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Rebecca Levene

Cardiac magnetic resonance of targeted annexin-iron oxide labeling detects cardiac cell death in vivo after doxorubicin and myocardial infarction. Background: Heart failure from myocardial infarction MI or doxorubicin DOX , used in cancer therapy, is preceded by significant cell apoptosis.

Real-time, non-invasive detection of early cardiac apoptosis might impact patient treatment and outcomes. Early apoptosis is detected by Annexin V protein ANX binding to externalized membrane phosphatidylserine. This conjugate specifically binds to early apoptotic cardiac cells in culture and is detectable by in vitro magnetic resonance imaging MRI. This molecular imaging strategy may help identify 'at risk' cardiac cell populations.

Integrated analysis of diastolic, systolic and pulmonary vascular function using MRI guided catheterization. Indroduction and purpose: An integrated approach for assessing ventricular pump function, diastolic compliance EDPVR , myocontractility and pulmonary vascular resistance would be of clinical interest. In addition to pump function, MRI guided catheterization was demonstrated to accurately measure myocontractility and vascular resistance.

Subsequently, this approach was applied in patients with Fontan hemodynamic in which abnormalities in pulmonary vascular, myocontractile and diastolic properties are debated. In the patients during dobutamine we noted failure to increase stroke volumes despite increased contractility and evidenced diastolic dysfunction. Conclusion: This novel MRI method provides differential information about diastolic, systolic and vascular function.

The method evidenced that in Fontan patients diastolic dysfunction is an important pathophysiologic cause of heart failure. Diffusion-prepared dark blood delayed enhancement imaging for improved detection of subendocardial infarcts. Introduction: Delayed enhancement MRI enables detection of non-viable myocardium with high spatial resolution and has.

Diffusion preparation-gradients have been used to create dark-blood vessel wall images [4], but have never been applied post-contrast for infarct imaging. Purpose: To develop a dark-blood delayed enhancement pulse sequence based on diffusion preparation which would not rely on complete blood exchange and would be relatively insensitive to changes in relaxation times.

Methods: A diffusion-prepared inversion recovery IR pulse sequence was developed by adding a driven equilibrium module using a BIR-4 0 degree radiofrequency pulse with motion sensitization gradients between its components prior to a segmented FLASH readout. The timing of this preparation was optimized to minimize the effects of strain-induced signal loss.

The technique was tested in a canine model of chronic infarction using a I. Images were obtained 5—10 minutes after injection of. There is an infarct in the inferior wall which is difficult to distinguish from the blood pool. Figure 1 b displays an image of the same slice that demonstrates the utility of the diffusion-prepared IR sequence.

Here, the diffusion preparation causes suppression of the blood pool, improving the ability to detect the subendocardial region of infarction. Conclusion: We have developed a new dark-blood delayed enhancement pulse sequence which attenuates the blood pool based on motion sensitization. In preliminary studies, this technique improves delineation of subendocardial infarcts.

However, subendocardial infarcts are sometimes difficult to detect as they may demonstrate similar image intensity as the ventricular cavity. A double inversion dark-blood pulse sequence to create black blood delayed enhancement images of myocardial infarction has been previously described [I, 2, 3]. The sequence relies on precise timing of non-selective and selective inversion pulses, and it is sensitive to incomplete exchange of blood and changes in TI relaxation of the blood and myocardium.

The technique significantly improves blood-infarct contrast at the. Introduction: Black-blood coronary vessel wall imaging is a powerful non-invasive tool for the quantitative assessment of positive arterial remodeling [1]. In addition, atherosclerosis is a spatially heterogeneous disease and therefore imaging at multiple anatomical levels of the coronary circulation is mandatory. However, this requirement of enhanced volumetric coverage typically comes at the expense of increased. Consequently, single-breath-hold multi-slice black-blood coronary vessel wall imaging is enabled using PS-DIR.

Reconstruction: A local region-growing reconstruction algorithm was developed and is summarized in Fig. Pixels with high. DIR pulse sequence. The phase values of these points are used to estimate the local signal phase inhomogeneity which is needed for local signed-magnitude image reconstruction [4]. A ring-shaped region-of-interest is selected around the coronary wall a. Phase points with high magnitude SNR are selected for region-growing b. Map of local phase inhomogeneity is created using bi-linear interpolation c.

Inhomogeneity is removed from the phase image d and a final signed image is calculated e. Data were acquired using a segmented k-space spiral acquisition with spectral spatial excitation [6]. Image processing was performed off-line on a personal computer. Experiments: Anatomical slices perpendicular to the proximal part of the right coronary artery RCA at end-systole were planned similar to a previously published methodology [7].

CNR was calculated on the signed-magnitude images reconstructed with the above algorithm. Finally, a dual-slice rather than a multi-phase version of the sequence Fig. Consistent with the visual findings, Fig. TI-insensitivity can be traded for enhanced volumetric coverage at no extra-cost in imaging time. Note the simliar shape and extent of the calcification in both the CT and MR results.

Inserts are zoomed images of the cross-section of the vessels in reformatted transverse images. Imaging the vessel wall in major peripheral arteries using susceptibility weighted imaging: visualizing calcifications. Introduction: Magnetic resonance imaging MRI has been used for many years to study atherosclerosis [I]. Black blood techniques are the most ubiquitous and are used to suppress the signal from flowing blood, making the vessel wall more conspicuous. The purpose of this study was to demonstrate a novel approach to imaging the vessel wall and vessel wall calcification using susceptibility weighted imaging [2] SWI with no need to suppress the signal from the blood.

Methods: Optimizing the imaging parameters: The SWI sequence parameters were optimized to allow for the best visualization of the femoral artery lumen in the magnitude images and the arterial wall in the phase images. Parameters such as resolution for time considerations , flip angle for contrast in the magnitude images and echo time for phase contrast were considered.

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Vessel wall magnitude and phase measurements: ROIs from the top to the bottom of the visible portions of the femoral artery were taken. Calcification Measurements: The area of calcification was manually measured on CT images and MR images both magnitude and phase images by an experienced radiologist. The magnitude contrast-to-noise ratio CNR between artery and vessel wall was The phase CNR between the arterial wall and the lumen was A total of 19 calcifications in the femoral vessel wall were identified with SWI in 8 subjects.

The mean area of calcification measured on CT, magnitude and phase images was 0. The Pearson correlation coefficient of the measured lesion area between CT and magnitude image is 0. A typical case having popliteal artery calcification is shown in Figure 1. Both magnitude and phase images show the calcifications clearly in the popliteal artery wall and correlate well with the CT image. Conclusion: SWI offers a means to image a large field-of-view over which the arterial wall can be clearly seen in both magnitude and SWI filtered phase images. These lesions were seen in CTand SWI and correlated well in both size and position with both methods.

We anticipate that SWI will play a complementary role to the current multi-contrast approach in studying atherosclerosis. Background: Cardiovascular magnetic resonance CMR has a broad range of clinical applications and is increasingly used in daily clinical practice in many European countries. Methods: Multicenter registry with consecutive enrolment of patients scanned in 29 German CMR centers using web based online case record forms.

Ninety-three percent of patients received a gadolinium based contrast agent. Severe complications occurred in a minority of patients 0. No patient died during or due to the CMR procedure. Conclusion: CMR is a frequently performed in German clinical practice. Sudden cardiac death with normal coronaries: cardiac MRI in the differential diagnosis of underlying disease in survivors. Introduction: Sudden cardiac death SCD is most commonly caused by acute myocardial infarction as a correlate of coronary artery disease.

However, in cases with normal coronary arteries SCD often remains unexplained. Diagnostic work-up in this collective is important to adjust and optimize therapy. Purpose: Aim of this study was to evaluate the use of contrast-enhanced cardiac MRI CMR in defining the underlying pathology of survived SCD in patients without coronary artery occlusion. Methods: More than 6. The CMR protocol I. Data sets for late gadolinium enhancement detection were acquired 8-I5 min after 0.

All cases of non-coronary SCD were reviewed and the different underlying pathologies as defined by MRI were collected. In I4 patients thereof, MRI could state the diagnosis based on the typical imaging features of myocardial disease: Primary cardio-myopathy was found in 7 patients arrhythmogenic right ventricular cardiomyopathy, 2; dilated cardiomyopathy, 3; hypertrophic cardiomyopathy, I; isolated left ventricular non-compaction, I. Acute myocarditis and acute cardiac sarcoidosis were found in 3 patients each.

Chronic aneurysm of the anterior wall most probably due to cardiac contusion was found in I patient. Conclusion: Contrast-enhanced CMR has unique features in detecting and differentiating myocardial disease with possibly fatal outcome. It has proven to be an utmost valuable tool for the diagnostic work-up of survivors of unclear SCD. This underlines the role of CMR as the first-line technique in myocardial disease.

Introduction: Diastolic dysfunction, often preclinical with no recognized CHF diagnosis, is associated with marked increases in all-cause mortality. Mean SD , correlation coefficients and multivariable regression coefficients were determined. Results: Table I illustrates associations between risk factors with diastolic LV function. I7] Fig I. Comapred to non-smokers smokers had lower peak filling rates. Compared to Whites, Hispanics were at a higher risk for diastolic dysfunction, Chinese ethnicity showed a relative protective effect after adjusting for all other risk factors.

Table l abstract Ol5 Multivariable regressions analysis of predictors of diastolic LV dysfunction. Conclusion: Impaired LV relaxation is associated with increasing age, male gender, obesity, diastolic hypertension, smoking, and varied by ethnicity. Correlation of pericardial and mediastinal fat with coronary artery disease, metabolic syndrome, and cardiac risk factors. Background: Obesity and abdominal fat have been shown to correlate with coronary artery disease CAD and may play a role in development of metabolic syndrome MS.

Methods: We enrolled consecutive patients, 63 male, who underwent CMR for cardiac evaluation and had coronary angiogram performed within 12 months. The baseline characteristics of these patients were as follows: Eighty had hypertension HTN , 42 had diabetes mellitus DM , 37 had hyperlipidemia and. The surface area of fat was measured by computer analysis from free-hand region of interest ROI curves. MS was considered positive if the patient had 3 or more of the.

Data in Table I. Prognostic significance of magnetic resonance imaging parameters in patients with idiopathic dilated cardiomyopathy. Background: Patients with idiopathic dilated cardiomyopathy IDC have a limited prognosis. Aim of this study was to evaluate the prognostic significance of novel magnetic resonance imaging CMR parameters in IDC patients. Interventricular dyssynchrony Primary endpoint was cardiac death, sudden death SCD and rehospitalization for pump failure, secondary endpoint cardiac death and SCD.

Apart from detecting reversible perfusion defect from flow-limiting coronary stenosis, CMR late enhancement imaging LGE is currently the most sensitive method in detecting clinically unrecognized subendocardial infarction UMI from prior ischemic injury. We therefore tested the hypothesis that, characterization of these 2 processes from coronary artery disease CAD by CMR can provide complementary patient prognostic values. Methods and Results: We performed CMR on patients referred with symptoms suspicious of myocardial ischemia. The readers were blinded to any clinical outcome in either session.

At a median follow up of 15 months, 13 cardiac deaths and 26 nonfatal events occurred. Kaplan-Meier curves illustrating. Figures 1 and 2.

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Initiation of statin therapy halts progression of atherosclerotic plaque burden in peripheral arterial disease. Introduction: Studies suggest lipid lowering therapy improves symptoms and exercise performance in patients with peripheral arterial disease PAD ; however, the mechanism of action is unclear. I5 had their most symptomatic leg studied with MRI to assess atherosclerotic plaque burden before and I year after being started on lipid lowering therapy. Lipid measurements were obtained as part of the VAP test. CMR was performed using fat-suppressed multi-slice turbo-spin-echo pulse sequence on a Siemens Avanto I.

A custom-built flexible, linear four-element I0 cm x I0 cm square element surface coil array was placed over the SFA to image Icm along the vessel beginning. Right - same location imaged one year later. Notice plaque regression in the SFA. Blood was suppressed through the multislice data set using spatial presaturation, with periodic excitation of upstream slices. Imaging parameters included: repetition time II00 ms, echo time 7.

Changes in all parameters between groups from baseline to year one were compared by unpaired t-test. Changes in LDL were compared with changes in plaque parameters by linear regression. The final total cholesterol was similar between groups. No between group changes in HDL or triglycerides were seen. See Table I for changes in vessel wall parameters in the 2 groups.

No between group differences in lumen volume or change in lumen volume over time was noted. Conclusion: Statin-naive patients with PAD who were begun on either simvastatin or simvastatin plus ezetimibe for one year had a halting of progression of atherosclerotic plaque volume in the superficial femoral artery when compared to those already treated with statin given ezetimibe. Reverse vessel wall remodeling was noted among PAD patients newly treated with statins compared to those with ezetimibe added to preexisting statin therapy. Characterization of tissue heterogeneity by contrast-enhanced cardiovascular magnetic resonance imaging is a powerful predictor of ventricular tachyarrhythmias on ambulatory holter ECG in hypertrophic cardiomyopathy.

The presence of LGE identifies HCM patients at risk for ventricular tachyarrhythmias, which is an independent predictor of sudden death in this disease. In this study, we sought to determine whether regions of abnormal tissue at the confluence of both viable myocardium and fibrosis "border zone" BZ are more predictive of ventricular tachyarrhythmias than areas of myocardial scarring alone. LGE was determined by a blinded, independent reader using grayscale thresholds of both 4 and 6 standard deviations SD above the mean of normal, remote myocardium.

All subjects underwent h ambulatory Holter electrocardiogram ECG within 7. Conclusion: These data demonstrate that in HCM patients, the extent of BZ may be a superior predictor of ventricular tachyarrhythmias than myocardial scarring alone using a grayscale thresholding technique of 6 SD above normal myocardium. This study indicates the need for follow-up studies evaluating the independent prognostic value of BZ in risk stratification strategies in HCM patients. Background: Cardiac transplant patients experience significant morbidity related to transplant vasculopathy and acute transplant rejection, both of which can cause scarring of the myocardium.

Contrast enhanced cardiovascular magnetic resonance CMR has the unique ability to visualize and quantify myocardial scarring. It is well understood that myocardial infarctions resulting from transplant vasculopathy adversely affect prognosis and modify therapy.

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There is a growing body of evidence from non-transplant disease states, that the presence of non-infarct myocardial scar is also correlated to poor prognosis. Currently there is very little data on the scarring patterns present in the cardiac transplant population and it is our goal to better describe this pathology. Methods: Thirty-nine transplant patients underwent contrast enhancement imaging at the time of routine myocardial biopsy at two hospital centers in Alberta, Canada.

Standard phase sensitive inversion recovery sequences were used on commercially available scanners Siemens Avanto and Sonata, Siemens, Erlangen, Germany. DE had to be cross-referenced in two orthogonal views. Disagreements were settled by consensus. The scores of the 17 individual myocardial segments were added together to give an aggregate DE burden. Example of non-ischemic fibrosis. Subepicardial delayed enhancement of the anteroseptal and anterior walls a and inferior wall b.

Overall, patients with DE had scores ranging from I to I9, with a mean of 5. Non-ischemic DE was most commonly seen in the anterolateral and inferior walls Figure 3. There was no significant association between the presence of DE and time since transplant or current biopsy result. Conclusion: DE is a common feature in the transplant population.

Most DE observed is in a non-ischemic pattern;. The relationship between DE and cumulative episodes of rejection, hospitalization, and long term prognosis needs to be explored in more detail. Comparison of a rapid visual algorithm for quantification of infarct size with direct planimetry of infarct size by delayed enhancement-CMR.

Introduction: Direct planimetry of myocardial infarct size by delayed enhancement CMR DE-CMR has been well validated as a technique for quantification of infarct size with both a high degree of accuracy and reproducibility. Direct planimetry however requires extensive post processing and is time consuming and therefore not ideal for performance in a routine clinical service. We propose a visual algorithm to quantify total infarct size that would be rapid and easily incorporated into routine clinical practice.

In this study we sought to: I compare the level of agreement between our visual scoring algorithm and direct planimetry of infarct size; 2 compare the time required for each method of quantifying infarct size. The total volume of the HE zone was then divided by the total volume of myocardium within the left ventricle calculated by planimetry of endocardial and epicardial borders on all DE-CMR images so as to include both HE and non-HE myocardium.

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Visual scoring of all studies was performed using a I7 segment model with scores assigned based on the visual extent of HE myocardium in each segment i. Agreement between the visual algorithm and planimetry was compared using Bland-Altman analysis. The total infarct size was not statistically different between the visual algorithm I I. Bland Altman analysis revealed a nonsignificant bias of 2.

The visual algorithm required less time than direct planimetry 5. Conclusion: The visual algorithm provides a rapid method to quantify total infarct size with good agreement to direct planimetry. This is a useful time-efficient alternative for quantifying chronic infarct size in a routine clinical setting. The utility of cardiovascular magnetic resonance imaging in Takotsubo Cardiomyopathy apical ballooning for differential diagnosis, pathophysiological insights and additional findings.

Introduction: Takotsubo cardiomyopathy TTC is an increasingly recognized acute cardiac syndrome ACS , which underlying pathophysiological mechanisms are still controversially discussed. Cardiovascular magnetic resonance imaging CMR contributes to an understanding and differential diagnosis of this new entity by demonstrating the absence of irreversible injury delayed enhancement but oedema formation on T2-weighted images. However, clinical experience with CMR in this entity is still limited and is based mainly on relatively small case series.

Methods: Between January and October 81 consecutive patients, showing a left ventricular dysfunction with apical ballooning not explainable by the coronary artery status and initially admitted with ACS underwent CMR using a 1. Left ventricular function, T2-weighted spin echo sequence for oedema and delayed enhancement DE images after administration of Gadobutrol were assessed. Additionally, in the last 20 patients T2-weighted triple-inversion-recovery imaging to calculate the edema ratio ER and T1-weighted imaging before and after contrast agent administration to calculate the myocardial global relative enhancement gRE were performed for detection of inflammation.

Results: CMR revealed diagnosis of myocardial infarction in 18 In all other 54 Follow-up CMR after three months showed complete normalization of left ventricular function and inflammatory parameters in the absence of DE, oedema and PE. Conclusion: CMR has incremental value for differential diagnosis, pathophysiological insights and detection of additional findings in TTC. Therefore CMR should be performed in all patients with suspected TTC for further differential diagnosis and guidance of medical therapy.

Moreover, our results support the probable underlying cause of inflammation in TTC. If inflammation is the primary cause or secondary phenomena due to sympathetic overdrive needs further research. Potential of multidetector computed tomography and magnetic resonance imaging in quantifying left ventricular function, perfusion and viability of chronic microinfarction. Magnetic resonance imaging MRI has become the reference method for non-invasive quantification of: 1 left ventricular LV function using cine imaging and 2 myocardial perfusion deficits using first pass perfusion and 3 acute and chronic myocardial contiguous infarction using delayed contrast enhancement DE technique.

Modern multidetector computed tomography MDCT has also been recently used in assessing the above parameters in contiguous infarction. To our knowledge both modalities have not been tested in the assessment of micro-infarction caused by microembolic agents. The perfusion territory was selectively microembolized in six pigs using a microembolic agent mm, , Multi-slice MDCT top , MR delayed enhancement images middle and TTC slices bottom from a representative animal show good correspondence between modalities in defining patchy microinfarction embedded in viable myocardium indicated by arrows.

Histochemical staining TTC was used as golden standard for quantification of microinfarction. Results: The LAD-territory was MRI, but not MDCT, detected a perfusion deficit in the microembolized territory significant decrease in max upslope and maximum signal intensity. Microinfarction size did not differ between MDCT 6.

The results of this study suggest that MDCT and MRI can be used for detecting the consequences of microinfarction following coronary interventions and evaluating the efficacy of new therapies and devices designed to prevent microemboliza-tion. Objective: To determine the overlap between the regions of delayed mechanical activation and regions of myocardial scar tissue. Introduction: The greatest benefit from Cardiac Resynchro-nization Therapy CRT is likely achieved when the LV pacing lead is placed in the area with the greatest mechanical activation delay [I].

However, in patients with ischemic heart failure, the region with the greatest mechanical activation delay may correspond to an area of myocardial scar. Placing the pacing leads in areas of scar tissue will result in nonresponse to CRT[2]. The co-localization of areas of delayed mechanical activation and myocardial scar in patients with previous MI is not known. Methods: I6 patients with a prior history of myocardial infarction MI were studied six-months post infarct.

Cine SSFP images were obtained in the 2-chamber, 4-chamber, and short axis orientations. Following a double-dose injection 0. Mechanical activation delays were calculated from the cine SSFP images. Radial displacement curves showing movement of the endocardial border toward the LV center of mass were calculated for chords around each short axis slice. Displacement curves were averaged throughout the entire myocardium to generate a global displacement curve.

Cross correlation analysis between the global curve and the curve for each chord was done to determine the mechanical activation delay. A cross correlation delay of 3I msec was used as the threshold to determine mechanical activation delay[3]. Mechanical activation delay information for the entire myocardium was mapped onto the standardized AHA Bullseye model. Endocardial and epicardial borders were traced on the short axis DCE images. A separate region of interest was drawn around the area of infarct.

Each slice of the myocardium was resampled at chords around the heart, and each chord was assigned a value between 0 and I depending on the scar burden at that. The analysis was repeated for each short axis slice. On average, only I4. See Figure I. Conclusion: The region of greatest mechanical activation delay does not necessarily correspond to the region of myocardial scar tissue. Papillary muscle involvement in acute and chronic myocardial infarction: an MRI study using multi-contrast delayed enhancement pulse sequence.

Introduction: The papillary muscle PM is an integral component of the mitral valve apparatus. Acute or chronic myocardial infarction MI with PM ischemia is a primary factor leading to the occurrence of mitral regurgitation, with associated substantial morbidity and mortality [I, 2]. PM-MI is also a source of ventricular arrhythmia in these patients [3]. Hypothesis: We hypothesize that multi-contrast delayed enhancement MCDE imaging will improve the identification of.

All studies were performed on a I. The in-plane resolution was I. The contrast between blood pool and hyper-enhanced LV infarct was rated as excellent 3 , good 2 or fair I based on their differentiation. The multi-contrast capability of MCDE facilitates an improved differentiation between blood pool and infarct.

Moreover, the MCDE sequence provided cine images that also facilitated the simultaneous. PM involvement in a subject with RCA territorial infarction. Conclusion: MCDE imaging provides better contrast between blood pool and infarcted myocardium, thus improving the determination of PM-MI that may help identify patients in whom the significant mitral regurgitation may affect morbidity and mortality. Introduction: Myocardial salvage can be assessed retrospectively by T2-weighted and delayed enhancement images as shown in animal studies [1].

Currently there are only limited data in humans with acquisition of T2-weighted images not covering the full ventricle [2]. Purpose: Aim of this trial was to establish myocardial salvage imaging by MR as a surrogate endpoint in a randomized single-blinded trial and to show that high-dose N-ACC reduces reperfusion injury by its antioxidative properties. Secondary endpoints were infarct size and microvascular obstruction, ST-resolution at 90 minutes and occurrence of MACE at 30 day follow-up. Results: Due to contraindications MRI could not be performed in 31 patients.

All images were assessable for the calculation of the myocardial salvage index. The area at risk was The primary endpoint reperfusion injury measured by myocardial salvage index was not different between both treatment groups In addition, no differences in infarct size Conclusion: MRI can reliably measure the area at risk and infarct size retrospectively and served as a surrogate endpoint in this randomized clinical trial which showed that high-dose N-ACC does not provide an additional clinical benefit to placebo with respect to prevention of myocardial reperfusion injury and CIN and in patients undergoing primary PCI.

Aletras AH, Tilak GS and Natanzon A, et al: Retrospective determination of the area at risk for reperfused acute myocardial infarction with T2-weighted cardiac magnetic resonance imaging: Histopathological and displacement encoding with stimulated echoes DENSE functional validations. Circulation , J Am Coll Cardiol , Clinical, angiographic, and electrocardiographic predictors of infarct size and microvascular obstruction sssessed by MRI. The time-to-reperfusion, electrocardio-graphic and angiographic parameters are also of prognostic relevance in STEMI patients.

Predictors of IS and MO occurrence have not been assessed so far. Methods: This study analyzed consecutive STEMI patients reperfused by primary percutaneous coronary intervention within 12 hours after symptom onset. This may explain why these clinical, angiographic and electrocardiographic measures are associated with survival. In contrast to other studies the time-to-reperfu-sion did not affect IS and MO, which might be a selection bias, as patients with larger infarctions will be treated earlier.

Myocardium at risk and myocardial salvage after acute infarction in humans; quantification by magnetic resonance imaging. Introduction: To assess reperfusion therapy, it is necessary to determine how much myocardium is salvaged by measuring the final infarct size in relation to the initial myocardium at risk MaR of the left ventricle LV. T2 weighted MRI has recently been shown to be able to identify MaR but has not been validated against an independent method in humans.

MRI was performed on a I. At I week, patients were injected with a gadolinium-based contrast agent for quantification of infarct size on delayed contrast enhanced DE MRI. I week. Both modalities identified MaR in the same perfusion territory and in concordance with coronary angiography. The results demonstrate that T2-STIR performed up to one week after reperfusion can accurately determine myocardium at risk as it was before opening of the occluded artery.

The result of reperfusion therapy can therefore be assessed clinically by calculating myocardial salvage as the difference between myocardium at risk and final infarct size using MR imaging. Impact of myocardial hemorrhage on left ventricular function and remodeling in patients with reperfused acute myocardial infarction.

Background: Myocardial hemorrhage is a common complication following reperfusion of ST-segment-elevation acute myocardial infarction MI. Although its presence is clearly related to infarct size, at present it is unknown whether post-reperfusion hemorrhage affects left ventricular LV remodeling. Magnetic resonance imaging MRI can be used to identify myocardial infarction, myocardial hemorrhage and microvascular obstruction MVO , as well as measure LV volumes, function and mass.

Methods and results: Ninety-eight patients I4 females, 84 males, mean age: T2-weighted MRI was used to differentiate between hemorrhagic i. In the acute phase, presence of myocardial hemorrhage was related to larger LV end-diastolic and end-systolic volumes and infarct transmurality, lower LV ejection fraction as. Infarct size, size of area at risk and size of MVO were significantly larger in patients with hermorrhagic MI. At 4 months, a significant improvement in LV ejection fraction in patients with non-hemorrhagic MI was seen baseline: LV ejection fraction did, however, not improve in patients with hemorrhagic MI baseline: Multivariate analysis showed myocardial hemorrhage to be an independent predictor of adverse LV remodeling at 4 months defined as an increase in LV end-systolic volume.

This pattern was independent of initial infarct size See Table 1. Conclusion: Myocardial hemorrhage, the presence of which can easily be detected with T2-weighted MRI, is a frequent complication after successful myocardial reperfusion, and an independent predictor of adverse LV remodeling regardless of initial infarct size. Strain-encoded imaging for prediction of functional recovery in patients after acute myocardial infarction. Introduction: Evaluation of reversible dysfunction after acute myocardial infarction AMI has important therapeutic and prognostic implications.

The role of impaired systolic function for evaluation of functional recovery has been extensively investigated. However, whether impaired regional diastolic function after AMI also has predictive implications has not been investigated so far in humans using magnetic resonance imaging MRI. Colour coded SENC-image of a patient with transmural myocardial infarction with corresponding strain-curve. Purpose: To evaluate the predictive value of regional systolic and diastolic function for improvement of regional myocardial function in patients after AMI.

Methods: MRI 1. True cine sequences of 3 long-axis views 2-,3- and 4-chamber and a short-axis SA view covering the ventricle from apex to basis were acquired using a Steady State Free Precession SSFP sequence. After that, SENC cine images were acquired on the same long-axis planes to measure circumferential strains. Finally, using the same plane. Receiver-operating characteristic ROC curve demonstrates that diastolic strain rate assessed with Strain-Encoded Imaging is more sensitive than peak systolic strain for prediction of functional recovery diastolic strain rate AUC 0.

Peak systolic circumferential strain and early diastolic strain rate were measured at each segment in a modified 17 segment model. Regional wall motion was evaluated at baseline and at follow-up semi-quantitatively from the SSFP cine sequences by consensus reading of two blinded observers as normokinetic, hypokinetic or akinetic to evaluate functional recovery.

Results: segments were analyzed. I57 segments showed normal resting function and II9 segments showed wall motion abnormalities at baseline. Diastolic strain rate was more sensitive for prediction of functional recovery than peak systolic strain figure 2. Conclusion: SENC allows mechanical characterization of regional myocardial injury. Susceptibility artifact arrowhead.

B The hemorrhage corresponds to the area of MVO red contour shown on the early enhancement image. C Myocardial necrosis red contour , and residual MVO black core , is shown on the late enhancement image. We also hypothesized that myocardial hemorrhage would lead to an underestimate of the AAR on T2-weighted imaging using conventional signal threshold criteria. Left ventricular function was assessed with conventional cine sequences. Microvascular obstruction MVO and late enhancement were imaged at I minute and I5 minute delays respectively using a 3 dimensional inversion-recovery sequence.

This was compared to a conventional signal intensity threshold method using 2, 3 and 5 standard deviations sd above the mean of remote normal myocardium. A salvage index was calculated as the proportion of the AAR that did not show late enhancement. Introduction: Occlusion of a coronary artery leads to myocardial tissue edema in the vascular bed downstream of the vessel. The extent of hyperintense edema on T2-weighted images allows the area at risk AAR from ischemic injury to be retrospectively determined.

However, reperfusion of severely. Boundary detection identifies the myocardial edema green line which represents the AAR. The AAR determined at each signal threshold is shown in red. At low thresholds non-specific signal noise results in bright pixels in non-ischemic territories causing an overestimation of the AAR. At higher thresholds the signal from the myocardial edema is masked by the presence of hemorrhage in the core of the infarct arrow and leads to an underestimation of the AAR. Results: The mean area of hemorrhage was 5. Estimation of myocardial salvage at 3 sd and 5 sd signal thresholds becomes unreliable in hemorrhagic infarcts as the apparent AAR becomes smaller than the actual infarct size.

Hemorrhage is frequently observed and is associated with large infarcts where MVO is present and is an indicator of poor myocardial salvage. Hemorrhage in the core of the infarct causes signal loss on T2-weighted imaging and boundary-detection is required to reliably assess the AAR. Conclusion: Studies using CMR to determine the AAR and myocardial salvage should use boundary detection methods for quantification as arbitrary signal thresholds are unreliable when hemorrhage is present.

T2-weighted MRI pulse sequences for imaging post-infarct edema in mice: comparison of spin echo and T2 preparation approaches. Introduction: An ongoing diagnostic challenge exists in reliably differentiating non-salvageable, acutely infarcted myocardium from surrounding stunned, yet viable, myocardium that defines the area at risk. T2w cardiac magnetic resonance CMR imaging has previously been used to image the edema characterizing the area-at-risk region in post myocardial infarcted MI canine, porcine and human hearts.

However, the rapid murine heart rate presents challenges to T2w CMR application in mice. The typical T2w echo time of 40—60 ms needed for the detection of edema occupies a significant portion of the murine cardiac cycle I00— I20 ms with significant periods of blood flow and cardiac motion. Purpose: Develop an effective T2w CMR sequence for mice that exhibits high immunity to flow and tissue motion artifacts while maintaining sufficient and consistent signal-to-noise SNR and contrast-to-noise CNR performance.

The SE sequence employed a slice-selective excitation RF pulse and a thicker slice-selective refocusing RF pulse, and applied readout and phase encoding. Gd-enhanced mid-ventricular image of the mouse heart I day after MI, showing enhanced infarct region A with corresponding thresholded infarct region D. Same slice T2w spin echo image B with corresponding thresholded edema E.

Same slice T2w T2prep image C with corresponding thresholded edema F. Comparison of detected infarct and edema region areas over Days I through 4 post-MI for each imaging method. The T2prep sequence employed non-selective MLEV-weighted composite RF pulses followed by a standard slice-selective gradient-echo readout. Each sequence was applied on an isoflurane-anesthetized mouse on Days I through 4 after reperfused MI induced by 60 min coronary occlusion, as described previously.

All scans of each sequence were performed consecutively at four identical contiguous slice positions from mid-ventricular toward the apex. Results: Panels A through C Fig. Panels D through F show the respective hyperintense regions identified by threshold analysis performed after manual segmentation of the left ventricular LV myocardium. Panel G tracks and compares the detected infarct and edema region areas over. Days 1 through 4 post-MI for each imaging method. However, the SE sequence had a higher occurrence of flow and motion artifacts that degraded the consistency and accuracy of threshold selection.

Conclusion: As shown panel G , the T2prep yielded a consistently higher edema area percentage of Meanwhile, the SE gave a slightly lower mean with higher variance edema area percentage of To our knowledge, this is the first study to demonstrate the feasibility of performing T2w CMR edema imaging in mice, which opens a variety of potential basic research applications investigating the role of individual genes in acute and chronic settings post-MI. Image quality, spatial and temporal resolution are limited by the need to acquire multiple slices as single shot acquisitions within a single heart beat, as the process of interest is transient and very rapid, especially under pharmacologically induced stress hyperaemia.

This is why accelerated acquisition methods, such as k-t BLAST [1], could provide a significant improvement in the assessment of myocardial perfusion by CMR. An optimised DCE-MRI sequence with eight-fold k-t BLAST acceleration was shown to provide a significant improvement in spatial resolution without loss of image quality[2], making these datasets very suitable for parametric mapping. Objective: To investigate the ability of eight-fold k-t BLAST accelerated stress perfusion DCE-MRI combined with a rapid parametric mapping algorithm to detect regions of ischaemia in a pilot cohort of patients with suspected coronary heart disease.

Methods: The regional ethics review board's permission was obtained and ten patients 9 male were recruited into this study age range , mean An optimised DCE-MRI sequence Table 1 allowed three uniformly prepared slices to be acquired in every heart beat, for heart rates of up to bpm [2]. Examples of parametric maps of enhancement ratios in single-vessel disease corresponding to images 2, 4 and 7 in Figure 1. Dark coloured voxels belong to the lowest part of the individual study's frequency distribution.

The algorithm for quantitative analysis and parametric mapping comprised the following steps: 1 automated detection of the target post-contrast frame 2 endocardial border detection in the target post-contrast frame using automated region-growing algorithm and a single spline fitting to define epicardial border 3 automated registration of pre- and post-contrast frames using incremental rigid translation 4 computation of percentage enhancement ratios ER on voxel-by-voxel basis 5 histogram analysis of the ER datasets and creation of percentile-based colour maps.

Example parametric maps of ER are presented in Figure 2. Global enhancement ratios were computed as a median percentage change over baseline Table 2. Motion correction was not required in three datasets, and voxel displacement was applied in seven datasets. In patients with no significant coronary stenoses, the average ratio was In a patient with two-vessel disease ERwas Discussion: The results of this pilot study suggest that the proposed methods for acquisition and analysis of first-pass myocardial perfusion are robust and ready for use in clinical studies, where its diagnostic utility needs to be assessed formally in a larger patient cohort.

The methods allow the assessment of regional differences in perfusion, as well as global changes in perfusion. There is scope for further improvement, notably in increasing resistance to motion artefact and reducing signal inhomogeneity, which can lead to the appearance of false positive lesions. Objective: The purpose of this study was to evaluate the feasibility and diagnostic accuracy of high spatial resolution stress myocardial perfusion MRI acquired at every heartbeat by using k-t SENSE and 3 Tesla MR imager. Background: High spatial and temporal resolutions are required for the accurate assessment of myocardial ischemia using stress perfusion MRI.

Methods: Thirty-three patients with suspected coronary artery disease were studied. Three short-axis sections of the left ventricle were imaged at every heart beat. Two observers determined the image quality score I:poor - 4:excellent and recorded the presence or absence of respiratory artifacts and endocardial dark rim artifacts using a segment model.

Results: All studies were successfully completed, with the averaged image quality score of 3. Endocardial dark rim artifacts were observed in 17 3. Rest perfusion MRI is normal and no endocardial banding artifact is observed. Stress perfusion MRI in the same patient with triple vessel disease.

Subendocardial ischemia is clearly demonstrated in the anteroseptal well, lateral wall and inferior wall on high resolution images. Respiratory artifacts were found in 1 1 2.

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In 14 patients who underwent coronary angiography within 2 weeks from MR study, stress-rest perfusion MRI demonstrated the sensitivity, specificity, positive and negative predictive values and accuracy of were Figures I and 2. This approach can substantially reduce endocardial dark rim artifacts and allows for an accurate detection of myocardial ischemia in patients with flow-limiting coronary artery disease. Adenosine magnetic resonance imaging versus dobutamine stress echocardiography in patients with low probability for coronary artery disease. Introduction: Accurate assessment of patients with chest pain without electrocardiographic changes or elevation of serum cardiac enzymes is challenging.

There is increased interest in the role of dobutamine stress echocardiography DSE and adenosine magnetic resonance imaging AMRI performed in the chest pain unit as a diagnostic method to rule out Coronary Artery Disease CAD as the cause of the chest pain in this population. Methods: Inclusion criteria for the study were patients with normal EKG no signs of cardiac ischemia and negative cardiac enzymes, who were admitted to the Cardiac Decision Unit. The diagnostic method used was chosen randomly by physician preference. T-test was used to assess differences in continuous variables, and X2 square to test differences in categorical variables between the two groups.

Logistic regression was used to assess the likelihood of detecting CAD after adjusting for technique used and baseline characteristics. Mean age was similar among groups 52 for AMRI vs. Data in Table 1. Although selection bias could account for part of these results, a higher sensitivity for AMRI is suggested. Blood oxygen level-dependent MRI in patients with coronary artery disease and normal volunteers: a validation study against PET. Background: Elevated deoxyhaemoglobin seen downstream in a territory subtended by a stenotic coronary artery can be.

Remote: myocardial segments subtended by arteries with minimal or no CAD. Normal: myocardial segments in normal volunteers. Previous animal and human BOLD studies at 1. Using PET with oxygen labeled water, myocardial blood flow MBF was measured at baseline and during adenosine hyperemia. SI values were corrected for differences in TI-weighting owing to heart rate changes at stress and rest. Results: Based on the coronary anatomy, 59 myocardial segments were supplied by significantly stenosed vessels stenosed segments and 73 segments were supplied by vessels with minimal or no disease remote to ischemia segments.

An example of a patient with significant disease in the right coronary artery. A SI drop was noted in the inferior wall black arrow. All other myocardial segments showed a rise in SI during stress. Minor off-resonance artifacts were found in 9 subjects 7 CAD patients and 2 normal volunteers. Figure I shows an example of a patient with significant disease in the right coronary artery.

Introduction: Restoration of normal epicardial coronary flow in acute ST segment elevation myocardial infarction does not ensure adequate perfusion at the myocardial tissue level. In the era of primary PCI, patients with acute myocardial infarction treated successfully with PCI are discharged mostly within first week.

There are no studies available regarding blood flow at the tissue level in these patients during effort. CMR was performed on I. Microvascular obstruction regions MVO were assessed on early enhancement images acquired I minutes after stress perfusion. Transmurality of myocardial perfusion defecits at rest and in stress condition, MVO were evaluated using 5 point scale in I6 segments. DE was also evaluated in segment The sum of scores were calculated for each variable. Scar size and MVO were additionally quantitatively analyzed using. Results: Only in 2 patients there was no evidence of at least subendocardial stress perfusion deficit.

Median stress perfusion sum of scores was I5 points ranged 0 to Median rest perfusion scores was significantly lower 3 points; ranged Regression model revealed that only DE and stress perfusion sum of scores have significantly contributed to the LV EF model at discharge. Conclusion: Despite TIMI 3 flow in coronary artery myocardial perfusion defects at the tissue level are very frequent. Only in patients with preserved LV function pharmacological stress have not induced or intensed perfusion deficits. The pathogenesis remains unknown. Further studies are needed. Evaluation of contrast wash-in and peak enhancement in adenosine first pass perfusion in patients post bypass surgery.

Introduction: CMR adenosine first pass perfusion yields excellent results for the detection of significant coronary artery disease. In this case, adenosine perfusion would be an inadequate diagnostic test. Purpose: Aim of the study was to evaluate semiquantitative perfusion parameters in patients after CABG in order to evaluate. Methods: 32 patients post CABG were included into the study consisting of adenosine first pass 0. In invasive angiography, areas perfused by native coronaries and the different bypasses were identified.

Only segments without vessel stenosis and without LGE were used for final analysis. Results: Results are displayed in Table I. Conclusion: Semiquantitaive parameters of first pass adeno-sine perfusion are similar in areas supplied by native vessels or by different bypass grafts. These parameters are indirectly used for visual analysis speed of contrast wash-in and peak signal intensity. Therefore the possible different contrast kinetic through grafts and native vessel does not seem to be a limiting factor for the accuracy of first pass adenosine perfusion in patients post CABG.

In-room treadmill exercise stress cardiac magnetic resonance in patients with suspected ischemic heart disease. Objective: To implement and demonstrate the feasibility of in-room treadmill exercise stress perfusion and cine CMR in patients with suspected coronary artery disease CAD. Background: Exercise is preferred to pharmacologic stress because it links physical activity to symptoms and ischemia and offers important information such as exercise capacity, blood pressure response, ECG changes, and the presence or absence of exercise-induced symptoms. The predicted receptor designs were experimentally confirmed to be strong and specific in vitro sensors, as well as in vivo cell-based biosensors.

Protein receptors, such as the ones discussed above, are typically membrane-bound; they trigger protein signalling cascades that ultimately result in a cellular response. However, several synthetic methods can be used to transmit captured sensory information in a tunable and desirable manner. Skerker et al. Alternatively, engineered protein scaffolds can be designed to physically recruit pathway modulators and synthetically reshape the dynamical response behaviour of a system Bashor et al.

This constitutes a modular method for programming protein-based biosensors to have any desired response, including accelerated, delayed or ultrasensitive responses, to upstream signals. Hybrid approaches. Combining synthetic transcriptional, translational and post-translational circuits into hybrid solutions and harnessing desired characteristics from each could lead to the creation of cell-based biosensors that are as robust as those of natural organisms.

Using a synthetic hybrid approach, Voigt and colleagues Levskaya et al. A synthetic sensor kinase was engineered to allow cells to identify and report the presence of red light. Specifically, a membrane-bound photoreceptor from cyanobacteria was fused to an E. In a clever example of its use, the bacterial optical sensor was applied in image edge detection Tabor et al. In this case, by wiring the optical sensor to transcriptional circuits that perform cell—cell communication the quorum-sensing 16 system from Vibrio fischeri and logical functions Box A , the researchers programmed only the cells that receive light and directly neighbour cells that do not receive light to produce a pigment, allowing.

In bacteria, they consist of two domains: a membrane-bound histidine kinase sensitive element that senses a specific environmental stimulus, and a cognate response regulator transducer domain that triggers a cellular response. A basic biosensor has two modules see the figure : the sensitive element recognizes and binds analytes, whereas the transducer module transmits and reports signals.

Post-translational biosensors part a consist of membrane-bound protein receptors that trigger signal transduction cascades through signalling proteins, such as response regulators of two-component systems. The hybrid example part b shows a synthetic genetic edge detection circuit Tabor et al. The sensitive element is a light—dark sensor, Cph8, made as a chimaera of the photoreceptor domain of the cyanobacteria phytochrome Cph1 and the kinase domain of Escherichia coli EnvZ.

Specifically, the absence of light triggers Cph8 kinase activity, which correspondingly activates the ompC promoter. Cells not receiving light will therefore produce the cell—cell communication molecule 3-oxohexanoyl-homoserine lactone AHL; yellow circle through expression of its biosynthetic enzyme LuxI. In addition, these cells will produce the transcriptional repressor CI grey oval. This work demonstrates that complex behaviour can emerge from properly wiring together smaller genetic programs, and that these programs can lead to unique real-world applications.

Human health is afflicted by new and old foes, including emergent drug-resistant microbes, cancer and obesity. Meanwhile, progress in medicine is faced with challenges at each stage of the therapeutic spectrum, ranging from the drying up of pharmaceutical pipelines to limited global access to viable medicines. In a relatively short amount of time, synthetic biology has made promising strides in reshaping and streamlining this spectrum Box A Indeed, the rational and model-guided construction of biological parts is enabling new therapeutic platforms, from the identification of disease mechanisms and drug targets to the production and delivery of small molecules.

Disease mechanism. This allows for the rigorous testing of submodules in an isolated, well-characterized environment. Similarly, synthetic biology provides a framework for synthetically reconstructing natural biological systems to explore how pathological behaviours may emerge. This strategy was used to give mechanistic insights into a primary immunodeficiency, agammaglobulinaemia, in which patients cannot generate mature B cells and as a result are unable to properly fight infections Ferrari et al. The researchers developed a synthetic testbed by systematically reconstructing the various components of the human B cell antigen receptor BCr signalling pathway in an orthogonal environment.

Pathogenic viral genomes can similarly be reconstructed for studying the molecular underpinnings of infectious disease pandemics. For instance, synthetic reconstruction of the severe acquired respiratory syndrome SARS coronavirus Becker et al. Drug-target identification. Building up synthetic pathways and systems from individual parts is one way of identifying disease mechanisms and therapeutic. Orthogonal hosts are often organisms with sufficient evolutionary distance from the native host.

Another is to deploy synthetic biology devices to systematically probe the function of individual components of a natural pathway. Our group, for instance, has engineered modular riboregulators that can be used to tune the expression of a toxic protein or any gene in a biological network Isaacs et al. Translational repression of this cis -repressed mRNA can be alleviated by an independently regulated transactivating RNA that targets the stem-loop for unfolding. Engineered riboregulators have been used to tightly regulate the expression of CcdB, a toxic bacterial protein that inhibits DNA gyrase, 18 to gain a better understanding of the sequence of events leading to induced bacterial cell death Dwyer et al.

These synthetic biology studies, in conjunction with systems biology studies of quinolones antibiotics that inhibit gyrase Dwyer et al. This work provided new insights into how bacteria respond to lethal stimuli and paved the way for the development of more effective antibacterial therapies. Drug discovery. After a faulty pathway component or target is identified, whole-cell screening assays can be designed using synthetic biology strategies for drug discovery.

As a demonstration of this approach, Fussenegger and colleagues Weber et al. However, due to transcriptional repression of ethA by the protein EthR, ethionamide-based therapy is often rendered ineffective. To address this problem, the researchers designed a synthetic mammalian gene circuit that featured an EthR-based transactivator of a reporter gene and used it to screen for and identify EthR inhibitors that could abrogate resistance to ethionamide.

Importantly, because the system is a cell-based assay, it intrinsically enriches for inhibitors that are non-toxic and membrane-permeable to mammalian cells, which are key drug criteria as M. This framework, in which drug discovery is applied to whole cells that have been engineered with circuits that highlight a pathogenic mechanism, could be extended to other diseases and phenotypes.

Control of chromosomal topological transitions is essential for DNA replication and transcription in bacteria, making gyrase an effective target for antimicrobial agents for example, the quinolone class of antibiotics. Part a of the figure shows a synthetic mammalian gene circuit that enabled drug discovery for antituberculosis compounds Weber et al. The antibiotic ethionamide is rendered cytotoxic to Mycobacterium tuberculosis by the enzyme EthA in infected cells. Because EthA is natively repressed by EthR, resistance to ethionamide treatment is common.

In the gene circuit, a fusion of EthR and the mammalian transactivator VP16 binds a minimal promoter P min with a synthetic EthR operator site and activates expression of the reporter gene SEAP human placental secreted alkaline phosphatase. This platform allows for the rapid screening of EthR inhibitors in mammalian cells. Part b shows a synthetic mammalian genetic switch for tight, tunable and reversible control of a desired gene for therapeutic or gene-delivery applications.

In the OFF configuration upper panel , expression of the gene of interest green is repressed at the levels of both transcription and translation. Constitutively expressed LacI repressor red binds to the lac operator sites in the transgene module of the gene of interest, therefore repressing its transcription. The discovery of drugs does not always translate to the people who need them the most because drug production processes can be difficult and costly. Antibiotics are industrially produced from microbes and fungi, and are therefore widespread and cheap.

Conversely, many other drugs are isolated from hosts that are not as amenable to large-scale production and are therefore costly and in short supply. Such drugs include the antimalaria drug artemisinin and the anticancer drug taxol. Fortunately, global access to drugs is being enabled by hybrid synthetic biology and metabolic engineering strategies for the microbial production of rare natural products. In the case of artemisinin part c , there exist two biosynthetic pathways for the synthesis of the universal precursors to all isoprenoids, the large and diverse family of natural products of which artemisinin is a member.

The native isoprenoid pathway found in Escherichia coli the deoxyxylulose 5-phosphate DXP pathway has been difficult to optimize, so instead researchers have synthetically constructed and tested the entire Saccharomyces cerevisiae mevalonate-dependent MEV pathway in E. The researchers initially used E.

They then linked the optimized heterologous pathway to a codon-optimized form of the plant terpene synthase ADS to funnel metabolic production to the specific terpene precursor to artemisinin. This work allowed them to build a full, optimized solution that could be ultimately and seamlessly deployed back into S.

FPP, farnesyl pyrophosphate. Therapeutic treatment. Synthetic biology devices have additionally been developed to serve as therapies themselves. Entire engineered viruses and organisms can be programmed to target specific pathogenic agents and pathological mechanisms. For instance, in two separate studies Lu and Collins, , researchers used engineered bacteriophages to combat antibiotic-resistant bacteria by endowing them with genetic mechanisms that target and thwart bacterial mechanisms for evading antibiotic action.

The first study was prompted by the observation that biofilms, 19 in which bacteria are encapsulated in an extracellular matrix, have inherent resistance to antimicrobial therapies and are sources of persistent infections. To more effectively penetrate this protective environment, T7 phage was engineered to express the biofilm matrix-degrading enzyme dispersin B DspB upon infection Lu and Collins, The two-pronged attack of T7 expressing DspB and phage-induced lysis fuelling the creation and spread of DspB resulted in the removal of In the second study Lu and Collins, , it was suggested that inhibition of certain bacterial genetic programs could improve the effectiveness of current antibiotic therapies.

In this case, bacteriophages were deliberately designed to be non-lethal so as not to elicit resistance mechanisms; instead, a non-lytic M13 phage was used to suppress the bacterial SOS DNA-damage response by overexpression of its repressor, lexA3. The engineered bacteriophage significantly enhanced killing by three major classes of antibiotics in traditional cell culture and in E. Synthetically engineered viruses and organisms that are able to sense and link their therapeutic activity to pathological cues may be useful in the treatment of cancer, in which current therapies often indiscriminately attack tumours and normal tissues.

For instance, adenoviruses were programmed to couple their replication to the state of the p53 pathway in human cells Ramachandra et al. Normal p53 production would result in inhibition of a crucial viral replication component, whereas a defunct p53 pathway, which is characteristic of tumour cells, would allow viral replication and cell killing. In another demonstration of translational synthetic biology applied to cancer therapy, Voigt and colleagues Anderson et al.

Constitutive expression of the heterologous invasin inv gene from Yersinia pseudotuberculosis can induce E. So, to preferentially invade cancer cells, the researchers placed inv under the control of transcriptional operons that are activated by environmental signals specific to the tumour microenvironment. These engineered bacteria could be made to carry or synthesize cancer therapies for the treatment of tumours. Biofilms are an antibiotic-resistant mode of microbial life found in natural and industrial settings. Therapeutic delivery. In addition to engineered therapeutic organisms, synthetic circuits and pathways can be used for the controlled delivery of drugs as well as for gene and metabolic therapy.

In some cases, sophisticated kinetic control over drug release in the body may yield therapeutic advantages and reduce undesired side effects. Most hormones in the body are released in time-dependent pulses. Glucocorticoid secretion, for instance, has a circadian and ultradian 20 pattern of release, with important transcriptional consequences for glucocorticoid-responsive cells Stavreva et al.

Faithfully mimicking these patterns in the administration of synthetic hormones to patients with glucocorticoid-responsive diseases, such as rheumatoid arthritis, may decrease known side effects and improve therapeutic response Stavreva et al, Periodic synthesis and release of biologic drugs can be autonomously achieved with synthetic oscillator circuits Elowitz and Leibler, ; Atkinson et al. In other cases, one may wish to place a limit on the amount of drug released by programming the synthetic system to self-destruct after a defined number of cell cycles or drug release pulses.

Our group has recently developed two variants of a synthetic gene counter Friedland et al. Gene therapy is beginning to make some promising advances in clinical areas in which traditional drug therapy is ineffective, such as in the treatment of many hereditary and metabolic diseases. Synthetic circuits offer a more controlled approach to gene therapy, such as the ability to dynamically silence, activate and tune the expression of desired genes.

In one such example Deans et al.

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Additionally, the construction of non-native pathways offers a unique and versatile approach to gene therapy, such as for the treatment of metabolic disorders. Operating at the interface of synthetic biology and metabolic engineering, Liao and colleagues Dean et al. Remarkably, the researchers found that when transplanted into mammals, the shunt actually increased fatty acid oxidation, evidently by creating an alternative cycle. Furthermore, mice expressing the shunt showed resistance to diet-induced obesity when.

This pathway diverts the tricarboxylic acid TCA cycle so that fatty acids are not completely oxidized and are instead converted into carbon energy sources. This work offers a new synthetic biology model for studying metabolic networks and disorders, and for developing treatments for the increasing problem of obesity. Finally, the discovery of drugs and effective treatments may not quickly — or ever — translate to the people who need them the most because drug production processes can be difficult and costly.

As discussed below, synthetic biology is allowing rare and costly drugs to be manufactured more cost-effectively Box A Recent excitement surrounding the production of biofuels, pharmaceuticals and biomaterials from engineered microorganisms is matched by the challenges that loom in bringing these technologies to production scale and quality. The most widely used biofuel is ethanol produced from corn or sugar cane Fortman et al.

Microorganisms engineered with optimized biosynthetic pathways to efficiently convert biomass into biofuels are an alternative and promising source of renewable energy. These strategies will succeed only if their production costs can be made to compete with, or even outcompete, current fuel production costs.

Similarly, there are many drugs for which expensive production processes preclude their capacity for a wider therapeutic reach. New synthetic biology tools would also greatly advance the microbial production of biomaterials and the development of novel materials. Constructing biosynthetic pathways. When engineering for biofuels, drugs or biomaterials, two of the first design decisions are choosing which biosynthetic pathway or pathways to focus on and which host organism to use.

Typically, these decisions begin with the search for organisms that are innately capable of achieving some desired biosynthetic activity or phenotype Alper and Stephanopoulos, For biofuel production, for instance, certain microorganisms have evolved to be proficient in converting lignocellulosic material to ethanol, biobutanol and other biofuels. These native isolates possess unique catabolic activity, heightened tolerances for toxic materials and a host of enzymes designed to break down the lignocellulosic components.

The example of the microbial production of biobutanol, a higher energy density alternative to ethanol, provides a useful glimpse into these design trade-offs. However, it is produced in low yields and as a mixture with acetone and ethanol, so substantial cellular engineering of a microorganism for which standard molecular biology techniques do not apply is needed to produce usable amounts of butanol Tummala et al. Furthermore, importing the biosynthetic genes into an industrial microbial host can lead to metabolic imbalances Inui et al.

In an altogether different approach, Liao and colleagues Atsumi et al. Indeed, complementary to efforts in traditional metabolic and genetic engineering is the use of engineering principles for constructing functional, predictable and non-native biological pathways de novo to control and improve microbial production. In an exemplary illustration of this, Keasling and colleagues engineered the microbial production of precursors to the antimalarial drug artemisinin to industrial levels Martin et al.

There are now many such examples of the successful application of synthetic approaches to biosynthetic pathway construction — such approaches have been used in the microbial production of fatty-acid-derived fuels and chemicals such as fatty esters, fatty alcohols and waxes Steen et al. Optimizing pathway flux.

After biosynthetic pathways have been constructed, the expression levels of all of the components need to be orchestrated to optimize metabolic flux 22 and achieve high product titres. A standard approach is to drive the expression of pathway components with strong and exogenously tunable promoters, such as the P L tet, P L lac, and P BAD promoters from the tet , lac and ara operons of E. To this end, there are ongoing synthetic biology efforts to create and characterize more reusable, biological control elements based on promoters for predictably tuning expression levels Alpert et al.

Further to this, synthetic biologists have devised a number of alternative methods for obtaining biological pathway balance, ranging from reconfiguring network connectivity to fine-tuning individual components. A richer. The rate is regulated by the enzymes in the pathway. In Box A and Box A we detail several synthetic biology strategies that specifically pertain to the optimization of metabolic pathway flux. These strategies range from those driven by evolutionary techniques, to those driven by rational design and in silico models, to those that combine both approaches.

Programming novel functionality and materials. Beyond facilitating metabolic tasks, synthetic systems can infuse novel functionality into engineered organisms for production purposes or for building new materials. Early work in the field laid the groundwork for constructing basic circuits that could sense and process signals, perform logic operations and actuate biological responses Voigt, Wiring these modules together to bring about reliable, higher-order functionality is one of the next major goals of synthetic biology Lu et al.

For instance, circuits designed to sense the bioreactor environment and shift metabolic phases accordingly would further improve biofuel production. Alternatively, autonomous timing circuits could be used to shut down metabolic processes after a prescribed duration of time.

Biological timers of this sort have been developed using genetic toggle switches that were deliberately rendered imbalanced through model-guided promoter engineering Ellis et al. These genetic timers were used to program the time-dependent flocculation 23 of yeast cells to facilitate the separation of cells from, for instance, the alcohol produced in industrial fermentation processes.

Synthetic control systems can also be used to extract and purify the synthesized product. This is particularly important in the production of recombinant proteins, bioplastics and other large biomaterials, which can accumulate inside cells, cause the formation of inclusion bodies and become toxic to cells if they are present at high titres. To export recombinant spider-silk monomers, Widmaier et al. The Salmonella type III secretion system T3SS not only fulfils these criteria but also possesses a natural regulatory scheme that ties expression of the protein to be secreted to the secretion capacity of the cell; as a result, the desired protein is only expressed when sufficient secretion complexes have been built.

To obtain superior secretion rates of recombinant silk protein, the researchers needed only to engineer a control circuit that hitches the heterologous silk-protein-producing genes to the innate genetic machinery for environmental sensing and secretion commitment. For example, flocculation occurs once the sugar in a beer brew has been fermented into ethanol. Finally, there is an emerging branch of synthetic biology that seeks to program coordinated behaviour in populations of cells, which could lead to the fabrication of novel biomaterials for various applications.

The engineering of synthetic multicellular systems is typically achieved with cell—cell communication and associated intracellular signal processing modules, as was elegantly used by Hasty and colleagues Danino et al. Weiss and colleagues Basu et al. These systems, which can be programmed to form intricate multicellular patterns from a solid-phase cellular lawn, would aid the development of fabrication-free scaffolds for tissue engineering.

The future of translational synthetic biology hinges on the development of reliable means for connecting smaller functional circuits to realize higher-order networks with predictable behaviours. In a previous article Lu et al. Beyond the challenge of improving the design cycle, applied synthetic biology would benefit from once again summoning the original inspiration of bio-computing. The ability to program higher-level decision-making into synthetic networks would yield more robust and dynamic organisms, including ones that can accomplish many tasks simultaneously.

Furthermore, as adaptive and predictive behaviours are naturally present in all organisms including microbes Mitchell et al. Finally, the majority of synthetic biology is currently practiced in microbes. However, many of the most pressing problems, and in particular those of human health, are inherently problems with mammalian systems. Therefore, a more concerted effort towards advancing mammalian synthetic biology will be crucial for next-generation therapeutic solutions, including the engineering of synthetic gene networks for stem-cell generation and differentiation.

By addressing such challenges, we will be limited not by the technicalities of construction or the robustness of synthetic gene networks but only by the imagination of researchers and the number of societal problems and applications that synthetic biology can resolve. In the production of artemisinin precursors, the native Escherichia coli isoprenoid pathway the deoxyxylulose 5-phosphate DXP pathway was eschewed in favour of a heterologous pathway so as to circumvent the complex regulatory control imposed by the host Box A In an alternative method of relieving regulatory control over the large number of DXP pathway components, Wang et al.

The researchers developed a rapid, automated method for the in vivo directed evolution of pathways, which they termed multiplex automated genome engineering MAGE. They then applied it to evolve the translational efficiencies of DXP pathway components to achieve maximal lycopene production. Specifically, cells were subjected to cycles of genetic modifications through oligo-mediated allelic replacement in an automated fashion to explore sufficient genomic diversity for optimizing biosynthetic pathways at laboratory timescales. At the other end of the spectrum are strategies that rely on quantitative models and blueprints for the rational design of optimized networks and pathways part b.

Finally, the optimized parameter set will be used to forward-engineer new networks and components. For example, stochastic biochemical models have been developed to capture the expression dynamics of synthetically engineered promoters; these models were subsequently used to predict the correct in vivo behaviour of different and more complex gene networks built from the modelled components Blake et al.

Similarly, at the level of translation, thermodynamic models that predict the relative translation initiation rates of proteins can be used to rationally forward-engineer synthetic RBS sequences to give desired expression levels Salis et al. Such techniques harness modelled genetic parameters transcriptional or translational to predict the level of expression of proteins and enzymes in a network. In a hybrid rational—combinatorial approach, Dueber et al. To construct the enzyme scaffolding, the researchers tethered protein—protein interaction domains for example, GBD, SH3 and PDZ domains from metazoan signalling proteins.

By varying the number of repeats of an interaction domain, the researchers could additionally control the stoichiometry of the enzymes recruited to the complex. Using the heterologous mevalonate-dependent MEV pathway in E. Finally, they showed that the optimized synthetic scaffold could substantially increase product titre while reducing the metabolic load on the host; in other words, their high product titres did not require the overexpression of biosynthetic enzymes in the cell. We thank members of the Collins laboratory for helpful discussions and K.

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Control of gene expression by a natural metabolite-responsive ribozyme. Regulation of bacterial gene expression by riboswitches. Yen, L. Exogenous control of mammalian gene expression through modulation of RNA self-cleavage. You, L. Programmed population control by cell—cell communication and regulated killing. Andrew D. Ellington and Jared Ellefson Recent years have marked a dramatic increase of our capabilities to sequence and synthesize nucleic acids. Ten years ago the first human genome was sequenced, and at the time it was a monumental undertaking requiring billions of dollars in funding and legions of dedicated researchers.

Today, the same genome sequence would cost a fraction of the price and soon personal genome sequencing will be commonplace. However, while our understanding of genomes has not yet. The same revolution is occurring in DNA synthesis, as prices for DNA drop and technologies are developed for large-scale and ever longer syntheses. As with sequencing, the technology revolution will lead understanding, and it can be argued that, although we can synthesize a microbial genome Gibson et al. Together, these emerging issues are the essence of what can be seen as the holy grail of both systems and synthetic biology, the genome as the unit of engineering.

Synthetic biology can, in many circumstances, be defined as the abstraction of biology to the point where it can be easily engineered. This is first and foremost an operational definition, not an intellectual one. Indeed, there hides in the background of synthetic biology the usually unstated hypothesis that biology was made to be engineered by engineers a hypothesis we take issue with below. Parallels are often drawn between electrical engineering and synthetic biology, where genetic information like promoters and proteins can be compared to components of a circuit board such as transistors or capacitors.

In the view of the new breed of synthetic biologists, standardized components can almost always be used to rationally design genetic elements to perform desired tasks. This view has much merit, since it has already yielded interesting products such as biofuels, pharmaceuticals, and biomaterials. For example, Bayer and Voigt coaxed yeast into making valuable methyl halides from biomass Bayer et al. It is not unreasonable to suspect that synthetic circuits will be introduced in human hosts at some point; for example, Fussenegger has created a synthetic circuit composed of a bacterial uric acid sensor and a fungal urate oxidase which converts uric acid into a more tolerable compound that can be used to control uric acid levels in mammalian hosts and thus ameliorate chronic disease states such as gout Kemmer et al.

However, it is nonetheless still the case that many of these systems must be optimized in order for them to function as intended, in large part because the constituent parts are neither truly modular nor is their function fully predictable in new contexts. The question thus becomes whether biology is really meant to be engineered the same way a circuit board is, whether engineers will learn to make biology a circuit board, or whether some composite view is more akin to reality.

Or, stated another way, the question is whether genetic tinkering which existed well before synthetic biology has somehow entered a new, more robust phase or if it has just been relabeled. Systems biology has taken a different approach to how to tinker with systems. This approach proceeds from an understanding of the system as a whole instead of as an amalgam of component parts. It is top down rather than bottom up. By integrating literally millions of points of data from genomes, transcriptomes, and proteomes across the phylogenetic spectra, systems biologists can draw remarkable conclusions, up to and including the identification of nonobvious and evolutionarily repurposed subsystems.

It is this sort of understanding that allows us to realize that the same interactions that govern resistance to antifungals in yeast also govern blood vessel formation in higher organisms McGary et al. While a comparable synthetic biology approach might be to repurpose a tractable signaling pathway for blood vessel formation, this approach would require extensive empirical testing.

As technology continues to develop for high-throughput analysis of DNA, RNA, and proteins, systems biologists will have the benefit of several billion years of empirical testing and, thus, will hold the intellectual high ground for understanding how organisms truly work. Quantitative modeling of the connectedness of extant systems will, in the end, be more likely to allow us to build a functional genome from scratch than the untested engineering hypothesis that organisms should work like we want them to.

In greater detail, this should inevitably lead to the following discussion. In fact, long before iGEM there were numerous biological engineers, and these biological engineers would, with different degrees of success, use genes not then known as biological parts to construct pathways not then known as circuits that had particular functions. It could be argued that there are two important milestones: First, many researchers in other engineering disciplines were somehow shut out of the biology because it lacked an engineering flair and was more dissective than synthetic.

In this view, the influx of engineers into biology is assisted by recasting biology in terms more familiar to engineers, and this is why we often see biological circuits represented incorrectly, as I will argue later as electronic circuits Khalil and Collins, Second, there is not just a quantitative but rather a qualitative or foundational difference in being able to make large amounts of DNA. The ability to remake a whole genome Gibson et al. I think there is some merit to this explanation, although it is a bit like saying that genomics is different than genetics.

Genomics further explains genetics; it does not remake it although individual concepts in genetics, such as the role of environment in inheritance, have certainly been remade radically. Nonetheless, it is true that techniques for the rational manipulation of whole genomes will prove to be of enormous value into the future. Where would they have turned for knowledge and inspiration? I contend that it would not have been electrical engineering—the frequent muse to whom many who call themselves synthetic biologists appeal—but rather to systems biology.

Systems biology attempts to understand the interrelationships between all of the molecular and cellular parts of system in a quantitative way, and it has as one of its ultimate goals the modeling of biological organisms down to the molecular level. Fortunately for both traditional biological engineers and synthetic biologists, our understanding of systems has increased dramatically in recent years, and thus there will come a point where systems biology will not only completely inform biological engineering, but it will overtake the utility of orthogonal, add-on circuits that were largely meant to operate independently of the systems in which they were embedded.

Systems biology will eat synthetic biology. This change in perspective and approach will be absolutely essential as biological engineering moves forward. The utility of orthogonal synthetic circuits is limited for a variety of reasons. First, by failing to take into account the unity of the system, the predictability of circuits must be limited. This can best be seen by thinking about a very simple synthetic circuit that has been around for.

You can in fact embed a plasmid in a strain and induce protein production. From this vantage, the synthetic circuit has worked. However, it is difficult if not impossible to predict protein yield at the outset with any degree of certainty. Different proteins will express to different extents, will form aggregates to different extents, and will hamper the growth of the organism to different extents, to mention just a few variables that ultimately affect yield. Outcomes will range from making huge amounts of a protein to killing the cell outright.

Of course the circuit can always be tinkered with to make more protein, but that was true before it was called a circuit and was merely a plasmid. What has synthetic biology brought to the modular, composable, scalable, and programmable overexpression of proteins that was not previously known? Nothing, because the tinkering that is now possible following the invention of the discipline is the same tinkering that was possible prior to its invention. Collins has pointed out that really tinkering is all that is necessary for engineering to prosper in biology, and he is of course correct.

However, understanding and progress in this area will come from an increasingly detailed understanding of systems biology and integrated models of metabolism. In discussions with Erik Winfree, a more charitable interpretation of the coexplosion of both systems and synthetic biology emerges, which is that these are different means to the same end—a predictable biology. The more complex system, the organism, is at present less predictable, and thus nominally orthogonal subsystems as built by synthetic biologists may allow us to initially develop better models.