This year we celebrate 50 years of the first interhuman heart transplantation. So, Ibelieve that it is interesting to review the most important steps of this gloriousjourney.th century, seen as a speculation for the future inthe middle of the same century, is now widely accepted by medical and laycommunities as a valuable therapeutic procedure.Heart transplantation first performed in the course of experiments of other nature inthe beginning of 20This history can be divided into an experimental and a clinical heart transplantationperiods, with the first one in heterotopic non-auxiliary, heterotopic auxiliary andorthotopic transplantations.. Inthe course of experiments to develop the technique of vascular anastomoses, theyhave done limb reimplantation, thyroid gland as well kidney and hearttransplantation. The description of their heterotopic heart transplantation is asfollows: "The heart of a small dog was extirpated and transplanted into the neck ofa larger one by anastomosing the cut ends of the jugular vein and the carotid arteryto the aorta, the pulmonary artery, one of the venae cavae and a pulmonary vein. Thecirculation was reestablished through the heart, about an hour and 15 minutes afterthe cessation of the beat; 20 minutes after the reestablishment of the circulation,the blood was actively circulating through the coronary system. A small openingbeing made through the wall of a small branch of the coronary vein, an abundant darkhemorrhage was produced. Then strong fibrillar contractions were seen. Afterwardcontractions appeared and about an hour after the operation, effective contractionsof the ventricles began. The transplanted heart beat at the rate of 88 per minute,while the rate of the normal heart was 100 per minute. A little later tracing wastaken. Owing to the fact that the operation was made without aseptic technique,coagulation occurred in the cavities of the heart after about two hours, and theexperiment was interrupted".The first heart transplant was performed by Alexis Carrel and Charles Guthrie in 1905at the University of Chicago. The problem with this model is that arterialinflow is in the atrium and aortic outflow is under venous pressure, in other words,totally not physiological. Despite these problems, the experiment demonstrated thatheart transplantation was possible, that the heart separated of its blood supplycould be sutured to the circulation of other animal and recover normal function.Although the exact details of experiments were not known the most probablearrangement of anastomosis are shown in  in 1933. Theobjective of the experiments was to obtain a totally denervated heart. In one oftheir models, the heart transplanted in the neck had pulmonary artery anastomosed toproximal jugular vein and aorta to distal carotid artery with a more adequatecoronary perfusion. They obtained a longest survival of eight days and mean survivalof four days. They made observations of recipient donor heart different response ofrate and furthermore studied the histology of rejection, and as Carrel stating thatsurgical technique is not the cause of failure of the grafted heart.The next reference to transplantation of heart in dogs was made by Mann etal. working at the Chicago Medical School, using a modifiedMann technique, studied complicated methods of cardiac preservation with poorsurvival. In 1953, Marcus et al.again modified the Mann preparation, so that the donor left ventricle would act as apump. The proximal end of the divided recipient common carotid artery wasanastomosed to the donor left atrium and the recipient distal common carotid to thedonor innominate artery, so that the donor left ventricle supplied blood to its owncoronary arteries and to the recipient cerebral circulation. Maximum survival withthis technique was 48 hours. Longer survival were obtained later on withheterotopic, non-functioning heart transplant.More than two decades later, Marcus et al.. In 1962, Demikhov published an extensive work he developed in Russia. Amonghis achievements, there are reports of heterotopic heart transplantation in theinguinal region in 1940, heterotopic intrathoracic heart transplants in 1946.Twenty-four variants were described and he mentions a 32 days survival of thegraft.The heterotopic abdominal and neck technique is a valuable tool for studies ofimmunosuppressors or other studies on the graftAnother technical amazing achievement by Demikhov was the transplantation of anadditional head in a dog and the intrathoracic heart - lung transplantation beforethe availability of cardiopulmonary bypass . Differe performed22 canine intrathoracic auxiliary heart transplants between 1951 and 1955. Aftergrafting, he excluded the recipient heart from circulation by ligating the greatvessels and mitral annulus. One such animal awoke from anesthesia stood up, drankbut died 15 hours later.In this field, DemikhovThis experiment was the first one to demonstrate that transplanted heart could assumetotal circulation and the death of the dog was attributed to superior vena cavathrombosis.In 1989, at the International Society for Heart-Lung Transplantation Annual Meeting,in Munich, the Organizing Committee could find Dr. Demikhov in the country of Russiaand payed a tribute to him at Gala Dinner. Apparently, he was bright and could notunderstand or speak a word in English, nevertheless, we could shake his hands., studied also intrathoracic auxiliary transplanted caninehearts with immunosuppression with a 4-hour survival with the recipient heartfibrillated. Another group, Johansson, Soderlund and William-Olsson working at the Karolinska Institutein Sweden performed intrathoracic auxiliary heart transplantation for assistance ofthe recipient left heart maintaining cardiac output for one hour.Reemtsma group, in 1964 and later in 1966 faced all theseproblems in their experiments. Their option was to use topical hypothermia for bothdonor and recipient and to avoid multiple venous anastomoses they performedheart-lung transplants. Three dogs survived six hours.By the beginning of 1950, there was evidence of the feasibility of hearttransplantation and recovery of adequate function of grafted heart. The step forwardwas the experimental orthotopic heart transplantation. In this phase, the majorconcern was the maintenance of the recipient animal, the preservation of the graftand the development of a surgical technique to minimize ischemia. In 1953, Neptuneet al.reported preservation of the heart in dogs by flushing the organ with cold potassiumcitrate and successfully transplant heterotopically. So, these authors in a certainway, opened the pathway for long distance procurement.In 1957, Webb and Howard at theUniversity of Maryland performed the first orthotopic heart transplantation in dog.They created the technique of left atrial anastomosis with a cuff of the orifices ofthe four pulmonary veins the conventional technique of cardiac transplantation thatwas used routinely for decades. The anastomosis of great vessels was performed bysutures and both vena cava were connected with methacrylate tubes. Recipient wasmaintained with cardiopulmonary bypass, but surprisingly the donor heart wasarrested with potassium citrate in normothermia. The longest survival was 20minutes.In 1959, Goldberg et al. performedorthotopic heart transplantation in dogs with hypothermic preservation butindividual pulmonary veins connection. Ten dogs survived from 30 to 450 minutes.Subsequently, in the same year, Cass and Brock working at the Guy's Hospital in London, performedautotransplantation and homologous transplantation using the left atrial cufftechnique described by Goldberg and introducing the right atrial anastomosis insteadof caval anastomosis. In other words, they established the standard technique oforthotopic heart transplantation used clinically until introduction of bicavaltechnique. In this scenario, late 1950, Dr. Shumway group, at Stanford, started toperform orthotopic heart transplantation in dogs with cardiopulmonary bypass,preservation of the donor heart by immersion for 5 minutes in a in 4°C saline. Theystandardized surgical technique incorporating the previous concepts of left andright atrial cuffs. Lower and Shumway published their experience in 1960 reporting that five ofeight animals survived for 6-21 days without immunosuppression. In sequence, Loweret al. made an importantcontribution in 1965, showing a decrease of electrocardiography voltage in rejectionand the reversal of process with a combination of azathioprine andmethylprednisolone achieving a survival of 250 days  to propose to Prof. Zerbini the performance of human transplant in1964. It was considered a premature indication . After Bhe heart and 9. Tth, 1968 Dr. Barnard organized a meeting called "Experiencewith human heart transplantation", in Cape Town, South Africa. Sixteen guestspeakers were invited; three from South Africa; four from USA; two from England; onefrom India; one from Argentina; 1 from Chile and Dr. Zerbini. Brazil was includedamong the pioneer countries ; Stanford-Shumway group; RichardLower-Richmond-Virginia (after leaving Stanford) and Christian Cabrol-LaPitié Salpetrieri in Paris. Nevertheless, in this period, we had severalimportant achievements mostly at Stanford. The introduction of immunologicalmonitoring, important in azathioprine/steroids era and the development ofendomyocardial biopsy by Phillip Caves, both at Stanford. The bioptome was developedbased on the Sakakibara clamp and made by Schultz. Schultz was a little old niceman. In 1977, when I spent three months and a half at Stanford following thetransplantation program, I went to Schultz home and bought a bioptome manufacturedin his garage. With this bioptome, I performed almost 400 biopsies in severalcardiomyopathies and transplanted patients.On April, 4In 1978, cyclosporine was introduced in kidney transplantation and, in 1980, incardiac transplantation at Stanford. In 1980, the modern era of hearttransplantation started with rapid increase of annual number of transplant centersand number of transplanted patients. It is amazing to look back and see suchprogress in an 80-year period.


Previous data has shown that patients in the indeterminate form of Chagasdisease may present myocardial fibrosis as shown on through magneticresonance imaging (MRI). However, there is little information availableregarding the degree of severity of myocardial fibrosis in theseindividuals. This variable has the potential to predict the evolution ofChagas’ disease into its cardiac form.To describe the frequency and extent of myocardial fibrosis evaluated usingan MRI in patients in the indeterminate form, and to compare it with otherforms of the disease.Patients were admitted one after another. Their clinical history wascollected and they were submitted to laboratory exams and an MRI.Sixty-one patients with Chagas’ disease, with an average age of 58 ± 9years old, 17 patients in the indeterminate form, 16 in the cardiac formwithout left ventricular (LV) dysfunction and 28 in the cardiac form with LVdysfunction were studied. P <0.05 was considered to be statisticallysignificant. Late enhancement was detected in 37 patients (64%). Myocardialfibrosis was identified in 6 individuals in indeterminate form  in a proportion similar to that observed in cardiac form without LVdysfunction (44%); p = 1.0. Among the individuals with fibrosis, the totalarea of the affected myocardium was 4.1% (IIQ: 2.1 - 10.7) in theindeterminate form versus 2.3% (IIQ: 1-5) in the cardiac form without LV (p= 0.18). The left ventricular fraction ejection in subjects in theindeterminate form was similar to that of the individuals in the cardiacform without ventricular dysfunction (p = 0.09).The presence of fibrosis in the indeterminate form of Chagas’ disease has afrequency and extension similar to that of in the cardiac form withoutdysfunction, suggesting that the former is part of a subclinical diseasespectrum, rather than lacking cardiac involvement. However, the disease does not manifest itself in thesepatients and one-third of them progress to some type of cardiac and/or digestivemanifestation, and thus are reclassified as chronic.7Chagas’ disease is a potentially debilitating endemic problem in Latin Americancountries and has led to an estimated loss of 750,000 years of productivelife.Identifying the indeterminate patients that are prone to progress to the chronic formserve as a basis for the research of preventive strategies and a betterunderstanding of the pathological processes that lead to this evolution. However,there are no predictive markers or models capable of estimating the risk of thischange.10 Theetiopathogenic process that promotes fibrosis involves a multifactorial relationshipbetween the aspects related to the etiologic agent and those related to thehost14As such, several researchers consider myocardial fibrosis to be a possible substratefor the development and progression of ventricular dysfunction, arrhythmia, anddeath.15 There isevidence that CMR is able to provide images with high spatial resolution and a highlevel accuracy in assessing ventricular function.16 Previous data have shown that even patients withthe indeterminate form may have myocardial fibrosis when tested using CMR.17 However, there is little dataavailable on the degree of myocardial fibrosis presented by these individuals, whichdemonstrates the potential of this variable in the prediction of evolution to thecardiac form. The purpose of this paper is to describe the frequency and extent ofmyocardial fibrosis evaluated using CMR in patients in the indeterminate form. Inorder to contextualize this situation, these patients were compared to otherindividuals in the chronic cardiac form with and without left ventriculardysfunction.The advent of cardiovascular magnetic resonance imaging (CMR), with the use of thelate enhancement technique allows for the identification of myocardial fibrosis.Furthermore, it has a gold standard rating with a close anatomopathologicalcorrelation.Individuals with Chagas’ disease were recruited between January of 2012 andDecember of 2013 at the Chagas’ disease outpatient clinic of the HospitalSão Rafael, a tertiary referral center in Salvador, Bahia, Brazil.Inclusion criteria were: between 18 and 70 years old and a diagnosis of Chagas’disease confirmed by two positive serological tests (indirect hemagglutinationand indirect immunofluorescence). The exclusion criteria were: an acute form ofChagas' disease; previous myocardial infarction, coronary artery disease, or thepresence of two risk factors; primary valve disease; terminal renal disease ondialysis; active hepatitis or cirrhosis; hematological, neoplastic or bonediseases and contraindication to magnetic resonance imaging.The study fulfilled its purpose from the Declaration of Helsinki and was approvedby the Ethics Committee of the São Rafael Hospital. All of theindividuals signed the Informed Consent Term prior to their participation.Trypanosomacruzi infection in the absence of clinical manifestations. Signs ofcardiac involvement were characterized by normal electrocardiograms, chestX-rays and echocardiograms. The cardiac form without ventricular dysfunction wasdefined by individuals with cardiac involvement known as abnormalelectrocardiograms  and without left ventricular dysfunction on the echocardiogram. Thecardiac form with ventricular dysfunction was composed of individuals with areduced left ventricular ejection fraction.The indeterminate form was defined by the presence of a 18All of the individuals underwent biochemical tests, a 12-lead electrocardiogram,a chest X-ray, 24-hour holter monitoring, a cardiac stress test, a Dopplerechocardiogram and a CMR. Scores were calculated as follows: functional classesIII and IV by the New York Heart Association (NYHA) (5 points), X-raycardiomegaly (5 points), left ventricular dysfunction, global or segmentalechocardiography (3 points), non-sustained ventricular tachycardia during the 24hour holter monitoring (3 points), low QRS voltage on the electrocardiogram (2points) and male (2 points). They were then classified into three risk groupsaccording to the score obtained: low risk (0 to 6 points), intermediate risk (7to 11 points) and high risk (12 to 20 points).A CMR was performed using the Sigma HDx1,5-T system . To evaluate how the functioning of the left ventricular, synchronizedimages were acquired from the electrocardiogram in the expiratory apnea,including in the short axis, long axis and the four chamber planes, in differentsequences. The acquisition parameters applied to the dynamic sequence were: arepetition time (RT) of 3.5 ms, an echo time (ET) of 1.5 ms, a flip angle of60º, a bandwidth of 125 kHz, a field of view of 35 x 35 cm, a matrix of 256 x148, a temporal resolution (RT) of 35 ms, a cut thickness of 8.0 mm without aninterval between cuts. Images from the late enhancement technique were acquiredwith each heart beat 10 to 20 minutes after the administration of agadolinium-based contrast agent (0.1 mmol/kg), using a 7.2 ms RT, a 3.1 ms ET,an angle of inclination of 20º, the first phase of cardiac cycle, 16/32 linesper follow-up, a matrix size of 256 x 192, a cut thickness of 8.0 mm, aninterval between 2 mm cuts, a field of view of 32 to 38 cm, an inversion time of150 to 300 ms, a bandwidth of 31,25 kHz, and 2 NEX (number of excitations). Thelate enhancement technique was used to investigate the presence of myocardialfibrosis, which was estimated by a qualitative  method in accordancewith the presence or absence of late enhancement, location and patternpresented; and quantitatively, using percentage values in relation to the totalmyocardial mass. All analyses were performed with the Siemens Argus program.The categorical data were expressed as numbers , and continuous data were expressed as mean ± standarddeviation (SD) or median and interquartile range (IIQ). Normality was determinedby the Kolmogorov-Smirnov test. The comparison of the continuous variables withnormal distribution was performed using Student's unpaired t test and Anova. The Bonferroni method was used for a post-hoc comparisonbetween the groups. Fisher's exact test was used to compare proportions. TheKruskal-Wallis test was used to analyze non-normal continuous variables. Simplelinear regression was used in the associations between fibrotic mass and thefraction of the left ventricular ejection. The analyses were performed using theSPSS program, version 20.0 (IBM), and p values below 0.05 (two-tailed test) wereconsidered statistically significant.As an a priori primary analysis, the frequency and extent of myocardial fibrosisin the indeterminate form was described, and cardiac forms were compared withand without dysfunction. As a post-hoc secondary analysis, the association offibrosis with the ejection fraction and the Rassi score was tested.Additionally, clinical and laboratory parameters were compared between theindeterminate form and the cardiac form without dysfunction.A total of 61 individuals with Chagas' disease, 56% female, with a mean age of 58± 9 years old were evaluated. Seventeen of them were in the indeterminateform, sixteen were in the cardiac form without left ventricular (LV) dysfunctionand twenty eight were in the cardiac form with LV dysfunction. The majority ofthe subjects (74%) were in the NYHA functional class I or II, and 4 (6.6%) hadconcurrent gastrointestinal involvement. 82% were from urban areas, 50individuals had previously lived in a mud house, and 44 reported contact withtriatomine bugs. 64% of the relatives of patients with Chagas' disease reporteda testimony. Eight patients used benzonidazole as an etiological treatment. Theprevalence of hypertension, diabetes mellitus, dyslipidemia and smoking wassimilar among the three groups. The median Rassi score was 5 (IIQ: 0 - 11), andwas distributed as follows: 36 individuals were classified as low risk (59%), 10were classified as intermediate risk (16%) and 15 as high risk (25%). Otherclinical and demographic characteristics are described in Late enhancement was found in 37 of the 58 subjects submitted to a CMR (64%). Thepercentage of the total area of myocardium affected by fibrosis was 9.4% (IIQ:2.4 - 18.4), and was located most frequently in the inferolateral and apical LVsegments. The location of the fibrosis in the subnedocardial and transmuralareas were the most prevalent (72%); however, transmural fibrosis occured morefrequently in those with ventricular dysfunction; p = 0.001. Myocardial fibrosiswas identified in 6 of the 17 individuals in the indeterminate form , which is a proportion similar to that observed in the cardiac formwithout LV dysfunction ; p = 1.0. Among theindividuals with fibrosis, the total area of the affected myocardium was 4.1%(IIQ: 2.1 - 10.7) in the indeterminate form versus 2.3% (IIQ: 1-5) in cardiacform without LV (p = 0.18). In those with ventricular dysfunction, thepercentage of fibrosis was higher than in the other groups, occurring in 23 ofthe 25 subjects (92%), with a compromised area of 15.2% (IIQ: 7.8-25); p <0.001 .The LV ejection fraction was significantly lower in individuals with lateenhancement when compared to subjects without enhancement (69 ± 13 versus48 ± 19%); p < 0.0001. A negative correlation was observed between theextent of fibrosis and ejection fraction . Throughlinear regression analysis, progressive reduction of the ejection fraction wasobserved at each percentage increase in the area affected by fibrosis. Thisanalysis showed a regression coefficient (b) of -0.968, which corresponds to theestimated reduction in the ejection fraction of the LV at each 1% increase inthe area of fibrosis .There was a progressive increase in the amount of fibrosis in the differentclasses of the Rassi score, when subdivided into low, intermediate and highrisk. The high-risk group had 13.8% (QI: 9 - 19) versus 4.9% (QI: 1 - 17) in themedium risk versus 0% (QI: 0 - 5) in the low risk group (p = 0.003). There wasno difference in fibrotic mass between the low and intermediate risk classes (p= 0.19), nor was there a difference between intermediate and high risk (p =1.0).The left ventricular ejection fraction (LVEF) in individuals in the indeterminateform was 72 ± 8%, which is similar to the LVEF of patients in the cardiacform without ventricular dysfunction 67 ± 6%; p = 0.09. There were noNT-proBNP levels , ultrasensitive CRP , TNF-alpha , interleukins (p = 0.35), IFN-gamma and MET  achieved through exercise testing and thelocation of late enhancement (p = 0.44) when comparing patients in theindeterminate form with those in the cardiac form without dysfunction .All 17 individuals in the indeterminate form were classified as low riskaccording to the Rassi score. While 16 individuals were shown to be in thecardiac form without dysfunction, 12 (75%) were considered low risk and 4 wereconsidered intermediate risk; p = 0.04.The present study highlighted the presence of myocardial fibrosis in patients in theindeterminate form of Chagas’ disease. It was present in a frequency and extensionsimilar to that of the group who had the disease in the cardiac form withoutventricular dysfunction. Additionally, it was shown that ventricular function andclinical parameters are similar between these two forms.21CMR has been used for decades for the anatomical and functional evaluation of theheart. It is important due to the fact that it is non-invasive, does not useionizing radiation, and has a high resolution, which allows for multiple studiesconcerning cardiac anatomy, function and tissue characterization with the lateenhancement technique.23 In 2005, Rochitteet al.17 evaluated CMR with theuse of the late enhancement technique in 51 patients with Chagas' heart disease, andfound fibrosis in 68.6% of all of the individuals evaluated, and in 100% in thosewith ventricular tachycardia.17Regueiro et al.23 found fibrosisin patients living outside the endemic area of the disease in a distribution of 7.4%of those in the indeterminate form, 15.8% of those in the cardiac form withoutventricular dysfunction, and 52.4% in those with ventricular dysfunction.23 We found a percentage of fibrosisinvolvement similar to the previous study (64%), also showing progressiveinvolvement in patients with LV dysfunction (92%).17 However, in addition to previous research, ourdata demonstrate a prevalence of enhancement as well as the percentage offibrosis-related area, which is similar between the indeterminate form and thenon-dysfunctional form of the LV. We found no difference in the subendocardial andtransmural location of fibrosis.Previous studies have validated the quantification of myocardial fibrosis using CMRin populations with Chagas' disease.12 However, its designation as an indeterminate formwas impaired since it was impossible to analyze the electrocardiographicalterations. In 1997, Andrade et al.,24 using a canine model, interpreted that the indeterminate formof the disease is characterized by a self-limited cycle of focal inflammatoryalterations, with modulation and suppression of immune responses mediated by cells.Thus, they considered that the indeterminate form of Chagas' disease ischaracterized by a host-parasite equilibrium instead of a progressive damagingprocess.24 As early as1978, Andrade et al.13 reportedthat chronic chagasic myocarditis lesions are not randomly distributed through theatrioventricular conduction system, but rather that there is a clear distribution oflesions in the conduction system.13 We now know that a large percentage of patients in theindeterminate form show evidence of cardiac involvement in the detailed,non-invasive evaluation.25 Thedata from the present study demonstrate that a CMR is not capable of differentiatingthe indeterminate form from the clinical form without LV dysfunction, since thepercentage of fibrosis is similar between the two clinical forms. In this study, thepercentage of fibrosis involvement in the indeterminate form (41.2%) was similar tothe percentage in the cardiac form without LV dysfunction (43.8%).The description of the cardiac ultrastructural changes that occur in theindeterminate phase of Chagas' disease were initially reported in individuals withpositive serology during necropsy after accidental death.Some limitations of the study should be recognized. No anatomical tests wereperformed to definitively rule out ischemic etiology as a cause of myocardialfibrosis. In order to minimize this possibility, an ergometric test was performedwith all of the individuals, in addition to including the exclusion criteria of thepresence of risk factors for atherosclerosis. Although it is recognized that to ruleout coronary artery disease definitively, a coronary angiography would be necessary,the negative predictive value of the exercise test in these circumstances is veryhigh. Coronary artery disease was excluded without performing a coronary angiographyin order to avoid radiation and complications resulting from the procedure.The presence of fibrosis in the indeterminate form of Chagas' disease has a frequencyand extension similar to that of the cardiac form without dysfunction, suggestingthat the former is part of a subclinical disease spectrum, rather than lackingcardiac involvement. Thus, indeterminate and cardiac forms without dysfunctionresemble each other and differ significantly from cardiac form with dysfunction.


Infarct heterogeneity, as assessed by determination of the peri-infarct zone (PIZ) by cardiac magnetic resonance imaging, has been shown to be an independent predictor for the development of cardiac arrhythmias and mortality post myocardial infarction (MI). The temporal evolution of the PIZ post MI is currently unknown. Thus, the main objective of our study was to describe the temporal evolution of the PIZ over a 6 month time period in contemporarily managed ST elevation myocardial infarction (STEMI) patients. Further, given the poor prognosis associated with microvascular obstruction (MVO) post STEMI, we sought to compare the temporal evolution of the PIZ in patients with and without MVO. We hypothesized that patients with MVO would show a relative persistence of PIZ over time when compared to those without MVO.Twenty-one patients post primary percutaneous coronary intervention were enrolled and treated with evidence based therapy. Each patient had three cardiac MRI scans at 48 h, 3 weeks and 6 months post infarction. Repeated Measures Analysis of Variance (ANOVA) was used to assess the evolution of core infarct size and peri-infarct zone size across the three time frames.p = 0.77). In contrast, non-MVO patients did have a significant decrease in PIZ size over time .The patients in this study were predominantly male, with ~40 % LAD territory infarction and a mean LVEF of 46 ± 7 %. Core infarct size and PIZ size both decreased significantly across the three time frames. The presence of microvascular obstruction (MVO), a known adverse prognostic factor, influenced PIZ size. Both patients with and without MVO had a significant reduction in core infarct size over time. Patients with MVO did not have a significant change in PIZ size over time (11.9 ± 6.8 %, 12.2 ± 7.5 %, 10.7 ± 6.6 % Peri-infarct zone size, like core infarct size, varies depending upon the timing of measurement. Patients with MVO displayed a persistence of the PIZ over time. Cardiac magnetic resonance (CMR) imaging is considered the gold standard in the assessment of left ventricular structure and function , 2. HoweTwenty-one patients who presented to Sunnybrook Health Sciences Centre with STEMI were prospectively enrolled in our study between the years 2009–2011. Consent was obtained from all patients. The main inclusion criteria was patients who had met the diagnosis of STEMI. Exclusion criteria included failure to provide consent, significant arrhythmias, severe renal impairment (GFR <30 mL/min) as well as contraindications to CMR imaging such as implantable cardioverter-defibrillators and pacemakers. Six patients who had initially agreed to participate in the study dropped out (not included in the 21 patients mentioned above). Of these, 3/6 dropped out due to claustrophobia, two patients were lost to follow up and one decided not to continue with the study after his first scan. All patients underwent primary percutaneous coronary intervention (primary PCI) of the culprit coronary vessel. Further, all 21 patients received standard of care post STEMI medications including dual anti-platelet therapy, a statin and a beta-blocker. The study was approved by the ethics review board of Sunnybrook Health Sciences Centre.Each patient was imaged at three time points: 48 h, 3 weeks, and 6 months post STEMI. Studies were performed on a 1.5-T clinical scanner  with image acquisition performed using electrocardiographic triggering and breath holds at end-expiration. Cardiac function was determined using contiguous short axis slices covering the left ventricle acquired with a standard SSFP (GE FIESTA) sequence with the following parameters: echo time = 1.6 ms, flip angle = 45°, acquisition matrix = 256 × 192, bandwidth = 125 kHz, and 20 cardiac phases per slice. An inversion recovery gradient echo T1-weighted sequence was used to assess for late gadolinium enhancement with the following parameters: TR = 5.5 msec, TE = 2.5 msec, FOV = 350 × 350 mm, matrix 192 × 128. The inversion time was manually adjusted to null signal from normal myocardium and short-axis images were obtained 10 min after intravenous administration of gadolinium-diethylene triamine pentaacetic acid  –17.Core infarct size, peri-infarct zone size and MVO were quantified from contrast-enhanced T1-weighted images utilizing the previously described full-width-half-maximum technique , 18, 19.T2 quantification was performed using a previously described free-breathing cardiac-gated spiral imaging sequence with T2 preparation. 5 short axis slices were equally spaced to cover the extent of the left ventricle from apex to base. The following parameters were utilized in this sequence; echo times = 2.9, 24.3, 88.2 and 184.2 milliseconds; 12 spiral interleaves with 4096 points each (16.4 millisecond duration) yielding a 1.4 mm in-plane resolution; slice thickness = 6 mm and bandwidth = 125kH , 20.Unpaired t-tests and chi-square tests were used to compare continuous and categorical variables between MVO and Non-MVO patients respectively. Results from these tests are displayed as means ± standard deviations (for t-tests) and as proportions (for chi-square tests). Repeated measures analysis of variance (ANOVA) was then used to compare the evolution of core infarct size and peri-infarct zone size across the three time frames. Tukey’s multiple comparison test was performed to compare preselected time frames (48 h vs. 3 weeks and 3 weeks vs. 6 months). MVO is a known predictor of adverse prognosis and infarct size. We therefore assessed the evolution of the peri-infarct zone, in patients with, and without MVO. In order to assess inter-observer variability in the assessment of the PIZ, a second experienced observer assessed one slice in 12 random patients (6 with MVO and 6 without) at the three time frames. This was done by calculating the Pearson correlation coefficient comparing the first and second observer. A Bland-Altman plot was also constructed comparing difference vs. average of observer 1 vs. observer 2. A p value <0.05 was considered statistically significant. Statistical analysis was done using GraphPad Prism 6.0 .p < 0.0001) and peri-infarct zone size  decreased significantly over the three time periods studied. Tukey’s multiple comparison test showed that in the case of both CIS and PIZ, there was no significant difference between the 48 h and 3 week time points. However, there was a significant difference between the 3 week and 6 month time points.Twenty-one patients were enrolled post STEMI. Table p = 0.03) and a significantly reduced LVEF  when compared to those without MVO. Otherwise, the two groups were similar with no significant differences in terms of mean age, gender, infarct territory, presence of multi-vessel coronary artery disease or rates of major cardiovascular risk factors  and 3 weeks post infarct . In contrast, there was no significant difference in T2 values in the peri-infarct zone of MVO vs. Non-MVO patients at the 6 month time frame . When comparing T2 values in the peri-infarct zone to the remote myocardium, both MVO and Non-MVO patients showed a significant increase in T2 values at 48 h  and 3 weeks  but not at 6 months .MVO patients had significantly elevated T2 values (in ms) in the peri-infarct zone when compared to the Non-MVO group at both 48 h  is an important arrhythmogenic substrate. Roes et al., assessed 91 patients post myocardial infarction who were scheduled for ICD implantation for primary prevention purposes. They found that the extent of the peri-infarct zone was the only significant predictor of appropriate ICD therapy, both on univariate and multivariate analysis . In anotMicrovascular obstruction is a known predictor of adverse outcomes, including cardiovascular death ost myocardial infarction. Cochet at al. assessed the prognostic significance of microvascular obstruction by cardiac magnetic resonance in 184 patients post acute myocardial infarction. CMR was performed within a week following the infarct. On univariate analysis, both core infarct size and MVO were associated with major adverse cardiac events (MACE). However, on multivariate analysis, only MVO was found to be an independent predictor of events . The mecThe finding of increased myocardial T2, likely representing edema in the peri-infarct zone, is driven in part by a marked local inflammatory response. In keeping with this, we noted that at 6 months, during the chronic remodeling phase, T2 values in the peri-infarct zone normalize to values similar to those in the remote myocardium. Further, at 6 months, T2 values in the PIZ are similar between MVO and Non-MVO patients. Whilst the exact mechanism behind the poor prognosis observed with MVO remains unclear, we speculate that a marked early inflammatory response (as measured by T2 and edema) results in greater tissue injury and excessive replacement fibrosis. This then provides a substrate for arrhythmias and ongoing remodeling. However, this theory must be evaluated by future pre-clinical and large multi-center clinical studies to better assess mechanism and the link between evolution and size of the peri-infarct zone, MVO and downstream adverse clinical outcomes.A recent systematic review and meta-analysis examining nine studies with a total of 4686 patients compared culprit only versus complete revascularization performed during the primary PCI procedure or during the index hospitalization in patients presenting with STEMI. The authors reported that there was no significant difference in major adverse cardiovascular events at 90 days and 1 year between the two strategies. However, there was a significant reduction in the rate of repeat revascularization at 1 year with a complete revascularization strategy . In our There are a number of important limitations of this study. First, our study was a single center one that involved a small number of patients. Despite the small numbers, this is a unique cohort, which has undergone contemporary management and comprehensive CMR evaluation at multiple time frames, utilizing well validated CMR sequences on the same magnet, thus reducing systematic bias in the reporting of the results. Nonetheless, future larger studies will be needed in order to determine if the presence of MVO is an independent predictor for the evolution of the peri-infarct zone over time, after adjustment for other relevant clinical factors. Second, there is inherent variability in contouring the myocardium in order to calculate the PIZ. We therefore assessed inter-observer variability in the determination of the peri-infarct zone. This showed excellent inter-observer variability and was similar to results reported by other groups . Third, Both infarct size and peri-infarct zone (‘gray zone’) evolve over time following STEMI. In patients with MVO, there was a lack of regression of the peri-infarct zone.


To determine references intervals for echocardiographic measurements in adult dairy goats.125 clinically healthy, adult dairy goats aged 2–9 years, belonging to two breeds–Polish Fawn Improved  and Polish White Improved , closely related to French Alpine and Saanen, respectively.Non-invasive transthoracic echocardiography examination was performed in unsedated goats in a standing position. Two-dimensional, M-mode and pulsed wave Doppler measurements were obtained. A non-parametric method was applied for determination of reference intervals. Measurements for the two breeds were compared using an analysis of covariance to control for their body weight. Repeatability was assessed using a between-day coefficient of variation and a coefficient of repeatability.Following reference intervals were determined: aortic diameter in diastole 2.2–3.3, left atrial diameter in systole 2.5–4.3cm cm, the ratio of the left atrial diameter to the aortic diameter 0.96–1.5, right ventricular internal diameter in diastole 0.4–1.7cm, left ventricular internal diameter in systole and diastole 1.8–3.2 and 3.2–5.6 cm, respectively, inter-ventricular septum thickness in systole and diastole 0.7–1.5 and 0.5–1.1cm, respectively, left ventricular posterior-wall in systole and diastole 0.8–1.6 and 0.5–1.2cm, respectively, E-point to septal separation 0.3–0.8cm, left ventricular fractional shortening 28–54%, left ventricular ejection fraction 55–86%, maximum Left and Right Ventricular Outflow Tract velocity 80–140 cm/s and 70–130 cm/s, respectively Left and Right Ventricular Outflow Tract pressure gradient 2.5–8.9mmHg and 1.9–6.5mmHg, respectively. Most of the differences between the two breeds could be attributed to different body weight.The study provides echocardiographic reference intervals determined on the highest sample of apparently healthy goats so far enrolled. Since the use of echocardiography in a goat was first reported in 1992, three oThe numbers of animals enrolled in so far published studies were low: 8 goats , 3, 10 gThe study was carried out between June and August 2013 and 2014, and was approved by the III Local Ethics Commission regarding Experiments in Animals at the Warsaw University of Life Sciences-SGGW .Female dairy goats from four dairy farms were enrolled in the study. Two farms were located in the Wielkopolska province  and two were located in Mazovia province . All the herd owners gave permission to conduct the study in their farms.They belonged to two breeds–Polish Fawn Improved (PFI) and Polish White Improved (PWI).To be initially enrolled in the study goats had to be more than 2 and less than 10 year-old, currently in lactation and apparently healthy at the moment of examination , the entire echocardiographic examination was repeated twice at one-day interval by the same echocardiographer (OSJ) to determine repeatability of measurements.The goats were examined in their own barns and they were held in standing position preferably by their owners to make them as relaxed as possible. No sedation was used.th intercostal spaces and the skin was cleaned with surgical spirit and covered with ultrasound coupling gel . All echocardiographic examinations were performed by one examiner (OSJ) using Mindray M7 diagnostic ultrasound system with Phased Array probe (P4-2s). Simultaneously, ECG was recorded using Mindray a built-in ECG module and on its basis diastolic and systolic frames were chosen. Diastolic left and right ventricular dimensions were measured at the beginning of the QRS complex, systolic dimensions were measured at the peak downward point of septal motion.Hair was clipped from small square areas on both sides of the thorax between the 4-6Following views were obtained: right parasternal long axis view optimized for viewing the atria, right parasternal long axis view optimized for viewing the aorta, right parasternal short axis view optimized for: papillary muscle, chordae tendineae, mitral valve, aorta and main pulmonary artery, left parasternal 4 and 5 chambers view , Fig 2. Primary measurements recorded from the two-dimensional echocardiography (2-D) included aortic (AoD) and left atrial diameter (LAD) The frame from right parasternal short axis views optimized for aorta, just after aortic valve closure was selected with three aortic valve cusps seen with symmetry.Primary measurements recorded from the one-dimensional (M-mode) echocardiography included right ventricular internal diameter in diastole (RVIDd), left ventricular internal diameter in systole (LVIDs) and diastole (LVIDd), inter-ventricular septum thickness in systole (IVSs) and diastole (IVSd), left ventricular posterior wall thickness in systole (LVPWs) and diastole (LVPWd) and E-point to septal separation (EPSS).Primary measurements recorded from the Pulsed wave Doppler echocardiography included maximum Left and maximum Right Ventricular Outflow Tract velocity (LVOT Vmax and RVOT Vmax).All measurements were taken in triplicate by the same person who performed echocardiography (OSJ) and then an averaged value was used in further calculations.Derived values computed using the aforementioned primary measurements were:the ratio of the left atrial diameter to the aortic diameter (LAD/AoD),left ventricular fractional shortening (FS) calculated in M-mode with the integrated calculation program according to an equation: FS = 100%×(LVIDd-LVIDs)/LVIDd,3)/(2.4+LVIDd)-(7×LVIDs3)/(2.4+ LVIDs)]×[100/(7×LVIDd3)/(2.4+LVIDd)].left ventricular ejection fraction (LVEF) calculated in M-mode with the integrated calculation program according to an equation: LVEF = [(7×LVIDd2×4 andmaximum Left Ventricular Outflow Tract pressure gradient (LVOT PGmax): LVOT PGmax = LVOT Vmax2×4.maximum Right Ventricular Outflow Tract pressure gradient (RVOT PGmax): RVOT PGmax = RVOT VmaxAnimals with valvulopathy, hydropericardium or hydrothorax, heart wall deformation, any inclusions in the heart cavities (whose origin was not a subject of any further investigation) and turbulent blood flow inside the heart visible in the echocardiography were excluded from the study.Guidelines for obtaining echocardiograms based on external anatomical landmarks were adapted from the ones previously described . Time reNumerical variables were given as an arithmetic mean and standard deviation (SD) if normally distributed, otherwise as a median and interquartile range). Normality of variable distribution was assessed using a Shapiro-Wilk W test. The range of values was always reported. Age and body weight were compared between the two goat breeds as well as between goats selected for repeatability assessment and the remaining goats using a Mann-Whitney U test. Analysis of covariance (ANCOVA) was used for comparison of echocardiographic measurements between the two breeds with control for the role of body weight. A p-value below 0.05 was considered to indicate statistical significance. Statistical analysis was performed in Statistica 10 (StatSoft Inc.).Each measurement was investigated for extreme observations (outliers) using Tukey's rule and an observation was identified as extreme if its value was <Q1–3×IQR or >Q3 + 3×IQR. If a goat lacked more than one primary measurement it was excluded from the analysis. Unless missing data constituted more than 10% of all observations for a given measurement they were replaced with a respective arithmetic mean. Reference intervals (RIs) were calculated in the Reference Value Advisor  with theTo assess the repeatability of the echocardiographic examination performed by the examiner involved in this study (OSJ) two approaches were applied. First, between-day coefficient of variation (CV) was calculated by dividing the overall standard deviation of the two repetitions by their mean  with itsSecondly, the coefficient of repeatability (CoR) was determined. CoR was calculated from the following equation: within-subject SD×√2×1.96 . Within-Total number of 150 goats were examined and 20 did not meet medical eligibility criteria: 10 had hydropericardium , 4 had mitral valve regurgitation, 3 had a heart rate over 150 bpm, 2 had aortic valve regurgitation, and one had a mediastinal tumor.In total 6 outliers were found  and they were removed from a database turning into missing data. As a result five goats were dropped due to missing data and 125 goats were eventually included in the analysis. Twenty two of them lacked one primary measurement– 1 LVOT Vmax , 4 RVOT Vmax (3.2%) and 17 EPSS (13.6%).One hundred twenty five goats were included in the final analysis. Sixty four (51%) belonged to PFI breed and sixty one (49%) to PWI. Goats’ age ranged from 2 to 9 years and was non-normally distributed (p<0.001) with the median of 4 years, IQR from 3 to 5 years and no difference between the breeds (p = 0.898). Neither did body weight have normal distribution (p<0.001). PWI goats weighed between 48–80 kg (median of 67.9) and were significantly heavier than PFI goats which weighed between 46 and 73 kg (median of 58.5) (p = 0.001).Twenty goats selected for the repeatability assessment had neither different body weight (p = 0.985) nor age (p = 0.491) compared to the remaining 105 goats.In 19 cases primary measurements could not be taken due to the low quality of views . RIs for echocardiographic measurements were calculated on 125 goats except for EPSS for which RI was determined on 108 goats as more than 10% of goats lacked this measurement.RIs with 90% CI were determined for all 17 echocardiographic measurements .Time required for image acquisition ranged from 11 to 39 min. with the median of 19 min.CVs for echocardiographic measurements ranged from 3.7% to 26%, however they mainly belonged to the class indicating low variability (10 of 17 CVs), followed by moderate variability (6 of 17). Only CV for EPSS indicated high variability . Mean diMost of differences observed between PWI and PFI goats proved to result from the higher body weight of the former breed . Only riThe study allowed to determine reference intervals for 17 main echocardiographic measurements. Given the large number of animals enrolled, and the use of both commonly accepted echocardiographic procedures and recommended statistical method, these RIs are very likely to show true normal echocardiographic measurements of clinically healthy goats. All the goats in this study were in lactation, however no differences between echocardiographic measurements taken during lactation, pregnancy or dry period have been so far revealed . TherefoGenerally, they are in agreement with RIs for a mixed sample of two breeds (Saanen and Golden Guernsey) previously reported by Hallowell et al. , althougVariability of measurements in the current study was comparable with results of Leroux et al. ,5 when eObviously, the technique of echocardiographic examination strongly affected the RIs and CoRs provided in our study and they are likely to differ when a different technique is used. To allow for their broadest use, we did our utmost to perform echocardiography according to widely accepted standards.th century by mating native goats with two popular dairy breeds–French Alpine (for PFI) and Saanen (for PWI) [Goats of two local Polish breeds were enrolled in the study, which may limit applicability of the results. However, both breeds were developed in last two decades of the 20for PWI) . Actuallfor PWI) . The anaThe study provides echocardiographic RIs for goats determined using the highest so far enrolled group of clinically healthy animals.


Endostatin is a circulating endogenous angiogenesis inhibitor preventingneovascularization. Previous studies demonstrated the prognostic value ofEndostatin among patients with heart failure with reduced ejection fraction(HFrEF). However, the role of Endostatin among patients with heart failurewith preserved ejection fraction (HFpEF) remains unclear.This study aimed to investigate the association between serum Endostatinlevels, natriuretic peptide levels and the severity of left ventriculardiastolic dysfunction and the diagnosis of HFpEF.Endostatin serum concentrations were measured in 301 patients comprising 77HFpEF patients, 169 patients with asymptomatic left ventricular diastolicdysfunction (ALVDD), and 55 controls with normal cardiac function.Endostatin serum levels were significantly elevated in patients with HFpEF(median/interquartile range 179.0 [159-220]) and ALVDD (163.8 [145.4-191.3])compared to controls (149.1 [130.6-176.9]), p < 0.001 and p = 0.004,respectively) and significant correlated with N-terminal pro B-typenatriuretic peptide (NT-proBNP).This hypothesis-generating pilot study gives first evidence that Endostatincorrelates with the severity of diastolic dysfunction and may become a novelbiomarker for HFpEF. We hypothesize a rise in Endostatin levels may reflectinhibition of adaptive angiogenesis and adverse cardiac remodeling. A major drawback regarding the development of new therapies forHFpEF, is the absence of clear diagnostic criteria.10 This makes the definition of patient populationsfor clinical studies difficult. At present, the diagnosis is solely basedechocardiography. Especially, the separation between HFpEF and HFrEF is even morechallenging and misleading in patients with newly diagnosed HF.11 Therefore new strategies fordisease phenotyping in HF are urgently needed. New biomarkers may achieve betterdisease phenotyping.12 Although,many reports have been published on HF biomarkers over the last decades, the impacton clinical decision making is still limited.13 BNP/NTproBNP demonstrated high clinical utility to identifypatients at high risk for heart failure hospitalization and death. However, in thiscontext these markers for clinical studies are only applicable in relatively stablepatients and not in terminal HF patients. Furthermore, the use of BNP/NTproBNP inclinical practice to optimize therapy with drugs, which are known to improvepatient`s outcome is suitable.14However, BNP/NTproBNP is not accepted as surrogate endpoint and can onlyexploratorily be used as endpoint in clinical trials. The appraisal of clinicalutility of BNP/NTproBNP manifests in the current guidelines for the management ofheart failure.15 A number ofpublications propose Endostatin, a potent angiogenesis inhibitor, known mostly fromoncology, as a potential new HF biomarker candidate.18 Mostimportantly Gouya et al. reported in a prospective observational cohort study in 151HF patients, a correlation between elevated circulating Endostatin levels andmortality. Furthermore, this study showed a clear association between Endostatinlevels and progressing diastolic dysfunction, the key characteristic ofHFpEF.19 This is why wehypothesize that Endostatin could potentially be a biomarker suitable to diagnoseand disease phenotype HFpEF patients. In the present study, we aimed to investigatethe sole role of Endostatin as a biomarker for HFpEF and diastolic dysfunction.The patient population affected by heart failure (HF) is growing in a constantmanner. This is because of an aging society, western lifestyle and improved acuteclinical care .The study protocol was approved by the Ethics Committee of the Private University ofWitten/Herdecke, Germany (project n°. 91/08) and conducted in accordance with theDeclaration of Helsinki. Signed written informed consent was obtained from allpatients.20 Thecontrol group consisted of 55 patients  with normal diastolic function(DF). The group with asymptomatic left ventricular diastolic dysfunction (ALVDD)contained 169 patients  with E medial < 8 cm/s, E/E' medial ratio8-15 and NT-proBNP levels < 220 pg/ml. The group with HFpEF comprised 77patients  displaying ALVDD Grad II - III with E/E´ratio> 15, NT-proBNP levels > 220 pg/mL and current or previous signs orsymptoms of heart failure.Participants of the prospective observational cohort study were patientscontacting the HELIOS Klinikum Wuppertal Heart Center  forelective coronary angiography or diagnostic work-up of heart failure. Patientswith a stable or suspected coronary artery disease (CAD) and/or a diagnosticworkup of CHF were included in the study. The exclusion criteria were: leftventricular ejection fraction (EF) < 50%, known CAD with progressive chestpain within the last month, coronary angioplasty or myocardial infarction within6 weeks, hypertrophic cardiomyopathy, moderate-to-severe valvular heart disease,uncontrolled hypertension, atrial fibrillation or other severe arrhythmias,serum-creatinine > 2,0 mg/dl. Patients selected for the control group had tohave no history or symptoms of CHF, a normal ejection fraction > 55%, a ratioof the early diastolic transmitral velocity (E) and the early diastolic tissueDoppler velocity (E´) of < 8, and normal NTproBNP values. A total of 301patients were recruited and assigned to three groups based on echocardiographicdiagnostic criteria as recommended by the European Society ofCardiology.22 The leftatrium volume index (LAVI) was calculated using the biplane area-length method.Left ventricular EF was measured by means of the modified biplane Simpson'smethod.23 Leftventricular mass index (LVMi) was computed with the Devereux formula indexed tothe body surface.22 HFpEF wasdefined in accordance with the EAE/ASE recommendation, based on the assessmentof left ventricular diastolic function.24 Primary measurements included mitral inflow peak early(E-wave) and late (A-wave) diastolic filling velocities as well as systolic (S)and early diastolic (E') mitral annular velocities whereat in each case threeconsecutive beats were measured and averaged. Conventional transmitral flow wasmeasured with Pulse-waved Doppler (PW). PW tissue Doppler imaging (DTI) wasperformed at the junction of the septal and lateral mitral annulus in the apical4-chamber view. Based on primary measurements E/A and E/E' ratios werecalculated.Echocardiography was performed using a standard ultrasound system . A complete transthoracic study wasperformed including 2D, M-mode, spectral and colour Doppler techniques followingcurrent recommendations and guidelines.Peripheral venous whole blood samples were taken after 5 minutes at rest forroutine laboratory testing . Blood was drawn into pyrogen-free tubes without anyadditives, centrifuged at room temperature, aliquoted and stored at -80°C. Alllaboratory analysis were outsourced to Roche Diagnostics  andperformed on blinded samples. For analysis of plasma NT-proBNP the Elecsys 2010NT-proBNP assay  was used. For measurementof Endostatin the ELISA assay of R&D Systems  was used.All assays were performed according to manufacturer's recommendations.th/75thpercentiles (interquartile range) for continuous variables or as absolutenumbers and corresponding percentages for categorical variables unless otherwisespecified. Log transformed values were used for analysis as appropriate. Ap-value < 0.05 was considered statistically significant. We used theKolmogorov-Smirnov test as appropriate to test for normal distribution. TheMann-Whitney U-test was used to analyze differences between the medians of twogroups and the Kruskal-Wallis test to test the equality of medians among morethan two distinct groups. Fisher's Test was used for the comparison of two setsof binary variables and the χ2 test to evaluate differences inproportions in more than 2 sets of categorical variables. Endostatin andNT-proBNP levels were compared across subjects with normal diastolic function,mild ALVDD and HFpEF by the Mann-Whitney U-test, and Jonckheere-Terpstra test.Spearman rank correlation was used to identify variables associated withEndostatin. A multivariable model was included to predict the presence of HFpEFand included the following covariates: Endostatin, age, gender, diabetes,hypertension, coronary artery disease and body mass index. Due to theexploratory nature of this study, there is no minimum required sample size.All analyses were performed using SPSS statistical software . The data are presented as median with 25Increasing quartiles of Endostatin were significantly associated with structuralchanges of the heart like the extent of LV- hypertrophy and left atrialenlargement, reflecting adverse cardiac remodeling. Moreover, increasingquartiles of Endostatin were significantly associated with worsening diastolicfunction measured by tissue Doppler imaging  . Thus, p25 Consistently, wefound that elevated Endostatin levels were associated with elevated NT-proBNPlevels, a well-recognized prognostic marker and indicator of elevated ventricularfilling pressures among patients, independent from LVEF.26We hypothesized that circulating Endostatin levels are altered in patients with ALVDDand HFpEF. Furthermore, elevated levels are associated with the presence andseverity of diastolic function abnormalities. To verify the hypothesis we performeda clinical observational study including 301 patients, which were assigned based ontheir echocardiographic characteristics to three different groups. To our knowledge,this is the first published report linking increased circulating Endostatin levelsto the presence and severity of diastolic function abnormalities and HFpEF in a wellphenotyped cohort of patients with normal systolic function. In the present study,Endostatin showed a graded increase from controls over ALVDD to HFpEF. Furthermore,higher Endostatin levels were significantly associated with established markers ofstructural cardiac abnormalities including the LAVi and increased LV mass as well asfunctional abnormalities like E/E' ratio. Particularly, an increased LAVi withoutconcomitant mitral valve disease reflects a chronic remodeling process typical forHFpEF.27Endostatin plays a role in the local balance of angiogenesis and shows potentanti-angiogenic activity by inhibiting proliferation and migration of endothelialcells in addition to inducing endothelial cell apoptosis.27 Endostatin is produced by the proteolytic cleavageof the C-terminal domain of collagen XVIII, a component of the extracellular matrix.The precise mechanism of conversion from collagen XVIII to Endostatin has not yetbeen fully elucidated.29 Recent studies of patients withcoronary artery disease (CAD) demonstrate that Endostatin protein levels correlatesignificantly with reduced angiogenesis and poorly developed cardiac collateralvasculature.30Endostatin, a 20-kDa proteolytic fragment from the C-terminal domain of collagenXVIII, was shown to have an inhibitory effect on tumor growth working as anantiangiogenic growth factor.33 Nonetheless, angiogenic andantiangiogenic growth factors often co-exist in tissues with angiogenesis.34 Thus, the status of endothelialcells and endothelial function is determined by a balance between these positive andnegative factors on angiogenesis, and the balance may in inappropriate shiftedtowards antiangiogenic factors in patients with HF. It was shown that the role ofmicrovascular dysfunction and microvascular inflammation is especial for patientswith the diagnosis of HFpEF.36 A new pathophysiologic model presented by Redfield etal.8 points frompro-inflammatory coexisting conditions to systemic endothelial inflammation andimpaired oxygen delivery.8 Globalventricular performance is highly dependent on oxygen supply and thus myocardialperfusion, and an essential component of myocardial perfusion during ventricularhypertrophy is the myocyte-microvascular balance and the myocyte/capillary ratio. Incardiac autopsy specimens, it has recently been shown that microvascular rarefactionis a downstream phenomenon in HFpEF.37 Furthermore, Kitzman et al.38 has demonstrated that HFpEF patients display significantabnormalities in the skeletal muscle as well as an abnormal capillary-to-fiberratio, probably building the basis for severe exercise intolerance in HFpEFpatients.38 In addition,Gouya et al.19 have shown in arelatively small HFrEF study population that high levels of serum Endostatin wereassociated with all-cause mortality and concluded that the effect of increasedangiogenesis is HF may be blunted by an overspill of anti-angiogenic factors such asEndostatin.19 Thus, wehypothesize that similar pathophysiological concepts may be involved in patientswith HFpEF, were a high proportion of patients has a coincidence of coronary arterydisease and diabetes, both damaging the endothelial structure.39 This was also true for our patientpopulation as shown in table 1. Endostatin could be moderator of the microvasculareffects seen in these patients.40The results from our study fit well to the pathophysiological model used to explainthe development of HFpEF. In general, HFpEF is a complex disease involving aninterplay of various factors. There is the hypothesis that a failure of oxygendelivery to the cardiomycytes triggers a pro-angiogenic response in patientssuffering from heart failure.Several limitations of this study must be acknowledged. The observational nature ofthe present study prohibits definitive determination of cause and effectrelationships. Second, the present study was a single-center experience with arelatively small number of subjects. Third, longitudinal follow-up data were notavailable to test associations between the Endostatin serum levels and clinicaloutcomes. Moreover, we enrolled consecutive patients referred for elective coronaryangiography and echocardiography, which may not represent a general populationcohort without evidence or suspicious for cardiovascular diseases.Further studies should include more patients from a broader population and capturelongitudinal data including information about hospitalization and mortality.In this exploratory hypothesis-generating study, we provide first evidence thatEndostatin correlates with the presence and severity of diastolic dysfunction andHFpEF and may become a novel biomarker for the diagnosis and stratification ofHFpEF. Increased Endostatin levels may reflect deterioration of diastolic functioncaused by adverse remodeling. Further prospective studies are needed to determinethe causal relationship as well as the diagnostic and prognostic value of Endostatinin HFpEF and the potential role as a therapeutic target.


Currently, many methodological approaches have been developed to assess peripheral endothelial function. However, a development of the noninvasive and automated technique for routinely assessing endothelial function is still required. We evaluated the potential value of a new method to measure peripheral endothelial function with reactive hyperemia peripheral arterial volume (RH-PAV) in patients with chest pain.We used a novel oximeter-like probe to detect the peripheral arterial volume (PAV) of the finger and compared it with brachial flow-mediated dilation (FMD) performed in 93 consecutive patients with chest pain. The RH-PAV index was defined as the ratio of the digital pulse volume during reactive hyperemia relative to the baseline.r = 0.69, p < 0.01). Similar to FMD, PAV was more impaired in patients who have more cardiovascular risk factors (CRFs). The subjects with CAD had lower PAV and FMD, compared with those without CAD , and the relationships between FMD and PAV were also significant in both CAD  and non-CAD  patients.Ninety-three patients  completed the study, and 53 patients demonstrated coronary artery disease (CAD) following scheduled coronary angiography. There was a moderate linear relationship between PAV and FMD (Endothelial function of digital artery assessed with the novel PAV method demonstrated a profile similar to that of brachial artery measured with FMD. The hyperemia PAV was decreased by factors which were considered to impair endothelial function, suggesting that PAV has the potential to be a novel method to study endothelial function. Cardiovascular diseases have become a global challenge in public health, and atherosclerosis is a major one that can lead to ischemia of the heart, brain or extremities, occasionally resulting in infarction . EndotheEndothelial function can be evaluated by detecting its capacity to perform the various functions, such as regulation of vasomotor tone which has become the most prevailing study endpoint , 4. The Nonetheless, the development of a noninvasive and automated technique for routinely assessing endothelial function is still required, from a clinical perspective. The measurement of pulsatile volume changes of index finger during reactive hyperemia has been proved to be an effective way to evaluate endothelial function . ForearmPatients were eligible for enrollment if they had chest pain, were scheduled to undergo coronary angiography (CAG) and were aged between 30 to 75 years. Exclusion criteria included acute coronary syndrome; left ventricular ejection fraction < 55%; Raynaud’s disease; atrial fibrillation; valvular heart disease; chronic respiratory; renal disease; thyroid disorders or sex hormone disorders. This study protocol conformed to the ethical guidelines of the Helsinki Declaration, approved by the ethical review board of China-Japan Union Hospital of Jilin University (2017040603), and the written informed consents were signed by all patients.The following cardiovascular risk factors (CRFs) were evaluated in each individual: male sex; hypertension; diabetes mellitus ; hyperlipidemia ; family history of CAD; postmenopause; and current smoking(have smoked 100 cigarettes in their lifetime and currently smoke cigarettes).In a dimly lit and temperature controlled room, PAV and FMD were performed on the early morning of the scheduled CAG. Participants were instructed to refrain from vasoactive medication, food, caffeine, tea, tobacco, or exercise for at least l2 hours before the measurement. CAD was defined as 50% or more luminal narrowing in one or more major epicardial vessels as determined by CAG.FMD was measured according to the guidelines . FollowiThe basic principle of PAV is the same as FMD and PAT, this technique  includes a photo-plethysmographically based index finger probe to measure digital arterial volume changes accompanying pulse waves. Light-emitting diodes (940 nm) in conjunction with a light-sensitive sensor were used to assess the absorption of the infrared light. The calculation was made by the difference in digital artery hemoglobin flow between pre- and post-occlusion of an upper limb.The blood pressure cuff was placed on the left upper arm undergoing hyperemia testing, while the contralateral arm served as a control. PAV was performed by placing the probes on the index finger of each Fig. . The occP ≤ 0.05 was considered statistically significant.Data are presented as mean ± standard deviation(SD). The two-sided t-test, Pearson rank correlation, and linear regression analysis were used to assess the relationship between FMD and PAV. Univariate analysis was used to evaluate the association between the PAV and various CRFs. Receiver operating characteristic (ROS) curves of FMD and PAV was performed to assess the discrimination performance of CAD. All analyses were done by SPSS 17.0 statistical software . Ninety-three individuals (58 ± 5 years) completed the study, and 53 of them had CAD. We describe the characteristics of our study population through Table r = 0.69, p < 0.01)  and non-CAD  populations .The average of FMD and PAV were 8.3% ± 3.4%(range 2.0-15.1%) and 1.21 ± 0.35(range 0.60-2.22), respectively. Linear regression analysis revealed that FMD and PAV were significantly correlated in all patients (01) Fig. . The sigons Fig. . The aver = 0.45, p < 0.01), as well as FMD , and the trend is similar . The PAV was significantly lower in patients with hypertension(1.10 ± 0.31), hyperlipidemia(1.11 ± 0.31), diabetes mellitus(1.02 ± 0.23) or family history of CAD(1.17 ± 0.25) than those without such CRF. Postmenopausal women (1.13 ± 0.34) had lower PAV than premenopausal ones(1.5 ± 0.32 P < 0.01). There was no significant relationship between the PAV and the gender, age, or current smoking.The average number of CRFs was 3.0 ± 1.2 in each patient. Spearman correlation analysis revealed a significant relationship between the number of CRFs and PAV (P < 0.01 for both). The area under the curve (AUC) of PAV and FMD was 0.785  and 0.810  for the prediction of CAD in ROC curves  with the pairwise comparison of ROC curve test. The sensitivity and specificity of PAV were 83 and 65%, while those of FMD were 85 and 63% for detection of CAD when 1.28 of PAV and 9.8% of FMD was identified as the cutoff points.The subjects with CAD had lower PAV(1.05 ± 0.23) and FMD (6.7 ± 2.9%) compared with those without CAD (1.41 ± 0.37, 10.4 ± 2.9%, Our pilot study was performed to evaluate the feasibility of reactive hyperemia PAV to assess endothelial function of the peripheral artery in patients with chest pain. The data demonstrated that the significant relationship between PAV hyperemia ratio and brachial artery FMD not only existed in all subjects but also in both CAD and non-CAD groups. Similar to FMD, peripheral arterial reactivity, defined as the PAV, also correlated with traditional CRFs and CAD. Since there is a significant relationship between abnormalities in the central and peripheral arteries in the process of atherosclerosis, peripheral vascular endothelial function testing has been evolving to be more feasible for daily test and screening large numbers of people . Our finThe PAV was lower in patients with diabetes mellitus, hypertension, hyperlipidemia, or family history of CAD, but there was no significant relationship between the decreased PAV and age, gender or current smoking. Previous studies have reported that the prevalence of reduced FMD increased substantially with advancing age, and the association attributed to alterations in the production of nitric oxide and oxidant species , 18. In Our study demonstrated that patients with CAD had attenuated PAV, compared to those without CAD, and the discrimination value of CAD existence was similar between PAV and FMD. Although it has been reported that peripheral vascular endothelial function measured by blood flow or reactive hyperemia after cuff release are not solely nitro oxygen-mediated, it possesses independent predictive value . VasodilThe absence of major muscle, high vascular density and good perfusion of the finger make it accessible for measuring changes in the volume of blood flow . The phoAs described above, the PAV technique is operator independent, analyzing automatically, requiring less training and the contralateral arm serves as its internal control that can be used to correct for any systemic changes during the test. Several limitations of this study and the technique deserve to be mentioned. First, only traditional CRFs were considered and other risk factors may also have potential to impair endothelial function. Gender, age and current smoking were acknowledged to impair endothelial function, but the relationships between them and PAV were not significant, the bias may affect the result. Second, reproducibility is important when vascular reactivity tests are compared . Future Our data indicated that the novel PAV method showed similar patterns of abnormality to FMD in patients with chest pain. PAV is influenced by factors proven to affect endothelial function, such as the presence of CRF and CAD. In addition, PAV is potentially useful for identifying patients at high risk for CAD, suggesting that PAV has the potential to be a novel noninvasive method in the field of cardiovascular research.


Myocardial Infarction with Non-Obstructive Coronary Arteries (MINOCA) is common with a prevalence of 6% of all patients fulfilling the diagnosis of myocardial infarction. MINOCA should be considered a working diagnosis. Cardiac Magnetic Resonance (CMR) imaging has recently been suggested to be of great value to determine the cause behind MINOCA. The objectives of this paper are to describe the rationale behind the second Stockholm Myocardial Infarction with Normal Coronaries (SMINC-2) study and to discuss the protocol for investigation of MINOCA patients in the light of the recently published position paper from the European Society of Cardiology.p-value of less than 0.05 when compared with CMR imaging in the SMINC-1 study. Furthermore, in addition to invasive coronary angiography, coronary arteries are evaluated by computed tomography angiography to investigate coronary causes and questionnaires are used to describe Quality-of-Life (QoL). By January 1st 2017, 75 patients have been included.The SMINC-2 study is an open non-randomised study using historical controls for comparison. The primary aim is to prove that MINOCA patients investigated with the latest CMR imaging technique can achieve a diagnosis in 70% of all cases entirely by imaging. By including 150 patients we will have >80% chance to prove that the diagnostic accuracy can be improved by 20 absolute % with a Whether CMR imaging can provide a diagnosis to an adequate proportion of MINOCA patients is unknown. Well-defined inclusion and exclusion criteria will be used to compare a MINOCA cohort from the population with an appropriate control group. Positive results are likely to influence future guidelines of the management of MINOCA. Furthermore, the study will give mechanistic insights into MINOCA in particular in patients with “true” myocardial infarction and describe QoL in this vulnerable group of patients.NCT02318498.Clinical Trials  Myocardial Infarction with Non-Obstructive Coronary Arteries (MINOCA) is common with a prevalence of 6% of all patients fulfilling the diagnosis of myocardial infarction . A valueprimary aim to give 70% of the patients a definite diagnosis purely on the basis of the CMR imaging results. To be able to do this, the protocol states that CMR imaging should be performed 2-4 days after hospitalisation because of MINOCA and to include T1 mapping in addition to T2 oedema sequences. An overall secondary aim is to test a protocol on how to investigate MINOCA patients including further characterisation of these patients with emphasis on coronary imaging and stress. The objectives of this paper are to describe the rationale behind the SMINC-2 study and to discuss the protocol of investigation of MINOCA patients in the light of the recently published position paper [The Stockholm Myocardial Infarction with Normal Coronaries (SMINC) study was a study with the primary aim to investigate the role of endothelial function in MINOCA . The reson paper .p-value of less than 0.05 when compared with our previous sample [The study design is an open non-randomised multi-center study using historical controls from the SMINC study for comparison. The primary aim is to prove that patients investigated with CMR imaging can achieve a definite diagnosis in 70% of all MINOCA cases entirely by imaging . By including 150 patients, we will have >80% chance to prove that the diagnostic accuracy can be improved by 20 absolute % with a Consecutive patients from the five major hospitals in Stockholm, 35-69 years old, fulfilling the existing diagnostic criteria of myocardial infarction [Patients without sinus rhythm on admission ECG, pulmonary embolism, COPD, previous myocardial infarction, known cardiomyopathy or severe renal impairment (serum creatinine above 150 umol/l) will be excluded.Patients with a pacemaker and claustrophobia will be excluded due to interference with interpretation or possibility to perform CMR imaging.An overview including the time-frame of investigations is shown in the Fig. . 10. Phase contrast-based flow measurement sequence, without breath-holding, through aorta ascendens and pulmonary artery.CMR is performed by a Siemens Aera 1,5 T® including the following steps: 1. Scout through thorax with transversal, coronar and sagital stacks with trueFISP-sequences, without ECG triggering. 2. Vertical long-axis through left ventricle with trueFISP without ECG triggering. 3. Horisontal long-axis through left ventricle with trueFISP cine, with ECG triggering. 4. Pre-contrast T1-mapping with MOLLI (Modified Look-Locker Inversion recovery) sequence containing several ECG-triggered single shot trueFISP in end-diastole with different times of inversion. Sequences should be sampled in short-axis, 2,3 and 4 chamber view stacks. 5. Pulmonary transition time, saturation recovery trueFISP perfusion, single slice short-axis through left atrium, 20 mm slice thickness, without ECG triggering, parallel -imagination with GRAPPA factor 4 with 2 ml bolus of contrast (Dotarem®) + 20 ml NaCl, 4 ml/s injected by automated injector. 6. Further contrast injection (Dotarem®) to double dose injected by automated injector. 7. Cine trueFISP in short-axis, 2,3 and 4 chamber view stacks to study left ventricular function. 8. TI-scout followed by phase sensitive inversion recovery (PSIR) late gadolinium enhancement (LGE) in short-axis, 2,3 and 4 chamber view stacks, to study infarction and microvascular obstruction. Should be made at least 10 min after contrast injection. 9. MOLLI post-contrast T1-mapping in short-axis, 2,3 and 4 chamber view stacksAcute myocardial infarction is defined as localized subendocardial late gadolinium enhancement (LGE) with corresponding oedema (T1 mapping).Acute myocarditis is defined as diffuse mainly subepicardial LGE with corresponding oedema (T1 mapping).Acute TCM is defined as presence of myocardial oedema (T1 mapping) and/or transient wall motion abnormalities with circumferential distribution with absence of corresponding LGE and coronary artery distribution.Computed tomography angiography (CTA) is performed using a 64-slice computed tomography scanner with ECG-triggered, prospective technique and with parameter settings resulting in submillimeter isotropic voxels. Patients with a heart rate > 65 beats/min will be given betablockers, orally or intravenously, before the scan. Patients with a systolic blood pressure > 100 mmHg will be given sublingual nitroglycerine before the scan. First, a non-enhanced scan will be performed enabling measurement of calciumscore. Second, nonionic iodinated contrast agent will be administered as a standard bolus volume but with individually measured time of delay. Data within the end-diastolic phase will be reconstructed in 5% steps and thereafter analyzed using dedicated software with post-processing tools such as multiplanar reformation, curved multiplanar reformation and maximum intensity projection. Images will be analyzed semi-quantitatively regarding number, location, severity and quality of coronary plaques by 2 observers who are blinded to the clinical information where disagreement will be solved by consensus.k, c, and x are calculated. These constants correlate closely to chamber stiffness, viscoelastic energy loss and load, respectively. Furthermore, the results are used to calculate the time constant of isovolumic chamber relaxation (tau), the stiffness and relaxation components of the deceleration time of the E wave, a load independent index of diastolic filling and stiffness.Echocardiography is performed on GE Vivid E9® scanners. All registrations besides CW/PW Doppler consist of three cardiac cycles. The ejection fraction will be calculated using Simpson’s rule. Regional wall motion will be classified by visual inspection of wall thickening, classifying segments with less than 30% thickening as hypokinetic, less than 10% as akinetic, and segments with systolic thinning or eccentric excursion as dyskinetic. 2D loops of from the three apical views are analyzed using the longitudinal 2D strain Q-analysis module in EchoPAC BT11.2® . After marking the myocardial borders as the region of interest (ROI), the application semi-automatically generates strain curves and peak systolic strains, and also automatically calculates the proportion of shortening that takes place after aortic valve closure (post systolic index). Furthermore the temporal derivation to strain rate can be performed. The software calculates global averages as well as segmental values for all variables, and both will be recorded. Color coded tissue Doppler loops from the same apical views will be analyzed in the TDI Q-analysis module. ROIs will be placed in the two basal segments in each view, in order to measure the peak systolic and early diastolic velocities, as well as the timing of the events of the cardiac cycle, which will be used to calculate the myocardial performance index for the left ventricle, and in the same manner for the right ventricle. The standard diastolic parameters of mitral inflow velocities, E/E’, and pulmonary venous flow will be measured. The E waves acquired during free breathing and after the Valsalva maneuver will be analyzed in a custom LabVIEW®  interface where, by fitting the curve generated by the equation for a simple harmonic oscillator to the maximum velocity envelope of each E wave, the equation constants Laboratory investigations are performed by routine clinical chemistry. Haematology, electrolytes, cystatin C, glucose, lipids and NT-proBNP, thyreoid function tests, hsTroponinT, D-dimer, metoxycatecholamines and urine albumin/creatinine ratio, oral glucose tolerance test (75 g with measurement of glucose after 2 h), HbA1C and cortisol in saliva (morning and evening). All patients will be investigated haematologically to exclude proneness to thromboembolism and systemic lupus erytematosus at 6 months according to the routine of Karolinska University Laboratory.Quality-of-Life is investigated: Rand 36 (former SF-36), Multidimensional Fatigue Inventory (MFI-20), Perceived Stress Scale (PSS-14) and Hospital and Anxiety Depression Scale (HADS).The primary objective of the present paper is to describe the rationale behind the SMINC-2 study that has the aim to test if early CMR imaging using the latest technique can give more than 70% of all MINOCA patients a CMR imaging-based diagnosis. The main diagnoses that are possible to acquire are myocarditis, TCM and “true” myocardial infarction. Our previous meta-analysis of CMR imaging findings in MINOCA showed that one third of the patients had myocarditis and 20% had “true” myocardial infarction . These nA secondary objective of the present paper is to discuss the protocol of investigation of MINOCA patients in the light of the recently published position paper by the ESC . In the The major strengths of SMINC-2 are the fairly high number of patients included from several hospitals being representative for a north European population and in comparison with several previous studies it is prospective and the first one with a control group. Limitations include the fairly strict age criteria and that it will not include patients with near significant coronary artery stenosis to be able to have the SMINC study as a control group. The reasons for the strict age criteria in the SMINC study were that it was unlikely that all patients would receive a coronary angiography if patients were below 35 and above 69 years of age. The reason for the lower age limit is that the likelihood is much higher for myocarditis than CHD and thus the diagnosis would be made without a coronary angiography. Regarding the upper age limit, many patients 70 years and older with suspected CHD were treated conservatively at the time of the start of the SMINC study 10 years ago. This together with the fairly strict inclusion and exclusion criteria may somewhat limit the representativity of the study.Whether CMR imaging can provide a diagnosis to an adequate proportion of MINOCA patients is unknown. Well-defined inclusion and exclusion criteria will be used to compare a MINOCA cohort from the population with an appropriate control group. Positive results are likely to influence future guidelines of the management of MINOCA. Furthermore, the study will give mechanistic insights into MINOCA in particular in patients with “true” myocardial infarction and describe QoL in this vulnerable group of patients.