Rapid detection of goose megrivirus using TaqMan real-time PCR technology.

The aim of this study was to develop an efficient and accurate platform for the detection of the newly identified goose megrivirus (GoMV). To achieve this goal, we developed a TaqMan real-time PCR technology for the rapid detection and identification of GoMV. Our data showed that the established TaqMan real-time PCR assay had high sensitivity, with the lowest detection limit of 67.3 copies/µL. No positive signal can be observed from other goose origin viruses (including AIV, GPV, GoCV, GHPyV, and GoAstV), with strong specificity. The coefficients of variation of repeated intragroup and intergroup tests were all less than 1.5%, with excellent repeatability. Clinical sample investigation data from domestic Minbei White geese firstly provided evidence that GoMV can be transmitted both horizontally and vertically. In conclusion, since the TaqMan real-time PCR method has high sensitivity, specificity, and reproducibility, it can be a useful candidate tool for GoMV epidemiological investigation.

Risk assessment of rectal anastomotic leakage (RAREAL) after DIXON in non-emergency patients with rectal cancer.

BACKGROUND: The routine establishment of a diverting stoma (DS) remains controversial in every patient undergoing Dixon operation. We aimed to establish a model for the risk assessment of rectal anastomotic leak (RAREAL) after Dixon in non-emergency patients with rectal cancer, using routinely available variables, by which surgeons could individualize their approach to DS. METHODS: 323 patients who underwent Dixon operation for rectal cancer from January 2015 to December 2018 were taken as the model group for retrospective study. Univariable and multivariable logistic regression analysis was used to determine the independent risk factors associated with anastomotic leakage. We constructed the RAREAL model. 150 patients who underwent Dixon operation due to rectal cancer from January 2019 to December 2020 were collected according to the uniform criteria as a validation group to validate the RAREAL model. RESULTS: In the model group, multivariable analysis identified the following variables as independent risk factors for AL: HbA1c (odds ratio (OR) = 4.107; P = 0.044), Left colic artery (LCA) non preservation (OR = 4.360; P = 0.026), Tumor distance from the anal margin (TD) (OR = 6.373; P = 0.002). In the model group, the area under the curve (AUC) of the receiver operating characteristic (ROC) for evaluating AL with RAREAL was 0.733, and when RAREAL score = 2.5, its sensitivity, specificity and Youden index were 0.385, 0.973, 0.358, respectively. The AUC was 0.722 in the validation group and its sensitivity and specificity were 0.333 and 0.985, respectively, when RAREAL score = 2.5. CONCLUSION: The RAREAL score can be used to assess the risk of AL after Dixon operation for rectal cancer, and prophylactic DS should be proactively done when the score is greater than 2.5.

Three-vessel coronary infusion of cardiosphere-derived cells for the treatment of heart failure with preserved ejection fraction in a pre-clinical pig model.

Heart failure with preserved ejection fraction (HFpEF) is a major public health concern. Its outcome is poor and, as of today, barely any treatments have been able to decrease its morbidity or mortality. Cardiosphere-derived cells (CDCs) are heart cell products with anti-fibrotic, anti-inflammatory and angiogenic properties. Here, we tested the efficacy of CDCs in improving left ventricular (LV) structure and function in pigs with HFpEF. Fourteen chronically instrumented pigs received continuous angiotensin II infusion for 5 weeks. LV function was investigated through hemodynamic measurements and echocardiography at baseline, after 3 weeks of angiotensin II infusion before three-vessel intra-coronary CDC (n = 6) or placebo (n = 8) administration and 2 weeks after treatment (i.e., at completion of the protocol). As expected, arterial pressure was significantly and similarly increased in both groups. This was accompanied by LV hypertrophy that was not affected by CDCs. LV systolic function remained similarly preserved during the whole protocol in both groups. In contrast, LV diastolic function was impaired (increases in Tau, LV end-diastolic pressure as well as E/A, E/E'septal and E/E'lateral ratios) but CDC treatment significantly improved all of these parameters. The beneficial effect of CDCs on LV diastolic function was not explained by reduced LV hypertrophy or increased arteriolar density; however, interstitial fibrosis was markedly reduced. Three-vessel intra-coronary administration of CDCs improves LV diastolic function and reduces LV fibrosis in this hypertensive model of HFpEF.

Cytotoxic CD8+ T cells promote granzyme B-dependent adverse post-ischemic cardiac remodeling.

Acute myocardial infarction is a common condition responsible for heart failure and sudden death. Here, we show that following acute myocardial infarction in mice, CD8+ T lymphocytes are recruited and activated in the ischemic heart tissue and release Granzyme B, leading to cardiomyocyte apoptosis, adverse ventricular remodeling and deterioration of myocardial function. Depletion of CD8+ T lymphocytes decreases apoptosis within the ischemic myocardium, hampers inflammatory response, limits myocardial injury and improves heart function. These effects are recapitulated in mice with Granzyme B-deficient CD8+ T cells. The protective effect of CD8 depletion on heart function is confirmed by using a model of ischemia/reperfusion in pigs. Finally, we reveal that elevated circulating levels of GRANZYME B in patients with acute myocardial infarction predict increased risk of death at 1-year follow-up. Our work unravels a deleterious role of CD8+ T lymphocytes following acute ischemia, and suggests potential therapeutic strategies targeting pathogenic CD8+ T lymphocytes in the setting of acute myocardial infarction.

Concomitant systolic and diastolic alterations during chronic hypertension in pig.

The mechanical and cellular relationships between systole and diastole during left ventricular (LV) dysfunction remain to be established. LV contraction-relaxation coupling was examined during LV hypertrophy induced by chronic hypertension. Chronically instrumented pigs received angiotensin II infusion for4weeks to induce chronic hypertension (133 ±â€¯7 mmHg vs 98 ±â€¯5 mmHg for mean arterial pressure at Day 28 vs 0, respectively) and LV hypertrophy. LV function was investigated with the instrumentation and echocardiography for LV twist-untwist assessment before and after dobutamine infusion. The cellular mechanisms were investigated by exploring the intracellular Ca2+ handling. At Day 28, pigs exhibited LV hypertrophy with LV diastolic dysfunction (impaired LV isovolumic relaxation, increased LV end-diastolic pressure, decreased and delayed LV untwisting rate) and LV systolic dysfunction (impaired LV isovolumic contraction and twist) although LV ejection fraction was preserved. Isolated cardiomyocytes exhibited altered shortening and lengthening. Interestingly, contraction-relaxation coupling remained preserved both in vivo and in vitro during LV hypertrophy. LV systolic and diastolic dysfunctions were associated to post-translational remodeling and dysfunction of the type 2 cardiac ryanodine receptor/Ca2+ release channel (RyR2), i.e., PKA hyperphosphorylation of RyR2, depletion of calstabin 2 (FKBP12.6), RyR2 leak and hypersensitivity of RyR2 to cytosolic Ca2+ during both contraction and relaxation phases. In conclusion, LV contraction-relaxation coupling remained preserved during chronic hypertension despite LV systolic and diastolic dysfunctions. This implies that LV diastolic dysfunction is accompanied by LV systolic dysfunction. At the cellular level, this is linked to sarcoplasmic reticulum Ca2+ leak through PKA-mediated RyR2 hyperphosphorylation and depletion of its stabilizing partner.

1H NMR-based metabonomic revealed protective effect of Moutan Cortex charcoal on blood-heat and hemorrhage rats.

Moutan Cortex charcoal (MCC), the processed root bark of Paeonia suff ;ruticosa Andrews (Paeoniaceae), is a kind of traditional Chinese medicine (TCM) and has been used for treating blood-heat and hemorrhage(BHH)syndrome in China for thousands of years. In order to explore potential metabolic mechanism, 1H NMR-based metabonomics technique was applied to evaluate the effect of MCC on metabolic changes in plasma and urine of BHH rat models. Serum and urine samples were obtained from male SD rats with normal group, model group and MCC group for study. Based on 1H NMR spectra obtained from plasma and urine samples, principal component analysis (PCA) and orthogonal partial least-squares discriminant analysis (OPLS-DA) models were capable of distinguishing the three group. And the 13 pharmacodynamic biomarkers of MCC were identified in the plasma and urine. The results showed that BHH induced great metabolic disorders in plasma and urine metabolisms. However, MCC could reverse the imbalanced metabolites by alanine, aspartate and glutamate metabolism and citrate cycle (TCA cycle) pathway, and its effect was also confirmed by the general signs and pharmacodynamics assessments. The results indicated that NMR-based metabolomic profiling method is sensitive and specific enough to evaluate the MCC efficacy and mechanism of action on BHH syndromes.

The cardiac renin-angiotensin system is responsible for high-salt diet-induced left ventricular hypertrophy in mice.

AIMS: This study aimed to determine the role of the renin-angiotensin system (RAS) in high-salt (HS) diet-induced left ventricular hypertrophy (LVH). METHODS AND RESULTS: Swiss mice were subjected to regular salt (RS) diet (0.6% NaCl), HS diet (4% NaCl), and HS plus irbesartan (50 mg/kg/day) or ramipril (1 mg/kg/day). After 8 weeks, arterial pressure was similar in all groups and similar to baseline, whereas left ventricle/body weight ratio was higher in HS mice than in RS mice (P < 0.005). There were also significant increases in collagen density, angiotensin-converting enzyme activity, angiotensin II type 1 receptor (AT1 receptor) density, and extracellular signal-regulated kinase (ERK1/2) phosphorylation in the left ventricle. Interestingly, increases in wall thickness and ERK1 phosphorylation were more marked in the septum than in the rest of the left ventricle. Irbesartan or ramipril treatment prevented LVH and the increase in ERK phosphorylation and reduced collagen content and AT1 up-regulation but up-regulated AT2 receptors. CONCLUSION: In normal mice, HS diet induces septum-predominant LVH and fibrosis through activation of the cardiac RAS-ERK pathway, which can be blocked by irbesartan or ramipril, indicating a key role of the cardiac RAS in HS diet-induced LVH.

The influence of diet on isotope ratio mass spectrometry values.

OBJECTIVES: Athletes have increasingly used testosterone (T) and other endogenous anabolic steroids that cannot be detected by conventional gas chromatography-mass spectrometry. This led to gas chromatography-combustion-isotope ratio mass spectrometry(GC/C/IRMS), which measures the relative amount of 13C in urinary steroids. Because exogenous testosterone is relatively low in 13C content, this study will determine if consuming a diet low in 13C plants, such as soy, can be confused with a GC/C/IRMS-positive test for exogenous testosterone. DESIGN: Cross-sectional study in which 22 vegetarians known to consume a diet depleted of 13C isotope were compared with a geographic control group of 14 subjects consuming a normal diet. SETTING: Two distinct subject populations with respect to diet. SUBJECTS: Subjects were recruited from a soy-based cooperative and control volunteers. Twenty-two of 24 research subjects completed the protocol compared with 14 of 22 control subjects. INTERVENTIONS: Independent variables were delta13C IRMS values,urinary steroid profile, and isoflavone analysis. MAIN OUTCOME MEASURES: Comparisons were made with respect to dietary analysis, isoflavones, and urinary steroid measurements using GC-C-IRMS. RESULTS: The delta13C values for 2 major metabolites of T (androsterone and etiocholanolone) were lower for the vegetarians than the controls (P = 0.005). The vegetarians excreted a median of 23 micromol/d of total isoflavones compared with 2.7 micromol/d for the control group (P =0.0002). CONCLUSIONS: The carbon isotope ratios of urinary testosterone metabolites of vegetarians consuming a diet that is markedly depleted of 13C content were lower than that of control subjects, but not low enough to result in World Anti-Doping Agency criteria for a positive IRMS analysis.

Endoventricular porcine autologous myoblast transplantation can be successfully achieved with minor mechanical cell damage.

OBJECTIVE: Transplantation of skeletal myogenic precursor cells (mpc) into the myocardium using a non-surgical procedure. METHODS: Closed-chest mpc transplantation was assessed in pigs using the NOGA-Biosense device allowing both electromechanical mapping of the left ventricle (LV), and guided mpc injections through endocardium. RESULTS: We successively established that: (1) adequate preimplantation handling of mpc can be achieved when mpc are kept in 0.1% serum albumin-containing medium until implantation; (2) mpc are neither retained nor destroyed in the catheter or the needle and their passage does not affect their survival, growth and differentiation; (3) large numbers of autologous mpc can be actually transplanted in the LV myocardium by transendocardial route, as assessed by post-mortem examination of pigs injected with iron-loaded mpc; (4) cell injection into the myocardium does not induce conspicuous cell mortality since more than 80% of mpc recovered from LV tissue are alive 15 min after injection; (5) mpc injections can be guided into circumscribed LV targets such as infarcted areas, as assessed by comparison of map injection sites with location of iron-loaded mpc at post-mortem examination of LV myocardium. CONCLUSION: This new approach may pave the way for a large spectrum of cell therapies targeting myocardial diseases.

Contributions of heart rate and contractility to myocardial oxygen balance during exercise.

The respective contributions of heart rate (HR) reduction and left ventricular (LV) negative inotropy to the effects of antianginal drugs are debated. Accordingly, eight instrumented dogs were investigated during exercise at spontaneous and paced HR (250 beats/min) after administration of either saline, atenolol, or ivabradine (selective pacemaker current channel blocker). During exercise, atenolol and ivabradine (both 1 mg/kg iv) similarly reduced HR (-30% from 222 +/- 5 beats/min), and LV mean ejection wall stress was not altered. LV dP/dt(max) was reduced by atenolol but not ivabradine. Diastolic time (DT) was increased by atenolol versus saline (195 +/- 6 vs. 123 +/- 4 ms, respectively) and to a greater extent by ivabradine (233 +/- 11 ms). Myocardial oxygen consumption (MVo(2)) was lower under ivabradine and atenolol versus saline (6.7 +/- 0.6 and 4.7 +/- 0.4 vs. 8.1 +/- 0.6 ml/min, respectively, P < 0.05). Under pacing, DT and MVo(2) were similar between ivabradine and saline but significantly reduced with atenolol. Thus HR reduction and negative inotropy equally contribute to the reduction in MVo(2) during exercise in the normal heart. The negative inotropy limits the increase in DT afforded by HR reduction.

Role of creatine kinase in cardiac excitation-contraction coupling: studies in creatine kinase-deficient mice.

To understand the role of creatine kinase (CK) in cardiac excitation-contraction coupling, CK-deficient mice (CK-/-) were studied in vitro and in vivo. In skinned fibers, the kinetics of caffeine-induced release of Ca2+ was markedly slowed in CK-/- mice with a partial restoration when glycolytic substrates were added. These abnormalities were almost compensated for at the cellular level: the responses of Ca2+ transient and cell shortening to an increased pacing rate from 1 Hz to 4 Hz were normal with a normal post-rest potentiation of shortening. However, the post-rest potentiation of the Ca2+ transient was absent and the cellular contractile response to isoprenaline was decreased in CK-/- mice. In vivo, echocardiographically determined cardiac function was normal at rest but the response to isoprenaline was blunted in CK-/- mice. Previously described compensatory pathways (glycolytic pathway and closer sarcoplasmic reticulum-mitochondria interactions) allow a quasi-normal SR function in isolated cells and a normal basal in vivo ventricular function, but are not sufficient to cope with a large and rapid increase in energy demand produced by beta-adrenergic stimulation. This shows the specific role of CK in excitation-contraction coupling in cardiac muscle that cannot be compensated for by other pathways.