Myocardial Histopathology Research Articles

Subendocardial ischemic-like state in patients with severe aortic stenosis: insights from myocardial histopathology and ultrastructure

Abstract Background myocardial adaptation to severe aortic stenosis (AS) is a complex process that involves myocardial fibrosis (MF) beyond cardiomyocyte hypertrophy. Perfusion impairment is believed to be involved in myocardial remodeling in chronic pressure overload. Aim to describe morphological and ultrastructural myocardial changes at EMB, possibly reflecting subendocardial ischemia, in a group of patients with severe AS referred to surgical aortic valve replacement (AVR), with no previous history of ischemic cardiomyopathy. Methods one-hundred-fifty-eight patients (73 [68-77] years, 50% women) referred for surgical AVR because of severe symptomatic AS with pre-operative clinical and imaging study and no previous history of ischemic cardiomyopathy. Intra-operative septal EMB was obtained in 129 patients. Tissue sections were stained with Masson ́s Trichrome for MF quantification and periodic acid-Schiff (PAS) staining was performed to assess the presence of intracellular glycogen. Ultrastructure was analyzed through Transmission electron microscopy (TEM). Results MF totalized a median fraction of 11.90% [6.54-19.97%] of EMB, with highly prevalent perivascular involvement (95.3%). None of the samples had histological evidence of myocardial infarction. In 58 patients (45%) we found subendocardial groups of cardiomyocytes with cytoplasmatic enlargement, vacuolization and myofiber derangement, surrounded by extensive interstitial fibrosis. These cardiomyocytes were PAS positive, PAS-diastase resistant and Alcian Blue/PAS indicative of the presence of neutral intracellular glyco-saccharides. At TEM there were signs of cardiomyocyte degeneration with sarcomere disorganization and reduction, organelle rarefaction but no signs of intracellular specific accumulation. Conclusion almost half of the patients with severe AS referred for surgical AVR have histological and ultrastructural signs of subendocardial cardiomyocyte ischemic insult. It might be inferred that local perfusion imbalance contributes to myocardial remodeling and fibrosis in chronic pressure overload.

Colchicine ameliorates myocardial injury induced by coronary microembolization through suppressing pyroptosis via the AMPK/SIRT1/NLRP3 signaling pathway

BackgroundCoronary microembolization(CME)is a common complication in acute coronary syndrome and percutaneous coronary intervention, which is closely related to poor prognosis. Pyroptosis, as an inflammatory programmed cell death, has been found to be associated with CME-induced myocardial injury. Colchicine (COL) has potential benefits in coronary artery disease due to its anti-inflammatory effect. However, the role of colchicine in pyroptosis-related CME-induced cardiomyocyte injury is unclear. This study was carried out to explore the effects and mechanisms of colchicine on myocardial pyroptosis induced by CME.MethodsThe CME animal model was constructed by injecting microspheres into the left ventricle with Sprague-Dawley rats, and colchicine (0.3 mg/kg) pretreatment seven days before and on the day of modeling or compound C(CC)co-treatment was given half an hour before modeling. The study was divided into 4 groups: Sham group, CME group, CME + COL group, and CME + COL + CC group (10 rats for each group). Cardiac function, serum myocardial injury markers, myocardial histopathology, and pyroptosis-related indicators were used to evaluate the effects of colchicine.ResultsColchicine pretreatment improved cardiac dysfunction and reduced myocardial injury induced by CME. The main manifestations were the improvement of left ventricular systolic function, the decrease of microinfarction area, and the decrease of mRNA and protein indexes related to pyroptosis. Mechanistically, colchicine increased the phosphorylation level of adenosine monophosphate-activated protein kinase (AMPK), promoted the expression of silent information regulation T1 (SIRT1), and inhibited the expression of NOD-like receptor pyrin containing 3 (NLRP3) to reduce myocardial pyroptosis. However, after CC co-treatment with COL, the effect of colchicine was partially reversed.ConclusionColchicine improves CME-induced cardiac dysfunction and myocardial injury by inhibiting cardiomyocyte pyroptosis through the AMPK/SIRT1/NLRP3 signaling pathway.

An eNAMPT-neutralizing mAb reduces post-infarct myocardial fibrosis and left ventricular dysfunction

Myocardial infarction (MI) triggers adverse ventricular remodeling (VR), cardiac fibrosis, and subsequent heart failure. Extracellular nicotinamide phosphoribosyltransferase (eNAMPT) is postulated to play a significant role in VR processing via activation of the TLR4 inflammatory pathway. We hypothesized that an eNAMPT specific monoclonal antibody (mAb) could target and neutralize overexpressed eNAMPT post-MI and attenuate chronic cardiac inflammation and fibrosis. We investigated humanized ALT-100 and ALT-300 mAb with high eNAMPT-neutralizing capacity in an infarct rat model to test our hypothesis. ALT-300 was 99mTc-labeled to generate 99mTc-ALT-300 for imaging myocardial eNAMPT expression at 2 hours, 1 week, and 4 weeks post-IRI. The eNAMPT-neutralizing ALT-100 mAb (0.4 mg/kg) or saline was administered intraperitoneally at 1 hour and 24 hours post-reperfusion and twice a week for 4 weeks. Cardiac function changes were determined by echocardiography at 3 days and 4 weeks post-IRI. 99mTc-ALT-300 uptake was initially localized to the ischemic area at risk (IAR) of the left ventricle (LV) and subsequently extended to adjacent non-ischemic areas 2 hours to 4 weeks post-IRI. Radioactive uptake (%ID/g) of 99mTc-ALT-300 in the IAR increased from 1 week to 4 weeks (0.54 ± 0.16 vs. 0.78 ± 0.13, P < 0.01). Rats receiving ALT-100 mAb exhibited significantly improved myocardial histopathology and cardiac function at 4 weeks, with a significant reduction in the collagen volume fraction (%LV) compared to controls (21.5 ± 6.1% vs. 29.5 ± 9.9%, P < 0.05). Neutralization of the eNAMPT/TLR4 inflammatory cascade is a promising therapeutic strategy for MI by reducing chronic inflammation, fibrosis, and preserving cardiac function.

Astragaloside IV-induced BMSC exosomes promote neovascularization and protect cardiac function in myocardial infarction mice via the miR-411/HIF-1α axis

This study focused on investigating the mechanism of the astragaloside IV-induced bone marrow mesenchymal stem cell exosome (AS-IV-MSC-exo)/microRNA(miR)-411/HIF-1α axis in affecting vascular neovascularization and protecting cardiac function in myocardial infarction (MI) mice. Exosomes (MSC-exo and AS-IV-MSC-exo) were separated by differential centrifugation and then characterized. MI mouse models were established by left anterior descending coronary artery ligation. Echocardiography was used to evaluate cardiac function. HE staining and Masson staining were performed to observe myocardial histopathology. Capillary density in the myocardium via immunohistochemistry and quantified the expression of vascular endothelial growth factor (VEGF) via RT-qPCR. The expression of miR-411 and HIF-1α was tested by RT-qPCR and western blot and the targeting relationship of miR-411 and HIF-1α was verified by bioinformatics website and dual luciferase reporter gene assay. Exosomes with lipid bi-layer membrane structure, expressing exosomal surface marker proteins, and being taken up by cardiomyocytes could be successfully isolated utilizing ultracentrifugation. Intramyocardial injection of MSC-exo could restore cardiac function, decrease myocardial pathological changes and collagen deposition, and promote neovascularization in MI mice; the effect of AS-IV-MSC-exo was more significant. The ability of AS-IV-MSC-exo to restore cardiac function, lower myocardial pathological changes and collagen deposition, and promote neovascularization in MI mice was diminished when miR-411 expression in AS-IV-MSC-exo was reduced. Mechanistically, miR-411 was found to target and inhibit HIF-1α expression. Overexpression of HIF-1α impaired the impact of AS-IV-MSC-exo on improving cardiac function and promoting neovascularization in MI mice. AS-IV-MSC-exo improves cardiac function and promoted neovascularization via the miR-411/HIF-1α axis, thereby ameliorating MI.

Myocardial edema, inflammation, and injury in human heart donated after circulatory death are sensitive to warm ischemia and subsequent cold storage

BackgroundSingle-dose del Nido solution was recently used in human donation after circulatory death (DCD) heart procurement. We compared the effect of del Nido cardioplegia on myocardial edema, inflammatory response, and injury in human DCD hearts and human donation after brain death (DBD) hearts with different warm ischemic times (WIT) and subsequent cold saline storage times (CST). MethodsA total of 24 human hearts, including 6 in the DBD group and 18 in the DCD group—were procured for the research study. The DCD group was divided into 3 subgroups based on WIT: 20, 40, and ≥60 minutes. All hearts received 1 L of del Nido cardioplegia before being placed in cold saline for 6 hours. Left ventricular biopsies were performed at 0, 2, 4, and 6 hours. Temporal changes in myocardial edema, inflammatory cytokines (TNF-α, IL-6, and IL-1β), and histopathology injury scores were compared between the DBD and DCD groups. ResultsDCD hearts showed more profound changes in myocardial edema, inflammation, and injury than DBD hearts at baseline and subsequent CST. The DCD heart with WIT of 20 and 40 minutes with CST of 4 and 2 hours, respectively, appeared to have limited myocardial edema, inflammation, and injury. DCD hearts with WIT ≥60 minutes showed severe myocardial edema, inflammation, and injury at baseline and subsequent CST. ConclusionsSingle-dose cold del Nido cardioplegia and subsequent cold normal saline storage can preserve both DCD and DBD hearts. DCD hearts have been shown to be able to tolerate a WIT of 20 minutes and subsequent CST of 4 hours without experiencing significant myocardial edema, inflammation, and injury.

Panax quinquefolius saponins combined with dual antiplatelet therapy enhanced platelet inhibition with alleviated gastric injury via regulating eicosanoids metabolism

BackgroundPanax quinquefolius saponin (PQS) was shown beneficial against platelet adhesion and for gastroprotection. This study aimed to investigate the integrated efficacy of PQS with dual antiplatelet therapy (DAPT) on platelet aggregation, myocardial infarction (MI) expansion and gastric injury in a rat model of acute MI (AMI) and to explore the mechanism regarding arachidonic acid (AA)-derived eicosanoids metabolism.MethodsWistar rats were subjected to left coronary artery occlusion to induce AMI model followed by treatment with DAPT, PQS or the combined therapy. Platelet aggregation was measured by light transmission aggregometry. Infarct size, myocardial histopathology was evaluated by TTC and H&E staining, respectively. Gastric mucosal injury was examined by scanning electron microscope (SEM). A comprehensive eicosanoids profile in plasma and gastric mucosa was characterized by liquid chromatography-mass spectrometer-based lipidomic analysis.ResultsPQS+DAPT further decreased platelet aggregation, lessened infarction and attenuated cardiac injury compared with DAPT. Plasma lipidomic analysis revealed significantly increased synthesis of epoxyeicosatrienoic acid (EET) and prostaglandin (PG) I2 (potent inhibitors for platelet adhesion and aggregation) while markedly decreased thromboxane (TX) A2 (an agonist for platelet activation and thrombosis) by PQS+DAPT, relative to DAPT. DAPT induced overt gastric mucosal damage, which was attenuated by PQS co-administration. Mucosal gastroprotective PGs (PGE2, PGD2 and PGI2) were consistently increased after supplementation of PQS+DAPT.ConclusionsCollectively, PQS+DAPT showed synergistic effect in platelet inhibition with ameliorated MI expansion partially through upregulation of AA/EET and AA/PGI2 synthesis while suppression of AA/TXA2 metabolism. PQS attenuated DAPT-induced gastric injury, which was mechanistically linked to increased mucosal PG production.

Drug-refractory Heart Failure in Female Carrier of Duchenne Muscular Dystrophy: A Case of X-linked Dilated Cardiomyopathy.

A 56-year-old woman was referred to our hospital for the further evaluation of drug-refractory heart failure with a reduced ejection fraction. A family history interview revealed that men in her family had died of Duchenne muscular dystrophy (DMD), whereas she had no skeletal muscle disorder. Myocardial histopathology revealed a reduced dystrophin expression in the cardiomyocyte membrane, and a dystrophin (DMD) gene analysis identified a duplication in exon 8-9 on Xp21, suggesting that she had a cardiac-specific phenotype of dystrophinopathy, i.e. X-linked dilated cardiomyopathy (XLDCM). In conclusion, careful family history interviews and an investigation of dystrophinopathy are required to detect XLDCM in women.

Pacemaker lead insertion sites contribute to abnormalities of myocardial function and histopathology

Ventricular pacing can cause myocardial dysfunction, but how lead anchoring to the myocardium affects function has not been studied. The purpose of this study was to evaluate patterns of regional and global ventricular function in patients with a ventricular lead using cine cardiac computed tomography (CCT) and histology. This was a single-center retrospective study with 2 groups of patients with a ventricular lead: (1) those who underwent cine CCT from September 2020 to June 2021 and (2) those whose cardiac specimen was analyzed histologically. Regional wall motion abnormalities on CCT were assessed in relation to lead characteristics. For the CCT group, 122 ventricular lead insertion sites were analyzed in 43 patients (47% female; median age 19 years; range 3-57 years). Regional wall motion abnormalities were present at 51 of 122 lead insertion sites (42%) in 23 of 43 patients (53%). The prevalence of a lead insertion-associated regional wall motion abnormality was higher with active pacing (55% vs 18%; P < .001). Patients with lead insertion-associated regional wall motion abnormalities had a lower systemic ventricular ejection fraction (median 38% vs 53%; P < .001) than did those without regional wall motion abnormalities. For the histology group, 3 patients with 10 epicardial lead insertion sites were studied. Myocardial compression, fibrosis, and calcifications were commonly present directly under active leads. Lead insertion site-associated regional wall motion abnormalities are common and associated with systemic ventricular dysfunction. Histopathological alterations including myocardial compression, fibrosis, and calcifications beneath active leads may explain this finding.

Dynamic evolution and mechanism of myocardial glucose metabolism in different functional phenotypes of diabetic cardiomyopathy — a study based on 18 F-FDG microPET myocardial metabolic imaging

PurposeTo use 18 F-FDG microPET dynamic imaging to preliminarily identify the changes of myocardial glucose metabolism corresponding to different functional phenotypes of diabetic cardiomyopathy (DCM) in mice and elucidate their relationships.MethodsLeft ventricular function was measured by echocardiography in C57BL/KsJ-db/db (db/db) mice and their controls at 8, 12, 16, and 20 weeks of age to divide DCM stages and functional phenotypes. Myocardial histopathology was used to verify the staging accuracy and list-mode microPET dynamic imaging was conducted. The myocardial metabolic rate of glucose (MRglu) and the glucose uptake rate constant (Ki) were derived via Patlak graphical analysis, and the differences in myocardial glucose metabolism levels in different DCM stages were compared. The key proteins involved in myocardial glucose metabolism signaling pathway were analyzed by Western blotting to elucidate the underlying mechanism of abnormal glucose metabolism in DCM.ResultsCompared with the controls, the ratio of early diastolic transmitral flow velocity to early diastolic mitral annular tissue velocity (E/e’) of db/db mice was significantly increased from the age of 12 weeks, while the left ventricular ejection fraction (LVEF) was significantly decreased from the age of 16 weeks (all P < 0.05). Based on the staging criteria, 8 and 12 weeks (8/12w) db/db mice were in DCM stage 1 (diastolic dysfunction with normal LVEF), and 16 and 20 weeks (16/20w) db/db mice were in DCM stage 2/3 (diastolic and systolic dysfunction). The degree of myocardial fibrosis, glycogen deposition and ultrastructural damage in 16/20w db/db mice were more obvious than those in 8/12w group. The myocardial MRglu, Ki of db/db mice in 8/12w group or 16/20w group were significantly lower than those in the control group (all P < 0.05), while the myocardial standard uptake value (SUV) was not significantly reduced in the 8/12w group compared with the control group (P > 0.05). MRglu and SUV were moderately negatively correlated with the E/e’ ratio (r=-0.539 and − 0.512, P = 0.007 and 0.011), which were not significantly correlated with LVEF (P > 0.05). Meanwhile, Ki was not significantly correlated with LVEF or E/e’ ratio. The decreased expression of glucose transporter (GLUT) -4 in db/db mice preceded GLUT-1 and was accompanied by decreased phosphorylated AMP-activated protein kinase (p-AMPK) expression. Myocardial MRglu, Ki and SUV were significantly positively correlated with the expression of GLUT-4 (MRglu: r = 0.537; Ki: r = 0.818; SUV: r = 0.491; P = 0.000 ~ 0.046), but there was no significant correlation with GLUT-1 expression (P = 0.238 ~ 0.780).ConclusionsDuring the progression of DCM, with the changes of left ventricular functional phenotype, abnormal and dynamic changes of myocardial glucose metabolism can occur in the early stage.

Cardiac sarcoidosis: phenotypes, diagnosis, treatment, and prognosis.

Cardiac sarcoidosis (CS) results from epithelioid cell granulomas infiltrating the myocardium and predisposing to conduction disturbances, ventricular tachyarrhythmias, and heart failure. Manifest CS, however, constitutes only the top of an iceberg as advanced imaging uncovers cardiac involvement 4 to 5 times more commonly than what is clinically detectable. Definite diagnosis of CS requires myocardial biopsy and histopathology, but a sufficient diagnostic likelihood can be achieved by combining extracardiac histology of sarcoidosis with clinical manifestations and findings on cardiac imaging. CS can appear as the first or only organ manifestation of sarcoidosis or on top of pre-existing extracardiac disease. Due to the lack of controlled trials, the care of CS is based on observational evidence of low quality. Currently, the treatment involves corticosteroid-based, tiered immunosuppression to control myocardial inflammation with medical and device-based therapy for symptomatic atrioventricular block, ventricular tachyarrhythmias, and heart failure. Recent outcome data indicate 90% to 96% 5-year survival in manifest CS with the 10-year figures ranging from 80% to 90%. Major progress in the care of CS awaits the key to its molecular-genetic pathogenesis and large-scale controlled clinical trials.

Propofol versus insulin cardioplegia in valvular heart surgeries assessed by myocardial histopathology and troponin I

ABSTRACT Background and Aims Despite of the effectiveness of on pump cardiac surgeries in valvular diseases, cardioplegic arrest and myocardial reperfusion injury are still an obstacle. Many cardioprotective additives are tried but the obtained laboratory results are mixed. Our objective was to compare the effects of supplementing the cardioplegia solution with propofol or insulin evaluated not only by laboratory biomarkers but also with papillary muscle biopsy in patients undergoing on pump valvular surgeries. Methods Sixty adult patients were randomly assigned into three equal groups to receive: cold blood cadioplegia (control (C) group), supplemented with either 9 mg/L propofol 10% (P group), or 10 IU/L regular insulin (I group) Results Propofol induced significant higher myocardial protection presented by better histopathological grading of the obtained muscle biopsies when compared to either group C (P = 0.0460) or group I (P = 0.014), lower postoperative dysrhythmia (P = 0.004), lower troponin I release (40.57 ± 8.5 vs 47.7 ± 6.22-fold increase), and more eukalemic state with lower need for K supplementation than insulin. Conclusion Propofol was superior to insulin in providing higher grade of myocardial protection with lower troponin I release, more steady K level and lower postoperative complications.

Protective effect of ischaemic postconditioning combined with nicorandil on myocardial ischaemia‒reperfusion injury in diabetic rats

BackgroundThe diabetic heart exhibits a high sensitivity to ischaemia/reperfusion (I/R) injury. Diabetes mellitus (DM) can affect the efficacy of cardioprotective interventions and reduce the therapeutic potential of existing treatment options. This study aimed to investigate the feasibility of shifting from monotherapy to combination therapy in diabetic myocardial I/R injury.Methods6–8 week rats were randomized into 10 groups: sham, I/R, ischaemia postconditioning (I-Post), nicorandil (Nic), combination therapy (I-Post + Nic), DM sham, DM I/R, DM I-Post, DM Nic and DM I-Post + Nic. The extent of myocardial injury was clarified by measuring CK-MB and NO levels in plasma, ROS content in myocardial tissues, and TTC/Evans Blue staining to assess the area of myocardial infarction. Pathological staining of cardiac tissue sections were performed to clarify the structural changes in myocardial histopathology. Finally, Western blotting was performed to detect the phosphorylation levels of some key proteins in the PI3K/Akt signalling pathway in myocardial tissues.ResultsWe confirms that myocardial injury in diabetic I/R rats remained at a high level after treatment with I-Post or nicorandil alone. I-Post combined with nicorandil showed better therapeutic effects in diabetic I/R rats, and the combined treatment further reduced the area of myocardial injury in diabetic I/R rats compared with I-Post or nicorandil treatment alone (P < 0.001), as well as the levels of the myocardial injury markers CK-MB and ROS (P < 0.001); it also significantly increased plasma NO levels. Pathological staining also showed that diabetic rats benefited significantly from the combination therapy. Further mechanistic studies confirmed this finding. The protein phosphorylation levels of PI3K/Akt signalling pathway in the heart tissue of diabetic I/R rats were significantly higher after the combination treatment than after one treatment alone (all P < 0.05).ConclusionI-Post combined with nicorandil treatment maintains effective cardioprotection against diabetic myocardial I/R injury by activating the PI3K/Akt signalling pathway.

Effective Echocardiography Parameters to Identify Radiation-Induced Heart Disease in the Mouse Model

<h3>Purpose/Objective(s)</h3> Radiation-induced heart disease (RIHD) may occur after excessive radiotherapy to thoracic cancer. However, the effect of cardiac irradiation on left ventricle (LV) function is still controversial and no previous studies has evaluated the importance of stroke volume (SV) in RIHD. The hypothesis is that diastolic dysfunction with decreased SV could be potentially effective echocardiography parameters to identify RIHD in the mouse model. <h3>Materials/Methods</h3> C57 mice (n=5-10 per group) were treated with a single heart X-ray radiation for 20Gy or 0Gy. Echocardiography was performed to evaluate changes in LV dimensions and function before radiation and after radiation 4, 8 weeks. Elisa kits were used to detect the level of serum BNP and cTnI. HE, Masson's trichrome and Sirius red staining was utilized to assess cardiomyocytes size and myocardial fibrosis. Data are expressed as mean values (means±SEM). Statistical analyses were performed using the two-way repeated measures ANOVA or one-way ANOVA for multiple comparisons and student's t-test for unpaired two groups. In all statistical analyses, significance was defined as P < 0.05. <h3>Results</h3> The Myocardial histopathology indicated a markably increased cardiac fibrosis area and cardiomyocytes size at 8 weeks after radiation. Echocardiography analyses revealed that SV (27.02±1.131 vs. 41.64±2.021ul, P<0.0001), the LV end-diastolic volume (45.64±5.054 vs. 72.24±3.758ul, P=0.0018), and cardiac output (12.58±0.6669 vs. 19.16±0.9942ml/min, P<0.0002) of the 20Gy-mice have significantly decreased than the 0Gy-mice at 8 weeks. The dead 20Gy-mice had lower SV (22.40±0.6110 vs. 30.48±0.1827ul, P=0.0153) as compared to the survival 20Gy-mice. Moreover, the LV diastolic anterior (1.042±0.05123 vs. 0.8456±0.02860mm, P=0.0398) and posterior (0.8745±0.02558 vs. 0.7701±0.02551mm, P=0.0290) wall thickness of 20Gy-mice also had significantly increased than the 0Gy-mice. However, there were no significant difference of the EF (P>0.05), fraction shortening (P>0.05), heart rate (P>0.05), and LV end-systolic volume (P>0.05) between two groups at all the time points. In addition, the result of Elisa kits showed that serum BNP level (427.3±34.05 vs. 279.8±24.84ng/ml, P=0.0127) of 20Gy-mice was significantly higher than the 0Gy-mice at 8 weeks while cTnI was not significantly changed. <h3>Conclusion</h3> Diastolic dysfunction and decreased SV instead of systolic dysfunction occurs firstly in radiation-induced heart disease, which could be effective indexes to detect cardiac toxicity induced by irradiation.

Tanshinone IIA reduces pyroptosis in rats with coronary microembolization by inhibiting the TLR4/MyD88/NF-κB/NLRP3 pathway.

Pyroptosis is an inflammatory form of programmed cell death that is linked with invading intracellular pathogens. Cardiac pyroptosis has a significant role in coronary microembolization (CME), thus causing myocardial injury. Tanshinone IIA (Tan IIA) has powerful cardioprotective effects. Hence, this study aimed to identify the effect of Tan IIA on CME and its underlying mechanism. Forty Sprague–Dawley (SD) rats were randomly grouped into sham, CME, CME + low-dose Tan IIA, and CME + high-dose Tan IIA groups. Except for the sham group, polyethylene microspheres (42 µm) were injected to establish the CME model. The Tan-L and Tan-H groups received intraperitoneal Tan IIA for 7 days before CME. After CME, cardiac function, myocardial histopathology, and serum myocardial injury markers were assessed. The expression of pyroptosis-associated molecules and TLR4/MyD88/NF-κB/NLRP3 cascade was evaluated by qRT-PCR, Western blotting, ELISA, and IHC. Relative to the sham group, CME group's cardiac functions were significantly reduced, with a high level of serum myocardial injury markers, and microinfarct area. Also, the levels of caspase-1 p20, GSDMD-N, IL-18, IL-1β, TLR4, MyD88, p-NF-κB p65, NLRP3, and ASC expression were increased. Relative to the CME group, the Tan-H and Tan-L groups had considerably improved cardiac functions, with a considerably low level of serum myocardial injury markers and microinfarct area. Tan IIA can reduce the levels of pyroptosis-associated mRNA and protein, which may be caused by inhibiting TLR4/MyD88/NF-κB/NLRP3 cascade. In conclusion, Tanshinone IIA can suppress cardiomyocyte pyroptosis probably through modulating the TLR4/MyD88/NF-κB/NLRP3 cascade, lowering cardiac dysfunction, and myocardial damage.

Resveratrol pretreatment improves mitochondrial function and alleviates myocardial ischemia-reperfusion injury by up-regulating mi R-20b-5p to inhibit STIM2

This study aimed to explore the mechanism of resveratrol(RES) pretreatment in improving mitochondrial function and alleviating myocardial ischemia-reperfusion(IR) injury by inhibiting stromal interaction molecule 2(STIM2) through microRNA-20 b-5 p(miR-20 b-5 p). Ninety rats were randomly assigned into sham group, IR group, IR+RES(50 mg·kg~(-1) RES) group, IR+RES+antagomir NC(50 mg·kg~(-1) RES+80 mg·kg~(-1) antagomir NC) group, and IR+RES+miR-20 b-5 p antagomir(50 mg·kg~(-1) RES+80 mg·kg~(-1) miR-20 b-5 p antagomir) group, with 18 rats/group. The IR rat model was established by ligation of the left anterior descending coronary artery. Two weeks before the operation, rats in the IR+RES group were intraperitoneally injected with 50 mg·kg~(-1) RES, and those in the sham and IR groups were injected with the same dose of normal saline, once a day. Ultrasonic instrument was used to detect the left ventricular internal diameter at end-diastole(LVIDd) and left ventricular internal diameter at end-systole(LVIDs) of rats in each group. The 2,3,5-triphenyte-trazoliumchloride(TTC) method and hematoxylin-eosin(HE) staining were employed to detect the myocardial infarction area and histopathology, respectively. Real-time quantitative PCR(qRT-PCR) was carried out to detect the expression of miR-20 b-5 p in myocardial tissue. Oxygen glucose deprivation/reoxygenation(OGD/R) was performed to establish an OGD/R model of H9 c2 cardiomyocytes. CCK-8 assay was employed to detect H9 c2 cell viability. H9 c2 cells were assigned into the control group, OGD/R group, OGD/R+RES group(25 μmol·L~(-1)), OGD/R+RES+inhibitor NC group, OGD/R+RES+miR-20 b-5 p inhibitor group, mimic NC group, miR-20 b-5 p mimic group, inhibitor NC group, and miR-20 b-5 p inhibitor group. Flow cytometry was employed to detect cell apoptosis. Western blot was employed to detect the expression of B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), cleaved-cysteine proteinase 3(cleaved-caspase-3), and STIM2 in cells. The mitochondrial membrane potential(MMP) assay kit, reactive oxygen species(ROS) assay kit, and adenosine triphosphate(ATP) assay kit were used to detect the MMP, ROS, and ATP levels, respectively. Dual luciferase reporter gene assay was adopted to verify the targeting relationship between miR-20 b-5 p and STIM2. Compared with the sham group, the modeling of IR increased the myocardial infarction area, LVIDd, LVIDs, and myocardial pathology and down-regulated the expression of miR-20 b-5 p(P&lt;0.05). These changes were alleviated in the IR+RES group(P&lt;0.05). The IR+RES+miR-20 b-5 p antagomir group had higher myocardial infarction area, LVIDd, LVIDs, and myocardial pathology and lower expression of miR-20 b-5 p than the IR+RES group(P&lt;0.05). The OGD/R group had lower viability of H9 c2 cells than the control group(P&lt;0.05) and the OGD/R+RES groups(25, 50, and 100 μmol·L~(-1))(P&lt;0.05). Additionally, the OGD/R group had higher H9 c2 cell apoptosis rate, protein levels of Bax and cleaved caspase-3, and ROS level and lower Bcl-2 protein, MMP, and ATP levels than the control group(P&lt;0.05) and the OGD/R+RES group(P&lt;0.05). The OGD/R+RES+miR-20 b-5 p inhibitor group had higher H9 c2 cell apoptosis rate, protein levels of Bax and cleaved-caspase 3, and ROS level and lower Bcl-2 protein, MMP, and ATP levels than the OGD/R+RES group(P&lt;0.05). miR-20 b-5 p had a targeting relationship with STIM2. The expression of STIM2 was lower in the miR-20 b-5 p mimic group than in the mimic NC group(P&lt;0.05) and lower in the inhibitor NC group than in the miR-20 b-5 p inhibitor group(P&lt;0.05). RES pretreatment can inhibit the expression of STIM2 by promoting the expression of miR-20 b-5 p, thereby improving the function of mitochondria and alleviating myocardial IR damage.

Histopathological and epigenetic changes in myocardium associated with cancer therapy-related cardiac dysfunction.

Cancer therapy-related cardiac dysfunction (CTRCD) is commonly reported, but its histopathology, mechanisms, and risk factors are not known. We aimed to clarify the histopathology and mechanisms of CTRCD to identify risk factors. We performed myocardial histopathological studies on 13 endomyocardial biopsies from CTRCD patients, 35 autopsied cancer cases with or without cardiac dysfunction, and controls without cancer (10 biopsies and 9 autopsies). Cardiotoxicity risk scores were calculated based on medication; and patient-related risk factors, fibrosis, and cardiomyocyte changes were scored; and p53 and H3K27ac histone modification were evaluated by histological score (H-score). In the biopsy cases, all histopathological changes and the p53 evaluation were significantly higher in the CTRCD group than in the controls [p53 H-score; 63 (9.109) vs. 33 (5.099), P<0.05]. In patients with a short time between drug and disease onset (<4.2years), fibrosis and p53 positively correlated (r=0.76, P<0.05), and in those with late onset disease (>4.2years), cellular abnormalities and p53 trended to a positive correlation and cardiotoxicity risk scores and p53 positively correlated (r=0.95, P<0.05). A year after biopsy, the short-term group had significant recovery of ejection fraction compared with the long-term group (P<0.05). The CTRCD group had a significantly worse overall survival prognosis than the control group [hazard ratio 7.61 (95% confidence interval 1.30-44.6), P<0.05]. Autopsy cases with cancer treatment also had a high grade of histopathological changes, with even more severe changes in patients with cardiac dysfunction, and had increased p53 and H3K27ac expression levels, compared with controls. H-scores of p53 and H3K27ac showed a positive correlation in the CTRCD group in biopsy cases (r=0.62, P<0.05) and a positive correlation in autopsy cases. Our results indicate distinct morphological characteristics in myocardial histopathology associated with CTRCD. p53 and H3K27ac histone modification could be sensitive markers of CTRCD and suggest a mechanistic involvement of epigenetic changes.

Protective mechanisms of 10-gingerol against myocardial ischemia may involve activation of JAK2/STAT3 pathway and regulation of Ca2+ homeostasis

10-Gingerol (10-Gin), an active ingredient extracted from ginger, has been reported to have beneficial effects on the cardiovascular system. However, its protective effects on myocardial ischemia (MI) and the underlying cellular mechanisms are still unclear. To investigate the protection conferred by 10-Gin against MI injury and its potential mechanisms in cardiomyocytes via patch-clamp and molecular biology techniques. A rat MI model was established using the subcutaneous injection of isoproterenol (85 mg/kg) administered on two consecutive days. 10-Gin was pre-administered to rats for seven days to assess its cardio-protection. The patch-clamp and IonOptix Myocam detection techniques were used to investigated 10-Gin’s effects on L-type Ca2+ channels (LTCCs), Ca2+ transients and cell contractility in isolated rat cardiomyocytes. 10-Gin administration alleviated MI injury, improved cardiac function and myocardial histopathology, reduced myocardial infarct area, downregulated oxidative stress and Ca2+ levels, and decreased the expression of apoptotic factors. Importantly, 10-Gin led to an increase in phosphorylated Janus kinase 2 and signal transducer and activator of transcription 3 (JAK2 and STAT3, respectively) expressions. Furthermore, 10-Gin inhibited LTCCs in a concentration-dependent manner with a half-maximal inhibitory concentration of 75.96 μM. Moreover, 10-Gin administration inhibited Ca2+ transients and cell contractility. Our results suggest that 10-Gin exerts cardioprotective effects on MI in vivo and in vitro in connection with the inhibition of oxidative stress and apoptosis via activation of the JAK2/STAT3 signalling pathway, and regulation of Ca2+ homeostasis by LTCCs.

Effect of Yanshiqiangxin Decoction on Cardiomyocyte Apoptosis in Heart Failure Rats Based on PI3K/Akt/Caspase-9 Signaling Pathway

<bold>Objective</bold> To detect the effect of Yanshiqiangxin decoction on apoptosis of cardiomyocytes in rats with heart failure based on PI3K/Akt/Caspase-9 signalling pathway and to explore the possible mechanism of Yanshiqiangxin decoction in the treatment of heart failure. <bold>Methods</bold> A total of 60 Wistar male rats were randomly divided into the normal group, the model group, the low dose Yanshiqiangxin decoction group, the medium dose Yanshiqiangxin decoction group,the high dose Yanshiqiangxin decoction group and the Perindopril group according to the random number table method, with 10 cases in each group. Except for the normal group, the remaining five groups were modelled by injecting adriamycin solution to induce myocardial injury and to establish the rat model of heart failure. In the low, medium and high dose Yanshiqiangxin decoction groups, 4.325, 8.65 and 17.3 g/(kg·d) of raw drug were administered by gavage respectively; the Perindopril group was given perindopril solution by gavage at the dose of 4 mg/(kg·d). The gavage dose was calculated as 10 mL/(kg·d) ,the normal group and the model group were given the same volume of distilled water. One day after the last gavage, M-mode echocardiography [left ventricular ejection fraction (LVEF), left ventricular short-axis shortening (LVFS), left ventricular internal dimension diastole (LVIDd) and left ventricular internal dimension systole (LVIDs)] was performed through an ultra-high resolution small animal ultrasound imaging system; B-type natriuretic peptide (BNP) was measured by ELISA method; the apoptosis of cardiomyocytes was observed through laser confocal microscopy and the apoptosis index was calculated; the expression of apoptosis-related proteins Bax, Bcl-2 and PI3K/Akt/Caspase-9 signalling pathway proteins were detected by Western blot method. <bold>Results</bold> ① Cardiac ultrasound test results: compared with the normal group, the left ventricular anterior wall motion waveform of the model group was not obvious, close to linear motion, and LVEF and LVFS decreased significantly (<italic>P</italic>&lt;0.05), LVIDd and LVIDs increased significantly (<italic>P</italic>&lt;0.05), the left ventricle was significantly enlarged and the contractility was reduced. Compared with the model group, the amplitude of left ventricular anterior wall motion waveform increased at different degrees in the low, medium and high dose Yanshiqiangxin decoction group, LVEF and LVFS gradually increased with the increase of administration concentration (<italic>P</italic>&lt;0.05), while LVIDd decreased only in the medium dose Yanshiqiangxin decoction group (<italic>P</italic>&lt;0.05), LVIDs of the medium dose Yanshiqiangxin decoction group and the high dose Yanshiqiangxin decoction group were significantly reduced, the medium dose Yanshiqiangxin decoction group decreased the most significantly (<italic>P</italic>&lt;0.05). ② Serum BNP results: compared with the normal group, BNP of the model group was significantly increased (<italic>P</italic>&lt;0.05). Compared with the model group, BNP of the low, medium and high dose Yanshiqiangxin decoction group was significantly decreased, BNP gradually decreased with the increase of administration concentration (<italic>P</italic>&lt;0.05). ③ Myocardial histopathology results: Cardiomyocytes had no apoptosis and almost no positive signal was detected in the normal group; compared with the model group,the number of cardiomyocyte apoptosis of the low, medium, high dose Yanshiqiangxin decoction group and the perindopril group was significantly decreased, the number of cardiomyocyte apoptosis gradually decreased with the increase of administration concentration (<italic>P</italic>&lt;0.05). ④ PI3K/Akt/Caspase-9 signaling pathway protein expression levels and apoptotic factors: compared with the model group, PI3K and Akt protein expression, Bcl-2 content and Bcl-2/Bax ratio of the low, medium and high dose Yanshiqiangxin decoction group were significantly increased, PI3K and Akt protein expression, Bcl-2 content and Bcl-2/Bax ratio gradually increased with the increase of administration concentration (<italic>P</italic>&lt;0.05); Caspase-9 and Caspase-3 protein expression of the low, medium and high dose Yanshiqiangxin decoction group were significantly decreased, Bax content of the medium and high dose Yanshiqiangxin decoction group were significantly decreased, and gradually increased with the increase of administration concentration (<italic>P</italic>&lt;0.05). <bold>Conclusions</bold> Yanshiqiangxin decoction can improve cardiac function, reduce the content of BNP in serum and cardiomyocyte apoptosis, the mechanism may be related to the regulation of the PI3K/Akt/Caspase-9 signalling pathway.

Propofol Protects Myocardium From Ischemia/Reperfusion Injury by Inhibiting Ferroptosis Through the AKT/p53 Signaling Pathway

The molecular mechanism underlying the protective role of propofol against myocardial ischemia/reperfusion (I/R) injury remains poorly understood. Previous studies have shown that ferroptosis is an imperative pathological process in myocardial I/R injury. We hypothesized that propofol prevents myocardial I/R injury by inhibiting ferroptosis via the AKT/p53 signaling pathway. The ferroptosis-inducing agent erastin (E) and AKT inhibitor MK2206 (MK) were used to investigate the role of propofol in myocardial I/R injury. H9C2 cells treated without any reagents, erastin for 24 h, propofol for 1 h before adding erastin were assigned as the control (C), E, and E + P group, respectively. Cell viability, reactive oxygen species (ROS), and the expression of antioxidant enzymes, including ferritin heavy chain 1 (FTH1), cysteine/glutamate transporter (XCT), and glutathione peroxidase 4 (GPX4) in H9C2 cells. Rat hearts from the I/R + P or I/R groups were treated with or without propofol for 20 min before stopping perfusion for 30 min and reperfusion for 60 min. Rat hearts from the I/R + P + MK or I/R + MK groups were treated with or without propofol for 20 min, with a 10-min treatment of MK2206 before stopping perfusion. Myocardial histopathology, mitochondrial structure, iron levels, and antioxidant enzymes expression were assessed. Our results demonstrated that erastin increased H9C2 cell mortality and reduced the expression of antioxidant enzymes. I/R, which reduced the expression of antioxidant enzymes and increased iron or p53 (p < 0.05), boosted myocardium pathological and mitochondrion damage. Propofol inhibited these changes; however, the effects of propofol on I/R injury were antagonized by MK (p < 0.05). In addition, AKT siRNA inhibited the propofol-induced expression of antioxidant enzymes (p < 0.05). Our findings confirm that propofol protects myocardium from I/R injury by inhibiting ferroptosis via the AKT/p53 signal pathway.

Effects of Qilong Capsules on myocardial fibrosis and insufficient blood circulation in ischemic cardiomyopathy with Qi deficiency and blood stasis

Protective effect of Qilong Capsules(QL) on the myocardial fibrosis and blood circulation of rats with coronary heart disease of Qi deficiency and blood stasis type was investigated. Sleep deprivation and coronary artery ligation were used to construct a disease-symptom combination model, and 60 SD rats were divided into sham operation(sham) group, syndrome(S) group, disease and syndrome(M) group and QL group randomly. The treatment group received administration of QL 0.4 g·kg~(-1)·d~(-1). Other groups were given the same amount of normal saline. The disease indexes of each group [left ventricular end diastolic diameter(LVESD), left ventricular end systolic diameter(LVEDD), left ventricular ejection fraction(LVEF), left ventricular axis shortening rate(LVFS), myocardial histopathology, platelet morphology, peripheral blood flow] and syndrome indexes(tongue color, pulse, grip power) were detected. In sham group, cardiomyocytes and myocardial fibers were arranged neatly and densely with clear structures. The tongues' color in sham were light red, and the pulse shape were regular. RGB is a parameter reflected the brightness of the image of the tongue. In the S group, the amplitude and frequency of the animal's pulse increased accompanied by decreasing R,G,B, however, the decreased R,G,B was accompanied by reduced pulse amplitude in M group. And in M group, we observed fuzzy cell morphology, hypertrophied myocytes, disordered arrangement of cardiomyocytes and myocardial fibers, reduced peripheral blood flow and increased collagen volume fraction(CVF). Increased LVESD and LVEDD, and decreased LVEF and LVFS represented cardiac function in S group was significantly lower than that in sham. In QL group, the tongue's color was red and the pulse was smooth. The myocardial fibers of the QL group were arranged neatly and secreted less collagen. It improved the blood circulation in the sole and tail, and reversed the increasing of LVEDD, LVESD and the decreasing of LVEF and LVFS of M group. Platelets in M and S group showed high reactivity, and QL could decrease aggregation risk. In conclusion, Qilong Capsules has an obvious myocardial protective effect on ischemic cardiomyopathy, which may inhibit the degree of myocardial fibrosis and reduce platelet reactivity.