VEGF enhances functional improvement of postinfarcted hearts by transplantation of ESC-differentiated cells.

Despite considerable advances in medicine, the incidence of heart failure remains high in patients after myocardial infarction (MI). This study investigated the effects of engrafted early-differentiated cells (EDCs) from mouse embryonic stem cells, with or without transfection of vascular endothelial growth factor (VEGF) cDNA (phVEGF(165)), on cardiac function in postinfarcted mice. EDCs were transfected with green fluorescent protein (GFP) cDNA and transplanted into infarcted myocardium. Compared with the MI mice receiving cell-free medium, cardiac function was significantly improved in the MI mice 6 wk after transplantation of EDCs. Moreover, improvement of heart function was significantly greater in the mice implanted with EDCs overexpressing VEGF (EDCs-VEGF) than with EDCs alone. Frozen sections of infarcted myocardium with EDCs or EDCs-VEGF transplantation showed GFP-positive tissue. The area with positive immunostaining for cardiac troponin I and alpha-myosin heavy chain was larger in injured myocardium with EDCs or EDCs-VEGF transplantation than with medium injection. Transplantation of EDCs or EDCs-VEGF significantly increased the number of blood vessels in the MI area. However, the density of capillaries was significantly higher in the EDCs-VEGF animals than in the EDC mice. Double staining for GFP and connexin-43 was positive in injured myocardium with EDC transplantation. Our data demonstrate that engrafted EDCs or EDCs-VEGF regenerated cardiac tissue and significantly improved cardiac function in postinfarcted hearts. The novel EDCs-VEGF synergistic approach may have an important impact on future cell therapy for patients experiencing MI or heart failure.     

Transplantation of embryonic stem cells improves cardiac function in postinfarcted rats.

Massive loss of cardiac myocytes after myocardial infarction (MI) is a common cause of heart failure. The present study was designed to investigate the improvement of cardiac function in MI rats after embryonic stem (ES) cell transplantation. MI in rats was induced by ligation of the left anterior descending coronary artery. Cultured ES cells used for cell transplantation were transfected with the marker green fluorescent protein (GFP). Animals in the treated group received intramyocardial injection of ES cells in injured myocardium. Compared with the MI control group injected with an equivalent volume of the cell-free medium, cardiac function in ES cell-implanted MI animals was significantly improved 6 wk after cell transplantation. The characteristic phenotype of engrafted ES cells was identified in implanted myocardium by strong positive staining to sarcomeric alpha-actin, cardiac alpha-myosin heavy chain, and troponin I. GFP-positive cells in myocardium sectioned from MI hearts confirmed the survival and differentiation of engrafted cells. In addition, single cells isolated from cell-transplanted MI hearts showed rod-shaped GFP-positive myocytes with typical striations. The present data demonstrate that ES cell transplantation is a feasible and novel approach to improve ventricular function in infarcted failing hearts.     

Cardiac telocytes were decreased during myocardial infarction and their therapeutic effects for ischaemic heart in rat

Recently, cardiac telocytes were found in the myocardium. However, the functional role of cardiac telocytes and possible changes in the cardiac telocyte population during myocardial infarction in the myocardium are not known. In this study, the role of the recently identified cardiac telocytes in myocardial infarction (MI) was investigated. Cardiac telocytes were distributed longitudinally and within the cross network of the myocardium, which was impaired during MI. Cardiac telocytes in the infarction zone were undetectable from approximately 4 days to 4 weeks after an experimental coronary occlusion was used to induce MI. Although cardiac telocytes in the non‐ischaemic area of the ischaemic heart experienced cell death, the cell density increased approximately 2 weeks after experimental coronary occlusion. The cell density was then maintained at a level similar to that observed 1–4 days after left anterior descending coronary artery (LAD)‐ligation, but was still lower than normal after 2 weeks. We also found that simultaneous transplantation of cardiac telocytes in the infarcted and border zones of the heart decreased the infarction size and improved myocardial function. These data indicate that cardiac telocytes, their secreted factors and microvesicles, and the microenvironment may be structurally and functionally important for maintenance of the physiological integrity of the myocardium. Rebuilding the cardiac telocyte network in the infarcted zone following MI may be beneficial for functional regeneration of the infarcted myocardium.     

Cardiomyogenic differentiation-independent improvement of cardiac function by human cardiomyocyte progenitor cell injection in ischaemic mouse hearts

We previously showed that human cardiomyocyte progenitor cells (hCMPCs) injected after myocardial infarction (MI) had differentiated into cardiomyocytes in vivo 3 months after MI. Here, we investigated the short-term (2 weeks) effects of hCMPCs on the infarcted mouse myocardium. MI was induced in immunocompromised (NOD/scid) mice, immediately followed by intramyocardial injection of hCMPCs labelled with enhanced green fluorescent protein (hCMPC group) or vehicle only (control group). Sham-operated mice served as reference. Cardiac performance was measured 2 and 14 days after MI by magnetic resonance imaging at 9.4 T. Left ventricular (LV) pressure-volume measurements were performed at day 15 followed by extensive immunohistological analysis. Animals injected with hCMPCs demonstrated a higher LV ejection fraction, lower LV end-systolic volume and smaller relaxation time constant than control animals 14 days after MI. hCMPCs engrafted in the infarcted myocardium, did not differentiate into cardiomyocytes, but increased vascular density and proliferation rate in the infarcted and border zone area of the hCMPC group. Injected hCMPCs engraft into murine infarcted myocardium where they improve LV systolic function and attenuate the ventricular remodelling process 2 weeks after MI. Since no cardiac differentiation of hCMPCs was evident after 2 weeks, the observed beneficial effects were most likely mediated by paracrine factors, targeting amongst others vascular homeostasis. These results demonstrate that hCMPCs can be applied to repair infarcted myocardium without the need to undergo differentiation into cardiomyocytes.     

The effect of hydrogel injection on cardiac function and myocardial mechanics in a computational post-infarction model

An emerging therapy to limit adverse heart remodelling following myocardial infarction (MI) is the injection of polymers into the infarcted left ventricle (LV). In the few numerical studies carried out in this field, the definition and distribution of the hydrogel in the infarcted myocardium were simplified. In this computational study, a more realistic biomaterial distribution was simulated after which the effect on cardiac function and mechanics was studied. A validated finite element heart model was used in which an antero-apical infarct was defined. Four infarct models were created representing different temporal phases in the progression of a MI. Hydrogel layers were simulated in the infarcted myocardium in each model. Biomechanical and functional improvement of the LV was found after hydrogel inclusion in the ischaemic models representing the early phases of MI. In contrast, only functional but no mechanical restitution was shown in the scar model due to hydrogel presence.     

培养于生物降解支架膜内的胚胎干细胞可修复小鼠的梗塞心肌

我们以往的研究表明,直接在心肌梗塞（myocardial infarction,MI）动物的心脏缺血区注射胚胎干细胞（embryonic stemceils,ESCs）可以提高其心肌功能,干细胞组织工程学可以使组织再生、修复。本研究旨在观察将ESCs接种到生物降解膜内并移植到梗塞部位的效果。通过结扎小鼠左冠状动脉制作MI模型,将培养3d的带有小鼠ESCs的聚羟基乙酸膜（polyglycolicacid,PGA）移植到心肌缺血及边缘区表面。实验小鼠分成4组：假手术组、MI组、MI＋PGA组、MI＋ESC组,移植操作8周后检测血流动力学和心肌功能。MI组的血压和左心室功能显著降低。与MI组和MI＋PGA组相比,MI＋ESC组的血压和心室功能显著改善,存活率也显著增高,在梗塞区检测到GFP阳性组织,表明ESCs存活,并可能有心肌再生。以上结果表明,移植生物降解膜内的ESCs可修复小鼠梗塞区心肌细胞并提高心脏功能。将ESCs和生物降解材料联合运用可能为修复受损心脏提供一个新的治疗方法。     

The effect of immune cell-derived exosomes in the cardiac tissue repair after myocardial infarction: Molecular mechanisms and pre-clinical evidence

After a myocardial infarction (MI), the inflammatory responses are induced and assist to repair ischaemic injury and restore tissue integrity, but excessive inflammatory processes promote abnormal cardiac remodelling and progress towards heart failure. Thus, a timely resolution of inflammation and a firmly regulated balance between regulatory and inflammatory mechanisms can be helpful. Molecular- and cellular-based approaches modulating immune response post-MI have emerged as a promising therapeutic strategy. Exosomes are essential mediators of cell-to-cell communications, which are effective in modulating immune responses and immune cells following MI, improving the repair process of infarcted myocardium and maintaining ventricular function via the crosstalk among immune cells or between immune cells and myocardial cells. The present review aimed to seek the role of immune cell-secreted exosomes in infarcted myocardium post-MI, together with mechanisms behind their repairing impact on the damaged myocardium. The exosomes we focus on are secreted by classic immune cells including macrophages, dendritic cells, regulatory T cells and CD4⁺ T cells; however, further research is demanded to determine the role of exosomes secreted by other immune cells, such as B cells, neutrophils and mast cells, in infarcted myocardium after MI. This knowledge can assist in the development of future therapeutic strategies, which may benefit MI patients.     

Resveratrol ameliorates myocardial damage by inducing vascular endothelial growth factor-angiogenesis and tyrosine kinase receptor Flk-1

In our study, resveratrol (polyphenol) has been identified as a very important stimulus/agent for the induction of new vessel growth. Occlusion of a main coronary depletes the blood supply to the myocardium and subsequently reduces cardiac function, which ultimately leads to heart failure. Progressive, chronic coronary artery occlusion has been shown to induce development of collateral arteries to re-establish and maintain blood flow to the myocardium at risk via the growth of new capillary vessels or angiogenesis. Studies from our laboratory, as well as from others, have already confirmed the protective role of collaterals against myocardial ischemia and cell death. We have successfully demonstrated in rat myocardial infarction (MI) model an effect of resveratrol on significant upregulation of the protein expression profiles of vascular endothelial growth factor (VEGF) and its tyrosine kinase receptor Flk-1, 3 wk after MI. Pretreatment with resveratrol also increased nitric-oxide synthase (inducible NOS and endothelial NOS) along with increased antiapoptotic and proangiogenic factors nuclear factor (NF)-kappaB and specificity protein (SP)-1. We also were able to demonstrate increased capillary density as well as improved left ventricular function by pharmacological preconditioning with resveratrol 3 wk after MI.     

Bone marrow stromal cells improve cardiac performance in healed infarcted rat hearts

Postinfarct congestive heart failure is one of the leading causes of morbidity and mortality in developed and developing countries. The main purpose of this study was to investigate whether transplantation of bone marrow stromal cells (BMSC) directly into the myocardium could improve the performance of healed infarcted rat hearts. Cell culture medium with or without BMSC was injected into borders of cardiac scar tissue 4 wk after experimental infarction. Cardiac performance was evaluated 2 wk after cellular (n = 10) or medium (n = 10) injection by electro- and echocardiography. Histological study was performed 3 wk after treatment. Electrocardiography of BMSC-treated infarcted rats showed electrical and mechanical parameters more similar to those in control than in medium-treated animals: a normal frontal QRS axis in 6 of 10 BMSC-treated and all control rats and a rightward deviation of the QRS axis in all 10 medium-treated animals. BMSC treatment, assessed by echocardiography, improved fractional shortening (39.00 +/- 4.03%) compared with medium-treated hearts (18.20 +/- 0.74%) and prevented additional changes in cardiac geometry. Immunofluorescence microscopy revealed colocalization of 4',6-diamidino-2-phenylindole-labeled nuclei of transplanted cells with cytoskeletal markers for cardiomyocytes and smooth muscle cells, indicating regeneration of damaged myocardium and angiogenesis. These data provide strong evidence that BMSC implantation can improve cardiac performance in healed infarctions and open new promising therapeutic opportunities for patients with postinfarction heart failure.     

MicroRNA-1 transfected embryonic stem cells enhance cardiac myocyte differentiation and inhibit apoptosis by modulating the PTEN/Akt pathway in the infarcted heart

microRNAs (miRs) have emerged as critical modulators of various physiological processes including stem cell differentiation. Indeed, miR-1 has been reported to play an integral role in the regulation of cardiac muscle progenitor cell differentiation. However, whether overexpression of miR-1 in embryonic stem (ES) cells (miR-1-ES cells) will enhance cardiac myocyte differentiation following transplantation into the infarcted myocardium is unknown. In the present study, myocardial infarction (MI) was produced in C57BL/6 mice by left anterior descending artery ligation. miR-1-ES cells, ES cells, or culture medium (control) was transplanted into the border zone of the infarcted heart, and 2 wk post-MI, cardiac myocyte differentiation, adverse ventricular remodeling, and cardiac function were assessed. We provide evidence demonstrating enhanced cardiac myocyte commitment of transplanted miR-1-ES cells in the mouse infarcted heart as compared with ES cells. Assessment of apoptosis revealed that overexpression of miR-1 in transplanted ES cells protected host myocardium from MI-induced apoptosis through activation of p-AKT and inhibition of caspase-3, phosphatase and tensin homolog, and superoxide production. A significant reduction in interstitial and vascular fibrosis was quantified in miR-1-ES cell and ES cell transplanted groups compared with control MI. However, no statistical significance between miR-1-ES cell and ES cell groups was observed. Finally, mice receiving miR-1-ES cell transplantation post-MI had significantly improved heart function compared with respective controls (P < 0.05). Our data suggest miR-1 drives cardiac myocyte differentiation from transplanted ES cells and inhibits apoptosis post-MI, ultimately giving rise to enhanced cardiac repair, regeneration, and function.     

Local activation of cardiac stem cells for post‐myocardial infarction cardiac repair

The prognosis of patients with myocardial infarction (MI) and resultant chronic heart failure remains extremely poor despite continuous advancements in optimal medical therapy and interventional procedures. Animal experiments and clinical trials using adult stem cell therapy following MI have shown a global improvement of myocardial function. The emergence of stem cell transplantation approaches has recently represented promising alternatives to stimulate myocardial regeneration. Regarding their tissue‐specific properties, cardiac stem cells (CSCs) residing within the heart have advantages over other stem cell types to be the best cell source for cell transplantation. However, time‐consuming and costly procedures to expanse cells prior to cell transplantation and the reliability of cell culture and expansion may both be major obstacles in the clinical application of CSC‐based transplantation therapy after MI. The recognition that the adult heart possesses endogenous CSCs that can regenerate cardiomyocytes and vascular cells has raised the unique therapeutic strategy to reconstitute dead myocardium via activating these cells post‐MI. Several strategies, such as growth factors, mircoRNAs and drugs, may be implemented to potentiate endogenous CSCs to repair infarcted heart without cell transplantation. Most molecular and cellular mechanism involved in the process of CSC‐based endogenous regeneration after MI is far from understanding. This article reviews current knowledge opening up the possibilities of cardiac repair through CSCs activation in situ in the setting of MI.     

Spaciotemporal alteration of 8-hydroxy-2'-deoxyguanosine levels in cardiomyocytes after myocardial infarction in rats

Temporary or persistent heart failure is one of the major complications after myocardial infarction (MI). In order to elucidate the pathogenesis of MI, we studied the spaciotemporal alteration of 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in cardiomyocytes in a rat model of ligation of the left anterior descending branch of the coronary artery. The lethality in this model was 18%. Hearts were dissected at 0, 3, 6, 12, 24, 48 h, and 1, 2, 4, 6 weeks after the operation. The cardiac level of 8-OHdG was evaluated biochemically as well as by immunohistochemistry with monoclonal antibody N45.1. Three to 6 h after ligation, the 8-OHdG levels were increased in the cardiomyocytes of MI (six-fold) and peri-MI (four-fold) areas. After 24 h, the myocardium in the MI area was necrotized, and thereafter the 8-OHdG level decreased. 8-OHdG levels in the myocardium of peri-MI areas returned once to a normal level, but were significantly increased at 2-4 weeks along with the appearance of apoptotic cardiomyocytes in this area. The heart after MI has been generally considered as clinically stable after four weeks. However, cardiomyocytes near the infarcted area were oxidatively stressed even after four weeks when the affected lesion was extensive. The present data support the use of supplementary antioxidant therapies to save functional myocardium after MI. (213 words)     

Increased cross-bridge cycling rate in stunned myocardium

It is well known that the implantation of bone marrow mononuclear cells (BM-MNCs) into ischemic hearts can induce angiogenesis and improve cardiac function after myocardial infarction, but the precise mechanisms of these actions are unclear. We hypothesize that the cytokines produced by BM-MNCs play a key role in this cell-based therapy. BM-MNCs from rats were cultured under normoxic or hypoxic (1% O2) conditions for 24 h, and then supernatants were collected for study. ELISA and Western blotting analysis showed that various cytokines, including VEGF, IL-1 beta, PDGF, and IGF-1, were produced from BM-MNCs, some of which were enhanced significantly under hypoxia stimulation. When compared with a control blank medium, the supernatants of BM-MNCs cultured under normoxic or hypoxic conditions inhibited apoptosis significantly and preserved the contractile capacity of isolated adult rat cardiomyocytes in vitro (P < 0.05). Using a rat model of acute myocardial infarction, we injected the supernatants of BM-MNCs or control medium intramyocardially on day 0 and then intraperitoneally on days 2, 4, and 6 after infarction. When compared with the control medium, the supernatants of BM-MNCs cultured under both normoxic or hypoxic conditions increased the microvessel density and decreased the fibrotic area in the infarcted myocardium significantly, contributing to remarkable improvement in cardiac function. Various cytokines were produced by BM-MNCs, and these cytokines contributed to functional improvement of the infarcted heart by directly preserving the contractile capacity of the myocardium, inhibiting apoptosis of cardiomyocytes, and inducing therapeutic angiogenesis of the infarcted heart.     

Age-related differences in postinfarct left ventricular rupture and remodeling

Cardiac rupture is more prevalent in elderly patients with first onset of acute myocardial infarct (MI), but the mechanism remains unexplored. We investigated the differences in the incidence of cardiac rupture and early left ventricular (LV) remodeling following coronary artery ligation between old (12-mo) and young (3-mo) C57Bl/6 male mice and explored responsible mechanisms. The incidence of rupture within 1 wk after MI was significantly higher in old than in young mice (40.7 vs. 18.3%, P = 0.013) despite a similar infarct size in both age groups. Old mice dying of rupture had more severe infarct expansion than young counterparts. Echocardiography and catheterization at day 7 revealed more profound LV chamber dilatation and dysfunction as well as higher blood pressures in aged mice. At day 3 after MI immediately before the peak of rupture occurrence, we observed significantly higher content of type I and III collagen, a greater density of macrophage and neutrophil, and markedly enhanced mRNA expression of inflammatory cytokines in the infarcted myocardium in old than in young mice. Furthermore, a more dramatic increment of matrix metalloproteinase (MMP)-9 activity was found in old than in young infarcted hearts, in keeping with enhanced inflammatory response. Collectively, these results revealed that old mice had a higher risk of post-MI cardiac rupture despite a higher level of collagen content and cross-linking. Enhanced inflammatory response and subsequent increase in MMP-9 activity together with higher blood pressure are important factors responsible for the higher risk of cardiac rupture and more severe LV remodeling in the aged heart following acute MI.     

Repeated sauna therapy attenuates ventricular remodeling after myocardial infarction in rats by increasing coronary vascularity of noninfarcted myocardium

Repeated sauna therapy (ST) increases endothelial nitric oxide synthase (eNOS) activity and improves cardiac function in heart failure as well as peripheral blood flow in ischemic limbs. The present study investigates whether ST can increase coronary vascularity and thus attenuate cardiac remodeling after myocardial infarction (MI). We induced MI by ligating the left coronary artery of Wistar rats. The rats were placed in a far-infrared dry sauna at 41°C for 15 min and then at 34°C for 20 min once daily for 4 wk. Cardiac hemodynamic, histopathological, and gene analyses were performed. Despite the similar sizes of MI between the ST and non-ST groups (51.4 ± 0.3 vs. 51.1 ± 0.2%), ST reduced left ventricular (LV) end-diastolic (9.7 ± 0.4 vs. 10.7 ± 0.5 mm, P < 0.01) and end-systolic (8.6 ± 0.5 vs. 9.6 ± 0.6 mm, P < 0.01) dimensions and attenuated MI-induced increases in LV end-diastolic pressure. Cross-sectional areas of cardiomyocytes were smaller in ST rats and associated with a significant reduction in myocardial atrial natriuretic peptide mRNA levels. Vascular density was reduced in the noninfarcted myocardium of non-ST rats, and the density of cells positive for CD31 and for α-smooth muscle actin was decreased. These decreases were attenuated in ST rats compared with non-ST rats and associated with increases in myocardial eNOS and vascular endothelial growth factor mRNA levels. In conclusion, ST attenuates cardiac remodeling after MI, at least in part, through improving coronary vascularity in the noninfarcted myocardium. Repeated ST might serve as a novel noninvasive therapy for patients with MI.     

Matrix metalloproteinase expression in cardiac myocytes following myocardial infarction in the rabbit

Myocardial infarction (MI), leads to cardiac remodeling, thinning of the ventricle wall, ventricular dilation, and heart failure, and is a leading cause of death. Interactions between the contractile elements of the cardiac myocytes and the extracellular matrix (ECM) help maintain myocyte alignment required for the structural and functional integrity of the heart. Following MI, reorganization of the ECM and the myocytes occurs, contributing to loss of heart function. In certain pathological circumstances, the ECM is modulated such that the structure of the tissue becomes damaged. The matrix metalloproteinases (MMPs) are a family of enzymes that degrade molecules of the ECM. The present experiments were performed to define the time-course, isozyme subtypes, and cellular source of increased MMP expression that occurs following MI in an experimental rabbit model. Heart tissue samples from infarcted and sham animals were analyzed over a time-course of 1-14 days. By zymography, it was demonstrated that, unlike the sham controls, MMP-9 expression was induced within 24 hours following MI. MMP-3 expression, also absent in sham controls, was induced 2 days after MI. MMP-2 expression was detected in both the sham and infarcted samples and was modestly up-regulated following MI. Tissue inhibitor of metalloproteinase-1 (TIMP-1) expression was evaluated and shown to be down-regulated following MI, inverse of MMP-9 and MMP-3 expression. Further, MMP-9 and MMP-3 expression was detected by immunohistochemistry in myocytes within the infarct. Additional studies were conducted in which cultured rat cardiac myocytes were exposed to a hypoxic environment (2% O2) for 24 hours and the media analyzed for MMP expression. MMP-9 and MMP-3 were induced following exposure to hypoxia. It is speculated that the net increase in proteolytic activity by myocytes is a contributing factor leading to myocyte misalignment and slippage. Additional studies with a MMP inhibitor would elucidate this hypothesis.     

高同型半胱氨酸血症对心肌梗死后心肌组织中SCF表达的影响

目的通过观察高同型半胱氨酸血症(HHCY)对心肌梗死后大鼠心肌组织中干细胞因子(SCF)表达的影响,探讨HHCY在心肌梗死(MI)后干细胞归巢并修复梗死心肌过程中的作用。方法饮食诱导大鼠HHCY的动物模型;随后建立大鼠MI模型;利用免疫组织化学染色及RT-PCR技术分别检测HHCY对MI后大鼠MI区域及MI灶周围心肌组织内SCF表达的影响。结果MI后大鼠MI灶周围心肌组织中SCF的表达增强(与假结扎组相比);高蛋氨酸饮食组大鼠MI灶周围心肌组织内SCF的表达显著减少(与对照组相比);补充叶酸降低了大鼠血清中同型半胱氨酸浓度,增强了MI灶周围心肌组织内SCF的表达。结论MI上调了MI灶周围心肌组织中SCF的表达,有利于干细胞的归巢并修复梗死的心肌;HHCY抑制了MI灶周围心肌组织内SCF的表达,从而减少由SCF所诱导的干细胞归巢到MI区域,进而抑制了梗死后心肌的修复。     

Significance of changes in TNF-alpha and IL-10 levels in the progression of heart failure subsequent to myocardial infarction

We tested whether a decrease in the ratio of interleukin-10 (IL-10) to tumor necrosis factor-alpha (TNF-alpha) correlates with the decrease in cardiac function in heart failure. It has been suggested that TNF-alpha plays a role in the progression of heart failure, and the effect of TNF-alpha in many tissues is modulated by IL-10. Any relation of these two cytokines to heart failure has never been examined. Cardiac function was assessed by echocardiographic and hemodynamic techniques in coronary artery-ligated rats at 1, 4, 8, and 16 wk after myocardial infarction (MI). Membrane-bound and soluble fractions of TNF-alpha and IL-10 proteins, the ratio of TNF-alpha to IL-10, and TNF-alpha and IL-10 mRNA levels were analyzed. Losartan was used to modify cardiac function in rats 4 wk after MI to further validate the relation between the IL-10-to-TNF-alpha ratio and cardiac function. Cardiac function deteriorated with time in all coronary artery-ligated groups, with severe failure at 16 wk after MI. Membrane-bound and soluble TNF-alpha protein fractions were increased 1 and 4 wk after MI, whereas TNF-alpha mRNA was increased 4 and 8 wk after MI. Membrane-bound IL-10 protein and mRNA levels were decreased 4, 8, and 16 wk after MI. The decrease in the IL-10-to-TNF-alpha protein ratio in all coronary artery-ligated groups correlated with the depressed cardiac function. Losartan improved cardiac function, membrane-bound and soluble TNF-alpha and IL-10 protein levels, the ratio of IL-10 to TNF-alpha, and IL-10 mRNA. This study suggests that a decrease in IL-10 and IL-10-to-TNF-alpha ratio correlates with depressed cardiac function.     

Cardiac adrenomedullin gene expression and peptide accumulation after acute myocardial infarction in rats

Plasma adrenomedullin (AM) has been shown to increase in the early phase of acute myocardial infarction (MI). However, little information is available regarding cardiac AM synthesis after MI. Accordingly, we examined the time course of ventricular AM production and potential stimulation of AM in the infarcted and noninfarcted regions in MI rats produced by coronary artery ligation. Compared with sham-operated rats, the ventricular AM peptide level 6 h after MI increased 1.5-fold in the infarcted region and 1.7-fold in the noninfarcted region in association with increased left ventricular end-diastolic pressure (EDP). Northern blot analysis also showed marked induction of AM gene expression in the infarcted region (11-fold) and the noninfarcted region (6-fold) 6 h after MI. The AM peptide level in the infarcted region reached its peak (2. 6-fold) 1 wk postinfarction and thereafter decreased to normal. In the noninfarcted region, however, the AM level remained elevated for at least 4 wk. Immunohistochemical studies demonstrated that intense immunostaining for AM was limited to myocytes in both the infarcted and noninfarcted regions. Interestingly, the AM level in the noninfarcted region correlated positively with infarct size (r = 0. 40, P < 0.01) and EDP (r = 0.52, P < 0.001). An oral angiotensin-converting enzyme inhibitor suppressed the overproduction of AM 1 wk postinfarction in association with decreases in EDP and mean arterial pressure. In summary, cardiac AM synthesis was rapidly induced in both the infarcted and noninfarcted regions after MI. The subsequent ventricular AM in the two regions demonstrated different time-concentration curves during 4 wk after MI. AM may be synthesized predominantly by cardiac myocytes, but not by fibroblasts, at least in part, in association with increased ventricular load after MI.     

The scar neovasculature after myocardial infarction in rats

A series of novel techniques, adapted from the field of tumor biology, were developed to quantify vascular structure and function and to explore the role of ANG II receptor AT1 in cardiac remodeling after myocardial infarction (MI). We examined the scar neovasculature at 1-4 wk post-MI in Sprague-Dawley rats with a view toward its ability to deliver and exchange oxygen. CD31 and DiOC7(3) staining was used to visualize anatomical vessels vs. those perfused. EF5/Cy3 immunohistochemical staining was used to quantify tissue hypoxia. We compared untreated controls with rats treated with losartan, an AT1 receptor antagonist. Our findings indicated that, at the infarct site, there was not only a 42-75% (1-4 wk post-MI) decrease in the number of anatomical vessels compared with controls but also a decrease in the fraction of perfused vessels from 70% in normal coronary vasculature to 48% at the infarct site. These changes were accompanied by progressive increases in diffusion distance and tissue hypoxia (100% increase in EF5/Cy3 staining at 4 wk post-MI). Losartan-treated rats exhibited a significantly less marked reduction in vascular perfusion and a significantly lesser extent of tissue hypoxia. Over the course of 4 wk post-MI, there is a reduction in coronary vasculature at the infarct site, the extent of which is attenuated by losartan. These findings implicate AT1 receptor upregulation, and perhaps angiotensin-related peptides, as being antiangiogenic.