The Arcanobacterium (Actinomyces) pyogenes hemolysin, pyolysin, is a novel member of the thiol-activated cytolysin family.

Arcanobacterium (Actinomyces) pyogenes, an animal pathogen, produces a hemolytic exotoxin, pyolysin (PLO). The gene encoding PLO was cloned, and sequence analysis revealed an open reading frame of 1,605 bp encoding a protein of 57.9 kDa. PLO has 30 to 40% identity with the thiol-activated cytolysins (TACYs) of a number of gram-positive bacteria. The activity of PLO was found to be very similar to those of other TACYs, except that it was not thiol activated. The highly conserved TACY undecapeptide is divergent in PLO; in particular, the cysteine residue required for thiol activation has been replaced with alanine. However, mutagenesis of the alanine residue to cysteine did not confer thiol activation on PLO, suggesting a conformational difference in the undecapeptide region of this toxin. Specific antibodies against purified, recombinant PLO completely neutralized the hemolytic activity of A. pyogenes, suggesting that this organism produces a single hemolysin. Furthermore, these antibodies could passively protect mice against lethal challenge with A. pyogenes, suggesting that like other TACYs PLO is an important virulence factor in the pathogenesis of this organism.     

BDNF-Dependent Accumulation of Palmitoleic Acid in CNS Neurons

Although it is known that brain-derived neurotrophic factor (BDNF) plays a critical role in neuronal survival and differentiation, its effect on lipid homeostasis is poorly understood. To understand them, we here investigated the effect of BDNF on the fatty acid composition of primary neurons. A detailed analysis of the fatty acid composition of BDNF-stimulated primary neurons revealed that BDNF treatment led to a significant and selective increase in intracellular palmitoleic acid (PLO) levels. Correspondingly, BDNF induced the expression of the enzyme responsible for PLO synthesis [stearoyl-CoA desaturase-1]. In addition, this increase was suppressed by K252a, an inhibitor for tropomyosin-related kinase (Trk) receptors, indicating that BDNF-dependent increase in the PLO was mediated through the activation of TrkB. Further, PLO in culture media was reduced by BDNF treatment. This result suggested that BDNF suppressed extracellular release of PLO. Taken together, these data indicate that BDNF increases intracellular PLO both by activating its biosynthesis and by suppressing its extracellular release.     

Expression of Dishevelled-1 in wound healing after acute myocardial infarction: possible involvement in myofibroblast proliferation and migration

One of our previous studies indicated that the expression of beta-catenin, which is the key factor of wnt-frizzled pathway, increased significantly in the ischemic area of the rat heart 7 days after myocardial infarction (MI). Together with the results of other recent studies, we made an assumption that wnt-frizzled pathway may be involved in the controlled cell proliferation and migration during repair processes after MI. To verify this assumption we tried to investigate the expression of another signal transduction molecule called Dishevelled in wnt-frizzled pathway during the wound healing process after MI. The left descending coronary arteries of rats were ligated to induce MI. Immunohistochemistry SABC method and in situ hybridization were performed to detect the expression of Dishevelled-1. The results showed, that one day after MI, Dishevelled-1 mRNA but not protein expression was detected in the cells at the border zone of the infarction area; 4 days after MI the expression of Dishevelled-1 increased exclusively and cytoplasmic Dishevelled-1 was observed not only at the border zone but also in the infarct area; 7 days after MI, it seems that the expression reached its peak, the positive staining even spread into the endothelial and smooth muscle cells of the newly formed and pre-existing blood vessels in the infarction area; after that the Dishevelled-1 expression decreased abruptly and could hardly be detected 28 days after MI. Thus cytoplasmic Dishevelled-1 may be involved in the controlled proliferation and migration of myofibroblasts and vascular endothelial cells, hence play a role during the wound healing process after MI.     

The effect of hyperglycemia on the development of experimental myocardial infarct

Experiment on 72 dogs has shown that myocardial infarction (M1) against a background of hyperglycemia proceeds as shifts healing hyperactive MI. Shifts in carbohydrate metabolism results in disturbances of dynamics of the necrotic infarction zone processes that induces complications of healing changes in the content of myoglobin, creatine kinase, aspartateaminotransferase with MI against a background of hyperglycemia greatly differ from those typical of noncomplicated and complicated hyperactive MI.     

Temporal and spatial characteristics of apoptosis in the infarcted rat heart

Following myocardial infarction (MI), tissue repair/remodeling occurs in both the infarcted and noninfarcted myocardium. Apoptosis has been demonstrated to play an important role in these processes. In the present study, we sought to determine the temporal and spatial characteristics of apoptosis in the infarcted heart as well as to identify cells undergoing programmed cell death at different stages of repair/remodeling and their relationship to the expression of anti-/pro-apoptotic genes following MI. Our study has shown that apoptosis appears in both infarcted and noninfarcted myocardium, and cells undergoing apoptosis depend on the stage of healing. In the infarcted myocardium, apoptosis contributes to the loss of cardiomyocytes during the early stage of healing, elimination of inflammatory cells during the inflammatory phase of healing, and reduction of myofibroblasts with the fibrogenic phase of repair in the infarcted myocardium. In noninfarcted myocardium, cardiomyocyte apoptosis was observed from day 3 to 28 postMI. Cardiac apoptosis following MI is correlated with the increase of Bax expression.     

Exogenous Administration of a Recombinant Variant of TWEAK Impairs Healing after Myocardial Infarction by Aggravation of Inflammation

Background

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) and its receptor fibroblast growth factor-inducible 14 (Fn14) are upregulated after myocardial infarction (MI) in both humans and mice. They modulate inflammation and the extracellular matrix, and could therefore be important for healing and remodeling after MI. However, the function of TWEAK after MI remains poorly defined.

Methods and results

Following ligation of the left coronary artery, mice were injected twice per week with a recombinant human serum albumin conjugated variant of TWEAK (HSA-Flag-TWEAK), mimicking the activity of soluble TWEAK. Treatment with HSA-Flag-TWEAK resulted in significantly increased mortality in comparison to the placebo group due to myocardial rupture. Infarct size, extracellular matrix remodeling, and apoptosis rates were not different after MI. However, HSA-Flag-TWEAK treatment increased infiltration of proinflammatory cells into the myocardium. Accordingly, depletion of neutrophils prevented cardiac ruptures without modulating all-cause mortality.

Conclusion

Treatment of mice with HSA-Flag-TWEAK induces myocardial healing defects after experimental MI. This is mediated by an exaggerated neutrophil infiltration into the myocardium.     

Inhibition of an opioid-evoked vagal reflex in rats by naloxone,SMS 201-995 and ICI 154,129

Intravenous injection of opioid agonists in rats evokes a vagal reflex resulting in a fall in heart rate and blood pressure. Three opioid antagonists, naloxone, SMS 201-995, and ICI 154,129 were used to assess the nature of the opioid receptors that mediate the vagal reflex. The agonists used were morphine, Tyr-Pro-NMePhe-d-Pro-NH₂ (PLO17), and d-Ala²-Leu⁵-enkephalin (DADL). At challenge doses of morphine, PLO17, and DADL at five times the ED50 for bradycardia, the naloxone ED50 for DADL was nine times greater than that for morphine and PLO17. The pA₂ value of naloxone against DADL was significantly less than that for morphine and PLO17. The antagonist properties of SMS 201-995 were similar to those of naloxone. ICI 154,129, a putative delta receptor antagonist, was not, however, selective in its antagonism of opioid bradycardia. Both SMS 201-995 and ICI 154,129, when injected alone, produced changes in heart rate and blood pressure. The cardiovascular actions of the peptide antagonists were not affected by naloxone hydrochloride at doses up to 4 mg/kg i.v.     

Dexamethasone treatment of post-MI rats attenuates sympathetic innervation of the infarct region.

Sympathetic fiber innervation of the damaged region following injury represents a conserved event of wound healing. The present study tested the hypothesis that impaired scar healing in post-myocardial infarction (post-MI) rats was associated with a reduction of sympathetic fibers innervating the infarct region. In 1-wk post-MI rats, neurofilament-M-immunoreactive fibers (1,116 +/- 250 microm(2)/mm(2)) were detected innervating the infarct region and observed in close proximity to a modest number of endothelial nitric oxide synthase-immunoreactive scar-residing vessels. Dexamethasone (Dex) treatment (6 days) of post-MI rats led to a significant reduction of scar weight (Dex + MI 38 +/- 4 mg vs. MI 63 +/- 2 mg) and a disproportionate nonsignificant decrease of scar surface area (Dex + MI 0.54 +/- 0.06 cm(2) vs. MI 0.68 +/- 0.06 cm(2)). In Dex-treated post-MI rats, the density of neurofilament-M-immunoreactive fibers (125 +/- 47 microm(2)/mm(2)) innervating the infarct region was significantly reduced and associated with a decreased expression of nerve growth factor (NGF) mRNA (Dex + MI 0.80 +/- 0.07 vs. MI 1.11 +/- 0.08; P < 0.05 vs. MI). Previous studies have demonstrated that scar myofibroblasts synthesize NGF and may represent a cellular target of Dex. The exposure of 1st passage scar myofibroblasts to Dex led to a dose-dependent suppression of [(3)H]thymidine uptake and a concomitant attenuation of NGF mRNA expression (untreated 3.47 +/- 0.35 vs. Dex treated 2.28 +/- 0.40; P < 0.05 vs. untreated). Thus the present study has demonstrated that impaired scar healing in Dex-treated post-MI rats was associated with a reduction of neurofilament-M-immunoreactive fibers innervating the infarct region. The attenuation of scar myofibroblast proliferation and NGF mRNA expression may represent underlying mechanisms contributing to the diminished neural response in the infarct region of Dex-treated post-MI rats.     

Effects of Triterpenes on the Mutagenicities of Various Mutagens toward Salmonella

The effects of different dietary levels of gamma-linolenic acid (GLA) on lipid metabolism was studied in rats using a combination of evening primrose oil (EPO) and palm oil (PLO). EPO compared to PLO significantly reduced liver cholesterol and triglyceride after 4 weeks of feeding, and the effect remained even when EPO was mixed with PLO at the same weight basis. The serum triglyceride level also tended to be low on feeding EPO. Neither liver Δ6-desaturase and phospholipase A₂ activities nor aortic production of prostacyclin and thromboxane A₂ production by platelets were influenced significantly by the fat type, suggesting a peculiar effect of PLO. The percentage of arachidonic acid in liver, serum, and aortic phosphatidylcholine depended on the dietary level of GLA. A more distinct increase in arachidonic acid was observed in tissue triglycerides of rats fed EPO. GLA appears to exert favorable effects on lipid metabolism even when the P/S ratio was lowered from 13.7 of EPO to 1.8 of the 1:1 mixture of EPO and PLO.     

Early moderate exercise benefits myocardial infarction healing via improvement of inflammation and ventricular remodelling in rats

Thus far, the cellular and molecular mechanisms related to early (especially within 24 hours after acute myocardial infarct (MI)) exercise‐mediated beneficial effects on MI have not yet been thoroughly established. In the present study, we demonstrated that acute MI rats that underwent early moderate exercise training beginning one day after MI showed no increase in mortality and displayed significant improvements in MI healing and ventricular remodelling, including an improvement in cardiac function, a decrease in infarct size, cardiomyocyte apoptosis, cardiac fibrosis and cardiomyocyte hypertrophy, and an increase in myocardial angiogenesis, left ventricular wall thickness and the number of cardiac telocytes in the border zone. Integrated miRNA‐mRNA profiling analysis performed by the ingenuity pathway analysis system revealed that the inhibition of the TGFB1 regulatory network, activation of leucocytes and migration of leucocytes into the infarct zone comprise the molecular mechanism underlying early moderate exercise‐mediated improvements in cardiac fibrosis and the pathological inflammatory response. The findings of the present study demonstrate that early moderate exercise training beginning one day after MI is safe and leads to significantly enhanced MI healing and ventricular remodelling. Understanding the mechanism behind the positive effects of this early training protocol will help us to further tailor suitable cardiac rehabilitation programmes for humans.     

Understanding cardiac extracellular matrix remodeling to develop biomarkers of myocardial infarction outcomes

Cardiovascular Disease (CVD) is the most common cause of death in industrialized countries, and myocardial infarction (MI) is a major CVD with significant morbidity and mortality. Following MI, the left ventricle (LV) undergoes a wound healing response to ischemia that results in extracellular matrix (ECM) scar formation to replace necrotic myocytes. While ECM accumulation following MI is termed cardiac fibrosis, this is a generic term that does not differentiate between ECM accumulation that occurs in the infarct region to form a scar that is structurally necessary to preserve left ventricle (LV) wall integrity and ECM accumulation that increases LV wall stiffness to exacerbate dilation and stimulate the progression to heart failure. This review focuses on post-MI LV ECM remodeling, targeting the discussion on ECM biomarkers that could be useful for predicting MI outcomes.     

Role of matricellular proteins in cardiac tissue remodeling after myocardial infarction

After onset of myocardial infarction (MI), the left ventricle (LV) undergoes a continuum of molecular, cellular, and extracellular responses that result in LV wall thinning, dilatation, and dysfunction. These dynamic changes in LV shape, size, and function are termed cardiac remodeling. If the cardiac healing after MI does not proceed properly, it could lead to cardiac rupture or maladaptive cardiac remodeling, such as further LV dilatation and dysfunction, and ultimately death. Although the precise molecular mechanisms in this cardiac healing process have not been fully elucidated, this process is strictly coordinated by the interaction of cells with their surrounding extracellular matrix (ECM) proteins. The components of ECM include basic structural proteins such as collagen, elastin and specialized proteins such as fibronectin, proteoglycans and matricellular proteins. Matricellular proteins are a class of non-structural and secreted proteins that probably exert regulatory functions through direct binding to cell surface receptors, other matrix proteins, and soluble extracellular factors such as growth factors and cytokines. This small group of proteins, which includes osteopontin, thrombospondin-1/2, tenascin, periostin, and secreted protein, acidic and rich in cysteine, shows a low level of expression in normal adult tissue, but is markedly upregulated during wound healing and tissue remodeling, including MI. In this review, we focus on the regulatory functions of matricellular proteins during cardiac tissue healing and remodeling after MI.     

Morphological characteristics of the microvasculature in healing myocardial infarcts.

Myocardial infarction (MI) is associated with an angiogenic response, critical for healing and cardiac repair. Using a canine model of myocardial ischemia and reperfusion, we examined the structural characteristics of the evolving microvasculature in healing MI. After 7 days of reperfusion, the infarcted territory was rich in capillaries and contained enlarged, pericyte-poor "mother vessels" and endothelial bridges. During scar maturation arteriolar density in the infarct increased, and a higher percentage of microvessels acquired a pericyte coat (60.4 +/- 6.94% after 28 days of reperfusion vs 30.17 +/- 3.65% after 7 days of reperfusion; p<0.05). The microvascular endothelium in the early stages of healing showed intense CD31/PECAM-1 and CD146/Mel-CAM immunoreactivity but weak staining with the Griffonia simplicifolia lectin I (GS-I). In contrast, after 28 days of reperfusion, most infarct microvessels demonstrated significant lectin binding. Our findings suggest that the infarct microvasculature undergoes a transition from an early phase of intense angiogenic activity to a maturation stage associated with pericyte recruitment and formation of a muscular coat. In addition, in the endothelium of infarct microvessels CD31 and CD146 expression appears to precede that of the specific sugar groups that bind the GS-I lectin. Understanding of the mechanisms underlying the formation and remodeling of the microvasculature after MI may be important in designing therapeutic interventions to optimize cardiac repair.     

左旋组氨酸对匹罗卡品致痫大鼠癫痫发作和海马区神经细胞凋亡的影响

目的:观察左旋组氨酸(L-His)对匹罗卡品(PLO)致痫大鼠皮质脑电图及大鼠海马各区神经细胞凋亡的影响。方法:雄性SD大鼠30只,随机平均分为对照组(匹罗卡品组)、干预组(匹罗卡品+左旋组氨酸组)。各组给予相关干预处理后腹腔注射匹罗卡品建立癫痫模型,行皮质脑电图观察及海马区细胞凋亡染色观察。结果:经皮质脑电图显示,干预组潜伏期延长、痫波频率及大发作次数较对照组明显降低,差异有统计学意义(P<0.05)。凋亡染色显示,对照组的海马各区细胞在各时间点的凋亡数明显高于干预组,差异具有统计学意义(P<0.05)。结论:左旋组氨酸可以延迟匹罗卡品致痫的形成并降低点燃后癫痫发作程度,降低海马各区细胞凋亡数值,提示左旋组氨酸具有抗痫作用。     

Factor XIIIA-V34L and factor XIIIB-H95R gene variants: effects on survival in myocardial infarction patients

It has been demonstrated recently that coagulation factor XIII (FXIII) plays an extraordinary role in myocardial healing after infarction, improving survival in a mouse model. Common FXIII gene variants (i.e. FXIIIA-V34L and FXIIIB-H95R) significantly influence the molecular activity. To evaluate whether there is a relationship between the two FXIII gene variants and survival in patients after myocardial infarction (MI), V34L and H95R were PCR-genotyped in a cohort of 560 MI cases and follow-up was monitored. Cases with ST-segment elevation MI (STEMI) were 416 (74.3%) and 374 of these were treated with primary percutaneous coronary intervention (PCI) (89.9%). The remaining 144 patients showed non-ST-segment elevation MI (NSTEMI) at enrollment. The combined endpoint was the occurrence of death, re-infarction, and heart failure. Kaplan-Meier analysis at one year yielded an overall rate for adverse events of 24.5% with a lower incidence in the L34-carriers (28.8% vs 17.1%; log-rank, P = 0.00025), similar to that of the 416 STEMI (23.8%) being (28.0% and 16.9%; VV34- and L34-carriers respectively; log-rank, P = 0.001). Primary PCI-group had a slight lower incidence (22.9%) of adverse events (26.8% and 17.1%; VV34- and L34-carriers respectively; log-rank, P = 0.009). During hospitalization, 506 patients received PCI (374 primary PCI and 132 elective PCI). Significance was conserved also in the overall PCI-group (28.6% and 17.8%; VV34- and L34-carriers respectively; log-rank, P = 0.001). Similar findings were observed at 30 days follow-up. Cases carrying both FXIII variants had improved survival rate (log-rank, P = 0.019). On the other hand, minor bleeding complications were found increased in L34-carriers (P = 0.0001) whereas major bleeding complications were not. Finally, more direct evidence on the role of FXIII molecule on survival might come from the fact that despite significant FXIII antigen reductions observed in cases after MI, regardless the FXIII genotype considered, L34-carriers kept almost normal FXIII activity (VV34- vs L34-carriers; P < 0.001). We conclude that FXIII L34-allele improves survival after MI in all the groups analyzed, possibly through its higher activity associated with assumable positive effects on myocardial healing and recovered functions. Genetically determined higher FXIII activity might influence post-MI outcome. This paves the way for using FXIII molecules to improve myocardial healing, recovery of functions, and survival after infarction.     

Cardiac fibroblast activation during myocardial infarction wound healing: Fibroblast polarization after MI

Cardiac wound healing after myocardial infarction (MI) evolves from pro-inflammatory to anti-inflammatory to reparative responses, and the cardiac fibroblast is a central player during the entire transition. The fibroblast mirrors changes seen in the left ventricle infarct by undergoing a continuum of polarization phenotypes that follow pro-inflammatory, anti-inflammatory, and pro-scar producing profiles. The development of each phenotype transition is contingent upon the MI environment into which the fibroblast enters. In this mini-review, we summarize our current knowledge regarding cardiac fibroblast activation during MI and highlight key areas where gaps remain.     

Differential cytokine expression in myocytes and non-myocytes after myocardial infarction in rats

The proinflammatory cytokines interleukin (IL)-1 and IL-6 are increased after acute myocardial infarction (MI). Moreover, serum IL-6 level is elevated after MI, but has also been associated with heart failure. In the present study, heart function was monitored in a rat model of chronic MI. Cytokine expression in the infarcted and non-infarcted myocardium as well as in hearts of sham-operated controls was measured by the ribonuclease-protection assay. To identify the cells contributing to the increased cytokine expression, we further analyzed myocytes and non-myocytes isolated in the acute phase as well as during congestive heart failure (CHF) after MI. There was a strong induction in cytokine expression in the myocytes of the infarct area 6 h after MI. In the non-infarcted myocardium, cytokine expression increased only slightly in the non-myocytes after 6 h. This was not different from sham-operated controls and may, therefore, be induced by stress and catecholamines. In CHF, however, cytokine expression level in myocytes was normal. It increased slightly but significantly in the non-myocytes 4 and 8 weeks after MI. In conclusion, we suggest that pro-inflammatory cytokines, produced by the ischemic myocytes may be involved in the initiation of wound healing of the necrotic area, whereas the effect of pro-inflammatory cytokines in CHF, if any, seems not to be crucial.     

A Comparative Characterization of the Mitochondrial Genomes of Paramoeba aparasomata and Neoparamoeba pemaquidensis (Amoebozoa,Paramoebidae)

Marine amebae of the genus Paramoeba (Amoebozoa, Dactylopodida) normally contain a eukaryotic endosymbiont known as Perkinsela‐like organism (PLO). This is one of the characters to distinguish the genera Neoparamoeba and Paramoeba from other Dactylopodida. It is known that the PLO may be lost, but PLO‐free strains of paramoebians were never available for molecular studies. Recently, we have described the first species of the genus Paramoeba which has no parasome—Paramoeba aparasomata. In this study, we present a mitochondrial genome of this species, compare it with that of Neoparamoeba pemaquidensis, and analyze the evolutionary dynamics of gene sequences and gene order rearrangements between these species. The mitochondrial genome of P. aparasomata is 46,254 bp long and contains a set of 31 protein‐coding genes, 19 tRNAs, two rRNA genes, and 7 open reading frames. Our results suggest that these two mitochondrial genomes within the genus Paramoeba have rather similar organization and gene order, base composition, codon usage, the composition and structure of noncoding, and overlapping regions.