biosynthetic procedures. On the other hand, dysfunctional mitochondria have detrimental effects on cells by inducing cellular damage, inflammation, and causing apoptosis in response to various stimuli. Therefore, a series of mitochondrial quality control pathways are required for the physiological state of cells to be maintained. Recent research has provided solid evidence to support that mitochondria are ejected from cells for transcellular degradation or transferred to other cells as metabolic support or regulatory messengers. In this review, we summarize the current understanding of the regulation of mitochondrial transmigration across the plasma membranes and discuss the functional significance of this unexpected phenomenon, with an additional focus on the impact on the pathogenesis of cardiovascular diseases. We also provide some perspective concerning the unrevealed mechanisms underlying mitochondrial ejection as well as existing problems and challenges concerning the therapeutic application of mitochondrial ejection.

Neutrophil extracellular traps-mediated Beclin-1 suppression aggravates atherosclerosis by inhibiting macrophage autophagy.

A growing body of evidence suggests that neutrophil extracellular traps (NETs) critically contribute to the development of atherosclerosis. However, the detailed mechanism of how NETs promote atherogenesis remains unknown. In this study, we explored the role of NETs for promoting atherosclerosis by modulating the activity of autophagy in macrophages. NETs were effectively induced by a nicotine administration to the HL-60 cell-derived neutrophil-like cells. Treatment with NETs markedly suppressed both autophagosome formation and autophagosome-lysosome fusion in 7-ketocholesterol-treated macrophages, which are accompanied by the enhancement of inflammasome activity. NETs upregulate epidermal growth factor receptor (EGFR) activity, which enhances Beclin-1 phosphorylation of the tyrosine residues of Beclin-1 by EGFR, inhibits the PI3 kinase activity of the Beclin1-Vps34 complex, and suppresses autophagosome formation in macrophages. Furthermore, NET-induced activation of EGFR allows Rubicon to increase its expression, thereby suppressing autophagosome-lysosome fusion. In vivo experiments revealed that the suppression of NET formation by ablating peptidyl arginine deiminase-4 in neutrophil leukocytes resulted in the attenuation of atherosclerotic plaques in a nicotine-administered HFD-fed ApoE -/- mice. Taken together, these results suggest that NET-mediated EGFR-Beclin-1 signaling in the macrophages promotes atherogenesis by autophagy inhibition-mediated inflammasome activation.

Inflammatory cardiomyopathy of possibly overlapping aetiology: a case posing treatment dilemma and potential association.

We report on a 52-year-old Brazilian immigrant woman with past histories of chronic kidney disease and uveitis, presenting with symptomatic atrioventricular block. Her country of origin being endemic for Trypanosoma cruzi infection, we suspected Chagas disease as the aetiology, diagnosis of which was confirmed by serological tests. Further systemic workup identified an emerging nodular lesion in the lung, which turned out to be a sarcoid epithelioid granuloma on biopsy. Involvement of the kidneys and eyes was suggestive of systemic extension of the lung sarcoidosis. Although imaging modalities did not detect inflammatory foci in the myocardium, the rare coexistence of histologically proven sarcoidosis raised the intriguing concept of cardiac manifestation having arisen from two possibly overlapping aetiologies: Chagas disease and cardiac sarcoidosis. The case highlights a treatment dilemma increasingly likely to be encountered in this globalized world, and also raises the potential, but intriguing, association of these two diseases.

Plasma apolipopotein C-2 elevation is associated with Takayasu arteritis.

Takayasu arteritis (TAK) is an autoimmune systemic arteritis of unknown etiology. Although a number of investigators have attempted to determine biomarkers for diagnosing TAK, there exist no specific serological markers of this intractable disease. We undertook the exploration of novel serological markers which could be useful for an accurate diagnosis of TAK using an unbiased proteomics approach. The purified plasma samples from untreated patients with TAK and healthy individuals were separated by two-dimensional electrophoresis. The differentially expressed protein spots were detected by gel comparison and identified using matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MS). Next, we validated plasma concentrations of identified proteins by enzyme-linked immunosorbent assay (ELISA). Two-dimensional electrophoresis and numerical analysis revealed 19 spots and 3 spot clusters whose sum of the sample averages was ≥ 0.01, and the average concentrations were ≥ 1.5 times in the patient group compared with the control group. Among them, 10 spots and spot clusters that met the condition of the average spot concentration being 2.5 times more than that in the control group were selected. After processing these spots using MS and conducting MS/MS ion search, we identified 10 proteins: apolipoprotein C-2 (ApoC-2), actin, apolipoprotein A-1, complement C3, kininogen-1, vitronectin, α2-macroglobulin, 14-3-3 protein ζ/δ, complement C4, and inter-α-trypsin inhibitor heavy chain H4 isoform 1 precursor. Finally, ELISA demonstrated that plasma ApoC-2 level was significantly elevated in patients with TAK compared with that in healthy individuals. Thus, ApoC-2 would be a promising candidate biomarker for TAK diagnosis.

Linagliptin, A Xanthine-Based Dipeptidyl Peptidase-4 Inhibitor, Ameliorates Experimental Autoimmune Myocarditis.

This study sought to show the mechanism of how to ameliorate experimental autoimmune myocarditis (EAM) by administering dipeptidyl peptidase (DPP)-4 inhibitor linagliptin. The number of RAR-related orphan nuclear receptor gamma-positive Th17 cells infiltrated to the EAM myocardium was significantly attenuated by linagliptin treatment. Tandem mass spectrometry-based analysis demonstrated that DPP-4 binds to cathepsin G in EAM hearts, thereby protecting cathepsin G activity through inhibiting SerpinA3N activity. Linagliptin suppresses oxidative stress in EAM hearts as well. Thus, we found that DPP-4 plays a detrimental role in the progression of EAM by interacting with cathepsin G, which, in turn, suppresses SerpinA3N activity.

Rivaroxaban, a Direct Oral Factor Xa Inhibitor, Attenuates Atherosclerosis by Alleviating Factor Xa-PAR2-Mediated Autophagy Suppression.

The authors showed a mechanism for attenuating atherosclerosis by directly administering an oral factor Xa inhibitor (ie, rivaroxaban [RIV]). The autophagy activity of macrophages was significantly suppressed by factor Xa and was alleviated by the administration of RIV. However, factor Xa failed to inhibit 7-ketocholesterol-induced autophagy and inflammasome activation in protease-activated receptor 2 (PAR2) knockout macrophages. The atherosclerotic area of apolipoprotein E knockout mice was significantly reduced by the genetic ablation of PAR2, which was partially reversed by chloroquine. Thus, the authors found that RIV attenuates atherogenesis by inhibiting the factor Xa-PAR2-mediated suppression of macrophage autophagy and abrogating inflammasome activity.

Relationship of systemic pentraxin-3 values with coronary plaque components on optical coherence tomography and post-percutaneous coronary intervention outcomes in patients with stable angina pectoris.

BACKGROUND AND AIMS: Elevated pentraxin-3 (PTX3) values are associated with vulnerable plaque existence and poor outcomes in acute coronary syndrome patients. The clinical significance of PTX3 values in stable angina pectoris (SAP) patients is, however, undetermined. We investigated the relationship of systemic PTX3 values and coronary plaque components and post-percutaneous coronary intervention (PCI) outcomes in SAP patients. METHODS: We included 93 consecutive de-novo lesions in 93 SAP patients with a normal pre-PCI high-sensitivity cardiac troponin-T (<0.014 ng/mL), undergoing pre- and post-PCI optical coherence tomography (OCT). Systemic PTX3 values were obtained immediately pre- and post-PCI, at 24-h and 9-month post-PCI. RESULTS: Peak post-PCI PTX3 values correlated with thinnest fibrous cap thickness (r = -0.23, p = 0.03) and lipid length (r = 0.24, p = 0.03), and were higher in patients with lesions having OCT-derived thin-cap fibroatheroma (6.67 (3.19-7.33) vs. 3.13 (2.34-4.11) ng/mL, p = 0.04) and post-stenting irregular tissue protrusion (4.76 (3.31-6.80) vs. 2.98 (2.23-4.06) ng/mL, p = 0.003) than in those without. At 9-month follow-up, cardiac event-free survival was poorer in patients with a peak post-PCI PTX3 value ≥ 4.08 ng/mL (upper tertile) (log-rank test χ2 = 9.0; p = 0.003). Multivariate Cox regression analysis showed a peak post-PCI PTX3 value ≥ 4.08 ng/mL as an independent predictor of MACE (hazard ratio, 3.915; 95% CI, 1.129-13.583; p = 0.03). CONCLUSIONS: Peak post-PCI PTX3 values correlated with pre-PCI plaque characteristics and post-PCI outcomes, providing a good prognostic factor of outcomes in SAP patients undergoing elective PCI.

Impact of lesion angle on optical coherence tomography findings and clinical outcomes after drug-eluting stent implantation in curved vessels.

Tortuous coronary lesions are associated with adverse outcomes after implantation of bare metal or first-generation drug-eluting stents (DESs). We investigated the impact of lesion angle on vessel wall injuries and stent apposition as assessed by optical coherence tomography (OCT) after second- and newer-generation DES implantation. We investigated 95 de novo lesions treated with a single DES (62 platinum-chromium everolimus-eluting stents and 33 bioresorbable-polymer sirolimus-eluting stents). Post-intervention OCT findings were compared between angled lesions (≥ 45°; n = 33) and non-angled lesions (< 45°; n = 62). The 12-month clinical outcomes were also compared between the groups. Cross-sectional OCT analysis revealed that compared to non-angled lesions, angled ones had a significantly higher incidence of intra-stent dissection around the centre of the angle (19.7% vs. 10.8%, p = 0.01) and incomplete stent apposition (ISA) in the distal and proximal sub-segments (10.0% vs. 4.1%, p = 0.002; 15.3% vs. 7.9%, p < 0.001, respectively). Strut-based analysis also showed that angled lesions demonstrated a higher rate of malapposed strut in the distal and proximal sub-segments (3.0% vs. 0.9%, p < 0.001; 4.3% vs. 1.8%, p < 0.001, respectively). The 12 month clinical outcomes were comparable between the groups. Compared to non-angled lesions, angled coronary lesions were associated with a higher incidence of intra-stent dissection and ISA on post-intervention OCT after implantation of second- and newer-generation DESs.

Intravascular Ultrasound and Angioscopy Assessment of Coronary Plaque Components in Chronic Totally Occluded Lesions.

BACKGROUND: The in vivo lesion morphologies and plaque components of coronary chronic total occlusion (CTO) lesions remain unclear.Methods and Results:We investigated 57 consecutive CTO lesions in 57 patients with stable angina pectoris undergoing elective percutaneous coronary intervention with intravascular ultrasound (IVUS) and coronary angioscopy (CAS) examination. All CTO lesions were classified according to the proximal angiographic lumen pattern; tapered-type (T-CTO) and abrupt-type (A-CTO). The differences in the intracoronary images of these lesion types were evaluated according to the location within the CTO segment. A total of 35 lesions (61.4%) were T-CTO. T-CTO lesions had higher frequencies of red thrombi (proximal 71.4%; middle 74.3%; distal 31.4%; P<0.001) and bright-yellow plaques (yellow-grade 2-3) (48.6%; 74.3%; 2.9%; P<0.001) at the proximal or middle than at the distal subsegment; A-CTO lesions showed no significant differences among the 3 sub-segments. At the middle subsegment, T-CTO lesions showed higher frequencies of positive remodeling (51.4% vs. 18.2%, P=0.01) and bright-yellow plaques (74.3% vs. 13.6%, P<0.001) compared with A-CTO lesions. Multivariate analysis identified bright-yellow plaque as an independent predictor (odds ratio, 7.25; 95% confidence interval, 1.25-42.04; P=0.03) of the occurrence of periprocedural myocardial necrosis. CONCLUSIONS: The combination of IVUS and CAS analysis may be useful for identifying lesion morphology and plaque components, which may help clarify the pathogenetic mechanism of CTO lesions.

The clinical significance of echo-attenuated plaque in stable angina pectoris compared with acute coronary syndromes: A combined intravascular ultrasound and optical coherence tomography study.

BACKGROUND: Echo-attenuated plaque (EA) on intravascular ultrasound (IVUS) is related to poor outcomes after percutaneous coronary intervention (PCI) in acute coronary syndrome (ACS) patients. However, the clinical significance of EA in stable angina pectoris (SAP) patients compared with that in ACS patients remains unclear. We assessed the relationships between EA and unstable plaque characteristics in patients with ACS and SAP. METHODS: We investigated 609 coronary lesions in 609 patients (234 with ACS; 375 with SAP) undergoing pre-intervention IVUS and optical coherence tomography (OCT). The differences in plaque morphology and post-PCI outcomes were assessed according to the clinical status of ACS or SAP and the presence or absence of EA. RESULTS: EA was more frequent in patients with ACS than in those with SAP (44.0% vs. 25.1%, p < 0.001). SAP-EA lesions showed thicker fibrous cap (157 ±â€¯97 µm vs. 100 ±â€¯58 µm, p < 0.001), smaller lipid arc (208 ±â€¯76° vs. 266 ±â€¯99°, p < 0.001), smaller plaque burden (83.0 ±â€¯6.1% vs. 86.5 ±â€¯4.1%, p < 0.001), and lower frequency of transient slow-reflow phenomenon during PCI (21.3% vs. 51.5%, p < 0.001) than ACS-EA lesions, but similar plaque vulnerability compared with ACS-non-EA lesions. SAP-EA lesions had less frequent OCT-thrombus than ACS-non-EA lesions (20.2% vs. 71.2%, p < 0.001). CONCLUSIONS: SAP-EA lesions had less plaque vulnerability than ACS-EA lesions, but were comparable to ACS-non-EA lesions. Less frequent thrombus formation might differentiate SAP-EA lesions from ACS-non-EA lesions. A combined IVUS and OCT approach might be useful to assess plaque vulnerability in SAP-EA lesions compared with ACS lesions.

Urinary Liver-Type Fatty Acid-Binding Protein Level as a Predictive Biomarker of Acute Kidney Injury in Patients with Acute Decompensated Heart Failure.

BACKGROUND: There are no biological markers to predict the onset of acute kidney injury (AKI) in patients with acute decompensated heart failure (ADHF). Liver-type fatty acid-binding protein (L-FABP) levels are markedly upregulated in the proximal tubules after renal ischemia. We investigated whether urinary L-FABP is a suitable marker to predict AKI in ADHF patients. METHODS: We examined 281 consecutive patients with ADHF. Serum creatinine (Cr) and L-FABP levels were measured at admission and 24 and 48 h after admission. RESULTS: AKI developed in 104 patients (37%). Urinary L-FABP levels at admission were significantly higher in patients with AKI than in those without (33.0 vs. 5.2 µg/g Cr; p < 0.001). Multivariate analysis showed that baseline urinary L-FABP level was an independent predictor of AKI in ADHF patients (odds ratio 1.08, 95% confidence interval 1.05-1.12; p < 0.001). Receiver operating characteristic analysis showed that baseline urinary L-FABP level exhibited 94.2% sensitivity and 87.0% specificity at a cutoff value of 12.5 µg/g Cr. CONCLUSIONS: Urinary L-FABP level is useful for predicting the onset of AKI in patients with ADHF. The results of our study could help clinicians diagnose AKI in ADHF patients earlier, leading to possible improvements in the treatment of this group of patients.

Usefulness of Lipoprotein(a) for Predicting Clinical Outcomes After Endovascular Therapy for Aortoiliac Atherosclerotic Lesions.

PURPOSE: To evaluate the usefulness of serum lipoprotein(a) as a biomarker of clinical outcomes after endovascular therapy (EVT) for atherosclerotic aortoiliac lesions. METHODS: Serum lipoprotein(a) concentrations were measured at admission in 189 consecutive patients (median age 72 years; 160 men) with peripheral artery disease who underwent EVT for aortoiliac occlusive disease. The patients were dichotomized into 2 groups based on serum lipoprotein(a) levels ≤40 mg/dL (LOW; n=135) or >40 mg/dL (HIGH; n=54). After EVT, the incidences of major adverse limb events (MALE) were analyzed. Predictors of MALE were sought with a Cox proportional hazards analysis; results are presented as the hazard ratio (HR) and 95% confidence interval. RESULTS: At the median follow-up of 33 months (interquartile range 11, 54), MALE occurred in 44 (23.3%) patients. The MALE-free survival estimate was significantly lower in patients in the HIGH group (55.6% vs 85.2%, p<0.001). Independent predictors of MALE after EVT were hemodialysis (HR 2.23, 95% CI 1.04 to 4.78, p=0.039) and high lipoprotein(a) levels (HR 2.80, 95% CI 1.44 to 5.45, p=0.003). CONCLUSION: High lipoprotein(a) levels were associated with a higher incidence of MALE after EVT for patients with aortoiliac lesions.

Impact of optical coherence tomography- and coronary angioscopy-assessed neointimal tissue characteristics on occurrence of periprocedural myonecrosis in patients with in-stent restenosis.

Several characteristics of neointimal tissues, including neoatherosclerotic progression, have been reported in lesions with in-stent restenosis (ISR). However, the effects of these characteristics on outcomes after percutaneous coronary intervention (PCI) for ISR lesions remain unclear. We assessed the relationships between neointimal tissue characteristics and the occurrence of periprocedural myonecrosis (PMN) after PCI in ISR lesions. We investigated 72 ISR lesions in 72 patients with stable angina pectoris (SAP) who underwent pre- and post-revascularization optical coherence tomography (OCT) and coronary angioscopy (CAS). All lesions were classified as with PMN, defined by an elevated peak high-sensitivity cardiac troponin-T level during the 24-h post-PCI period, and without PMN. PMN was observed in 23 (31.9 %) lesions. PMN lesions had higher frequencies of OCT-derived thin-cap fibroatheroma (26.1 vs. 6.1 %, P = 0.03), CAS-derived intensive yellow neointima (30.4 vs. 10.2 %, P = 0.04), neointima with complex surface (60.9 vs. 28.6 %, P = 0.01), and CAS-derived atheromatous appearance (CAS-AAP), defined as yellow plaque including complex thrombi underneath disrupted neointimal coverage after ballooning (47.8 vs. 16.3 %, P = 0.008) at the most stenotic sites inside stents, compared to lesions without PMN. Multivariate logistic regression analysis identified CAS-AAP (odds ratio: 3.568, 95 % confidence interval: 1.109-11.475, P = 0.033) as an independent predictor of PMN. For ISR lesions in SAP patients, an OCT- and CAS-based assessment of neointimal tissue characteristics might help to predict the occurrence of PMN.