妊娠相关血浆蛋白A的原核表达及蛋白纯化

目的构建原核表达PAPP-A重组蛋白,并对蛋白质进行纯化。方法根据DNAstar软件分析获得的编码PAPP-A特异抗原表位,利用Primer Premier 5.0软件设计引物,以PAPP-A c DNA-质粒转化菌液为模板,进行PCR扩增。PAPP-A的DNA和p ET42a质粒分别双酶切,经琼脂糖凝胶电泳及胶回收后进行连接反应,转化至大肠埃希菌Top10,筛选阳性克隆经测序鉴定正确后,转化至大肠埃希菌BL21,诱导产生PAPP-A重组蛋白,利用镍柱初步纯化及离子交换层析柱进一步纯化,用SDS-PAGE及DEAE层析鉴定重组抗原蛋白。结果筛选出抗原表位集中区,在大肠埃希菌BL21中高效表达了重组的PAPP-A蛋白,经过镍柱及离子交换层析纯化后,PAPP-A浓度为0.22 g/L,DEAE层析分析证实其纯度较高。结论成功筛选出PAPP-A的抗原表位集中区,并制备了重组抗原蛋白,为PAPP-A的后续临床研究提供依据。     

Clinical experience with electroanatomic mapping of ectopic atrial tachycardia

The aim of this study was to evaluate the clinical use of a new three-dimensional mapping system as a guide for catheter ablation of ectopic atrial tachycardia. A series of 42 consecutive patients with drug refractory ectopic atrial tachycardia was studied in a prospective observational trial with the electroanatomic mapping system CARTO. The arrhythmogenic focus was found in the right atrium in 30 patients and in the left atrium in 12 patients. The construction of a complete electroanatomic map of the right or left atrium was possible in 37 of 42 consecutive patients with ectopic atrial tachycardia. Mean activation time of the right atrium, including the proximal coronary sinus, was 94 +/- 25 ms for right atrial tachycardias; left atrial activation time during left atrial tachycardias was 86 +/- 17 ms. Average mapping time was 30 minutes for right atrial tachycardias and 22 minutes for left atrial tachycardias, allowing the collection of 86 +/- 50 and 65 +/- 28 catheter positions, respectively. The size of the area of earliest atrial activation calculated from the electroanatomic map amounted to 0.6 +/- 0.4 cm2 in right atrial tachycardias and 1.0 +/- 0.9 cm2 in left atrial tachycardias. In the right atrium the most common locations of the 33 arrhythmogenic foci in 30 patients were the high or mid-lateral right atrium (n = 10) and the inferoparaseptal region near the coronary sinus ostium (n = 7). Ectopic left atrial foci were most commonly located in an inferior position near the mitral annulus (n = 5) and in proximity to the ostium of the pulmonary veins (n = 4). Biatrial electroanatomic mapping allowed visualization of earliest right atrial activation during left atrial tachycardia at the high interatrial septum or near the coronary sinus ostium. Catheter ablation was successful in 85% of right atrial tachycardias and 82% of left atrial tachycardias. In patients with ectopic atrial tachycardia electroanatomic mapping is a safe and feasible technique that allows three-dimensional visualization of the automatic focus in a precise anatomic reconstruction of the atria. This novel mapping technology facilitates catheter ablation of complex ectopic atrial tachycardia.     

Arrhythmia vulnerability assessment using magnetic field maps and body surface potential maps

Magnetic field maps and body surface potential maps can be used to measure cardiac activity. The ability of magnetic and potential body surface maps to identify patients' vulnerable to recurrent sustained ventricular arrhythmia (VA) were compared. Magnetic field maps (MFM) and body surface potential mapping (BSPM) were obtained from 76 normal (N) subjects, 15 myocardial infarct (MI) patients, and 15 VA patients. QRST integral maps were calculated for each subject and nondipolar content was determined using Karhunen-Loeve transform eigen-maps. Although differences in nondipolar content were significant between the normal and patient groups (P = 2.4 x 10(-5) for BSPM and P = 6.0 x 10(-8) for MFM), differences in nondipolar content between MI and VA patients using QRST integral BSPM and MFM maps were not significant. The trajectory of the location of the maxima and minima on the map area during the QRS and ST-T intervals were also constructed. Discrimination between MI and VA patients was based on intergroup differences in the amount of fragmentation of the trajectory plots. The ST-T trajectory plots were significantly more fragmented (P < 0.0001 and P < 0.05 for MFM and BSPM, respectively) for VA than for MI patients. The ST-T interval MFM and BSPM trajectory plots enabled separation of MI and VA patients with accuracies of 83% and 73%, respectively. These results suggest that repolarization MFM and BSPM extrema trajectory plots can be used effectively as a means of identifying patients at risk for VA.     

Wide QRS Complex Arrhythmia with Alternating QRS Morphology: What Is the Mechanism?

A 75‐year‐old man was admitted due to an electrical storm with appropriate recurrent implantable cardioverter defibrillator (ICD) discharges. The patient had had an extensive anterolateral myocardial infarction with associated severe left ventricular dysfunction 10 years earlier (left ventricular ejection fraction, 25%), and an ICD was placed 9 years before admission for primary prevention of sudden cardiac death. A first invasive study induced up to five ventricular tachycardias and an extensive endocardial substrate ablation was performed. Despite intravenous β‐blockers, general anesthesia and procainamide infusion, the patient continued to have recurrent episodes of very slow sustained ventricular tachycardia with a right bundle branch block pattern. In a subsequent invasive study, no mid‐diastolic activity was found despite careful mapping during the induced clinical ventricular tachycardia and ablation attempts inside the apical endocardial scar were unsuccessful. A percutaneous epicardial approach with navigation system support (EnSite Precision^TM Cardiac Mapping System v. 2.0, St. Jude Medical, St. Paul, MN, USA) without antiarrhythmic infusion was planned. A wide QRS complex rhythm with alternating QRS morphology was readily induced by epicardial ventricular pacing trains (Fig. 1, top) that elicited both arrhythmia QRS patterns with very long stimulus QRS intervals (Fig. 1, bottom). What is the possible mechanism of this arrhythmia? Do we need further pacing maneuvers during the arrhythmia to localize critical sites at which ablation pulses can predictably be successful?     

Deep weakly-supervised breast tumor segmentation in ultrasound images with explicit anatomical constraints

Breast tumor segmentation is an important step in the diagnostic procedure of physicians and computer-aided diagnosis systems. We propose a two-step deep learning framework for breast tumor segmentation in breast ultrasound (BUS) images which requires only a few manual labels. The first step is breast anatomy decomposition handled by a semi-supervised semantic segmentation technique. The input BUS image is decomposed into four breast anatomical structures, namely fat, mammary gland, muscle and thorax layers. Fat and mammary gland layers are used as constrained region to reduce the search space for breast tumor segmentation. The second step is breast tumor segmentation performed in a weakly-supervised learning scenario where only image-level labels are available. Breast tumors are first recognized by a classification network and then segmented by the proposed class activation mapping and deep level set (CAM-DLS) method. For breast anatomy decomposition, the proposed framework achieves Dice similarity coefficient (DSC) of 83.0 ± 11.8%, 84.3 ± 10.0%, 80.7 ± 15.4% and 91.0 ± 11.4% for fat, mammary gland, muscle and thorax layers, respectively. For breast tumor recognition, the proposed framework achieves sensitivity of 95.8%, precision of 92.4%, specificity of 93.9%, accuracy of 94.8% and F1-score of 0.941. For breast tumor segmentation, the proposed framework achieves DSC of 77.3% and intersection-over-union (IoU) of 66.0%. In conclusion, the proposed framework could efficiently perform breast tumor recognition and segmentation simultaneously in a weakly-supervised setting with anatomical constraints.     

Improved Spatial Resolution for Cardiac Mapping Using Current Source Density-Based Electrode Arrays

The time of the minimum slope (i.e., the fastest negative deflection) in monopolar (MP) electrograms from normal hearts compares closely with time of phase 0 of the action potential in cells underlying the electrode, but poor rejection of far-field activity may limit the utility ofMP electrode technology in dense arrays used for the study of ventricular tachycardia and fibrillation. The purpose of this study is to evaluate more myopic discrete bipolar (BP) and nondirectional, two-dimensional current source density (CSD) based arrays for rejection of far-field potentials and precision of activation time determination. Simultaneous recordings of the CSD, MP, and multiple BP electrograms were performed on normal dog epicardium. The time of the minimum slope in MP electrograms was compared to activation times in CSD and BP derivations using: (1) peak; (2) steepest slope; (3) zero crossing of the steepest sloping segment in either direction; and (4) waveform morphology. In vivo, CSD amplitude was reduced significantly more than MP and BP amplitudes by insertion of inert media between the heart and the electrodes. The time of the steepest slope in CSD electrograms designated activation times closest to the time of the minimum slope in MP electrograms (0.9 ± 1.3 msec). We conclude that CSD provides a nondirectional electrode system that accurately defines the time of local activation and possesses better spatial specificity than MP electrode systems and BP electrode systems having the same interelectrode distances.